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The Role of Public Sector Innovation in COVID-19 Management: A Systematic Review Approach
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Strong leadership in public sector innovation can empower governments to address community challenges in new ways in light of the challenges posed by the global coronavirus pandemic. Coronavirus management policy, pandemic responses, needs, and options are reflected in various Asian countries in respective published literature, but a summarized synthesis is not available. Using a systematic review approach (PRISMA), this study has analyzed the role of leadership in public sector innovation in COVID-19 management and synthesized 23 articles from 23 different Asian countries. In the light of available data, public sector innovation (PSI) and the role played by the leadership of each country' have been found to be largely inter-dependent. The current review provides a cross-section of the ongoing nature of the pandemic, as management responses and trend data in the countries are still emerging or evolving. Additionally, our study contributes a current state report regarding the barriers facing the leadership of Asian countries in mitigating the global pandemic through PSI. Our study found that a strong political leadership presence combined with a technocratic approach and a highly-skilled public sector workforce, could lead to more tremendous success in managing the outbreak. Furthermore, religious leadership was also found to have a potentially significant role in COVID-19 management strategies.
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[
"A novel coronavirus (2019-nCoV) is causing an outbreak of viral pneumonia that started in Wuhan, China. Using the travel history and symptom onset of 88 confirmed cases that were detected outside Wuhan in the early outbreak phase, we estimate the mean incubation period to be 6.4 days (95% credible interval: 5.6-7.7), ranging from 2.1 to 11.1 days (2.5th to 97.5th percentile). These values should help inform 2019-nCoV case definitions and appropriate quarantine durations.",
"We report two cases of coronavirus disease 2019 (COVID-19) in travellers from Wuhan, China to Thailand. Both were independent introductions on separate flights, discovered with thermoscanners and confirmed with RT-PCR and genome sequencing. Both cases do not seem directly linked to the Huanan Seafood Market in Hubei but the viral genomes are identical to four other sequences from Wuhan, suggesting early spread within the city already in the first week of January."
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Hemojuvelin and the BMP-SMAD signaling pathway in the regulation of hepcidin expression in the liver
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Mutations in hemojuvelin (HJV) are the most common cause of the juvenile-onset form of the iron overload disorder hereditary hemochromatosis. The discovery that HJV functions as a co-receptor for the bone morphogenetic protein (BMP) family of signaling molecules helped to identify this signaling pathway as a central regulator of the key iron hormone hepcidin in the control of systemic iron homeostasis. This review highlights recent work uncovering the mechanism of action of HJV and the BMP-SMAD signaling pathway in regulating hepcidin expression in the liver, as well as additional studies investigating possible extra-hepatic functions of HJV. This review also explores the interaction between HJV, the BMP-SMAD signaling pathway and other regulators of hepcidin expression in systemic iron balance.
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[
"Bone morphogenetic proteins (BMPs) are members of the transforming growth factor beta superfamily that exert their effects via type I and type II serine threonine kinase receptors and the SMAD intracellular signaling pathway to regulate diverse biologic processes. Recently, we discovered that the repulsive guidance molecule (RGM) family, including RGMA, RGMB, and RGMC/hemojuvelin (HJV), function as co-receptors that enhance cellular responses to BMP ligands. Here, we use surface plasmon resonance to quantitate the binding kinetics of RGM proteins for BMP ligands. We show that among the RGMs, HJV exhibits the highest affinity for BMP6, BMP5, and BMP7 with K(D) 8.1, 17, and 20 nM respectively, versus 28, 33, and 166 nM for RGMB, and 55, 83, and 63 nM for RGMA. Conversely, RGMB exhibits preferential binding to BMP4 and BMP2 with K(D) 2.6 and 5.5 nM respectively, versus 4.5 and 9.4 nM for HJV, and 14 and 22 nM for RGMA, while RGMA exhibits the lowest binding affinity for most BMPs tested. Among the BMP ligands, RGMs exhibit the highest relative affinity for BMP4 and the lowest relative affinity for BMP7, while none of the RGMs bind to BMP9. Thus, RGMs exhibit preferential binding for distinct subsets of BMP ligands. The preferential binding of HJV for BMP6 is consistent with the functional role of HJV and BMP6 in regulating systemic iron homeostasis. Our data may help explain the mechanism by which BMPs exert cell-context specific effects via a limited number of type I and type II receptors.",
"In the present study, we correlate homozygosity for the very recently identified HJV p.R176C substitution with a juvenile hemochromatosis phenotype. We also show that the p.R176C variant fails to up-regulate the hepcidin promoter activity. Altogether, our results definitively show the R176C amino-acid change to be a novel hemojuvelin loss-of-function mutation.",
"Hereditary haemochromatosis (HH), which affects some 1 in 400 and has an estimated carrier frequency of 1 in 10 individuals of Northern European descent, results in multi-organ dysfunction caused by increased iron deposition, and is treatable if detected early. Using linkage-disequilibrium and full haplotype analysis, we have identified a 250-kilobase region more than 3 megabases telomeric of the major histocompatibility complex (MHC) that is identical-by-descent in 85% of patient chromosomes. Within this region, we have identified a gene related to the MHC class I family, termed HLA-H, containing two missense alterations. One of these is predicted to inactivate this class of proteins and was found homozygous in 83% of 178 patients. A role of this gene in haemochromatosis is supported by the frequency and nature of the major mutation and prior studies implicating MHC class I-like proteins in iron metabolism.",
"Anemia is very common in patients suffering from infections or chronic inflammation and can add substantially to the morbidity of the underlying disease. It is mediated by excessive production of the iron-regulatory peptide hepcidin, but the signaling pathway responsible for hepcidin up-regulation in the inflammatory context is still not understood completely. In the present study, we show that activin B has an unexpected but crucial role in the induction of hepcidin by inflammation. There is a dramatic induction of Inhbb mRNA, encoding the activin β(B)-subunit, in the livers of mice challenged with lipopolysaccharide, slightly preceding an increase in Smad1/5/8 phosphorylation and Hamp mRNA. Activin B also induces Smad1/5/8 phosphorylation in human hepatoma-derived cells and, synergistically with IL-6 and STAT-3 signaling, up-regulates hepcidin expression markedly, an observation confirmed in mouse primary hepatocytes. Pretreatment with a bone morphogenic protein type I receptor inhibitor showed that the effect of activin B on hepcidin expression is entirely attributable to its effect on bone morphogenetic protein signaling, most likely via activin receptor-like kinase 3. Activin B is therefore a novel and specific target for the treatment of anemia of inflammation.",
"Hepcidin is a central regulator of iron homeostasis. HFE and transferrin receptor 2 (TFR2) are mutated in adult-onset forms of hereditary hemochromatosis and regulate the expression of hepcidin in response to iron. Whether they act through the same or parallel pathways is unclear. To investigate this, we generated a mouse model with deletion of both Hfe and Tfr2 genes by crossing Hfe and Tfr2 null mice on a genetically identical background. Tissue and serum from wildtype, single-, and double-null mice were analyzed. Serum transferrin saturation and hepatic iron concentrations were determined. The expression of iron-related messenger RNA (mRNA) transcripts was analyzed by real-time polymerase chain reaction (PCR). Levels of the iron-related proteins Tfr1, Tfr2, ferritin, and prohepcidin, and the phosphorylation status of the cell signaling proteins extracellular signal-regulated kinase 1/2 (Erk1/2) and Smad1/5/8, were analyzed by immunoblotting. Double-null mice had more severe iron loading than mice lacking either Hfe or Tfr2; Tfr2 null mice had a greater iron burden than Hfe-null mice. Hepcidin expression relative to iron stores was reduced in the Hfe-null mice, with significantly lower values in the Tfr2-null mice. In the absence of both Hfe and Tfr2, hepcidin expression was reduced even further. A significant decrease in phospho-Erk1/2 in the livers of null mice and a reduction in phospho-Smad1/5/8 suggest that both the mitogen-activated protein kinase (MAPK) and bone morphogenetic protein / mothers against decapentaplegic homolog (BMP/SMAD) signaling pathways may be involved in Hfe- and Tfr2-mediated regulation of hepcidin. These studies demonstrate that iron overload due to deletion of Tfr2 is more severe than that due to Hfe, and that loss of both molecules results in pronounced iron overload. Analysis of Hfe/Tfr2 double-null mice suggests that Hfe and Tfr2 regulate hepcidin through parallel pathways involving Erk1/2 and Smad1/5/8.",
"Apoptotic cell death in pancreatic beta-cells is involved in the pathogenesis of diabetes. Signals from death receptors and DNA damage have been widely accepted as being triggers of apoptosis in beta-cells. Recent studies indicated that the endoplasmic reticulum (ER) can sense and transduce apoptotic signals. Various genetic and environmental stresses interfere with protein folding in the ER and induce ER stress. In mammals, ER stress transducer proteins IRE1, PERK and ATF6 activate both survival and apoptotic pathways. The former includes transcriptional induction of ER chaperones, translational attenuation, and ER-associated degradation (ERAD) while the latter includes transcriptional induction of CHOP/GADD153, the activation of cJUN NH(2)-terminal kinase, and the activation of caspase-12. A characteristic feature of beta-cells is the highly developed ER apparently due to a heavy engagement in insulin secretion. beta-cells are most susceptible to ER stress. The recent studies reviewed in this article revealed that ER stress-mediated apoptosis in beta-cells plays an important role in the development of diabetes.",
"Originally identified as a gene up-regulated by iron overload in mouse liver, the HEPC gene encodes hepcidin, the first mammalian liver-specific antimicrobial peptide and potential key regulator of iron metabolism. Here we demonstrate that during rat liver development, amounts of HEPC transcripts were very low in fetal liver, strongly and transiently increased shortly after birth, and reappeared in adult liver. To gain insight into mechanisms that regulate hepatic expression of hepcidin, 5'-flanking regions of human and mouse HEPC genes were isolated and analyzed by functional and DNA binding assays. Human and mouse HEPC promoter-luciferase reporter vectors exhibited strong basal activity in hepatoma HuH-7 and mouse hepatocytes, respectively, but not in non-hepatic U-2OS cells. We found that CCAAT/enhancer-binding protein alpha (C/EBPalpha) and C/EBPbeta were respectively very potent and weak activators of both human and mouse promoters. In contrast, co-expression of hepatocyte nuclear factor 4alpha (HNF4alpha) failed to induce HEPC promoter activity. By electrophoretic mobility shift assay we demonstrated that one putative C/EBP element found in the human HEPC promoter (-250/-230) predominantly bound C/EBPalpha from rat liver nuclear extracts. Hepatic deletion of the C/EBPalpha gene resulted in reduced expression of HEPC transcripts in mouse liver. In contrast, amounts of HEPC transcripts increased in liver-specific HNF4alpha-null mice. Decrease of hepcidin mRNA in mice lacking hepatic C/EBPalpha was accompanied by iron accumulation in periportal hepatocytes. Finally, iron overload led to a significant increase of C/EBPalpha protein and HEPC transcripts in mouse liver. Taken together, these data demonstrate that C/EBPalpha is likely to be a key regulator of HEPC gene transcription and provide a novel mechanism for cross-talk between the C/EBP pathway and iron metabolism.",
"Hemojuvelin (HJV), encoded by the gene HFE2, is a critical upstream regulator of hepcidin expression. Hepcidin, the central iron regulatory hormone, is secreted from hepatocytes, whereas HFE2 is highly expressed in skeletal muscle and liver. Previous studies demonstrated that HJV is a GPI-anchored protein, binds the proteins neogenin and bone morphogenetic proteins (BMP2 and BMP4), and can be released from the cell membrane (shedding). In this study, we investigated the physiological significance and the underlying mechanism of HJV shedding. In acutely iron-deficient rats with markedly suppressed hepatic hepcidin expression, we detected an early phase increase of serum HJV with no significant change of either HFE2 mRNA or protein levels in gastrocnemius muscle. Studies in both C2C12 (a mouse myoblast cell line) and HepG2 (a human hepatoma cell line) cells showed active HJV shedding, implying that both skeletal muscle and liver could be the source of serum HJV. In agreement with the observations in iron-deficient rats, HJV shedding in these cell lines was down-regulated by holo-transferrin in a concentration-dependent manner. Our present study showing that knock-down of endogenous neogenin, a HJV receptor, in C2C12 cells suppresses HJV shedding and that overexpression of neogenin in HEK293 cells markedly enhances this process, suggests that membrane HJV shedding is mediated by neogenin. The finding that neither BMP4 nor its antagonist, noggin, was able to alter HJV shedding support the lack of involvement of BMP signaling pathway in this process.",
"Iron is essential for the human being, involving in oxygen transport, energy metabolism and DNA synthesis. Iron homeostasis is tightly governed by the hepcidin-ferroportin axis, of which hepcidin is the master regulator. Excess iron is associated with various diseases including osteopenia and osteoporosis, which are closely related to the alternation of the endogenous estrogen level. To verify the biological effect of estrogen on iron metabolism, we established a mouse model of estrogen deficiency by ovariectomy. We demonstrated that the hemoglobin content and serum iron level decreased, whereas the tissue iron level in liver and spleen increased in the ovariectomized mice. Moreover, the transcription of hepatic hepcidin was elevated in ovariectomized mice compared to the control mice. We further demonstrated that there was an estrogen response element (ERE) in the promoter region of the hepcidin gene. The assay using the luciferase reporter system confirmed the existence of a functional ERE in the hepcidin promoter, as the estradiol treatment reduced hepcidin expression in cells transfected with ERE-intact construct, with no response to estradiol in cells transfected with ERE-devoid construct. In conclusion, estrogen greatly contributes to iron homeostasis by regulating hepatic hepcidin expression directly through a functional ERE in the promoter region of hepcidin gene. These findings might help build a better understanding towards the etiology of postmenopausal osteoporosis accompanied by excess tissue iron (such as iron retention of osteoclasts in bone) under estrogen deficiency.",
"The endoplasmic reticulum is responsible for much of a cell's protein synthesis and folding, but it also has an important role in sensing cellular stress. Recently, it has been shown that the endoplasmic reticulum mediates a specific set of intracellular signalling pathways in response to the accumulation of unfolded or misfolded proteins, and these pathways are collectively known as the unfolded-protein response. New observations suggest that the unfolded-protein response can initiate inflammation, and the coupling of these responses in specialized cells and tissues is now thought to be fundamental in the pathogenesis of inflammatory diseases. The knowledge gained from this emerging field will aid in the development of therapies for modulating cellular stress and inflammation."
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Mitochondrial-initiated events protect the neurovascular unit against lethal stress
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Mitochondrial-initiated events protect the neurovascular unit against lethal stress via a process called preconditioning, which independently promotes changes in cerebrovascular tone through shared signaling pathways. Activation of adenosine triphosphate (ATP)-dependent potassium channels on the inner mitochondrial membrane (mitoKATP channels) is a specific and dependable way to induce protection of neurons, astroglia, and cerebral vascular endothelium. Through the opening of mitoKATP channels, mitochondrial depolarization leads to activation of protein kinases and transient increases in cytosolic calcium (Ca(2+)) levels that activate terminal mechanisms that protect the neurovascular unit against lethal stress. The release of reactive oxygen species from mitochondria has similar protective effects. Signaling elements of the preconditioning pathways also are involved in the regulation of vascular tone. Activation of mitoKATP channels in cerebral arteries causes vasodilation, with cell-specific contributions from the endothelium, vascular smooth muscles, and nerves. Preexisting chronic conditions, such as insulin resistance and/or diabetes, prevent preconditioning and impair relaxation to mitochondrial-centered responses in cerebral arteries. Surprisingly, mitochondrial activation after anoxic or ischemic stress appears to protect cerebral vascular endothelium and promotes the restoration of blood flow; therefore, mitochondria may represent an important, but underutilized target in attenuating vascular dysfunction and brain injury in stroke patients.
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[
"Age-related disease, not aging per se, causes most morbidity in older humans. Here we report that skeletal muscle respiratory uncoupling due to UCP1 expression diminishes age-related disease in three mouse models. In a longevity study, median survival was increased in UCP mice (animals with skeletal muscle-specific UCP1 expression), and lymphoma was detected less frequently in UCP female mice. In apoE null mice, a vascular disease model, diet-induced atherosclerosis was decreased in UCP animals. In agouti yellow mice, a genetic obesity model, diabetes and hypertension were reversed by induction of UCP1 in skeletal muscle. Uncoupled mice had decreased adiposity, increased temperature and metabolic rate, elevated muscle SIRT and AMP kinase, and serum characterized by increased adiponectin and decreased IGF-1 and fibrinogen. Accelerating metabolism in skeletal muscle does not appear to impact aging but may delay age-related disease.",
"Heart cells contain ATP-sensitive potassium (KATP) channels in both the sarcolemma and the inner mitochondrial membrane. The sarcolemmal channels are believed to be heteromultimeric complexes of sulfonylurea receptors (SUR) and potassium inward rectifier (Kir) gene products, but the molecular identity of mitochondrial KATP (mitoKATP) channels remains unclear. To probe the molecular composition of KATP channels, we used adenoviral gene transfer to express wild-type (WT) and dominant-negative (AFA) constructs of Kir6.1 and 6.2 in rabbit ventricular myocytes. None of the Kir6.1 or 6.2 constructs affected mitoKATPchannel activity as assayed by confocal imaging of flavoprotein fluorescence, contradicting the proposal, based on subcellular antibody localization, that Kir6.1 forms part of mitoKATP channels. As previously reported, dominant-negative Kir6.2 gene transfer suppressed sarcolemmal KATP current, while Kir6.1 constructs had no effect on sarcolemmal activity. Immunohistochemistry with an anti-Kir6.1 antibody revealed expression of this protein in heart but no apparent co-localization with mitochondria. Thus, the available evidence indicates that both Kir6.1 and 6.2 are expressed in ventricular myocytes, but neither plays a discernible functional role in the mitoKATP channel.",
"Following focal cerebral ischemia (\"stroke\") a complex and dynamic interaction of vascular cells, glial cells, and neurons determines the extent of the ensuing lesion. Traditionally, the focus has been on mechanisms of damage, while recently it has become clear that endogenous mechanisms of protection are equally important for the final outcome. Glial cells, in particular astrocytes, have always been viewed as supporters of neuronal function. Only recently a very active role for glial cells has been emerging in physiology and pathophysiology. Not surprisingly, then, specific protective pathways have been identified by which these cells can protect or even help to regenerate brain tissue after acute insults. However, as exemplified by the existence of the glial scar, which forms around lesioned brain tissue, is composed mainly of astrocytes and plays a key role in regeneration failure, it is an oversimplification to assign merely protective functions to astrocytes. The present review will discuss the role of astrocytes in ischemic brain injury with a focus on neuroprotection in general. In this context we will consider particularly the phenomenon of \"ischemic tolerance,\" which is an experimental paradigm helpful in discriminating destructive from protective mechanisms after cerebral ischemia.",
"Pancreatic beta cells secrete insulin in response to raised blood glucose levels. This glucose-stimulated insulin secretion (GSIS) depends on mitochondrial function and is regulated by the efficiency with which oxidative metabolism is coupled to ATP synthesis. Uncoupling protein-2 (UCP2) affects this coupling efficiency and is therefore a plausible pathological and physiological regulator of GSIS. In this respect, it is important to be able to measure coupling efficiencies accurately. Here, we describe experimental protocols to determine the coupling efficiency of trypsinized INS-1E cells, a popular beta cell model, and we present practical details of our RNA interference studies to probe the effect of UCP2 knockdown on this efficiency. We also introduce a method to determine coupling efficiencies noninvasively in attached cells and discuss theoretical and practical aspects of a modular-kinetic approach to describe and understand cellular bioenergetics.",
"Mitochondria regulate intracellular calcium (Ca2+) signals in smooth muscle cells, but mechanisms mediating these effects, and the functional relevance, are poorly understood. Similarly, antihypertensive ATP-sensitive potassium (KATP) channel openers (KCOs) activate plasma membrane KATP channels and depolarize mitochondria in several cell types, but the contribution of each of these mechanisms to vasodilation is unclear. Here, we show that cerebral artery smooth muscle cell mitochondria are most effectively depolarized by diazoxide (-15%, tetramethylrhodamine [TMRM]), less so by levcromakalim, and not depolarized by pinacidil. KCO-induced mitochondrial depolarization increased the generation of mitochondria-derived reactive oxygen species (ROS) that stimulated Ca2+ sparks and large-conductance Ca2+-activated potassium (KCa) channels, leading to transient KCa current activation. KCO-induced mitochondrial depolarization and transient KCa current activation were attenuated by 5-HD and glibenclamide, KATP channel blockers. MnTMPyP, an antioxidant, and Ca2+ spark and KCa channel blockers reduced diazoxide-induced vasodilations by >60%, but did not alter dilations induced by pinacidil, which did not elevate ROS. Data suggest diazoxide drives ROS generation by inducing a small mitochondrial depolarization, because nanomolar CCCP, a protonophore, similarly depolarized mitochondria, elevated ROS, and activated transient KCa currents. In contrast, micromolar CCCP, or rotenone, an electron transport chain blocker, induced a large mitochondrial depolarization (-84%, TMRM), reduced ROS, and inhibited transient KCa currents. In summary, data demonstrate that mitochondria-derived ROS dilate cerebral arteries by activating Ca2+ sparks, that some antihypertensive KCOs dilate by stimulating this pathway, and that small and large mitochondrial depolarizations lead to differential regulation of ROS and Ca2+ sparks.",
"Mitochondrial proton and electron leak have a major impact on mitochondrial coupling efficiency and production of reactive oxygen species. In the first part of this chapter, we address the molecular nature of the basal and inducible proton leak pathways, and their physiological importance. The basal leak is unregulated, and a major proportion can be attributed to mitochondrial anion carriers, whereas the proton leak through the lipid bilayer appears to be minor. The basal proton leak is cell-type specific and correlates with metabolic rate. The inducible leak through the ANT (adenine nucleotide translocase) and UCPs (uncoupling proteins) can be activated by fatty acids, superoxide or lipid peroxidation products. The physiological role of inducible leak through UCP1 in mammalian brown adipose tissue is heat production, whereas the roles of non-mammalian UCP1 and its paralogous proteins, in particular UCP2 and UCP3, are not yet resolved. The second part of the chapter focuses on the electron leak that occurs in the mitochondrial electron transport chain. Exit of electrons prior to the reduction of oxygen to water at cytochrome c oxidase causes superoxide production. As the mechanisms of electron leak are crucial to understanding their physiological relevance, we summarize the mechanisms and topology of electron leak from complexes I and III in studies using isolated mitochondria. We also highlight recent progress and challenges of assessing electron leak in the living cell. Finally, we emphasize the importance of proton and electron leak as therapeutic targets in body mass regulation and insulin secretion.",
"Pulmonary arterial hypertension (PAH) is characterized by a hyperproliferative and anti-apoptotic diathesis within the vascular wall of the resistance pulmonary arteries, leading to vascular lumen occlusion, right ventricular failure, and death. Most current therapies show poor efficacy due to emphasis on vasodilation (rather than proliferation/apoptosis) and a lack of specificity to the pulmonary circulation. The multiple molecular abnormalities described in PAH are diverse and seemingly unrelated, calling for therapies that attack comprehensive, integrative mechanisms. Similar abnormalities also occur in cancer where a cancer-specific metabolic switch toward a non-hypoxic glycolytic phenotype is thought to be not only a result of several primary molecular or genetic abnormalities but also underlie many aspects of its resistance to apoptosis. In this paper, we review the evidence and propose that a metabolic, mitochondria-based theory can be applied in PAH. A pulmonary artery smooth muscle cell mitochondrial remodeling could integrate a number of diverse molecular abnormalities described in PAH and respond by orchestrating a switch toward a cancer-like glycolytic phenotype that drives resistance to apoptosis; via redox and calcium signals, this mitochondrial remodeling may also regulate critical transcription factors like HIF-1 and nuclear factor of activated T cells that have been described to play an important role in PAH. Because mitochondria in pulmonary arteries are quite different from mitochondria in systemic arteries, they could form the basis of relatively selective PAH therapies. This metabolic theory of PAH could facilitate the development of novel diagnostic and selective therapeutic approaches in this disease that remains deadly."
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How to generate a query for Gallia-Alumina (Ga,Al)
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Gallia-alumina (Ga,Al)
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[
"A novel catalyst material for the selective dehydrogenation of propane is presented. The catalyst consists of 1000 ppm Pt, 3 wt% Ga, and 0.25 wt% K supported on alumina. We observed a synergy between Ga and Pt, resulting in a highly active and stable catalyst. Additionally, we propose a bifunctional active phase, in which coordinately unsaturated Ga(3+) species are the active species and where Pt functions as a promoter.",
"By hydrolysis of an ethanolic gallium nitrate solution, gamma-Ga2O3 was prepared as a single-phase polymorph having a specific surface area of 160 m2 g(-1). Surface acidity and basicity of this material was studied by IR spectroscopy, using pyridine, 2,6-dimethylpyridine, acetonitrile, and carbon dioxide as spectroscopic probe molecules. For comparison, a gamma-Al2O3 sample having a surface area of 290 m2 g(-1) was also studied. On partially hydroxylated gamma-Ga2O3, the main O-H stretching bands were found at 3693 (sharp) and at 3660-3630 cm(-1) (broad), and the material proved (by adsorbed dimethylpyridine) to have a weak Brønsted acidity. Surface Lewis acidity of gamma-Ga2O3 was revealed (mainly) by adsorbed pyridine, which gave the characteristic IR absorption bands of Lewis-type adducts at 1612, 1579, 1488, and 1449 cm(-1) (values noted under an equilibrium pressure of 1 Torr at room temperature); the corresponding Lewis acid centers (coordinatively unsaturated Ga3+ ions) were found to be weaker, although more abundant, than those present on the surface of gamma-Al2O3 (unsaturated Al3+ ions). Another significant difference between gamma-Ga2O3 and gamma-Al2O3 is the smaller thermal stability of pyridine and 2,6-dimethylpyridine Lewis adducts formed on the gallium oxide. The surface basicity of gamma-Ga2O3 was studied by using carbon dioxide and deuterated acetonitrile as IR probe molecules. Adsorbed CO2 gave carbonate and hydrogen-carbonate surface species similar to those formed by gamma-Al2O3. Adsorbed acetonitrile gave rise to acetamide species, which revealed the basic character of surface O2- ions. These acetamide species were found to be more abundant on gamma-Ga2O3 than on gamma-Al2O3."
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Hydroxychloroquine and lupus nephritis
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Hydroxychloroquine (HCQ) has been recommended as a basic treatment for lupus nephritis (LN) during this decade based on its ability to improve LN-related renal immune-mediated inflammatory lesions. As a classical lysosomal inhibitor, HCQ may inhibit lysosomal degradation and disrupt protective autophagy in proximal tubular epithelial cells (PTECs). Therefore, the final renal effects of HCQ on LN need to be clarified.
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[
"The efficacy of both chloroquine and hydroxychloroquine in rheumatoid arthritis (RA) has been proved in controlled clinical trials. Despite similar chemical characteristics, it is believed the clinical efficacy of chloroquine is superior to that of hydroxychloroquine in patients with RA. Excessive production of proinflammatory cytokines was shown to contribute to the pathogenesis of RA. From different studies testing either chloroquine or hydroxychloroquine, it could be concluded that both drugs differentially inhibit cytokine production. We compared the effects of both chloroquine and hydroxychloroquine on stimulated peripheral blood mononuclear cells (PBMC) with respect to cytokine production. Therefore, PBMC were tested for tumor necrosis factor-alpha (TNF-alpha), interleukin 6 (IL-6), and interferon-gamma (IFN-gamma) by specific ELISA, after stimulation with phytohemagglutinin (PHA) or lipopolysaccharide (LPS) in the presence or absence of different concentrations of chloroquine or hydroxychloroquine. We observed that chloroquine and hydroxychloroquine equally inhibit PHA induced TNF-alpha and IFN-gamma production, and LPS induced TNF-alpha and IL-6 production, while PHA induced IL-6 production was not affected. Chloroquine and hydroxychloroquine display similar effects on PHA and LPS induced cytokine production by PBMC under identical in vitro conditions. These findings may help in understanding the mechanism of action of these drugs on RA.",
"Despite widespread clinical use of antimalarial drugs such as hydroxychloroquine and chloroquine in the treatment of rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) and other inflammatory rheumatic diseases, insights into the mechanism of action of these drugs are still emerging. Hydroxychloroquine and chloroquine are weak bases and have a characteristic 'deep' volume of distribution and a half-life of around 50 days. These drugs interfere with lysosomal activity and autophagy, interact with membrane stability and alter signalling pathways and transcriptional activity, which can result in inhibition of cytokine production and modulation of certain co-stimulatory molecules. These modes of action, together with the drug's chemical properties, might explain the clinical efficacy and well-known adverse effects (such as retinopathy) of these drugs. The unknown dose-response relationships of these drugs and the lack of definitions of the minimum dose needed for clinical efficacy and what doses are toxic pose challenges to clinical practice. Further challenges include patient non-adherence and possible context-dependent variations in blood drug levels. Available mechanistic data give insights into the immunomodulatory potency of hydroxychloroquine and provide the rationale to search for more potent and/or selective inhibitors.",
"In 2016, the American Academy of Ophthalmology (AAO) changed the recommended daily dose of hydroxychloroquine (HCQ) from 6.5 mg/kg to <5 mg/kg. However, it is not clear that the lower prescribed dose of HCQ will have the same efficacy for systemic lupus erythematosus (SLE) activity or the same role in protecting against cardiovascular risk factors and thrombosis. This study was undertaken to address the frequency of HCQ retinopathy and the role of HCQ blood levels in identifying those individuals who are at a greater future risk of retinopathy. HCQ blood levels in 537 patients with SLE from a large clinical cohort were repeatedly measured, and patients were tested for HCQ retinopathy. We assessed the risk of retinopathy according to clinical characteristics and blood levels of HCQ. The overall frequency of retinopathy was 4.3% (23 of 537 patients). There was a 1% risk of retinopathy in the first 5 years of HCQ treatment, 1.8% from 6 to 10 years, 3.3% from 11 to 15 years, 11.5% from 16 to 20 years, and 8.0% after 21 years of use. We found that older age (P < 0.0001), higher body mass index (P for trend = 0.0160), and longer duration of HCQ intake (P = 0.0024 and P for trend = 0.0006) were associated with a higher risk of HCQ toxicity. Higher blood levels of HCQ predicted later HCQ retinopathy (P = 0.0124 and P = 0.0340 for mean and maximum HCQ blood levels, respectively). Our data prove the utility of assessing blood levels of HCQ in the prediction of retinopathy. This would allow clinicians to either decrease the dose or increase monitoring in those patients with high HCQ blood levels.",
"Reduced renal function is associated with worse renal outcome in patients with lupus nephritis (LN). However, there is insufficient knowledge regarding renal function recovery in patients with LN with reduced baseline renal function. Therefore, the present study aimed to investigate renal function recovery and related factors in patients with reduced baseline renal function. The present retrospective longitudinal cohort study included patients with LN and reduced renal function. Reduced renal function was defined as an estimated glomerular filtration rate (eGFR) of <60 mL/min/1.73 m2. Recovery of renal function was determined by an eGFR of >60 mL/min/1.73 m2 at six months after baseline, and factors associated with it were evaluated using logistic regression analysis. We included 90 patients with LN, with a mean eGFR value of 37.2 ± 13.9 mL/min/1.73 m2. Forty-six (51.1%) patients recovered their renal function after six months. On multivariate analysis, hydroxychloroquine use (odds ratio (OR) = 3.891, 95% confidence interval (CI) 1.196-12.653, p = 0.024), prolonged LN (OR = 0.926, 95% CI 0.874-0.981, p = 0.009) and high-grade tubular atrophy (OR = 0.451, 95% CI 0.208-0.829, p = 0.013) were associated with renal function recovery. During follow up, 25 patients were on end-stage renal disease (ESRD). Kaplan-Meier analysis revealed that renal function recovery after six months and lower probability of ESRD are associated. In patients with LN and reduced renal function, renal function recovery at six months was associated with use of hydroxychloroquine and inversely related to longer duration of LN and higher grade of tubular atrophy.",
"Autophagy is a catabolic process that facilitates nutrient recycling via degradation of damaged organelles and proteins through lysosomal mediated degradation. Alterations in this complex, and tightly regulated process, lead to disease. Autophagy is widely accepted as cytoprotective against neurodegenerative diseases and a variety of clinical interventions are moving forward to increase autophagy as a therapeutic intervention. Autophagy has both positive and negative roles in cancer and this has led to controversy over whether or how autophagy manipulation should be attempted in cancer therapy. Nevertheless, cancer is the disease where most current activity in trying to manipulate autophagy for therapy is taking place and dozens of clinical trials are using autophagy inhibition with Chloroquine or Hydroxychloroquine in combination with other drugs for the treatment of multiple neoplasms. Here, we review recent literature implicating autophagy in neurodegenerative diseases and cancer and highlight some of the opportunities, controversies and potential pitfalls of therapeutically targeting autophagy.",
"To determine the relationship between hydroxychloroquine concentration and effect in patients with rheumatoid arthritis (RA). Using a cross sectional study design, drug concentration and effect were measured at one time. Forty-three patients with RA, receiving hydroxychloroquine therapy for at least 6 months and not receiving glucocorticosteroids, gold or penicillamine therapy were enrolled. The main outcome measures were hydroxychloroquine concentration and disease activity measured as degree of synovitis, pain, duration and intensity of morning stiffness, impairment of activities of daily living, patients' and physicians' subjective assessment of disease, erythrocyte sedimentation rate, and rheumatoid factor (RF). Hemoglobin and albumin concentrations were also recorded. Patients with none or mild intensity of morning stiffness, those with stiffness lasting less than 0.5 h and those with negative RF had significantly higher hydroxychloroquine blood concentrations than those in whom these measures indicated a more active disease (Mann-Whitney U test, p < 0.05). Similar trends were recorded for 3 other disease activity measures (p = 0.12-0.24). Analysis of all 9 individual disease activity measures indicated that the groups with less active disease had higher mean blood concentrations of hydroxychloroquine than those with measures indicating more active disease (p < 0.01). Our data provide the first evidence of a concentration-response relationship for hydroxychloroquine in RA for individual disease activity measures. However, an unweighted summed score of disease activity did not correlate significantly with drug blood concentrations. A prospective study is necessary to confirm the relationship and to determine a therapeutic concentration range.",
"Acute kidney injury (AKI) is a contributing factor in the development and progression of chronic kidney disease (CKD). Despite rapid progresses, the mechanism underlying AKI-CKD transition remains largely unclear. Animal models recapitulating this process are crucial to the research of the pathophysiology of AKI-CKD transition and the development of effective therapeutics. In this review, we present the commonly used rodent models of AKI-CKD transition, including bilateral ischemia-reperfusion injury (IRI), unilateral IRI, unilateral IRI with contralateral nephrectomy, multiple episodes of IRI, and repeated treatment of low-dose cisplatin, diphtheria toxin, aristolochic acid, or folic acid. The main merits and pitfalls of these models are also discussed. This review provides helpful information for establishing reliable and clinically relevant models for studying post-AKI development of chronic renal pathologies and the progression to CKD."
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Two distinct oligomer species of lysozyme formed during the assembly of amyloid fibrils and plaques
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There is compelling evidence that small oligomeric aggregates, emerging during the assembly of amyloid fibrils and plaques, are important molecular pathogens in many amyloid diseases. While significant progress has been made in revealing the mechanisms underlying fibril growth, understanding how amyloid oligomers fit into the fibril assembly process, and how they contribute to the pathogenesis of amyloid diseases, has remained elusive. Commonly, amyloid oligomers are considered to be metastable, early-stage precursors to fibril formation that are either on- or off-pathway from fibril growth. In addition, amyloid oligomers have been reported to colocalize with late-stage fibrils and plaques. Whether these early and late-stage oligomer species are identical or distinct, and whether both are relevant to pathogenesis remains unclear. Here we report on the formation of two distinct oligomer species of lysozyme, formed either during the early or late-stages of
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[
"Soluble Aβ oligomers are now widely recognized as key pathogenic structures in Alzheimer's disease. They inhibit synaptic function, leading to early memory deficits and synaptic degeneration, and they trigger the downstream neuronal signaling responsible for phospho-tau Alzheimer's pathology. The marginal effects observed in recent clinical studies of solanezumab, targeting monomeric Aβ, and bapineuzumab, targeting amyloid plaques, prompted expert comments that drug discovery efforts in Alzheimer's disease should focus on soluble forms of Aβ rather than fibrillar Aβ deposits found in amyloid plaques. Accumulating scientific data suggest that soluble Aβ oligomers represent the optimal intervention target within the amyloid manifold. Active drug discovery approaches include antibodies that selectively capture soluble Aβ oligomers, selective modifiers of oligomer assembly, and receptor antagonists. The onset of symptomatic clinical benefit is expected to be rapid for such agents, because neuronal memory signaling should normalize on blockage of soluble Aβ oligomers. This key feature is not shared by amyloid-lowering therapeutics, and it should translate into streamlined clinical development for oligomer-targeting drugs. Oligomer-targeting drugs should also confer long-term disease modification and slowing of disease progression, because they prevent the downstream signaling responsible for phospho-tau mediated cytoskeletal degeneration.",
"Polymorphism is a key feature of amyloid fibril structures but it remains challenging to explain these variations for a particular sample. Here, we report electron cryomicroscopy-based reconstructions from different fibril morphologies formed by a peptide fragment from an amyloidogenic immunoglobulin light chain. The observed fibril morphologies vary in the number and cross-sectional arrangement of a structurally conserved building block. A comparison with the theoretically possible constellations reveals the experimentally observed spectrum of fibril morphologies to be governed by opposing sets of forces that primarily arise from the β-sheet twist, as well as peptide-peptide interactions within the fibril cross-section. Our results provide a framework for rationalizing and predicting the structure and polymorphism of cross-β fibrils, and suggest that a small number of physical parameters control the observed fibril architectures.",
"Point mutations in the amyloid-β (Aβ) coding region produce a combination of mutant and WT Aβ isoforms that yield unique clinicopathologies in familial Alzheimer's disease (fAD) and cerebral amyloid angiopathy (fCAA) patients. Here, we report a method to investigate the structural variability of amyloid deposits found in fAD, fCAA, and sporadic AD (sAD). Using this approach, we demonstrate that mutant Aβ determines WT Aβ conformation through prion template-directed misfolding. Using principal component analysis of multiple structure-sensitive fluorescent amyloid-binding dyes, we assessed the conformational variability of Aβ deposits in fAD, fCAA, and sAD patients. Comparing many deposits from a given patient with the overall population, we found that intrapatient variability is much lower than interpatient variability for both disease types. In a given brain, we observed one or two structurally distinct forms. When two forms coexist, they segregate between the parenchyma and cerebrovasculature, particularly in fAD patients. Compared with sAD samples, deposits from fAD patients show less intersubject variability, and little overlap exists between fAD and sAD deposits. Finally, we examined whether E22G (Arctic) or E22Q (Dutch) mutants direct the misfolding of WT Aβ, leading to fAD-like plaques in vivo. Intracerebrally injecting mutant Aβ40 fibrils into transgenic mice expressing only WT Aβ induced the deposition of plaques with many biochemical hallmarks of fAD. Thus, mutant Aβ40 prions induce a conformation of WT Aβ similar to that found in fAD deposits. These findings indicate that diverse AD phenotypes likely arise from one or more initial Aβ prion conformations, which kinetically dominate the spread of prions in the brain.",
"Age-related neurodegenerative diseases share a number of important pathological features, such as accumulation of misfolded proteins as amyloid oligomers and fibrils. Recent evidence suggests that soluble amyloid oligomers and not the insoluble amyloid fibrils may represent the primary pathological species of protein aggregates. We have produced several monoclonal antibodies that specifically recognize prefibrillar oligomers and do not recognize amyloid fibrils, monomer or natively folded proteins. Like the polyclonal antisera, the individual monoclonals recognize generic epitopes that do not depend on a specific linear amino acid sequence, but they display distinct preferences for different subsets of prefibrillar oligomers. Immunological analysis of a number of different prefibrillar Aβ oligomer preparations show that structural polymorphisms exist in Aβ prefibrillar oligomers that can be distinguished on the basis of their reactivity with monoclonal antibodies. Western blot analysis demonstrates that the conformers defined by the monoclonal antibodies have distinct size distributions, indicating that oligomer structure varies with size. The different conformational types of Aβ prefibrillar oligomers can serve as they serve as templates for monomer addition, indicating that they seed the conversion of Aβ monomer into more prefibrillar oligomers of the same type. These results indicate that distinct structural variants or conformers of prefibrillar Aβ oligomers exist that are capable of seeding their own replication. These conformers may be analogous to different strains of prions.",
"Amyloid diseases, including Alzheimer's, Parkinson's, and the prion conditions, are each associated with a particular protein in fibrillar form. These amyloid fibrils were long suspected to be the disease agents, but evidence suggests that smaller, often transient and polymorphic oligomers are the toxic entities. Here, we identify a segment of the amyloid-forming protein αB crystallin, which forms an oligomeric complex exhibiting properties of other amyloid oligomers: β-sheet-rich structure, cytotoxicity, and recognition by an oligomer-specific antibody. The x-ray-derived atomic structure of the oligomer reveals a cylindrical barrel, formed from six antiparallel protein strands, that we term a cylindrin. The cylindrin structure is compatible with a sequence segment from the β-amyloid protein of Alzheimer's disease. Cylindrins offer models for the hitherto elusive structures of amyloid oligomers.",
"Despite significant advances, the molecular identity of the cytotoxic species populated during in vivo amyloid formation crucial for the understanding of neurodegenerative disorders is yet to be revealed. In this study lysozyme prefibrillar oligomers and fibrils in both mature and sonicated states have been isolated through an optimized ultrafiltration/ultracentrifugation method and characterized with various optical spectroscopic techniques, atomic force microscopy, and transmission electron microscopy. We examined their level and mode of toxicity on rat pheochromocytoma (PC12) cells in both differentiated and undifferentiated states. We find that oligomers and fibrils display cytotoxic capabilities toward cultured cells in vitro, with oligomers producing elevated levels of cellular injury toward undifferentiated PC12 cells (PC12(undiff)). Furthermore, dual flow cytometry staining experiments demonstrate that the oligomers and mature fibrils induce divergent cellular death pathways (apoptosis and secondary necrosis, respectively) in these PC12 cells. We have also shown that oligomers but not sonicated mature fibrils inhibit hippocampal long term potentiation, a form of synaptic plasticity implicated in learning and memory, in vivo. We conclude that our in vitro and in vivo findings confer a level of resistance toward amyloid fibrils, and that the PC 12-based comparative cytotoxicity assay can provide insights into toxicity differences between differently aggregated protein species.",
"Solid-state nuclear magnetic resonance (NMR) measurements have made major contributions to our understanding of the molecular structures of amyloid fibrils, including fibrils formed by the beta-amyloid peptide associated with Alzheimer's disease, by proteins associated with fungal prions, and by a variety of other polypeptides. Because solid-state NMR techniques can be used to determine interatomic distances (both intramolecular and intermolecular), place constraints on backbone and side-chain torsion angles, and identify tertiary and quaternary contacts, full molecular models for amyloid fibrils can be developed from solid-state NMR data, especially when supplemented by lower-resolution structural constraints from electron microscopy and other sources. In addition, solid-state NMR data can be used as experimental tests of various proposals and hypotheses regarding the mechanisms of amyloid formation, the nature of intermediate structures, and the common structural features within amyloid fibrils. This review introduces the basic experimental and conceptual principles behind solid-state NMR methods that are applicable to amyloid fibrils, reviews the information about amyloid structures that has been obtained to date with these methods, and discusses how solid-state NMR data provide insights into the molecular interactions that stabilize amyloid structures, the generic propensity of polypeptide chains to form amyloid fibrils, and a number of related issues that are of current interest in the amyloid field."
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Environmental factors associated with uveal coloboma.
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Uveal coloboma is a condition defined by missing ocular tissues and is a significant cause of childhood blindness. It occurs from a failure of the optic fissure to close during embryonic development and may lead to missing parts of the iris, ciliary body, retina, choroid, and optic nerve. Because there is no treatment for coloboma, efforts have focused on prevention. While several genetic causes of coloboma have been identified, little definitive research exists regarding the environmental causes of this condition. We review the current literature on environmental factors associated with coloboma in an effort to guide future research and preventative counseling related to this condition.
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[
"In a recent study, high maternal periconceptional intake of vitamin E was found to be associated with risk of congenital heart defects (CHDs). To explore this association further, we investigated the association between total daily vitamin E intake and selected birth defects. We analyzed data from 4525 controls and 8665 cases from the 1997 to 2005 National Birth Defects Prevention Study. We categorized estimated periconceptional energy-adjusted total daily vitamin E intake from diet and supplements into quartiles (referent, lowest quartile). Associations between quartiles of energy-adjusted vitamin E intake and selected birth defects were adjusted for demographic, lifestyle, and nutritional factors. We observed a statistically significant association with the third quartile of vitamin E intake (odds ratio [OR], 1.17; 95% confidence interval [CI], 1.01-1.35) and all CHDs combined. Among CHD sub-types, we observed associations with left ventricular outflow tract obstruction defects, and its sub-type, coarctation of the aorta and the third quartile of vitamin E intake. Among defects other than CHDs, we observed associations between anorectal atresia and the third quartile of vitamin E intake (OR, 1.66; 95% CI, 1.01-2.72) and hypospadias and the fourth quartile of vitamin E intake (OR, 1.42; 95% CI, 1.09-1.87). Selected quartiles of energy-adjusted estimated total daily vitamin E intake were associated with selected birth defects. However, because these few associations did not exhibit exposure-response patterns consistent with increasing risk associated with increasing intake of vitamin E, further studies are warranted to corroborate our findings.",
"Recent Zika virus (ZIKV) outbreaks have been associated with an increased incidence of neonatal microcephaly. Subsequently, tropism for the brain was established in human fetal brain tissue. We present the first congenital ZIKV infection in the United States, confirmed by high ZIKV immunoglobulin M antibody titers in serum and cerebrospinal fluid. The phenotypic characteristics of the patient fall within fetal brain disruption sequence, suggesting impaired brain development in the second half of gestation. Brain imaging revealed an almost agyric brain with diffuse parenchymal calcifications, hydrocephalus ex vacuo, and cerebellar hypoplasia. Ophthalmologic examination revealed macular pigment stippling and optic nerve atrophy. Liver, lungs, heart, and bone marrow were not affected. The patient had progressive neurologic deterioration in the first month of life. The discovery of ZIKV infection in human fetal brain tissue along with serologic confirmation proves the vertical transmission of ZIKV. Therefore, ZIKV has joined the group of congenital infections.",
"The eye is a very complex structure derived from the neural tube, surface ectoderm, and migratory mesenchyme from a neural crest origin. Because structures that evolve from the neural tube may be affected by a folate/folic acid (FA) deficiency, the aim of this work was to investigate whether a maternal folic acid-deficient diet may cause developmental alterations in the mouse eye. Female C57BL/6J mice (8 weeks old) were assigned into two different folic acid groups for periods ranging between 2 and 16 weeks. Animals were killed at gestation day 17. Hepatic folate was analyzed, and the eyes from 287 fetuses were macroscopically studied, sectioned and immunolabeled with anti-transforming growth factor (TGF)-β2 and anti-TGF-βRII. Mice exposed to a FA-deficient diet exhibited numerous eye macroscopic anomalies, such as anophthalmia and microphthalmia. Microscopically, the eye was the most affected organ (43.7% of the fetuses). The highest incidence of malformations occurred from the 8th week onward. A statistically significant linear association between the number of maternal weeks on the FA-deficient diet and embryonic microscopic eye malformations was observed. The optic cup derivatives and structures forming the eye anterior segment showed severe abnormalities. In addition, TGF-β2 and TGF-βRII expression in the eye was also altered. This study suggests that an adequate folic acid/folate status plays a key role in the formation of ocular tissues and structures, whereas a vitamin deficiency is negatively associated with a normal eye development even after a short-term exposure.",
"Clinical hyperthyroidism has been associated with an increased risk of maternal, fetal, and neonatal complications. The available antithyroid drugs are methimazole/carbimazole and propylthiouracil. Several case reports and some epidemiologic studies suggest that methimazole/carbimazole exposure during the first trimester of pregnancy is associated with an increased risk of congenital malformations, including ectodermal anomalies, choanal atresia, esophageal atresia, and omphalocele. However, the absolute risk appears to be very small, and it remains unclear whether the association is driven by the maternal disease, the medication, or the combination of both factors. Propylthiouracil exposure has not been associated with an increased risk of congenital malformations and is the recommended drug during the first trimester of pregnancy. Since propylthiouracil-induced hepatotoxicity has been reported in approximately 0.1% of exposed adults and the number of case-reports of severe liver injury is increasing, treatment with low dose methimazole during the second and third trimesters should be considered. Until now, there has been no evidence that children prenatally exposed to methimazole/carbimazole or propylthiouracil have an increased risk of neurodevelopmental delay.",
"Studies comparing risk of specific congenital malformations (CM) between multiple pregnancies resulting from IVF/intracytoplasmic sperm injection (ICSI) and those conceived naturally report conflicting results; furthermore, there is a lack of a complete overview. This meta-analysis aimed to address which types of CM are increased in IVF/ICSI multiple pregnancies compared with those conceived naturally. All studies testing the association between IVF/ICSI multiple pregnancies and specific CM identified in various databases were considered. The literature search yielded 856 records, of which 21 cohort studies were included for analysis. Overall, multiple pregnancies achieved with IVF/ICSI experienced a significantly higher risk of chromosomal defects (relative risk [RR] = 1.36; 95% confidence interval [CI]: 1.04-1.77), urogenital (RR = 1.18; 95% CI: 1.03-1.36) and circulatory (RR = 1.22; 95% CI: 1.01-1.47) system malformations. However, the remaining specific CM, such as cleft lip and/or palate, eye, ear, face and neck, respiratory, musculoskeletal, nervous and digestive system malformations, were similar in the two groups. No substantial heterogeneity was observed for most outcomes except for digestive (P = 0.094; I2 = 38.3%) and circulatory (P = 0.070; I2 = 35.2%) system malformations. These findings provide additional information on risks of IVF/ICSI for use when counselling patients.",
"Zika virus infection can be prenatally passed from a pregnant woman to her fetus. There is sufficient evidence to conclude that intrauterine Zika virus infection is a cause of microcephaly and serious brain anomalies, but the full spectrum of anomalies has not been delineated. To inform pediatric clinicians who may be called on to evaluate and treat affected infants and children, we review the most recent evidence to better characterize congenital Zika syndrome. We reviewed published reports of congenital anomalies occurring in fetuses or infants with presumed or laboratory-confirmed intrauterine Zika virus infection. We conducted a comprehensive search of the English literature using Medline and EMBASE for Zika from inception through September 30, 2016. Congenital anomalies were considered in the context of the presumed pathogenetic mechanism related to the neurotropic properties of the virus. We conclude that congenital Zika syndrome is a recognizable pattern of structural anomalies and functional disabilities secondary to central and, perhaps, peripheral nervous system damage. Although many of the components of this syndrome, such as cognitive, sensory, and motor disabilities, are shared by other congenital infections, there are 5 features that are rarely seen with other congenital infections or are unique to congenital Zika virus infection: (1) severe microcephaly with partially collapsed skull; (2) thin cerebral cortices with subcortical calcifications; (3) macular scarring and focal pigmentary retinal mottling; (4) congenital contractures; and (5) marked early hypertonia and symptoms of extrapyramidal involvement. Although the full spectrum of adverse reproductive outcomes caused by Zika virus infection is not yet determined, a distinctive phenotype-the congenital Zika syndrome-has emerged. Recognition of this phenotype by clinicians for infants and children can help ensure appropriate etiologic evaluation and comprehensive clinical investigation to define the range of anomalies in an affected infant as well as determine essential follow-up and ongoing care.",
"We describe Zika virus (ZIKV) vertical transmission in an imported case in Spain, in a 17-week pregnant woman. ZIKV IgG, IgM and RNA were detected in serum in week 17. At 19 weeks, ultrasound scan revealed fetal malformations and ZIKV was detected in the amniotic fluid. Pregnancy was terminated at week 21; autopsy of the fetus revealed bilateral hydrocephalus, brain microcalcifications and arthrogryposis multiplex congenita. ZIKV was detected in the umbilical cord and brain tissue."
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Mononuclear phagocytes in atherosclerotic plaques
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Mononuclear phagocytes (MPs) relevant to atherosclerosis include monocytes, macrophages, and dendritic cells. A decade ago, studies on macrophage behavior in atherosclerotic lesions were often limited to quantification of total macrophage area in cross-sections of plaques. Although technological advances are still needed to examine plaque MP populations in an increasingly dynamic and informative manner, innovative methods to interrogate the biology of MPs in atherosclerotic plaques developed in the past few years point to several mechanisms that regulate the accumulation and function of MPs within plaques. Here, I review the evolution of atherosclerotic plaques with respect to changes in the MP compartment from the initiation of plaque to its progression and regression, discussing the roles that recruitment, proliferation, and retention of MPs play at these different disease stages. Additional work in the future will be needed to better distinguish macrophages and dendritic cells in plaque and to address some basic unknowns in the field, including just how cholesterol drives accumulation of macrophages in lesions to build plaques in the first place and how macrophages as major effectors of innate immunity work together with components of the adaptive immune response to drive atherosclerosis. Answers to these questions are sought with the goal in mind of reversing disease where it exists and preventing its development where it does not.
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"Leukocytosis is associated with increased cardiovascular disease risk in humans and develops in hypercholesterolemic atherosclerotic animal models. Leukocytosis is associated with the proliferation of hematopoietic stem and multipotential progenitor cells (HSPCs) in mice with deficiencies of the cholesterol efflux-promoting ABC transporters ABCA1 and ABCG1 in BM cells. Here, we have determined the role of endogenous apolipoprotein-mediated cholesterol efflux pathways in these processes. In Apoe⁻/⁻ mice fed a chow or Western- type diet, monocytosis and neutrophilia developed in association with the proliferation and expansion of HSPCs in the BM. In contrast, Apoa1⁻/⁻ mice showed no monocytosis compared with controls. ApoE was found on the surface of HSPCs, in a proteoglycan-bound pool, where it acted in an ABCA1- and ABCG1-dependent fashion to decrease cell proliferation. Accordingly, competitive BM transplantation experiments showed that ApoE acted cell autonomously to control HSPC proliferation, monocytosis, neutrophilia, and monocyte accumulation in atherosclerotic lesions. Infusion of reconstituted HDL and LXR activator treatment each reduced HSPC proliferation and monocytosis in Apoe⁻/⁻ mice. These studies suggest a specific role for proteoglycanbound ApoE at the surface of HSPCs to promote cholesterol efflux via ABCA1/ABCG1 and decrease cell proliferation, monocytosis, and atherosclerosis. Although endogenous apoA-I was ineffective, pharmacologic approaches to increasing cholesterol efflux suppressed stem cell proliferative responses.",
"Inflammation underlies a wide variety of physiological and pathological processes. Although the pathological aspects of many types of inflammation are well appreciated, their physiological functions are mostly unknown. The classic instigators of inflammation - infection and tissue injury - are at one end of a large range of adverse conditions that induce inflammation, and they trigger the recruitment of leukocytes and plasma proteins to the affected tissue site. Tissue stress or malfunction similarly induces an adaptive response, which is referred to here as para-inflammation. This response relies mainly on tissue-resident macrophages and is intermediate between the basal homeostatic state and a classic inflammatory response. Para-inflammation is probably responsible for the chronic inflammatory conditions that are associated with modern human diseases.",
"Atherosclerosis is an inflammatory disease in which leukocytes and oxidatively modified lipids accumulate in the arterial intima. Previously, we showed that dendritic cells (DCs) accumulate preferentially in regions predisposed to atherosclerosis in the normal murine aortic intima. The function of these cells in atherogenesis is unknown. Our goal was to determine the role of resident intimal DCs in the initiation of atherosclerosis. En face immunostaining of nascent atherosclerotic lesions in low-density lipoprotein receptor-deficient (Ldlr(-/-)) mice fed a cholesterol-rich diet for 5 or 10 days demonstrated that foam cells expressed DC markers CD11c, 33D1, and major histocompatibility complex class II. Transmission electron microscopy revealed that the majority of intimal lipid was intracellular. The role of resident intimal DCs in lesion formation was verified by their conditional depletion using transgenic mice expressing the simian diphtheria toxin receptor in CD11c(+) cells. A single injection of diphtheria toxin depleted intimal CD11c(+) DCs by >98% within 24 hours, with 25% and 75% recovery at 1 and 3 weeks, respectively. When bred onto the Ldlr(-/-) background, intimal DC depletion with diphtheria toxin during 5 days of lesion formation reduced the intimal lipid area by 55% relative to undepleted controls. Transmission electron microscopy revealed few foam cells in DC-depleted mice and abundant accumulation of subendothelial extracellular lipid. Induction of hypercholesterolemia in mice triggers rapid ingestion of lipid by resident intimal DCs, which initiate nascent foam cell lesion formation.",
"We have previously shown that mouse atherosclerosis regression involves monocyte-derived (CD68+) cell emigration from plaques and is dependent on the chemokine receptor CCR7. Concurrent with regression, mRNA levels of the gene encoding LXRalpha are increased in plaque CD68+ cells, suggestive of a functional relationship between LXR and CCR7. To extend these results, atherosclerotic Apoe-/- mice sufficient or deficient in CCR7 were treated with an LXR agonist, resulting in a CCR7-dependent decrease in plaque CD68+ cells. To test the requirement for LXR for CCR7-dependent regression, we transplanted aortic arches from atherosclerotic Apoe-/- mice, or from Apoe-/- mice with BM deficiency of LXRalpha or LXRbeta, into WT recipients. Plaques from both LXRalpha and LXRbeta-deficient Apoe-/- mice exhibited impaired regression. In addition, the CD68+ cells displayed reduced emigration and CCR7 expression. Using an immature DC line, we found that LXR agonist treatment increased Ccr7 mRNA levels. This increase was blunted when LXRalpha and LXRbeta levels were reduced by siRNAs. Moreover, LXR agonist treatment of primary human immature DCs resulted in functionally significant upregulation of CCR7. We conclude that LXR is required for maximal effects on plaque CD68+ cell expression of CCR7 and monocyte-derived cell egress during atherosclerosis regression in mice.",
"Both reductions in atherogenic lipoproteins and increases in high-density lipoprotein (HDL) levels may affect atherosclerosis regression. Here, the relative potential of low-density lipoprotein (LDL) lowering and HDL raising gene transfer strategies to induce regression of complex murine atherosclerotic lesions was directly compared. Male C57BL/6 LDL receptor (LDLr)(-/-) mice were fed an atherogenic diet (1.25% cholesterol and 10% coconut oil) to induce advanced atherosclerotic lesions. A baseline group was killed after 6 months and remaining mice were randomized into a control progression (Adnull or saline), an apolipoprotein (apo) A-I (AdA-I), an LDLr (AdLDLr), or a combined apo A-I/LDLr (AdA-I/AdLDLr) adenoviral gene transfer group and followed-up for another 12 weeks with continuation of the atherogenic diet. Gene transfer with AdLDLr decreased non-HDL cholesterol levels persistently by 95% (p < 0.001) compared with baseline. This drastic reduction of non-HDL cholesterol levels induced lesion regression by 28% (p < 0.001) in the aortic root and by 25% (p < 0.05) in the brachiocephalic artery at 12 weeks after transfer. Change in lesion size was accompanied by enhanced plaque stability, as evidenced by increased collagen content, reduced lesional macrophage content, a drastic reduction of necrotic core area, and decreased expression of inflammatory genes. Elevated HDL cholesterol following AdA-I transfer increased collagen content in lesions, but did not induce regression. Apo A-I gene transfer on top of AdLDLr transfer resulted in additive effects, particularly on inflammatory gene expression. In conclusion, drastic lipid lowering induced by a powerful gene transfer strategy leads to pronounced regression and stabilization of advanced murine atherosclerosis.",
"HDL cholesterol (HDL-C) plasma levels are inversely related to cardiovascular disease risk. Previous studies have shown in animals and humans that HDL promotes regression of atherosclerosis. We hypothesized that this was related to an ability to promote the loss of monocyte-derived cells (CD68(+), primarily macrophages and macrophage foam cells) from plaques. To test this hypothesis, we used an established model of atherosclerosis regression in which plaque-bearing aortic arches from apolipoprotein E-deficient (apoE(-/-)) mice (low HDL-C, high non-HDL-C) were transplanted into recipient mice with differing levels of HDL-C and non-HDL-C: C57BL6 mice (normal HDL-C, low non-HDL-C), apoAI(-/-) mice (low HDL-C, low non-HDL-C), or apoE(-/-) mice transgenic for human apoAI (hAI/apoE(-/-); normal HDL-C, high non-HDL-C). Remarkably, despite persistent elevated non-HDL-C in hAI/apoE(-/-) recipients, plaque CD68(+) cell content decreased by >50% by 1 wk after transplantation, whereas there was little change in apoAI(-/-) recipient mice despite hypolipidemia. The decreased content of plaque CD68(+) cells after HDL-C normalization was associated with their emigration and induction of their chemokine receptor CCR7. Furthermore, in CD68(+) cells laser-captured from the plaques, normalization of HDL-C led to decreased expression of inflammatory factors and enrichment of markers of the M2 (tissue repair) macrophage state. Again, none of these beneficial changes were observed in the apoAI(-/-) recipients, suggesting a major requirement for reverse cholesterol transport for the beneficial effects of HDL. Overall, these results establish HDL as a regulator in vivo of the migratory and inflammatory properties of monocyte-derived cells in mouse atherosclerotic plaques, and highlight the phenotypic plasticity of these cells.",
"Atherosclerosis is a chronic inflammatory disease of large and medium-sized arteries characterized by leukocyte accumulation in the vessel wall. Both innate and adaptive immune responses contribute to atherogenesis, but the identity of atherosclerosis-relevant antigens and the role of antigen presentation in this disease remain poorly characterized. We developed live-cell imaging of explanted aortas to compare the behavior and role of APCs in normal and atherosclerotic mice. We found that CD4+ T cells were capable of interacting with fluorescently labeled (CD11c-YFP+) APCs in the aortic wall in the presence, but not the absence, of cognate antigen. In atherosclerosis-prone Apoe-/-CD11c-YFP+ mice, APCs extensively interacted with CD4+ T cells in the aorta, leading to cell activation and proliferation as well as secretion of IFN-γ and TNF-α. These cytokines enhanced uptake of oxidized and minimally modified LDL by macrophages. We conclude that antigen presentation by APCs to CD4+ T cells in the arterial wall causes local T cell activation and production of proinflammatory cytokines, which promote atherosclerosis by maintaining chronic inflammation and inducing foam cell formation.",
"Chronic inflammation is a fundamental aspect of metabolic disorders such as obesity, diabetes and cardiovascular disease. Cholesterol crystals are metabolic signals that trigger sterile inflammation in atherosclerosis, presumably by activating inflammasomes for IL-1β production. We found here that atherogenesis was mediated by IL-1α and we identified fatty acids as potent inducers of IL-1α-driven vascular inflammation. Fatty acids selectively stimulated the release of IL-1α but not of IL-1β by uncoupling mitochondrial respiration. Fatty acid-induced mitochondrial uncoupling abrogated IL-1β secretion, which deviated the cholesterol crystal-elicited response toward selective production of IL-1α. Our findings delineate a previously unknown pathway for vascular immunopathology that links the cellular response to metabolic stress with innate inflammation, and suggest that IL-1α, not IL-1β, should be targeted in patients with cardiovascular disease.",
"A defined role in the atherogenic sequence is proposed for the circulating monocyte. The author has been able to demonstrate a \"monocyte clearance system\" in which large numbers of circulating monocytes invade the intima of lesion-prone areas in arteries, become phagocytic, and accumulate lipid. A fatty cell lesion results. Once lipid-laden, foam cells migrate back into the bloodstream by crossing the arterial endothelium. The ratio of penetrating monocytes to emerging foam cells decreases as fatty cell lesions develop until a one-to-one ratio is achieved in late fatty cell lesions, which do not progress further. Advanced fibroatherosclerotic plaques in the same animals do not show the same characteristics and have smooth muscle cell involvement. It would appear that advancement of the lesion is at least partially a result of failure of the monocyte clearance system to remove sufficient lipid. The invasion of monocytes and endothelial damage caused by foam cell clearance may, in late fatty lesions, contribute to plaque evolution by introducing growth factors from macrophages and platelets and allowing greater lipid influx. Elucidation of this system was facilitated by the examination of vessels from diet initiation onwards and by the observation of late nonprogressing fatty cell lesions. It is possible that this system exists in other models but has been overlooked by a predilection for the study of advanced lesions that prevails in the literature.",
"Work in recent years has shown an essential role for Toll-like receptors (TLRs) in the activation of innate and adaptive immunity in vertebrate animals. These germ-line encoded receptors, expressed on a diverse variety of cells and tissues, recognize conserved molecular products derived from various classes of pathogens, including Gram-positive and -negative bacteria, DNA and RNA viruses, fungi and protozoa. Ligand recognition induces a conserved host defense program, which includes production of inflammatory cytokines, upregulation of costimulatory molecules, and induction of antimicrobial defenses. Importantly, activation of dendritic cells by TLR ligands is necessary for their maturation and consequent ability to initiate adaptive immune responses. How responses are tailored by individual TLRs to contain specific classes of pathogens is not yet clear."
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Changes in gene expression in blood cells during egg oral immunotherapy in children
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We previously reported the results of a randomized, open-label trial of egg oral immunotherapy (OIT) in 50 children where 44% were desensitized and 46% were partially desensitized after 8 months of treatment. Here we focus on cell-mediated molecular mechanisms driving desensitization during egg OIT. We sought to determine whether changes in genome-wide gene expression in blood cells during egg OIT correlate with humoral responses and the clinical outcome. The blood cell transcriptome of 50 children receiving egg OIT was profiled using peripheral blood mononuclear cell (PBMC) samples obtained at baseline and after 3 and 8 months of OIT. We identified 467 differentially expressed genes (DEGs) after 3 or 8 months of egg OIT. At 8 months, 86% of the DEGs were downregulated and played a role in the signaling of TREM1, IL-6, and IL-17. In correlation analyses, Gal d 1-4-specific IgG4 antibodies associated positively with DEGs playing a role in pathogen recognition and antigen presentation and negatively with DEGs playing a role in the signaling of IL-10, IL-6, and IL-17. Desensitized and partially desensitized patients had differences in their antibody responses, and although most of the transcriptomic changes were shared, both groups had also specific patterns, which suggest slower changes in partially desensitized and activation of NK cells in the desensitized group. OIT for egg allergy in children inhibits inflammation and activates innate immune responses regardless of the clinical outcome at 8 months. Changes in gene expression patterns first appear as posttranslational protein modifications, followed by more sustained epigenetic gene regulatory functions related to successful desensitization.
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[
"Long non-coding RNAs (lncRNAs) are diverse transcription products emanating from thousands of loci in mammalian genomes. Cis-acting lncRNAs, which constitute a substantial fraction of lncRNAs with an attributed function, regulate gene expression in a manner dependent on the location of their own sites of transcription, at varying distances from their targets in the linear genome. Through various mechanisms, cis-acting lncRNAs have been demonstrated to activate, repress or otherwise modulate the expression of target genes. We discuss the activities that have been ascribed to cis-acting lncRNAs, the evidence and hypotheses regarding their modes of action, and the methodological advances that enable their identification and characterization. The emerging principles highlight lncRNAs as transcriptional units highly adept at contributing to gene regulatory networks and to the generation of fine-tuned spatial and temporal gene expression programmes.",
"We introduce CIBERSORT, a method for characterizing cell composition of complex tissues from their gene expression profiles. When applied to enumeration of hematopoietic subsets in RNA mixtures from fresh, frozen and fixed tissues, including solid tumors, CIBERSORT outperformed other methods with respect to noise, unknown mixture content and closely related cell types. CIBERSORT should enable large-scale analysis of RNA mixtures for cellular biomarkers and therapeutic targets (http://cibersort.stanford.edu/).",
"The development of microarrays permits us to monitor transcriptomes on a genome-wide scale. To validate microarray measurements, quantitative-real time-reverse transcription PCR (Q-RT-PCR) is one of the most robust and commonly used approaches. The new challenge in gene quantification analysis is how to explicitly incorporate statistical estimation in such studies. In the realm of statistical analysis, the various available methods of the probe level normalization for microarray analysis may result in distinctly different target selections and variation in the scores for the correlation between microarray and Q-RT-PCR. Moreover, it remains a major challenge to identify a proper internal control for Q-RT-PCR when confirming microarray measurements. Sixty-six Affymetrix microarray slides using lung adenocarcinoma tissue RNAs were analyzed by a statistical re-sampling method in order to detect genes with minimal variation in gene expression. By this approach, we identified DDX5 as a novel internal control for Q-RT-PCR. Twenty-three genes, which were differentially expressed between adjacent normal and tumor samples, were selected and analyzed using 24 paired lung adenocarcinoma samples by Q-RT-PCR using two internal controls, DDX5 and GAPDH. The percentage correlation between Q-RT-PCR and microarray were 70% and 48% by using DDX5 and GAPDH as internal controls, respectively. Together, these quantification strategies for Q-RT-PCR data processing procedure, which focused on minimal variation, ought to significantly facilitate internal control evaluation and selection for Q-RT-PCR when corroborating microarray data.",
"We propose a new class of support vector algorithms for regression and classification. In these algorithms, a parameter nu lets one effectively control the number of support vectors. While this can be useful in its own right, the parameterization has the additional benefit of enabling us to eliminate one of the other free parameters of the algorithm: the accuracy parameter epsilon in the regression case, and the regularization constant C in the classification case. We describe the algorithms, give some theoretical results concerning the meaning and the choice of nu, and report experimental results."
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Biomechanics of tomato fruit cuticle
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The cuticle is the most external layer that protects fruits from the environment and constitutes the first shield against physical impacts. The preservation of its mechanical integrity is essential to avoid the access to epidermal cell walls and to prevent mass loss and damage that affect the commercial quality of fruits. The rheology of the cuticle is also very important to respond to the size modification along fruit growth and to regulate the diffusion of molecules from and toward the atmosphere. The mechanical performance of cuticles is regulated by the amount and assembly of its components (mainly cutin, polysaccharides, and waxes). In tomato fruit cuticles, phenolics, a minor cuticle component, have been found to have a strong influence on their mechanical behavior. To fully characterize the biomechanics of tomato fruit cuticle, transient creep, uniaxial tests, and multi strain dynamic mechanical analysis (DMA) measurements have been carried out. Two well-differentiated stages have been identified. At early stages of growth, characterized by a low phenolic content, the cuticle displays a soft elastic behavior. Upon increased phenolic accumulation during ripening, a progressive stiffening is observed. The increment of viscoelasticity in ripe fruit cuticles has also been associated with the presence of these compounds. The transition from the soft elastic to the more rigid viscoelastic regime can be explained by the cooperative association of phenolics with both the cutin and the polysaccharide fractions.
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"Rheological properties were determined for cuticular membranes (CMs) enzymatically isolated from mature tomato (Lycopersicon esculentum Mill. cv Pik Red) fruit. The cuticle responded as a viscoelastic polymer in stress-strain studies. Both CM and dewaxed CM expanded and became more elastic and susceptible to fracture when hydrated, suggesting that water plasticized the cuticle. Dewaxing of the CM caused similar changes in elasticity and fracturing, indicating that wax may serve as a supporting filler in the cutin matrix. Exposure of the cuticle to the surfactant Triton X-100 did not significantly affect its rheological properties.",
"The cuticle covers the aerial epidermis of land plants and plays a primary role in water regulation and protection from external stresses. Remarkable species diversity in the structure and composition of its components, cutin and wax, have been catalogued, but few functional or genetic correlations have emerged. Tomato (Solanum lycopersicum) is part of a complex of closely related wild species endemic to the northern Andes and the Galapagos Islands (Solanum Sect. Lycopersicon). Although sharing an ancestor <7 million years ago, these species are found in diverse environments and are subject to unique selective pressures. Furthermore, they are genetically tractable, since they can be crossed with S. lycopersicum, which has a sequenced genome. With the aim of evaluating the relationships between evolution, structure and function of the cuticle, we characterized the morphological and chemical diversity of fruit cuticles of seven species from Solanum Sect. Lycopersicon. Striking differences in cuticular architecture and quantities of cutin and waxes were observed, with the wax coverage of wild species exceeding that of S. lycopersicum by up to seven fold. Wax composition varied in the occurrence of wax esters and triterpenoid isomers. Using a Solanum habrochaites introgression line population, we mapped triterpenoid differences to a genomic region that includes two S. lycopersicum triterpene synthases. Based on known metabolic pathways for acyl wax compounds, hypotheses are discussed to explain the appearance of wax esters with atypical chain lengths. These results establish a model system for understanding the ecological and evolutionary functional genomics of plant cuticles.",
"Improving crop productivity and quality while promoting sustainable agriculture have become major goals in plant breeding. The cuticle is a natural film covering the aerial organs of plants and consists of lipid polyesters covered and embedded with wax. The cuticle protects plants against water loss and pathogens and affects traits with strong impacts on crop quality such as, for horticultural crops, fruit brightness, cracking, russeting, netting, and shelf life. Here we provide an overview of the most important cuticle-associated traits that can be targeted for crop improvement. To date, most studies on cuticle-associated traits aimed at crop breeding have been done on fleshy fruits. Less information is available for staple crops such as rice, wheat or maize. Here we present new insights into cuticle formation and properties resulting from the study of genetic resources available for the various crop species. Our review also covers the current strategies and tools aimed at exploiting available natural and artificially induced genetic diversity and the technologies used to transfer the beneficial alleles affecting cuticle-associated traits to commercial varieties.",
"The cuticle covering plants' aerial surfaces is a unique structure that plays a key role in organ development and protection against diverse stress conditions. A detailed analysis of the tomato colorless-peel y mutant was carried out in the framework of studying the outer surface of reproductive organs. The y mutant peel lacks the yellow flavonoid pigment naringenin chalcone, which has been suggested to influence the characteristics and function of the cuticular layer. Large-scale metabolic and transcript profiling revealed broad effects on both primary and secondary metabolism, related mostly to the biosynthesis of phenylpropanoids, particularly flavonoids. These were not restricted to the fruit or to a specific stage of its development and indicated that the y mutant phenotype is due to a mutation in a regulatory gene. Indeed, expression analyses specified three R2R3-MYB-type transcription factors that were significantly down-regulated in the y mutant fruit peel. One of these, SlMYB12, was mapped to the genomic region on tomato chromosome 1 previously shown to harbor the y mutation. Identification of an additional mutant allele that co-segregates with the colorless-peel trait, specific down-regulation of SlMYB12 and rescue of the y phenotype by overexpression of SlMYB12 on the mutant background, confirmed that a lesion in this regulator underlies the y phenotype. Hence, this work provides novel insight to the study of fleshy fruit cuticular structure and paves the way for the elucidation of the regulatory network that controls flavonoid accumulation in tomato fruit cuticle.",
"Genetics of traits related to fruit cuticle deposition and composition was studied in two red-fruited tomato species. Two mapping populations derived from the cross between the cultivated tomato (Solanum lycopersicum L.) and its closest relative wild species Solanum pimpinellifolium L. were employed to conduct a QTL analysis. A combination of fruit cuticle deposition, components and anatomical traits were investigated and the individual effect of each QTL evaluated. A total of 70 QTLs were identified, indicating that all the cuticle traits analyzed have a complex polygenic nature. A combination of additive and epistatic interactions was observed for all the traits, with positive contribution of both parental lines to most of them. Colocalization of QTLs for various traits uncovered novel genomic regions producing extensive changes in the cuticle. Cuticle density emerges as an important trait since it can modulate cuticle thickness and invagination thus providing a strategy for sustaining mechanical strength without compromising palatability. Two genomic regions, located in chromosomes 1 and 12, are responsible for the negative interaction between cuticle waxes and phenolics identified in tomato fruit. Several candidate genes, including transcription factors and structural genes, are postulated and their expression analyzed throughout development.",
"The plant cuticle is an extracellular hydrophobic layer that covers the aerial epidermis of all land plants, providing protection against desiccation and external environmental stresses. The past decade has seen considerable progress in assembling models for the biosynthesis of its two major components, the polymer cutin and cuticular waxes. Most recently, two breakthroughs in the long-sought molecular bases of alkane formation and polyester synthesis have allowed construction of nearly complete biosynthetic pathways for both waxes and cutin. Concurrently, a complex regulatory network controlling the synthesis of the cuticle is emerging. It has also become clear that the physiological role of the cuticle extends well beyond its primary function as a transpiration barrier, playing important roles in processes ranging from development to interaction with microbes. Here, we review recent progress in the biochemistry and molecular biology of cuticle synthesis and function and highlight some of the major questions that will drive future research in this field.",
"Mutation of the ECERIFERUM9 (CER9) gene in Arabidopsis (Arabidopsis thaliana) causes elevated amounts of 18-carbon-length cutin monomers and a dramatic shift in the cuticular wax profile (especially on leaves) toward the very-long-chain free fatty acids tetracosanoic acid (C₂₄) and hexacosanoic acid (C₂₆). Relative to the wild type, cer9 mutants exhibit elevated cuticle membrane thickness over epidermal cells and cuticular ledges with increased occlusion of the stomatal pore. The cuticular phenotypes of cer9 are associated with delayed onset of wilting in plants experiencing water deficit, lower transpiration rates, and improved water use efficiency measured as carbon isotope discrimination. The CER9 protein thus encodes a novel determinant of plant drought tolerance-associated traits, one whose deficiency elevates cutin synthesis, redistributes wax composition, and suppresses transpiration. Map-based cloning identified CER9, and sequence analysis predicted that it encodes an E3 ubiquitin ligase homologous to yeast Doa10 (previously shown to target endoplasmic reticulum proteins for proteasomal degradation). To further elucidate CER9 function, the impact of CER9 deficiency on interactions with other genes was examined using double mutant and transcriptome analyses. For both wax and cutin, cer9 showed mostly additive effects with cer6, long-chain acyl-CoA synthetase1 (lacs1), and lacs2 and revealed its role in early steps of both wax and cutin synthetic pathways. Transcriptome analysis revealed that the cer9 mutation affected diverse cellular processes, with primary impact on genes associated with diverse stress responses. The discovery of CER9 lays new groundwork for developing novel cuticle-based strategies for improving the drought tolerance and water use efficiency of crop plants."
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Serum levels of pro- and anti-inflammatory bioactive lipids in severe acute respiratory syndrome coronavirus 2 patients
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Specialized proresolution molecules (SPMs) halt the transition to chronic pathogenic inflammation. We aimed to quantify serum levels of pro- and anti-inflammatory bioactive lipids in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) patients, and to identify potential relationships with innate responses and clinical outcome.
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"An excessive, non-resolving inflammatory response underlies severe COVID-19 that may have fatal outcomes. Therefore, the investigation of endogenous pathways leading to resolution of inflammation is of interest to uncover strategies for mitigating inflammation in people with SARS-CoV-2 infection. This becomes particularly urgent in individuals with preexisting pathologies characterized by chronic respiratory inflammation and prone to bacterial infection, such as cystic fibrosis (CF). Here, we analyzed the immune responses to SARS-CoV-2 virion spike 1 glycoprotein (S1) of macrophages (MΦ) from volunteers with and without CF and tested the efficacy of resolvins (Rv) D1 and D2 in regulating the inflammatory and antimicrobial functions of MΦ exposed to S1. S1 significantly increased chemokine release, including interleukin (IL)-8, in CF and non-CF MΦ, while it enhanced IL-6 and tumor necrosis factor (TNF)-α in non-CF MΦ, but not in CF cells. S1 also triggered the biosynthesis of RvD1 and modulated microRNAs miR-16, miR-29a, and miR-103, known to control the inflammatory responses. RvD1 and RvD2 treatment abated S1-induced inflammatory responses in CF and non-CF MΦ, significantly reducing the release of select chemokines and cytokines including IL-8 and TNF-α. RvD1 and RvD2 both restored the expression of miR-16 and miR-29a, while selectively increasing miR-223 and miR-125a, which are involved in NF-κB activation and MΦ inflammatory polarization. During Pseudomonas aeruginosa infection, S1 stimulated the MΦ phagocytic activity that was further enhanced by RvD1 and RvD2. These results provide a map of molecular responses to SARS-CoV-2 in MΦ, key determinants of COVID-19-related inflammation, unveiling some peculiarity in the response of cells from individuals with CF. They also demonstrate beneficial, regulatory actions of RvD1 and RvD2 on SARS-CoV-2-induced inflammation.",
"Inflammation plays a critical role in cystic fibrosis (CF) lung pathology and disease progression making it an active area of research and important therapeutic target. In this review, we explore the most recent research on the major contributors to the exuberant inflammatory response seen in CF as well as potential therapeutics to combat this response. Absence of functional cystic fibrosis transmembrane conductance regulator (CFTR) alters anion transport across CF airway epithelial cells and ultimately results in dehydration of the airway surface liquid. The dehydrated airway surface liquid in combination with abnormal mucin secretion contributes to airway obstruction and subsequent infection that may serve as a trigger point for inflammation. There is also evidence to suggest that airway inflammation may be excessive and sustained relative to the infectious stimuli. Studies have shown dysregulation of both pro-inflammatory mediators such as IL-17 and pro-resolution mediators including metabolites of the eicosanoid pathway. Recently, CFTR potentiators and correctors have garnered much attention in the CF community. Although these modulators address the underlying defect in CF, their impact on downstream consequences such as inflammation are not known. Here, we review pre-clinical and clinical data on the impact of CFTR modulators on inflammation. In addition, we examine other cell types including neutrophils, macrophages, and T-lymphocytes that express CFTR and contribute to the CF inflammatory response. Finally, we address challenges in developing anti-inflammatory therapies and highlight some of the most promising anti-inflammatory drugs under development for CF.",
"Severe coronavirus disease (COVID-19) is characterized by pulmonary hyper-inflammation and potentially life-threatening \"cytokine storms\". Controlling the local and systemic inflammatory response in COVID-19 may be as important as anti-viral therapies. Endogenous lipid autacoid mediators, referred to as eicosanoids, play a critical role in the induction of inflammation and pro-inflammatory cytokine production. SARS-CoV-2 may trigger a cell death (\"debris\")-induced \"eicosanoid storm\", including prostaglandins and leukotrienes, which in turn initiates a robust inflammatory response. A paradigm shift is emerging in our understanding of the resolution of inflammation as an active biochemical process with the discovery of novel endogenous specialized pro-resolving lipid autacoid mediators (SPMs), such as resolvins. Resolvins and other SPMs stimulate macrophage-mediated clearance of debris and counter pro-inflammatory cytokine production, a process called inflammation resolution. SPMs and their lipid precursors exhibit anti-viral activity at nanogram doses in the setting of influenza without being immunosuppressive. SPMs also promote anti-viral B cell antibodies and lymphocyte activity, highlighting their potential use in the treatment of COVID-19. Soluble epoxide hydrolase (sEH) inhibitors stabilize arachidonic acid-derived epoxyeicosatrienoic acids (EETs), which also stimulate inflammation resolution by promoting the production of pro-resolution mediators, activating anti-inflammatory processes, and preventing the cytokine storm. Both resolvins and EETs also attenuate pathological thrombosis and promote clot removal, which is emerging as a key pathology of COVID-19 infection. Thus, both SPMs and sEH inhibitors may promote the resolution of inflammation in COVID-19, thereby reducing acute respiratory distress syndrome (ARDS) and other life-threatening complications associated with robust viral-induced inflammation. While most COVID-19 clinical trials focus on \"anti-viral\" and \"anti-inflammatory\" strategies, stimulating inflammation resolution is a novel host-centric therapeutic avenue. Importantly, SPMs and sEH inhibitors are currently in clinical trials for other inflammatory diseases and could be rapidly translated for the management of COVID-19 via debris clearance and inflammatory cytokine suppression. Here, we discuss using pro-resolution mediators as a potential complement to current anti-viral strategies for COVID-19.",
"Resolvins (Rvs), endogenous lipid mediators, play a key role in the resolution of inflammation. Sickle cell disease (SCD), a genetic disorder of hemoglobin, is characterized by inflammatory and vaso-occlusive pathologies. We document altered proresolving events following hypoxia/reperfusion in humanized SCD mice. We demonstrate novel protective actions of 17R-resolvin D1 (17R-RvD1; 7S, 8R, 17R-trihydroxy-4Z, 9E, 11E, 13Z, 15E, 19Z-docosahexaenoic acid) in reducing ex vivo human SCD blood leukocyte recruitment by microvascular endothelial cells and in vivo neutrophil adhesion and transmigration. In SCD mice exposed to hypoxia/reoxygenation, oral administration of 17R -RvD1 reduces systemic/local inflammation and vascular dysfunction in lung and kidney. The mechanism of action of 17R-RvD1 involves (1) enhancement of SCD erythrocytes and polymorphonuclear leukocyte efferocytosis, (2) blunting of NF-κB activation, and (3) a reduction in inflammatory cytokines, vascular activation markers, and E-selectin expression. Thus, 17R-RvD1 might represent a new therapeutic strategy for the inflammatory vasculopathy of SCD.",
"Resolvin D1 (RvD1; 7S,8R,17S-trihydroxy-4Z,9E,11E,13Z,15E,19Z-docosahexaenoic acid) is an endogenous immunoresolvent that regulates acute inflammation and orchestrates resolution. Here, we investigated anti-inflammatory and proresolving actions of RvD1 after oral administration. RvD1 rapidly accumulated in the mouse plasma after oral delivery and dose-dependently (1-100 ng/mouse) reduced leukocyte infiltration in zymosan A-induced acute peritonitis. Using mathematical resolution indices, RvD1 reduced Ψmax by ∼50%, shortened the resolution interval by 3 h, and significantly reduced total leukocyte (by ∼30-45%) and polymorphonuclear neutrophil (by ∼40-55%) accumulation when administered at the peak of peritonitis. RvD1 also improved course and outcome of severe peritonitis, shifting it toward resolution. In peritoneal macrophages (MΦs) from the resolution phase of peritonitis, RvD1 down-regulated (by 2- to 3-fold) select genes that control gene transcription, namely coactivator-associated arginine methyltransferase 1 (CARM1), and downstream genes, such as colony-stimulating factor 3, intercellular adhesion molecule 1, and monocyte inflammatory protein 2, which promote neutrophil infiltration and reduce MΦ phagocytosis. Congruently, CARM1 knockdown in human and murine MΦs induced a proresolving phenotype, recapitulating in vivo actions of RvD1. These results establish novel properties of RvD1 and demonstrate that RvD1 modifies the transcription control machinery in MΦs, as part of its mechanisms of action during the resolution of acute inflammation.-Recchiuti, A., Codagnone, M., Pierdomenico, A. M., Rossi, C., Mari, V. C., Cianci, E., Simiele, F., Gatta, V., Romano, M. Immunoresolving actions of oral resolvin D1 include selective regulation of the transcription machinery in resolution-phase mouse macrophages.",
"Coronavirus disease 2019 (COVID-19), caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), has currently led to a global pandemic with millions of confirmed and increasing cases around the world. The novel SARS-CoV-2 not only affects the lungs causing severe acute respiratory dysfunction but also leads to significant dysfunction in multiple organs and physiological systems including the cardiovascular system. A plethora of studies have shown the viral infection triggers an exaggerated immune response, hypercoagulation and oxidative stress, which contribute significantly to poor cardiovascular outcomes observed in COVID-19 patients. To date, there are no approved vaccines or therapies for COVID-19. Accordingly, cardiovascular protective and supportive therapies are urgent and necessary to the overall prognosis of COVID-19 patients. Accumulating literature has demonstrated the beneficial effects of n-3 polyunsaturated fatty acids (n-3 PUFA) toward the cardiovascular system, which include ameliorating uncontrolled inflammatory reactions, reduced oxidative stress and mitigating coagulopathy. Moreover, it has been demonstrated the n-3 PUFAs, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), are precursors to a group of potent bioactive lipid mediators, generated endogenously, which mediate many of the beneficial effects attributed to their parent compounds. Considering the favorable safety profile for n-3 PUFAs and their metabolites, it is reasonable to consider n-3 PUFAs as potential adjuvant therapies for the clinical management of COVID-19 patients. In this article, we provide an overview of the pathogenesis of cardiovascular complications secondary to COVID-19 and focus on the mechanisms that may contribute to the likely benefits of n-3 PUFAs and their metabolites.",
"MicroRNAs are key regulators of many biological processes, including cell differentiation. Here we show that during human monocyte-macrophage differentiation, expression of the microRNAs miR-223, miR-15a and miR-16 decreased considerably, which led to higher expression of the serine-threonine kinase IKKalpha in macrophages. In macrophages, higher IKKalpha expression in conjunction with stabilization of the kinase NIK induced larger amounts of p52. Because of low expression of the transcription factor RelB in untreated macrophages, high p52 expression repressed basal transcription of both canonical and noncanonical NF-kappaB target genes. However, proinflammatory stimuli in macrophages resulted in greater induction of noncanonical NF-kappaB target genes. Thus, a decrease in certain microRNAs probably prevents macrophage hyperactivation yet primes the macrophage for certain responses to proinflammatory stimuli."
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Effect of high-fat diet consumption and exercise intervention on the level of tumor necrosis factor alpha in the spleen of rats.
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High-fat diet (HFD) consumption can trigger chronic inflammation in some tissues. However, it remains unclear if HFD induces chronic inflammation in the spleen. This investigation aims to address the effect of HFD consumption and exercise intervention on the level of tumor necrosis factor alpha (TNF-α) in the spleen. Rats were subjected to HFD feeding and/or moderate-intensity treadmill running. The TNF-α levels in plasma and spleen were detected by ELISA. The mass and total cell numbers of the spleen were measured. In addition, the expression of TNF-α and its relevant gene mRNAs in macrophages from the spleen were analyzed by qRT-PCR. We found that HFD consumption did not significantly affect the mass and total cell numbers of the spleen. However, HFD consumption significantly increased splenic TNF-α level, the expression of TNF-α, toll-like receptor 4, and nuclear factor κB p65 mRNAs. In contrast, the expression of nicotinic acetylcholine receptor alpha 7 subunit (α7nAChR) mRNA in macrophages was downregulated. Additionally, exercise abolished the increase in splenic TNF-α level as well as the abnormal expression of TNF-α and related gene mRNAs in macrophages in HFD-fed rats. In conclusion, our results reveal that HFD consumption increases TNF-α level in the spleen, which is along with upregulation of the expression of TLR4 and NF-κB mRNAs as well as downregulation of the expression of α7nAChR mRNA in splenic macrophages in rats. Exercise abolished detrimental effects of HFD on TNF-α level in the spleen and prevented abnormal expression of these genes in the macrophages from rat spleen.
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"High levels of free fatty acids (FFA) have been suggested to be one of the underlying mechanisms for adipose tissue (AT) inflammation and dysfunction in obesity. Human AT produces several adipokines including monocyte chemoattractant protein-1 (MCP-1), which are involved in the pathogenesis of obesity-mediated inflammation. In this study, we investigated the effects of lipopolysaccharide (LPS) and a panel of dietary FFA on MCP-1 gene and protein expression in adipocytes and macrophages. Furthermore, we investigated whether the effect of LPS and FFA were mediated through the toll-like receptor 4 (TLR4). 3T3-L1 adipocytes and THP-1 macrophages were incubated for 24 h with the following FFA: monounsaturated fatty acid (oleic acid), saturated fatty acid (palmitic acid) and trans fatty acid (elaidic acid; 500 μM) with and without LPS (2 ng ml(-1)), and MCP-1 and TLR4 mRNA expression and MCP-1 protein secretion was determined. The results showed that LPS significantly increased MCP-1 and TLR4 expression and MCP-1 secretion in 3T3-L1 adipocytes, and that the MCP-1 expression was blocked by a TLR4 inhibitor (CLI095). The effects of the various FFA on MCP-1 mRNA expression and protein secretion in the adipocytes showed no significant changes either alone or in combination with LPS. In macrophages, palmitic acid increased MCP-1 mRNA expression by 1.8-fold (P<0.05), but oleic acid and elaidic acid had no effects. In conclusion, in 3T3-L1 adipocyte, the TLR4-agonist, LPS, stimulates the proinflammatory chemokine MCP-1. The different classes of FFA did not induce MCP-1 mRNA expression or protein secretion in the adipocytes, but the saturated FFA, palmitic acid, induced MCP-1 mRNA expression in macrophages, possibly because of the higher expression level of TLR4 in the macrophages than the adipocytes. Our results indicate that FFA may induce AT inflammation through proinflammatory stimulation of macrophages.",
"Oxidative stress basically defines a condition in which prooxidant-antioxidant balance in the cell is disturbed; cellular biomolecules undergo severe oxidative damage, ultimately compromising cells viability. In recent years, a number of studies have shown that oxidative stress could cause cellular apoptosis via both the mitochondria-dependent and mitochondria-independent pathways. Since these pathways are directly related to the survival or death of various cell types in normal as well as pathophysiological situations, a clear picture of these pathways for various active molecules in their biological functions would help designing novel therapeutic strategy. This review highlights the basic mechanisms of ROS production and their sites of formation; detail mechanism of both mitochondria-dependent and mitochondria-independent pathways of apoptosis as well as their regulation by ROS. Emphasis has been given on the redox-sensitive ASK1 signalosome and its downstream JNK pathway. This review also describes the involvement of oxidative stress under various environmental toxin- and drug-induced organ pathophysiology and diabetes-mediated apoptosis. We believe that this review would provide useful information about the most recent progress in understanding the mechanism of oxidative stress-mediated regulation of apoptotic pathways. It will also help to figure out the complex cross-talks between these pathways and their modulations by oxidative stress. The literature will also shed a light on the blind alleys of this field to be explored. Finally, readers would know about the ROS-regulated and apoptosis-mediated organ pathophysiology which might help to find their probable remedies in future.",
"Oxidative stress and inflammation are important pathogenic mediators in diabetes-related organ damage. Accumulating evidence suggests that immunodeficiency in diabetes is associated with diabetes-induced spleen damage. Sleeve gastrectomy (SG) has been proved to improve diabetes and its multiple associated complications. However, the ameliorative role of SG against spleen damage in diabetes has not been investigated. Animal model of diabetic obese rats induced by high-fat diet (HFD) combined with streptozotocin (STZ) was treated with sham operation, caloric restriction, and SG. Metabolic parameters were measured, and the morphological and histopathological changes, status of oxidative stress, and levels of inflammatory factors were evaluated. SG reduced body weight and improved glucose tolerance and insulin sensitivity in diabetic obese rats. SG significantly reversed splenic atrophy and alleviated abnormalities of white and red pulp. Additionally, SG also reversed the increased splenocyte apoptosis (P < 0.001). Meanwhile, indicators of oxidative stress including reactive oxygen species (ROS), nitric oxide (NO), malondialdehyde (MDA), and protein carbonylation were reduced, and the activity and expression of antioxidant enzymes including SOD and CAT were improved after SG. The mRNA expression of inflammatory factors in SG groups such as TNF-α (P < 0.001), IL-6 (P < 0.001), MCP-1 (P < 0.01), and ICAM-1 (P < 0.001) was also significantly reduced. SG ameliorates diabetes-related splenic injury by restoring the balance between oxidative stress process and antioxidant defense systems as well as reducing inflammation in the spleen. These findings indicate that SG is an appropriate therapeutic strategy for diabetes-related spleen damage.",
"The role of inflammation in the pathogenesis of type 2 diabetes and associated complications is now well established. Several conditions that are driven by inflammatory processes are also associated with diabetes, including rheumatoid arthritis, gout, psoriasis and Crohn's disease, and various anti-inflammatory drugs have been approved or are in late stages of development for the treatment of these conditions. This review discusses the rationale for the use of some of these anti-inflammatory treatments in patients with diabetes and what we could expect from their use. Future immunomodulatory treatments may not target a specific disease, but could instead act on a dysfunctional pathway that causes several conditions associated with the metabolic syndrome.",
"Diabetic retinopathy remains the major cause of blindness among working age adults. Although a number of metabolic abnormalities have been associated with its development, due to complex nature of this multi-factorial disease, a link between any specific abnormality and diabetic retinopathy remains largely speculative. Diabetes increases oxidative stress in the retina and its capillary cells, and overwhelming evidence suggests a bidirectional relationship between oxidative stress and other major metabolic abnormalities implicated in the development of diabetic retinopathy. Due to increased production of cytosolic reactive oxygen species, mitochondrial membranes are damaged and their membrane potentials are impaired, and complex III of the electron transport system is compromised. Suboptimal enzymatic and nonenzymatic antioxidant defense system further aids in the accumulation of free radicals. As the duration of the disease progresses, mitochondrial DNA (mtDNA) is damaged and the DNA repair system is compromised, and due to impaired transcription of mtDNA-encoded proteins, the integrity of the electron transport system is encumbered. Due to decreased mtDNA biogenesis and impaired transcription, superoxide accumulation is further increased, and the vicious cycle of free radicals continues to self-propagate. Diabetic milieu also alters enzymes responsible for DNA and histone modifications, and various genes important for mitochondrial homeostasis, including mitochondrial biosynthesis, damage and antioxidant defense, undergo epigenetic modifications. Although antioxidant administration in animal models has yielded encouraging results in preventing diabetic retinopathy, controlled longitudinal human studies remain to be conducted. Furthermore, the role of epigenetic in mitochondrial homeostasis suggests that regulation of such modifications also has potential to inhibit/retard the development of diabetic retinopathy.",
"Obesity and metabolic syndrome are disorders that correlate with the activation of pro-inflammatory pathways and cytokine production, to which Toll like receptors (TLR) contribute. Exercise may act as an anti-inflammatory modulator, but there is no consensus about the role of the TLR in this tuning. The present styudy aims to systematically review the current evidence on exercise-induced TLR regulation in animals and humans suffering from obesity and metabolic syndrome. Pubmed and Scopus databases were searched for publications from 1990 to September 2015. Search terms included: \"Toll like Receptor\", \"TLR\", \"exercise\", \"obesity\", \"diabetes\", and \"metabolic syndrome\". Elegibility criteria comprised: randomized control trials, cross-sectional and cohort studies; human or animal models with metabolic syndrome; any type of exercise; TLR expression measurement in any tissue by a clearly reported technique. The quality of selected studies was assessed using a modified version of the Downs and Black Quality Assessment Checklist. Data of study design; population; exercise type, timing and training elements; measurement technique, tissue analyzed and main outcome were extracted and categorized to facilitate data synthesis. 17 studies were included, of which 11 publications obtained a high, 5 a moderate and 1 a low score for quality assessment. A total of 8 human studies were analyzed: 6 studies used endurance continuous or interval training protocols, 1 study resistance training and the remaining study was performed following a marathon race. Blood cells were analyzed in seven studies, of which four studies sampled peripheral blood mononuclear cells (PBMC), three analyzed whole blood and one study sampled skeletal muscle. Nine animal studies were included: 8 used endurance training and 1 acute aerobic exercise. A variety of tissues samples were explored such as PBMC, skeletal muscle, adipose, vascular and nervous tissue. Globally, the animal studies showed a marked tendency towards a down-regulation of TLR2 and 4 expression accompagnied with, a reduced activation of nuclear factorkappaB (NF-κB) signaling and cytokine production, and an improvement in insulin sensitivity and body composition. While animal studies showed a marked tendency towards TLR2 and 4 down-regulation after chronic endurance exercise, the current evidence in human is not sufficiently robust to conclude any role of TLR in the anti-inflammatory properties of exercise.",
"A decrease in skeletal muscle mass and motor function occurs in diabetic patients. In type 1 diabetic patients, in particular, fast-type fiber-dominated muscle atrophy occurs due to increased oxidative stress and inflammation. Juzentaihoto is a herbal medicine that has been found to be effective in reducing oxidative stress. In this study, juzentaihoto hot water extract (JTT) was administered prophylactically to mice with diabetic oxidative stress, which was induced by an injection of streptozotocin, and the effects on skeletal muscle mass, motor function, and antioxidant activity were evaluated. In mice that were administered JTT, skeletal muscle atrophy and loss of motor function were suppressed. Additionally, the administration of JTT increased the mRNA expression level of Sirt1 and the activity of superoxide dismutase in the gastrocnemius. In addition to skeletal muscle atrophy, atrophy of the liver, spleen and thymus gland, and kidney hypertrophy were also suppressed. Furthermore, in order to evaluate the antioxidant activity of 10 constituent crude drugs that comprise juzentaihoto, Sirt1 transcriptional activity in C2C12 cells was evaluated. The Sirt1 transcriptional activity was increased by Cinnamomi Cortex, Astragali Radix, and Glycyrrhizae Radix extracts. These three constituent crude drugs play an important function in the antioxidant action of juzentaihoto, suggesting that juzentaihoto can prevent muscle atrophy by decreasing oxidative stress."
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Optimization of fluoxetine laden orodispersible film in enhancing dosage forms options for pediatric patients with selective mutism and obsessive-compulsive disorder
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The objective of this research was to fabricate, characterize, and optimize fluoxetine laden orodispersible film (ODF), in enhancing dosage forms options for the pediatric population suffering from incapacitating psychotic disorders of selective mutism and obsessive-compulsive disorder, which will be ultimately beneficial in enhancing compliance factor and the quality of pharmacotherapy.
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"Little is known about the neural correlates of tics and associated urges. In the present study, we aimed to explore the neural basis of tics in patients with Tourette syndrome by using event-related functional MRI (fMRI). Ten patients (6 women, 4 men; age: mean +/- SD = 31 +/- 11.2) were studied while spontaneously exhibiting a variety of motor and vocal tics. On the basis of synchronized video/audio recordings, fMRI activities were analysed 2 s before and at tic onset irrespective of the clinical phenomenology. We identified a brain network of paralimbic areas such as anterior cingulate and insular cortex, supplementary motor area (SMA) and parietal operculum (PO) predominantly activated before tic onset (P < 0.05, corrected for multiple comparisons). In contrast, at the beginning of tic action, significant fMRI activities were found in sensorimotor areas including superior parietal lobule bilaterally and cerebellum. The results of this study indicate that paralimbic and sensory association areas are critically implicated in tic generation, similar to movements triggered internally by unpleasant sensations, as has been shown for pain or itching.",
"To comprehend psychosomatic processes, it will be necessary to understand the brain's influences on bodily functions and also the body's afferent sensory input to the central nervous system, including the effects of this input on behavior and cognitive functions, especially emotion. The objective of this Presidential Address is to review what is known circa the year 2000 of the processes and mechanisms of visceral sensory psychobiology, often called interoception. Over 1000 publications that have appeared since the 19th century were reviewed to prepare this review, including a group that are specifically cited here. Factors and data were reviewed that were identified as germane to understanding interoception. These included definitional issues, historical roots, the neural basis, studies and results in the cardiovascular-respiratory and alimentary-gastrointestinal systems, studies of emotion, and studies in people with mental disorders. Drug and hormone effects, pain, proprioception, and phantom limb or organ factors, and the role of awareness were briefly described. Methodological issues, methods of study including functional imaging, and possible future directions for study were identified. Understanding the physical basis of psychosomatic processes, including the so-called mind-body problem, will require a detailed understanding the psychobiology of interoception.",
"Recently, fast dissolving films are gaining interest as an alternative of fast dissolving tablets. The films are designed to dissolve upon contact with a wet surface, such as the tongue, within a few seconds, meaning the consumer can take the product without need for additional liquid. This convenience provides both a marketing advantage and increased patient compliance. As the drug is directly absorbed into systemic circulation, degradation in gastrointestinal tract and first pass effect can be avoided. These points make this formulation most popular and acceptable among pediatric and geriatric patients and patients with fear of choking. Over-the-counter films for pain management and motion sickness are commercialized in the US markets. Many companies are utilizing transdermal drug delivery technology to develop thin film formats. In the present review, recent advancements regarding fast dissolving buccal film formulation and their evaluation parameters are compiled.",
"In this issue, Terasawa and colleagues used functional neuroimaging to test for common neural substrates supporting conscious appraisal of subjective bodily and emotional states and explored how the relationship might account for personality and experience of anxiety symptoms. Their study highlights a role for the same region of anterior insula cortex in appraisal of emotions and bodily physiology. The reactivity of this region also mediated the relationship between 'bodily sensibility' and social fear, translating a cognitive representation of subjective physical state into an individual personality trait that influences social interaction. The task used by Terasawa and colleagues taps into conscious aspects to the expression of this dynamic. These findings add to increasing evidence for the role of anterior insula as the interface between physiologically driven internal motivational states, emotional awareness and interpersonal behaviour.",
"The ultimate goal of any drug delivery system is the successful delivery of the drug to the body; however, patient compliance must not be overlooked. Fast dissolving drug delivery systems, such as, Mouth Dissolving Films (MDF), offer a convenient way of dosing medications, not only to special population groups with swallowing difficulties such as children and the elderly, but also to the general population. MDF are the novel dosage forms that disintegrate and dissolve within the oral cavity. Intra-oral absorption permits rapid onset of action and helps by-pass first-pass effects, thereby reducing the unit dose required to produce desired therapeutic effect. The present review provides an overview of various polymers that can be employed in the manufacture of MDF and highlights the effect of polymers and plasticizers on various physico-mechanical properties of MDF. It further gives a brief account of formulation of MDF and problems faced during its manufacture.",
"The orbitofrontal cortex (OFC)-striatal circuit, which is important for motivational behavior, is assumed to be involved in the pathophysiology of obsessive-compulsive disorder (OCD) according to current neurobiological models of this disorder. However, the engagement of this neural loop in OCD has not been tested directly in a cognitive activation imaging paradigm so far. To determine whether the OFC and the ventral striatum show abnormal neural activity in OCD during cognitive challenge. A reversal learning task was employed in 20 patients with OCD who were not receiving medication and 27 healthy controls during an event-related functional magnetic resonance imaging experiment using a scanning sequence sensitive to OFC signal. This design allowed investigation of the neural correlates of reward and punishment receipt as well as of \"affective switching,\" ie, altering behavior on reversing reinforcement contingencies. Patients with OCD exhibited an impaired task end result reflected by a reduced number of correct responses relative to control subjects but showed adequate behavior on receipt of punishment and with regard to affective switching. On reward outcome, patients showed decreased responsiveness in right medial and lateral OFC as well as in the right caudate nucleus (border zone ventral striatum) when compared with controls. During affective switching, patients recruited the left posterior OFC, bilateral insular cortex, bilateral dorsolateral, and bilateral anterior prefrontal cortex to a lesser extent than control subjects. No areas were found for which patients exhibited increased activity relative to controls, and no differential activations were observed for punishment in a direct group comparison. These data show behavioral impairments accompanied by aberrant OFC-striatal and dorsal prefrontal activity in OCD on a reversal learning task that addresses this circuit's function. These findings not only confirm previous reports of dorsal prefrontal dysfunction in OCD but also provide evidence for the involvement of the OFC-striatal loop in the pathophysiology of OCD.",
"This study examined the utility of fluoxetine in the treatment of 5 children, aged 5 to 14 years, diagnosed with selective mutism who also demonstrated symptoms of social anxiety. A nonconcurrent, randomized, multiple-baseline, single-case design with a single-blind placebo-controlled procedure was used. Parents and the study psychiatrist completed multiple methods of assessment including Direct Behavior Ratings and questionnaires. Treatment outcomes were evaluated by calculating effect sizes for each participant as an individual and for the participants as a group. Information regarding adverse effects with an emphasis on behavioral disinhibition and ratings of parental acceptance of the intervention was gathered. All 5 children experienced improvement in social anxiety, responsive speech, and spontaneous speech with medium to large effect sizes; however, children still met criteria for selective mutism at the end of the study. Adverse events were minimal, with only 2 children experiencing brief occurrences of minor behavioral disinhibition. Parents found the treatment highly acceptable.",
"As humans, we perceive feelings from our bodies that relate our state of well-being, our energy and stress levels, our mood and disposition. How do we have these feelings? What neural processes do they represent? Recent functional anatomical work has detailed an afferent neural system in primates and in humans that represents all aspects of the physiological condition of the physical body. This system constitutes a representation of 'the material me', and might provide a foundation for subjective feelings, emotion and self-awareness.",
"Previous neuroimaging research has contributed insights regarding the neural substrates of specific psychiatric disorders. The purpose of this study was to determine the shared mediating neuroanatomy of anxiety symptoms across three different anxiety disorders. Data were pooled from 23 right-handed adult outpatients meeting criteria for obsessive-compulsive disorder, simple phobia, or posttraumatic stress disorder. Relative regional cerebral blood flow (rCBF) was measured using positron emission tomography in the context of symptom provocation paradigms. Symptom severity was measured via self-reports. The analysis of pooled imaging data indicated activation in right inferior frontal cortex, right posterior medial orbitofrontal cortex, bilateral insular cortex, bilateral lenticulate nuclei, and bilateral brain stem foci during the symptomatic versus control conditions. A positive correlation was found between rCBF at one brain stem locus and subjective anxiety scores (r = .744, p < .001). These findings suggest that elements of the paralimbic belt together with right inferior frontal cortex and subcortical nuclei mediate symptoms across different anxiety disorders. In addition, activation at one brain stem locus appears to be associated with the subjective severity of anxiety. Further studies are warranted to determine whether these same brain systems mediate normal anxiety states as well.",
"Over the past century, much research has investigated how the brain processes signals from the body (interoception) and how this processing may be disturbed in patients with psychiatric disorders. In this paper, I discuss the literature examining the relationship between interoceptive awareness and emotional and cognitive processes, and review the evidence suggesting that anxiety and obsessive-compulsive disorder (OCD) are characterized by abnormal interoception. A network of cortical and subcortical brain regions centered on the insula has repeatedly been implicated in interoception and is abnormal in anxiety and OCD. The investigation of interoception provides a framework for understanding behavioral and neural mechanisms of anxiety and OCD, although additional research is needed to directly link insula functioning to aberrant interoception in these disorders. Future work targeting interoception may be useful for the development of novel treatments."
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MicroRNA-Mediated Endothelial Progenitor Cell Dysfunction in Diabetic Retinopathy
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Endothelial progenitor cells (EPCs) are involved in the pathogenesis of microvascular dysfunction in diabetic retinopathy (DR). MicroRNAs (miRNAs) serve as crucial regulators in many biological process and human diseases. Herein, to investigate the expression profile and possible role of miRNAs in EPCs, small RNA sequencing was conducted to identify EPC dysfunction-related miRNAs in DR. A total of 72 miRNAs were differentially expressed in EPCs following high glucose stress. Based on Gene Ontology (GO) analysis, the target genes of differentially expressed miRNAs were targeted to "protein binding," "cell differentiation," and "cytoskeleton." Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that cGMP-PKG signaling pathway was tightly associated with miRNA-mediated EPC function. Furthermore, miR-375-3p was verified to be up-regulated in the clinical samples of DR patients. Inhibition of miR-375-3p protected against hyperglycemic stress- or hypoxic stress-induced EPC injury, which increased the viability, proliferation, migration, and tube formation ability of EPCs and retarded the development of apoptosis. Collectively, this study provides a novel insight into the pathogenesis of EPC dysfunction in DR. miR-375-3p is a potential target for the diagnosis or treatment of DR.
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"Endothelial progenitor cells (EPCs) have potential for promoting vascular repair and revascularization of ischemic retina. However, the highly heterogeneous nature of these cells causes confusion when assessing their biological functions. The purpose of this study was to provide a comprehensive comparison between the two main EPC subtypes, early EPCs (eEPCs) and outgrowth endothelial cells (OECs), and to establish the potential of OECs as a novel cell therapy for ischemic retinopathy. Two types of human blood-derived EPCs were isolated and compared using immunophenotyping and multiple in vitro functional assays to assess interaction with retinal capillary endothelial cells and angiogenic activity. OECs were delivered intravitreally in a mouse model of ischemic retinopathy, and flat mounted retinas were examined using confocal microscopy. These data indicate that eEPCs are hematopoietic cells with minimal proliferative capacity that lack tube-forming capacity. By contrast, OECs are committed to an endothelial lineage and have significant proliferative and de novo tubulogenic potential. Furthermore, only OECs are able to closely interact with endothelial cells through adherens and tight junctions and to integrate into retinal vascular networks in vitro. The authors subsequently chose OECs to test a novel cell therapy approach for ischemic retinopathy. Using a murine model of retinal ischemia, they demonstrated that OECs directly incorporate into the resident vasculature, significantly decreasing avascular areas, concomitantly increasing normovascular areas, and preventing pathologic preretinal neovascularization. As a distinct EPC population, OECs have potential as therapeutic cells to vascularize the ischemic retina.",
"Diabetes mellitus is a complex metabolic disorder with aggressive and progressive cardiovascular expression. This article reviews the main percutaneous techniques and their results in the diabetic population, from the early days of balloon angioplasty to the recently advent of drug eluting stents. As the latter devices may eradicate restenosis, plaque progression will remain the cornerstone for interventional cardiologists and the medical community. Attempts to modify life habits, and a more accurate control of the components of the metabolic syndrome should be the main therapeutic objective to slow down the progression and expansion of the disease.",
"To investigate the role of endothelial progenitor cell (EPC) in the pathogenesis of diabetic retinopathy (DR) in streptozotocin (STZ)-induced type 1 diabetes mellitus (T1DM) rats. A total of 160 male Wistar rats (12 weeks old; 250-350 g) were randomly assigned into four groups (n = 40 for each), including control (group 1, no treatment), T1DM1 (group 2, 1 month after 50 mg/kg of STZ, single i.p.), T1DM3 (group 3, 3 months after 50 mg/kg of STZ, single i.p.), T1DM6 (group 4, 6 months after 50 mg/kg of STZ, single i.p.). Enumeration of circulating EPC from peripheral blood was measured by flow cytometry. EPC from bone marrow of rats was cultured in vitro to evaluate its function of proliferation, adhesion, and migration activities. Plasma levels of vascular endothelial growth factor (VEGF) and nitric oxide (NO) were measured by enzyme-linked immunosorbent assay (ELISA). Retinal sections were imaged by light microscopy and a transmission electron microscope (TEM). The numbers of circulating EPC were significantly decreased in diabetic groups compared with the control group. Impaired proliferation, adhesion, and migratory activities of cultured EPC were observed in diabetic groups. There were significantly higher levels of plasma VEGF but lower levels of plasma NO in diabetic groups than those in non-diabetic controls. The significantly reduced thickness and obvious disorganized retinal cell layers were seen in T1DM DR rats. In the diabetic groups, we also found that T1DM rats developed telangiectatic vessels, vacuolar degeneration of ganglion cells, and thickened capillary basement membrane with capillary lumen stenosis in the retina. Significantly raised EPC numbers during DR formation and progression were also found. The reduced numbers and impaired function of circulating EPC may contribute to the pathogenesis of DR in T1DM rats.",
"Diabetic retinopathy, a secondary complication of diabetes mellitus, can lead to irreversible vision loss. Currently, no treatment is approved for early phases of diabetic retinopathy. Modifications of the expression pattern of miRNAs could be involved in the early retinal damage of diabetic subjects. Therefore, we aimed at identification of dysregulated miRNAs-mRNA interactions that might be biomarkers and pharmacological targets for diagnosis and treatment of early diabetic retinopathy. A focused set of miRNAs was predicted through a bioinformatic analysis accessing to Gene Expression Omnibus dataset and enrichment of information approach (GENEMANIA-Cytoscape). Identification of miRNAs-mRNA interactions was carried out with miRNET analysis. Diabetes was induced in C57BL6J mice by streptozotocin and samples analysed at 5 and 10 weeks after diabetes induction. Retinal ultrastructure of diabetic mice was analysed through electron microscopy. We used Real-time PCR, western blot analysis, elisa, and immunohistochemistry to study expression of miRNAs and possible targets of dysregulated miRNAs. We found that miR-20a-5p, miR-20a-3p, miR-20b, miR-106a-5p, miR-27a-5p, miR-27b-3p, miR-206-3p, and miR-381-3p were dysregulated in the retina and serum of diabetic mice. VEGF, brain-derived neurotrophic factor (BDNF), PPAR-α, and cAMP response element-binding protein 1 (CREB1) are targets of dysregulated miRNAs, which then modulated protein expression in diabetic retina. We found structural modifications in retinas from diabetic mice. Serum and retina of diabetic mice express eight dysregulated miRNAs, which modified the expression of VEGF, BDNF, PPAR-α, and CREB1, before vasculopathy in diabetic retinas.",
"Atherosclerosis is the most common cause for cardiovascular diseases and is based on endothelial dysfunction. A growing body of evidence suggests the contribution of bone marrow-derived endothelial progenitor cells, monocytic cells, and mature endothelial cells to vessel formation and endothelial rejuvenation. To this day, various subsets of these endothelial-regenerating cells have been identified according to cellular origin, phenotype, and properties in vivo and in vitro. However, the definition and biology, especially of endothelial progenitor cells, is complex and under heavy debate. In this review, we focus on current definitions of endothelial progenitor cells, highlight the clinical relevance of endothelial-regenerating cells, and provide new insights into cell-cell interactions involved in endothelial cell rejuvenation.",
"Endothelial progenitor cells (EPCs) are decreased in number and function in type 2 diabetes. Mechanisms by which this dysfunction occurs are largely unknown. We tested the hypothesis that a chronic inflammatory environment leads to insulin signaling defects in EPCs and thereby reduces their survival. Modifying EPCs by a knockdown of nuclear factor-κB (NF-κB) can reverse the insulin signaling defects, improve EPC survival, and decrease neointimal hyperplasia in Zucker fatty rats postangioplasty. EPCs from Zucker fatty insulin-resistant rats were cultured and exposed to tumor necrosis factor-α (TNF-α). Insulin signaling defects and apoptosis were measured in the presence and absence of an NF-κB inhibitor, BAY11. Then, EPCs were modified by a knockdown of NF-κB (RelA) and exposed to TNF-α. For in vivo experiments, Zucker fatty rats were given modified EPCs post-carotid angioplasty. Tracking of EPCs was done at various time points, and neointimal hyperplasia was measured 3 weeks later. Insulin signaling as measured by the phosphorylated-to-total AKT ratio was reduced by 56% in EPCs exposed to TNF-α. Apoptosis was increased by 71%. These defects were reversed by pretreatment with an NF-κB inhibitor, BAY11. Modified EPCs exposed to TNF-α showed a lesser reduction (RelA 20%) in insulin-stimulated AKT phosphorylation versus a 55% reduction in unmodified EPCs. Apoptosis was 41% decreased for RelA knockdown EPCs. Noeintimal hyperplasia postangioplasty was significantly less in rats receiving modified EPCs than in controls (intima-to-media ratio 0.58 vs. 1.62). In conclusion, we have shown that insulin signaling and EPC survival is impaired in Zucker fatty insulin resistant rats. For the first time, we have shown that this defect can be significantly ameliorated by a knockdown of NF-κB and that these EPCs given to Zucker fatty rats decrease neointimal hyperplasia post-carotid angioplasty.",
"MicroRNAs (miRNAs) constitute a large family of noncoding RNAs that function as guide molecules in diverse gene silencing pathways. Current efforts are focused on the regulatory function of miRNAs, while little is known about how these unusual genes themselves are regulated. Here we present the first direct evidence that miRNA genes are transcribed by RNA polymerase II (pol II). The primary miRNA transcripts (pri-miRNAs) contain cap structures as well as poly(A) tails, which are the unique properties of class II gene transcripts. The treatment of human cells with alpha-amanitin decreased the level of pri-miRNAs at a concentration that selectively inhibits pol II activity. Furthermore, chromatin immunoprecipitation analyses show that pol II is physically associated with a miRNA promoter. We also describe, for the first time, the detailed structure of a miRNA gene by determining the promoter and the terminator of mir-23a approximately 27a approximately 24-2. These data indicate that pol II is the main, if not the only, RNA polymerase for miRNA gene transcription. Our study offers a basis for understanding the structure and regulation of miRNA genes."
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The potential for people to engage in remote work and occupational social distancing
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Since late 2019, COVID-19 has devastated the global economy, with indirect implications for the environment. As governments' prioritized health and implemented measures such as the closure of non-essential businesses and social distancing, many workers have lost their jobs, been furloughed, or started working from home. Consequently, the world of work has drastically transformed and this period is likely to have major implications for mobility, transportation and the environment. This paper estimates the potential for people to engage in remote work and social distancing using O*NET data and Irish Census data and calculates the potential emission savings, by commuter type from a switch to remote working and occupational social distancing. The results show that while those who commute by car have a relatively high potential for remote work, they are less likely to be able to engage in social distancing in their workplace. While this may be negative for employment prospects in the short run, our analysis indicates that this pattern has the potential for positive environmental implications in the short and long run.
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"To review the effects of school closures on pandemic and seasonal influenza outbreaks. Systematic review. MEDLINE and EMBASE, reference lists of identified articles, hand searches of key journals and additional papers from the authors' collections. Studies were included if they reported on a seasonal or pandemic influenza outbreak coinciding with a planned or unplanned school closure. Of 2579 papers identified through MEDLINE and EMBASE, 65 were eligible for inclusion in the review along with 14 identified from other sources. Influenza incidence frequently declined after school closure. The effect was sometimes reversed when schools reopened, supporting a causal role for school closure in reducing incidence. Any benefits associated with school closure appeared to be greatest among school-aged children. However, as schools often closed late in the outbreak or other interventions were used concurrently, it was sometimes unclear how much school closure contributed to the reductions in incidence. School closures appear to have the potential to reduce influenza transmission, but the heterogeneity in the data available means that the optimum strategy (eg, the ideal length and timing of closure) remains unclear.",
"The COVID-19 pandemic and the measures taken to combat it led to severe constraints for various areas of life, including mobility. To study the effects of this disruptive situation on the mobility behaviour of entire subgroups, and how they shape their mobility in reaction to the special circumstances, can help to better understand, how people react to external changes. Aim of the study presented in this article was to investigate to what extent, how and in what areas mobility behaviour has changed during the outbreak of SARS-CoV-2 in Germany. In addition, a focus was put on the comparison of federal states with and without lockdown in order to investigate a possible contribution of this measure to changes in mobility. We asked respondents via an online survey about their trip purposes and trip frequency, their choice of transport mode and the reasons for choosing it in the context of the COVID-19 crisis. For the analyses presented in this paper, we used the data of 4157survey participants (2512 without lockdown, 1645 with lockdown). The data confirmed a profound impact on the mobility behaviour with a shift away from public transport and increases in car usage, walking and cycling. Comparisons of federal states with and without lockdown revealed only isolated differences. It seems that, even if the lockdown had some minor effects, its role in the observed behavioural changes was minimal.",
"In Switzerland, strict measures as a response to the Covid-19 pandemic were imposed on March 16, 2020, before being gradually relaxed from May 11 onwards. We report the impact of these measures on mobility behaviour based on a GPS tracking panel of 1439 Swiss residents. The participants were also exposed to online questionnaires. The impact of both the lockdown and the relaxation of the measures up until the middle of August 2020 are presented. Reductions of around 60% in the average daily distance were observed, with decreases of over 90% for public transport. Cycling increased in mode share drastically. Behavioural shifts can even be observed in response to the announcement of the measures and relaxation, a week before they came in to place. Long-term implications for policy are discussed, in particular the increased preference for cycling as a result of the pandemic.",
"Although from a societal point of view a modal shift from car to bicycle may have beneficial health effects due to decreased air pollution emissions, decreased greenhouse gas emissions, and increased levels of physical activity, shifts in individual adverse health effects such as higher exposure to air pollution and risk of a traffic accident may prevail. We describe whether the health benefits from the increased physical activity of a modal shift for urban commutes outweigh the health risks. We have summarized the literature for air pollution, traffic accidents, and physical activity using systematic reviews supplemented with recent key studies. We quantified the impact on all-cause mortality when 500,000 people would make a transition from car to bicycle for short trips on a daily basis in the Netherlands. We have expressed mortality impacts in life-years gained or lost, using life table calculations. For individuals who shift from car to bicycle, we estimated that beneficial effects of increased physical activity are substantially larger (3-14 months gained) than the potential mortality effect of increased inhaled air pollution doses (0.8-40 days lost) and the increase in traffic accidents (5-9 days lost). Societal benefits are even larger because of a modest reduction in air pollution and greenhouse gas emissions and traffic accidents. On average, the estimated health benefits of cycling were substantially larger than the risks relative to car driving for individuals shifting their mode of transport.",
"The COVID-19 pandemic poses a great challenge for contemporary public transportation worldwide, resulting from an unprecedented decline in demand and revenue. In this paper, we synthesize the state-of-the-art, up to early June 2020, on key developments regarding public transportation and the COVID-19 pandemic, including the different responses adopted by governments and public transportation agencies around the world, and the research needs pertaining to critical issues that minimize contagion risk in public transportation in the so-called post-lockdown phase. While attempts at adherence to physical distancing (which challenges the very concept of mass public transportation) are looming in several countries, the latest research shows that for closed environments such as public transportation vehicles, the proper use of face masks has significantly reduced the probability of contagion. The economic and social effects of the COVID-19 outbreak in public transportation extend beyond service performance and health risks to financial viability, social equity, and sustainable mobility. There is a risk that if the public transportation sector is perceived as poorly transitioning to post-pandemic conditions, that viewing public transportation as unhealthy will gain ground and might be sustained. To this end, this paper identifies the research needs and outlines a research agenda for the public health implications of alternative strategies and scenarios, specifically measures to reduce crowding in public transportation. The paper provides an overview and an outlook for transit policy makers, planners, and researchers to map the state-of-affairs and research needs related to the impacts of the pandemic crisis on public transportation. Some research needs require urgent attention given what is ultimately at stake in several countries: restoring the ability of public transportation systems to fulfill their societal role."
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In vitro human sensory neuron models in preclinical pain research
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In vitro models fill a vital niche in preclinical pain research, allowing detailed study of molecular pathways, and in the case of humanised systems, providing a translational bridge between in vivo animal models and human patients. Significant advances in cellular technology available to basic pain researchers have occurred in the last decade, including developing protocols to differentiate sensory neuron-like cells from stem cells and greater access to human dorsal root ganglion tissue. In this review, we discuss the use of both models in preclinical pain research: What can a human sensory neuron in a dish tell us that rodent in vivo models cannot? How similar are these models to their endogenous counterparts, and how should we judge them? What limitations do we need to consider? How can we leverage cell models to improve translational success? In vitro human sensory neuron models equip pain researchers with a valuable tool to investigate human nociception. With continual development, consideration for their advantages and limitations, and effective integration with other experimental strategies, they could become a driving force for the pain field's advancement.
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"See Basbaum (doi:10.1093/brain/awx227) for a scientific commentary on this article. Peripheral neuropathic pain arises as a consequence of injury to sensory neurons; the development of ectopic activity in these neurons is thought to be critical for the induction and maintenance of such pain. Local anaesthetics and anti-epileptic drugs can suppress hyperexcitability; however, these drugs are complicated by unwanted effects on motor, central nervous system and cardiac function, and alternative more selective treatments to suppress hyperexcitability are therefore required. Here we show that a glutamate-gated chloride channel modified to be activated by low doses of ivermectin (but not glutamate) is highly effective in silencing sensory neurons and reversing neuropathic pain-related hypersensitivity. Activation of the glutamate-gated chloride channel expressed in either rodent or human induced pluripotent stem cell-derived sensory neurons in vitro potently inhibited their response to both electrical and algogenic stimuli. We have shown that silencing is achieved both at nerve terminals and the soma and is independent of membrane hyperpolarization and instead likely mediated by lowering of the membrane resistance. Using intrathecal adeno-associated virus serotype 9-based delivery, the glutamate-gated chloride channel was successfully targeted to mouse sensory neurons in vivo, resulting in high level and long-lasting expression of the channel selectively in sensory neurons. This enabled reproducible and reversible modulation of thermal and mechanical pain thresholds in vivo; analgesia was observed for 3 days after a single systemic dose of ivermectin. We did not observe any motor or proprioceptive deficits and noted no reduction in cutaneous afferent innervation or upregulation of the injury marker ATF3 following prolonged glutamate-gated chloride channel expression. Established mechanical and cold pain-related hypersensitivity generated by the spared nerve injury model of neuropathic pain was reversed by ivermectin treatment. The efficacy of ivermectin in ameliorating behavioural hypersensitivity was mirrored at the cellular level by a cessation of ectopic activity in sensory neurons. These findings demonstrate the importance of aberrant afferent input in the maintenance of neuropathic pain and the potential for targeted chemogenetic silencing as a new treatment modality in neuropathic pain.",
"TRPA1 is a receptor expressed by sensory neurons, that is activated by low temperature (<17 degrees C) and plant derivatives such as cinnamaldehyde and isoeugenol, to elicit sensations including pain. Using immunohistochemistry, we have, for the first time, localised TRPA1 in human DRG neurons, spinal cord motoneurones and nerve roots, peripheral nerves, intestinal myenteric plexus neurones, and skin basal keratinocytes. TRPA1 co-localised with a subset of hDRG neurons positive for TRPV1, the heat and capsaicin receptor. The number of small/medium TRPA1 positive neurons (< or =50 microm) was increased after hDRG avulsion injury [percentage of cells, median (range): controls 16.5 (7-23); injured 46 (34-55); P<0.005], but the number of large TRPA1 neurons was unchanged [control 19.5 (13-31); injured 21 (11-35)]. Similar TRPA1 changes were observed in cultured hDRG neurons, after exposure to a combination of key neurotrophic factors NGF, GDNF and NT-3 (NTFs) in vitro. We used calcium imaging to examine responses of HEK cells transfected with hTRPA1 cDNA, and of human and rat DRG neurons cultured with or without added NTFs, to cinnamaldehyde (CA) and isoeugenol (IE). Exposure to NTFs in vitro sensitized cultured human sensory neuronal responses to CA; repeated CA exposure produced desensitisation. In rDRG neurons, low (225 microM) CA preincubation enhanced capsaicin responses, while high (450 microM and 2mM) CA caused inhibition which was partially reversed in the presence of 8 bromo cAMP, indicating receptor dephosphorylation. While TRPA1 localisation is more widespread than TRPV1, it represents a promising novel drug target for the treatment of chronic pain and hypersensitivity.",
"Peripheral neuropathy is dose limiting in paclitaxel cancer chemotherapy and can result in both acute pain during treatment and chronic persistent pain in cancer survivors. The hypothesis tested was that paclitaxel produces these adverse effects at least in part by sensitizing transient receptor potential vanilloid subtype 1 (TRPV1) through Toll-like receptor 4 (TLR4) signaling. The data show that paclitaxel-induced behavioral hypersensitivity is prevented and reversed by spinal administration of a TRPV1 antagonist. The number of TRPV1(+) neurons is increased in the dorsal root ganglia (DRG) in paclitaxel-treated rats and is colocalized with TLR4 in rat and human DRG neurons. Cotreatment of rats with lipopolysaccharide from the photosynthetic bacterium Rhodobacter sphaeroides (LPS-RS), a TLR4 inhibitor, prevents the increase in numbers of TRPV1(+) neurons by paclitaxel treatment. Perfusion of paclitaxel or the archetypal TLR4 agonist LPS activated both rat DRG and spinal neurons directly and produced acute sensitization of TRPV1 in both groups of cells via a TLR4-mediated mechanism. Paclitaxel and LPS sensitize TRPV1 in HEK293 cells stably expressing human TLR4 and transiently expressing human TRPV1. These physiological effects also are prevented by LPS-RS. Finally, paclitaxel activates and sensitizes TRPV1 responses directly in dissociated human DRG neurons. In summary, TLR4 was activated by paclitaxel and led to sensitization of TRPV1. This mechanism could contribute to paclitaxel-induced acute pain and chronic painful neuropathy. Significance statement: In this original work, it is shown for the first time that paclitaxel activates peripheral sensory and spinal neurons directly and sensitizes these cells to transient receptor potential vanilloid subtype 1 (TRPV1)-mediated capsaicin responses via Toll-like receptor 4 (TLR4) in multiple species. A direct functional interaction between TLR4 and TRPV1 is shown in rat and human dorsal root ganglion neurons, TLR4/TRPV1-coexpressing HEK293 cells, and in both rat and mouse spinal cord slices. Moreover, this is the first study to show that this interaction plays an important role in the generation of behavioral hypersensitivity in paclitaxel-related neuropathy. The key translational implications are that TLR4 and TRPV1 antagonists may be useful in the prevention and treatment of chemotherapy-induced peripheral neuropathy in humans.",
"Nociceptive sensory neurons are unusual in expressing voltage-gated inward currents carried by sodium channels resistant to block by tetrodotoxin (TTX) as well as currents carried by conventional TTX-sensitive sodium channels and voltage-dependent calcium channels. To examine how currents carried by each of these helps to shape the action potential in small-diameter dorsal root ganglion cell bodies, we voltage clamped cells by using the action potential recorded from each cell as the command voltage. Using intracellular solutions of physiological ionic composition, we isolated individual components of current flowing during the action potential with the use of channel blockers (TTX for TTX-sensitive sodium currents and a mixture of calcium channel blockers for calcium currents) and ionic substitution (TTX-resistant current measured by the replacement of extracellular sodium by N-methyl-D-glucamine in the presence of TTX, with correction for altered driving force). TTX-resistant sodium channels activated quickly enough to carry the largest inward charge during the upstroke of the nociceptor action potential (approximately 58%), with TTX-sensitive sodium channels also contributing significantly ( approximately 40%), especially near threshold, and high voltage-activated calcium currents much less (approximately 2%). Action potentials had a prominent shoulder during the falling phase, characteristic of nociceptive neurons. TTX-resistant sodium channels did not inactivate completely during the action potential and carried the majority (58%) of inward current flowing during the shoulder, with high voltage-activated calcium current also contributing significantly (39%). Unlike calcium current, TTX-resistant sodium current is not accompanied by opposing calcium-activated potassium current and may provide an effective mechanism by which the duration of action potentials (and consequently calcium entry) can be regulated."
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What is the name of a medicinal plant?
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The medicinal plant,
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[
"In Kenya, most people especially in rural areas use traditional medicine and medicinal plants to treat many diseases including malaria. Malaria is of national concern in Kenya, in view of development of resistant strains of Plasmodium falciparum to drugs especially chloroquine, which had been effective and affordable. There is need for alternative and affordable therapy. Many antimalarial drugs have been derived from medicinal plants and this is evident from the reported antiplasmodial activity. The present study reports on the in vivo antimalarial activity and brine shrimp lethality of five medicinal plants traditionally used to treat malaria in Msambweni district, Kenya. A total of five aqueous crude extracts from different plant parts used in traditional medicine for the treatment of malaria were evaluated for their in vivo antimalarial activity using Plasmodium berghei infected Swiss mice and for their acute toxicity using Brine shrimp lethality test. The screened crude plant extracts suppressed parasitaemia as follows: Azadirachta indica (L) Burm. (Meliaceae), 3.1%; Dichrostachys cinerea (L) Wight et Arn (Mimosaceae), 6.3%; Tamarindus indica L. (Caesalpiniaceae), 25.1%; Acacia seyal Del. (Mimosaceae) 27.8% and Grewia trichocarpa Hochst ex A.Rich (Tiliaceae) 35.8%. In terms of toxicity, A.indica root bark extract had an LC50 of 285.8 µg/ml and was considered moderately toxic. T.indica stem bark extract and G.trichocarpa root extract had an LC50 of 516.4 and 545.8 µg/ml respectively and were considered to be weakly toxic while A.seyal and D.cinerea root extracts had a LC50>1000 µg/ml and were therefore considered to be non toxic. The results indicate that the aqueous extracts of the tested plants when used alone as monotherapy had antimalarial activity which was significantly different from that of chloroquine (P≤0.05). The results also suggest that the anecdotal efficacy of the above plants reported by the study community is related to synergism of phytoconstituents since the assayed plant parts are used in combination with others to treat malaria. It is also evident that none of the screened plant extracts is toxic to the arthropod invertebrate, Artemia salina L. (Artemiidae) larvae, justifying the continued use of the plant parts to treat malaria. A.seyal, G.trichocarpa and T.indica have not been reported before for in vivo antimalarial activity and brine shrimp lethality.",
"Indoor residual spraying (IRS) and long-lasting insecticidal nets (LLINs) are the primary tools for malaria prevention in Africa. It is not known whether reductions in malaria can be sustained after IRS is discontinued. Our aim in this study was to assess changes in malaria morbidity in an area of Uganda with historically high transmission where IRS was discontinued after a 4-year period followed by universal LLIN distribution. Individual-level malaria surveillance data were collected from 1 outpatient department and 1 inpatient setting in Apac District, Uganda, from July 2009 through November 2015. Rounds of IRS were delivered approximately every 6 months from February 2010 through May 2014 followed by universal LLIN distribution in June 2014. Temporal changes in the malaria test positivity rate (TPR) were estimated during and after IRS using interrupted time series analyses, controlling for age, rainfall, and autocorrelation. Data include 65 421 outpatient visits and 13 955 pediatric inpatient admissions for which a diagnostic test for malaria was performed. In outpatients aged <5 years, baseline TPR was 60%-80% followed by a rapid and then sustained decrease to 15%-30%. During the 4-18 months following discontinuation of IRS, absolute TPR values increased by an average of 3.29% per month (95% confidence interval, 2.01%-4.57%), returning to baseline levels. Similar trends were seen in outpatients aged ≥5 years and pediatric admissions. Discontinuation of IRS in an area with historically high transmission intensity was associated with a rapid increase in malaria morbidity to pre-IRS levels.",
"Malaria is one of the most important diseases in the world. Because of the devastating nature of the disease there is an urgent need to develop new drugs or vaccines for the treatment, prevention and management of the disease. The objective of the present study was to collect and document information on herbal remedies traditionally used for the treatment of malaria in the Dangme West District of Ghana. Data was collected from 67 indigenous households in ten communities in the district using a validated questionnaire. In total, 30 species of plants belonging to 28 genera in 20 families were reported to be used in the preparation of the herbal remedies. Mature leaves were the most (55%) common plant part used and 73.3% of the herbal remedies involved a single plant. Most of the herbal remedies were prepared by boiling and administered orally. The majority (47%) of the species of plants used were collected from their compounds or home gardens. Knowledge about malaria and treatment practices exists in the study area. Herbal remedies were commonly used by people for the treatment of malaria because they were cost-effective. They are also more accessible. Many of the species of plants used have been documented for the treatment of malaria as well as investigated for their phytochemical and antimalarial and/or antiplasmodial activity confirming the results of previous studies as well as rationalization of their traditional use. Five species of plants used in the study area, namely, Bambusa vulgaris Schrad. ex J.C. Wendl. (Poaceae), Deinbollia pinnata Schum. &Thonn. (Sapindaceae), Elaeis guineensis Jacq. (Arecaceae), Greenwayodendron sp. (Annonaceae) and Solanum torvum Sw (Solanaceae), are documented for the first time for their use in the treatment of malaria. \"The result of this study provides the basis for further pharmacological studies on the herbal remedies used\".",
"Malaria is one of the most rampant diseases today not only in Uganda but also throughout Africa. Hence, it needs very close attention as it can be severe, causing many deaths, especially due to the rising prevalence of pathogenic resistance to current antimalarial drugs. The majority of the Ugandan population relies on traditional herbal medicines for various health issues. Thus, herein, we review various plant resources used to treat malaria across communities in Uganda so as to provide comprehensive and valuable ethnobotanical data about these plants. Approximately 182 plant species from 63 different plant families are used for malaria treatment across several communities in Uganda, of which 112 plant species have been investigated for antimalarial activities and 96% of the plant species showing positive results. Some plants showed very strong antimalarial activities and could be investigated further for the identification and validation of potentially therapeutic antimalarial compounds. There is no record of an investigation of antimalarial activity for approximately 39% of the plant species used for malaria treatment, yet these plants could be potential sources for potent antimalarial remedies. Thus, the review provides guidance for areas of further research on potential plant resources that could be sources of compounds with therapeutic properties for the treatment of malaria. Some of the plants were investigated for antimalarial activities, and their efficacy, toxicity, and safety aspects still need to be studied.",
"Plants develop unorganized cell masses like callus and tumors in response to various biotic and abiotic stimuli. Since the historical discovery that the combination of two growth-promoting hormones, auxin and cytokinin, induces callus from plant explants in vitro, this experimental system has been used extensively in both basic research and horticultural applications. The molecular basis of callus formation has long been obscure, but we are finally beginning to understand how unscheduled cell proliferation is suppressed during normal plant development and how genetic and environmental cues override these repressions to induce callus formation. In this review, we will first provide a brief overview of callus development in nature and in vitro and then describe our current knowledge of genetic and epigenetic mechanisms underlying callus formation.",
"Fifty-five organic and aqueous extracts of 11 plants used in malaria therapy in Kisii District, Kenya were tested in vitro against chloroquine (CQ)-sensitive and resistant strains of Plasmodium falciparum. Of the plants tested, 73% were active (IC(50) < 100 microg/ml). Three plants, Vernonia lasiopus, Rhamnus prinoides and Ficus sur afforded extracts with IC(50) values ranging less than 30 microg/ml against both CQ-sensitive and resistant strains. Combination of some extracts with CQ against the multi-drug resistant P. falciparum isolate V1/S revealed some synergistic effect. The plant extracts with low IC(50) values may be used as sources for novel antimalarial compounds to be used alone or in combination with CQ.",
"In Tanzania and elsewhere, medicinal plants, including Maytenus senegalensis, are still widely used in the treatment of malaria and other ailments. The aim of the present study was to investigate the in vivo antiplasmodial and toxic effects in mice. Oral antiplasmodial and acute toxicity of the ethanolic root extract of M. senegalensis was evaluated in mice. The Peters 4-day in vivo antiplasmodial effect against early rodent malaria infection in chloroquine-sensitive Plasmodium berghei NK 65 strain in mice. The M. senegalensis extract was found non-toxic and the oral median lethal dose in mice was determined to be greater than 1,600 mg/kg body weight. The findings revealed a significant (P = 0.001) daily increase in the level of parasitaemia in the parasitized untreated groups and a significant (P < 0.001) dose dependent decrease in parasitaemia in the parasitized groups treated with varying doses ranging from 25 to 100 mg/kg body weight of M. senegalensis extract and the standard drug sulphadoxine/pyrimethamine at 25/1.25 mg/kg body weight. Overall, the dose dependent parasitaemia suppression effects were in the order of: 25/1.25 mg/kg body weight of sulphadoxine/pyrimethamine > 100 mg/kg > 75 mg/kg > 50 mg/kg > 25 mg/kg body weight of M. senegalensis extract. The implications of these findings is that M. senegalensis ethanolic root bark extract possess potent antiplasmodial effect and may, therefore, serve as potential sources of safe, effective and affordable anti-malarial drugs. The displayed high in vivo antiplasmodial activity and lack of toxic effect render M. senegalensis a candidate for the bioassay-guided isolation of compounds which could develop into new lead structures and candidates for drug development programmes against human malaria."
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Targeting ATP-binding cassette transporters in colorectal cancer using CRISPR/Cas9
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Multidrug resistance (MDR) has been extensively reported in colorectal cancer patients, which remains a major cause of chemotherapy failure. One of the critical mechanisms of MDR in colorectal cancer is the reduced intracellular drug level led by the upregulated expression of the ATP-binding cassette (ABC) transporters, particularly, ABCB1/P-gp. In this study, the CRISPR/Cas9 system was utilized to target
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[
"Multi-drug resistance (MDR) remains a significant obstacle to successful chemotherapy treatment for osteosarcoma patients. One of the central causes of MDR is the overexpression of the membrane bound drug transporter protein P-glycoprotein (P-gp), which is the protein product of the MDR gene ABCB1. Though several methods have been reported to reverse MDR in vitro and in vivo when combined with anticancer drugs, they have yet to be proven useful in the clinical setting. The meta-analysis demonstrated that a high level of P-gp may predict poor survival in patients with osteosarcoma. The expression of P-gp can be efficiently blocked by the clustered regularly interspaced short palindromic repeats (CRISPR)-associated Cas9 system (CRISPR-Cas9). Inhibition of ABCB1 was associated with reversing drug resistance in osteosarcoma MDR cell lines (KHOSR2 and U-2OSR2) to doxorubicin. We performed a meta-analysis to investigate the relationship between P-gp expression and survival in patients with osteosarcoma. Then we adopted the CRISPR-Cas9, a robust and highly efficient novel genome editing tool, to determine its effect on reversing drug resistance by targeting endogenous ABCB1 gene at the DNA level in osteosarcoma MDR cell lines. These results suggest that the CRISPR-Cas9 system is a useful tool for the modification of ABCB1 gene, and may be useful in extending the long-term efficacy of chemotherapy by overcoming P-gp-mediated MDR in the clinical setting.",
"Indeed, multi-drug resistance (MDR) is a significant obstacle to effective chemotherapy. The overexpression of ATP-binding cassette (ABC) membrane transporters is a principal cause of enhanced cytotoxic drug efflux and treatment failure in various types of cancers. At cellular level, the pumps of ABC family regulate the transportation of numerous substances including drugs in and out of the cells. In past, the overexpression of ABC pumps suggested a well-known mechanism of drug resistance in cancers as well as infectious diseases. In oncology, the search for new compounds for the inhibition of these hyperactive ABC pumps either genetically or functionally, growing interest to reverse multi-drug resistance and increase chemotherapeutic effects. Several ABC pump inhibitor/modulators has been explored to address the cancer associated MDR. However, the clinical results are still disappointing and conventional chemotherapies are constantly failed in successful eradication of MDR tumors. In this context, the structural and functional understanding of different ATP pumps is most important. In this concise review, we elaborated basic crystal structure of ABC transporter proteins as well as its critical elements such as different domains, motifs as well as some important amino acids which are responsible for ATP binding and drug efflux as well as demonstrated an ATP-switch model employed by various ABC membrane transporters. Furthermore, we briefly summarized different newly identified MDR inhibitors/modulators, deployed alone or in combination with cytotoxic agents to deal with MDR in different types of cancers.",
"In recent years, gene editing, especially that using clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9, has made great progress in the field of gene function. Rapid development of gene editing techniques has contributed to their significance in the field of medicine. Because the CRISPR/Cas9 gene editing tool is not only powerful but also has features such as strong specificity and high efficiency, it can accurately and rapidly screen the whole genome, facilitating the administration of gene therapy for specific diseases. In the field of tumor research, CRISPR/Cas9 can be used to edit genomes to explore the mechanisms of tumor occurrence, development, and metastasis. In these years, this system has been increasingly applied in tumor treatment research. CRISPR/Cas9 can be used to treat tumors by repairing mutations or knocking out specific genes. To date, numerous preliminary studies have been conducted on tumor treatment in related fields. CRISPR/Cas9 holds great promise for gene-level tumor treatment. Personalized and targeted therapy based on CRISPR/Cas9 will possibly shape the development of tumor therapy in the future. In this study, we review the findings of CRISPR/Cas9 for tumor treatment research to provide references for related future studies on the pathogenesis and clinical treatment of tumors.",
"Survival of osteosarcoma patients is currently limited by the development of metastases and multidrug resistance (MDR). A well-established cause of MDR involves overexpression of P-glycoprotein (Pgp) in tumor cells. However, some discrepancies still exist as to the clinical significance of Pgp in osteosarcoma. We sought to elucidate further whether the Pgp expression correlated with clinical behavior in a series of patients with osteosarcoma via high-throughput tissue microarray (TMA) analysis. Immunohistochemical analysis of Pgp expression in a TMA of 114 specimens with a retrospective review of 70 osteosarcoma patients admitted to the Massachusetts General Hospital (MGH) was performed. High Pgp expression was correlated with metastasis development and poor response to pre-operative chemotherapy in osteosarcoma patients. Eighteen of the fifty-seven patients initially admitted with primary osteosarcoma showed high Pgp expression. Among these 18 patients with high Pgp expression, 13 of 18 (72%) patients eventually developed metastases. There was no significant clinical relevance between Pgp expression and osteosarcoma survival. These results support that high expression of Pgp is important, but cannot be assigned as, an individual predictor in the development of human osteosarcoma. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1606-1612, 2016."
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Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Symptom Trajectories and Oropharyngeal Viral Shedding in Outpatients with Uncomplicated Coronavirus Disease of 2019
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Although the vast majority of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infections are uncomplicated, our understanding of predictors of symptom resolution and viral shedding cessation remains limited. We characterized symptom trajectories and oropharyngeal viral shedding among 120 outpatients with uncomplicated Coronavirus Disease of 2019 (COVID-19) enrolled in a clinical trial of Peginterferon Lambda, which demonstrated no clinical or virologic benefit compared with placebo. In the combined trial cohort, objective fever was uncommon, inflammatory symptoms (myalgias, fatigue) peaked at 4 to 5 days postsymptom onset, and cough peaked at 9 days. The median time to symptom resolution from earliest symptom onset was 17 days (95% confidence interval 14-18). SARS-CoV-2 IgG seropositivity at enrollment was associated with hastened resolution of viral shedding (hazard ratio 1.80, 95% confidence interval 1.05-3.1, P = 0.03), but not with symptom resolution. Inflammatory symptoms were associated with a significantly greater odds of oropharyngeal SARS-CoV-2 RNA detection; respiratory symptoms were not. These findings have important implications for COVID-19 screening approaches and trial design.
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"Coronavirus disease 2019 (COVID-19), currently a worldwide pandemic, is a viral illness caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The suspected contribution of thrombotic events to morbidity and mortality in COVID-19 patients has prompted a search for novel potential options for preventing COVID-19-associated thrombotic disease. In this article by the Global COVID-19 Thrombosis Collaborative Group, we describe novel dosing approaches for commonly used antithrombotic agents (especially heparin-based regimens) and the potential use of less widely used antithrombotic drugs in the absence of confirmed thrombosis. Although these therapies may have direct antithrombotic effects, other mechanisms of action, including anti-inflammatory or antiviral effects, have been postulated. Based on survey results from this group of authors, we suggest research priorities for specific agents and subgroups of patients with COVID-19. Further, we review other agents, including immunomodulators, that may have antithrombotic properties. It is our hope that the present document will encourage and stimulate future prospective studies and randomized trials to study the safety, efficacy, and optimal use of these agents for prevention or management of thrombosis in COVID-19.",
"Xanthine oxidase and xanthine dehydrogenase have been implicated in producing myocardial damage following reperfusion of an occluded coronary artery. We investigated and compared the effect of febuxostat and allopurinol in an experimental model of ischemia-reperfusion (IR) injury with a focus on the signaling pathways involved. Male Wistar rats were orally administered vehicle (CMC) once daily (sham and IR + control), febuxostat (10 mg/kg/day; FEB10 + IR), or allopurinol (100 mg/kg/day; ALL100 + IR) for 14 days. On the 15th day, the IR-control and treatment groups were subjected to one-stage left anterior descending (LAD) coronary artery ligation for 45 minutes followed by a 60-minute reperfusion. Febuxostat and allopurinol pretreatment significantly improved cardiac function and maintained morphological alterations. They also attenuated oxidative stress and apoptosis by suppressing the expression of proapoptotic proteins (Bax and caspase-3), reducing TUNEL-positive cells, and increasing the level of antiapoptotic proteins (Bcl-2). The MAPK-based molecular mechanism revealed suppression of active JNK and p38 proteins concomitant with the rise in ERK1/ERK2, a prosurvival kinase. Additionally, a reduction in the level of inflammatory markers (TNF-α, IL-6, and NF-κB) was also observed. The changes observed with febuxostat were remarkable in comparison with those observed with allopurinol. Febuxostat protects relatively better against IR injury than allopurinol by suppressing inflammation and apoptosis mediating the MAPK/NF-κBp65/TNF-α pathway.",
"The aim of this clinical trial was to evaluate the effects of febuxostat (FBX) in comparison with hydroxychloroquine (HCQ) on clinical symptoms, laboratory tests and chest CT findings in outpatients with moderate symptoms of COVID-19 infection. We conducted a clinical trial involving adult outpatients with the moderate respiratory illness following COVID-19 infection. Patients were randomly assigned to receive either FBX or HCQ for 5 days. The measured variables were needs to hospitalisation, clinical and laboratory data including fever, cough, breathing rate, C-Reactive Protein level, lymphocytes count at onset of admission and was well as at 5 days of treatments. In addition, CT findings were evaluated on admission and 14 days after initiation of treatment. Sixty subjects were enrolled in the study with a 1 to 1 ratio in FBX and HCQ groups. On admission, fever (66.7%), cough (87%), tachypnoea (44.4%), dyspnoea (35%), elevated CRP value (94.4%) and lung involvement according to chest CT (100%) were documented in enrolled patients with insignificant difference between FBX and HCQ groups. Fever, cough and tachypnoea were significantly mitigated in both groups after five days of treatments without any significant differences between groups. The mean percentages of lung involvement were significantly reduced to 7.3% and 8% after 14 days of treatment with FBX and HCQ, respectively. In adult outpatients with moderate COVID-19 infection, the effectiveness of FBX and HCQ was not different in terms of resolution of clinical manifestations, laboratory tests and lung CT findings. This trial suggests that FBX is as an alternative treatment to HCQ for COVID-19 infection and may be considered in patients with a contraindication or precaution to HCQ."
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Enhancing Critical Thinking Skills Among Arab College Students
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Critical thinking is a requisite skill for college success, employability, and conducive active civic participation. Empirical studies have noted to the low achievement of Arab students on critical thinking assessments. Insufficient endeavors have attempted to propose effective interventions enhancing critical thinking abilities among Arab students. The current analysis provides a preliminary overview of a special course designed to improve critical thinking skills among Arab college students. Results indicated a great improvement in all areas of critical thinking including explanation of information, identification of strategies, implementing solutions, and formulating logical inferences. Students' scores on a critical thinking assessment increased from sufficient to good as a result of participating in the program. The gains are consistent after controlling for gender, major, class seniority, and nationality. Notwithstanding these promising results, this paper is limited in several respects including the choice of critical thinking assessments represented by two questions, the highly contextualized setting making it difficult to be replicated, and the convenient sampling strategy used to recruit participants. This set of limitations, however, does not discourage proactive attempts like designing special courses to enhance students' critical thinking acquisition in the Middle East.
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"Scientific framework is important in designing curricula and evaluating students in the field of education and clinical practice. The purpose of this study was to examine the effectiveness of non-traditional educational methods on critical thinking skills. A systematic review approach was applied. Studies published in peer-reviewed journals from January 2001 to December 2014 were searched using electronic databases and major education journals. A meta-analysis was performed using Review Manager 5.2. Reviewing the included studies, the California Critical Thinking Dispositions Inventory (CCTDI) and California Critical Thinking Skills Test (CCTST) were used to assess the effectiveness of critical thinking in the meta-analysis. The eight CCTDI datasets showed that non- traditional teaching methods (i.e., no lectures) were more effective compared to control groups (standardized mean difference [SMD]: 0.42, 95 % confidence interval [CI]: 0.26-0.57, p < .00001). And six CCTST datasets showed the teaching and learning methods in these studies were also had significantly more effects when compared to the control groups (SMD: 0.29, 95 % CI: 0.10-0.48, p = 0.003). This research showed that new teaching and learning methods designed to improve critical thinking were generally effective at enhancing critical thinking dispositions.",
"A quasi-experimental, two-group pretest-post-test design was conducted to examine the effect of problem-based learning on the critical thinking skills of 46 Year 2 undergraduate nursing students in the People's Republic of China. The California Critical Thinking Skills Test Form A, Chinese-Taiwanese version was used as both a pretest and as a post-test for a semester-long nursing course. There was no significant difference in critical thinking skills at pretest, whereas, significant differences in critical thinking skills existed between the problem-based learning and lecture groups at post-test. The problem-based learning students had a significantly greater improvement on the overall California Critical Thinking Skills Test, analysis, and induction subscale scores compared with the lecture students. Problem-based learning fostered nursing students' critical thinking skills."
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Temporal prediction error processing in human primary somatosensory cortex
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The human brain continuously generates predictions of incoming sensory input and calculates corresponding prediction errors from the perceived inputs to update internal predictions. In human primary somatosensory cortex (area 3b), different cortical layers are involved in receiving the sensory input and generation of error signals. It remains unknown, however, how the layers in the human area 3b contribute to the temporal prediction error processing. To investigate prediction error representation in the area 3b across layers, we acquired layer-specific functional magnetic resonance imaging (fMRI) data at 7T from human area 3b during a task of index finger poking with no-delay, short-delay and long-delay touching sequences. We demonstrate that all three tasks increased activity in both superficial and deep layers of area 3b compared to the random sensory input. The fMRI signal was differentially modulated solely in the deep layers rather than the superficial layers of area 3b by the delay time. Compared with the no-delay stimuli, activity was greater in the deep layers of area 3b during the short-delay stimuli but lower during the long-delay stimuli. This difference activity features in the superficial and deep layers suggest distinct functional contributions of area 3b layers to tactile temporal prediction error processing. The functional segregation in area 3b across layers may reflect that the excitatory and inhibitory interplay in the sensory cortex contributions to flexible communication between cortical layers or between cortical areas.
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"Improvements in the spatial resolution of structural and functional MRI are beginning to enable analysis of intracortical structures such as heavily myelinated layers in 3D, a prerequisite for in-vivo parcellation of individual human brains. This parcellation can only be performed precisely if the profiles used in cortical analysis are anatomically meaningful. Profiles are often constructed as traverses that are perpendicular to computed laminae. In this case they are fully determined by these laminae. The aim of this study is to evaluate models for cortical laminae used so far and to establish a new model. Methods to model the laminae used so far include constructing laminae that keep a constant distance to the cortical boundaries, so-called equidistant laminae. Another way is to compute equipotentials between the cortical boundary surfaces with the Laplace equation. The Laplace profiles resulting from the gradients to the equipotentials were often-used because of their nice mathematical properties. However, the equipotentials these Laplacian profiles are constructed from and the equidistant laminae do not follow the anatomical layers observed using high resolution MRI of cadaver brain. To remedy this problem, we introduce a novel equi-volume model that derives from work by Bok (1929). He argued that cortical segments preserve their volume, while layer thickness changes to compensate cortical folding. We incorporate this preservation of volume in our new equi-volume model to generate a three-dimensional well-adapted undistorted coordinate system of the cortex. When defined by this well-adapted coordinate system, cortical depth is anatomically meaningful. We compare isocontours from these cortical depth values to locations of myelinated bands on high-resolution ex-vivo and in-vivo three-dimensional MR images. A similar comparison was performed with equipotentials computed with the Laplace equation and with equidistant isocontours. A quantitative evaluation of the equi-volume model using measured image intensities confirms that it provides a much better fit to observed cortical layering.",
"This paper demonstrates that nonnegative matrix factorisation is mathematically related to a class of neural networks that employ negative feedback as a mechanism of competition. This observation inspires a novel learning algorithm which we call Divisive Input Modulation (DIM). The proposed algorithm provides a mathematically simple and computationally efficient method for the unsupervised learning of image components, even in conditions where these elementary features overlap considerably. To test the proposed algorithm, a novel artificial task is introduced which is similar to the frequently-used bars problem but employs squares rather than bars to increase the degree of overlap between components. Using this task, we investigate how the proposed method performs on the parsing of artificial images composed of overlapping features, given the correct representation of the individual components; and secondly, we investigate how well it can learn the elementary components from artificial training images. We compare the performance of the proposed algorithm with its predecessors including variations on these algorithms that have produced state-of-the-art performance on the bars problem. The proposed algorithm is more successful than its predecessors in dealing with overlap and occlusion in the artificial task that has been used to assess performance.",
"In recent years, two-photon imaging has become an invaluable tool in neuroscience, as it allows for chronic measurement of the activity of genetically identified cells during behavior(1-6). Here we describe methods to perform two-photon imaging in mouse cortex while the animal navigates a virtual reality environment. We focus on the aspects of the experimental procedures that are key to imaging in a behaving animal in a brightly lit virtual environment. The key problems that arise in this experimental setup that we here address are: minimizing brain motion related artifacts, minimizing light leak from the virtual reality projection system, and minimizing laser induced tissue damage. We also provide sample software to control the virtual reality environment and to do pupil tracking. With these procedures and resources it should be possible to convert a conventional two-photon microscope for use in behaving mice.",
"Processing in cortical circuits is driven by combinations of cortical and subcortical inputs. These inputs are often conceptually categorized as bottom-up, conveying sensory information, and top-down, conveying contextual information. Using intracellular recordings in mouse primary visual cortex, we measured neuronal responses to visual input, locomotion, and visuomotor mismatches. We show that layer 2/3 (L2/3) neurons compute a difference between top-down motor-related input and bottom-up visual flow input. Most L2/3 neurons responded to visuomotor mismatch with either hyperpolarization or depolarization, and the size of this response was correlated with distinct physiological properties. Consistent with a subtraction of bottom-up and top-down input, visual and motor-related inputs had opposing influence on L2/3 neurons. In infragranular neurons, we found no evidence of a difference computation and responses were consistent with positive integration of visuomotor inputs. Our results provide evidence that L2/3 functions as a bidirectional comparator of top-down and bottom-up input.",
"The neural basis of the internal models used in sensorimotor transformations is beginning to be uncovered. Sensorimotor learning involves the modification of such models. Different stages of sensory-motor processing have been explored with a continuum of experimental tasks, from learning arbitrary associations of sensory cues to movements, to adapting to altered kinematic and dynamic environments. Several groups have been studying changes in neuronal activity in cortical and subcortical areas that may be related to the acquisition and consolidation processes. We discuss the progress and challenges in understanding how these learning-related neural changes are involved in the modification of internal models, and offer future directions.",
"The capacity to predict future events permits a creature to detect, model, and manipulate the causal structure of its interactions with its environment. Behavioral experiments suggest that learning is driven by changes in the expectations about future salient events such as rewards and punishments. Physiological work has recently complemented these studies by identifying dopaminergic neurons in the primate whose fluctuating output apparently signals changes or errors in the predictions of future salient and rewarding events. Taken together, these findings can be understood through quantitative theories of adaptive optimizing control.",
"Local microcircuits within neocortical columns form key determinants of sensory processing. Here, we investigate the excitatory synaptic neuronal network of an anatomically defined cortical column, the C2 barrel column of mouse primary somatosensory cortex. This cortical column is known to process tactile information related to the C2 whisker. Through multiple simultaneous whole-cell recordings, we quantify connectivity maps between individual excitatory neurons located across all cortical layers of the C2 barrel column. Synaptic connectivity depended strongly upon somatic laminar location of both presynaptic and postsynaptic neurons, providing definitive evidence for layer-specific signaling pathways. The strongest excitatory influence upon the cortical column was provided by presynaptic layer 4 neurons. In all layers we found rare large-amplitude synaptic connections, which are likely to contribute strongly to reliable information processing. Our data set provides the first functional description of the excitatory synaptic wiring diagram of a physiologically relevant and anatomically well-defined cortical column at single-cell resolution."
] |
Bipartite motifs learning: a parameter-free web server for gene regulation analysis
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Motif discovery and characterization are important for gene regulation analysis. The lack of intuitive and integrative web servers impedes the effective use of motifs. Most motif discovery web tools are either not designed for non-expert users or lacking optimization steps when using default settings. Here we describe bipartite motifs learning (BML), a parameter-free web server that provides a user-friendly portal for online discovery and analysis of sequence motifs, using high-throughput sequencing data as the input. BML utilizes both position weight matrix and dinucleotide weight matrix, the latter of which enables the expression of the interdependencies of neighboring bases. With input parameters concerning the motifs are given, the BML achieves significantly higher accuracy than other available tools for motif finding. When no parameters are given by non-expert users, unlike other tools, BML employs a learning method to identify motifs automatically and achieve accuracy comparable to the scenario where the parameters are set. The BML web server is freely available at http://motif.t-ridership.com/ (https://github.com/Mohammad-Vahed/BML).
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[
"The purpose of this article is to provide a brief history of the development and application of computer algorithms for the analysis and prediction of DNA binding sites. This problem can be conveniently divided into two subproblems. The first is, given a collection of known binding sites, develop a representation of those sites that can be used to search new sequences and reliably predict where additional binding sites occur. The second is, given a set of sequences known to contain binding sites for a common factor, but not knowing where the sites are, discover the location of the sites in each sequence and a representation for the specificity of the protein.",
"BIOLOGY IS ENCODED IN MOLECULAR SEQUENCES: deciphering this encoding remains a grand scientific challenge. Functional regions of DNA, RNA, and protein sequences often exhibit characteristic but subtle motifs; thus, computational discovery of motifs in sequences is a fundamental and much-studied problem. However, most current algorithms do not allow for insertions or deletions (indels) within motifs, and the few that do have other limitations. We present a method, GLAM2 (Gapped Local Alignment of Motifs), for discovering motifs allowing indels in a fully general manner, and a companion method GLAM2SCAN for searching sequence databases using such motifs. glam2 is a generalization of the gapless Gibbs sampling algorithm. It re-discovers variable-width protein motifs from the PROSITE database significantly more accurately than the alternative methods PRATT and SAM-T2K. Furthermore, it usefully refines protein motifs from the ELM database: in some cases, the refined motifs make orders of magnitude fewer overpredictions than the original ELM regular expressions. GLAM2 performs respectably on the BAliBASE multiple alignment benchmark, and may be superior to leading multiple alignment methods for \"motif-like\" alignments with N- and C-terminal extensions. Finally, we demonstrate the use of GLAM2 to discover protein kinase substrate motifs and a gapped DNA motif for the LIM-only transcriptional regulatory complex: using GLAM2SCAN, we identify promising targets for the latter. GLAM2 is especially promising for short protein motifs, and it should improve our ability to identify the protein cleavage sites, interaction sites, post-translational modification attachment sites, etc., that underlie much of biology. It may be equally useful for arbitrarily gapped motifs in DNA and RNA, although fewer examples of such motifs are known at present. GLAM2 is public domain software, available for download at http://bioinformatics.org.au/glam2.",
"The motif discovery problem consists of finding recurring patterns of short strings in a set of nucleotide sequences. This classical problem is receiving renewed attention as most early motif discovery methods lack the ability to handle large data of recent genome-wide ChIP studies. New ChIP-tailored methods focus on reducing computation time and pay little regard to the accuracy of motif detection. Unlike such methods, our method focuses on increasing the detection accuracy while maintaining the computation efficiency at an acceptable level. The major advantage of our method is that it can mine diverse multiple motifs undetectable by current methods. The repulsive parallel Markov chain Monte Carlo (RPMCMC) algorithm that we propose is a parallel version of the widely used Gibbs motif sampler. RPMCMC is run on parallel interacting motif samplers. A repulsive force is generated when different motifs produced by different samplers near each other. Thus, different samplers explore different motifs. In this way, we can detect much more diverse motifs than conventional methods can. Through application to 228 transcription factor ChIP-seq datasets of the ENCODE project, we show that the RPMCMC algorithm can find many reliable cofactor interacting motifs that existing methods are unable to discover.",
"In some dimeric cases of transcription factor (TF) binding, the specificity of dimeric motifs has been observed to differ notably from what would be expected were the two factors to bind to DNA independently of each other. Current motif discovery methods are unable to learn monomeric and dimeric motifs in modular fashion such that deviations from the expected motif would become explicit and the noise from dimeric occurrences would not corrupt monomeric models. We propose a novel modeling technique and an expectation maximization algorithm, implemented as software tool MODER, for discovering monomeric TF binding motifs and their dimeric combinations. Given training data and seeds for monomeric motifs, the algorithm learns in the same probabilistic framework a mixture model which represents monomeric motifs as standard position-specific probability matrices (PPMs), and dimeric motifs as pairs of monomeric PPMs, with associated orientation and spacing preferences. For dimers the model represents deviations from pure modular model of two independent monomers, thus making co-operative binding effects explicit. MODER can analyze in reasonable time tens of Mbps of training data. We validated the tool on HT-SELEX and ChIP-seq data. Our findings include some TFs whose expected model has palindromic symmetry but the observed model is directional.",
"Identifying regulatory elements is a fundamental problem in the field of gene transcription. Motif discovery-the task of identifying the sequence preference of transcription factor proteins, which bind to these elements-is an important step in this challenge. MEME is a popular motif discovery algorithm. Unfortunately, MEME's running time scales poorly with the size of the dataset. Experiments such as ChIP-Seq and DNase-Seq are providing a rich amount of information on the binding preference of transcription factors. MEME cannot discover motifs in data from these experiments in a practical amount of time without a compromising strategy such as discarding a majority of the sequences. We present EXTREME, a motif discovery algorithm designed to find DNA-binding motifs in ChIP-Seq and DNase-Seq data. Unlike MEME, which uses the expectation-maximization algorithm for motif discovery, EXTREME uses the online expectation-maximization algorithm to discover motifs. EXTREME can discover motifs in large datasets in a practical amount of time without discarding any sequences. Using EXTREME on ChIP-Seq and DNase-Seq data, we discover many motifs, including some novel and infrequent motifs that can only be discovered by using the entire dataset. Conservation analysis of one of these novel infrequent motifs confirms that it is evolutionarily conserved and possibly functional. All source code is available at the Github repository http://github.com/uci-cbcl/EXTREME.",
"Prediction of transcription factor binding sites is an important challenge in genome analysis. The advent of next generation genome sequencing technologies makes the development of effective computational approaches particularly imperative. We have developed a novel training-based methodology intended for prokaryotic transcription factor binding site prediction. Our methodology extends existing models by taking into account base interdependencies between neighbouring positions using conditional probabilities and includes genomic background weighting. This has been tested against other existing and novel methodologies including position-specific weight matrices, first-order Hidden Markov Models and joint probability models. We have also tested the use of gapped and ungapped alignments and the inclusion or exclusion of background weighting. We show that our best method enhances binding site prediction for all of the 22 Escherichia coli transcription factors with at least 20 known binding sites, with many showing substantial improvements. We highlight the advantage of using block alignments of binding sites over gapped alignments to capture neighbouring position interdependencies. We also show that combining these methods with ChIP-on-chip data has the potential to further improve binding site prediction. Finally we have developed the ungapped likelihood under positional background platform: a user friendly website that gives access to the prediction method devised in this work."
] |
Sleep health in alternative care settings: a call to action
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Sleep health is a critical but under-recognized area of concern for the more than 650,000 children served by the US child welfare system each year. While sleep is vital to optimal child health and development, it is likely harmed by the multiple adversities and traumas experienced among children and youth residing in alternative care settings (ie, kinship care, nonrelative foster care, group homes). Children residing in alternative care settings have experienced, at a minimum, the trauma of removal from a biological parent's care and would benefit from holistic, comprehensive care approaches inclusive of sleep health. Furthermore, few studies are currently available to guide practitioners and policymakers in promoting sleep health among these children. In this Call to Action, our goal is to draw attention to the sleep health of children residing in alternative care settings. We highlight the need for a more robust evidence base to address major knowledge gaps and outline concrete steps toward building future promising sleep health-promoting practices and policies supporting children residing in alternative care settings.
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[
"Sleep is important for overall health and well-being. Insufficient sleep and sleep disorders are highly prevalent among adults and children and therefore a public health burden, particularly because poor sleep is associated with adverse health outcomes. Emerging evidence has demonstrated that environmental factors at the household- and neighborhood-level can alter healthy sleep. This paper will (1) review recent literature on the environmental determinants of sleep among adults as well as children and adolescents; and (2) discuss the opportunities and challenges for advancing research on the environment and sleep. Epidemiologic research has shown that social features of environments, family, social cohesion, safety, noise, and neighborhood disorder can shape and/or impact sleep patterns; and physical features such as light, noise, traffic, pollution, and walkability can also influence sleep and is related to sleep disorders among adults and children. Prior research has mainly measured one aspect of the environment, relied on self-reported sleep, which does not correlate well with objective measures, and investigated cross-sectional associations. Although most studies are conducted among non-Hispanic white populations, there is growing evidence that indicates that minority populations are particularly vulnerable to the effects of the environment on insufficient sleep and sleep disorders. There is clear evidence that environmental factors are associated with insufficient sleep and sleep disorders. However, more research is warranted to evaluate how and which environmental factors contribute to sleep health. Interventions that target changes in the environment to promote healthy sleep should be developed, tested, and evaluated as a possible pathway for ameliorating sleep health disparities and subsequently health disparities.",
"African American children are overrepresented in foster care at twice to three times the rate of white children. Scholars argue that racism and oppression underlie disproportionality (Križ & Skivenes, 2011). This study explored disproportionality as seen through the eyes of African American parents in the child welfare system. The aim was to understand why African American families are over-represented in child custody statistics and to improve family and parenting support for African American communities. Participants included twenty-one African Americans--12 women and 9 men, two of whom were foster parents and 19 of whom were parents involved with child welfare services. All participants reside in two impoverished areas in southern United States. Focus groups were used to collect data and were conducted at a community center. The method of analysis was constant comparison analysis (Strauss) and thematic analysis of the focus group discussions in the context of institutional policy. Six themes (profound lack of trust; overwhelming trauma; severe and persistent poverty; health and mental health; socio-economic conditions; and sense of social isolation were identified, along with three participant suggestions to improve child welfare services (family support services, economic revival, and better communication). In the current study we note the strong link between poverty, child maltreatment, and child removal and conclude with an exploration of practice and policy implications with recommendations for a way forward. The need for culturally competent and trauma informed child welfare services is also discussed.",
"Reducing health disparities requires an understanding of the mechanisms that generate disparities. Life course approaches to health disparities leverage theories that explain how socially patterned physical, environmental, and socioeconomic exposures at different stages of human development shape health within and across generations and can therefore offer substantial insight into the etiology of health disparities. Life course approaches are informed by developmental and structural perspectives. Developmental perspectives emphasize how socially patterned exposures to risk factors during sensitive life stages shift health trajectories, whereas structural perspectives emphasize how social identity and position within socially patterned environments disproportionately allocate risk factors and resources, resulting in altered health trajectories. We conclude that the science of health disparities will be advanced by integrating life course approaches into etiologic and intervention research on health disparities. The following 4 strategies are offered to guide in this process: (1) advance the understanding of multiple exposures and their interactions, (2) integrate life course approaches into the understanding of biological mechanisms, (3) explore transgenerational transmission of health disparities, and (4) integrate life course approaches into health disparities interventions.",
"Multilevel interventions can be uniquely effective at addressing minority health and health disparities, but they pose substantial methodological, data analytic, and assessment challenges that must be considered when designing and applying interventions and assessment. To facilitate the adoption of multilevel interventions to reduce health disparities, we outline areas of need in filling existing operational challenges to the design and assessment of multilevel interventions. We discuss areas of development that address overarching constructs inherent in multilevel interventions, with a particular focus on their application to minority health and health disparities. Our approach will prove useful to researchers, as it allows them to integrate information related to health disparities research into the framework of broader constructs with which they are familiar. We urge researchers to prioritize building transdisciplinary teams and the skills needed to overcome the challenges in designing and assessing multilevel interventions, as even small contributions can accelerate progress toward improving minority health and reducing health disparities. To make substantial progress, however, a concerted and strategic effort, including work to advance analytic techniques and measures, is needed.",
"We investigated physical activity as a moderator of relations between sleep duration and quality and adolescents' internalizing and externalizing problems. The study used a cross-sectional design. Participants were recruited from small towns and semi-urban communities in Alabama. The sample was comprised of 235 adolescents (Mage = 15.78 years, SD = 9.60 months) who were diverse in sex (53% female), race/ethnicity (34% Black/African American, 66% White), and socioeconomic status. Sleep duration (actual sleep minutes), efficiency (minutes/total sleep period), and latency (minutes from sleep attempt to onset) were examined with actigraphs for 1 week. Youth reported on their physical activity levels and internalizing and externalizing problems. Interactions between sleep and physical activity emerged in the prediction of adolescents' internalizing and externalizing problems. Supportive of moderation effects, adolescents with shorter or poorer-quality sleep in conjunction with less physical activity showed the highest levels of internalizing and externalizing problems. Demonstrative of protective effects, adolescents with more physical activity had lower levels of internalizing and externalizing problems regardless of their sleep duration or quality. Findings illustrate that not all youth are at equal risk for adjustment problems when they experience short or poor-quality sleep, suggesting the importance of examining both bioregulatory and environmental factors in understanding adolescent adjustment.",
"Sleep deficiencies, which include insufficient or long sleep duration, poor sleep quality, and irregular timing of sleep, are disproportionately distributed among populations that experience health disparities in the United States. Sleep deficiencies are associated with a wide range of suboptimal health outcomes, high-risk health behaviors, and poorer overall functioning and well-being. This report focuses on sleep health disparities (SHDs), which is a term defined as differences in one or more dimensions of sleep health on a consistent basis that adversely affect designated disadvantaged populations. SHDs appear to share many of the same determinants and causal pathways observed for health outcomes with well-known disparities. There also appears to be common behavioral and biological mechanisms that connect sleep with poorer health outcomes, suggesting a link between SHDs and other health disparities observed within these designated populations. In 2018, the National Institute on Minority Health and Health Disparities, the National Heart, Lung, and Blood Institute, and the Office of Behavioral and Social Sciences Research convened a workshop with experts in sleep, circadian rhythms, and health disparities to identify research gaps, challenges, and opportunities to better understand and advance research to address SHDs. The major strategy to address SHDs is to promote integration between health disparity causal pathways and sleep and circadian-related mechanisms in research approaches and study designs. Additional strategies include developing a comprehensive, integrative conceptual model, building transdisciplinary training and research infrastructure, and designing as well as testing multilevel, multifactorial interventions to address SHDs.",
"The relationship between income and life expectancy is well established but remains poorly understood. To measure the level, time trend, and geographic variability in the association between income and life expectancy and to identify factors related to small area variation. Income data for the US population were obtained from 1.4 billion deidentified tax records between 1999 and 2014. Mortality data were obtained from Social Security Administration death records. These data were used to estimate race- and ethnicity-adjusted life expectancy at 40 years of age by household income percentile, sex, and geographic area, and to evaluate factors associated with differences in life expectancy. Pretax household earnings as a measure of income. Relationship between income and life expectancy; trends in life expectancy by income group; geographic variation in life expectancy levels and trends by income group; and factors associated with differences in life expectancy across areas. The sample consisted of 1,408,287,218 person-year observations for individuals aged 40 to 76 years (mean age, 53.0 years; median household earnings among working individuals, $61,175 per year). There were 4,114,380 deaths among men (mortality rate, 596.3 per 100,000) and 2,694,808 deaths among women (mortality rate, 375.1 per 100,000). The analysis yielded 4 results. First, higher income was associated with greater longevity throughout the income distribution. The gap in life expectancy between the richest 1% and poorest 1% of individuals was 14.6 years (95% CI, 14.4 to 14.8 years) for men and 10.1 years (95% CI, 9.9 to 10.3 years) for women. Second, inequality in life expectancy increased over time. Between 2001 and 2014, life expectancy increased by 2.34 years for men and 2.91 years for women in the top 5% of the income distribution, but by only 0.32 years for men and 0.04 years for women in the bottom 5% (P < .001 for the differences for both sexes). Third, life expectancy for low-income individuals varied substantially across local areas. In the bottom income quartile, life expectancy differed by approximately 4.5 years between areas with the highest and lowest longevity. Changes in life expectancy between 2001 and 2014 ranged from gains of more than 4 years to losses of more than 2 years across areas. Fourth, geographic differences in life expectancy for individuals in the lowest income quartile were significantly correlated with health behaviors such as smoking (r = -0.69, P < .001), but were not significantly correlated with access to medical care, physical environmental factors, income inequality, or labor market conditions. Life expectancy for low-income individuals was positively correlated with the local area fraction of immigrants (r = 0.72, P < .001), fraction of college graduates (r = 0.42, P < .001), and government expenditures (r = 0.57, P < .001). In the United States between 2001 and 2014, higher income was associated with greater longevity, and differences in life expectancy across income groups increased over time. However, the association between life expectancy and income varied substantially across areas; differences in longevity across income groups decreased in some areas and increased in others. The differences in life expectancy were correlated with health behaviors and local area characteristics."
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what are brevinins
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Brevinins are an important antimicrobial peptide (AMP) family identified in the skin of
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[
"Toll-like receptor 4 (TLR4) is activated by lipopolysaccharide (LPS), a component of Gram-negative bacteria to induce production of pro-inflammatory mediators aiming at eradication of the bacteria. Dysregulation of the host responses to LPS can lead to a systemic inflammatory condition named sepsis. In a typical scenario, activation of TLR4 is preceded by binding of LPS to CD14 protein anchored in cholesterol- and sphingolipid-rich microdomains of the plasma membrane called rafts. CD14 then transfers the LPS to the TLR4/MD-2 complex which dimerizes and triggers MyD88- and TRIF-dependent production of pro-inflammatory cytokines and type I interferons. The TRIF-dependent signaling is linked with endocytosis of the activated TLR4, which is controlled by CD14. In addition to CD14, other raft proteins like Lyn tyrosine kinase of the Src family, acid sphingomyelinase, CD44, Hsp70, and CD36 participate in the TLR4 signaling triggered by LPS and non-microbial endogenous ligands. In this review, we summarize the current state of the knowledge on the involvement of rafts in TLR4 signaling, with an emphasis on how the raft proteins regulate the TLR4 signaling pathways. CD14-bearing rafts, and possibly CD36-rich rafts, are believed to be preferred sites of the assembly of a multimolecular complex which mediates the endocytosis of activated TLR4.",
"Six antimicrobial peptides, named gaegurins, were isolated from the skin of a Korean frog, Rana rugosa, and their amino acid sequences were determined by automated Edman degradation. All peptides contain two invariant cysteine residues, one at their C-terminus and the second at the seventh position from the C-terminus. The heptapeptides containing these two cysteine residues, which we designate 'Rana boxes', are conserved in the antimicrobial peptides derived from other Rana species. Each peptide manifested a broad spectrum of antimicrobial activity against Gram positive and Gram negative bacteria, fungi and protozoa with slightly different specific activities. All gaegurins manifest very little or no hemolytic activity. These properties provide the potential for application of these peptides to effective therapeutic agents for control of pathogenic microorganisms.",
"Bacterial infections are known to cause severe health-threatening conditions, including sepsis. All attempts to get this disease under control failed in the past, and especially in times of increasing antibiotic resistance, this leads to one of the most urgent medical challenges of our times. We designed a peptide to bind with high affinity to endotoxins, one of the most potent pathogenicity factors involved in triggering sepsis. The peptide Pep19-2.5 reveals high endotoxin neutralization efficiency in vitro, and here, we demonstrate its antiseptic/anti-inflammatory effects in vivo in the mouse models of endotoxemia, bacteremia, and cecal ligation and puncture, as well as in an ex vivo model of human tissue. Furthermore, we show that Pep19-2.5 can bind and neutralize not only endotoxins but also other bacterial pathogenicity factors, such as those from the Gram-positive bacterium Staphylococcus aureus. This broad neutralization efficiency and the additive action of the peptide with common antibiotics makes it an exceptionally appropriate drug candidate against bacterial sepsis and also offers multiple other medication opportunities.",
"Antimicrobial peptides form part of the innate immune response and play a vital role in host defense against pathogens. Here we report a new antimicrobial peptide belonging to the cathelicidin family, cathelicidin-MH (cath-MH), from the skin of Microhyla heymonsivogt frog. Cath-MH has a single α-helical structure in membrane-mimetic environments and is antimicrobial against fungi and bacteria, especially Gram-negative bacteria. In contrast to other cathelicidins, cath-MH suppresses coagulation by affecting the enzymatic activities of tissue plasminogen activator, plasmin, β-tryptase, elastase, thrombin, and chymase. Cath-MH protects against lipopolysaccharide (LPS)- and cecal ligation and puncture-induced sepsis, effectively ameliorating multiorgan pathology and inflammatory cytokine through its antimicrobial, LPS-neutralizing, coagulation suppressing effects as well as suppression of MAPK signaling. Taken together, these data suggest that cath-MH is an attractive candidate therapeutic agent for the treatment of septic shock.",
"Most patients with sepsis and acute lung injury require mechanical ventilation to improve oxygenation and facilitate organ repair. Mast cells are important in response to infection and resolution of tissue injury. Since tryptase secreted from mast cells has been associated with tissue fibrosis, we hypothesized that tryptase would be involved in the early development of ventilator-induced pulmonary fibrosis in a clinically relevant model of sepsis-induced lung injury. Prospective, randomized, controlled animal study using Sprague-Dawley rats. Sepsis was induced by cecal ligation and perforation. Animals were randomized to spontaneous breathing or two ventilatory strategies for 4 h: protective ventilation with tidal volume (VT) = 6 ml/kg plus 10 cmH₂O positive end-expiratory pressure (PEEP) or injurious ventilation with VT = 20 ml/kg plus 2 cmH₂O PEEP. Healthy, non-ventilated animals served as non-septic controls. We studied the following end points: histology, serum cytokine levels, hydroxyproline content, tryptase and proteinase-activated receptor-2 (PAR-2) protein level in lung homogenates, and tryptase and PAR-2 immunohistochemical localization in the lungs. All septic animals developed acute lung injury. Animals ventilated with high VT had a significant increase of pulmonary fibrosis, hydroxyproline content, tryptase and PAR-2 protein levels compared to septic controls (P <0.0001). However, protective ventilation attenuated sepsis-induced lung injury and decreased lung tryptase and PAR-2 protein levels. Immunohistochemical staining confirmed the presence of tryptase and PAR-2 in the lungs. Mechanical ventilation modified tryptase and PAR-2 in injured lungs. Increased levels of these proteins were associated with development of sepsis and ventilator-induced pulmonary fibrosis early in the course of sepsis-induced lung injury.",
"Sepsis is an exacerbated inflammatory reaction induced by severe infection. As important defensive molecules in innate immunity, several AMPs are reported to prevent septic shock. In this study, we characterized a novel cathelicidin, FM-CATH, from the frog skin of F. multistriata. FM-CATH was found to adopt an amphipathic α-helix structural in membrane-mimetic environments and possess favorable antimicrobial effects against bacteria and fungus. In addition, it triggered the agglutination of bacteria. It could also strongly bind to LPS and LTA. Additionally, FM-CATH affected the enzymatic activities of thrombin, plasmin, β-tryptase, and tPA, leading to coagulation inhibition in vitro and in vivo. Finally, we observed that FM-CATH improved survival rate and inhibited pathological alteration, bacterial count, serum biochemistry, and pro-inflammatory cytokine expression in the cecal ligation and puncture-induced sepsis mice. Taken together, these findings suggest that FM-CATH might be served as a promising agent for the treatment of sepsis.",
"Host defence peptides (HDPs) are evolutionarily conserved components of innate immunity. Human HDPs, produced by a variety of immune cells of hematopoietic and epithelial origin, are generally grouped into two families: beta structured defensins and variably-structured cathelicidins. We report the characterization of a very promising cryptic human HDP, here called GVF27, identified in 11-hydroxysteroid dehydrogenase-1 β-like protein. Conformational analysis of GVF27 and its propensity to bind endotoxins were performed by NMR, Circular Dichroism, Fluorescence and Dynamic Light Scattering experiments. Crystal violet and WST-1 assays, ATP leakage measurement and colony counting procedures were used to investigate antimicrobial, anti-biofilm, cytotoxicity and hemolytic activities. Anti-inflammatory properties were evaluated by ELISA. GVF27 possesses significant antibacterial properties on planktonic cells and sessile bacteria forming biofilm, as well as promising dose dependent abilities to inhibit attachment or eradicate existing mature biofilm. It is unstructured in aqueous buffer, whereas it tends to assume a helical conformation in mimic membrane environments as well as it is able to bind lipopolysaccharide (LPS) and lipoteichoic acid (LTA). Notably it is not toxic towards human and murine cell lines and triggers a significant innate immune response by attenuating expression levels of pro-inflammatory interleukins and release of nitric oxide in LPS induced macrophages. Human GVF27 may offer significant advantages as leads for the design of human-specific therapeutics. Human cryptic host defence peptides are naturally no immunogenic and for this they are a real alternative for solving the lack of effective antibiotics to control bacterial infections."
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Immune-related adverse events (irAEs) in emerging immunotherapy
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Emerging immunotherapeutic agents, including immune checkpoint inhibitors targeting cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), programmed cell death protein 1 (PD-1), and programmed cell death protein ligand 1 (PD-L1), have revolutionized cancer treatment. The first immune checkpoint inhibitor (ICI) ipilimumab, an anti-CTLA-4, was approved in 2011. Since then, the US Food and Drug Administration (FDA) has approved more than half a dozen immune checkpoint inhibitors to treat various malignancies. These agents are part of a broader class of chemotherapy agents termed immunotherapy, which selectively target different steps in the immune response cascade to upregulate the body's normal response to cancer. While the effects of traditional chemotherapy are well known, the toxicity profile of emerging immune therapies is not fully elucidated. They have been associated with atypical side effects labeled collectively as immune-related adverse events (irAEs).
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[
"Oral mucositis is a clinically important and sometimes dose-limiting complication of cancer therapy. Mucositis lesions can be painful, affect nutrition and quality of life, and have a significant economic impact. The pathogenesis of oral mucositis is multifactorial and complex. This review discusses the morbidity, economic impact, pathogenesis and clinical course of mucositis. Current clinical management of oral mucositis is largely focused on palliative measures such as pain management, nutritional support and maintenance of good oral hygiene. However, several promising therapeutic agents are in various stages of clinical development for the management of oral mucositis. These agents are discussed in the context of recently updated evidence-based clinical management guidelines.",
"Antibacterial prophylaxis with quinolone antibiotics has resulted in an increase in streptococcal infections among bone marrow transplantation (BMT) recipients with myelosuppression. Oral ulceration (mucositis), which frequently occurs as a consequence of chemotherapy, has been implicated as a significant portal of entry for streptococci. The objectives of this study were to confirm the correlation between mucositis and streptococcal bacteremia, determine the risk associated with this correlation, and evaluate the impact of mucositis and streptococcal bacteremia on hospital course and costs associated with autologous BMT. This was a retrospective, case-control study in which the charts of autologous BMT recipients treated for hematologic malignancies between 1990 and 1996 were reviewed. Twenty-four patients were identified who met the criteria of autologous BMT; their blood cultures confirmed (x2) alpha-hemolytic streptococcal sepsis. A control group of 45 without positive cultures was matched by gender, age, diagnosis, and treatment to the study group. The results confirm that ulcerative mucositis is a significant risk factor for alpha-hemolytic streptococcal bacteremia among autologous BMT patients. Of the 24 patients with bacteremia, 15 of 24 (62%) had ulcerative mucositis, compared with 16 of 45 (36%) of patients in the control population (P < 0.05). Patients with ulcerative mucositis were found to be three times as likely to develop alpha-hemolytic streptococcal bacteremia as those without ulcerative mucositis (odds ratio=3.02). Both independently and as a cofactor associated with bacteremia, mucositis adversely affected the length of hospital stay (LOS). Of all the patients studied, those with oral ulcerations had a LOS of 34 days, compared with 29 days for patients without oral ulcerations (P < 0.05). Of patients in the study group, those with oral ulcerations stayed in the hospital 6 days longer than patients without oral ulcerations (40 days vs. 34 days, P < 0.05). Oral ulcerative mucositis is a significant, common, and important risk factor for alpha-hemolytic streptococcal bacteremia in BMT recipients with myelosuppression; it results in longer hospital stay and increased costs.",
"Recent advances in the immunotherapeutic treatment of cancer have led to the development of multiple new directed therapies including monoclonal antibodies that block the immune checkpoint T-cell receptor programmed death 1 (PD-1) and the PD-1 ligand, programmed death ligand 1 (PD-L1). Various immune-related toxicities have been associated with these drugs including, most commonly, skin rashes. Five cases of lichenoid dermatitis, including one case of lichenoid mucositis and one case of lichen sclerosus, associated with anti-PD-L1 and anti-PD1 therapy were compared with three biopsies of non-drug-related lichen planus (LP) and three lichen planus-like keratoses (LPLK) used as controls. Histopathologic and immunophenotypic analysis of these lichenoid lesions demonstrated significantly greater histiocytic infiltrates than observed in control lichenoid reactions (p = 0.0134). We also observed increased spongiosis and epidermal necrosis. No significant differences were seen in expression of CD3, CD4:CD8, CD20, PD-1, CD25, Foxp3, CXCL13 and PD-L1 expression. These findings expand the literature of immune-related toxicities of PD-L1 and PD-1 blockade to include lichenoid dermatitis and lichenoid mucositis. Of note, these cutaneous side effects were amenable to topical treatment, without the need for medication dose reduction or discontinuation.",
"Oral sequelae of high-dose chemotherapy, including conditioning chemotherapy for hematopoietic stem cell transplantation, can lead to significant morbidity including potentially serious systemic infectious complications. These complications reduce the quality of life, are associated with higher treatment and supportive care costs, and can affect overall outcome of cancer therapy. Pretreatment oral assessment and intervention followed by the provision of supportive oral care during and after cancer therapy can reduce at least some of the adverse impact of oral complications. A definitive supportive oral care program is strongly recommended for patients scheduled for high-dose chemotherapeutic regimens. This article reviews recent insights into the pathogenesis of common oral problems in patients treated with high-dose chemotherapy regimens and discusses a multidisciplinary approach to management of these complications.",
"Oral complications in patients being treated for malignancies that were not in the head and neck were studied. Age, type of therapy, and type of malignancy were factors related to the prevalence of oral complications. Mucosal ulcerations, xerostomia, and bacterial and fungal infections were the most frequently encountered oral problems. The frequency of oral complications in these patients indicates the need for an awareness and involvement of dental practitioners in their management.",
"To identify predictors of oral mucositis and gastrointestinal toxicity after high-dose therapy. Mucositis and gastrointestinal toxicity were prospectively evaluated in 202 recipients of high-dose therapy and autologous or allogeneic stem-cell rescue. Of 10 outcome variables, three were selected as end points: the peak value for the University of Nebraska Oral Assessment Score (MUCPEAK), the duration of parenteral nutritional support, and the peak daily output of diarrhea. Potential covariates included patient age, sex, diagnosis, treatment protocol, transplantation type, stem-cell source, and rate of neutrophil recovery. The three selected end points were also examined for correlation with blood infections and transplant-related mortality. A diagnosis of leukemia, use of total body irradiation, allogeneic transplantation, and delayed neutrophil recovery were associated with increased oral mucositis and longer parenteral nutritional support. No factors were associated with diarrhea. Also, moderate to severe oral mucositis (MUCPEAK > or = 18 on a scale of 8 to 24) was correlated with blood infections and transplant-related mortality: 60% of patients with MUCPEAK > or = 18 had positive blood cultures versus 30% of patients with MUCPEAK less than 18 (P =.001); 24% of patients with MUCPEAK > or = 8 died during the transplantation procedure versus 4% of patients with MUCPEAK less than 18 (P =.001). Gastrointestinal toxicity is a major cause of transplant-related morbidity and mortality, emphasizing the need for corrective strategies. The peak oral mucositis score and the duration of parenteral nutritional support are useful indices of gastrointestinal toxicity because these end points are correlated with clinically significant events, including blood infections and treatment-related mortality.",
"Use of the low-energy helium-neon laser (LEL) appears to be a simple atraumatic technique for the prevention and treatment of mucositis of various origins. Preliminary findings, and significant results obtained for chemotherapy-induced mucositis in a previous phase III study, prompted a randomized multicenter double-blind trial to evaluate LEL in the prevention of acute radiation-induced stomatitis. Irradiation by LEL corresponds to local application of a high-photon-density monochromatic light source. Activation of epithelial healing for LEL-treated surfaces, the most commonly recognized effect, has been confirmed by numerous in vitro studies. The mechanism of action at a molecular and enzymatic level is presently being studied. From September 1994 to March 1998, 30 patients were randomized. Technical specification: 60 mW (25 mW at Reims, 1 patient), He-Ne, wavelength 632.8 nm. The trial was open to patients with carcinoma of the oropharynx, hypopharynx and oral cavity, treated by radiotherapy alone (65 Gy at a rate of 2 Gy/fraction, 5 fractions per week) without prior surgery or concomitant chemotherapy. The malignant tumor had to be located outside the tested laser application areas (9 points): posterior third of the internal surfaces of the cheeks, soft palate and anterior tonsillar pillars. Patients were randomized to LEL or placebo light treatment, starting on the first day of radiotherapy and before each session. The treatment time (t) for each application point was given by the equation : t(s)= energy (J/cm2) x surface (cm2)/Power (W). Objective assessment of the degree of mucositis was recorded weekly by a physician blinded to the type of treatment, using the WHO scale for grading of mucositis and a segmented visual analogue scale for pain evaluation. Protocol feasibility and compliance were excellent. Grade 3 mucositis occured with a frequency of 35.2% without LEL and of 7.6% with LEL (P<0.01). The frequency of \"severe pain\" (grade 3) was 23.8% without LEL, falling to 1.9% with LEL (P<0.05). Pain relief was significantly reduced throughout the treatment period (weeks 2-7). LEL therapy is capable of reducing the severity and duration of oral mucositis associated with radiation therapy. In addition, there is a tremendous potential for using LEL in combined treatment protocols utilizing concomitant chemotherapy and radiotherapy."
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Opsin 3 is present in enteric neurons in the muscularis propria of the murine colon
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Opsin photoreceptors outside of the central nervous system have been shown to mediate smooth muscle photorelaxation in several organs. We hypothesized that opsin receptor activation in the colon would have a similar effect and influence colonic motility. We detected Opsin 3 (OPN3) protein expression in the colonic wall and demonstrated that OPN3 was present in enteric neurons in the muscularis propria of the murine colon. Precontracted murine colon segments demonstrated blue light (BL) -mediated relaxation
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"The spontaneous colonic migrating motor complex (CMMC) is a cyclical contractile and electrical event that is the primary motor pattern underlying fecal pellet propulsion along the murine colon. We have combined Ca(2+) imaging with immunohistochemistry to determine the role of different classes of myenteric neurons during the CMMC. Between CMMCs, myenteric neurons usually displayed ongoing but uncoordinated activity. Stroking the mucosa at the oral or anal end of the colon resulted in a CMMC (latency: 6 to 10 s; duration: 28 s) that consisted of prolonged increases in activity in many myenteric neurons that was correlated to Ca(2+) transients in and displacement of the muscle. These neurons were likely excitatory motor neurons. Activity in individual neurons during the CMMC was similar regardless of whether the CMMC occurred spontaneously or was evoked by anal or oral mucosal stimulation. This suggests that convergent interneuronal pathways exist which generate CMMCs. Interestingly, Ca(2+) transients in a subset of NOS +ve neurons were substantially reduced during the CMMC. These neurons are likely to be inhibitory motor neurons that reduce their activity during a complex (disinhibition) to allow full excitation of the muscle. Local stimulation of the mucosa evoked synchronized Ca(2+) transients in Dogiel Type II (mitotracker/calbindin-positive) neurons after a short delay (1-2 s), indicating they were the sensory neurons underlying the CMMC. These local responses were observed in hexamethonium, but were blocked by ondansetron (5-HT(3) antagonist), suggesting Dogiel Type II neurons were activated by 5-HT release from enterochromaffin cells in the mucosa. In fact, removal of the mucosa yielded no spontaneous CMMCs, although many neurons (NOS +ve and NOS ve) exhibited ongoing activity, including Dogiel Type II neurons. These results suggest that spontaneous or evoked 5-HT release from the mucosa is necessary for the activation of Dogiel Type II neurons that generate CMMCs.",
"1. Intracellular microelectrodes have been used to record the electrical activity of smooth muscle cells of the circular layer from full length strips of mouse colon in vitro. The membrane potential was unstable and showed slow depolarizations (mean amplitude, 10.9 mV; mean frequency, 0.008 Hz; mean duration, 56.4 s). 2. A variable number (mean fifty-six) of rapid oscillations in membrane potential (mean amplitude, 10.2 mV) with a frequency of approximately 2 Hz and a duration of approximately 400 ms were superimposed on each slow depolarization. Occasionally, action potentials arose from the rapid oscillations. The action potentials, but neither the slow depolarizations nor the rapid oscillations, were abolished by 1.0 microM-nifedipine. 3. The majority of the slow depolarizations and the associated rapid oscillations migrated aborally along the colon at a velocity of between 0.5 and 1.5 mm s-1; in the distal colon the slow depolarization was often preceded by a small hyperpolarization. 4. During the rising and plateau phase of the slow depolarization the amplitude of electronic potentials was decreased. Hyperpolarization induced by passing current during the slow depolarization increased the amplitude of the rapid oscillations. 5. Transmural electrical stimulation (single pulses) in the presence of nifedipine evoked (1 mm anal to the stimulating electrodes) an inhibitory junction potential which was sometimes preceded by an excitatory junction potential. The amplitude, of the evoked inhibitory junction potential was decreased during the rising and plateau phase of the slow depolarization. 6. The slow depolarization and the rapid oscillations were abolished by hexamethonium (500 microM), morphine (1-10 microM) and tetrodotoxin (3.1 microM). Atropine (3.5 microM) abolished the rapid oscillations and reduced the amplitude of the slow depolarization. 7. Atropine (3.5 microM) and morphine (10 microM) abolished the evoked excitatory junction potential whilst tetrodotoxin (3.1 microM) abolished both the excitatory and the inhibitory junction potential. 8. It is suggested that the migrating depolarization and accompanying oscillations, which are neurogenic in origin, represent the electrical correlate in the circular muscle layer of the migrating colonic motor complex which has been associated with the propulsion of faecal pellets along the colon.",
"Spontaneous excitability and contractions of colonic smooth muscle cells (SMCs) are normally suppressed by inputs from inhibitory motor neurons, a behavior known as tonic inhibition. The post-junctional cell(s) mediating tonic inhibition have not been elucidated. We investigated the post-junctional cells mediating tonic inhibition in the proximal colon and whether tonic inhibition results from suppression of the activity of Ano1 channels, which are expressed exclusively in interstitial cells of Cajal (ICC). We found that tetrodotoxin (TTX), an inhibitor of nitric oxide (NO) synthesis, L-NNA, and an inhibitor of soluble guanylyl cyclase, ODQ, greatly enhanced colonic contractions. Ano1 antagonists, benzbromarone and Ani9 inhibited the effects of TTX, L-NNA and ODQ. Ano1 channels are activated by Ca2+ release from the endoplasmic reticulum (ER) in ICC, and blocking Ca2+ release with a SERCA inhibitor (thapsigargin) or a store-operated Ca2+ entry blocker (GSK 7975 A) reversed the effects of TTX, L-NNA and ODQ. Ca2+ imaging revealed that TTX, L-NNA and ODQ increased Ca2+ transient firing in colonic ICC. Our results suggest that tonic inhibition in the proximal colon occurs through suppression of Ca2+ release events in ICC. Suppression of Ca2+ release in ICC limits the open probability of Ano1 channels, reducing the excitability of electrically-coupled SMCs.",
"The colonic migrating motor complex (CMMC) is a major pattern of motility that is entirely generated and organized by the enteric nervous system. We have previously demonstrated that the Nav1.9 channel underlies a tetrodotoxin-resistant sodium current which modulates the excitability of enteric neurons. The aim of this study was to observe the effect of loss of the Nav1.9 channel in enteric neurons on mouse colonic motility in vitro. The mechanical activity of the circular muscle was simultaneously recorded from three sites, namely, proximal, mid- and distal, along the whole colon of male, age-matched wild-type and Nav1.9 null mice. Spontaneous CMMCs were observed in all preparations. The mean frequency of CMMCs was significantly higher in the Nav1.9 null mice (one every 2.87 ± 0.1 min compared to one every 3.96 ± 0.23 min in the wild type). The mean duration of CMMCs was shorter and the mean area-under-contraction was larger in the Nav1.9 null mice compared to the wild type. In addition, CMMCs propagated preferentially in an aboral direction in the Nav1.9 null mice. Our study demonstrates that CMMCs do occur in mice lacking the Nav1.9 channel, but their characteristics are significantly different from controls. Up to now, the Nav1.9 channel was mainly associated with nociceptive neurons and involved in their hyperexcitability after inflammation. Our result shows for the first time a role for the Nav1.9 channel in a complex colonic motor pattern.",
"5-HT3 antagonists, such as ondansetron (Zofran), retard colonic transit and provide effective relief of symptoms of chronic diarrhea and diarrhea-predominant irritable bowel syndrome (IBS), but the mechanism by which ondansetron retards transit is unclear. What is clear is that the frequency of colonic migrating motor complexes (CMMCs) is reduced by ondansetron, which could account for reduced transit. Our aim was to determine whether an acute depletion of 5-HT from enteric neurons would inhibit spontaneous CMMCs; and determine whether the sensitivity of ondansetron to reduce CMMC frequency would change in a 5-HT-depleted preparation. Mice were injected with reserpine, 24 h prior to euthanasia to deplete neuronally synthesized 5-HT. Mechanical recordings were made from proximal and mid-distal regions of isolated whole mouse colon. Immunohistochemical staining for 5-HT was used to detect neuronal 5-HT. Reserpine depleted all detectable 5-HT from enteric nerves. In whole colons, with mucosa and submucosal plexus removed, the frequency and amplitude of spontaneous CMMCs was not different between groups treated with or without reserpine. Surprisingly, in mucosa and submucosal plexus-free preparations, ondansetron was equally or significantly more effective at inhibiting CMMC frequency compared with control preparations (containing 5-HT). Reserpine pretreatment had no effect on the sensitivity of ondansetron to inhibit CMMCs. Endogenous 5-HT in enteric neurons (or the mucosa) is not required for the spontaneous generation or propagation of CMMCs. Furthermore, the primary mechanism by which ondansetron inhibits CMMC frequency is not mediated via the mucosa, submucosal plexus or 5-HT in myenteric neurons.",
"Intracellular recordings have been made in vitro from the myenteric neurons of the distal colon of normal littermates of the piebald-lethal mouse. Out of a total of 90 neurons, 82 were classified as S/type 1 cells and 8 as AH/type 2 cells. Seventy-eight out of 82 S cells showed spontaneous fast excitatory postsynaptic potentials (EPSPs) sensitive to d-tubocurarine (dTC, 280 microM), and 22 S cells showed spontaneous action potentials (APs). Six S cells and 1 AH cell showed spontaneous nonnicotinic slow depolarizations associated with an increase in the input resistance of the cells; during the spontaneous slow depolarization in the S cells there was an increase in the frequency of nicotinic fast EPSPs and APs. Three S cells showed spontaneously occurring regular oscillations of the membrane potential (approximately mV in amplitude and approximately 4/min). Transmural nerve stimulation produced fast EPSPs with a wide range of latencies (3 ms to 20 s) in S cells; the fast EPSPs were blocked by dTC (280 microM) or solutions containing low Ca2+ (0.25 mM) and high Mg2+ (12 mM) but not by atropine (ATR, 14 microM). Single or repetitive transmural stimulation produced slow EPSPs in 24 S cells and 3 AH cells; these were not blocked by dTC (280 microM) nor ATR (14 microM). During the slow EPSPs there was an increase in the input resistance of the cells. In those S cells that showed slow EPSPs there were many long-latency fast EPSPs; long-latency fast EPSPs were also observed in 11 other S cells that did not show a slow EPSP following repetitive transmural nerve stimulation. Long-latency fast EPSPs may be related to the firing of other neurons during their slow EPSPs. The myenteric neurons in the mouse colon have similar properties to the myenteric neurons in the guinea pig small intestine. However, the colonic myenteric neurons show more ongoing synaptic activity and more prolonged activity after nerve stimulation than myenteric neurons in the guinea pig small intestine. This activity may be due to regional differences, species differences, or preparation differences (in this study the myenteric plexus was adherent to the underlying circular muscle layer).",
"Understanding the role of voltage-gated sodium channels in nociception may provide important insights into pain mechanisms. Voltage-gated sodium channels are critically important for electrogenesis and nerve impulse conduction, and a target for important clinically relevant analgesics such as lidocaine. Furthermore, within the last decade studies have shown that certain sodium channel isoforms are predominantly expressed in peripheral sensory neurons associated with pain sensation, and that the expression and functional properties of voltage-gated sodium channels in peripheral sensory neurons can be dynamically regulated following axonal injury or peripheral inflammation. These data suggest that specific voltage-gated sodium channels may play crucial roles in nociception. Experiments with transgenic mice lines have clearly implicated Na(v)1.7, Na(v)1.8 and Na(v)1.9 in inflammatory, and possibly neuropathic, pain. However the most convincing and perhaps most exciting results regarding the role of voltage-gated sodium channels have come out recently from studies on human inherited disorders of nociception. Point mutations in Na(v)1.7 have been identified in patients with two distinct autosomal dominant severe chronic pain syndromes. Electrophysiological experiments indicate that these pain-associated mutations cause small yet significant changes in the gating properties of voltage-gated sodium channels that are likely to contribute substantially to the development of chronic pain. Equally exciting, recent studies indicate that recessive mutations in Na(v)1.7 that eliminate functional current can result in an apparent complete, and possibly specific, indifference to pain in humans, suggesting that isoform specific blockers could be very effective in treating pain. In this review we will examine what is known about the roles of voltage-gated sodium channels in nociception."
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Ephrin-A2 and Ephrin-A5 are important for lens morphometrics, nucleus formation, and refractive index
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Our studies in mouse eye lenses demonstrate that ephrin-A5 and EphA2 are needed for normal epithelial cells and lens transparency. We sought to determine whether EphA2 and ephrin-A5 are important for lens morphometrics, nucleus formation, and refractive index.
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"The retina sends spatially ordered visual information to the superior colliculus (SC) directly and indirectly via the thalamus and primary visual cortex (V1). Gradients of Ephs and ephrins are present in all of these regions, and have been shown to be involved in establishing topography of at least some of these interconnected visual pathways. Studies in ephrin-A knockout mice show that abnormal retinotectal termination zones (TZs) are present in a majority of mice lacking (-/-) ephrin-A2 (57%), and ephrin-A2 and -A5 (89%). A similar but seemingly less disordered pattern is detected in the retina-to-dorsal lateral geniculate nucleus (dLGN) and dLGN-to-V1 projections. Here we analyse the dLGN-to-V1 and V1-to-SC projections in ephrin-A(-/-) mice to determine the extent to which topographic errors are transmitted across synaptic relays. Fluorescent tracers were injected into V1 of wild-type (WT), ephrin-A2(-/-) or ephrin-A2A5(-/-) mice. We examined the number, location and size of anterograde TZs in SC, and mapped the distribution of retrogradely labelled neurons in dLGN. Compared with WT and ephrin-A2(-/-) mice, the volume of individual TZs in the SC was smaller in ephrin-A2A5(-/-) mice (P = 0.002). Single V1 injections labelled two foci of dLGN neurons in 70%, and two SC TZs in 80% of ephrin-A2A5(-/-) mice. Abnormalities in one or other of the projections were detected in 10% of ephrin-A2(-/-) mice. Importantly, there was no consistent correspondence between the organization of geniculocortical and corticotectal projections in either genotype, suggesting a role for ephrin-As in maintaining topographic organization in register across multiple interconnected central visual pathways.",
"Ephrin-A5 (AL-1/RAGS), a ligand for Eph receptor tyrosine kinases, repels retinal axons in vitro and has a graded expression in the superior colliculus (SC), the major midbrain target of retinal ganglion cells. These properties implicate ephrin-A5 in the formation of topographic maps, a fundamental organizational feature of the nervous system. To test this hypothesis, we generated mice lacking ephrin-A5. The majority of ephrin-A5-/- mice develop to adulthood, are morphologically intact, and have normal anterior-posterior patterning of the midbrain. However, within the SC, retinal axons establish and maintain dense arborizations at topographically incorrect sites that correlate with locations of low expression of the related ligand ephrin-A2. In addition, retinal axons transiently overshoot the SC and extend aberrantly into the inferior colliculus (IC). This defect is consistent with the high level of ephrin-A5 expression in the IC and the finding that retinal axon growth on membranes from wild-type IC is inhibited relative to that on membranes from ephrin-A5-/- IC. These findings show that ephrin-A5 is required for the proper guidance and mapping of retinal axons in the mammalian midbrain.",
"To identify the disease-causing mutation in a five-generation Chinese family affected with bilateral congenital nuclear cataract. Linkage analysis was performed for the known candidate genes and whole-exome sequencing was used in two affected family members to screen for potential genetic mutations; Sanger sequencing was used to verify the mutations throughout family. A novel beaded filament structural protein 1 (BFSP1) gene missense mutation was identified. Direct sequencing revealed a heterozygous G>A transversion at c.1042 of the coding sequence in exon 7 of BFSP1 (c.1042G>A) in all affected members, which resulted in the substitution of a wild-type aspartate to an asparagine (D348N). This mutation was neither seen in unaffected family members nor in 200 unrelated people as controls. A novel mutation (c.1042G>A) at exon 7 of BFSP1, which creates a substitution of an aspartate to an asparagine (p.D348N) was identified to be associated with autosomal dominant congenital cataract in a Chinese family. This is the first report of autosomal dominant congenital cataract being associated with a mutation in BFSP1, highlighting the important role of BFSP1 for physiological lens function and optical properties.",
"Microphthalmia, anophthalmia and coloboma (MAC) are distinct phenotypes that represent a continuum of structural developmental eye defects. In severe bilateral cases (anophthalmia or severe microphthalmia) the genetic cause is now identifiable in approximately 80 percent of cases, with de novo heterozygous loss-of-function mutations in SOX2 or OTX2 being the most common. The genetic cause of other forms of MAC, in particular isolated coloboma, remains unknown in the majority of cases. This review will focus on MAC phenotypes that are associated with mutation of the genes SOX2, OTX2, PAX6, STRA6, ALDH1A3, RARB, VSX2, RAX, FOXE3, BMP4, BMP7, GDF3, GDF6, ABCB6, ATOH7, C12orf57, TENM3 (ODZ3), and VAX1. Recently reported mutation of the SALL2 and YAP1 genes are discussed in brief. Clinical and genetic features were reviewed in a total of 283 unrelated MAC cases or families that were mutation-positive from these 20 genes. Both the relative frequency of mutations in MAC cohort screens and the level of confidence in the assignment of disease-causing status were evaluated for each gene.",
"Advances in genetic technology and analytical algorithms have greatly accelerated elucidation of the genetic contribution to cataractogenesis. Currently, 27 isolated or primary cataract loci have been identified by linkage analysis or mutational screening, and 20 are associated with specific genes. These are beginning to provide a framework for thinking of congenital cataracts. In addition to clues provided by the study of congenital and childhood cataracts, new experimental approaches to age-related cataracts are beginning to provide insights into its genetic origins.",
"Fine focusing of light by the eye lens onto the retina relies on the ability of the lens to change shape during the process of accommodation. Little is known about the cellular structures that regulate elasticity and resilience. We tested whether Eph-ephrin signaling is involved in lens biomechanical properties. We used confocal microscopy and tissue mechanical testing to examine mouse lenses with genetic disruption of EphA2 or ephrin-A5. Confocal imaging revealed misalignment of the suture between each shell of newly added fiber cells in knockout lenses. Despite having disordered sutures, loss of EphA2 or ephrin-A5 did not affect lens stiffness. Surprisingly, knockout lenses were more resilient and recovered almost completely after load removal. Confocal microscopy and quantitative image analysis from live lenses before, during, and after compression revealed that knockout lenses had misaligned Y-sutures, leading to a change in force distribution during compression. Knockout lenses displayed decreased separation of fiber cell tips at the anterior suture at high loads and had more complete recovery after load removal, which leads to improved whole-lens resiliency. EphA2 and ephrin-A5 are needed for normal patterning of fiber cell tips and the formation of a well-aligned Y-suture with fiber tips stacked on top of previous generations of fiber cells. The misalignment of lens sutures leads to increased resilience after compression. The data suggest that alignment of the Y-suture may constrain the overall elasticity and resilience of the lens.",
"Interlocking membrane domains are specialized membrane interdigitations in the form of ball-and-sockets and protrusions between lens fibre cells of all species. They are believed to play a key role in maintaining fibre-fibre stability and are therefore, important for normal lens function. Here we report the specific association of the clathrin/AP-2 adaptor complex and the branching F-actin network with the development of interlocking domains in rats and several other species. By thin-section electron microscopy we consistently observed a layer of distinct coating (approximately 25-nm thick) on the concave membrane surface of small and intermediate-sized developing interlocking domains. These membrane coats remarkably resembled the clathrin-coat of endocytic vesicles in which clathrin and the AP-2 adaptor are involved in the induction of coated pit formation during receptor-mediated endocytosis. We hypothesize that the clathrin/AP-2 complex is directly involved in the induction of interlocking domains in fibre cells. By immunoconfocal microscopy, co-labelling of a dotted-pattern of clathrin and AP-2 adaptor antibodies was seen along the cortical fibre cells. Immunoblot analysis further confirmed that clathrin and AP-2 adaptor antibodies specifically stained a polypeptide band of 180 and 106kD, respectively, in the membrane fractions prepared separately from the outer and inner cortical fibres where interlocking domains are abundant but endocytic vesicles are absent. Immunoelectron microscopy showed that the clathrin antibody was localized along the interlocking membrane. In addition, branching actin filament networks were frequently observed within the cytoplasmic compartment of developing interlocking domains by TEM, in consistent with the findings by fluorescence and immunogold labelling of the F-actin antibody in the domains. These results demonstrate for the first time that the clathrin/AP-2 complex plays a new role for the formation of interlocking domains in lens fibre cells. Branching actin networks and possibly other cytoskeletal components are also associated with the development and maintenance of these interlocking domains. The coordinated 'pulling and pushing' actions generated by the clathrin/AP-2 complex and branching actin networks during interlocking domain formation are discussed.",
"Crystalline lenses are often simply described as inside-out stratified epithelial-like organs composed of uniform (hexagonal cross-section profiles) crescent-like cells, arranged end-to-end in concentric shells around a polar axis. In this manner, as light is transmitted through lenses, their highly ordered architecture contributes to transparency by effectively transforming the multicellular organ into a series of coaxial refractive surfaces. This review will attempt to demonstrate that such a description seriously understates the structural complexity that produces lenses of variable optical quality in different species as a function of development, growth, and age. Embryological development of the lens occurs in a similar manner in all species. However, the growth patterns and effects of aging on lens fibers varies significantly among species. The terminally differentiated fiber cells of all lenses are generally hexagonal in cross section and crescent shaped along their length. But, while the fibers of all lenses are arranged in both highly ordered radial cell columns and concentric growth shells, only avian lens fibers are meridian-like, extending from pole to pole. In all other species, two types of fibers defined by different shapes are continuously formed throughout life. The majority of fibers are s-shaped, with ends that do not extend to the poles. Rather, the ends of these fibers are arranged as latitudinal arc lengths within and between growth shells. The overlap of the ends of specifically defined groups of such fibers constitutes the lens suture branches. The location, number, and extent of suture branches within and between growth shells are important considerations in lens function because the shapes of fiber ends, unlike that along fiber length, are very irregular. Consequently, as light is transmitted through sutures, spherical aberration (i.e., focal length variation) is increased. The degree of focal length variability depends on the arrangement of suture branches within and between growth shells, and this architecture varies significantly between species. The lifelong production of additional fibers at the circumference of the lens, culminating in new growth shells, neither proceeds equally around the lens equator, nor features identical fibers formed around the equator. Suture formation commences in the inferonasal quadrant, and continues sequentially in the superotemporal, inferotemporal, and finally the superonasal quadrants. During this process, lens growth produces fibers of specifically defined length and shape as a function of their equatorial location.(ABSTRACT TRUNCATED AT 250 WORDS)",
"This study aimed to investigate the genetic effects underlying non-familial sporadic congenital cataract (SCC). We collected DNA samples from 74 patients with SCC and 20 patients with traumatic cataract (TC) in an age-matched group and performed genomic sequencing of 61 lens-related genes with target region capture and next-generation sequencing (NGS). The suspected SCC variants were validated with MassARRAY and Sanger sequencing. DNA samples from 103 healthy subjects were used as additional controls in the confirmation examination. By filtering against common variants in public databases and those associated with TC cases, we identified 23 SCC-specific variants in 17 genes from 19 patients, which were predicted to be functional. These mutations were further confirmed by examination of the 103 healthy controls. Among the mutated genes, CRYBB3 had the highest mutation frequency with mutations detected four times in four patients, followed by EPHA2, NHS, and WDR36, the mutation of which were detected two times in two patients. We observed that the four patients with CRYBB3 mutations had three different cataract phenotypes. From this study, we concluded the clinical and genetic heterogeneity of SCC. This is the first study to report broad spectrum genotyping for patients with SCC."
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Regulation of Gasdermin Pore Formation
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Gasdermins are proteins that can self-assemble into membrane channels (also known as pores). These pores can serve as conduits for the secretion of cytosolic molecules, with the most commonly studied being members of the interleukin-1 family of cytokines. However, gasdermin pore forming activities must be tightly regulated, as the channels that they form can lead to a lytic form of cell death known as pyroptosis. Recent studies have revealed multiple mechanisms that control gasdermin activities within cells and identified gasdermin proteins in organisms as diverse as bacteria, humans and yeast. In this Review, we discuss the molecular and cellular mechanisms that regulate gasdermin pore formation. These mechanisms of gasdermin regulation likely explain the flexibility of these proteins to display cell type specific (and potentially organism specific) functions.
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"Programmed cell death (PCD) in filamentous fungi prevents cytoplasmic mixing following fusion between conspecific genetically distinct individuals (allorecognition) and serves as a defense mechanism against mycoparasitism, genome exploitation, and deleterious cytoplasmic elements (i.e., senescence plasmids). Recently, we identified regulatorof cell death-1 (rcd-1), a gene controlling PCD in germinated asexual spores in the filamentous fungus Neurospora crassarcd-1 alleles are highly polymorphic and fall into two haplogroups in N. crassa populations. Coexpression of alleles from the two haplogroups, rcd-1-1 and rcd-1-2, is necessary and sufficient to trigger a cell death reaction. Here, we investigated the molecular bases of rcd-1-dependent cell death. Based on in silico analyses, we found that RCD-1 is a remote homolog of the N-terminal pore-forming domain of gasdermin, the executioner protein of a highly inflammatory cell death reaction termed pyroptosis, which plays a key role in mammalian innate immunity. We show that RCD-1 localizes to the cell periphery and that cellular localization of RCD-1 was correlated with conserved positively charged residues on predicted amphipathic α-helices, as shown for murine gasdermin-D. Similar to gasdermin, RCD-1 binds acidic phospholipids in vitro, notably, cardiolipin and phosphatidylserine, and interacts with liposomes containing such lipids. The RCD-1 incompatibility system was reconstituted in human 293T cells, where coexpression of incompatible rcd-1-1/rcd-1-2 alleles triggered pyroptotic-like cell death. Oligomers of RCD-1 were associated with the cell death reaction, further supporting the evolutionary relationship between gasdermin and rcd-1 This report documents an ancient transkingdom relationship of cell death execution modules involved in organismal defense.",
"The death of a cell is an inevitable part of its biology. During homeostasis, most cells die through apoptosis. If homeostasis is disturbed, cell death can switch to proinflammatory forms of death, such as necroptosis, pyroptosis, or NETosis. We demonstrate that the formation of neutrophil extracellular traps (NETs), a special form of neutrophil cell death that releases chromatin structures to the extracellular space, is dependent on gasdermin D (GSDMD). GSDMD is a pore-forming protein and an executor of pyroptosis. We screened a chemical library and found a small molecule based on the pyrazolo-oxazepine scaffold that efficiently blocks NET formation and GSDMD-mediated pyroptotic cell death in human cells. During NETosis, GSDMD is proteolytically activated by neutrophil proteases and, in turn, affects protease activation and nuclear expansion in a feed-forward loop. In addition to the central role of GSDMD in pyroptosis, we propose that GSDMD also plays an essential function in NETosis.",
"Protozoan infections are a serious global health problem. Natural killer (NK) cells and cytolytic T lymphocytes (CTLs) eliminate pathogen-infected cells by releasing cytolytic granule contents--granzyme (Gzm) proteases and the pore-forming perforin (PFN)--into the infected cell. However, these cytotoxic molecules do not kill intracellular parasites. CD8(+) CTLs protect against parasite infections in mice primarily by secreting interferon (IFN)-γ. However, human, but not rodent, cytotoxic granules contain the antimicrobial peptide granulysin (GNLY), which selectively destroys cholesterol-poor microbial membranes, and GNLY, PFN and Gzms rapidly kill intracellular bacteria. Here we show that GNLY delivers Gzms into three protozoan parasites (Trypanosoma cruzi, Toxoplasma gondii and Leishmania major), in which the Gzms generate superoxide and inactivate oxidative defense enzymes to kill the parasite. PFN delivers GNLY and Gzms into infected cells, and GNLY then delivers Gzms to the intracellular parasites. Killer cell-mediated parasite death, which we term 'microbe-programmed cell death' or 'microptosis', is caspase independent but resembles mammalian apoptosis, causing mitochondrial swelling, transmembrane potential dissipation, membrane blebbing, phosphatidylserine exposure, DNA damage and chromatin condensation. GNLY-transgenic mice are protected against infection by T. cruzi and T. gondii, and survive infections that are lethal to wild-type mice. Thus, GNLY-, PFN- and Gzm-mediated elimination of intracellular protozoan parasites is an unappreciated immune defense mechanism.",
"Pyroptosis is a form of lytic inflammatory cell death driven by inflammatory caspase-1, caspase-4, caspase-5 and caspase-11. These caspases cleave and activate the pore-forming protein gasdermin D (GSDMD) to induce membrane damage. By contrast, apoptosis is driven by apoptotic caspase-8 or caspase-9 and has traditionally been classified as an immunologically silent form of cell death. Emerging evidence suggests that therapeutics designed for cancer chemotherapy or inflammatory disorders such as SMAC mimetics, TAK1 inhibitors and BH3 mimetics promote caspase-8 or caspase-9-dependent inflammatory cell death and NLRP3 inflammasome activation. However, the mechanism by which caspase-8 or caspase-9 triggers cell lysis and NLRP3 activation is still undefined. Here, we demonstrate that during extrinsic apoptosis, caspase-1 and caspase-8 cleave GSDMD to promote lytic cell death. By engineering a novel Gsdmd D88A knock-in mouse, we further demonstrate that this proinflammatory function of caspase-8 is counteracted by caspase-3-dependent cleavage and inactivation of GSDMD at aspartate 88, and is essential to suppress GSDMD-dependent cell lysis during caspase-8-dependent apoptosis. Lastly, we provide evidence that channel-forming glycoprotein pannexin-1, but not GSDMD or GSDME promotes NLRP3 inflammasome activation during caspase-8 or caspase-9-dependent apoptosis.",
"Inflammatory caspases (caspases 1, 4, 5 and 11) are activated in response to microbial infection and danger signals. When activated, they cleave mouse and human gasdermin D (GSDMD) after Asp276 and Asp275, respectively, to generate an N-terminal cleavage product (GSDMD-NT) that triggers inflammatory death (pyroptosis) and release of inflammatory cytokines such as interleukin-1β. Cleavage removes the C-terminal fragment (GSDMD-CT), which is thought to fold back on GSDMD-NT to inhibit its activation. However, how GSDMD-NT causes cell death is unknown. Here we show that GSDMD-NT oligomerizes in membranes to form pores that are visible by electron microscopy. GSDMD-NT binds to phosphatidylinositol phosphates and phosphatidylserine (restricted to the cell membrane inner leaflet) and cardiolipin (present in the inner and outer leaflets of bacterial membranes). Mutation of four evolutionarily conserved basic residues blocks GSDMD-NT oligomerization, membrane binding, pore formation and pyroptosis. Because of its lipid-binding preferences, GSDMD-NT kills from within the cell, but does not harm neighbouring mammalian cells when it is released during pyroptosis. GSDMD-NT also kills cell-free bacteria in vitro and may have a direct bactericidal effect within the cytosol of host cells, but the importance of direct bacterial killing in controlling in vivo infection remains to be determined.",
"The epidermis forms a critical barrier that is maintained by orchestrated programs of proliferation, differentiation, and cell death. Gene mutations that disturb this turnover process may cause skin diseases. Human GASDERMIN A (GSDMA) is frequently silenced in gastric cancer cell lines and its overexpression has been reported to induce apoptosis. GSDMA has also been linked with airway hyperresponsiveness in genetic association studies. The function of GSDMA in the skin was deduced by dominant mutations in mouse gasdermin A3 (Gsdma3), which caused skin inflammation and hair loss. However, the mechanism for the autosomal dominance of Gsdma3 mutations and the mode of Gsdma3's action remain unanswered. We demonstrated a novel function of Gsdma3 in modulating mitochondrial oxidative stress. We showed that Gsdma3 is regulated by intramolecular fold-back inhibition, which is disrupted by dominant mutations in the C-terminal domain. The unmasked N-terminal domain of Gsdma3 associates with Hsp90 and is delivered to mitochondrial via mitochondrial importer receptor Tom70, where it interacts with the mitochondrial chaperone Trap1 and causes increased production of mitochondrial reactive oxygen species (ROS), dissipation of mitochondrial membrane potential, and mitochondrial permeability transition (MPT). Overexpression of the C-terminal domain of Gsdma3 as well as pharmacological interventions of mitochondrial translocation, ROS production, and MPT pore opening alleviate the cell death induced by Gsdma3 mutants. Our results indicate that the genetic mutations in the C-terminal domain of Gsdma3 are gain-of-function mutations which unmask the N-terminal functional domain of Gsdma3. Gsdma3 regulates mitochondrial oxidative stress through mitochondrial targeting. Since mitochondrial ROS has been shown to promote epidermal differentiation, we hypothesize that Gsdma3 regulates context-dependent response of keratinocytes to differentiation and cell death signals by impinging on mitochondria.",
"Cell death is a fundamental physiological process in all living organisms. Its roles extend from embryonic development, organ maintenance, and aging to the coordination of immune responses and autoimmunity. In recent years, our understanding of the mechanisms orchestrating cellular death and its consequences on immunity and homeostasis has increased substantially. Different modalities of what has become known as 'programmed cell death' have been described, and some key players in these processes have been identified. We have learned more about the intricacies that fine tune the activity of common players and ultimately shape the different types of cell death. These studies have highlighted the complex mechanisms tipping the balance between different cell fates. Here, we summarize the latest discoveries in the three most well understood modalities of cell death, namely, apoptosis, necroptosis, and pyroptosis, highlighting common and unique pathways and their effect on the surrounding cells and the organism as a whole."
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Exosomal miR-222 promotes hepatic stellate cell activation and promotes liver fibrosis in mouse Exosomal LO2 cells
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Exosomal miRNAs activates hepatic stellate cell (HSC) and promote fibrosis. miR-222 was found to be increased in hepatitis B virus (HBV)-infected hepatocytes, and ferroptosis was reported to ameliorate liver fibrosis (LF). Although miR-222 and ferroptosis have been implicated in LF, the association between miR-222 and ferroptosis and how they coordinate to regulate LF are still not explicit. This study investigates the roles of miR-222 and transferrin receptor (TFRC) in LF. Lipid reactive oxygen species (ROS) level was analyzed by flow cytometry. FerroOrange staining was used to measure intracellular iron level. Luciferase reporter assay was adopted to confirm the binding of miR-222 and TFRC. Real-time quantitative PCR and immunoblots were applied to analyze gene and protein expression. The results showed that supplementation of exosomes derived from HBV-infected LO2 cells remarkably enhanced LX-2 cell activation, evidenced by elevated hydroxyprolin (Hyp) secretion and α-SMA and COL1A2 expression. miR-222 was significantly increased in HBV-Exo. Overexpressing miR-222 upregulated cell viability, secretion of Hpy, and expression of α-SMA and COL1A2, which were all blocked by overexpression of TFRC. Further study showed that TFRC was a target of miR-222, and miR-222 promoted LX-2 cell activation through suppressing TFRC-induced ferroptosis in LX-2 cells. Exosomal miR-222 derived from HBV-infected hepatocytes promoted LF through inhibiting TFRC and TFRC-induced ferroptosis. This study emphasizes the significance of miR-222/TFRC axis in LF and suggests new insights in clinical decision making while treating LF. Exosomes derived from HBV-infected LO2 cells promote LX-2 cell activation and liver fibrosis in mouse Exosomal miR-222 derived from HBV-infected LO2 cells promotes LX-2 cell activation TFRC is a target of miR-222 and inhibits LX-2 cell activation induced by miR-222 miR-222 promotes LX-2 cell activation through inhibiting TFRC-induced ferroptosis.
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"Several recent studies demonstrated that microRNAs (miRNAs) are stably detectable in plasma/serum. We hypothesised that plasma miRNAs concentrations contributed to potential biomarkers in patients with oesophageal squamous cell carcinoma (ESCC). We selected three oncogenic miRNAs (miR-21, miR-184, miR-221) and one tumour suppressive miRNA (miR-375), which are frequently reported in squamous cell carcinoma, as candidate targets for this plasma miRNA assay. This study was divided into three steps: (1) Determination of appropriate plasma miRNAs in preliminary tests. (2) Evaluation of whether the plasma miRNA assays could monitor tumour dynamics. (3) Validation study on the clinical application of plasma miRNA assays in 50 ESCC patients and 20 healthy volunteers. (1) In preliminary tests, the plasma level of miR-21 was significantly higher (P=0.0218) and that of miR-375 (P=0.0052) was significantly lower in ESCC patients than controls. (2) The high plasma miR-21 levels reflected tumour levels in all cases (100%). The plasma level of miR-21 was significantly reduced in postoperative samples (P=0.0058). (3) On validation analysis, the plasma level of miR-21 tended to be higher in ESCC patients (P=0.0649), while that of miR-375 was significantly lower (P<0.0001) and the miR-21/miR-375 ratio was significantly higher (P<0.0001) in ESCC patients than in controls. The value of the area under the receiver-operating characteristic curve (AUC) was 0.816 for the miR-21/miR-375 ratio assay. Patients with a high plasma level of miR-21 tended to have greater vascular invasion (P=0.1554) and to show a high correlation with recurrence (P=0.0164). Detection of circulating miRNAs might provide new complementary tumour markers for ESCC.",
"Cancer-secreted microRNAs (miRNAs) are emerging mediators of cancer-host crosstalk. Here we show that miR-105, which is characteristically expressed and secreted by metastatic breast cancer cells, is a potent regulator of migration through targeting the tight junction protein ZO-1. In endothelial monolayers, exosome-mediated transfer of cancer-secreted miR-105 efficiently destroys tight junctions and the integrity of these natural barriers against metastasis. Overexpression of miR-105 in nonmetastatic cancer cells induces metastasis and vascular permeability in distant organs, whereas inhibition of miR-105 in highly metastatic tumors alleviates these effects. miR-105 can be detected in the circulation at the premetastatic stage, and its levels in the blood and tumor are associated with ZO-1 expression and metastatic progression in early-stage breast cancer.",
"Activated hepatic stellate cells (HSCs) are the major source of extracellular matrix in fibrosis and cirrhosis. In this study, we have investigated the role of hepatitis C virus (HCV) core protein induced immortalized human hepatocytes (IHH) on HSC growth. Preferential growth of IHH and apoptosis of activated human hepatic stellate cells (LX2) were observed upon coculture of these two cell types in a dual chamber or in the presence of conditioned medium (CM) from IHH. CM did not display a growth inhibitory role on other hepatic (Huh-7, HepG2, Hep3B and THLE) and non-hepatic (HeLa, MCF-7, and BHK) epithelial cells, indicating that the soluble mediator from IHH does not have a generalized effect on cell lines examined in our study. Further studies suggested that CM from IHH increased the expression of TRAIL receptors on LX2 cell surface, and induced apoptosis by a caspase dependent mechanism. Peptide mass fingerprinting of the purified soluble mediator from CM suggested that gelsolin fragments may play a role in apoptosis of LX2 cells. Taken together, our results suggested that a soluble mediator secreted from immortalized human hepatocytes plays an important role in hepatic stellate cell growth regulation.",
"Cells release into the extracellular environment diverse types of membrane vesicles of endosomal and plasma membrane origin called exosomes and microvesicles, respectively. These extracellular vesicles (EVs) represent an important mode of intercellular communication by serving as vehicles for transfer between cells of membrane and cytosolic proteins, lipids, and RNA. Deficiencies in our knowledge of the molecular mechanisms for EV formation and lack of methods to interfere with the packaging of cargo or with vesicle release, however, still hamper identification of their physiological relevance in vivo. In this review, we focus on the characterization of EVs and on currently proposed mechanisms for their formation, targeting, and function.",
"Liver fibrosis is the excessive accumulation of extracellular matrix proteins including collagen that occurs in most types of chronic liver diseases. Advanced liver fibrosis results in cirrhosis, liver failure, and portal hypertension and often requires liver transplantation. Our knowledge of the cellular and molecular mechanisms of liver fibrosis has greatly advanced. Activated hepatic stellate cells, portal fibroblasts, and myofibroblasts of bone marrow origin have been identified as major collagen-producing cells in the injured liver. These cells are activated by fibrogenic cytokines such as TGF-beta1, angiotensin II, and leptin. Reversibility of advanced liver fibrosis in patients has been recently documented, which has stimulated researchers to develop antifibrotic drugs. Emerging antifibrotic therapies are aimed at inhibiting the accumulation of fibrogenic cells and/or preventing the deposition of extracellular matrix proteins. Although many therapeutic interventions are effective in experimental models of liver fibrosis, their efficacy and safety in humans is unknown. This review summarizes recent progress in the study of the pathogenesis and diagnosis of liver fibrosis and discusses current antifibrotic strategies.",
"The role of the extracellular non-coding RNAs, particularly microRNAs present in tumor-derived extravesicles, has been intensively exploited in human cancer as a promising tool for diagnostic and prognostic purposes. Current knowledge on exosomes shows an important role not only as vehicles in the intercellular communication, but the transfer of their content can specifically modulate the surrounding microenvironment, leading to tumor development and progression and affecting therapy response. Based on this, much effort has focused on understanding the mechanisms behind the biology of exosomes and their closely interaction with non-coding RNAs as an efficient tool in tumor diagnostic and therapy. Here we summarize the current knowledge on extracellular and exosomes-enclosed non-coding RNAs, and their importance as potential biomarkers and mediators of intercellular communication in tumor biology.",
"Recent studies suggest both normal and cancerous cells secrete vesicles into the extracellular space. These extracellular vesicles (EVs) contain materials that mirror the genetic and proteomic content of the secreting cell. The identification of cancer-specific material in EVs isolated from the biofluids (e.g., serum, cerebrospinal fluid, urine) of cancer patients suggests EVs as an attractive platform for biomarker development. It is important to recognize that the EVs derived from clinical samples are likely highly heterogeneous in make-up and arose from diverse sets of biologic processes. This article aims to review the biologic processes that give rise to various types of EVs, including exosomes, microvesicles, retrovirus like particles, and apoptotic bodies. Clinical pertinence of these EVs to neuro-oncology will also be discussed."
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Emergent variants of SARS-CoV-2 and the efficacy of developed vaccines
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The emergence of SARS-CoV-2 variants may cause resistance at the immunity level against current vaccines. Some emergent new variants have increased transmissibility, infectivity, hospitalization, and mortality. Since the administration of the first SARS-CoV-2 vaccine to a human in March 2020, there is an ongoing global race against SARS-CoV-2 to control the current pandemic situation. Spike (S) glycoprotein of SARS-CoV-2 is the main target for current vaccine development, which can neutralize the infection. Companies and academic institutions have developed vaccines based on the S glycoprotein, as well as its antigenic domains and epitopes, which have been proven effective in generating neutralizing antibodies. The effectiveness of SARS-CoV-2 vaccines and other therapeutics developments are limited by the new emergent variants at the global level. We have discussed the emergent variants of SARS-CoV-2 on the efficacy of developed vaccines. Presently, most of the vaccines have been tremendously effective in severe diseases. However, there are still noteworthy challenges in certifying impartial vaccines; the stories of re-infections are generating more stressful conditions, and this needs further clinical evaluation.
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"The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants threatens progress toward control of the coronavirus disease 2019 (Covid-19) pandemic. In a phase 1-2 trial involving healthy adults, the NVX-CoV2373 nanoparticle vaccine had an acceptable safety profile and was associated with strong neutralizing-antibody and antigen-specific polyfunctional CD4+ T-cell responses. Evaluation of vaccine efficacy was needed in a setting of ongoing SARS-CoV-2 transmission. In this phase 2a-b trial in South Africa, we randomly assigned human immunodeficiency virus (HIV)-negative adults between the ages of 18 and 84 years or medically stable HIV-positive participants between the ages of 18 and 64 years in a 1:1 ratio to receive two doses of either the NVX-CoV2373 vaccine (5 μg of recombinant spike protein with 50 μg of Matrix-M1 adjuvant) or placebo. The primary end points were safety and vaccine efficacy against laboratory-confirmed symptomatic Covid-19 at 7 days or more after the second dose among participants without previous SARS-CoV-2 infection. Of 6324 participants who underwent screening, 4387 received at least one injection of vaccine or placebo. Approximately 30% of the participants were seropositive for SARS-CoV-2 at baseline. Among 2684 baseline seronegative participants (94% HIV-negative and 6% HIV-positive), predominantly mild-to-moderate Covid-19 developed in 15 participants in the vaccine group and in 29 in the placebo group (vaccine efficacy, 49.4%; 95% confidence interval [CI], 6.1 to 72.8). Vaccine efficacy among HIV-negative participants was 60.1% (95% CI, 19.9 to 80.1). Of 41 sequenced isolates, 38 (92.7%) were the B.1.351 variant. Post hoc vaccine efficacy against B.1.351 was 51.0% (95% CI, -0.6 to 76.2) among the HIV-negative participants. Preliminary local and systemic reactogenicity events were more common in the vaccine group; serious adverse events were rare in both groups. The NVX-CoV2373 vaccine was efficacious in preventing Covid-19, with higher vaccine efficacy observed among HIV-negative participants. Most infections were caused by the B.1.351 variant. (Funded by Novavax and the Bill and Melinda Gates Foundation; ClinicalTrials.gov number, NCT04533399.).",
"A vaccine against coronavirus disease 2019 (COVID-19) is urgently needed. To evaluate the safety and immunogenicity of an investigational inactivated whole-virus COVID-19 vaccine in China. In the phase 1 trial, 96 participants were assigned to 1 of the 3 dose groups (2.5, 5, and 10 μg/dose) and an aluminum hydroxide (alum) adjuvant-only group (n = 24 in each group), and received 3 intramuscular injections at days 0, 28, and 56. In the phase 2 trial, 224 adults were randomized to 5 μg/dose in 2 schedule groups (injections on days 0 and 14 [n = 84] vs alum only [n = 28], and days 0 and 21 [n = 84] vs alum only [n = 28]). Interim analysis of ongoing randomized, double-blind, placebo-controlled, phase 1 and 2 clinical trials to assess an inactivated COVID-19 vaccine. The trials were conducted in Henan Province, China, among 96 (phase 1) and 224 (phase 2) healthy adults aged between 18 and 59 years. Study enrollment began on April 12, 2020. The interim analysis was conducted on June 16, 2020, and updated on July 27, 2020. The primary safety outcome was the combined adverse reactions 7 days after each injection, and the primary immunogenicity outcome was neutralizing antibody response 14 days after the whole-course vaccination, which was measured by a 50% plaque reduction neutralization test against live severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Among 320 patients who were randomized (mean age, 42.8 years; 200 women [62.5%]), all completed the trial up to 28 days after the whole-course vaccination. The 7-day adverse reactions occurred in 3 (12.5%), 5 (20.8%), 4 (16.7%), and 6 (25.0%) patients in the alum only, low-dose, medium-dose, and high-dose groups, respectively, in the phase 1 trial; and in 5 (6.0%) and 4 (14.3%) patients who received injections on days 0 and 14 for vaccine and alum only, and 16 (19.0%) and 5 (17.9%) patients who received injections on days 0 and 21 for vaccine and alum only, respectively, in the phase 2 trial. The most common adverse reaction was injection site pain, followed by fever, which were mild and self-limiting; no serious adverse reactions were noted. The geometric mean titers of neutralizing antibodies in the low-, medium-, and high-dose groups at day 14 after 3 injections were 316 (95% CI, 218-457), 206 (95% CI, 123-343), and 297 (95% CI, 208-424), respectively, in the phase 1 trial, and were 121 (95% CI, 95-154) and 247 (95% CI, 176-345) at day 14 after 2 injections in participants receiving vaccine on days 0 and 14 and on days 0 and 21, respectively, in the phase 2 trial. There were no detectable antibody responses in all alum-only groups. In this interim report of the phase 1 and phase 2 trials of an inactivated COVID-19 vaccine, patients had a low rate of adverse reactions and demonstrated immunogenicity; the study is ongoing. Efficacy and longer-term adverse event assessment will require phase 3 trials. Chinese Clinical Trial Registry Identifier: ChiCTR2000031809.",
"As the global severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic expands, genomic epidemiology and whole genome sequencing are being used to investigate its transmission and evolution. Against the backdrop of the global emergence of \"variants of concern\" (VOCs) during December 2020 and an upsurge in a state in the western part of India since January 2021, whole genome sequencing and analysis of spike protein mutations using sequence and structural approaches were undertaken to identify possible new variants and gauge the fitness of the current circulating strains. Phylogenetic analysis revealed that newly identified lineages B.1.617.1 and B.1.617.2 were predominantly circulating. The signature mutations possessed by these strains were L452R, T478K, E484Q, D614G and P681R in the spike protein, including within the receptor-binding domain (RBD). Of these, the mutations at residue positions 452, 484 and 681 have been reported in other globally circulating lineages. The structural analysis of RBD mutations L452R, T478K and E484Q revealed that these may possibly result in increased ACE2 binding while P681R in the furin cleavage site could increase the rate of S1-S2 cleavage, resulting in better transmissibility. The two RBD mutations, L452R and E484Q, indicated decreased binding to select monoclonal antibodies (mAbs) and may affect their neutralization potential. Further in vitro/in vivo studies would help confirm the phenotypic changes of the mutant strains. Overall, the study revealed that the newly emerged variants were responsible for the second wave of COVID-19 in Maharashtra. Lineage B.1.617.2 has been designated as a VOC delta and B.1.617.1 as a variant of interest kappa, and they are being widely reported in the rest of the country as well as globally. Continuous monitoring of these and emerging variants in India is essential.",
"Severe Acute Respiratory Syndrome Coronavirus 2 is a novel strain of human beta-coronavirus that has produced over two million deaths and affected one hundred million individuals worldwide. As all the proposed drugs proved to be unstable, inducing side effects, the need to develop a vaccine crystallized in a short time. As a result, we searched the databases for articles in which the authors reported the efficacy and safety of the use of several vaccines vaccines by sex, age group, and frequency of adverse reactions. We identified a total of 19 relevant articles that were discussed throughout this manuscript. We concluded that from all eleven vaccines, three had an efficacy >90% (Pfizer-BioNTech (~95%), Moderna (~94%), and Sputnik V (~92%)) except for Oxford-AstraZeneca (~81%). However, Moderna, Sputnik V, and Oxford-AstraZeneca also alleviate severe adverse reactions, whereas in Pfizer-BioNTech this was not revealed. The remaining five (Convidicea (AD5-nCOV); Johnson & Johnson (Ad26.COV2.S); Sinopharm (BBIBP-CorV); Covaxin (BBV152), and Sinovac (CoronaVac)) were discussed based on their immunogenicity, and safety reported by the recipients since only phases 1 and 2 were conducted without clear evidence published regarding their efficacy. CoviVac and EpiVacCorona have just been approved, which is why no published article could be found. All adverse events reported following the administration of one of the four vaccines ranged from mild to moderate; limited exceptions in which the patients either developed severe forms or died, because most effects were dose-dependent. It can be concluded that aforementioned vaccines are efficient and safe, regardless of age and sex, being well-tolerated by the recipients.",
"A heterologous recombinant adenovirus (rAd)-based vaccine, Gam-COVID-Vac (Sputnik V), showed a good safety profile and induced strong humoral and cellular immune responses in participants in phase 1/2 clinical trials. Here, we report preliminary results on the efficacy and safety of Gam-COVID-Vac from the interim analysis of this phase 3 trial. We did a randomised, double-blind, placebo-controlled, phase 3 trial at 25 hospitals and polyclinics in Moscow, Russia. We included participants aged at least 18 years, with negative SARS-CoV-2 PCR and IgG and IgM tests, no infectious diseases in the 14 days before enrolment, and no other vaccinations in the 30 days before enrolment. Participants were randomly assigned (3:1) to receive vaccine or placebo, with stratification by age group. Investigators, participants, and all study staff were masked to group assignment. The vaccine was administered (0·5 mL/dose) intramuscularly in a prime-boost regimen: a 21-day interval between the first dose (rAd26) and the second dose (rAd5), both vectors carrying the gene for the full-length SARS-CoV-2 glycoprotein S. The primary outcome was the proportion of participants with PCR-confirmed COVID-19 from day 21 after receiving the first dose. All analyses excluded participants with protocol violations: the primary outcome was assessed in participants who had received two doses of vaccine or placebo, serious adverse events were assessed in all participants who had received at least one dose at the time of database lock, and rare adverse events were assessed in all participants who had received two doses and for whom all available data were verified in the case report form at the time of database lock. The trial is registered at ClinicalTrials.gov (NCT04530396). Between Sept 7 and Nov 24, 2020, 21 977 adults were randomly assigned to the vaccine group (n=16 501) or the placebo group (n=5476). 19 866 received two doses of vaccine or placebo and were included in the primary outcome analysis. From 21 days after the first dose of vaccine (the day of dose 2), 16 (0·1%) of 14 964 participants in the vaccine group and 62 (1·3%) of 4902 in the placebo group were confirmed to have COVID-19; vaccine efficacy was 91·6% (95% CI 85·6-95·2). Most reported adverse events were grade 1 (7485 [94·0%] of 7966 total events). 45 (0·3%) of 16 427 participants in the vaccine group and 23 (0·4%) of 5435 participants in the placebo group had serious adverse events; none were considered associated with vaccination, with confirmation from the independent data monitoring committee. Four deaths were reported during the study (three [<0·1%] of 16 427 participants in the vaccine group and one [<0·1%] of 5435 participants in the placebo group), none of which were considered related to the vaccine. This interim analysis of the phase 3 trial of Gam-COVID-Vac showed 91·6% efficacy against COVID-19 and was well tolerated in a large cohort. Moscow City Health Department, Russian Direct Investment Fund, and Sberbank.",
"In March 2020, the World Health Organization (WHO) declared coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)1, a pandemic. With rapidly accumulating numbers of cases and deaths reported globally2, a vaccine is urgently needed. Here we report the available safety, tolerability and immunogenicity data from an ongoing placebo-controlled, observer-blinded dose-escalation study (ClinicalTrials.gov identifier NCT04368728) among 45 healthy adults (18-55 years of age), who were randomized to receive 2 doses-separated by 21 days-of 10 μg, 30 μg or 100 μg of BNT162b1. BNT162b1 is a lipid-nanoparticle-formulated, nucleoside-modified mRNA vaccine that encodes the trimerized receptor-binding domain (RBD) of the spike glycoprotein of SARS-CoV-2. Local reactions and systemic events were dose-dependent, generally mild to moderate, and transient. A second vaccination with 100 μg was not administered because of the increased reactogenicity and a lack of meaningfully increased immunogenicity after a single dose compared with the 30-μg dose. RBD-binding IgG concentrations and SARS-CoV-2 neutralizing titres in sera increased with dose level and after a second dose. Geometric mean neutralizing titres reached 1.9-4.6-fold that of a panel of COVID-19 convalescent human sera, which were obtained at least 14 days after a positive SARS-CoV-2 PCR. These results support further evaluation of this mRNA vaccine candidate.",
"We developed a heterologous COVID-19 vaccine consisting of two components, a recombinant adenovirus type 26 (rAd26) vector and a recombinant adenovirus type 5 (rAd5) vector, both carrying the gene for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein (rAd26-S and rAd5-S). We aimed to assess the safety and immunogenicity of two formulations (frozen and lyophilised) of this vaccine. We did two open, non-randomised phase 1/2 studies at two hospitals in Russia. We enrolled healthy adult volunteers (men and women) aged 18-60 years to both studies. In phase 1 of each study, we administered intramuscularly on day 0 either one dose of rAd26-S or one dose of rAd5-S and assessed the safety of the two components for 28 days. In phase 2 of the study, which began no earlier than 5 days after phase 1 vaccination, we administered intramuscularly a prime-boost vaccination, with rAd26-S given on day 0 and rAd5-S on day 21. Primary outcome measures were antigen-specific humoral immunity (SARS-CoV-2-specific antibodies measured by ELISA on days 0, 14, 21, 28, and 42) and safety (number of participants with adverse events monitored throughout the study). Secondary outcome measures were antigen-specific cellular immunity (T-cell responses and interferon-γ concentration) and change in neutralising antibodies (detected with a SARS-CoV-2 neutralisation assay). These trials are registered with ClinicalTrials.gov, NCT04436471 and NCT04437875. Between June 18 and Aug 3, 2020, we enrolled 76 participants to the two studies (38 in each study). In each study, nine volunteers received rAd26-S in phase 1, nine received rAd5-S in phase 1, and 20 received rAd26-S and rAd5-S in phase 2. Both vaccine formulations were safe and well tolerated. The most common adverse events were pain at injection site (44 [58%]), hyperthermia (38 [50%]), headache (32 [42%]), asthenia (21 [28%]), and muscle and joint pain (18 [24%]). Most adverse events were mild and no serious adverse events were detected. All participants produced antibodies to SARS-CoV-2 glycoprotein. At day 42, receptor binding domain-specific IgG titres were 14 703 with the frozen formulation and 11 143 with the lyophilised formulation, and neutralising antibodies were 49·25 with the frozen formulation and 45·95 with the lyophilised formulation, with a seroconversion rate of 100%. Cell-mediated responses were detected in all participants at day 28, with median cell proliferation of 2·5% CD4+ and 1·3% CD8+ with the frozen formulation, and a median cell proliferation of 1·3% CD4+ and 1·1% CD8+ with the lyophilised formulation. The heterologous rAd26 and rAd5 vector-based COVID-19 vaccine has a good safety profile and induced strong humoral and cellular immune responses in participants. Further investigation is needed of the effectiveness of this vaccine for prevention of COVID-19. Ministry of Health of the Russian Federation."
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Pannexin channels during immune cell migration and recruitment
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The role of Pannexin (PANX) channels during collective and single cell migration is increasingly recognized. Amongst many functions that are relevant to cell migration, here we focus on the role of PANX-mediated adenine nucleotide release and associated autocrine and paracrine signaling. We also summarize the contribution of PANXs with the cytoskeleton, which is also key regulator of cell migration. PANXs, as mechanosensitive ATP releasing channels, provide a unique link between cell migration and purinergic communication. The functional association with several purinergic receptors, together with a plethora of signals that modulate their opening, allows PANX channels to integrate physical and chemical cues during inflammation. Ubiquitously expressed in almost all immune cells, PANX1 opening has been reported in different immunological contexts. Immune activation is the epitome coordination between cell communication and migration, as leukocytes (i.e., T cells, dendritic cells) exchange information while migrating towards the injury site. In the current review, we summarized the contribution of PANX channels during immune cell migration and recruitment; although we also compile the available evidence for non-immune cells (including fibroblasts, keratinocytes, astrocytes, and cancer cells). Finally, we discuss the current evidence of PANX1 and PANX3 channels as a both positive and/or negative regulator in different inflammatory conditions, proposing a general mechanism of these channels contribution during cell migration.
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"T cells must migrate to encounter antigen-presenting cells and perform their roles in host defense. Here, we found that autocrine stimulation of the purinergic receptor P2Y11 regulates the migration of human CD4 T cells. P2Y11 receptors redistributed from the front to the back of polarized cells where they triggered intracellular cAMP/PKA signals that attenuated mitochondrial metabolism at the back. The absence of P2Y11 receptors at the front of cells resulted in hotspots of mitochondrial metabolism and localized ATP production that stimulated P2X4 receptors, Ca2+ influx, and pseudopod protrusion at the front. This regulatory function of P2Y11 receptors depended on their subcellular redistribution and autocrine stimulation by cellular ATP release and was perturbed by indiscriminate global stimulation. We conclude that excessive extracellular ATP-such as in response to inflammation, sepsis, and cancer-disrupts this autocrine feedback mechanism, which results in defective T cell migration, impaired T cell function, and loss of host immune defense.",
"Engagement of T cells with antigen-presenting cells requires T-cell receptor (TCR) stimulation at the immune synapse. We previously reported that TCR stimulation induces the release of cellular adenosine-5'-triphosphate (ATP) that regulates T-cell activation. Here we tested the roles of pannexin-1 hemichannels, which have been implicated in ATP release, and of various P2X receptors, which serve as ATP-gated Ca(2+) channels, in events that control T-cell activation. TCR stimulation results in the translocation of P2X1 and P2X4 receptors and pannexin-1 hemichannels to the immune synapse, while P2X7 receptors remain uniformly distributed on the cell surface. Removal of extracellular ATP or inhibition, mutation, or silencing of P2X1 and P2X4 receptors inhibits Ca(2+) entry, nuclear factors of activated T cells (NFAT) activation, and induction of interleukin-2 synthesis. Inhibition of pannexin-1 hemichannels suppresses TCR-induced ATP release, Ca(2+) entry, and T-cell activation. We conclude that pannexin-1 hemichannels and P2X1 and P2X4 receptors facilitate ATP release and autocrine feedback mechanisms that control Ca(2+) entry and T-cell activation at the immune synapse.",
"Organization of immune responses requires exchange of information between cells. This is achieved through either direct cell-cell contacts and establishment of temporary synapses or the release of soluble factors, such as cytokines and chemokines. Here we show a novel form of cell-to-cell communication based on adenosine triphosphate (ATP). ATP released by stimulated T cells induces P2X4/P2X7-mediated calcium waves in the neighboring lymphocytes. Our data obtained in lymph node slices suggest that, during T-cell priming, ATP acts as a paracrine messenger to reduce the motility of lymphocytes and that this may be relevant to allow optimal tissue scanning by T cells.",
"Calcium ion (Ca2+) is an essential second messenger involved in multiple cellular and subcellular processes. Ca2+ can be released and sensed globally or locally within cells, providing complex signals of variable amplitudes and time-scales. The key function of Ca2+ in the regulation of acto-myosin contractility has provided a simple explanation for its role in the regulation of immune cell migration. However, many questions remain, including the identity of the Ca2+ stores, channels and upstream signals involved in this process. Here, we focus on dendritic cells (DCs), because their immune sentinel function heavily relies on their capacity to migrate within tissues and later on between tissues and lymphoid organs. Deciphering the mechanisms by which cytoplasmic Ca2+ regulate DC migration should shed light on their role in initiating and tuning immune responses.",
"Chemotaxis, the movement of cells along chemical gradients, is critical for the recruitment of immune cells to sites of inflammation; however, how cells navigate in chemotactic gradients is poorly understood. Here, we show that macrophages navigate in a gradient of the chemoattractant C5a through the release of adenosine triphosphate (ATP) and autocrine \"purinergic feedback loops\" that involve receptors for ATP (P2Y(2)), adenosine diphosphate (ADP) (P2Y(12)), and adenosine (A2a, A2b, and A3). Whereas macrophages from mice deficient in pannexin-1 (which is part of a putative ATP release pathway), P2Y(2), or P2Y(12) exhibited efficient chemotactic navigation, chemotaxis was blocked by apyrase, which degrades ATP and ADP, and by the inhibition of multiple purinergic receptors. Furthermore, apyrase impaired the recruitment of monocytes in a mouse model of C5a-induced peritonitis. In addition, we found that stimulation of P2Y(2), P2Y(12), or adenosine receptors induced the formation of lamellipodial membrane protrusions, causing cell spreading. We propose a model in which autocrine purinergic receptor signaling amplifies and translates chemotactic cues into directional motility.",
"The immune system is the most diffuse cellular system in the body. Accordingly, long-range migration of cells and short-range communication by local chemical signaling and by cell-cell contacts are vital to the control of an immune response. Cellular homing and migration within lymphoid organs, antigen recognition, and cell signaling and activation are clearly vital during an immune response, but these events had not been directly observed in vivo until recently. Introduced to the field of immunology in 2002, two-photon microscopy is the method of choice for visualizing living cells deep within native tissue environments, and it is now revealing an elegant cellular choreography that underlies the adaptive immune response to antigen challenge. We review cellular dynamics and molecular factors that contribute to basal motility of lymphocytes in the lymph node and cellular interactions leading to antigen capture and recognition, T cell activation, B cell activation, cytolytic effector function, and antibody production.",
"ATP and hydrolysis products of ATP like adenosine regulate the chemotaxis of neutrophils by activating purinoceptors and adenosine receptors. The present study was designed to examine exogenous ATP, activation of purinoceptors, and activation of A(3) adenosine receptor as key steps in the signal cascades that control cell orientation and migration of rat neutrophils. One or more of those steps might be potential therapeutic targets for treatment of inflammatory diseases. The chemotaxis of rat neutrophils was stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP) and measured with an EZ-TAXIScan apparatus. The effects of apyrase, exogenous ATP, suramin (P2X and P2Y blocker), PPADS (a P2X blocker), TNP-ATP (P2X(1) and P2X(3) antagonist), and Reactive Blue 2 (a P2Y blocker) on the chemotactic response were also investigated. Rat neutrophil chemotaxis was significantly suppressed by apyrase. fMLP induced rat neutrophil chemotaxis was potentiated by ATP, blocked by suramin, not affected by PPADS or TNP-ATP, and significantly inhibited by RB-2. Western blotting showed that A(3), P2Y(2), and P2Y(11) were expressed in rat neutrophils. The chemotactic response of rat neutrophils to fMLP stimulation is potentiated by ATP via P2Y(11) purinoceptors but not P2X purinoceptors or A(3) adenosine receptor, and that the response plays a critical role in host defense and pathogenicity.",
"The P2Y11 receptor is a G protein-coupled receptor that is stimulated by endogenous purine nucleotides, particularly ATP. Amongst P2Y receptors it has several unique properties; (1) it is the only human P2Y receptor gene that contains an intron in the coding sequence; (2) the gene does not appear to be present in the rodent genome; (3) it couples to stimulation of both phospholipase C and adenylyl cyclase. Its absence in mice and rats, along with a limited range of selective pharmacological tools, has hampered the development of our knowledge and understanding of its properties and functions. Nonetheless, through a combination of careful use of the available tools, suppression of receptor expression using siRNA and genetic screening for SNPs, possible functions of native P2Y11 receptors have been identified in a variety of human cells and tissues. Many are in blood cells involved in inflammatory responses, consistent with extracellular ATP being a damage-associated signalling molecule in the immune system. Thus proposed potential therapeutic applications relate, in the main, to modulation of acute and chronic inflammatory responses.",
"Mathematical and computational models can assist in gaining an understanding of cell behavior at many levels of organization. Here, we review models in the literature that focus on eukaryotic cell motility at 3 size scales: intracellular signaling that regulates cell shape and movement, single cell motility, and collective cell behavior from a few cells to tissues. We survey recent literature to summarize distinct computational methods (phase-field, polygonal, Cellular Potts, and spherical cells). We discuss models that bridge between levels of organization, and describe levels of detail, both biochemical and geometric, included in the models. We also highlight links between models and experiments. We find that models that span the 3 levels are still in the minority.",
"Cell migration involves dynamic and spatially regulated changes to the cytoskeleton and cell adhesion. The Rho GTPases play key roles in coordinating the cellular responses required for cell migration. Recent research has revealed new molecular links between Rho family proteins and the actin cytoskeleton, showing that they act to regulate actin polymerization, depolymerization and the activity of actin-associated myosins. In addition, studies on integrin signalling suggest that the substratum continuously feeds signals to Rho proteins in migrating cells to influence migration rate. There is also increasing evidence that Rho proteins affect the organization of the microtubule and intermediate filament networks and that this is important for cell migration."
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Heuristic Planning with Sensing in Human Behavior
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From cooking a meal to finding a route to a destination, many real life decisions can be decomposed into a hierarchy of sub-decisions. In a hierarchy, choosing which decision to think about requires planning over a potentially vast space of possible decision sequences. To gain insight into how people decide what to decide on, we studied a novel task that combines perceptual decision making, active sensing and hierarchical and counterfactual reasoning. Human participants had to find a target hidden at the lowest level of a decision tree. They could solicit information from the different nodes of the decision tree to gather noisy evidence about the target's location. Feedback was given only after errors at the leaf nodes and provided ambiguous evidence about the cause of the error. Despite the complexity of task (with 107 latent states) participants were able to plan efficiently in the task. A computational model of this process identified a small number of heuristics of low computational complexity that accounted for human behavior. These heuristics include making categorical decisions at the branching points of the decision tree rather than carrying forward entire probability distributions, discarding sensory evidence deemed unreliable to make a choice, and using choice confidence to infer the cause of the error after an initial plan failed. Plans based on probabilistic inference or myopic sampling norms could not capture participants' behavior. Our results show that it is possible to identify hallmarks of heuristic planning with sensing in human behavior and that the use of tasks of intermediate complexity helps identify the rules underlying human ability to reason over decision hierarchies.
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"The time course of perceptual choice is discussed in a model of gradual, leaky, stochastic, and competitive information accumulation in nonlinear decision units. Special cases of the model match a classical diffusion process, but leakage and competition work together to address several challenges to existing diffusion, random walk, and accumulator models. The model accounts for data from choice tasks using both time-controlled (e.g., response signal) and standard reaction time paradigms and its adequacy compares favorably with other approaches. A new paradigm that controls the time of arrival of information supporting different choice alternatives provides further support. The model captures choice behavior regardless of the number of alternatives, accounting for the log-linear relation between reaction time and number of alternatives (Hick's law) and explains a complex pattern of visual and contextual priming in visual word identification.",
"The gradual and noisy accumulation of evidence is a fundamental component of decision-making, with noise playing a key role as the source of variability and errors. However, the origins of this noise have never been determined. We developed decision-making tasks in which sensory evidence is delivered in randomly timed pulses, and analyzed the resulting data with models that use the richly detailed information of each trial's pulse timing to distinguish between different decision-making mechanisms. This analysis allowed measurement of the magnitude of noise in the accumulator's memory, separately from noise associated with incoming sensory evidence. In our tasks, the accumulator's memory was noiseless, for both rats and humans. In contrast, the addition of new sensory evidence was the primary source of variability. We suggest our task and modeling approach as a powerful method for revealing internal properties of decision-making processes.",
"When multiple pieces of information bear on a decision, the best approach is to combine the evidence provided by each one. Evidence integration models formalize the computations underlying this process [1-3], explain human perceptual discrimination behavior [4-9], and correspond to neuronal responses elicited by discrimination tasks [10-14]. These findings suggest that evidence integration is key to understanding the neural basis of decision making [15-18]. But while evidence integration has most often been studied with simple tasks that limit deliberation to relatively brief periods, many natural decisions unfold over much longer durations. Neural network models imply acute limitations on the timescale of evidence integration [19-23], and it is currently unknown whether existing computational insights can generalize beyond rapid judgments. Here, we introduce a new psychophysical task and report model-based analyses of human behavior that demonstrate evidence integration at long timescales. Our task requires probabilistic inference using brief samples of visual evidence that are separated in time by long and unpredictable gaps. We show through several quantitative assays how decision making can approximate a normative integration process that extends over tens of seconds without accruing significant memory leak or noise. These results support the generalization of evidence integration models to a broader class of behaviors while posing new challenges for models of how these computations are implemented in biological networks.",
"The mesostriatal dopamine system is prominently implicated in model-free reinforcement learning, with fMRI BOLD signals in ventral striatum notably covarying with model-free prediction errors. However, latent learning and devaluation studies show that behavior also shows hallmarks of model-based planning, and the interaction between model-based and model-free values, prediction errors, and preferences is underexplored. We designed a multistep decision task in which model-based and model-free influences on human choice behavior could be distinguished. By showing that choices reflected both influences we could then test the purity of the ventral striatal BOLD signal as a model-free report. Contrary to expectations, the signal reflected both model-free and model-based predictions in proportions matching those that best explained choice behavior. These results challenge the notion of a separate model-free learner and suggest a more integrated computational architecture for high-level human decision-making.",
"Several widely accepted models of decision making suggest that, during simple decision tasks, neural activity builds up until a threshold is reached and a decision is made. These models explain error rates and reaction time distributions in a variety of tasks and are supported by neurophysiological studies showing that neural activity in several cortical and subcortical regions gradually builds up at a rate related to task difficulty and reaches a relatively constant level of discharge at a time that predicts movement initiation. The mechanism responsible for this buildup is believed to be related to the temporal integration of sequential samples of sensory information. However, an alternative mechanism that may explain the neural and behavioral data is one in which the buildup of activity is instead attributable to a growing signal related to the urgency to respond, which multiplicatively modulates updated estimates of sensory evidence. These models are difficult to distinguish when, as in previous studies, subjects are presented with constant sensory evidence throughout each trial. To distinguish the models, we presented human subjects with a task in which evidence changed over the course of each trial. Our results are more consistent with \"urgency gating\" than with temporal integration of sensory samples and suggest a simple mechanism for implementing trade-offs between the speed and accuracy of decisions.",
"Planning allows actions to be structured in pursuit of a future goal. However, in natural environments, planning over multiple possible future states incurs prohibitive computational costs. To represent plans efficiently, states can be clustered hierarchically into \"contexts\". For example, representing a journey through a subway network as a succession of individual states (stations) is more costly than encoding a sequence of contexts (lines) and context switches (line changes). Here, using functional brain imaging, we asked humans to perform a planning task in a virtual subway network. Behavioral analyses revealed that humans executed a hierarchically organized plan. Brain activity in the dorsomedial prefrontal cortex and premotor cortex scaled with the cost of hierarchical plan representation and unique neural signals in these regions signaled contexts and context switches. These results suggest that humans represent hierarchical plans using a network of caudal prefrontal structures. VIDEO ABSTRACT.",
"The ability to manipulate neural activity with precision is an asset in uncovering neural circuits for decision-making. Diverse tools for manipulating neurons are available for mice, but their feasibility remains unclear, especially when decisions require accumulating visual evidence. For example, whether mice' decisions reflect leaky accumulation is unknown, as are the relevant/irrelevant factors that influence decisions. Further, causal circuits for visual evidence accumulation are poorly understood. To address this, we measured decisions in mice judging the fluctuating rate of a flash sequence. An initial analysis (>500,000 trials, 29 male and female mice) demonstrated that information throughout the 1000 ms trial influenced choice, with early information most influential. This suggests that information persists in neural circuits for ∼1000 ms with minimal accumulation leak. Next, in a subset of animals, we probed strategy more extensively and found that although animals were influenced by stimulus rate, they were unable to entirely suppress the influence of stimulus brightness. Finally, we identified anteromedial (AM) visual area via retinotopic mapping and optogenetically inhibited it using JAWS. Light activation biased choices in both injected and uninjected animals, demonstrating that light alone influences behavior. By varying stimulus-response contingency while holding stimulated hemisphere constant, we surmounted this obstacle to demonstrate that AM suppression biases decisions. By leveraging a large dataset to quantitatively characterize decision-making behavior, we establish mice as suitable for neural circuit manipulation studies. Further, by demonstrating that mice accumulate visual evidence, we demonstrate that this strategy for reducing uncertainty in decision-making is used by animals with diverse visual systems.SIGNIFICANCE STATEMENT To connect behaviors to their underlying neural mechanism, a deep understanding of behavioral strategy is needed. This understanding is incomplete for mice. To surmount this, we measured the outcome of >500,000 decisions made by 29 mice trained to judge visual stimuli and performed behavioral/optogenetic manipulations in smaller subsets. Our analyses offer new insights into mice' decision-making strategies and compares them with those of other species. We then disrupted neural activity in a candidate neural structure and examined the effect on decisions. Our findings establish mice as suitable for visual accumulation of evidence decisions. Further, the results highlight similarities in decision-making strategies across very different species."
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Molecular basis for avirulence in Shigella sonnei
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Shigella sonnei is a major cause of bacillary dysentery and an increasing concern due to the spread of multidrug resistance. S. sonnei harbors pINV, an ∼210 kb plasmid that encodes a type III secretion system (T3SS), which is essential for virulence. During growth in the laboratory, avirulence arises spontaneously in S. sonnei at high frequency, hampering studies on and vaccine development against this important pathogen. Here, we investigated the molecular basis for the emergence of avirulence in S. sonnei and showed that avirulence mainly results from pINV loss, which is consistent with previous findings. Ancestral deletions have led to the loss from S. sonnei pINV of two toxin-antitoxin (TA) systems involved in plasmid maintenance, CcdAB and GmvAT, which are found on pINV in Shigella flexneri. We showed that the introduction of these TA systems into S. sonnei pINV reduced but did not eliminate pINV loss, while the single amino acid polymorphisms found in the S. sonnei VapBC TA system compared with S. flexneri VapBC also contributed to pINV loss. Avirulence also resulted from deletions of T3SS-associated genes in pINV through recombination between insertion sequences (ISs) on the plasmid. These events differed from those observed in S. flexneri due to the different distribution and repertoire of ISs. Our findings demonstrated that TA systems and ISs influenced plasmid dynamics and loss in S. sonnei and could be exploited for the design and evaluation of vaccines.
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"Construction of a stable Shigella sonnei vaccine has been complicated by the instability of the virulence phenotype caused by the spontaneous loss of the invasion plasmid. To select a suitable candidate for vaccine construction, 16 S. sonnei strains were screened for stability of the virulence phenotype. A stable strain, S. sonnei Mosely, was selected for further work. pDeltavirG2, a deletion derivative of the virG gene in the sacB suicide vector pCVD442, was used to generate an S. sonnei virG deletion strain, WRSS1, which was invasive in HeLa cells but negative in the Sereny test. WRSS1 was found to be both immunogenic and protective in the guinea pig keratoconjunctivitis model.",
"Shigella sonnei is the most common agent of shigellosis in high-income countries, and causes a significant disease burden in low- and middle-income countries. Antimicrobial resistance is increasingly common in all settings. Whole genome sequencing (WGS) is increasingly utilised for S. sonnei outbreak investigation and surveillance, but comparison of data between studies and labs is challenging. Here, we present a genomic framework and genotyping scheme for S. sonnei to efficiently identify genotype and resistance determinants from WGS data. The scheme is implemented in the software package Mykrobe and tested on thousands of genomes. Applying this approach to analyse >4,000 S. sonnei isolates sequenced in public health labs in three countries identified several common genotypes associated with increased rates of ciprofloxacin resistance and azithromycin resistance, confirming intercontinental spread of highly-resistant S. sonnei clones and demonstrating the genomic framework can facilitate monitoring the spread of resistant clones, including those that have recently emerged, at local and global scales.",
"Shigella is the major cause of bacillary dysentery world-wide. It is divided into four species, named S. flexneri, S. sonnei, S. dysenteriae, and S. boydii, which are distinct genomically and in their ability to cause disease. Shigellosis, the clinical presentation of Shigella infection, is characterized by watery diarrhea, abdominal cramps, and fever. Shigella's ability to cause disease has been attributed to virulence factors, which are encoded on chromosomal pathogenicity islands and the virulence plasmid. However, information on these virulence factors is not often brought together to create a detailed picture of infection, and how this translates into shigellosis symptoms. Firstly, Shigella secretes virulence factors that induce severe inflammation and mediate enterotoxic effects on the colon, producing the classic watery diarrhea seen early in infection. Secondly, Shigella injects virulence effectors into epithelial cells via its Type III Secretion System to subvert the host cell structure and function. This allows invasion of epithelial cells, establishing a replicative niche, and causes erratic destruction of the colonic epithelium. Thirdly, Shigella produces effectors to down-regulate inflammation and the innate immune response. This promotes infection and limits the adaptive immune response, causing the host to remain partially susceptible to re-infection. Combinations of these virulence factors may contribute to the different symptoms and infection capabilities of the diverse Shigella species, in addition to distinct transmission patterns. Further investigation of the dominant species causing disease, using whole-genome sequencing and genotyping, will allow comparison and identification of crucial virulence factors and may contribute to the production of a pan-Shigella vaccine.",
"A plasmid of about 140 megadaltons has been associated with the invasiveness of Shigella flexneri. Upon subculturing in liquid media of fully virulent isolates of Shigella flexneri 2a YSH6000, which contains only a 230-kilobase-pair (kbp) plasmid in addition to 3.3- and 4.2-kbp cryptic plasmids characteristic to all S. flexneri strains, loss of invasiveness, loss of Congo red binding activity (Pcr), and complete loss of, or a deletion, or even a single-site IS insertion in the plasmid occurred simultaneously. This was ascribed to the fact that, once a noninvasive Pcr- cell has emerged, it overgrows the wild type as a consequence of its selective advantage in artificial media. A deletion map of the 230-kbp plasmid was made by analyzing SalI digests of 39 deletion derivatives plus 1 formed by insertion of an IS1-like element in independently isolated, noninvasive Pcr- mutants. Of 39 deletion derivatives, 16 belonged to a single type, and 6 belonged to another, suggesting deletion hot spots. The deletion map was confirmed and extended by analyzing 359 SalI-generated partial digests of the wild-type plasmid cloned into pBR322. Three copies of IS1-like elements were found on three different SalI fragments by Southern hybridization. Segments required for the Pcr+ phenotype seemed to occur at several different locations in the plasmid. Each of 28 representative Pcr- mutants were negative by the Sereny test. Hence, many, or possibly all, Pcr determinants were required for full virulence.",
"Transformation of E. coli cells treated with CaCl(2) to multiple antibiotic resistance by purified R-factor DNA is reported. Drug resistance is expressed in a small fraction of the recipient bacterial population almost immediately after uptake of DNA, but full genetic expression of resistance requires subsequent incubation in drugfree medium before antibiotic challenge. Transformed bacteria acquire a closed circular, transferable DNA species having the resistance, fertility, and sedimentation characteristics of the parent R factor. Covalently-closed, catenated, and open (nicked) circular forms of R-factor DNA are all effective in transformation, but denaturation and sonication abolish the transforming ability of R-factor DNA in this system.",
"Expression of the predominantly plasmid-encoded virulence regulon of Shigella flexneri 2a is induced by growth at 37 degrees C and repressed by growth at 30 degrees C. During growth at 37 degrees C, spontaneous S. flexneri mutants arise which have undergone virulence plasmid curing or rearrangement and no longer display the virulent phenotype. In the laboratory, the unstable nature of the virulence plasmid causes complete loss of virulence in a growing population. We have undertaken an analysis of virulence plasmid instability, classifying events which produced individual avirulent derivatives within a virulent population and identifying the factor(s) which controlled conversion. Multiplex PCR analysis of DNA obtained from spontaneous avirulent derivatives indicated that virF and virB were deleted or otherwise inactivated in over 97% of the isolates. The virF and virB loci encode regulatory proteins required for transcriptional activation of the virulence regulon. Inactivation of these key regulatory loci in the vast majority of avirulent derivatives which arose during growth at 37 degrees C suggested that virulence gene expression induced virulence plasmid instability. Consistent with this hypothesis, we observed stable virulence plasmid maintenance during growth of a wild-type strain at 30 degrees C where virulence gene expression was repressed. The virulence plasmid was also stably maintained in virF and virB mutants grown at 37 degrees C. Conversely, virulence plasmid destabilization was induced at 30 degrees C and accelerated at 37 degrees C through expression of VirF or VirB from multicopy plasmids. These results indicate that exposure of S. flexneri to conditions favoring induction of the virulent phenotype also favor its loss. The significance of this paradox of Shigella pathogenicity is discussed.",
"Transcriptional fusions between the phage lambda promotor pR and ATP synthase genes, atp, on plasmid pBR322 were constructed in order to study the effects upon growth and physiology of Escherichia coli of induced overproduction of H+-ATPase subunits. Constitutive overproduction of the complete enzyme had earlier been found to result in decreased growth rate and cytological defects. When a 15-fold overproduction of subunit a alone, or together with subunit c, or with all other ATP synthase subunits was suddenly induced, the following effects were observed. Inhibition of growth and protein synthesis within 10 min of induction, which effect was suppressed by N,N'-dicyclohexylcarbodiimide, also when the chromosomal atp genes coding for the Fo subunits a, b and c were deleted. Partial collapse of the membrane potential delta psi at 4-6 min after induction paralleled by inhibition of thiomethylgalactoside and guanosine transport. Respiration and alpha-methylglucoside transport was not affected. The partial collapse of delta psi, and the specific inhibition of proton-driven transport systems is taken to show that the subunit a has--when suddenly overproduced and inserted into the membrane--a protonophoric activity. It is suggested that this protonophoric activity of subunit a is related to the function of this subunit in the Fo sector in H+-ATPases."
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Engineering Noble Metal Nanoparticles for Lateral Flow Immunoassay
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Lateral flow immunoassay (LFIA) with gold nanoparticles (AuNPs) as signal reporters is a popular point-of-care diagnostic technique. However, given the weak absorbance of traditional 20-40 nm spherical AuNPs, their sensitivity is low, which greatly limits the wide application of AuNP-based LFIA. With the rapid advances in materials science and nanotechnology, the synthesis of noble metal nanoparticles (NMNPs) has enhanced physicochemical properties such as optical, plasmonic, catalytic, and multifunctional activity by simply engineering their physical parameters, including the size, shape, composition, and external structure. Using these engineered NMNPs as an alternative to traditional AuNPs, the sensitivity of LFIA has been significantly improved, thereby greatly expanding the working range and application scenarios of LFIA, particularly in trace analysis. Therefore, in this review, we will focus on the design of engineered NMNPs and their demonstration in improving LFIA. We highlight the strategies available for tailoring NMNP designs, the effect of NMNP engineering on their performance, and the working principle of each engineering design for enhancing LFIA. Finally, current challenges and future improvements in this field are briefly discussed.
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"The authors describe the preparation of gold-platinum nanoflower (AuPt NFs) and show that they can be simultaneously used as a label and as an enzyme mimic in lateral flow immunoassays (LFIs). The AuPt NFs were prepared by growing Pt nanowires on the surface of gold nanoparticle. The assay involves the capture of target proteins (here: rabbit IgG as a model analyte) by the immobilized capture antibody, and by using AuPt NF-labeled secondary antibody. The AuPt NFs are thus captured by the test zone and produce a characteristic black band for visual detection of the antigen (IgG). The coloration of the test line can be further enhanced by addition of the chromogenic substrate 3-amino-9-ethyl-carbazole which is catalytically oxidized by the captured Pt nanowires on the AuPt NF and produce a red coloration. Quantitative results were obtained by reading the test line intensities with a portable strip reader. The LFI has a 5 pg mL-1 detection limit for IgG under optimized experimental conditions. This is 100 times lower than that of the conventional AuNP-based LFI. Conceivably, this assay has a wide scope in that it may be applied to numerous other targets for which appropriate antibodies are available. Graphical abstract Gold-platinum nanoflowers are used as a label and as an enzyme mimic in a highly sensitive lateral flow immunoassay for IgG. The detection limit of gold-platinum nanoflower-based lateral flow assay is 100 times lower than that of the conventional gold nanopaticle-based lateral flow assay.",
"Lateral flow immunoassay (LFIA) has emerged as an effective technique in the field of food safety and environmental monitoring. However, sensitive and quantitative detection is still challenging for LFIAs in complex environments. In this work, a dual-model colorimetric/SERS lateral flow immunoassay for ultrasensitive determination of clenbuterol was constructed based on a metallic core-shell Au/Au nanostar acting as a multifunction tag. Raman reporter molecules are located between the core (AuNP) and shell (Au nanostar) to form a sandwich structure, which contributes to eliminate the environmental interference and improve the detection stability. In addition, the Au/Au nanostar provides a much higher Raman enhancement due to the presence of sharp tips and larger surface roughness in comparison with gold nanoparticles (AuNPs). Thus, on the basis of the antibody-antigen interaction, the dual-model immunoassay can produce strong colorimetric and surface-enhanced Raman spectroscopy (SERS) signals for highly sensitive detection of the target analyte, clenbuterol. Under optimal conditions, clenbuterol could be detected by the colorimetric model with a visual detection limit of 5 ng/mL. Meanwhile, the SERS signal of the Au/Au nanostar was accumulated on the test line for the SERS model detection with a quantitative detection limit as low as 0.05 ng/mL, which is at least 200-fold lower than that of the traditional AuNPs-based immunoassay. Furthermore, recovery rates of the proposed method in food samples were 86-110%. This dual-model immunoassay provides an effective tool for antibiotic residues analysis and demonstrates a broad potential for future applications in food safety monitoring.",
"Lateral flow immunoassay (LFIA) is a widely used express method and offers advantages such as a short analysis time, simplicity of testing and result evaluation. However, an LFIA based on gold nanospheres lacks the desired sensitivity, thereby limiting its wide applications. In this study, spherical nanogold labels along with new types of nanogold labels such as gold nanopopcorns and nanostars were prepared, characterized, and applied for LFIA of model protein antigen procalcitonin. It was found that the label with a structure close to spherical provided more uniform distribution of specific antibodies on its surface, indicative of its suitability for this type of analysis. LFIA using gold nanopopcorns as a label allowed procalcitonin detection over a linear range of 0.5-10 ng mL-1 with the limit of detection of 0.1 ng mL-1, which was fivefold higher than the sensitivity of the assay with gold nanospheres. Another approach to improve the sensitivity of the assay included the silver enhancement method, which was used to compare the amplification of LFIA for procalcitonin detection. The sensitivity of procalcitonin determination by this method was 10 times better the sensitivity of the conventional LFIA with gold nanosphere as a label. The proposed approach of LFIA based on gold nanopopcorns improved the detection sensitivity without additional steps and prevented the increased consumption of specific reagents (antibodies).",
"The present study describes a lateral-flow-based dipstick immunoassay format using a novel hapten-protein-gold conjugate for the rapid screening of atrazine in water samples. The immunoassay is based on the competitive inhibition, in which a newly developed hapten-protein-gold conjugate competes with the free antigen present in the sample, for the limited antibody binding sites available at test zone on dipstick membrane, housed in a plastic cartridge. The tracer used as the detection reagent was prepared by first conjugating hapten (a derivative of atrazine) molecules to a carrier protein (bovine serum albumin) via its surface lysine residues and then linking colloidal gold nanoparticles to the hapten-protein conjugate via cysteine residues of the carrier protein. The developed conjugate showed a high level of stability as it did not show any significant loss of activity even after 8 weeks of storage at ambient conditions. The color developed due to conjugate, based on competitive inhibition approach, is correlated with the concentration of atrazine sample. The sensitivity of the developed dipstick was enhanced by gold nanoparticles, as an amplification tag, presenting detection limit of atrazine in standard water samples down to 1.0 ppb level. The kit could serve as a rapid screening methodology for visual screening of atrazine contamination of water samples within 5 min of analysis time, and, when coupled with a portable colorimeter, as an inexpensive semi-quantitative assay. The method reported can be useful for screening a large number of pesticides samples in a very short time in the field.",
"Lateral flow assay (LFA) has been applied in many fields due to its relative ease of use and cost-effectiveness. However, it has low sensitivity and its applications are limited. Probe materials play a significant role in improving the detection efficiency and sensitivity of LFA. In this study, by using concave palladium-platinum (Pd-Pt) nanoparticles as a nanozyme probe, we developed a sensitive LFA based on the sandwich format for qualitative and quantitative detection of Escherichia coli O157:H7. The sensitivity of the LFA was improved by applying the 3,3',5,5'-tetramethylbenzidine (TMB) substrate onto the test line where the nanozyme was accumulated in the presence of analytes. The nanozyme showed high catalytic performance toward TMB and greatly enhanced the signal intensity of the test line. The sensitivity of the nanozyme-based LFA was 9.0 × 102 cfu/mL in milk, which was 111-fold higher than that of traditional colloidal gold-based LFA. The proposed method has remarkable potential in the detection of various pathogens in real samples.",
"The conventional colloidal gold immunochromatographic assay (AuNP-ICA) cannot meet the requirements for the rapid and sensitive detection of Escherichia coli (E. coli) O157:H7 because of its poor sensitivity. Herein, a novel two-step cascade signal amplification strategy that integrates in situ gold growth and nanozyme-mediated catalytic deposition was proposed to enhance the detection sensitivity of conventional AuNP-ICA dramatically. The enhanced strip displayed ultrahigh sensitivity in E. coli O157:H7 detection and had a detection limit of 1.25 × 101 CFU/mL, which is approximately 400-fold lower than that of traditional AuNP-ICA (5 × 103 CFU/mL). The amplified strip had no background signal in the T-line zone in the absence of E. coli O157:H7 even after one round of cascade signal amplification. The enhanced strip demonstrated excellent selectivity against E. coli O157:H7 with a negligible cross-reaction to nine other common pathogens. Intra-assays and interassays showed that the improved strip has acceptable accuracy and precision for determining E. coli O157:H7. The average recoveries in a real milk sample ranged from 87.33 to 112.15%, and the coefficients of variation were less than 10%. The enhanced strip also showed great potential in detecting a single E. coli O157:H7 cell in a 75 μL solution.",
"Phenylethanolamine A (PA) is a new kind of β-agonist, which was illegally used as a feed additive for growth promotion in China. In this study, a novel immunochromatographic assay (ICA) based on surface-enhanced Raman scattering (SERS) for the ultrasensitive and quantitative detection of phenylethanolamine A is presented. The principle of this new ICA is similar to that based on colloidal gold particles, but using Au(MBA)@Ag-Ab [e.g., polyclonal antibody of PA labeled Au@Ag core-shell nanoparticles (NPs) sandwiched with a Raman reporter (4-mercaptobenzoic acid, MBA)] as a probe. After ICA procedures, the specific Raman scattering intensity of MBA on the test line was measured for quantitative detection of PA. This assay was completed within 15 min. The IC50 and limit of detection (LOD) values of the ICA for PA detection were 0.06 ng mL(-1) and 0.32 pg mL(-1), respectively, which were 1-3 orders of magnitude lower than those obtained by other immunoassays, indicating the ultrasensitivity of this ICA. There was no cross-reactivity (CR) of the assay with another three β-agonists (ractopamine, clenbuterol, and salbutamol), suggesting high specificity of the SERS-based ICA. A spiking experiment revealed that the recoveries of PA from pig urine samples were in range of 99.9- 101.2% with relative standard deviations (RSDs) of 3.6-5.8%. The results demonstrated that this SERS-based ICA was able to quantitatively detect PA in urine samples with high sensitivity, specificity, precision, and accuracy and might be a powerful method for the analysis of other target analytes in the food area."
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Spontaneous bone regeneration after extensive resection of the jaw bone
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Few cases of spontaneous bone regeneration after extensive resection of the jaw bone have been reported, but it is more common in young adults or children. In this case, we report spontaneous bone healing in a 73-year-old female patient. On radiological examination, necrotic regions were seen in the right mandible. She was diagnosed with medication-related osteonecrosis of the jaw due to previous bisphosphonate use. After segmental resection, stabilization achieved using a reconstruction plate. The periosteum was preserved during the procedure. Twelve months later, panoramic radiography was taken and bone formation was seen both horizontally and vertically around the plate. If the periosteum is preserved and stabilization is achieved after resection in benign lesions, the bone may regenerate spontaneously regardless of age. Therefore, instead of simultaneous autogenous bone application, such patients may be followed to determine whether spontaneous bone healing will occur. This should improve patient comfort and reduce surgical cost.
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[
"While century old clinical reports document the periosteum's remarkable regenerative capacity, only in the past decade have scientists undertaken mechanistic investigations of its regenerative potential. At a Workshop at the 2012 Annual Meeting of Orthopaedic Research Society, we reviewed the molecular, cellular, and tissue scale approaches to elucidate the mechanisms underlying the periosteum's regenerative potential as well as translational therapies engineering solutions inspired by its remarkable regenerative capacity. The entire population of osteoblasts within periosteum, and at endosteal and trabecular bone surfaces within the bone marrow, derives from the embryonic perichondrium. Periosteal cells contribute more to cartilage and bone formation within the callus during fracture healing than do cells of the bone marrow or endosteum, which do not migrate out of the marrow compartment. Furthermore, a current healing paradigm regards the activation, expansion, and differentiation of periosteal stem/progenitor cells as an essential step in building a template for subsequent neovascularization, bone formation, and remodeling. The periosteum comprises a complex, composite structure, providing a niche for pluripotent cells and a repository for molecular factors that modulate cell behavior. The periosteum's advanced, \"smart\" material properties change depending on the mechanical, chemical, and biological state of the tissue. Understanding periosteum development, progenitor cell-driven initiation of periosteum's endogenous tissue building capacity, and the complex structure-function relationships of periosteum as an advanced material are important for harnessing and engineering ersatz materials to mimic the periosteum's remarkable regenerative capacity.",
"One of the key challenges in bone tissue engineering is the timely formation of blood vessels that promote the survival of the implanted cells in the construct. Fracture healing largely depends on the presence of an intact periosteum but it is still unknown whether periosteum-derived cells (PDC) are critical for bone repair only by promoting bone formation or also by inducing neovascularization. We first established a protocol to specifically isolate murine PDC (mPDC) from long bones of adult mice. Mesenchymal stem cells were abundantly present in this cell population as more than 50% of the mPDC expressed mesenchymal markers (CD73, CD90, CD105, and stem cell antigen-1) and the cells exhibited trilineage differentiation potential (chondrogenic, osteogenic, and adipogenic). When transplanted on a collagen-calcium phosphate scaffold in vivo, mPDC attracted numerous blood vessels and formed mature bone which comprises a hematopoiesis-supportive stroma. We explored the proangiogenic properties of mPDC using in vitro culture systems and showed that mPDC promote the survival and proliferation of endothelial cells through the production of vascular endothelial growth factor. Coimplantation with endothelial cells demonstrated that mPDC can enhance vasculogenesis by adapting a pericyte-like phenotype, in addition to their ability to stimulate blood vessel ingrowth from the host. In conclusion, these findings demonstrate that periosteal cells contribute to fracture repair, not only through their strong osteogenic potential but also through their proangiogenic features and thus provide an ideal cell source for bone regeneration therapies.",
"Bone regeneration relies on the activation of skeletal stem cells (SSCs) that still remain poorly characterized. Here, we show that periosteum contains SSCs with high bone regenerative potential compared to bone marrow stromal cells/skeletal stem cells (BMSCs) in mice. Although periosteal cells (PCs) and BMSCs are derived from a common embryonic mesenchymal lineage, postnatally PCs exhibit greater clonogenicity, growth and differentiation capacity than BMSCs. During bone repair, PCs can efficiently contribute to cartilage and bone, and integrate long-term after transplantation. Molecular profiling uncovers genes encoding Periostin and other extracellular matrix molecules associated with the enhanced response to injury of PCs. Periostin gene deletion impairs PC functions and fracture consolidation. Periostin-deficient periosteum cannot reconstitute a pool of PCs after injury demonstrating the presence of SSCs within periosteum and the requirement of Periostin in maintaining this pool. Overall our results highlight the importance of analyzing periosteum and PCs to understand bone phenotypes.",
"Bone formation is a continuous process that begins during fetal development and persists throughout life as a remodeling process. In the event of injury, bones heal by generating new bone rather than scar tissue; thus, it can accurately be described as a regenerative process. To elucidate the extent to which fetal skeletal development and skeletal regeneration are similar, we performed a series of detailed expression analyses using a number of genes that regulate key stages of endochondral ossification. They included genes in the indian hedgehog (ihh) and core binding factor 1 (cbfa1) pathways, and genes associated with extracellular matrix remodeling and vascular invasion including vascular endothelial growth factor (VEGF) and matrix metalloproteinase 13 (mmp13). Our analyses suggested that even at the earliest stages of mesenchymal cell condensation, chondrocyte (ihh, cbfa1 and collagen type II-positive) and perichondrial (gli1 and osteocalcin-positive) cell populations were already specified. As chondrocytes matured, they continued to express cbfa1 and ihh whereas cbfa1, osteocalcin and gli1 persisted in presumptive periosteal cells. Later, VEGF and mmp13 transcripts were abundant in chondrocytes as they underwent hypertrophy and terminal differentiation. Based on these expression patterns and available genetic data, we propose a model where Ihh and Cbfa1, together with Gli1 and Osteocalcin participate in establishing reciprocal signal site of injury. The persistence of cbfa1 and ihh, and their targets osteocalcin and gli1, in the callus suggests comparable processes of chondrocyte maturation and specification of a neo-perichondrium occur following injury. VEGF and mmp13 are expressed during the later stages of healing, coincident with the onset of vascularization of the callus and subsequent ossification. Taken together, these data suggest the genetic mechanisms regulating fetal skeletogenesis also regulate adult skeletal regeneration, and point to important regulators of angiogenesis and ossification in bone regeneration."
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The Priority of Emergency Care in Low and Middle Income Countries
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The high burden of emergency medical conditions has not been met with adequate financial and political prioritisation especially in low and middle-income countries. We examined the factors that have shaped the priority of global emergency care and highlight potential responses by emergency care advocates.
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[
"We sought to compare National Institutes of Health (NIH) funding received by Emergency Medicine (EM) to the specialties of Family Medicine, Neurology, Orthopedics, Pediatrics and Psychiatry over the 10-year period from 2008 to 2017. The NIH database of both submitted and funded NIH applications were queried and crossed with the departmental affiliation of the principal investigator. Research Grants were defined by the following activity codes: R, P, M, S, K, U (excluding UC6), DP1, DP2, DP3, DP4, DP5, D42 and G12. Derived data were further analyzed using information from the Association of American Medical Colleges to determine the relationship between the number of awards and the size of respective teaching and research faculty. From 2008 to 2017, there were a total of 14,676 funded grants across included specialties with total monetary support of $6.002 billion. Of these funded grants, 250 (1.7%) were from EM principal investigators which corresponded to total support of $89,453,635 (1.5% of overall dollars). There was an increase in total support after 2012 in EM, however when compared to the other specialties, EM investigators submitted relatively fewer grants and awarded grants were funded by a wider distribution of NIH Institutes and Centers (ICs). Compared to other select specialties, EM investigators accounted for a small proportion of grants submitted and funded over the past decade. Though findings illustrate promising trends, to foster success, more submitted grant applications are needed from within EM along with systematic approaches to support faculty members in their pursuit of NIH funding.",
"The recent adoption of World Health Assembly Resolution 60.22, titled \"Health Systems: Emergency Care Systems,\" has established an important health care policy tool for improving emergency care access and availability globally. The resolution highlights the role that strengthened emergency care systems can play in reducing the increasing burden of disease from acute illness and injury in populations across the socioeconomic spectrum and calls on governments and the World Health Organization to take specific and concrete actions to make this happen. This resolution constitutes recognition by the World Health Assembly of the growing public health role of emergency care systems and is the highest level of international attention ever devoted to emergency care systems worldwide. Emergency care systems for secondary prevention of acute illnesses and injury remain inadequately developed in many low- and middle-income countries, despite evidence that basic strategies for improving emergency care systems can reduce preventable mortality and morbidity and can in many cases also be cost-effective. Emergency care providers and their professional organizations have used their comprehensive expertise to strengthen emergency care systems worldwide through the development of tools for emergency medicine education, systems assessment, quality improvement, and evidence-based clinical practice. World Health Assembly 60.22 represents a unique opportunity for emergency care providers and other advocates for improved emergency care to engage with national and local health care officials and policymakers, as well as with the World Health Organization, and leverage the expertise within the international emergency medicine community to make substantial improvements in emergency care delivery in places where it is most needed.",
"Despite the high burden of surgical conditions, the provision of surgical services has been a low global health priority. We examined factors that have shaped priority for global surgical care. We undertook semi-structured interviews by telephone with members of global surgical networks and ministries of health to explore the challenges and opportunities surgeons, anaesthesiologists, and other proponents face in increasing global priority for surgery. We did a literature review and collected information from reports from organisations involved in surgery. We used a policy framework consisting of four categories-actor power, ideas, political contexts, and characteristics of the issue itself-to analyse factors that have shaped global political priority for surgery. We did a thematic analysis on the collected information. Several factors hinder the acquisition of attention and resources for global surgery. With respect to actor power, the global surgery community is fragmented, does not have unifying leadership, and is missing guiding institutions. Regarding ideas, community members disagree on how to address and publicly position the problem. With respect to political contexts, the community has made insufficient efforts to capitalise on political opportunities such as the Millennium Development Goals. Regarding issue characteristics, data on the burden of surgical diseases are limited and public misperceptions surrounding the cost and complexity of surgery are widespread. However, the community has several strengths that portend well for the acquisition of political support. These include the existence of networks deeply committed to the cause, the potential to link with global health priorities, and emerging research on the cost-effectiveness of some procedures. To improve global priority for surgery, proponents will need to create an effective governance structure that facilitates achievement of collective goals, generate consensus on solutions, and find an effective public positioning of the issue that attracts political support. None.",
"Emergency medicine (EM) is a global discipline that provides secondary disease prevention and is also a tool for primary prevention. It is a horizontally integrated system of emergency care consisting of access to EM care; provision of EM care in the community and during transportation of patients; and provision of care at the receiving facility or hospital emergency department. EM can offer many tools to improve public health. These tools include primary disease prevention; interventions for addressing substance abuse and interpersonal violence; education about safety practices; epidemiological surveillance; enrolment of patients in clinical research trials focusing on acute interventions; education and clinical training of health-care providers; and participation in local and regional responses to natural and man-made disasters. Public health advocates and health policy-makers can benefit from the opportunities of EM and can help overcome its challenges. Advocating the establishment and recognition of the specialty of EM worldwide can result in benefits for health-care education, help in incorporating the full scope of EM care into the system of public health, and expand the capabilities of EM for primary and secondary prevention for the benefit of the health of the public.",
"The Ebola virus disease outbreak in West Africa was unprecedented in both its scale and impact. Out of this human calamity has come renewed attention to global health security--its definition, meaning, and the practical implications for programmes and policy. For example, how does a government begin to strengthen its core public health capacities, as demanded by the International Health Regulations? What counts as a global health security concern? In the context of the governance of global health, including WHO reform, it will be important to distil lessons learned from the Ebola outbreak. The Lancet invited a group of respected global health practitioners to reflect on these lessons, to explore the idea of global health security, and to offer suggestions for next steps. Their contributions describe some of the major threats to individual and collective human health, as well as the values and recommendations that should be considered to counteract such threats in the future. Many different perspectives are proposed. Their common goal is a more sustainable and resilient society for human health and wellbeing.",
"The international burden of injury is an increasing concern in global healthcare. Developed trauma care systems have reduced death and disability following injury. The ideal platform for surveillance and clinical governance in trauma care quality improvement is the trauma registry. There is a great disparity in the prevalence of active trauma registries between developed and developing countries. More detailed information on lessons learnt would guide those settings, hospitals and regions looking to establish a sustainable trauma registry. The aim of this study was to explore the experiences and perceptions of trauma registry custodians regarding the development of successful and sustainable trauma registries. This was a qualitative study using semi-structured interviews of trauma registry custodians. Trauma registries were selected from a wide range of jurisdictions, including single hospital and multi-hospital registries, based in developed and developing countries. Interview transcripts were analysed using thematic analysis; recurrent themes were identified, and a coding frame developed. Quotes were identified to illustrate the themes in the participants' own words. Twenty-seven interviews, representing 29 registries, were completed. Fourteen of the source registries were based in developed countries (6 single hospital, 8 multi-hospital) and 15 were based in developing countries (9 single hospital, 6 multi-hospital). The analysis generated 15 themes covering resources, data and strategies. The themes dealing with resources were: funding, staffing, information technology and tools for guidance. The themes dealing with data were: data quality, simplicity, injury coding and data utilisation. The themes dealing with strategies were: having a local champion and a clear purpose, stakeholder buy-in, governance, integration, getting started and persistence. For developing countries, the need for a local champion, dealing with data quality through prospective data collection, integration into local resources and keeping it simple were considered particularly important. The general consensus was that, for a trauma registry to be successful, in addition to adequate funding and trained staff, it needs to be led by a local champion with engagement of key local stakeholders. It should have a clear purpose, pay close attention to data quality and ensure that the data is well used.",
"Barriers to global emergency care development include a critical lack of data in several areas, including limited documentation of the acute disease burden, lack of agreement on essential components of acute care systems, and a lack of consensus on key analytic elements, such as diagnostic classification schemes and regionally appropriate metrics for impact evaluation. These data gaps obscure the profound health effects of lack of emergency care access in low- and middle-income countries (LMICs). As part of the Academic Emergency Medicine consensus conference \"Global Health and Emergency Care: A Research Agenda,\" a breakout group sought to develop a priority research agenda for data collection and management within global emergency care systems.",
"Quality emergency medical care is critical to reducing the burden of disease in low-income and middle-income countries (LMICs) and protecting the health of populations during disasters and epidemics. However, conducting research in emergency care settings in LMIC settings entails unique methodological and operational challenges. Therefore, new approaches and strategies that address these challenges need to be developed and will require increased attention from scientists, academic institutions and the global health research funding community. Research priorities to address emergency care in LMICs have also not been well defined, resulting in limited research output from LMICs. This manuscript frames the efforts of four multidisciplinary working groups, which were established under the auspices of the Fogarty International Center as part of the Collaborative on Enhancing Emergency Care Research in LMICs and serves as an introduction to this series, which identifies challenges and solutions in the context of emergency care research in LMICs. The objective of this introductory paper is to articulate the need for emergency care research in LMICs and underscore its future promise. We present public health arguments for greater investment in emergency care research, identify barriers to develop and conduct research, and present a list of research priorities for community organizations, academic institutions and funding agencies. We conclude that advances in emergency care research will be critical to achieve national and global health targets, such as the Sustainable Development Goals (SDGs), and to ensure that evidence informs how such research is best conducted.",
"Emergency medicine is one of the youngest recognized specialties in Nepal, and its growth in clinical practice and academic development has been challenging. In this paper, we reviewed the current state of emergency medicine in Nepal based on review of the literature, personal observations and experience, and interviews with many Nepali and foreign emergency physicians. Most hospitals in Nepal have adopted a multi-specialist approach, where emergency room physicians are primarily general practitioners/family physicians or house officers. As physicians are receiving their training via various pathways, national standards in training and certification have not been developed. As a result, the scope of practice for emergency physicians and the quality of care vary greatly among hospitals. Difficult working conditions, physician recruitment, compensation, and academic enrichment remain major challenges in the development of emergency medicine. For the sustainable development of this specialty, more international guidance and local leadership is needed to standardize the training curriculum, to provide adequate funding opportunities for academic development and to promote the overall development of the emergency care system."
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Colonization in Chronic Kidney Disease
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Chronic Kidney Disease (CKD) is a growing public-health concern worldwide. Patients exhibit compromised immunity and are more prone to infection than other populations. Therefore, oral colonization by clinically relevant members of the
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[
"Profiling phylogenetic marker genes, such as the 16S rRNA gene, is a key tool for studies of microbial communities but does not provide direct evidence of a community's functional capabilities. Here we describe PICRUSt (phylogenetic investigation of communities by reconstruction of unobserved states), a computational approach to predict the functional composition of a metagenome using marker gene data and a database of reference genomes. PICRUSt uses an extended ancestral-state reconstruction algorithm to predict which gene families are present and then combines gene families to estimate the composite metagenome. Using 16S information, PICRUSt recaptures key findings from the Human Microbiome Project and accurately predicts the abundance of gene families in host-associated and environmental communities, with quantifiable uncertainty. Our results demonstrate that phylogeny and function are sufficiently linked that this 'predictive metagenomic' approach should provide useful insights into the thousands of uncultivated microbial communities for which only marker gene surveys are currently available.",
"A 16S rRNA gene database (http://greengenes.lbl.gov) addresses limitations of public repositories by providing chimera screening, standard alignment, and taxonomic classification using multiple published taxonomies. It was found that there is incongruent taxonomic nomenclature among curators even at the phylum level. Putative chimeras were identified in 3% of environmental sequences and in 0.2% of records derived from isolates. Environmental sequences were classified into 100 phylum-level lineages in the Archaea and Bacteria.",
"An understanding of the relation of commensal microbiota to health is essential in preventing disease. Here we studied the oral microbial composition of children (N = 74, aged 3 - 18 years) in natural transition from their deciduous to a permanent dentition and related the microbial profiles to their oral health status. The microbial composition of saliva was assessed by barcoded pyrosequencing of the V5-V6 hypervariable regions of the 16 S rRNA, as well as by using phylogenetic microarrays. Pyrosequencing reads (126174 reads, 1045 unique sequences) represented 8 phyla and 113 higher taxa in saliva samples. Four phyla--Firmicutes, Bacteriodetes, Proteobacteria and Actinobacteria--predominated in all groups. The deciduous dentition harboured a higher proportion of Proteobacteria (Gammaproteobacteria, Moraxellaceae) than Bacteroidetes, while in all other groups Bacteroidetes were at least as abundant as Proteobacteria. Bacteroidetes (mainly genus Prevotella), Veillonellaceae family, Spirochaetes and candidate division TM7 increased with increasing age, reflecting maturation of the microbiome driven by biological changes with age. Microarray analysis enabled further analysis of the individual salivary microbiota. Of 350 microarray probes, 156 gave a positive signal with, on average, 77 (range 48-93) probes per individual sample. A caries-free oral status significantly associated with the higher signal of the probes targeting Porphyromonas catoniae and Neisseria flavescens. The potential role of P. catoniae and N. flavescens as oral health markers should be assessed in large-scale clinical studies. The combination of both open-ended and targeted molecular approaches provides us with information that will increase our understanding of the interplay between the human host and its microbiome.",
"Seventy children with chronic renal failure (CRF) aged 4-13.6 years were recruited from the Renal Unit of the Great Ormond Street Hospital for Children. Indices were recorded for dental caries, dental plaque, gingival inflammation, gingival enlargement, and enamel defects. Salivary urea, buffering capacity, and the oral streptococcal flora were determined for 25 of the children. A significantly greater proportion of the CRF children was caries free, 40% compared with 8.5% of the controls. The mean plaque score was significantly greater in the CRF group for both the primary 12.7 (16) and permanent dentition 22.0 (18.2) compared with the controls, 5.3 (7.6) and 15.5 (13.3), respectively. Eight CRF children had gingival enlargement. Enamel defects affecting the permanent teeth were observed in 57% of the CRF children compared with 33% of the controls. The buffering capacity was significantly greater in the CRF group, pH 6.4 (0.5) compared with the controls pH 5.6 (0.8). The mean salivary urea level (mmol/l) was significantly greater in the CRF children, 11.6 (5.9) compared with 3.6 (1.4) for the controls. The isolation frequency of Streptococcus mutans was significantly greater from controls compared with the CRF children ( P=0.002). An integrated dental service needs to be developed with emphasis on tooth brushing to prevent gingival hyperplasia and periodontal disease after puberty.",
"Growing evidence suggests the existence of new antibiotic resistance mechanisms. Recent studies have revealed that quorum-quenching enzymes, such as MacQ, are involved in both antibiotic resistance and cell-cell communication. Furthermore, some small bacterial regulatory RNAs, classified into RNA attenuators and small RNAs, modulate the expression of resistance genes. For example, small RNA sprX, can shape bacterial resistance to glycopeptide antibiotics via specific downregulation of protein SpoVG. Moreover, some bacterial lipocalins capture antibiotics in the extracellular space, contributing to severe multidrug resistance. But this defense mechanism may be influenced by Agr-regulated toxins and liposoluble vitamins. Outer membrane porin proteins and efflux pumps can influence intracellular concentrations of antibiotics. Alterations in target enzymes or antibiotics prevent binding to targets, which act to confer high levels of resistance in respiratory/oral bacteria. As described recently, horizontal gene transfer, including conjugation, transduction and transformation, is common in respiratory/oral microflora. Many conjugative transposons and plasmids discovered to date encode antibiotic resistance proteins and can be transferred from donor bacteria to transient recipient bacteria. New classes of mobile genetic elements are also being identified. For example, nucleic acids that circulate in the bloodstream (circulating nucleic acids) can integrate into the host cell genome by up-regulation of DNA damage and repair pathways. With multidrug resistant bacteria on the rise, new drugs have been developed to combate bacterial antibiotic resistance, such as innate defense regulators, reactive oxygen species and microbial volatile compounds. This review summaries various aspects and mechanisms of antibiotic resistance in the respiratory/oral microbiota. A better understanding of these mechanisms will facilitate minimization of the emergence of antibiotic resistance.",
"The dysbiosis of the oral microbiome is associated with both localized and systemic diseases. Modulating the resident microbial communities by the dietary consumption of probiotics has become an appealing means to promote host health by either restoring host-microbe balance or preventing dysbiosis. Most probiotics strategies target the intestinal microbiome, but little is known about their impact on the oral microbiome. We analyzed here the saliva microbiome from 21 volunteers, longitudinally collected before, during, and after consumption of a commercial probiotic and a standard yoghurt using 16S amplicon sequencing. The alpha diversity of the saliva microbiome had a statistically significant increase (P-value = 0.0011) in one of the groups that consumed the probiotic. The overall structure of the microbiome was however not significantly impacted by the probiotic, although oligotyping analysis revealed that both Streptococci and Lactobacilli present in the probiotic product persisted in the saliva microbiome. In contrast, non-probiotic yoghurt consumption had a lesser impact on the overall diversity and Lactobacillus and Streptococcus persistence. Our results suggest that consumption of commercial probiotics in healthy subjects increase the overall diversity of the oral cavity microbiome in the short term, but such dietary interventions are not able to substantially modify the structure of the microbiome.",
"This article reviews the current understanding of the oral and dental aspects of chronic renal disease (CRD). A PubMed literature search was performed and all relevant studies were assessed. As the number of people suffering from CRD increases worldwide, dentists are expected to encounter more patients with CRD who need oral care. In children, CRD can elicit a wide spectrum of oral manifestations in the hard and soft tissues. Bleeding, altered drug metabolism, impaired immune function, and an increased risk of dentally induced bacterial endocarditis are some important features that require attention. Dental management of patients with CRD requires that clinicians appreciate that multiple systems can be affected by the disease. Dentists should consult with nephrologists regarding the specific precautions required for each patient. Medical treatments in these patients may need to be postponed due to an unfavorable oral health status or potential risk of life-threatening infection after surgery. Improving oral hygiene and performing necessary dental and oral treatment before hemodialysis or transplantation may prevent endocarditis and septicemia in these patients. Hence, treatment plans should be formulated to restore the patient's dentition and protect them from potentially severe infections of dental origin.",
"Poor oral health has broad consequences that can be seen at personal as well as societal levels, especially in developing countries like India. We have limited information on the healthy oral cavity's inhabitant microorganisms that play a crucial role in overall oral health. In a comprehensive culture-independent approach, the bacterial composition of healthy human oral cavities was determined from a sub-population of northern India. During this study, 20 mouthwash-derived metagenomes were explored for identifying bacterial diversity using the 16S rRNA hypervariable V3 region with the MiSeq Illumina platform. On the taxonomy assignment of operational taxonomic units (OTUs), 20 assigned phyla and 162 genera were recovered among the participants. The mean relative abundance revealed that Streptococcus was the dominant genera among the participants. However, at inter-individual analysis, Neisseria and Haemophilus exhibited first-order dominance among five and three healthy individuals, respectively. Correlation studies indicate that Streptococcus shares a strong relationship with Rothia, Corynebacterium, Prevotella, and Veillonella, whereas it was negatively correlated with Neisseria, Aggregatibacter, Porphyromonas, and Fusobacteria like Gram-negative bacteria. Bacterial diversity showed insignificant differences at the level of age and gender within and between the participants. The results support several of the major findings of previous reports on the healthy oral microbiome of the Indian population, however, the present investigation further illustrates that demographic region leaves an impact on overall bacterial composition. The study will assist in a better understanding of the oral microbiome from region-specific Indian population that was otherwise highly under-represented.",
"Population-based epidemiologic studies can provide important insight regarding the role of the microbiome in human health and disease. Buccal cells samples using commercial mouthwash have been obtained in large prospective cohorts for the purpose of studying human genomic DNA. We aimed to better understand if these mouthwash samples are also a valid resource for the study of the oral microbiome. We collected one saliva sample and one Scope mouthwash sample from 10 healthy subjects. Bacterial 16S rRNA genes from both types of samples were amplified, sequenced, and assigned to bacterial taxa. We comprehensively compared these paired samples for bacterial community composition and individual taxonomic abundance. We found that mouthwash samples yielded similar amount of bacterial DNA as saliva samples (p from Student's t-test for paired samples = 0.92). Additionally, the paired samples had similar within sample diversity (p from = 0.33 for richness, and p = 0.51 for Shannon index), and clustered as pairs for diversity when analyzed by unsupervised hierarchical cluster analysis. No significant difference was found in the paired samples with respect to the taxonomic abundance of major bacterial phyla, Bacteroidetes, Firmicutes, Proteobacteria, Fusobacteria, and Actinobacteria (FDR adjusted q values from Wilcoxin signed-rank test = 0.15, 0.15, 0.87, 1.00 and 0.15, respectively), and all identified genera, including genus Streptococcus (q = 0.21), Prevotella (q = 0.25), Neisseria (q = 0.37), Veillonella (q = 0.73), Fusobacterium (q = 0.19), and Porphyromonas (q = 0.60). These results show that mouthwash samples perform similarly to saliva samples for analysis of the oral microbiome. Mouthwash samples collected originally for analysis of human DNA are also a resource suitable for human microbiome research.",
"More than 700 bacterial species or phylotypes, of which over 50% have not been cultivated, have been detected in the oral cavity. Our purposes were (i) to utilize culture-independent molecular techniques to extend our knowledge on the breadth of bacterial diversity in the healthy human oral cavity, including not-yet-cultivated bacteria species, and (ii) to determine the site and subject specificity of bacterial colonization. Nine sites from five clinically healthy subjects were analyzed. Sites included tongue dorsum, lateral sides of tongue, buccal epithelium, hard palate, soft palate, supragingival plaque of tooth surfaces, subgingival plaque, maxillary anterior vestibule, and tonsils. 16S rRNA genes from sample DNA were amplified, cloned, and transformed into Escherichia coli. Sequences of 16S rRNA genes were used to determine species identity or closest relatives. In 2,589 clones, 141 predominant species were detected, of which over 60% have not been cultivated. Thirteen new phylotypes were identified. Species common to all sites belonged to the genera Gemella, Granulicatella, Streptococcus, and Veillonella. While some species were subject specific and detected in most sites, other species were site specific. Most sites possessed 20 to 30 different predominant species, and the number of predominant species from all nine sites per individual ranged from 34 to 72. Species typically associated with periodontitis and caries were not detected. There is a distinctive predominant bacterial flora of the healthy oral cavity that is highly diverse and site and subject specific. It is important to fully define the human microflora of the healthy oral cavity before we can understand the role of bacteria in oral disease."
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Influence of Crack on Plaque Rupture: A Finite Element Numerical Simulation Study
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Atherosclerotic plaque rupture is the main cause of many cardiovascular diseases, and biomechanical factors play an important role in the process of plaque rupture. In the study of plaque biomechanics, there are relatively few studies based on fatigue fracture failure theory, and most of them mainly focus on the whole fatigue propagation process from crack initiation to plaque rupture, while there are few studies on the influence of crack on plaque rupture at a certain time in the process of fatigue propagation. In this paper, a two-dimensional plaque model with crack was established. Based on the theory of fracture mechanics and combined with the finite element numerical simulation method, the stress intensity factor (SIF) and related influencing factors at the crack tip in the plaque were studied. The SIF was used to measure the influence of crack on plaque rupture. The results show that the existence of crack can lead to local stress concentration, which increases the risk of plaque rupture. The SIF at the crack tip in the plaque was positively correlated with blood pressure, but negatively correlated with fibrous cap thickness and lipid pool stiffness. The effect of the thickness and angle of lipid pool on the SIF at the crack tip in the plaque was less than 4%, which could be ignored. This study provides a theoretical basis for the risk assessment of plaque rupture with cracks.
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[
"A 'thin' fibrous cap atheroma is the typical morphological characteristic of vulnerable plaque. Yet the very pathological studies that have provided these descriptions have also shown the actual prediction of plaque rupture to be rather less exact. Other relevant characteristics must be involved in the mechanisms of plaque rupture--blood pressure loading (P) and the material properties of the soft atheromatous core--as predictors of the distribution of the peak circumferential stress (PCS) locations. We used a computational structural analysis based on three typical in-vivo intravascular ultrasound images of fibrous cap atheroma in which we decreased the cap thickness (CTh). With different soft atheromatous core Young's moduli (Ecore), 414 simulations were performed under eight different physiological loading blood pressures. The transition from plaque stability to plaque instability was defined by a threshold of 300 kPa and is a feature of vulnerability. It was found that (1) irrespective of plaque geometry and composition, CTh < 60 microm exposed the plaque to PCSs in excess of 300 kPa; (2) the exponential variations in PCS with change in CTh and Ecore values show that very slight structural changes are enough to tilt a vulnerable plaque from stability to instability or vice versa; and (3) the relationship between P and PCS is proportional with P acting as trigger or as protector. The present study shows why, in clinical practice, mere morphological detection by imaging techniques of thin-cap fibro-atheroma is not in itself enough for the prediction of future rupture.",
"A cohesive zone model (CZM) approach is applied to simulate atherosclerotic plaque delamination experiments in mouse abdominal aorta specimens. A three-dimensional finite element model is developed for the experiments. The aortic wall is treated as a fiber-reinforced, highly deformable, incompressible material, and the Holzapfel-Gasser-Ogden (HGO) model is adopted for the aortic bulk material behavior. Cohesive elements are placed along the plaque-media interface along which delamination occurs. The 3D specimen geometry is created based on images from the experiments and certain simplifying approximations. A set of HGO and CZM parameter values is determined based on values suggested in the literature and through matching simulation predictions of the load vs. load-point displacement curve with experimental measurements for one loading-delamination-unloading cycle. Using this set of parameter values, simulation predictions for four other loading-delamination-unloading cycles are obtained, which show good agreement with experimental measurements. The findings of the current study demonstrate the applicability of the CZM approach in arterial tissue failure simulations.",
"Fibrous cap delamination is a critical process during the rupture of atherosclerotic plaque, which often leads to severe life-threatening clinical consequences such as myocardial infarction or stroke. In this study a finite element modeling and simulation approach is presented that enables the study of fibrous cap delamination experiments for the purpose of understanding the fibrous cap delamination process. A cohesive zone model (CZM) approach is applied to simulate delamination of the fibrous cap from the underlying plaque tissue. A viscoelastic anisotropic (VA) model for the bulk arterial material behavior is extended from existing studies so that the hysteresis phenomenon observed in the fibrous cap delamination experiments can be captured. A finite element model is developed for the fibrous cap delamination experiments, in which arterial layers (including the fibrous cap and the underlying plaque tissue) are represented by solid elements based on the VA model and the fibrous cap-underlying plaque tissue interface is characterized by interfacial CZM elements. In the CZM, the delamination process is governed by an exponential traction-separation law which utilizes critical energy release rates obtained directly from the fibrous cap delamination experiments. A set of VA model parameter values and CZM parameter values is determined based on values suggested in the literature and through matching simulation predictions of the load vs. load-point displacement curve with one set of experimental measurements. Using this set of parameter values, simulation predictions for other sets of experimental measurements are obtained and good agreement between simulation predictions and experimental measurements is observed. Results of this study demonstrate the applicability of the viscoelastic anisotropic model and the CZM approach for the simulation of diseased arterial tissue failure processes."
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The complete pathway for bacterial ceramide synthesis
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The bacterial domain produces numerous types of sphingolipids with various physiological functions. In the human microbiome, commensal and pathogenic bacteria use these lipids to modulate the host inflammatory system. Despite their growing importance, their biosynthetic pathway remains undefined since several key eukaryotic ceramide synthesis enzymes have no bacterial homolog. Here we used genomic and biochemical approaches to identify six proteins comprising the complete pathway for bacterial ceramide synthesis. Bioinformatic analyses revealed the widespread potential for bacterial ceramide synthesis leading to our discovery of a Gram-positive species that produces ceramides. Biochemical evidence demonstrated that the bacterial pathway operates in a different order from that in eukaryotes. Furthermore, phylogenetic analyses support the hypothesis that the bacterial and eukaryotic ceramide pathways evolved independently.
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[
"Labeled C18-phytosphingosine was detected in a glyco-lipid-rich fraction from rat intestine 4 and 24 h after injection of D-erythro[4,5-3H]C18-dihydrosphingosine. The criteria for identification were: (a) co-migration on gas-liquid chromatography of the trimethylsilyl ethers of the free and N-acetylated base with authentic standard, (b) co-migration on thin layer chromatography of the dinitrophenyl derivative with authentic standard, (c) labeled pentadecanol detection by gas-liquid radiochromatography after subjecting intact sphingolipids or a mixture of free long chain bases to periodate oxidation followed by sodium borohydride reduction, and (d) obtainment of the same fragmentation pattern as with authentic standard when the trimethylsilyl ethers of the N-acetylated base were subjected to gas-liquid chromatography-mass spectrometry. Smaller amounts of labeled C18-phytosphingosine were also detected in ceramide- and sphingomyelin-rich fractions of intestine and in liver and kidney. Experiments with germ-free rats strongly suggest that the intestinal flora is not the (sole) site for the biosynthesis of C18-phytosphingosine.",
"The diversity of cell shapes across the bacterial kingdom reflects evolutionary pressures that have produced physiologically important morphologies. While efforts have been made to understand the regulation of some prototypical cell morphologies such as that of rod-shaped Escherichia coli, little is known about most cell shapes. For Caulobacter crescentus, polar stalk synthesis is tied to its dimorphic life cycle, and stalk elongation is regulated by phosphate availability. Based on the previous observation that C. crescentus stalks are lysozyme-resistant, we compared the composition of the peptidoglycan cell wall of stalks and cell bodies and identified key differences in peptidoglycan crosslinking. Cell body peptidoglycan contained primarily DD-crosslinks between meso-diaminopimelic acid and D-alanine residues, whereas stalk peptidoglycan had more LD-transpeptidation (meso-diaminopimelic acid-meso-diaminopimelic acid), mediated by LdtD. We determined that ldtD is dispensable for stalk elongation; rather, stalk LD-transpeptidation reflects an aging process associated with low peptidoglycan turnover in the stalk. We also found that lysozyme resistance is a structural consequence of LD-crosslinking. Despite no obvious selection pressure for LD-crosslinking or lysozyme resistance in C. crescentus, the correlation between these two properties was maintained in other organisms, suggesting that DAP-DAP crosslinking may be a general mechanism for regulating bacterial sensitivity to lysozyme.",
"Sphingolipid biosynthesis begins with the condensation of L-serine and palmitoyl-CoA catalyzed by the PLP-dependent enzyme serine palmitoyltransferase (SPT). Mutations in human SPT cause hereditary sensory autonomic neuropathy type 1, a disease characterized by loss of feeling in extremities and severe pain. The human enzyme is a membrane-bound hetereodimer, and the most common mutations are located in the enzymatically incompetent monomer, suggesting a \"dominant\" or regulatory effect. The molecular basis of how these mutations perturb SPT activity is subtle and is not simply loss of activity. To further explore the structure and mechanism of SPT, we have studied the homodimeric bacterial enzyme from Sphingomonas paucimobilis. We have analyzed two mutants (N100Y and N100W) engineered to mimic the mutations seen in hereditary sensory autonomic neuropathy type 1 as well as a third mutant N100C designed to mimic the wild-type human SPT. The N100C mutant appears fully active, whereas both N100Y and N100W are significantly compromised. The structures of the holoenzymes reveal differences around the active site and in neighboring secondary structure that transmit across the dimeric interface in both N100Y and N100W. Comparison of the l-Ser external aldimine structures of both native and N100Y reveals significant differences that hinder the movement of a catalytically important Arg(378) residue into the active site. Spectroscopic analysis confirms that both N100Y and N100W mutants subtly affect the chemistry of the PLP. Furthermore, the N100Y and R378A mutants appear less able to stabilize a quinonoid intermediate. These data provide the first experimental insight into how the most common disease-associated mutations of human SPT may lead to perturbation of enzyme activity.",
"Membrane fatty acid desaturases are responsible for inserting double bonds into specific positions in fatty acids. We have cloned a new member of the human membrane fatty acid (lipid) desaturase gene family, MLD. The derived amino acid sequence of MLD contains three consensus motifs, HX3H, HX2HH, and HX2HHXFP, that are characteristic of a group of membrane fatty acid desaturases. MLD is predicted to be a multiple membrane-spanning protein and is found to be extractable from particulate fractions with detergent but not salt or urea. MLD is widely expressed in human tissues and is localized to the endoplasmic reticulum. Cotransfection of MLD with the epidermal growth factor (EGF) receptor resulted in decreased expression of the receptor but did not affect platelet-derived growth factor receptor expression. MLD overexpression inhibited biosynthesis of the EGF receptor, suggesting a possible role of a fatty acid desaturase in regulating biosynthetic processing of the EGF receptor.",
"The introduction of the double bond in the sphingoid backbone of sphingolipids occurs at the level of dihydroceramide via an NADPH-dependent desaturase, as discovered in permeabilized rat hepatocytes. In the rat, the enzyme activity, which has now been further characterized, appeared to be mostly enriched in liver and Harderian gland. By means of subcellular fractionation of rat liver homogenates and density gradient separation of microsomal fractions, the desaturase was localized to the endoplasmic reticulum. Various detergents were inhibitory to the enzyme, and maximal activities were obtained in the presence of NADPH and when the substrate was complexed to albumin. In the presence of albumin, the chain length of the fatty acid of the truncated dihydroceramides hardly affected the activity. Finally, in view of a likely evolutionary relationship between desaturases and hydroxylases, the formation of hydroxylated intermediates was analyzed. No evidence for their presence was found under our assay conditions.",
"The C-4 hydroxylation of sphinganine and dihydroceramide is a rate-limiting reaction in the biosynthesis of phytosphingolipids. Mouse DES1 (MDES1) cDNA homologous to the Drosophila melanogaster degenerative spermatocyte gene-1 (des-1) cDNA leads to sphingosine Delta4-desaturase activity, and another mouse homologue, MDES2, has bifunctional activity, producing C-4 hydroxysphinganine and Delta4-sphingenine in yeast [Ternes, Franke, Zahringer, Sperling and Heinz (2002) J. Biol. Chem. 277, 25512-25518]. Here, we report the characterization of mouse DES2 (MDES2) using an in vitro assay with a homogenate of COS-7 cells transfected with MDES2 cDNA and N -octanoyl-sphinganine and sphinganine as substrates. MDES2 protein prefers dihydroceramide as a substrate to sphinganine, and exhibits dihydroceramide Delta4-desaturase and C-4 hydroxylase activities. MDES2 mRNA content was high in the small intestine and abundant in the kidney. In situ hybridization detected signals of MDES2 mRNA in the crypt cells. Immunohistochemistry using an anti-MDES2 peptide antibody stained the crypt cells and the adjacent epithelial cells. These results suggest that MDES2 is the dihydroceramide C-4 hydroxylase responsible for the biosynthesis of enriched phytosphingoglycolipids in the microvillous membranes of intestinal epithelial cells.",
"In mammalian cells, there are two major classes of sphingolipids---sphingomyelin and glycosphingolipids (GSLs)--both of which are synthesized from the hydrophobic molecule ceramide. The synthesis of most GSLs begins with glucosylation of ceramide to form glucosylceramide (GlcCer), which, in turn, serves as the source of 300-400 GSLs. Although most of these GSLs have been characterized chemically, the biological functions of ceramide glycosylation and GSLs still remain enigmatic. The recent description of a GSL-deficient cell line and isolation of cDNA for GlcCer synthase provide new insights into GSL functions."
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MORC2 O-GlcNAcylation Is Required for Transforming Growth Factor-1-Induced Breast Cancer Cell Migration and Invasion
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MORC family CW-type zinc finger 2 (MORC2) is a newly identified chromatin-remodeling enzyme involved in DNA damage response and gene transcription, and its dysregulation has been linked with Charcot-Marie-Tooth disease, neurodevelopmental disorder, and cancer. Despite its functional importance, how MORC2 is regulated remains enigmatic. Here, we report that MORC2 is O-GlcNAcylated by O-GlcNAc transferase (OGT) at threonine 556. Mutation of this site or pharmacological inhibition of OGT impairs MORC2-mediated breast cancer cell migration and invasion in vitro and lung colonization in vivo. Moreover, transforming growth factor-β1 (TGF-β1) induces MORC2 O-GlcNAcylation through enhancing the stability of glutamine-fructose-6-phosphate aminotransferase (GFAT), the rate-limiting enzyme for producing the sugar donor for OGT. O-GlcNAcylated MORC2 is required for transcriptional activation of TGF-β1 target genes connective tissue growth factor (CTGF) and snail family transcriptional repressor 1 (SNAIL). In support of these observations, knockdown of GFAT, SNAIL or CTGF compromises TGF-β1-induced, MORC2 O-GlcNAcylation-mediated breast cancer cell migration and invasion. Clinically, high expression of OGT, MORC2, SNAIL, and CTGF in breast tumors is associated with poor patient prognosis. Collectively, these findings uncover a previously unrecognized mechanistic role for MORC2 O-GlcNAcylation in breast cancer progression and provide evidence for targeting MORC2-dependent breast cancer through blocking its O-GlcNAcylation.
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[
"It has been reported that epithelial-mesenchymal transition (EMT) mediates multiple physiological and pathological processes. However, the occurrence and the pathogenic role of high glucose-induced EMT in retinal pigment epithelial cells (RPE cells) is unknown. The aim of this study was to examine the effects of high glucose on EMT in RPE cells. Cultured RPE cells were exposed to 25 mM D-glucose. A vector encoding the Snail gene and siRNA targeting Snail (Snail siRNA) were transfected into the cells to induce the overexpression or silencing of Snail, respectively. AKT and extracellular signal-regulated kinase (ERK) inhibitors were used to block the activation of AKT and ERK, respectively. The levels of EMT markers, fibrogenic factors, phosphorylated ERK and phosphorylated AKT were determined by western blot analysis and immunofluorescence staining. Cell migration was evaluated by wound healing assay. Our results revealed that high glucose elevated the expression of the key EMT transcriptional factor, Snail, and that of other mesenchymal makers, and promoted cell migration. Moreover, the overexpression of Snail elevated the levels of fibronectin and connective tissue growth factor (CTGF), whereas the silencing of Snail decreased the expression of fibronectin and CTGF induced by high glucose in the cells. Mechanistically, the AKT inhibitor (AKT inhibitor IV) and ERK inhibitor (U0126) significantly decreased the expression of Snail, as well as the levels of fibronectin and CTGF which were induced by high glucose. On the whole, and to the best of our knowedge, the present study is the first to demonstrate the upregulation of mesenchymal markers in RPE cells induced by high glucose, and suggest that mesenchymal transition may be involved in the pathological processes of retinal diseases.",
"Increased flux through the hexosamine biosynthetic pathway with glutamine:fructose-6-phosphate aminotransferase (GFAT) as a rate-limiting enzyme has been linked to the enhanced bioactivity of the prosclerotic cytokine TGF-beta1, a key mediator in the development of diabetic nephropathy and possibly other diabetic angiopathies. In this study we investigated the effect of enhanced expression of wild-type GFAT and two enzymatically inactive GFAT mutants on TGF-beta1 synthesis in mesangial cells. Mutated human GFAT expression vectors were prepared by PCR-site directed mutagenesis. Wild-type and mutated vectors were transfected into human embryonic kidney 293 cells and mesangial cells and GFAT enzyme activity was assessed by formation of glucosamine-6-phosphate. Production of TGF-beta1 and fibronectin protein was examined by ELISA. Mutation of histidine 577 or lysine 676 to alanine led to a complete loss of GFAT enzyme activity. An increased concentration of wild-type GFAT in mesangial cells enhanced both TGF-beta1 and fibronectin production 1.5-fold, while mesangial cells transfected with the mutated GFAT constructs showed no effect. The data indicate that the hexosamine pathway-mediated induction of TGF-beta1 synthesis in mesangial cells is dependent on GFAT enzyme activity. Our results add to previous observations showing that the hexosamine pathway could increase the transcriptional activity of nuclear proteins leading to enhanced cytokine synthesis.",
"Microrchidia 2 (MORC2) is important in DNA damage repair and lipogenesis, however, the clinical and functional role of MORC2 in liver cancer remains to be fully elucidated. The aim the present study was to clarify the role of MORC2 in liver cancer. Expression profile analysis, immunohistochemical staining, reverse transcription-quantitative polymerase chain reaction analysis and western blot analysis were performed to evaluate the levels of MORC2 in liver cancer patient specimens and cell lines; subsequently the expression of MORC2 was suppressed or increased in liver cancer cells and the effects of MORC2 on the cancerous transformation of liver cancer cells were examined in vitro and in vivo. MORC2 was upregulated in liver cancer tissues, and the upregulation was associated with certain clinicopathologic features of patients with liver cancer. MORC2 knockdown caused marked inhibition of liver cancer cell proliferation and clonogenicity, whereas the overexpression of MORC2 substantially promoted liver cancer cell proliferation. In addition, the knockdown of MORC2 inhibited the migratory and invasive ability of liver cancer cells, whereas increased migration and invasion rates were observed in cells with ectopic expression of MORC2. In a model of nude mice, the overexpression of MORC2 promoted tumorigenicity and markedly enhanced pulmonary metastasis of liver cancer. Furthermore, MORC2 regulated apoptosis and its expression level had an effect on the sensitivity of liver cancer cells to doxorubicin, 5-fluorouracil and cisplatin. Mechanically, MORC2 modulated the mitochondrial apoptotic pathway, possibly in a p53-dependent manner, and its dysregulation also resulted in the abnormal activation of the Hippo pathway. For the first time, to the best of our knowledge, the present study confirmed that MORC2 was a novel oncogene in liver cancer. These results provide useful insight into the mechanism underlying the tumorigenesis and progression of liver cancer, and offers clues into potential novel liver cancer therapies.",
"Transposable elements (TEs) and DNA repeats are commonly targeted by DNA and histone methylation to achieve epigenetic gene silencing. We isolated mutations in two Arabidopsis genes, AtMORC1 and AtMORC6, which cause derepression of DNA-methylated genes and TEs but no losses of DNA or histone methylation. AtMORC1 and AtMORC6 are members of the conserved Microrchidia (MORC) adenosine triphosphatase (ATPase) family, which are predicted to catalyze alterations in chromosome superstructure. The atmorc1 and atmorc6 mutants show decondensation of pericentromeric heterochromatin, increased interaction of pericentromeric regions with the rest of the genome, and transcriptional defects that are largely restricted to loci residing in pericentromeric regions. Knockdown of the single MORC homolog in Caenorhabditis elegans also impairs transgene silencing. We propose that the MORC ATPases are conserved regulators of gene silencing in eukaryotes."
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what is polycystic ovary syndrome
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Polycystic ovary syndrome (PCOS) is characterized by ovulatory dysfunction and hyperandrogenism and can be associated with cardiometabolic dysfunction, but it remains unclear which of these features are inciting causes and which are secondary consequences.
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[
"Because hair loss may be a surrogate measure of androgenic activity-possibly a determinant of coronary atherosclerosis-several studies have explored the presence and magnitude of an association between male pattern baldness and myocardial infarction (MI). In particular, vertex baldness, but not frontal baldness alone, was strongly associated with incident MI in a large, hospital-based, case-control study. The authors examined these associations in a cross-sectional sample of 5,056 men aged 52-75 years, of whom 767 had a history of MI. The sample was derived from the Atherosclerosis Risk in Communities (ARIC) Study (1987-1998). As compared with a baldness-free reference group, the estimated odds ratios for prevalent MI from a multivariable model were 1.28 (frontal baldness), 1.02 (mild vertex baldness), 1.40 (moderate vertex baldness), and 1.18 (severe vertex baldness). Other regression models have yielded similar results, including the absence of a monotonic \"dose-response relation\" between the extent of vertex baldness and prevalent MI. The authors also examined the relation of baldness pattern to carotid intimal-medial thickness, a measure of atherosclerosis, among those who were free of clinical cardiovascular disease. The estimated mean differences in carotid intimal-medial thickness between groups of men with various types of baldness and their baldness-free counterparts were all close to zero. The results of this study suggest that male pattern baldness is not a surrogate measure of an important risk factor for myocardial infarction or asymptomatic atherosclerosis.",
"Male pattern baldness (MPB) or androgenetic alopecia is one of the most common conditions affecting men, reaching a prevalence of ~50% by the age of 50; however, the known genes explain little of the heritability. Here, we present the results of a genome-wide association study including more than 70,000 men, identifying 71 independently replicated loci, of which 30 are novel. These loci explain 38% of the risk, suggesting that MPB is less genetically complex than other complex traits. We show that many of these loci contain genes that are relevant to the pathology and highlight pathways and functions underlying baldness. Finally, despite only showing genome-wide genetic correlation with height, pathway-specific genetic correlations are significant for traits including lifespan and cancer. Our study not only greatly increases the number of MPB loci, illuminating the genetic architecture, but also provides a new approach to disentangling the shared biological pathways underlying complex diseases.",
"Male-pattern baldness (MPB) is a common and highly heritable trait characterized by androgen-dependent, progressive hair loss from the scalp. Here, we carry out the largest GWAS meta-analysis of MPB to date, comprising 10,846 early-onset cases and 11,672 controls from eight independent cohorts. We identify 63 MPB-associated loci (P<5 × 10-8, METAL) of which 23 have not been reported previously. The 63 loci explain ∼39% of the phenotypic variance in MPB and highlight several plausible candidate genes (FGF5, IRF4, DKK2) and pathways (melatonin signalling, adipogenesis) that are likely to be implicated in the key-pathophysiological features of MPB and may represent promising targets for the development of novel therapeutic options. The data provide molecular evidence that rather than being an isolated trait, MPB shares a substantial biological basis with numerous other human phenotypes and may deserve evaluation as an early prognostic marker, for example, for prostate cancer, sudden cardiac arrest and neurodegenerative disorders.",
"The many functional partnerships and interactions that occur between proteins are at the core of cellular processing and their systematic characterization helps to provide context in molecular systems biology. However, known and predicted interactions are scattered over multiple resources, and the available data exhibit notable differences in terms of quality and completeness. The STRING database (http://string-db.org) aims to provide a critical assessment and integration of protein-protein interactions, including direct (physical) as well as indirect (functional) associations. The new version 10.0 of STRING covers more than 2000 organisms, which has necessitated novel, scalable algorithms for transferring interaction information between organisms. For this purpose, we have introduced hierarchical and self-consistent orthology annotations for all interacting proteins, grouping the proteins into families at various levels of phylogenetic resolution. Further improvements in version 10.0 include a completely redesigned prediction pipeline for inferring protein-protein associations from co-expression data, an API interface for the R computing environment and improved statistical analysis for enrichment tests in user-provided networks."
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Ablation and Immunotherapy for Hepatocellular Carcinoma
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Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related deaths worldwide and is increasing in incidence. Local ablative therapy plays a leading role in HCC treatment. Radiofrequency (RFA) is one of the first-line therapies for early local ablation. Other local ablation techniques (e.g., microwave ablation, cryoablation, irreversible electroporation, phototherapy.) have been extensively explored in clinical trials or cell/animal studies but have not yet been established as a standard treatment or applied clinically. On the one hand, single treatment may not meet the needs. On the other hand, ablative therapy can stimulate local and systemic immune effects. The combination strategy of immunotherapy and ablation is reasonable. In this review, we briefly summarized the current status and progress of ablation and immunotherapy for HCC. The immune effects of local ablation and the strategies of combination therapy, especially synergistic strategies based on biomedical materials, were discussed. This review is hoped to provide references for future researches on ablative immunotherapy to arrive to a promising new era of HCC treatment.
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[
"Tremelimumab is a monoclonal antibody that blocks cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4), an inhibitory co-receptor that interferes with T cell activation and proliferation. The purpose of this pilot clinical trial was to test the antitumor and antiviral effect of tremelimumab in patients with hepatocellular carcinoma (HCC) and chronic hepatitis C virus (HCV) infection; and to study the safety of its administration to cirrhotic patients. Tremelimumab at a dose of 15 mg/kg IV every 90 days was administered until tumor progression or severe toxicity. Twenty patients were assessable for toxicity and viral response and 17 were assessable for tumor response. Most patients were in the advanced stage and 43% had an altered liver function (Child-Pugh class B). A good safety profile was recorded and no patient needed steroids because of severe immune-mediated adverse events. Some patients had a transient albeit intense elevation of transaminases after the first dose, but not following subsequent cycles. Partial response rate was 17.6% and disease control rate was 76.4%. Time to progression was 6.48 months (95% CI 3.95-9.14). A significant drop in viral load was observed while new emerging variants of the hypervariable region 1 of HCV replaced the predominant variants present before therapy, particularly in those patients with a more prominent drop in viral load. This antiviral effect was associated with an enhanced specific anti-HCV immune response. Tremelimumab safety profile and antitumor and antiviral activity, in patients with advanced HCC developed on HCV-induced liver cirrhosis, support further investigation.",
"Dendritic cell (DC)-based immunotherapies are believed to help eradicate residual tumor cells, including hepatocellular carcinoma (HCC). Here, we assessed the safety and clinical response to OK432-stimulated monocyte-derived DCs (MoDCs) in treating HCC after radiofrequency ablation (RFA). MoDCs were derived from 30 HCC patients in the presence of interleukin-4 and granulocyte-macrophage colony stimulating factor for 5 days and then cultured for 2 more days in the medium (basic protocol) or stimulated with OK432. On day 7, DCs were harvested and percutaneously injected into HCC tumors after RFA. We observed no grade 3 or 4 National Cancer Institute Common Toxicity Criteria adverse events. Kaplan-Meier analysis indicated that patients treated with RFA + OK432-stimulated DCs transfer had longer recurrence-free survival than those treated with RFA + basic-protocol DCs (median: 24.8 vs 13.0 months; P = .003). RFA with DC infusion can enhance various tumor-associated antigen (TAA)-specific T-cell responses. Additionally, the 5-year RFS rate for patients with significantly increased TAA-specific T-cell responses was much higher than for other patients (50.0% vs. 7.7%; P = .030). Our study provides useful information for development of HCC immunotherapies (trial registration: UMIN000001701).",
"No adjuvant therapy has been shown to extend the survival of patients with hepatocellular carcinoma (HCC) receiving curative treatment. We investigated whether injections of activated cytokine-induced killer (CIK) cells (CD3+/CD56+ and CD3+/CD56- T cells and CD3-/CD56+ natural killer cells) prolongs recurrence-free survival of patients after curative therapy for HCC. We performed a multicenter, randomized, open-label, phase 3 trial of the efficacy and safety of adjuvant immunotherapy with activated CIK cells (created by incubation of patients' peripheral blood mononuclear cells with interleukin 2 and an antibody against CD3). The study included 230 patients with HCC treated by surgical resection, radiofrequency ablation, or percutaneous ethanol injection at university-affiliated hospitals in Korea. Patients were assigned randomly to receive immunotherapy (injection of 6.4 × 10(9) autologous CIK cells, 16 times during 60 weeks) or no adjuvant therapy (controls). The primary end point was recurrence-free survival; secondary end points included overall survival, cancer-specific survival, and safety. The median time of recurrence-free survival was 44.0 months in the immunotherapy group and 30.0 months in the control group (hazard ratio with immunotherapy, 0.63; 95% confidence interval [CI], 0.43-0.94; P = .010 by 1-sided log-rank test). Hazard ratios also were lower in the immunotherapy than in the control group for all-cause death (0.21; 95% CI, 0.06-0.75; P = .008) and cancer-related death (0.19; 95% CI, 0.04-0.87; P = .02). A significantly higher proportion of patients in the immunotherapy group than in the control group had an adverse event (62% vs 41%; P = .002), but the proportion of patients with serious adverse events did not differ significantly between groups (7.8% vs 3.5%; P = .15). In patients who underwent curative treatment for HCC, adjuvant immunotherapy with activated CIK cells increased recurrence-free and overall survival. ClinicalTrials.gov number: NCT00699816.",
"The combination of atezolizumab and bevacizumab showed encouraging antitumor activity and safety in a phase 1b trial involving patients with unresectable hepatocellular carcinoma. In a global, open-label, phase 3 trial, patients with unresectable hepatocellular carcinoma who had not previously received systemic treatment were randomly assigned in a 2:1 ratio to receive either atezolizumab plus bevacizumab or sorafenib until unacceptable toxic effects occurred or there was a loss of clinical benefit. The coprimary end points were overall survival and progression-free survival in the intention-to-treat population, as assessed at an independent review facility according to Response Evaluation Criteria in Solid Tumors, version 1.1 (RECIST 1.1). The intention-to-treat population included 336 patients in the atezolizumab-bevacizumab group and 165 patients in the sorafenib group. At the time of the primary analysis (August 29, 2019), the hazard ratio for death with atezolizumab-bevacizumab as compared with sorafenib was 0.58 (95% confidence interval [CI], 0.42 to 0.79; P<0.001). Overall survival at 12 months was 67.2% (95% CI, 61.3 to 73.1) with atezolizumab-bevacizumab and 54.6% (95% CI, 45.2 to 64.0) with sorafenib. Median progression-free survival was 6.8 months (95% CI, 5.7 to 8.3) and 4.3 months (95% CI, 4.0 to 5.6) in the respective groups (hazard ratio for disease progression or death, 0.59; 95% CI, 0.47 to 0.76; P<0.001). Grade 3 or 4 adverse events occurred in 56.5% of 329 patients who received at least one dose of atezolizumab-bevacizumab and in 55.1% of 156 patients who received at least one dose of sorafenib. Grade 3 or 4 hypertension occurred in 15.2% of patients in the atezolizumab-bevacizumab group; however, other high-grade toxic effects were infrequent. In patients with unresectable hepatocellular carcinoma, atezolizumab combined with bevacizumab resulted in better overall and progression-free survival outcomes than sorafenib. (Funded by F. Hoffmann-La Roche/Genentech; ClinicalTrials.gov number, NCT03434379.).",
"Microwave ablation (MWA) has been widely used in the treatment of solid tumors. Studies have been less conducted on the efficacy of MWA used with cell immunotherapy in treating hepatocellular carcinoma (HCC). The current study aimed at exploring the efficacy of MWA in combination with cell immunotherapy in treating HCC. Hepa1-6 HCC mice were treated by MWA, blockade, or the combined therapy (MWA used with blockade), or left untreated. Survival rates of the mice were plotted by Kaplan-Meier Curve, followed by log-rank test. 25 days after the operation, surviving mice were monitored for tumor recurrence, and tumor volumes were calculated every 5 days. Immunohistochemistry and flow cytometry were performed to detect the numbers of CD4+ and CD8+ cells in the tumors and spleens of mice. The expressions of related cytokines were detected and measured by ELISPOT and ELISA. The results showed that MWA combined with anti-PD-1/anti-CTLA-4 not only increased the survival time, protected the mice against tumor recurrence, but also enhanced the intratumoral infiltration of cytotoxic T lymphocyte and systemic T-cell immune responses induced by MWA through activation of synergistically specific antitumor immunity. In addition, the combined therapy increased T-helper 1 cell (Th1-type) cytokines, but reduced Th2-type cytokines, resulting in the polarization of Th1 cells. T-cell immune responses of HCC cells were activated by MWA. In addition, the combined therapy of MWA and anti-PD-1/anti-CTLA-4 induced Th1-type immune response, and showed specific antitumor immunity.",
"Radiofrequency ablation (RFA) is one of the standards of care for early stage hepatocellular carcinoma (HCC). However, rapid progression of residual tumor after RFA has been confirmed. The aim of this study was to investigate the underlying mechanism of this phenomenon. Human HCC cell lines HCCLM3 and HepG2 were employed to establish insufficient RFA models in vivo and in vitro, respectively. The effects of insufficient RFA on metastatic potential of residual tumors were evaluated. The molecular changes after insufficient RFA were evaluated by PCR array, western blot, immunofluorescence, and immunohistochemistry. Results showed that insufficient RFA significantly promoted lung and intrahepatic residual tumor cells in vivo, and heat intervention promoted migration and invasion of hepatoma cells in vitro. PCR array revealed that the expression of integrin β3 (ITGB3) and MMP2 were up-regulated in the residual tumors of HCCLM3 xenograft model. The up-regulation of ITGB3 was confirmed by qRT-PCR, Western blot and immunohistochemistry. Knockdown ITGB3 expression in HCCLM3 cells by shRNA significantly lowered the pro-metastatic effects of insufficient RFA. Mechanism studies indicated that ITGB3 mediated the expression of MMP2 by activing FAK/PI3K/AKT signaling pathway. The up-regulation of ITGB3 contributed to enhanced metastatic potential of residual cancer in HCCLM3 model after insufficient RFA. Targeting ITGB3 expression may further improve the clinical effects of RFA.",
"Up-to-date evidence on levels and trends for age-sex-specific all-cause and cause-specific mortality is essential for the formation of global, regional, and national health policies. In the Global Burden of Disease Study 2013 (GBD 2013) we estimated yearly deaths for 188 countries between 1990, and 2013. We used the results to assess whether there is epidemiological convergence across countries. We estimated age-sex-specific all-cause mortality using the GBD 2010 methods with some refinements to improve accuracy applied to an updated database of vital registration, survey, and census data. We generally estimated cause of death as in the GBD 2010. Key improvements included the addition of more recent vital registration data for 72 countries, an updated verbal autopsy literature review, two new and detailed data systems for China, and more detail for Mexico, UK, Turkey, and Russia. We improved statistical models for garbage code redistribution. We used six different modelling strategies across the 240 causes; cause of death ensemble modelling (CODEm) was the dominant strategy for causes with sufficient information. Trends for Alzheimer's disease and other dementias were informed by meta-regression of prevalence studies. For pathogen-specific causes of diarrhoea and lower respiratory infections we used a counterfactual approach. We computed two measures of convergence (inequality) across countries: the average relative difference across all pairs of countries (Gini coefficient) and the average absolute difference across countries. To summarise broad findings, we used multiple decrement life-tables to decompose probabilities of death from birth to exact age 15 years, from exact age 15 years to exact age 50 years, and from exact age 50 years to exact age 75 years, and life expectancy at birth into major causes. For all quantities reported, we computed 95% uncertainty intervals (UIs). We constrained cause-specific fractions within each age-sex-country-year group to sum to all-cause mortality based on draws from the uncertainty distributions. Global life expectancy for both sexes increased from 65.3 years (UI 65.0-65.6) in 1990, to 71.5 years (UI 71.0-71.9) in 2013, while the number of deaths increased from 47.5 million (UI 46.8-48.2) to 54.9 million (UI 53.6-56.3) over the same interval. Global progress masked variation by age and sex: for children, average absolute differences between countries decreased but relative differences increased. For women aged 25-39 years and older than 75 years and for men aged 20-49 years and 65 years and older, both absolute and relative differences increased. Decomposition of global and regional life expectancy showed the prominent role of reductions in age-standardised death rates for cardiovascular diseases and cancers in high-income regions, and reductions in child deaths from diarrhoea, lower respiratory infections, and neonatal causes in low-income regions. HIV/AIDS reduced life expectancy in southern sub-Saharan Africa. For most communicable causes of death both numbers of deaths and age-standardised death rates fell whereas for most non-communicable causes, demographic shifts have increased numbers of deaths but decreased age-standardised death rates. Global deaths from injury increased by 10.7%, from 4.3 million deaths in 1990 to 4.8 million in 2013; but age-standardised rates declined over the same period by 21%. For some causes of more than 100,000 deaths per year in 2013, age-standardised death rates increased between 1990 and 2013, including HIV/AIDS, pancreatic cancer, atrial fibrillation and flutter, drug use disorders, diabetes, chronic kidney disease, and sickle-cell anaemias. Diarrhoeal diseases, lower respiratory infections, neonatal causes, and malaria are still in the top five causes of death in children younger than 5 years. The most important pathogens are rotavirus for diarrhoea and pneumococcus for lower respiratory infections. Country-specific probabilities of death over three phases of life were substantially varied between and within regions. For most countries, the general pattern of reductions in age-sex specific mortality has been associated with a progressive shift towards a larger share of the remaining deaths caused by non-communicable disease and injuries. Assessing epidemiological convergence across countries depends on whether an absolute or relative measure of inequality is used. Nevertheless, age-standardised death rates for seven substantial causes are increasing, suggesting the potential for reversals in some countries. Important gaps exist in the empirical data for cause of death estimates for some countries; for example, no national data for India are available for the past decade. Bill & Melinda Gates Foundation."
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How to create a query for a narrative review?
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Narrative Review.
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"Ensuring that the patient's voice is routinely incorporated in all aspects of health care in oncology is essential to provide quality care. Patient reported outcomes (PROs) are standardized measures that are used to obtain the patient's perspective and are increasingly used in all aspects of health care to ensure optimal delivery of patient-centered care. The US Food and Drug Administration encourages that PROs be used in studies for label indications. There are no uniform standardized methods to use PROs nor is there consensus on which PROs are best for regulatory approval, comparative effectiveness research, toxicity assessment, health-related quality of life, or symptom monitoring. For this review, we conducted a literature search using PubMed and Google Scholar, and herein summarize the evidence related to the use of PROs in clinic care and research. Using valid, reliable, and easily interpretable PROs developed in comparable populations will provide the most useful results. Various ways that PROs can be used successfully in oncology have been exemplified in this overview to provide clinicians and researchers practical guidance.",
"To test the impact of method of administration (MOA) on the measurement characteristics of items developed in the Patient-Reported Outcomes Measurement Information System (PROMIS). Two non-overlapping parallel 8-item forms from each of three PROMIS domains (physical function, fatigue, and depression) were completed by 923 adults (age 18-89) with chronic obstructive pulmonary disease, depression, or rheumatoid arthritis. In a randomized cross-over design, subjects answered one form by interactive voice response (IVR) technology, paper questionnaire (PQ), personal digital assistant (PDA), or personal computer (PC) on the Internet, and a second form by PC, in the same administration. Structural invariance, equivalence of item responses, and measurement precision were evaluated using confirmatory factor analysis and item response theory methods. Multigroup confirmatory factor analysis supported equivalence of factor structure across MOA. Analyses by item response theory found no differences in item location parameters and strongly supported the equivalence of scores across MOA. We found no statistically or clinically significant differences in score levels in IVR, PQ, or PDA administration as compared to PC. Availability of large item response theory-calibrated PROMIS item banks allowed for innovations in study design and analysis.",
"All nurses are interested in the effects of diseases and treatments on individuals. Patient-reported outcome (PRO) measures are used to obtain self-reported information about symptoms, function, perceptions, and experiences. However, there are challenges to their use, including multiple measures of the same concept, widely varying quality, excessive length and complexity, and difficulty comparing findings across studies and conditions. To address these challenges, the National Institutes of Health funded the Patient-Reported Outcomes Measurement Information System (PROMIS), a web-based repository of valid and reliable PRO measures of health concepts relevant to clinician and researchers. Through the PROMIS Assessment Center, clinicians and researchers can access PRO measures, administer computerized adaptive tests, collect self-report data, and report instant health assessments. The purpose of this article was to summarize the development and validation of the PROMIS measures and to describe its current functionality as it relates to nursing science.",
"Cohort study of patients with cervical radiculopathy undergoing physical therapy. Examine the test-retest reliability, construct validity, and minimum levels of detectable and clinically important change for the Neck Disability Index (NDI) and Patient Specific Functional Scale (PSFS) in cohort of patients with cervical radiculopathy. To date, no studies have investigated the psychometric properties of the NDI or PSFS in a cohort of patients with cervical radiculopathy. Thirty-eight patients with cervical radiculopathy undergoing physical therapy completed the NDI and PSFS, and Numerical Pain Rating Scale (NPRS) at the baseline examination and at a follow-up. In addition, at follow-up, patients completed a 15-point global rating of change (GROC), which was used to dichotomize patients as improved or stable. Changes in the NDI and PSFS were then used to assess test-retest reliability, construct validity, and minimal levels of detectable and clinically important change. Test-retest reliability was moderate for the NDI (intraclass correlation coefficient [ICC] = 0.68; 95% confidence interval [CI], 0.30-0.90) and high for the PSFS (ICC = 0.82; 95% CI, 0.54-0.93). The PSFS was more responsive to change than the NDI. The minimal detectable change for the NDI was 10.2 and for the PSFS 2.1. The minimally clinically important change for the NDI was 7.0 and PSFS 2.0. Our results suggest that the PSFS exhibits superior reliability, construct validity, and responsiveness in this cohort of patients with cervical radiculopathy compared with the NDI. Further research is needed to examine the ability of these measures to accurately reflect changes in individuals, as well as large samples of patients.",
"The goal of comparative effectiveness research (CER) is to explain the differential benefits and harms of alternate methods to prevent, diagnose, treat, and monitor a clinical condition or to improve the delivery of care. To inform decision making, information from the patient's perspective that reflects outcomes that patients care about are needed and can be collected rigorously using appropriate patient-reported outcomes (PRO). It can be challenging to select the most appropriate PRO measure given the proliferation of such questionnaires over the past 20 years. In this paper, we discuss the value of PROs within CER, types of measures that are likely to be useful in the CER context, PRO instrument selection, and key challenges associated with using PROs in CER. We delineate important considerations for defining the CER context, selecting the appropriate measures, and for the analysis and interpretation of PRO data. Emerging changes that may facilitate CER using PROs as an outcome are also reviewed including implementation of electronic and personal health records, hospital and population-based registries, and the use of PROs in national monitoring initiatives. The potential benefits of linking the information derived from PRO endpoints in CER to decision making is also reviewed. The recommendations presented for incorporating PROs in CER are intended to provide a guide to researchers, clinicians, and policy makers to ensure that information derived from PROs is applicable and interpretable for a given CER context. In turn, CER will provide information that is necessary for clinicians, patients, and families to make informed care decisions.",
"Adverse events in chemotherapy clinical trials are assessed and reported by clinicians, yet clinician accuracy in assessing symptoms has been questioned. We compared patient reporting of eight symptoms using a validated instrument, the European Organization for the Research and Treatment of Cancer Quality-of-Life Questionnaire C30 (QLQ-C30 or QLQ) with physicians' reporting of the same symptoms in the study's adverse events log. Thirty-seven men with metastatic, androgen-independent prostate cancer enrolled onto a phase II trial of weekly calcitriol and docetaxel completed the QLQ every 4 weeks for up to 28 weeks. A patient-reported symptom was defined as an increase in a QLQ symptom score by at least 10 points (0 to 100 scale), sustained for at least 4 weeks. A physician-reported symptom was considered present if it was ever documented in the adverse event log. Forty-nine (new or worsened) symptoms were detected by both physician and QLQ, 48 symptoms were detected by the physician alone, and 55 symptoms were detected by the QLQ alone. They agreed on the absence of a symptom in 102 instances of 254 possible opportunities. Their uncorrected agreement was 59.4%, but Cohen's kappa, a coefficient of agreement that corrects for chance, was 0.15, indicating only slight agreement. Using the QLQ as the standard, overall physician sensitivity and specificity was 47% and 68%, respectively, although it varied considerably among symptoms. Even in a tightly controlled clinical trial, physician reporting was neither sensitive nor specific in detecting common chemotherapy adverse effects. Tools for collecting patient-reported adverse event data in chemotherapy clinical trials should be developed.",
"Degenerative cervical myelopathy represents the most common form of non-traumatic spinal cord injury. This trial investigated whether riluzole enhances outcomes in patients undergoing decompression surgery for degenerative cervical myelopathy. This multicentre, double-blind, placebo-controlled, randomised, phase 3 trial was done at 16 university-affiliated centres in Canada and the USA. Patients with moderate-to-severe degenerative cervical myelopathy aged 18-80 years, who had a modified Japanese Orthopaedic Association (mJOA) score of 8-14, were eligible. Patients were randomly assigned (1:1) to receive either oral riluzole (50 mg twice a day for 14 days before surgery and then for 28 days after surgery) or placebo. Randomisation was done using permuted blocks stratified by study site. Patients, physicians, and outcome assessors remained masked to treatment group allocation. The primary endpoint was change in mJOA score from baseline to 6 months in the intention-to-treat (ITT) population, defined as all individuals who underwent randomisation and surgical decompression. Adverse events were analysed in the modified intention-to-treat (mITT) population, defined as all patients who underwent randomisation, including those who did not ultimately undergo surgical decompression. This study is registered with ClinicalTrials.gov, NCT01257828. From Jan 31, 2012, to May 16, 2017, 408 patients were screened. Of those screened, 300 were eligible (mITT population); 290 patients underwent decompression surgery (ITT population) and received either riluzole (n=141) or placebo (n=149). There was no difference between the riluzole and placebo groups in the primary endpoint of change in mJOA score at 6-month follow-up: 2·45 points (95% CI 2·08 to 2·82 points) versus 2·83 points (2·47 to 3·19), difference -0·38 points (-0·90 to 0·13; p=0·14). The most common adverse events were neck or arm or shoulder pain, arm paraesthesia, dysphagia, and worsening of myelopathy. There were 43 serious adverse events in 33 (22%) of 147 patients in the riluzole group and 34 serious adverse events in 29 (19%) of 153 patients in the placebo group. The most frequent severe adverse events were osteoarthrosis of non-spinal joints, worsening of myelopathy, and wound complications. In this trial, adjuvant treatment for 6 weeks perioperatively with riluzole did not improve functional recovery beyond decompressive surgery in patients with moderate-to-severe degenerative cervical myelopathy. Whether riluzole has other benefits in this patient population merits further study. AOSpine North America."
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The International Database of SARS-CoV-2 Variations (IDbSV)
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Ending COVID-19 pandemic requires a collaborative understanding of SARS-CoV-2 and COVID-19 mechanisms. Yet, the evolving nature of coronaviruses results in a continuous emergence of new variants of the virus. Central to this is the need for a continuous monitoring system able to detect potentially harmful variants of the virus in real-time. In this manuscript, we present the International Database of SARS-CoV-2 Variations (IDbSV), the result of ongoing efforts in curating, analyzing, and sharing comprehensive interpretation of SARS-CoV-2's genetic variations and variants. Through user-friendly interactive data visualizations, we aim to provide a novel surveillance tool to the scientific and public health communities. The database is regularly updated with new records through a 4-step workflow (1-Quality control of curated sequences, 2-Call of variations, 3-Functional annotation, and 4-Metadata association). To the best of our knowledge, IDbSV provides access to the largest repository of SARS-CoV-2 variations and the largest analysis of SARS-CoV-2 genomes with over 60 thousand annotated variations curated from the 1,808,613 genomes alongside their functional annotations, first known appearance, and associated genetic lineages, enabling a robust interpretation tool for SARS-CoV-2 variations to help understanding SARS-CoV-2 dynamics across the world.
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[
"Understanding the spread and evolution of pathogens is important for effective public health measures and surveillance. Nextstrain consists of a database of viral genomes, a bioinformatics pipeline for phylodynamics analysis, and an interactive visualization platform. Together these present a real-time view into the evolution and spread of a range of viral pathogens of high public health importance. The visualization integrates sequence data with other data types such as geographic information, serology, or host species. Nextstrain compiles our current understanding into a single accessible location, open to health professionals, epidemiologists, virologists and the public alike. All code (predominantly JavaScript and Python) is freely available from github.com/nextstrain and the web-application is available at nextstrain.org.",
"The COVID-19 outbreak has become a global emergency since December 2019. Analysis of SARS-CoV-2 sequences can uncover single nucleotide variants (SNVs) and corresponding evolution patterns. The Global Evaluation of SARS-CoV-2/hCoV-19 Sequences (GESS, https://wan-bioinfo.shinyapps.io/GESS/) is a resource to provide comprehensive analysis results based on tens of thousands of high-coverage and high-quality SARS-CoV-2 complete genomes. The database allows user to browse, search and download SNVs at any individual or multiple SARS-CoV-2 genomic positions, or within a chosen genomic region or protein, or in certain country/area of interest. GESS reveals geographical distributions of SNVs around the world and across the states of USA, while exhibiting time-dependent patterns for SNV occurrences which reflect development of SARS-CoV-2 genomes. For each month, the top 100 SNVs that were firstly identified world-widely can be retrieved. GESS also explores SNVs occurring simultaneously with specific SNVs of user's interests. Furthermore, the database can be of great help to calibrate mutation rates and identify conserved genome regions. Taken together, GESS is a powerful resource and tool to monitor SARS-CoV-2 migration and evolution according to featured genomic variations. It provides potential directive information for prevalence prediction, related public health policy making, and vaccine designs.",
"In a P.1 coronavirus disease 2019 (COVID-19) outbreak in a long-term care home, vaccine effectiveness against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection was 52.5% (95% confidence interval: 26.9%-69.1%) in residents and 66.2% (2.3%-88.3%) in staff. Vaccine effectiveness against severe illness was 78.6% (47.9%-91.2%) in residents. Two of 19 vaccinated resident case patients died. Outbreak management required both vaccination and infection control measures.",
"Rapid analysis of SARS-CoV-2 genomic data plays a crucial role in surveillance and adoption of measures in controlling spread of Covid-19. Fast, inclusive and adaptive methods are required for the heterogenous SARS-CoV-2 sequence data generated at an unprecedented rate. We present an updated version of the SARS-CoV-2 analysis module of our automated computational pipeline, Infectious Pathogen Detector (IPD) 2.0, to perform genomic analysis to understand the variability and dynamics of the virus. It adopts the recent clade nomenclature and demonstrates the clade prediction accuracy of 92.8%. IPD 2.0 also contains a SARS-CoV-2 updater module, allowing automatic upgrading of the variant database using genome sequences from GISAID. As a proof of principle, analyzing 208,911 SARS-CoV-2 genome sequences, we generate an extensive database of 2.58 million sample-wise variants. A comparative account of lineage-specific mutations in the newer SARS-CoV-2 strains emerging in the UK, South Africa and Brazil and data reported from India identify overlapping and lineages specific acquired mutations suggesting a repetitive convergent and adaptive evolution. A novel and dynamic feature of the SARS-CoV-2 module of IPD 2.0 makes it a contemporary tool to analyze the diverse and growing genomic strains of the virus and serve as a vital tool to help facilitate rapid genomic surveillance in a population to identify variants involved in breakthrough infections. IPD 2.0 is freely available from http://www.actrec.gov.in/pi-webpages/AmitDutt/IPD/IPD.html and the web-application is available at http://ipd.actrec.gov.in/ipdweb/ .",
"The novel coronavirus disease (COVID-19) has claimed lots of lives, posing a dire threat to global health. It was predicted that the coronavirus outbreak in the African population would be very lethal and result to economic devastation owing to the prevalence of immune-compromised population, poverty, low lifespan, fragile health care systems, poor economy, and lifestyle factors. Accumulation of mutations gives virus selective advantage for host invasion and adaptation, higher transmissibility of more virulent strains, and drug resistance. The present study determined the severe acute respiratory syndrome-2 (SARS-CoV-2) genomic variability and the contributory factors to the low COVID-19 fatality in Africa. To assess the SARS-CoV-2 mutational landscape, 924 viral sequences from the Africa region with their sociobiological characteristics mined from the Global Initiative on Sharing All Influenza Data (GISAID) database were analyzed. Mutational analysis of the SARS-CoV-2 sequences revealed highly recurrent mutations in the SARS-CoV-2 spike glycoprotein D614G (97.2%), concurrent R203K, and G204R (65.2%) in the nucleocapsid phosphoprotein, and P4715L (97.2%) in the RNA-dependent RNA polymerase flagging these regions as SARS-CoV-2 mutational hotspots in the African population. COVID-19 is more severe in older people (> 65 years); Africa has a low percentage of people within this age group (4.36%). The average age of the infected patients observed in this study is 46 years with only 47 infected patients (5.1%) above 65 years in Africa in comparison to 13.12% in countries in other continents with the highest prevalence of COVID-19. Africa's young generation, the late incidence of the disease, and adherence to public health guidelines are important indicators that may have contributed to the observed low COVID-19 deaths in Africa. However, with the easing of lockdown and regulatory policies, daily increasing incidence in most countries, and low testing and sequencing rate, the epidemiology and the true impact of the pandemic in Africa remain to be unraveled.",
"The COVID-19 pandemic is due to infection caused by the novel SARS-CoV-2 virus that impacts the lower respiratory tract. The spectrum of symptoms ranges from asymptomatic infections to mild respiratory symptoms to the lethal form of COVID-19 which is associated with severe pneumonia, acute respiratory distress, and fatality. To address this global crisis, up-to-date information on viral genomics and transcriptomics is crucial for understanding the origins and global dispersion of the virus, providing insights into viral pathogenicity, transmission, and epidemiology, and enabling strategies for therapeutic interventions, drug discovery, and vaccine development. Therefore, this review provides a comprehensive overview of COVID-19 epidemiology, genomic etiology, findings from recent transcriptomic map analysis, viral-human protein interactions, molecular diagnostics, and the current status of vaccine and novel therapeutic intervention development. Moreover, we provide an extensive list of resources that will help the scientific community access numerous types of databases related to SARS-CoV-2 OMICs and approaches to therapeutics related to COVID-19 treatment."
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Lysosomes: Physiology, Functions, and Targeting
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For a long time, lysosomes were considered as mere waste bags for cellular constituents. Thankfully, studies carried out in the past 15 years were brimming with elegant and crucial breakthroughs in lysosome research, uncovering their complex roles as nutrient sensors and characterizing them as crucial multifaceted signaling organelles. This review presents the scientific knowledge on lysosome physiology and functions, starting with their discovery and reviewing up to date ground-breaking discoveries highlighting their heterogeneous functions as well as pending questions that remain to be answered. We also review the roles of lysosomes in anti-cancer drug resistance and how they undergo a series of molecular and functional changes during malignant transformation which lead to tumor aggression, angiogenesis, and metastases. Finally, we discuss the strategy of targeting lysosomes in cancer which could lead to the development of new and effective targeted therapies.
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"The view of the lysosome as the terminal end of cellular catabolic pathways has been challenged by recent studies showing a central role of this organelle in the control of cell function. Here we show that a lysosomal Ca2+ signalling mechanism controls the activities of the phosphatase calcineurin and of its substrate TFEB, a master transcriptional regulator of lysosomal biogenesis and autophagy. Lysosomal Ca2+ release through mucolipin 1 (MCOLN1) activates calcineurin, which binds and dephosphorylates TFEB, thus promoting its nuclear translocation. Genetic and pharmacological inhibition of calcineurin suppressed TFEB activity during starvation and physical exercise, while calcineurin overexpression and constitutive activation had the opposite effect. Induction of autophagy and lysosomal biogenesis through TFEB required MCOLN1-mediated calcineurin activation. These data link lysosomal calcium signalling to both calcineurin regulation and autophagy induction and identify the lysosome as a hub for the signalling pathways that regulate cellular homeostasis.",
"Lysosome-related organelles (LROs) comprise a group of cell type-specific subcellular compartments with unique composition, morphology and structure that share some features with endosomes and lysosomes and that function in varied processes such as pigmentation, hemostasis, lung plasticity and immunity. In recent years, studies of genetic diseases in which LRO functions are compromised have provided new insights into the mechanisms of LRO biogenesis and the regulated secretion of LRO contents. These insights have revealed previously unappreciated specialized endosomal sorting processes in all cell types, and are expanding our views of the plasticity of the endosomal and secretory systems in adapting to cell type-specific needs.",
"Acidification of endocytic compartments occurs within 2-3 minutes following internalization. In CHO cells, three kinetically and morphologically distinct endocytic or recycling compartments can be identified. Measurements of the pH in these compartments show that the acidity of the compartments is regulated independently, although some parts of the acidification mechanism may be shared. The pH values found in these compartments provide a satisfactory explanation for various required steps in the endocytosis and recycling pathways. Further work is required to understand the biochemical basis for the pH regulation in various intracellular organelles.",
"The Hermansky-Pudlak syndrome is a disorder affecting endosome sorting. Disease is triggered by defects in any of 15 mouse gene products, which are part of five distinct cytosolic molecular complexes: AP-3, homotypic fusion and vacuole protein sorting, and BLOC-1, -2, and -3. To identify molecular associations of these complexes, we used in vivo cross-linking followed by purification of cross-linked AP-3 complexes and mass spectrometric identification of associated proteins. AP-3 was co-isolated with BLOC-1, BLOC-2, and homotypic fusion and vacuole protein sorting complex subunits; clathrin; and phosphatidylinositol-4-kinase type II alpha (PI4KIIalpha). We previously reported that this membrane-anchored enzyme is a regulator of AP-3 recruitment to membranes and a cargo of AP-3 ( Craige, B., Salazar, G., and Faundez, V. (2008) Mol. Biol. Cell 19, 1415-1426 ). Using cells deficient in different Hermansky-Pudlak syndrome complexes, we identified that BLOC-1, but not BLOC-2 or BLOC-3, deficiencies affect PI4KIIalpha inclusion into AP-3 complexes. BLOC-1, PI4KIIalpha, and AP-3 belong to a tripartite complex, and down-regulation of either PI4KIIalpha, BLOC-1, or AP-3 complexes led to similar LAMP1 phenotypes. Our analysis indicates that BLOC-1 complex modulates the association of PI4KIIalpha with AP-3. These results suggest that AP-3 and BLOC-1 act, either in concert or sequentially, to specify sorting of PI4KIIalpha along the endocytic route.",
"The phosphoinositide PI(3,5)P2, generated exclusively by the PIKfyve lipid kinase complex, is key for lysosomal biology. Here, we explore how PI(3,5)P2 levels within cells are regulated. We find the PIKfyve complex comprises five copies of the scaffolding protein Vac14 and one copy each of the lipid kinase PIKfyve, generating PI(3,5)P2 from PI3P and the lipid phosphatase Fig4, reversing the reaction. Fig4 is active as a lipid phosphatase in the ternary complex, whereas PIKfyve within the complex cannot access membrane-incorporated phosphoinositides due to steric constraints. We find further that the phosphoinositide-directed activities of both PIKfyve and Fig4 are regulated by protein-directed activities within the complex. PIKfyve autophosphorylation represses its lipid kinase activity and stimulates Fig4 lipid phosphatase activity. Further, Fig4 is also a protein phosphatase acting on PIKfyve to stimulate its lipid kinase activity, explaining why catalytically active Fig4 is required for maximal PI(3,5)P2 production by PIKfyve in vivo.",
"Dynactin is a multisubunit complex that plays an accessory role in cytoplasmic dynein function. Overexpression in mammalian cells of one dynactin subunit, dynamitin, disrupts the complex, resulting in dissociation of cytoplasmic dynein from prometaphase kinetochores, with consequent perturbation of mitosis (Echeverri, C.J., B.M. Paschal, K.T. Vaughan, and R.B. Vallee. 1996. J. Cell Biol. 132:617-634). Based on these results, dynactin was proposed to play a role in linking cytoplasmic dynein to kinetochores and, potentially, to membrane organelles. The current study reports on the dynamitin interphase phenotype. In dynamitin-overexpressing cells, early endosomes (labeled with antitransferrin receptor), as well as late endosomes and lysosomes (labeled with anti-lysosome-associated membrane protein-1 [LAMP-1]), were redistributed to the cell periphery. This redistribution was disrupted by nocodazole, implicating an underlying plus end-directed microtubule motor activity. The Golgi stack, monitored using sialyltransferase, galactosyltransferase, and N-acetylglucosaminyltransferase I, was dramatically disrupted into scattered structures that colocalized with components of the intermediate compartment (ERGIC-53 and ERD-2). The disrupted Golgi elements were revealed by EM to represent short stacks similar to those formed by microtubule-depolymerizing agents. Golgi-to-ER traffic of stack markers induced by brefeldin A was not inhibited by dynamitin overexpression. Time-lapse observations of dynamitin-overexpressing cells recovering from brefeldin A treatment revealed that the scattered Golgi elements do not undergo microtubule-based transport as seen in control cells, but rather, remain stationary at or near their ER exit sites. These results indicate that dynactin is specifically required for ongoing centripetal movement of endocytic organelles and components of the intermediate compartment. Results similar to those of dynamitin overexpression were obtained by microinjection with antidynein intermediate chain antibody, consistent with a role for dynactin in mediating interactions of cytoplasmic dynein with specific membrane organelles. These results suggest that dynamitin plays a pivotal role in regulating organelle movement at the level of motor-cargo binding.",
"Our study was aimed at examinating whether or not the human genome encodes for previously unreported cysteine cathepsins. To this end, we used analyses of the genome sequence and mRNA expression levels. The program TBLASTN was employed to scan the draft sequence of the human genome for the 11 known cysteine cathepsins. The cathepsin-like segments in the genome were inspected, filtered, and annotated. In addition to the known cysteine cathepsins, the scan identified three pseudogenes, closely related to cathepsin L, on chromosome 10, as well as two remote homologs, tubulointerstitial protein antigen and tubulointerstitial protein antigen-related protein. No new members of the family were identified. mRNA expression profiles for 10 known human cysteine cathepsins showed varying expression levels in 46 different human tissues and cell lines. No expression of any of the three cathepsin L-like pseudogenes was found. Based on these results, it is likely that to date all human cysteine cathepsins are known."
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Megalichthyids (Tetrapodomorpha: Megalichthyidae)
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The megalichthyids are one of several clades of extinct tetrapodomorph fish that lived throughout the Devonian-Permian periods. They are advanced "osteolepidid-grade" fishes that lived in freshwater swamp and lake environments, with some taxa growing to very large sizes. They bear cosmine-covered bones and a large premaxillary tusk that lies lingually to a row of small teeth. Diagnosis of the family remains controversial with various authors revising it several times in recent works. There are fewer than 10 genera known globally, and only one member definitively identified from Gondwana.
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"Complete, exceptionally-preserved skulls of the Permian lungfish Persephonichthys chthonica gen. et sp. nov. are described. Persephonichthys chthonica is unique among post-Devonian lungfishes in preserving portions of the neurocranium, permitting description of the braincase of a stem-ceratodontiform for the first time. The completeness of P. chthonica permits robust phylogenetic analysis of the relationships of the extant lungfish lineage within the Devonian lungfish diversification for the first time. New analyses of the relationships of this new species within two published matrices using both maximum parsimony and Bayesian inference robustly place P. chthonica and modern lungfishes within dipterid-grade dipnoans rather than within a clade containing Late Devonian 'phaneropleurids' and common Late Paleozoic lungfishes such as Sagenodus. Monophyly of post-Devonian lungfishes is not supported and the Carboniferous-Permian taxon Sagenodus is found to be incidental to the origins of modern lungfishes, suggesting widespread convergence in Late Paleozoic lungfishes. Morphology of the skull, hyoid arch, and pectoral girdle suggests a deviation in feeding mechanics from that of Devonian lungfishes towards the more dynamic gape cycle and more effective buccal pumping seen in modern lungfishes. Similar anatomy observed previously in 'Rhinodipterus' kimberyensis likely represents an intermediate state between the strict durophagy observed in most Devonian lungfishes and the more dynamic buccal pump seen in Persephonichthys and modern lungfishes, rather than adaptation to air-breathing exclusively.",
"Actinopterygians (ray-finned fishes) are the most diverse living osteichthyan (bony vertebrate) group, with a rich fossil record. However, details of their earliest history during the middle Palaeozoic (Devonian) 'Age of Fishes' remains sketchy. This stems from an uneven understanding of anatomy in early actinopterygians, with a few well-known species dominating perceptions of primitive conditions. Here we present an exceptionally preserved ray-finned fish from the Late Devonian (Middle Frasnian, ca 373 Ma) of Pas-de-Calais, northern France. This new genus is represented by a single, three-dimensionally preserved skull. CT scanning reveals the presence of an almost complete braincase along with near-fully articulated mandibular, hyoid and gill arches. The neurocranium differs from the coeval Mimipiscis in displaying a short aortic canal with a distinct posterior notch, long grooves for the lateral dorsal aortae, large vestibular fontanelles and a broad postorbital process. Identification of similar but previously unrecognized features in other Devonian actinopterygians suggests that aspects of braincase anatomy in Mimipiscis are apomorphic, questioning its ubiquity as stand-in for generalized actinopterygian conditions. However, the gill skeleton of the new form broadly corresponds to that of Mimipiscis, and adds to an emerging picture of primitive branchial architecture in crown gnathostomes. The new genus is recovered in a polytomy with Mimiidae and a subset of Devonian and stratigraphically younger actinopterygians, with no support found for a monophyletic grouping of Moythomasia with Mimiidae.",
"One of the first endocasts of a dipnoan (lungfish) to be realised was that of the Upper Devonian taxon Chirodipterus australis. This early interpretation was based on observations of the shape of the cranial cavity alone and was not based on a natural cast or 'steinkern' nor from serial sectioning. The validity of this reconstruction is therefore questionable and continued reference to and use of this interpretation in analyses of sarcopterygian cranial evolution runs the risk of propagation of error. Here we present a new detailed anatomical description of the endocast of 'Chirodipterus' australis from the Upper Devonian Gogo Formation of Western Australia, known for exceptional 3D preservation which enables fine-scale scrutiny of endocranial anatomy. We show that it exhibits a suite of characters more typical of Lower and Middle Devonian dipnoan taxa. Notably, the small utricular recess is unexpected for a taxon of this age, whereas the ventral expansion of the telencephalon is more typical of more derived taxa. The presence of such 'primitive' characters in 'C.' australis supports its relatively basal position as demonstrated in the most recent phylogenies of Devonian Dipnoi.",
"For the past 25 years NIH Image and ImageJ software have been pioneers as open tools for the analysis of scientific images. We discuss the origins, challenges and solutions of these two programs, and how their history can serve to advise and inform other software projects."
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Transfer RNAs and Cancer: A Review
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The dysregulation of transfer RNA (tRNA) expression contributes to the diversity of proteomics, heterogeneity of cell populations, and instability of the genome, which may be related to human cancer susceptibility. However, the relationship between tRNA dysregulation and cancer susceptibility remains elusive because the landscape of cancer-associated tRNAs has not been portrayed yet. Furthermore, the molecular mechanisms of tRNAs involved in tumorigenesis and cancer progression have not been systematically understood. In this review, we detail current knowledge of cancer-related tRNAs and comprehensively summarize the basic characteristics and functions of these tRNAs, with a special focus on their role and involvement in human cancer. This review bridges the gap between tRNAs and cancer and broadens our understanding of their relationship, thus providing new insights and strategies to improve the potential clinical applications of tRNAs for cancer diagnosis and therapy.
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"Breast milk is a complex liquid rich in immunological components that affect the development of the infant's immune system. Exosomes are membranous vesicles of endocytic origin that are found in various body fluids and that can mediate intercellular communication. MicroRNAs (miRNAs), a well-defined group of non-coding small RNAs, are packaged inside exosomes in human breast milk. Here, we identified 602 unique miRNAs originating from 452 miRNA precursors (pre-miRNAs) in human breast milk exosomes using deep sequencing technology. We found that, out of 87 well-characterized immune-related pre-miRNAs, 59 (67.82%) are presented and enriched in breast milk exosomes (P < 10(-16), χ(2) test). In addition, compared with exogenous synthetic miRNAs, these endogenous immune-related miRNAs are more resistant to relatively harsh conditions. It is, therefore, tempting to speculate that these exosomal miRNAs are transferred from the mother's milk to the infant via the digestive tract, and that they play a critical role in the development of the infant immune system.",
"Posttranscriptional gene regulation by small RNAs and its crucial impact on development, apoptosis, stem cell self-renewal and differentiation gained tremendous scientific attention since the discovery of RNA interference (RNAi) and microRNAs (miRNAs). However, in the last few years, many more examples for regulatory small RNAs were discovered, some of them even with miRNA-like functions. Even though these small RNA molecules were previously thought to be mere artifacts accumulating during the preparation of RNA libraries, advances in sequencing technology revealed that small RNAs derive from hairpin-fold RNA structures, for example. Mirtrons, short hairpin RNAs or small RNAs that are processed from longer non-coding RNAs such as tRNAs or snoRNAs have been found recently and some of them might be involved in the regulation of gene expression in different organisms. Furthermore, small RNAs originating from transposable elements, heterochromatic regions or convergent transcription units forming endogenous short interfering RNAs (endo-siRNAs) are the somatic equivalents of the germline-specific Piwi-interacting RNAs (piRNAs) in mediating transposon silencing. This review will focus on several recent findings that have added new aspects to small RNA-guided gene silencing.",
"Over the past decades, tRNA was found to be a rich hub of RNA modifications such as 1-methyladenosine and 5-methycytosine modifications and others, holding more than half of all modifications occurring in RNA molecules. Moreover, tRNA was discovered to be a source of various small noncoding RNA species, such as the stress induced angiogenin cleaved tRNA halves (tiRNA) or the miRNA like tRNA derived fragments. tRNA cleavage under stress was fist discovered in bacteria and later was found to be conserved across different species, including mammals. Under cellular stress conditions, tRNA undergoes conformational changes and angiogenin cleaves it into 3' and 5' halves. 5'tiRNA halves were shown to repress protein translations. tRNA cleavage is thought of to be a cytoprotective mechanism by which cells evade apoptosis, however some data hints to the opposite; that tiRNA are cytotoxic or at least related to apoptosis initiation. tRNA cleavage also was shown to be affected by tRNA modifications via different enzymes in the cytosol and mitochondria. In this review, we will highlight the biology of tRNA cleavage, show the evidence of it being cytoprotective or a marker of cell death and shed a light on its role in disease models and human diseases as well as possible future directions in this field of RNA research.",
"Recently, it has been shown that tRNA molecules can be processed into small RNAs that are derived from both the 5' and 3' termini. To date, the function of these tRNA fragments (tRFs) derived from the 5' end of tRNA has not been investigated in depth. We present evidence that conserved residues in tRNA, present in all 5' tRFs, can inhibit the process of protein translation without the need for complementary target sites in the mRNA. These results implicate 5' tRFs in a new mechanism of gene regulation by small RNAs in human cells.",
"Small RNAs complex with proteins to mediate a variety of functions in animals and plants. Some small RNAs, particularly miRNAs, circulate in mammalian blood and may carry out a signaling function by entering target cells and modulating gene expression. The subject of this study is a set of circulating 30-33 nt RNAs that are processed derivatives of the 5' ends of a small subset of tRNA genes, and closely resemble cellular tRNA derivatives (tRFs, tiRNAs, half-tRNAs, 5' tRNA halves) previously shown to inhibit translation initiation in response to stress in cultured cells. In sequencing small RNAs extracted from mouse serum, we identified abundant 5' tRNA halves derived from a small subset of tRNAs, implying that they are produced by tRNA type-specific biogenesis and/or release. The 5' tRNA halves are not in exosomes or microvesicles, but circulate as particles of 100-300 kDa. The size of these particles suggest that the 5' tRNA halves are a component of a macromolecular complex; this is supported by the loss of 5' tRNA halves from serum or plasma treated with EDTA, a chelating agent, but their retention in plasma anticoagulated with heparin or citrate. A survey of somatic tissues reveals that 5' tRNA halves are concentrated within blood cells and hematopoietic tissues, but scant in other tissues, suggesting that they may be produced by blood cells. Serum levels of specific subtypes of 5' tRNA halves change markedly with age, either up or down, and these changes can be prevented by calorie restriction. We demonstrate that 5' tRNA halves circulate in the blood in a stable form, most likely as part of a nucleoprotein complex, and their serum levels are subject to regulation by age and calorie restriction. They may be produced by blood cells, but their cellular targets are not yet known. The characteristics of these circulating molecules, and their known function in suppression of translation initiation, suggest that they are a novel form of signaling molecule.",
"Small noncoding RNAs circulating in the blood may serve as signaling molecules because of their ability to carry out a variety of cellular functions. We have previously described tRNA- and YRNA-derived small RNAs circulating as components of larger complexes in the blood of humans and mice; the characteristics of these small RNAs imply specific processing, secretion, and physiological regulation. In this study, we have asked if changes in the serum abundance of these tRNA and YRNA fragments are associated with a diagnosis of cancer. We used deep sequencing and informatics analysis to catalog small RNAs in the sera of breast cancer cases and normal controls. 5' tRNA halves and YRNA fragments are abundant in both groups, but we found that a breast cancer diagnosis is associated with changes in levels of specific subtypes. This prompted us to look at existing sequence datasets of serum small RNAs from 42 breast cancer cases, taken at the time of diagnosis. We find significant changes in the levels of specific 5' tRNA halves and YRNA fragments associated with clinicopathologic characteristics of the cancer. Although these findings do not establish causality, they suggest that circulating 5' tRNA halves and YRNA fragments with known cellular functions may participate in breast cancer syndromes and have potential as circulating biomarkers. Larger studies with multiple types of cancer are needed to adequately evaluate their potential use for the development of noninvasive cancer screening.",
"Higher eukaryotes employ extensive post-transcriptional gene regulation to accomplish fine control of gene expression. The microRNA (miRNA) family plays important roles in the post-transcriptional gene regulation of broad networks of target mRNA expression. Most miRNAs are generated by a conserved mechanism involving two RNase III enzymes Drosha and Dicer. However, work from the past few years has uncovered diverse noncanonical miRNA pathways, which exploit a variety of other RNA processing enzymes. In addition, the discovery of another abundant small RNA family, endogenous short interfering RNAs (endo-siRNAs), has also broadened the catalogs of short regulatory RNAs. This review highlights recent studies that revealed novel small RNA biogenesis pathways, and discusses their relevance to gene regulatory networks.",
"Exosomes are small (30-150 nm) vesicles containing unique RNA and protein cargo, secreted by all cell types in culture. They are also found in abundance in body fluids including blood, saliva, and urine. At the moment, the mechanism of exosome formation, the makeup of the cargo, biological pathways, and resulting functions are incompletely understood. One of their most intriguing roles is intercellular communication--exosomes function as the messengers, delivering various effector or signaling macromolecules between specific cells. There is an exponentially growing need to dissect structure and the function of exosomes and utilize them for development of minimally invasive diagnostics and therapeutics. Critical to further our understanding of exosomes is the development of reagents, tools, and protocols for their isolation, characterization, and analysis of their RNA and protein contents. Here we describe a complete exosome workflow solution, starting from fast and efficient extraction of exosomes from cell culture media and serum to isolation of RNA followed by characterization of exosomal RNA content using qRT-PCR and next-generation sequencing techniques. Effectiveness of this workflow is exemplified by analysis of the RNA content of exosomes derived from HeLa cell culture media and human serum, using Ion Torrent PGM as a sequencing platform.",
"Cells use secreted signals (e.g. chemokines and growth factors) and sophisticated vehicles such as argosomes, cytonemes, tunneling nanotubes and exosomes to relay important information to other cells, often over large distances. Exosomes, 30-100-nm intraluminal vesicles of multivesicular bodies (MVB) released upon exocytic fusion of the MVB with the plasma membrane, are increasingly recognized as a novel mode of cell-independent communication. Exosomes have been shown to function in antigen presentation and tumor metastasis, and in transmitting infectious agents. However, little is known about the biogenesis and function of exosomes in polarized cells. In this review, we discuss new evidence suggesting that exosomes participate in the transport of morphogens and RNA, and thus influence cell polarity and developmental patterning of tissues.",
"The proto-oncogene product c-Myc has a direct role in both metazoan cell growth and division. RNA polymerase III (pol III) is involved in the generation of transfer RNA and 5S ribosomal RNA, and these molecules must be produced in bulk to meet the need for protein synthesis in growing cells. We demonstrate here that c-Myc binds to TFIIIB, a pol III-specific general transcription factor, and directly activates pol III transcription. Chromatin immunoprecipitation reveals that endogenous c-Myc is present at tRNA and 5S rRNA genes in cultured mammalian cells. These results suggest that activation of pol III may have a role in the ability of c-Myc to stimulate cell growth."
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Methyl-CpG-binding protein 2 promotes osteogenic differentiation of bone marrow mesenchymal stem cells during osteoporosis
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Postmenopausal osteoporosis is characterized by inadequate bone formation of osteoblasts and excessive bone resorption of osteoclasts. Bone marrow mesenchymal stem cells (BMSCs), with the potential of osteogenic differentiation, have been widely used in the bone tissues engineering for the treatment of bone diseases, including postmenopausal osteoporosis. Methyl-CpG-binding protein 2 (MECP2) has been reported to be implicated in bone formation during the development of Rett syndrome. However, the influence of MeCP2 on osteogenic differentiation of BMSCs during osteoporosis remains unclear. Firstly, mice model with estrogen deficiency-induced osteoporosis was established through ovariectomy (OVX). MeCP2 was found to be down-regulated in bone tissues and BMSCs of OVX-induced osteoporosis mice. Secondly, over-expression of MeCP2 enhanced the calcium deposition of BMSCs isolated from the OVX-induced osteoporosis mice. Moreover, expression of osteogenic biomarkers including alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2), collagen type I alpha 1 (COL1A1), and osteocalcin (OCN) was increased in BMSCs by overexpression of MeCP2. Thirdly, over-expression of MeCP2 reduced protein expression of forkhead box F1 (FOXF1) and adenomatous polyposis coli (APC), while enhanced Wnt5a and β-catenin expression in BMSCs. Over-expression of FOXF1 attenuated MeCP2 over-expression-induced decrease of FOXF1 and APC, as well as increase of Wnt5a and β-catenin. Finally, the increased calcium deposition, protein expression of ALP, RUNX2COL1A1 and OCN induced by concomitant overexpression of MeCP2 were also restored by FOXF1 over-expression. In conclusion, MeCP2 promoted osteogenic differentiation of BMSCs through regulating FOXF1/Wnt/β-Catenin axis to attenuate osteoporosis. MeCP2 over-expression reduced FOXF1 to promote the activation of Wnt5a/β-Catenin and promote osteogenic differentiation of BMSCs during the prevention of postmenopausal osteoporosis.
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"Skeletal unloading induced by hindlimb suspension in rats reduces bone formation and induces osteopenia, but its effect on adipogenesis is unknown. We assessed the effects of unloading and transforming growth factor (TGF) beta2 on bone marrow stromal cell adipocyte differentiation in relation with osteoblast differentiation. Skeletal unloading rapidly (4-7 days) decreased osteoblast transcription factor Runx2, osteocalcin (OC), and type I collagen messenger RNA (mRNA) levels and reduced bone formation in the long bone metaphysis. Conversely, unloading increased expression of the adipocyte transcription factor peroxisome proliferator-activated receptor gamma2 (PPARgamma2) at 4 days and increased expression of the adipocyte differentiation genes lipoprotein lipase (LPL) and aP2 in the bone marrow stroma at 7 days. Consistently, unloading increased the number and volume of adipocytes in the bone marrow stroma. Continuous (0-7 days) and late (4-7 days) treatments with TGF-beta2 corrected the abnormal expression of Cbfa1/Runx2, OC, and type I collagen mRNAs and normalized bone formation in unloaded metaphyseal bone. Moreover, both TGF-beta2 treatments decreased PPARy2 and C/EBPalpha mRNA levels at 4 days and normalized aP2 and LPL expression and adipocyte number and volume at 7 days. These results show that skeletal unloading increases adipocyte differentiation concomitantly with inhibition of osteoblast differentiation. These abnormalities are prevented and reversed by TGF-beta2, suggesting a role for TGF-beta in the control of adipogenic differentiation in the bone marrow stroma.",
"Little is known about the implications of circRNAs in the effects of melatonin (MEL) on bone marrow mesenchymal stem cell (BMSC) osteogenic differentiation and osteoporosis (OP) progression. The aim of our study was to investigate circRNAs in MEL-regulated BMSC differentiation and OP progression. BMSC osteogenic differentiation was measured by qRT-PCR, western blot (WB), Alizarin Red, and alkaline phosphatase (ALP) staining. Differential circRNA and mRNA profiles of BMSCs treated by MEL were characterized by deep sequencing, followed by validation using RT-PCR, Sanger sequencing, and qRT-PCR. Silencing and overexpression of circ_0003865 were conducted for functional investigations. The sponged microRNAs and targeted mRNAs were predicted by bioinformatics and validated by qRT-PCR, RNA pull-down, and dual-luciferase reporter assay. The function of miR-3653-3p and circ_0003865/miR-3653-3p/growth arrest-specific gene 1 (GAS1) cascade was validated for the osteogenic differentiation of BMSCs by CCK-8, qRT-PCR, WB, Alizarin Red, and ALP staining. The effects of circ_0003865 on OP development were tested in murine OP model. MEL promoted osteogenic differentiation of BMSCs. RNA sequencing revealed significant alterations in circRNA and mRNA profiles associated with multiple biological processes and signaling pathways. Circ_0003865 expression in BMSCs was significantly decreased by MEL treatment. Silencing of circ_0003865 had no effect on proliferation while promoted osteogenic differentiation of BMSCs. Overexpression of circ_0003865 abrogated the promotion of BMSC osteogenic differentiation induced by MEL, but proliferation of BMSCs induced by MEL had no change whether circ_0003865 was overexpression or not. Furthermore, circ_0003865 sponged miR-3653-3p to promote GAS1 expression in BMSCs. BMSC osteogenic differentiation was enhanced by miR-3653-3p overexpression while BMSC proliferation was not affected. By contrast, miR-3653-3p silencing mitigated the promoted BMSC osteogenic differentiation caused by circ_0003865 silencing, but had no effect on proliferation. Finally, circ_0003865 silencing repressed OP development in mouse model. MEL promotes BMSC osteogenic differentiation and inhibits OP pathogenesis by suppressing the expression of circ_0003865, which regulates GAS1 gene expression via sponging miR-3653-3p.",
"Chondrocytes and osteoblasts are two primary cell types in the skeletal system that are differentiated from common mesenchymal progenitors. It is believed that osteoblast differentiation is controlled by distinct mechanisms in intramembranous and endochondral ossification. We have found that ectopic canonical Wnt signaling leads to enhanced ossification and suppression of chondrocyte formation. Conversely, genetic inactivation of beta-catenin, an essential component transducing the canonical Wnt signaling, causes ectopic formation of chondrocytes at the expense of osteoblast differentiation during both intramembranous and endochondral ossification. Moreover, inactivation of beta-catenin in mesenchymal progenitor cells in vitro causes chondrocyte differentiation under conditions allowing only osteoblasts to form. Our results demonstrate that beta-catenin is essential in determining whether mesenchymal progenitors will become osteoblasts or chondrocytes regardless of regional locations or ossification mechanisms. Controlling Wnt/beta-catenin signaling is a common molecular mechanism underlying chondrocyte and osteoblast differentiation and specification of intramembranous and endochondral ossification.",
"To investigate the effect of icariin on proliferation of bone marrow mesenchymal stem cells (BMSCs) in Sprague-Dawley (SD) rats. BMSCs were obtained from SD rat bone marrow with differential time adherent method. Its characteristic was identified through differentiation cell surface antigens and the multi-lineage (osteo/adipo/chondo) differentiation potential. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method and 5-Bromo-2-Deoxyuridine (BrdU) incorporation were applied to detect the effect of icariin on BMSCs proliferation. Flow cytometry was used to detect proliferation index of BMSCs. The RNA level and the distribution of β-catenin were evaluated by Real-time Polymerase Chain Reaction (PCR) and Immunofluorescent staining respectively. Western blot was used to detect protein expression levels of β-catenin, glycogen synthase kinase-3 beta (GSK-3β), phospho-glycogen synthase kinase-3 beta (pGSK-3β) and cyclinD1. Icariin promoted BMSCs proliferation at the concentration of 0.05-2.0 mg/L. The percentage of BrdU positive cells of BMSCs was increased from 40.98% to 70.42%, and the proliferation index value was increased from 8.9% to 17.5% with the treatment of 0.05 mg/L icariin, which significance values were both less than 0.05. Compared with the control group, total and nuclear β-catenin proteins, as well as β-catenin mRNA expression, were all increased with icariin treatment. Meanwhile, the phosphorylation level of GSK-3β and cyclinD1 protein expressions were also increased in BMSCs with icariin treatment. The findings of the present study demonstrated that low dosage of icariin could promote BMSCs proliferation. The activation of Wnt/β-catenin pathways was involved in this process.",
"Wnt signaling is essential for osteogenesis and also functions as an adipogenic switch, but it is not known if interrupting wnt signaling via knockout of β-catenin from osteoblasts would cause bone marrow adiposity. Here, we determined whether postnatal deletion of β-catenin in preosteoblasts, through conditional cre expression driven by the osterix promoter, causes bone marrow adiposity. Postnatal disruption of β-catenin in the preosteoblasts led to extensive bone marrow adiposity and low bone mass in adult mice. In cultured bone marrow-derived cells isolated from the knockout mice, adipogenic differentiation was dramatically increased, whereas osteogenic differentiation was significantly decreased. As myoblasts, in the absence of wnt/β-catenin signaling, can be reprogrammed into the adipocyte lineage, we sought to determine whether the increased adipogenesis we observed partly resulted from a cell-fate shift of preosteoblasts that had to express osterix (lineage-committed early osteoblasts), from the osteoblastic to the adipocyte lineage. Using lineage tracing both in vivo and in vitro we showed that the loss of β-catenin from preosteoblasts caused a cell-fate shift of these cells from osteoblasts to adipocytes, a shift that may at least partly contribute to the bone marrow adiposity and low bone mass in the knockout mice. These novel findings indicate that wnt/β-catenin signaling exerts control over the fate of lineage-committed early osteoblasts, with respect to their differentiation into osteoblastic versus adipocytic populations in bone, and thus offers potential insight into the origin of bone marrow adiposity.",
"The cellular constituents forming the haematopoietic stem cell (HSC) niche in the bone marrow are unclear, with studies implicating osteoblasts, endothelial and perivascular cells. Here we demonstrate that mesenchymal stem cells (MSCs), identified using nestin expression, constitute an essential HSC niche component. Nestin(+) MSCs contain all the bone-marrow colony-forming-unit fibroblastic activity and can be propagated as non-adherent 'mesenspheres' that can self-renew and expand in serial transplantations. Nestin(+) MSCs are spatially associated with HSCs and adrenergic nerve fibres, and highly express HSC maintenance genes. These genes, and others triggering osteoblastic differentiation, are selectively downregulated during enforced HSC mobilization or beta3 adrenoreceptor activation. Whereas parathormone administration doubles the number of bone marrow nestin(+) cells and favours their osteoblastic differentiation, in vivo nestin(+) cell depletion rapidly reduces HSC content in the bone marrow. Purified HSCs home near nestin(+) MSCs in the bone marrow of lethally irradiated mice, whereas in vivo nestin(+) cell depletion significantly reduces bone marrow homing of haematopoietic progenitors. These results uncover an unprecedented partnership between two distinct somatic stem-cell types and are indicative of a unique niche in the bone marrow made of heterotypic stem-cell pairs.",
"Inactivation of beta-catenin in mesenchymal progenitors prevents osteoblast differentiation; inactivation of Lrp5, a gene encoding a likely Wnt coreceptor, results in low bone mass (osteopenia) by decreasing bone formation. These observations indicate that Wnt signaling controls osteoblast differentiation and suggest that it may regulate bone formation in differentiated osteoblasts. Here, we study later events and find that stabilization of beta-catenin in differentiated osteoblasts results in high bone mass, while its deletion from differentiated osteoblasts leads to osteopenia. Surprisingly, histological analysis showed that these mutations primarily affect bone resorption rather than bone formation. Cellular and molecular studies showed that beta-catenin together with TCF proteins regulates osteoblast expression of Osteoprotegerin, a major inhibitor of osteoclast differentiation. These findings demonstrate that beta-catenin, and presumably Wnt signaling, promote the ability of differentiated osteoblasts to inhibit osteoclast differentiation; thus, they broaden our knowledge of the functions Wnt proteins have at various stages of skeletogenesis."
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what is ccsk
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Clear cell sarcoma of the kidney (CCSK) is the second most common pediatric renal tumor.
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[
"Clear cell sarcoma of the kidney (CCSK) is a rare paediatric renal malignant tumour. The majority of CCSKs have internal tandem duplications (ITDs) of the BCOR gene, whereas a minority have the YWHAE-NUTM2 gene fusion. A third 'double-negative' (DN) category comprises CCSKs with neither BCOR ITDs nor YWHAE-NUTM2 fusion. The aim of this study was to characterise 11 histologically diagnosed CCSKs immunohistochemically (with CCND1, BCOR and CCNB3 stains) and genetically. By next-generation sequencing, 10 cases (90.9%) had BCOR exon 15 ITDs, with positive BCOR immunoreactivity being found in four (36%) or eight (72%) cases, depending on the antibody clone. By reverse transcription polymerase chain reaction, none had the YWHAE-NUTM2 fusion. The DN case had a BCOR-CCNB3 fusion and strong nuclear CCNB3 and BCOR immunoreactivity. Quantitative polymerase chain reaction showed markedly elevated BCOR expression in this case, whereas BCOR ITD cases had lower levels of elevated BCOR expression. The majority of the CCSKs in our cohort had BCOR ITDs, and none had the YWHAE-NUTM2 fusion. We verified the strong, diffuse cyclin D1 (CCND1) immunoreactivity in CCSKs described in recent reports. BCOR immunoreactivity was not consistently positive in all CCSKs with BCOR ITDs, and therefore cannot be used as a diagnostic immunohistochemical stain to identify BCOR ITD cases. The DN case was a BCOR-CCNB3 fusion sarcoma. BCOR-CCNB3 sarcoma is typically a primary bone sarcoma affecting male adolescents, and this is the first report of it presenting in a kidney of a young child as a CCSK. The full spectrum of DN CCSKs awaits more comprehensive characterisation.",
"Clear Cell Sarcoma of the Kidney (CCSK) is a rare childhood renal tumour. Only a few homogeneously treated CCSK cohorts have been reported. This study aims to describe clinical characteristics and survival of CCSK patients treated according to recent International Society of Pediatric Oncology (SIOP) protocols. We analysed the prospectively collected data of patients with a histologically verified CCSK, entered onto SIOP 93-01/2001 trials. A total of 191 CCSK patients (64% male) were analysed, with a median age at diagnosis of 2.6 years. Stage distribution for stages I, II, III and IV was 42%, 23%, 28% and 7%, respectively. Pre-operative chemotherapy was administered to 169/191 patients. All patients underwent total nephrectomy and 189/191 patients received post-operative chemotherapy. Radiotherapy was applied in 2/80 stage I, 33/44 stage II, 44/54 stage III and 6/13 stage IV patients. Five year event-free survival (EFS) and overall survival (OS) were 79% (95% confidence interval (CI): 73-85%) and 86% (95% CI: 80-92%) respectively. Stage IV disease and young age were significant adverse prognostic factors for event-free survival. Factors such as gender, tumour volume and type of initial treatment were not found to be prognostic for EFS and OS. In this largest SIOP cohort described so far, overall outcome of CCSK is reasonable, although treatment of young and advanced-stage disease patients is challenging. As further intensification of treatment is hampered by direct and late toxicity, future directions should include the development of targeted therapy based on specific molecular aberrations of CCSK.",
"To explore the role of NQO1 overexpression for prognostic implication in hepatocellular carcinoma (HCC), NQO1 mRNA levels were detected in HCC fresh tissue samples of HCC and nontumor tissues, respectively. One hundred fifty-six cases of HCC meeting strict follow-up criteria were selected for immunohistochemical staining of NQO1 protein. Correlations between NQO1 overexpression and clinicopathological features of HCC were evaluated using χ2 tests, survival rates were calculated using the Kaplan-Meier method, and the relationship between prognostic factors and patient 5-year survival was analyzed using Cox proportional hazards analysis. In results, the levels of NQO1 mRNA were significantly up-regulated in 14 fresh tissue samples of HCC. Immunohistochemical analysis showed that the NQO1 expression and overexpression rates were significantly higher in HCC samples compared with either adjacent nontumor tissues or normal liver tissues. NQO1 overexpression correlated to tumor size, venous infiltration and late pTNM stage of HCC. NQO1 overexpression was also related to low disease-free survival and 5-year survival rates. In the late-stage group, disease-free and 5-year survival rates of patients with NQO1 overexpression were significantly lower than those of patients without NQO1 expression. Further analysis using a Cox proportional hazards regression model revealed that NQO1 expression emerged as a significant independent hazard factor for the 5-year survival rate of patients with HCC. Therefore, NQO1 plays an important role in the progression of HCC. NQO1 may potentially be used as an independent biomarker for prognostic evaluation of HCC.",
"Although hepatoblastoma is the most common pediatric liver cancer, its genetic heterogeneity and therapeutic targets are not well elucidated. Therefore, we conducted a multiomics analysis, including mutatome, DNA methylome, and transcriptome analyses, of 59 hepatoblastoma samples. Based on DNA methylation patterns, hepatoblastoma was classified into three clusters exhibiting remarkable correlation with clinical, histological, and genetic features. Cluster F was largely composed of cases with fetal histology and good outcomes, whereas clusters E1 and E2 corresponded primarily to embryonal/combined histology and poor outcomes. E1 and E2, albeit distinguishable by different patient age distributions, were genetically characterized by hypermethylation of the HNF4A/CEBPA-binding regions, fetal liver-like expression patterns, upregulation of the cell cycle pathway, and overexpression of NQO1 and ODC1. Inhibition of NQO1 and ODC1 in hepatoblastoma cells induced chemosensitization and growth suppression, respectively. Our results provide a comprehensive description of the molecular basis of hepatoblastoma and rational therapeutic strategies for high-risk cases.",
"Clear cell sarcoma of kidney (CCSK) is classified as a tumour of unfavourable histology by the National Wilms' Tumor Study Group. It has worse clinical outcomes than Wilms' tumour. Virtually nothing is known about CCSK biology, as there have been very few genetic aberrations identified to act as pointers in this cancer. Three cases of CCSK bearing a chromosomal translocation, t(10;17)(q22;p13), have been individually reported but not further investigated to date. The aim of this research was to characterize t(10;17)(q22;p13) in CCSK to identify the genes involved in the translocation breakpoints. Using fluorescently labelled bacterial artificial chromosomes (BACs) and a chromosome-walking strategy on an index case of CCSK with t(10;17)(q22;p13) by karyotype, we identified the chromosomal breakpoints on 17p13.3 and 10q22.3. The translocation results in rearrangement of YWHAE on chromosome 17 and FAM22 on chromosome 10, producing an in-frame fusion transcript of ∼3 kb, incorporating exons 1-5 of YWHAE and exons 2-7 of FAM22, as determined by RT-PCR using YWHAE- and FAM22-specific primers. The YWHAE-FAM22 transcript was detected in six of 50 further CCSKs tested, therefore showing an overall incidence of 12% in our cohort. No transcript-positive cases presented with stage I disease, despite this being the stage for 31% of our cohort. Tumour cellularity was significantly higher in the cases that were transcript-positive. Based on the chromosome 10 breakpoint identified by FISH and the sequences of the full-length transcripts obtained, the FAM22 members involved in the translocation in these CCSK cases include FAM22B and FAM22E. Elucidation of the role of YWHAE-FAM22 in CCSK will assist development of more efficient and targeted therapies for this childhood cancer, which currently has poor outcomes.",
"Despite being the most common liver cancer in children, hepatoblastoma (HB) is a rare neoplasm. Consequently, few pretreatment tumors have been molecularly profiled, and there are no validated prognostic or therapeutic biomarkers for HB patients. We report on the first large-scale effort to profile pretreatment HBs at diagnosis. Our analysis of 88 clinically annotated HBs revealed three risk-stratifying molecular subtypes that are characterized by differential activation of hepatic progenitor cell markers and metabolic pathways: high-risk tumors were characterized by up-regulated nuclear factor, erythroid 2-like 2 activity; high lin-28 homolog B, high mobility group AT-hook 2, spalt-like transcription factor 4, and alpha-fetoprotein expression; and high coordinated expression of oncofetal proteins and stem-cell markers, while low-risk tumors had low lin-28 homolog B and lethal-7 expression and high hepatic nuclear factor 1 alpha activity. Analysis of immunohistochemical assays using antibodies targeting these genes in a prospective study of 35 HBs suggested that these candidate biomarkers have the potential to improve risk stratification and guide treatment decisions for HB patients at diagnosis; our results pave the way for clinical collaborative studies to validate candidate biomarkers and test their potential to improve outcome for HB patients. (Hepatology 2017;65:104-121).",
"Deletion of all or part of chromosome 9q is the most common genetic alteration in all stages and grades of bladder cancer. DBCCR1 (deleted in bladder cancer chromosome region candidate 1) maps to the chromosome region 9q32-33, a candidate tumour suppressor locus for bladder cancer. Although no mutations of DBCCR1 have been detected in bladder tumours, expression of DBCCR1 is silenced by promoter hypermethylation in 50% of bladder cancer cell lines analysed. Here we sought to provide functional evidence to authenticate DBCCR1 as a tumour suppressor using gene-transfer methods. Exogenous expression of DBCCR1 protein or an HA epitope-tagged fusion protein, HA-DBCCR1 in NIH3T3 cells and human bladder tumour cell lines resulted in suppression of proliferation. Cell cycle analyses in NIH3T3 cells revealed that DBCCR1-mediated growth inhibition was due to an increase in the number of cells in the G(1) phase of the cell cycle. The levels of apoptosis were not altered. These results demonstrate a role for DBCCR1 in cell cycle control, thereby supporting the hypothesis that this is the tumour suppressor gene targeted by 9q32-33 deletion in bladder cancer."
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Peri-menarchal screening for asymptomatic Müllerian anomalies in girls with contralateral solitary functioning kidney.
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Unilateral renal agenesis and multicystic dysplastic kidney, resulting in a contralateral solitary functioning kidney (SFK), are part of the broad spectrum of congenital anomalies of the kidney and urinary tract (CAKUT). In girls with SFK, screening for asymptomatic Müllerian anomalies of uterus and vagina is not yet routinely performed, and therefore often overlooked until clinical complications in the menstrual cycle or fertility process occur. In this case series, we report on four teenagers with congenital SFK presenting with menstrual problems due to a Müllerian anomaly. Routine peri-menarchal screening for Müllerian anomalies in girls with SFK may provide timely counseling, surgical treatment and prevention of associated complications such as endometriosis, infertility and miscarriages.
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[
"Müllerian duct anomalies encompass a wide variety of disorders resulting from abnormalities in the embryological development of the Müllerian ducts. In the prepubertal pediatric population, Müllerian duct anomalies are often incidental findings on studies obtained for other reasons. The onset of menses can prompt more clinical symptoms. Proper characterization of Müllerian duct anomalies is important because these anomalies can affect the development of gynecological disorders as well as fertility. Müllerian duct anomalies also carry a high association with other congenital anomalies, particularly renal abnormalities. MRI is widely considered the best modality for assessing Müllerian duct anomalies; it provides multiplanar capability, clear anatomical detail and tissue characterization without ionizing radiation. MRI allows for careful description of Müllerian duct anomalies, often leading to classification into the most widely accepted classification system for Müllerian duct anomalies. This system, developed by the American Society of Reproductive Medicine, includes seven subtypes: uterine agenesis/hypoplasia, unicornuate, didelphys, bicornuate, septate, arcuate, and diethylstilbestrol (DES) drug-related uterus. In cases of complex anomalies that defy classification, MRI allows detailed depiction of all components of the anatomical abnormality, allowing for proper management and surgical planning.",
"To describe congenital malformations and coexisting disorders occurring in 125 Polish women with Mayer-Rokitansky-Küster-Hauser syndrome (MRKHS). The syndrome is defined as uterovaginal aplasia in female with normal 46,XX karyotype. A retrospective analysis of the clinical data of MRKHS patients diagnosed or treated at the Gynecology and Obstetrics Clinical Hospital of Poznan University of Medical Sciences between 2010 and 2015. Sixty-eight patients (54,4%) were found to have one or more coexisting anomalies. Thirty-eight patients (55,9% of cases with concomitant malformations, 30,4% of the entire study group) had coexisting anomalies of at least two organ systems. The most frequent extragenital malformations were skeletal anomalies found in 40 patients (32%) and renal anomalies found in 36 patients (28,8%). Fifty-seven patients (45,6%) were diagnosed with typical form (type 1) and 16 (12,8%) with the atypical form (type 2) of MRKHS. In the other 52 patients (41,6%) we diagnosed MURCS association. Five of our patients (4%) had karyotype abnormalities. Our study confirms complexity and clinical heterogeneity of MRKHS. Concomitant congenital malformations are present in about half of MRKHS women. A significant proportion of patients have coexisting anomalies of at least two organ systems. The most common coexisting findings are musculoskeletal and renal abnormalities. Chromosomal aberrations may be present in patients with either typical or atypical form of MRKHS.",
"Herlyn-Werner-Wunderlich syndrome (HWWS) is a rare variant of Mullerian ductal anomaly associated with ipsilateral renal agenesis. Most patients are diagnosed after menarche with complications of uterovaginal obstruction, genitourinary infection and pelvic adhesions. Patients often undergo additional operations for misdiagnosis or treatment of complications. Our institution manages several HWWS patients diagnosed before symptoms by screening for antenatally-diagnosed renal agenesis. This study aims to improve the presymptomatic management of HWWS patients. We carried out retrospective case review of patients diagnosed with HWWS from 2010 to 2017 on patient demographics, symptoms, clinical course and operative management and summarize the sparse literature published to date. There were 8 patients with HWWS but only 2 symptomatic patients presented acutely with hematocolpos requiring urgent vaginal surgery. The other six patients had early diagnosis through postnatal ultrasound screening. No patient required further operation for diagnosis or complications related to obstructed hemivagina. Our case series and literature review show that the majority of prepubertal patients with HWWS do not require early gynecological surgery. We recommend that female babies with renal agenesis should be screened for HWWS syndrome with ultrasound. Early diagnosis and presymptomatic elective surgery may prevent urogynecological complications that cause fertility and renal impairment. Case series, level IV evidence.",
"Mayer-Rokitansky-Küster-Hauser syndrome causes absence or underdevelopment of uterus and vagina, but women's subjective experience remains understudied. This systematic review was conducted to examine the psychological and health-related quality-of-life outcomes of Mayer-Rokitansky-Küster-Hauser syndrome. In total, 22 articles identified through electronic search matched the inclusion criteria and were included in our review. Mayer-Rokitansky-Küster-Hauser syndrome may be associated with psychological symptoms and impaired quality of life, but especially with poor sexual esteem and genital image. Women may experience difficulties managing intimacy and disclosing to partners. Mothers may be perceived as overinvolved, with consequent negative emotions in women with the disease.",
"Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome is characterized by congenital absence of the vagina and uterus. We conducted genome-wide SNP analyses and exome sequencing to detect the causes of MRKH syndrome. We identified de novo variants of MYCBP2, NAV3, and PTPN3 in three families and a variant of MYCBP2 in a sporadic case. Here, we demonstrated the partial genetic makeup of Japanese MRKH syndrome.",
"Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome, also referred to as Müllerian aplasia, is a congenital disorder characterized by aplasia of the uterus and upper part of the vagina in females with normal secondary sex characteristics and a normal female karyotype (46,XX). The diagnosis is often made during adolescence following investigations for primary amenorrhea and has an estimated prevalence of 1 in 5000 live female births. MRKH syndrome is classified as type I (isolated uterovaginal aplasia) or type II (associated with extragenital manifestations). Extragenital anomalies typically include renal, skeletal, ear, or cardiac malformations. The etiology of MRKH syndrome still remains elusive, however increasing reports of familial clustering point towards genetic causes and the use of various genomic techniques has allowed the identification of promising recurrent genetic abnormalities in some patients. The psychosexual impact of having MRKH syndrome should not be underestimated and the clinical care foremost involves thorough counselling and support in careful dialogue with the patient. Vaginal agenesis therapy is available for mature patients following therapeutical counselling and education with non-invasive vaginal dilations recommended as first-line therapy or by surgery. MRKH syndrome involves absolute uterine factor infertility and until recently, the only option for the patients to achieve biological motherhood was through gestational surrogacy, which is prohibited in most countries. However, the successful clinical trial of uterus transplantation (UTx) by a Swedish team followed by the first live-birth in September, 2014 in Gothenburg, proofed the first available fertility treatment in MRKH syndrome and UTx is now being performed in other countries around the world allowing women with MRKH syndrome to carry their own child and achieve biological motherhood. Several advances in research across multiple disciplines have been made in the recent years and this kaleidoscopic review provides a current status of various key aspects in MRKH syndrome and provides perspectives for future research and improved clinical care.",
"To examine the process and emotional effect of disclosing a personal diagnosis of Mayer-Rokitansky-Küster-Hauser syndrome (MRKH) to peers during adolescence and young adulthood. Qualitative study using semistructured telephone interviews. Nine women diagnosed with MRKH, aged 21-31 years, recruited via patient support groups. Motivators and barriers to self-disclosure of a diagnosis of MRKH to peers and partners. Motivators to tell peers about a diagnosis included significant trust in the relationship (whether platonic or romantic), needing to unload the experienced burden of diagnosis, and a sense of responsibility to be forthcoming if a long-term romantic future was desired. The most common barrier to telling others was fear of rejection or being labeled a \"freak.\" Although most participants did not receive guidance from a health care provider regarding approaches to sharing diagnostic information with others, almost all participants reported wishing they had received such counseling. A diagnosis of MRKH elicits recurring anxieties about disclosure and the effect on relationships that are inadequately addressed by health care providers. Guidance and support on disclosure to friends and romantic partners should be provided whenever possible."
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Health-related quality of life and clinical and imaging outcome measures in secondary progressive MS
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Health-related quality of life (HRQOL) outcomes are often included as secondary outcomes in clinical trials in secondary progressive MS (SPMS), but little is known about the longitudinal association of HRQOL and clinical and imaging outcome measures in SPMS.
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[
"The timed 25-foot walk (T25FW) is a key clinical outcome measure in multiple sclerosis patient management and clinical research. To evaluate T25FW performance and factors associated with its change in the Multiple Sclerosis Outcome Assessments Consortium (MSOAC) Placebo Database (n = 2465). We created confirmed disability progression (CDP) variables for T25FW and Expanded Disability Status Scale (EDSS) outcomes. We used intraclass correlation coefficients (ICCs) and Bland Altman plots to evaluate reliability. We evaluated T25FW changes and predictive validity using a mixed-effects model, survival analysis, and nested case-control analysis. The mean baseline score for the T25FW in this study population was 9.2 seconds, median = 6.1 (standard deviation = 11.0, interquartile range (IQR) = 4.8, 9.0). The T25FW measure demonstrated excellent test-retest reliability (ICC = 0.98). Walk times increased with age, disability, disease type, and disease duration; relapses were not associated with an increase. Patients with T25FW progression had a faster time to EDSS-CDP compared to those without (hazards ratio (HR): 2.6; confidence interval (CI): 2.2, 3.1). Changes in the T25FW were more likely to precede changes in EDSS. This research confirms the association of the T25FW with disability and provides some evidence of predictive validity. Our findings support the continued use of the T25FW in clinical practice and clinical trials.",
"Cognitive impairment in MS impacts negatively on many patients at all disease stages and in all subtypes. Full clinical cognitive assessment is expensive, requiring expert staff and special equipment. Test versions and normative data are not available for all languages and cultures. To recommend a brief cognitive assessment for multiple sclerosis (MS) that is optimized for small centers, with one or few staff members, who may not have neuropsychological training and constructed to maximize international use. An expert committee of twelve members representing the main cultural groups that have so far contributed considerable data about MS cognitive dysfunction was convened. Following exhaustive literature review, peer-reviewed articles were selected to cover a broad spectrum of cultures and scales that targeted cognitive domains vulnerable to MS. Each was rated by two committee members and candidates scales were rated on psychometric qualities (reliability, validity, and sensitivity), international application, ease of administration, feasibility in the specified context, and acceptability to patients. The committee recommended the Symbol Digit Modalities Test, if only 5 minutes was available, with the addition of the California Verbal Learning Test - Second Edition and the Brief Visuospatial Memory Test - Revised learning trials if a further 10 minutes could be allocated for testing. A brief cognitive assessment for MS has been recommended. A validation protocol has been prepared for language groups and validation studies have commenced.",
"The association of pathology and neurological deficit with quality of life (QoL) in multiple sclerosis (MS) is not fully understood. In this study, magnetic resonance imaging (MRI) measures of pathology--T1 and T2 lesion volume and ratio; active T2 lesion number; global and regional brain volume and atrophy; magnetization transfer ratio (MTR) for lesions, normal appearing grey and white matter (NAGM, NAWM); and spinal cord cross-sectional area-and measures of neurological disability (expanded disability status scale, EDSS), deficit (MS functional composite, MSFC) and inflammatory activity (relapse rate) were compared with the MS impact scale (MSIS-29), in participants in a trial of lamotrigine in secondary progressive MS. Data were collected from 118 people (85 female:33 male) aged 30-61 years (mean 50.6 years)--median EDSS 6.0 (range 4.0-7.5); mean disease duration 20.1 years (range 3-41)--at baseline and 2 years. Regression analysis was used to identify independently significant cross-sectional and longitudinal correlates of the physical (MSIS-phys) and psychological (MSIS-psych) components of the MSIS-29; longitudinal analysis using the 57 people in the placebo arm. The only independently significant correlate of MSIS-phys was 1/timed walk (TW) (p < 0.0001, R (2) = 0.13; p = 0.047, R (2) = 0.09); cross-sectionally the best model for MSIS-psych was the paced auditory serial addition test (PASAT-3) (p = 0.041) and T1-to-T2 lesion volume ratio (p = 0.009) (R (2) = 0.13); longitudinally it was change in 1/TW (p = 0.007), mean NAWM MTR (p = 0.003) and NAGM peak height (p = 0.048) (R (2) = 0.32). These data show that MRI measures and clinical measures do impact on quality of life, but the association is limited.",
"Cognitive and motor performance measures are commonly employed in multiple sclerosis (MS) research, particularly when the purpose is to determine the efficacy of treatment. The increasing focus of new therapies on slowing progression or reversing neurological disability makes the utilization of sensitive, reproducible, and valid measures essential. Processing speed is a basic elemental cognitive function that likely influences downstream processes such as memory. The Multiple Sclerosis Outcome Assessments Consortium (MSOAC) includes representatives from advocacy organizations, Food and Drug Administration (FDA), European Medicines Agency (EMA), National Institute of Neurological Disorders and Stroke (NINDS), academic institutions, and industry partners along with persons living with MS. Among the MSOAC goals is acceptance and qualification by regulators of performance outcomes that are highly reliable and valid, practical, cost-effective, and meaningful to persons with MS. A critical step for these neuroperformance metrics is elucidation of clinically relevant benchmarks, well-defined degrees of disability, and gradients of change that are deemed clinically meaningful. This topical review provides an overview of research on one particular cognitive measure, the Symbol Digit Modalities Test (SDMT), recognized as being particularly sensitive to slowed processing of information that is commonly seen in MS. The research in MS clearly supports the reliability and validity of this test and recently has supported a responder definition of SDMT change approximating 4 points or 10% in magnitude.",
"The Multiple Sclerosis Outcome Assessments Consortium (MSOAC) was formed by the National MS Society to develop improved measures of multiple sclerosis (MS)-related disability. (1) To assess the current literature and available data on functional performance outcome measures (PerfOs) and (2) to determine suitability of using PerfOs to quantify MS disability in MS clinical trials. (1) Identify disability dimensions common in MS; (2) conduct a comprehensive literature review of measures for those dimensions; (3) develop an MS Clinical Data Interchange Standards Consortium (CDISC) data standard; (4) create a database of standardized, pooled clinical trial data; (5) analyze the pooled data to assess psychometric properties of candidate measures; and (6) work with regulatory agencies to use the measures as primary or secondary outcomes in MS clinical trials. Considerable data exist supporting measures of the functional domains ambulation, manual dexterity, vision, and cognition. A CDISC standard for MS ( http://www.cdisc.org/therapeutic#MS ) was published, allowing pooling of clinical trial data. MSOAC member organizations contributed clinical data from 16 trials, including 14,370 subjects. Data from placebo-arm subjects are available to qualified researchers. This integrated, standardized dataset is being analyzed to support qualification of disability endpoints by regulatory agencies.",
"The need for more robust outcomes in multiple sclerosis (MS) clinical trials has been a main priority of the field for decades. Dissatisfaction with existing measures has led to several consensus meetings and initiatives over the past few decades in hopes of defining and gaining acceptance of measures that are valid, reliable, sensitive to change and progression, and most importantly, relevant to those living with MS. The Multiple Sclerosis Outcome Assessments Consortium (MSOAC) was formed for this purpose. The objective of this paper is to describe the results of the MSOAC plan to obtain qualification for a cognitive performance measure that meets these requirements. Using data from 14 MS disease-modifying registration trials, we completed a comprehensive examination of the psychometric qualities of the Symbol Digit Modalities Test (SDMT) and the Paced Auditory Serial Addition Test (PASAT) with the goal of compiling evidence to support the utilization of one of these measures in future clinical trials. Consistent with the published literature, the SDMT proved superior to the PASAT. The SDMT should be considered the measure of choice for MS trials in assessing cognitive processing speed."
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Sialyltransferases in cancer biology and immunity
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Aberrant glycosylation is a key feature of malignant transformation. Hypersialylation, the enhanced expression of sialic acid-terminated glycoconjugates on the cell surface, has been linked to immune evasion and metastatic spread, eventually by interaction with sialoglycan-binding lectins, including Siglecs and selectins. The biosynthesis of tumor-associated sialoglycans involves sialyltransferases, which are differentially expressed in cancer cells. In this review article, we provide an overview of the twenty human sialyltransferases and their roles in cancer biology and immunity. A better understanding of the individual contribution of select sialyltransferases to the tumor sialome may lead to more personalized strategies for the treatment of cancer.
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"Potent, cell-permeable, and subtype-selective sialyltransferase inhibitors represent an attractive family of substances that can potentially be used for the clinical treatment of cancer metastasis. These substances operate by specifically inhibiting sialyltransferase-mediated hypersialylation of cell surface glycoproteins or glycolipids, which then blocks the sialic acid recognition pathway and leads to deterioration of cell motility and invasion. A vast amount of evidence for the in vitro and in vivo effects of sialyltransferase inhibition or knockdown on tumor progression and tumor cell metastasis or colonization has been accumulated over the past decades. In this regard, this review comprehensively discusses the results of studies that have led to the recent discovery and development of sialyltransferase inhibitors, their potential biomedical applications in the treatment of cancer metastasis, and their current limitations and future opportunities.",
"Our study deals with the interaction of CD33 related-siglecs-5,-7,-8,-9,-10 with gangliosides GT1b, GQ1b, GD3, GM2, GM3 and GD1a. Siglec-5 bound preferentially to GQ1b, but weakly to GT1b, whereas siglec-10 interacted only with GT1b ganglioside. Siglec-7 and siglec-9 displayed binding to gangliosides GD3, GQ1b and GT1b bearing a disialoside motif, though siglec-7 was more potent; besides, siglec-9 interacted also with GM3. Siglec-8 demonstrated low affinity to the gangliosides tested compared with other siglecs. Despite high structural similarity of CD33 related siglecs, they demonstrated different ganglioside selectivity, in particular to the Neu5Acalpha2-8Neu5Ac motif.",
"The sialyltransferase activities of 10 human colorectal specimens were compared with those of the corresponding adjacent normal mucosa. Using asialofetuin as an acceptor we found, in tumor tissues of 9 out of 10 patients, an increased sialyltransferase activity towards the N-linked chains as determined upon peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase (PNGase) treatment. On the contrary, the activity towards the O-linked chains was not significantly changed. When the specificity of the sialyltransferase acting on N-linked chains was investigated by using N-acetyl-lactosamine (Gal beta 1,4GlcNAc) as an acceptor, we found that the alpha 2,6 sialyltransferase activity expressed by both normal and tumor colorectal tissues was far higher than the alpha 2,3-activity and that alpha 2,6 was the only sialyltransferase activity increased in tumor tissues. Kinetic analysis revealed that normal and tumor alpha 2,6 sialyltransferases have the same apparent Km for the acceptor substrate (469 and 465 microns), but normal enzyme has a higher Km for CMP-NeuAc (303 microns) than the tumor enzyme (50 microns). The higher affinity of tumor enzyme for the nucleotide-sugar might partially explain its increased activity in tumor tissues. In addition, tumor tissues contain a lower amount of sialic acid despite the increase in alpha 2,6 sialyltransferase activity.",
"Hypersialylation of tumor cell surface proteins along with a marked upregulation of sialyltransferase (ST) activity is a well-established hallmark of cancer. Due to the critical role of STs in tumor growth and progression, ST inhibition has emerged as a potential new antimetastatic strategy for a range of cancers including pancreatic and ovarian. Human STs are divided into subtypes based on their linkage and acceptor molecule, with each subtype controlling the synthesis of specific sialylated structures with unique biological roles. This has important implications for inhibitor development, as STs also play significant roles in immune responses, inflammation, viral infection, and neurological disorders. Thus, the current goal in order to advance to the clinic is the development of subtype selective, cell-permeable and synthetically accessible, small-molecule ST inhibitors. Herein is a comprehensive review of the latest developments in ST inhibitors from design, Nature, and high-throughput screening, addressing both the challenges and opportunities in targeting cell surface sialylation. The review features an overview of the biological evaluation methods, computational and imaging tools, inhibitor molecular diversity, and selectivity toward ST subtypes, along with the emerging role of ST inhibitors as diagnostic tools for disease imaging.",
"Emerging evidence suggests an immunosuppressive role of altered tumor glycosylation due to downregulation of innate immune responses via immunoregulatory Siglecs. In contrast, human T cells, a major anticancer effector cell, only rarely express Siglecs. However, here, we report that the majority of intratumoral, but not peripheral blood, cytotoxic CD8+ T cells expressed Siglec-9 in melanoma. We identified Siglec-9+ CD8+ T cells as a subset of effector memory cells with high functional capacity and signatures of clonal expansion. This cytotoxic T-cell subset was functionally inhibited in the presence of Siglec-9 ligands or by Siglec-9 engagement by specific antibodies. TCR signaling pathways and key effector functions (cytotoxicity, cytokine production) of CD8+ T cells were suppressed by Siglec-9 engagement, which was associated with the phosphorylation of the inhibitory protein tyrosine phosphatase SHP-1, but not SHP-2. Expression of cognate Siglec-9 ligands was observed on the majority of tumor cells in primary and metastatic melanoma specimens. Targeting the tumor-restricted, glycosylation-dependent Siglec-9 axis may unleash this intratumoral T-cell subset, while confining T-cell activation to the tumor microenvironment.",
"The alpha2,3 sialyltransferase, alpha2,3 SAT (O), catalyzes the transfer of sialic acid to Galbeta1,3 N-acetyl-D-galactosamine (GalNAc) (core-1) in mucin type O-glycosylation, and thus terminates chain extension. A Core-2 branch can also be formed from core-1 by the core-2 beta1,6 N-acetyl-d-glucosamine transferase (beta1,6 GlcNAc T) that leads to chain extension. Increased levels of the alpha2,3 SAT (O) and decreased levels of the core-2 beta1,6 GlcNAc T are seen in breast cancer cells and correlate with differences in the structure of the O-glycans synthesized (Brockhausen et al., 1995; Lloyd et al., 1996). Since in mucin type O-glycosylation sugars are added individually and sequentially in the Golgi apparatus, the position of the transferases, as well as their activity, can determine the final structure of the O-glycans synthesized. A cDNA coding for the human alpha2,3 SAT (O) tagged with an immunoreactive epitope from the myc gene has been used to map the position of the glycosyltransferase in nontumorigenic (MTSV1-7) and malignant (T47D) breast epithelial cell lines. Transfectants were analyzed for expression of the enzyme at the level of message and protein, as well as for enzymic activity. In T47D cells, which do not express core-2 beta1,6 GlcNAc T, the increased activity of the sialyltransferase correlated with increased sialylation of core-1 O-glycans on the epithelial mucin MUC1. Furthermore, in MTSV1-7 cells, which do express core-2 beta1,6 GlcNAc T, an increase in sialylated core-1 structures is accompanied by a reduction in the ratio of GlcNAc: GalNAc in the O-glycans attached to MUC1, implying a decrease in branching. Using quantitative immunoelectron microscopy, the sialyltransferase was mapped to the medial- and trans-Golgi cisternae, with some being present in the TGN. The data represent the first fine mapping of a sialyltransferase specifically active in O-glycosylation and demonstrate that the structure of O-glycans synthesized by a cell can be manipulated by transfecting with recombinant glycosyltransferases.",
"Human gliomas express very high levels of cell-surface alpha2,3-linked terminal sialic acids on glycoproteins bearing N-linked oligosaccharides, most notably on alpha3beta1 integrin, which is the predominant integrin found in these tumors. Alpha2,6-linked terminal sialic acids, however, are not expressed. Two stable transfectants were made using a tumorigenic human glioma cell line, U-373 MG. Galbeta1,4GlcNAc alpha2,6-sialyltransferase (ST6Gal I) transfectants were made to replace the endogenous alpha2,3-linked sialic acids with alpha2,6-linked sialic acids. And Galbeta1,3(4)GlcNAc alpha2,3-sialyltransferase (ST3Gal III) transfectants were made to increase further the expression of cell-surface, N-glycan, alpha2,3-linked sialic acids. Although ST3Gal III transfection resulted in increased invasivity when compared with parental U-373 MG and vector-transfected control cells in vitro, ST6Gal I transfection abolished invasion in vitro and induced alterations in both cell morphology, cell-spreading, and adhesion-mediated protein tyrosine phosphorylation. Furthermore, the ST6Gal I transfectants produced no intracranial tumors in severe combined immunodeficient mice, whereas parental U-373 MG cells, the vector-transfected control cells, and ST3Gal III-transfected U-373 MG cells did. These results suggest that both the linkage and expression levels of the terminal sialic acids of alpha3beta1 integrin N-glycans play an important role in glioma cell-extracellular matrix interactions. Thus, manipulating ST6Gal I gene expression may have therapeutic potential for the treatment of malignant gliomas.",
"Metastasis of tumor cells is the most deadly attribute of breast cancer patients. Aberrant sialylation is closely associated with malignant phenotype of tumor cells, including invasiveness and metastasis. The objective of this study is to clarify the possible role and mechanism of ST6GalNAcII in the metastasis process of breast carcinoma. Real-time PCR, Western blot, and immunohistochemical were used to analyze differential expression of ST6GalNAc II in breast carcinoma cell lines and tissue specimens. PI3K/AKt signaling pathway was also analyzed. The high expression level of ST6GalNAcII was corresponding to invasive phenotype of breast cancer cells both in vitro and in vivo. Further data indicated that manipulation of ST6GalNAcII gene expression led to alter the activity of phosphoinositide-3 kinase (PI3K)/Akt signaling pathway. Blocking the PI3K/Akt pathway resulted in reduced capacity in invasion of MDA-MB-231 cells. ST6GalNAcII elucidated the unusual properties of invasion in breast cancer cell via modulating the PI3K/AKt signaling pathway.",
"Sialic acids are common monosaccharides that are widely expressed as outer terminal units on all vertebrate cell surfaces, and play fundamental roles in cell-cell and cell-microenvironment interactions. The predominant sialic acids on most mammalian cells are N-glycolylneuraminic acid (Neu5Gc) and N-acetylneuraminic acid (Neu5Ac). Neu5Gc is notable for its deficiency in humans due to a species-specific and species-universal inactivating deletion in the CMAH gene encoding the hydroxylase that converts CMP-Neu5Ac to CMP-Neu5Gc. However, Neu5Gc is metabolically incorporated into human tissues from dietary sources (particularly red meat), and detected at even higher levels in some human cancers. Early life exposure to Neu5Gc-containing foods in the presence of certain commensal bacteria that incorporate dietary Neu5Gc into lipooligosaccharides can lead to generation of antibodies that are also cross-reactive against Neu5Gc-containing glycans in human tissues (\"xeno-autoantigens\"). Such anti-Neu5Gc \"xeno-autoantibodies\" are found in all humans, although ranging widely in levels among individuals, and displaying diverse and variable specificities for the underlying glycan. Experimental evidence in a human-like Neu5Gc-deficient Cmah(-) (/) (-) mouse model shows that inflammation due to \"xenosialitis\" caused by this antigen-antibody interaction can promote tumor progression, suggesting a likely mechanism for the well-known epidemiological link between red meat consumption and carcinoma risk. In this review, we discuss the history of this field, mechanisms of Neu5Gc incorporation into tissues, the origin and specificities of human anti-Neu5Gc antibodies, their use as possible cancer biomarkers, implications of xenosialitis in cancer initiation and progression, and current and future approaches toward immunotherapy that could take advantage of this unusual human-specific phenomenon.",
"Gliomas are the most common intracranial primary tumors, for which very few therapeutic options are available. The most malignant subtype is the glioblastoma, a disease associated with a 5-year survival rate lower than 5%. Given that research in glycobiology continues highlighting the role of glycans in tumor cell biology, it offers an interesting niche for the search of new therapeutic targets. In this study, we characterized aberrant glycosylation and its impact on cell biology over a broad panel of high- and low-grade glioma cell lines. Results show high expression of terminal Lewis glycans, mainly SLex, and overexpression of sialyl- and fucosyltransferases involved in their biosynthesis in high-grade glioma cell lines. Moreover, we report an association of complex multi-antennary N-glycans presenting β1,6-GlcNAc branches with the high-grade glioma cells, which also overexpressed the gene responsible for these assemblies, MGAT5. In addition, downmodulation of N-glycosylation by treatment with the inhibitors Tunicamycin/Swainsonine or MGAT5 silencing decreased SLex expression, adhesion and migration in high-grade glioma cells. In contrast, no significant changes in these cell capacities were observed in low-grade glioma after treatment with the N-glycosylation inhibitors. Furthermore, inhibition of histone deacetylases by Trichostatin A provoked an increase in the expression of SLex and its biosynthetic related glycosyltransferases in low-grade glioma cells. Our results describe that aggressive glioma cells show high expression of Lewis glycans anchored to complex multi-antennary N-glycans. This glycophenotype plays a key role in malignant cell behavior and is regulated by histone acetylation dependent mechanisms."
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Curcumin: a dietary dye extracted from turmeric
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Curcumin is one of the dietary dyes extracted from turmeric and used for prevention and treatment of various illnesses. However, the low bioavailability and poor stability of curcumin limits its relevant applications. Therefore, different metal ions including Cu
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"Dysregulations in hepatic lipid synthesis are often associated with obesity and type 2 diabetes, and therefore a perfect understanding of the regulation of this metabolic pathway appears essential to identify potential therapeutic targets. Recently, the transcription factor ChREBP (carbohydrate-responsive element-binding protein) has emerged as a major mediator of glucose action on lipogenic gene expression and as a key determinant of lipid synthesis in vivo. Indeed, liver-specific inhibition of ChREBP improves hepatic steatosis and insulin resistance in obese ob/ob mice. Since ChREBP cellular localization is a determinant of its functional activity, a better knowledge of the mechanisms involved in regulating its nucleo-cytoplasmic shuttling and/or its post-translational activation is crucial in both physiology and physiopathology. Here, we review some of the studies that have begun to elucidate the regulation and function of this key transcription factor in liver.",
"A novel polymer of poloxamer188-b-PCL was synthesized via a ring-opening polymerization. Fourier transform infrared spectroscopy (FTIR), Raman, and 1H nuclear magnetic resonance (1H NMR) spectra were used to study the structures of obtained poloxamer188-b-PCL. The thermo-stability of poloxamer188 -b-PCL was carried out with a thermal gravimetric analyzer (TGA), and cytotoxicity was obtained using the CCK8 method. Cargo-free and curcumin (CUR)-loaded poloxamer188-b-PCL NPs were fabricated via the solvent evaporation method. The morphology, particle size distribution, and stability of cargo-free NPs were studied with a scanning electron microscope (SEM) and laser particle analyzer. The in vitro radioprotection activity of CUR-loaded NPs was performed. FTIR, Raman, and 1H NMR spectra confirmed that poloxamer188-b-PCL was obtained. TGA curves suggested poloxamer188-b-PCL had better thermo-stability than original poloxamer188. Cell tests suggested that the cargo-free NPs had no cytotoxicity. SEM image showed that the cargo-free NPs were spherical with a diameter of 100 nm. Free radical scavenging experiments proved that CUR-loaded NPs had better antioxidant activity than CUR solutions. CUR-loaded NPs could be detected in all tissues, including liver, kidneys and lung. In summary, this work demonstrated a feasibility of developing an injective formulation of CUR and provided a protection agent in caner radiotherapy.",
"Numerous natural products available over the counter are commonly consumed by healthy, sub-healthy or ill people for the treatment and prevention of various chronic diseases. Among them, a few dietary polyphenols, including the curry compound curcumin, have been attracting the most attention from biomedical researchers and drug developers. Unlike many so-called \"good drug candidates\", curcumin and several other dietary polyphenols do not have a single known therapeutic target or defined receptor. In addition, the bioavailability of these polyphenols is usually very low due to their poor absorption in the gut. These recently debated features have created enormous difficulties for drug developers. In this review, I do not discuss how to develop curcumin, other dietary polyphenols or their derivatives into pharmaceutical agents. Instead, I comment on how curcumin and dietary polyphenol research has enriched our knowledge of insulin signaling, including the presentation of my perspectives on how these studies will add to our understanding of the famous hepatic insulin function paradox.",
"Two stoichiometrically different copper(II) complexes of curcumin (stoichiometry, 1:1 and 1:2 for copper:curcumin), were examined for their superoxide dismutase (SOD) activity, free radical-scavenging ability and antioxidant potential. Both the complexes are soluble in lipids and DMSO. The formation constants of the complexes were determined by voltammetry. EPR spectra of the complexes in DMSO at 77K showed that the 1:2 Cu(II)-curcumin complex is square planar and the 1:1 Cu(II)-curcumin complex is distorted orthorhombic. Cu(II)-curcumin complex (1:1) with larger distortion from square planar structure shows higher SOD activity. These complexes inhibit gamma-radiation induced lipid peroxidation in liposomes and react with DPPH acting as free radical scavengers. One-electron oxidation of the two complexes by radiolytically generated azide radicals in Tx-100 micellar solutions produced phenoxyl radicals, indicating that the phenolic moiety of curcumin in the complexes participates in free radical reactions. Depending on the structure, these two complexes possess different SOD activities, free radical neutralizing abilities and antioxidant potentials. In addition, quantum chemical calculations with density functional theory have been performed to support the experimental observations.",
"Type 2 diabetes in humans is associated with increased de novo lipogenesis (DNL), increased fatty acid (FA) fluxes, decreased FA oxidation, and hepatic steatosis. In this condition, VLDL production is increased and resistant to suppressive effects of insulin. The relationships between hepatic FA metabolism, steatosis, and VLDL production are incompletely understood. We investigated VLDL-triglyceride and -apolipoprotein (apo)-B production in relation to DNL and insulin sensitivity in female ob/ob mice. Hepatic triglyceride (5-fold) and cholesteryl ester (15-fold) contents were increased in ob/ob mice compared with lean controls. Hepatic DNL was increased approximately 10-fold in ob/ob mice, whereas hepatic cholesterol synthesis was not affected. Basal rates of hepatic VLDL-triglyceride and -apoB100 production were similar between the groups. Hyperinsulinemic clamping reduced VLDL-triglyceride and -apoB100 production rates by approximately 60% and approximately 75%, respectively, in lean mice but only by approximately 20% and approximately 20%, respectively, in ob/ob mice. No differences in hepatic expression of genes encoding apoB and microsomal triglyceride transfer protein were found. Hepatic expression and protein phosphorylation of insulin receptor and insulin receptor substrate isoforms were reduced in ob/ob mice. Thus, strongly induced hepatic DNL is not associated with increased VLDL production in ob/ob mice, possibly related to differential hepatic zonation of apoB synthesis (periportal) and lipid accumulation (perivenous) and/or relatively low rates of cholesterogenesis. Insulin is unable to effectively suppress VLDL-triglyceride production in ob/ob mice, presumably because of impaired insulin signaling.",
"The transcription factor carbohydrate response element-binding protein (ChREBP) mediates insulin-independent, glucose-stimulated gene expression of multiple liver enzymes responsible for converting excess carbohydrate to fatty acids for long-term storage. To investigate ChREBP's role in the development of obesity and obesity-associated metabolic dysregulation, ChREBP-deficient mice were intercrossed with ob/ob mice. As a result of deficient leptin expression, ob/ob mice overeat, become obese and resistant to insulin, and display marked elevations in hepatic lipogenesis, gluconeogenesis, and plasma glucose and triglycerides. mRNA expression of all hepatic lipogenic enzymes was significantly lower in ob/ob-ChREBP-/- than in ob/ob mice, resulting in decreased hepatic fatty acid synthesis and normalization of plasma free fatty acid and triglyceride levels. Overall weight gain in addition to adiposity was reduced in the doubly deficient mice. The former was largely attributable to decreased food intake and may result from decreased hypothalamic expression of the appetite-stimulating neuropeptide agouti-related protein. mRNA expression and activity of gluconeogenic enzymes also was lower in the doubly deficient mice, contributing to significantly lower blood glucose levels. The results of this study suggest that inactivation of ChREBP expression not only reduces fat synthesis and obesity in ob/ob mice but also results in improved glucose tolerance and appetite control.",
"The onset of diabetic nephropathy (DN) is associated with both systemic and renal changes. Fibroblast growth factor (FGF)-21 prevents diabetic complications mainly by improving systemic metabolism. In addition, low-dose radiation (LDR) protects mice from DN directly by preventing renal oxidative stress and inflammation. In the present study, we tried to define whether the combination of FGF21 and LDR could further prevent DN by blocking its systemic and renal pathogeneses. To this end, type 2 diabetes was induced by feeding a high-fat diet for 12 wk followed by a single dose injection of streptozotocin. Diabetic mice were exposed to 50 mGy LDR every other day for 4 wk with and without 1.5 mg/kg FGF21 daily for 8 wk. The changes in systemic parameters, including blood glucose levels, lipid profiles, and insulin resistance, as well as renal pathology, were examined. Diabetic mice exhibited renal dysfunction and pathological abnormalities, all of which were prevented significantly by LDR and/or FGF21; the best effects were observed in the group that received the combination treatment. Our studies revealed that the additive renal protection conferred by the combined treatment against diabetes-induced renal fibrosis, inflammation, and oxidative damage was associated with the systemic improvement of hyperglycemia, hyperlipidemia, and insulin resistance. These results suggest that the combination treatment with LDR and FGF21 prevented DN more efficiently than did either treatment alone. The mechanism behind these protective effects could be attributed to the suppression of both systemic and renal pathways.",
"Recent advances have been made in the understanding of pharmacological and dietary agents that contribute to browning of white adipose tissue in order to combat obesity by promoting energy expenditure. Here, we show that curcumin induces browning of 3T3-L1 and primary white adipocytes via enhanced expression of brown fat-specific genes. Curcumin-induced browning in white adipocytes was investigated by determining expression levels of brown adipocyte-specific genes/proteins by real-time reverse transcriptase polymerase chain reaction, immunoblot analysis and immunocytochemical staining. Curcumin increased mitochondrial biogenesis, as evidenced by transmission electronic microscopic detection and enhanced expression of proteins involved in fat oxidation. Cucurmin also increased protein levels of hormone-sensitive lipase and p-acyl-CoA carboxylase, suggesting its possible role in augmentation of lipolysis and suppression of lipogenesis. Increased expression of UCP1 and other brown adipocyte-specific markers was possibly mediated by curcumin-induced activation of AMP-activated protein kinase (AMPK) based on the fact that inhibition of AMPK by dorsomorphin abolished expression of PRDM16, UCP1 and peroxisome proliferator-activated receptor gamma co-activator 1-alpha while the activator 5-Aminoimidazole-4-carboxamide ribonucleotide elevated expression of these brown marker proteins. Our findings suggest that curcumin plays a dual modulatory role in inhibition of adipogenesis as well as induction of the brown fat-like phenotype and thus may have potential therapeutic implications for treatment of obesity.",
"The method of dispersion correction as an add-on to standard Kohn-Sham density functional theory (DFT-D) has been refined regarding higher accuracy, broader range of applicability, and less empiricism. The main new ingredients are atom-pairwise specific dispersion coefficients and cutoff radii that are both computed from first principles. The coefficients for new eighth-order dispersion terms are computed using established recursion relations. System (geometry) dependent information is used for the first time in a DFT-D type approach by employing the new concept of fractional coordination numbers (CN). They are used to interpolate between dispersion coefficients of atoms in different chemical environments. The method only requires adjustment of two global parameters for each density functional, is asymptotically exact for a gas of weakly interacting neutral atoms, and easily allows the computation of atomic forces. Three-body nonadditivity terms are considered. The method has been assessed on standard benchmark sets for inter- and intramolecular noncovalent interactions with a particular emphasis on a consistent description of light and heavy element systems. The mean absolute deviations for the S22 benchmark set of noncovalent interactions for 11 standard density functionals decrease by 15%-40% compared to the previous (already accurate) DFT-D version. Spectacular improvements are found for a tripeptide-folding model and all tested metallic systems. The rectification of the long-range behavior and the use of more accurate C(6) coefficients also lead to a much better description of large (infinite) systems as shown for graphene sheets and the adsorption of benzene on an Ag(111) surface. For graphene it is found that the inclusion of three-body terms substantially (by about 10%) weakens the interlayer binding. We propose the revised DFT-D method as a general tool for the computation of the dispersion energy in molecules and solids of any kind with DFT and related (low-cost) electronic structure methods for large systems.",
"Lung carcinogenesis is a complex process in an unregulated inflammatory environment. Curcumin has been extensively investigated as a multi-target anti-tumor and anti-inflammation compound. In this paper, we demonstrate a novel inflammation-related mechanism for curcumin-induced inhibition of lung tumor growth. We found that neutrophil elastase, an important regulator of inflammatory processes, directly triggered tumor cell proliferation in human lung adenocarcinoma A549 cells, and curcumin could completely suppress the excess tumor proliferation induced by neutrophil elastase. α1-antitrypsin is synthesized by tumor cells and is the natural inhibitor of neutrophil elastase. We found that curcumin counteracted the decrease of α1-antitrypsin induced by neutrophil elastase by inducing the promoter activity of α1-antitrypsin and promoting its expression in A549 cells. The inhibition of neutrophil elastase-induced proliferation by curcumin was dependent on the PI3K/Akt pathway. Knockdown of α1-antitrypsin by siRNA further enhanced the tumor cell proliferation induced by neutrophil elastase and significantly blocked the anti-proliferation effect of curcumin against neutrophil elastase. Curcumin remarkably inhibited the primary tumor growth of Lewis lung carcinoma (LLC) in C57BL/6 mice. We further showed that curcumin upregulated the level of α1-antitrypsin in primary tumor tissue by promoting its local expression, and the protein level of neutrophil elastase in tumor tissue was obviously decreased in mice treated with curcumin. Overall, our results suggest that neutrophil elastase and α1-antitrypsin play important roles in modulating lung tumor proliferation in inflammatory microenvironment and curcumin inhibits neutrophil elastase-induced tumor proliferation via upregulating α1-antitrypsin expression in vitro and in vivo."
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Toxic freshwater benthic cyanobacteria: ecology, toxin production, and impacts
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1. This review summarises knowledge on the ecology, toxin production, and impacts of toxic freshwater benthic cyanobacterial proliferations. It documents monitoring, management, and sampling strategies, and explores mitigation options. 2. Toxic proliferations of freshwater benthic cyanobacteria (taxa that grow attached to substrates) occur in streams, rivers, lakes, and thermal and meltwater ponds, and have been reported in 19 countries. Anatoxin- and microcystin-containing mats are most commonly reported (eight and 10 countries, respectively). 3. Studies exploring factors that promote toxic benthic cyanobacterial proliferations are limited to a few species and habitats. There is a hierarchy of importance in environmental and biological factors that regulate proliferations with variables such as flow (rivers), fine sediment deposition, nutrients, associated microbes, and grazing identified as key drivers. Regulating factors differ among colonisation, expansion, and dispersal phases. 4. New -omics-based approaches are providing novel insights into the physiological attributes of benthic cyanobacteria and the role of associated microorganisms in facilitating their proliferation. 5. Proliferations are commonly comprised of both toxic and non-toxic strains, and the relative proportion of these is the key factor contributing to the overall toxin content of each mat. 6. While these events are becoming more commonly reported globally, we currently lack standardised approaches to detect, monitor, and manage this emerging health issue. To solve these critical gaps, global collaborations are needed to facilitate the rapid transfer of knowledge and promote the development of standardised techniques that can be applied to diverse habitats and species, and ultimately lead to improved management.
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"Cyanobacterial microcystins (MCs), potent serine/threonine-phosphatase inhibitors, pose an increasing threat to humans. Current detection methods are optimised for water matrices with only a few MC congeners simultaneously detected. However, as MC congeners are known to differ in their toxicity, methods are needed that simultaneously quantify the congeners present, thus allowing for summary hazard and risk assessment. Moreover, detection of MCs should be expanded to complex matrices, e.g., blood and tissue samples, to verify in situ MC concentrations, thus providing for improved exposure assessment and hazard interpretation. To achieve this, we applied two synthetic deuterated MC standards and optimised the tissue extraction protocol for the simultaneous detection of 14 MC congeners in a single ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) run. This procedure was validated using plasma and liver homogenates of mice (male and female) spiked with deuterated MC standards. For proof of concept, tissue and plasma samples from mice i.p. injected with MC-LR and MC-LF were analysed. While MC-LF was detected in all tissue samples of both sexes, detection of MC-LR was restricted to liver samples of male mice, suggesting different toxicokinetics in males, e.g., transport, conjugation or protein binding. Thus, deconjugation/-proteinisation steps should be employed to improve detection of bound MC.",
"Nostoc is a cyanobacterial genus, common in soils and a prolific producer of natural products. This research project aimed to explore and characterize Brazilian cyanobacteria for new bioactive compounds. Here we report the production of hepatotoxins and new protease inhibitors from benthic Nostoc sp. CENA543 isolated from a small, shallow, saline-alkaline lake in the Nhecolândia, Pantanal wetland area in Brazil. Nostoc sp. CENA543 produces exceptionally high amounts of nodularin-R. This is the first free-living Nostoc that produces nodularin at comparable levels as the toxic, bloom-forming, Nodularia spumigena. We also characterized pseudospumigins A-F, which are a novel family of linear tetrapeptides. Pseudospumigins are structurally related to linear tetrapeptide spumigins and aeruginosins both present in N. spumigena but differ in respect to their diagnostic amino acid, which is Ile/Leu/Val in pseudospumigins, Pro/mPro in spumigins, and Choi in aeruginosins. The pseudospumigin gene cluster is more similar to the spumigin biosynthetic gene cluster than the aeruginosin gene cluster. Pseudospumigin A inhibited trypsin (IC50 4.5 μM after 1 h) in a similar manner as spumigin E from N. spumigena but was almost two orders of magnitude less potent. This study identifies another location and environment where the hepatotoxic nodularin has the potential to cause the death of eukaryotic organisms.",
"The first identification of anatoxin-a in a French lotic system is reported. Rapid deaths of dogs occurred in 2003 after the animals drank water from the shoreline of the La Loue River in eastern France. Sediments, stones and macrophytes surfaces at the margin of the river were covered by a thick biofilm containing large quantities of several benthic species of filamentous, non-heterocystous cyanobacteria. Known cyanotoxins, such as microcystins, saxitoxins and anatoxins were screened from biofilm samples by biochemical and analytical assays. A compound with similar UV spectra to the anatoxin-a standard was detected by high-performance liquid chromatography (HPLC) coupled with photo-diode array detector. This toxin was further identified by HPLC coupled with a UV detector and by electrospray ionisation-Quadrupole-Time-Of-Flight mass spectrometer, and confirmed by tandem mass spectrometry. These two techniques were necessary to discriminate anatoxin-a in phenylalanine-containing matrices such as liver samples of poisoned dogs. The toxin and the aromatic amino acid, phenylalanine, present the same pseudomolecular ion at m/z 166, but have differing fragmentation patterns, retention times and UV spectra. Finally, several cyanobacterial strains were isolated from the green biofilm and tested for anatoxin-a production. Phormidium favosum was identified as a new anatoxin-a producing species.",
"Saxitoxin and its analogs cause the paralytic shellfish-poisoning syndrome, adversely affecting human health and coastal shellfish industries worldwide. Here we report the isolation, sequencing, annotation, and predicted pathway of the saxitoxin biosynthetic gene cluster in the cyanobacterium Lyngbya wollei. The gene cluster spans 36 kb and encodes enzymes for the biosynthesis and export of the toxins. The Lyngbya wollei saxitoxin gene cluster differs from previously identified saxitoxin clusters as it contains genes that are unique to this cluster, whereby the carbamoyltransferase is truncated and replaced by an acyltransferase, explaining the unique toxin profile presented by Lyngbya wollei. These findings will enable the creation of toxin probes, for water monitoring purposes, as well as proof-of-concept for the combinatorial biosynthesis of these natural occurring alkaloids for the production of novel, biologically active compounds.",
"In November 2005, at least five dogs died rapidly after contact with water from the Hutt River (lower North Island, New Zealand). Necropsy performed 24h later on one of the dogs (a 20-month-old Labrador) revealed few findings of interest, except for copious amounts of froth in the respiratory tract down to the bifurcation of the trachea and large quantities of algal material in the dog's stomach. Low and relatively stable flows in the Hutt River during spring had resulted in the proliferation of benthic cyanobacteria that formed large black/brown mats along the river edge. Samples from the Labrador's stomach contents and cyanobacterial mats were analysed microscopically and screened using chemical and biochemical assays for cyanotoxins: anatoxin-a, homoanatoxin-a, cylindrospermopsins, saxitoxins and microcystins. Liquid chromatography-mass spectrometry (LC-MS) confirmed the presence of the neurotoxic cyanotoxins anatoxin-a and homoanatoxin-a and their degradation products, dihydro-anatoxin-a and dihydro-homoanatoxin-a. This is the first report of homoanatoxin-a and associated degradation product in New Zealand. Based on morphology, the causative species was identified as Phormidium sp. Subsequent phylogenetic analysis of 16S rRNA gene sequences demonstrated that the causative organism was most similar to Phormidium autumnale. Further investigations led to the detection of homoanatoxin-a and anatoxin-a in cyanobacterial mats from four other rivers in the Wellington region (lower North Island, New Zealand). Access restrictions were placed on over 60% of river catchments in the western Wellington region, severely affecting recreational users.",
"This study has been performed in the framework of a research program aiming to develop a low-cost aerial sensor for the monitoring of cyanobacteria in freshwater ecosystems that could be used for early detection. Several empirical and mechanistic remote-sensing tools have been already developed and tested at large scales and have proven useful in monitoring cyanobacterial blooms. However, the effectiveness of these tools for early detection is hard to assess because such work requires the detection of low concentrations of characteristic pigments amid complex ecosystems exhibiting several confounding factors (turbidity, blooms of other species, etc.). We developed a framework for performing high-throughput measurements of the absorbance and reflectance of small volumes (∼ = 20 mL) of controlled mixtures of phytoplankton species and studied the potential of this framework to validate remote-sensing proxies of cyanobacteria concentration. The absorption and reflectance spectra of single and multiple cultures carried a specific signal that allowed for the quantitative analysis of culture mixes. This specific signal was shown to be related to known pigment absorbance spectra. The concentrations of chlorophyll-a and -b, phycocyanin and phycoerythrin could be obtained from direct absorbance measurements and were correlated with the concentration obtained after pigment extraction (R2 ≥ 0.96 for all pigments). A systematic test of every possible two-band and three-band normalized difference between optical indices was then performed, and the coincidental correlation with chlorophyll-b (absent in cyanobacteria) was used as an indicator of non-specificity. Two-band indices were shown to suffer from non-specificity issues and could not yield strong and specific relationships with phycocyanin or phycoerythrin (maximum R2 < 0.5). On the other hand, the three-band modified normalized difference indices yielded strong specific relationships (R2 > 0.8).",
"Matrix-dependent signal suppression or enhancement represents a major drawback in quantitative analysis with liquid chromatography coupled to atmospheric pressure ionization mass spectrometry (LC-API-MS). Because matrix effects (ME) might exert a detrimental impact on important method parameters (limit of detection, limit of quantification, linearity, accuracy, and precision), they have to be tested and evaluated during validation procedure. This review gives a detailed description on when these phenomena might be expected, and how they can be evaluated. The major sources of ME are discussed and illustrated with examples from bioanalytical, pharmaceutical, environmental, and food analysis. Because there is no universal solution for ME, the main strategies to overcome these phenomena are described in detail. Special emphasis is devoted to the sample-preparation procedures as well as to the recent improvements on chromatographic and mass spectrometric conditions. An overview of the main calibration techniques to compensate for ME is also presented. All these solutions can be used alone or in combination to retrieve the performance of the LC-MS for a particular matrix-analyte combination."
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Gamma entrainment therapy in Alzheimer's disease.
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Despite decades of research, Alzheimer's Disease (AD) remains a lethal neurodegenerative disorder for which there are no effective treatments. This review examines the latest evidence of a novel and newly introduced perspective, which focuses on the restoration of gamma oscillations and investigates their potential role in the treatment of AD. Gamma brain activity (∼25-100 Hz) has been well-known for its role in cognitive function, including memory, and it is fundamental for healthy brain activity and intra-brain communication. Aberrant gamma oscillations have been observed in both mice AD models and human AD patients. A recent line of work demonstrated that gamma entrainment, through auditory and visual sensory stimulation, can effectively attenuate AD pathology and improve cognitive function in mice models of the disease. The first evidence from AD patients indicate that gamma entrainment therapy can reduce loss of functional connectivity and brain atrophy, improve cognitive function, and ameliorate several pathological markers of the disease. Even though research is still in its infancy, evidence suggests that gamma-based therapy may have a disease-modifying effect and has signified a new and promising era in AD research.
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[
"The sensory evoked neuromagnetic response consists of superimposition of an immediately stimulus-driven component and induced changes in the autonomous brain activity, each having distinct functional relevance. Commonly, the strength of phase locking in neural activities has been used to differentiate the different responses. The steady-state response is a strong oscillatory neural activity, which is evoked with rhythmic stimulation, and provides an effective tool to investigate oscillatory brain networks. In this case, both the sensory response and intrinsic activity, representing higher order processes, are highly synchronized to the stimulus. In this study we hypothesized that temporal dynamics of oscillatory activities would characterize the differences between the two types of activities and that beta and gamma oscillations are differently involved in this distinction. We used magnetoencephalography (MEG) for studying how ongoing steady-state responses elicited by a 20-Hz vibro-tactile stimulus to the right index finger were affected by a concurrent isolated touch stimulus to the same hand ring finger. SI source activity showed oscillations at multiples of 20 Hz with characteristic differences in the beta band and the gamma band. The response amplitudes were largest at 20 Hz (beta) and significantly reduced at 40 Hz and 60 Hz (gamma), although synchronization strength, indicated by inter-trial coherence (ITC), did not substantially differ between 20 Hz and 40 Hz. Moreover, the beta oscillations showed a fast onset, whereas the amplitude of gamma oscillations increased slowly and reached the steady state 400 ms after onset of the vibration stimulus. Most importantly, the pulse stimuli interacted only with gamma oscillations in a way that gamma oscillations decreased immediately after the concurrent stimulus onset and recovered slowly, resembling the initial slope. Such time course of gamma oscillations is similar to our previous observations in the auditory system. The time constant is in line with the time required for conscious perception of the sensory stimulus. Based on the observed different spectro-temporal dynamics, we propose that while beta activities likely relate to independent representation of the sensory input, gamma oscillation likely relates to binding of sensory information for higher order processing.",
"Acutely developing lesions of the brain have been highly instructive in elucidating the neural systems underlying memory in humans and animal models. Much less has been learned from chronic neurodegenerative disorders that insidiously impair memory. But the advent of a detailed molecular hypothesis for the development of Alzheimer's disease and the creation of compelling mouse models thereof have begun to change this situation. Experiments in rodents suggest that soluble oligomers of the amyloid beta protein (Abeta) may discretely interfere with synaptic mechanisms mediating aspects of learning and memory, including long-term potentiation. In humans, memory impairment correlates strongly with cortical levels of soluble Abeta species, which include oligomers. Local inflammatory changes, neurofibrillary degeneration, and neurotransmitter deficits all contribute to memory impairment, but available evidence suggests that these develop as a consequence of early Abeta accumulation. Accordingly, attempts to slow memory and cognitive loss by decreasing cerebral Abeta levels have entered human trials.",
"Statistical interdependencies between magnetoencephalographic signals recorded over different brain regions may reflect the functional connectivity of the resting-state networks. We investigated topographic characteristics of disturbed resting-state networks in Alzheimer's disease patients in different frequency bands. Whole-head 151-channel MEG was recorded in 18 Alzheimer patients (mean age 72.1 years, SD 5.6; 11 males) and 18 healthy controls (mean age 69.1 years, SD 6.8; 7 males) during a no-task eyes-closed resting state. Pair-wise interdependencies of MEG signals were computed in six frequency bands (delta, theta, alpha1, alpha2, beta and gamma) with the synchronization likelihood (a nonlinear measure) and coherence and grouped into long distance (intra- and interhemispheric) and short distance interactions. In the alpha1 and beta band, Alzheimer patients showed a loss of long distance intrahemispheric interactions, with a focus on left fronto-temporal/parietal connections. Functional connectivity was increased in Alzheimer patients locally in the theta band (centro-parietal regions) and the beta and gamma band (occipito-parietal regions). In the Alzheimer group, positive correlations were found between alpha1, alpha2 and beta band synchronization likelihood and MMSE score. Resting-state functional connectivity in Alzheimer's disease is characterized by specific changes of long and short distance interactions in the theta, alpha1, beta and gamma bands. These changes may reflect loss of anatomical connections and/or reduced central cholinergic activity and could underlie part of the cognitive impairment.",
"High frequency (30-70 Hz) gamma band oscillations in the human electro-encephalogram (EEG) are thought to reflect perceptual and cognitive processes. It is therefore interesting to study these measures in cognitive impairment and dementia. To evaluate gamma band oscillations as a diagnostic biomarker in Alzheimer's disease (AD) and mild cognitive impairment (MCI), 15 psychoactive drug naïve AD patients, 20 MCI patients and 20 healthy controls participated in this study. Gamma band power (GBP) was measured in four conditions viz. resting state, music listening, story listening and visual stimulation. To evaluate test-retest reliability (TRR), subjects underwent a similar assessment one week after the first. The overall TRR was high. Elevated GBP was observed in AD when compared to MCI and control subjects in all conditions. The results suggest that elevated GBP is a reproducible and sensitive measure for cognitive dysfunction in AD in comparison with MCI and controls.",
"The Alzheimer's Disease Assessment Scale (ADAS) is often used in international multicenter trials. Use across countries presupposes correct translation and adaptation of the scale, and maintenance of its psychometric properties. To compare the various translations of the ADAS used in Western Europe, to design internationally harmonized translations and to validate these. International cooperative study in eight European countries. An inventory was made of existing versions of the ADAS-Cog used in eight European countries, and adaptations were made. The concurrent validity of the harmonized versions of the ADAS was tested in 283 patients with probable or possible Alzheimer's disease. The Nurses Observation Scale for Geriatrics (NOSGER), CAMCOG-R and MMSE was used to assess concordance between cognitive and behavioral measures. Differences between the versions mainly involved object naming, items for verbal memory, such as the number of trials allowed, the imagery value of the words selected as targets or distractors, and the number of parallel versions. These differences were eliminated by adapting and harmonizing the various versions of the ADAS-Cog. Thereafter, only small differences between the different countries were found, and patterns of correlation between ADAS-Cog, and the NOSGER, CAMCOG-R and MMSE were consistent. The study underlines the need to use harmonized versions of instruments for rating dementia in multinational studies. The findings indicate that the harmonization of the ADAS-Cog was successful.",
"We experience objects as whole, complete entities irrespective of whether they are perceived by our sensory systems or are recalled from memory. However, it is also known that many of the properties of objects are encoded and processed in different areas of the brain. How then, do coherent representations emerge? One theory suggests that rhythmic synchronization of neural discharges in the gamma band (around 40 Hz) may provide the necessary spatial and temporal links that bind together the processing in different brain areas to build a coherent percept. In this article we propose that this mechanism could also be used more generally for the construction of object representations that are driven by sensory input or internal, top-down processes. The review will focus on the literature on gamma oscillatory activities in humans and will describe the different types of gamma responses and how to analyze them. Converging evidence that suggests that one particular type of gamma activity (induced gamma activity) is observed during the construction of an object representation will be discussed.",
"Factors related to physical health have been implicated in both normal and pathological aging of cognitive abilities. To substantiate this notion, we studied existing morbidity, as diagnosed by the general practitioner according to well-defined criteria, as a potential predictor of cognitive test performance. A sample of 1360 individuals, aged 24-81 years and living in the community, was stratified for age, sex, and general ability. Active and total morbidity in this group were classified according to the International Classification of Primary Care. Neurocognitive tests were used to assess the domains of verbal memory, sensorimotor speed, and cognitive flexibility. Multiple regression analyses with adjustment for age, sex, and educational level showed both insulin-dependent and noninsulin-dependent diabetes to be negatively associated with all cognitive measures. More specific negative associations were found for chronic bronchitis (performance speed) and presbyacusia (memory). Single or aggregated cardiovascular morbidity (including hypertension) was unrelated to test performance. Existing morbidity as a whole contributes only modestly (up to 3.5%) to total variance in cognitive function. However, some specific, relatively common diseases of the elderly, such as diabetes and chronic bronchitis, may aggravate the age-related decline in cognitive ability."
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Emotional regulation and the quality of romantic relationships
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In romantic relationships, individual differences are determinant factors for relational quality. Specifically, romantic attachment (RA) and difficulties in emotional regulation influence each other and may have predictive potential for the perceived dyadic adjustment (DA) level. This paper aims to identify the developmental parallel between behavioral patterns built since childhood and the construction of the emotional regulation skills that characterize them. Our analysis was based on the attachment theory and the concepts of romantic relationship and DA. In this way, we sought to further the understanding of relationship dynamics, beyond the usual focus on a single element and on associative relationships, and by exploring other effects among the different dimensions of relational functioning. In particular, we explored the predictive ability of emotional regulation patterns (more flexible individual characteristics) in discriminating between RA styles (more perennial influences), and their impact on the quality of romantic relationships, in the anticipation of dyadic adjustment variations.
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[
"This paper reviews the research on relationships and mental health. Individuals who are more mentally healthy are more likely to select into relationships, but relationships are also demonstrably associated with mental health. The type of relationship matters - evidence suggests that more established, committed relationships, such as marriage, are associated with greater benefits than less committed unions such as cohabitation. The association between relationships and mental health is clearly bidirectional, however, stronger effects are observed when mental health is the outcome and relationships are the predictor, suggesting that the causal arrow flows more strongly from relationships to mental health than vice versa. Moreover, improving relationships improves mental health, but improving mental health does not reliably improve relationships. Our review of research corroborates the view that relationships are a keystone component of human functioning that have the potential to influence a broad array of mental health outcomes.",
"This research tested whether adult attachment orientations predict use of emotion regulation strategies in theoretically consistent ways, and whether associations among attachment orientations and emotion regulatory strategies are moderated by critical features of the relationship context. Ninety-six couples (192 individuals) reported on their attachment orientations, habitual use of emotion regulation strategies (cognitive reappraisal, expressive suppression, negative emotion expressivity), and perceptions of relationship closeness and negative partner behaviors. Highly secure individuals reported greater use of cognitive reappraisal, especially when they felt closer to their partners, and engaged in less suppression when their partners behaved more negatively toward them. Highly avoidant individuals reported greater use of suppression, especially when they perceived more negative partner behaviors, and when their partners were more avoidant. Highly anxious individuals also used more suppression when their partners were more avoidant, but they expressed more negative emotions when they were paired with less avoidant partners. Fearful-avoidant individuals' emotion regulation patterns resembled those of both highly secure and dismissive-avoidant individuals. This study illustrates how attending to moderating effects within specific relationships and testing joint effects of both partners' personality characteristics can help identify contextual boundaries of emotion regulation strategies and clarify emotional response patterns in couples.",
"Emotions are powerful determinants of behaviour, thought and experience, and they may be regulated in various ways. Neuroimaging studies have implicated several brain regions in emotion regulation, including the ventral anterior cingulate and ventromedial prefrontal cortices, as well as the lateral prefrontal and parietal cortices. Drawing on computational approaches to value-based decision-making and reinforcement learning, we propose a unifying conceptual framework for understanding the neural bases of diverse forms of emotion regulation.",
"Romantic relationship, a widespread feature of human society, is one of the most influential factors in daily life. Although stimuli related to romantic love or being in a romantic relationship commonly result in enhancement of activation or functional connectivity of the reward system, including the striatum, the structure underlying romantic relationship-related regions remain unclear. Because individual experiences can alter gray matter within the adult human brain, we hypothesized that romantic relationship is associated with structural differences in the striatum related to the positive subjective experience of being in a romantic relationship. Because intimate romantic relationships contribute to perceived subjective happiness, this subjective enhancement of happiness might be accompanied by the experience of positive events related to being in a romantic relationship. To test this hypothesis and elucidate the structure involved, we compared subjective happiness, an indirect measure of the existence of positive experiences caused by being in a romantic relationship, of participants with or without romantic partners (N = 68). Furthermore, we also conducted a voxel-based morphometry study of the effects of being in a romantic relationship (N = 113). Being in a romantic relationship was associated with greater subjective happiness and reduced gray matter density within the right dorsal striatum. These results suggest that being in a romantic relationship enhances perceived subjective happiness via positive experiences. Furthermore, the observed reduction in gray matter density in the right dorsal striatum may reflect an increase in saliency of social reward within a romantic relationship. Thus, being in a romantic relationship is associated with positive experiences and a reduction of gray matter density in the right dorsal striatum, representing a modulation of social reward.",
"This study examined links between two distinct facets of empathy-empathic accuracy and perceived empathic effort-and one's own and one's partner's relationship satisfaction. Using a video recall procedure, participants (n = 156 couples in committed relationships) reported on their own emotions and their perceptions of partners' emotions and partners' empathic intentions during moments of high affect in laboratory-based discussions of upsetting events. Partners' data were correlated as a measure of how accurately they were able to read what the other was feeling and to what degree they felt the other was trying to be empathic at those moments. The perception of empathic effort by one's partner was more strongly linked with both men's and women's relationship satisfaction than empathic accuracy. Men's relationship satisfaction was related to the ability to read their partners' positive emotions accurately, whereas women's relationship satisfaction was related to their partners' ability to read women's negative emotions accurately. Women's ability to read their husbands' negative emotions was positively linked to both men's and women's relationship satisfaction. Findings suggest that the perception of a partner's empathic effort-as distinct from empathic accuracy-is uniquely informative in understanding how partners may derive relationship satisfaction from empathic processes. When working with couples in treatment, heightening partners' perceptions of each other's empathic effort, and helping partners learn to demonstrate effort, may represent particularly powerful opportunities for improving satisfaction in relationships.",
"This study examined the indirect effects of relationship status (single vs. in a relationship) on life satisfaction through social and emotional (romantic and family) loneliness and perceived social support from significant others, family, and friends. Five hundred and fifty three Polish young adults (335 females and 218 males), ranging in age from 20-30 years (M = 23.42), completed the Polish versions of the Satisfaction With Life Scale, the Social and Emotional Loneliness Scale for Adults, and the Multidimensional Scale of Perceived Social Support. The results indicated that single individuals reported significantly lower satisfaction with life and social support from a significant other, but higher romantic and social loneliness, and higher family support compared to participants in a relationship. A path analysis revealed no direct effect of relationship status on satisfaction with life. However, there were significant indirect effects from relationship status to life satisfaction though romantic, family, and social loneliness, and through perceived social support from significant others and from family. Therefore, singlehood may be deleterious to life satisfaction because of the higher loneliness and lower social support from a significant other.",
"The convergent and discriminant validities of well-being concepts were examined using multitrait-multimethod matrix analyses (D. T. Campbell & D. W. Fiske, 1959) on 3 sets of data. In Study 1, participants completed measures of life satisfaction, positive affect, negative affect, self-esteem, and optimism on 2 occasions 4 weeks apart and also obtained 3 informant ratings. In Study 2, participants completed each of the 5 measures on 2 occasions 2 years apart and collected informant reports at Time 2. In Study 3, participants completed 2 different scales for each of the 5 constructs. Analyses showed that (a) life satisfaction is discriminable from positive and negative affect, (b) positive affect is discriminable from negative affect, (c) life satisfaction is discriminable from optimism and self-esteem, and (d) optimism is separable from trait measures of negative affect."
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Molecular genetics of phototropin and phytochrome
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Changes in environmental conditions like temperature and light critically influence crop production. To deal with these changes, plants possess various photoreceptors such as Phototropin (PHOT), Phytochrome (PHY), Cryptochrome (CRY), and UVR8 that work synergistically as sensor and stress sensing receptors to different external cues. PHOTs are capable of regulating several functions like growth and development, chloroplast relocation, thermomorphogenesis, metabolite accumulation, stomatal opening, and phototropism in plants. PHOT plays a pivotal role in overcoming the damage caused by excess light and other environmental stresses (heat, cold, and salinity) and biotic stress. The crosstalk between photoreceptors and phytohormones contributes to plant growth, seed germination, photo-protection, flowering, phototropism, and stomatal opening. Molecular genetic studies using gene targeting and synthetic biology approaches have revealed the potential role of different photoreceptor genes in the manipulation of various beneficial agronomic traits. Overexpression of PHOT2 in
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[
"Heading date is a key trait in rice domestication and adaption, and a number of quantitative trait loci (QTLs) have been identified. The rice (Oryza sativa L.) cultivars in the Heilongjiang Province, the northernmost region of China, have to flower extremely early to fulfill their life cycle. However, the critical genes or different gene combinations controlling early flowering in this region have not been determined. QTL and candidate gene analysis revealed that Hd2/Ghd7.1/OsPRR37 plays a major role in controlling rice distribution in Heilongjiang. Further association analysis with a collection of rice cultivars demonstrated that another three major QTL genes (Hd4/Ghd7, Hd5/DTH8/Ghd8, and Hd1) also participate in regulating heading date under natural long day (LD) conditions. Hd2/Ghd7.1/OsPRR37 and Hd4/Ghd7 are two major QTLs and function additively. With the northward rice cultivation, the Hd2/Ghd7.1/OsPRR37 and Hd4/Ghd7 haplotypes became non-functional alleles. Hd1 might be non-functional in most Heilongjiang rice varieties, implying that recessive hd1 were selected during local rice breeding. Non-functional Hd5/DTH8/Ghd8 is very rare, but constitutes a potential target for breeding extremely early flowering cultivars. Our results indicated that diverse genetic combinations of Hd1, Hd2, Hd4, and Hd5 determined the different distribution of rice varieties in this northernmost province of China.",
"Mutations in a component of phytochrome A (phyA)-specific light signal transduction, SPA1, result in enhanced responsiveness of Arabidopsis seedlings to red and far-red light. Here, we have examined the effects of spa1 mutations on the two known modes of phyA function, the high-irradiance responses (HIRs) to continuous irradiation with far-red light and the very-low-fluence responses (VLFRs) to inductive pulses of light that establish only a small proportion of active phyA. spa1 mutants exhibited an enhanced VLFR under hourly pulses of far-red light for hypocotyl growth inhibition, cotyledon unfolding, anthocyanin accumulation, block of greening in subsequent white light and negative regulation of phyB signaling. We provide evidence that the phenotype of spa1 mutants in red light is also caused by an increase in the VLFR. Taken together, our results indicate that light-induced hypocotyl growth inhibition in spa1 mutants is primarily due to a VLFR. While wild-type seedlings required hourly pulses of far-red light to induce a VLFR, infrequent irradiation with far-red pulses (every 12 h) was sufficient to induce a strong VLFR of hypocotyl elongation in spa1 mutants. This shows that the effect of the VLFR was more persistent in spa1 mutants than in the wild type. We, therefore, propose that SPA1 has an important function in reducing the persistence of phyA signaling. spa1 mutations also enhanced the HIRs of anthocyanin accumulation and of phyA-mediated responsivity amplification towards phyB. Thus, our results suggest that spa1 mutations amplify both the phyA-mediated VLFR and the HIR.",
"Light and temperature are major environmental factors that coordinately control plant growth and survival. However, how plants integrate light and temperature signals to better adapt to environmental stresses is poorly understood. PHYTOCHROME-INTERACTING FACTOR 3 (PIF3), a key transcription factor repressing photomorphogenesis, has been shown to play a pivotal role in mediating plants' responses to various environmental signals. In this study, we found that PIF3 functions as a negative regulator of Arabidopsis freezing tolerance by directly binding to the promoters of C-REPEAT BINDING FACTOR (CBF) genes to down-regulate their expression. In addition, two F-box proteins, EIN3-BINDING F-BOX 1 (EBF1) and EBF2, directly target PIF3 for 26S proteasome-mediated degradation. Consistently, ebf1 and ebf2 mutants were more sensitive to freezing than were the wild type, and the pif3 mutation suppressed the freezing-sensitive phenotype of ebf1 Furthermore, cold treatment promoted the degradation of EBF1 and EBF2, leading to increased stability of the PIF3 protein and reduced expression of the CBF genes. Together, our study uncovers an important role of PIF3 in Arabidopsis freezing tolerance by negatively regulating the expression of genes in the CBF pathway.",
"We investigated the effect of supplemental LED inter-lighting (80% red, 20% blue; 70 W m-2; light period 04:00-22:00) on the productivity and physiological traits of tomato plants (Flavance F1) grown in an industrial greenhouse with high pressure sodium (HPS) lamps (235 W m-2, 420 µmol m-2 s-1 at canopy). Physiological trait measurements included diurnal photosynthesis and fruit relative growth rates, fruit weight at specific positions in the truss, root pressure, xylem sap hormone and ion compositions, and fruit quality. In the control treatment with HPS lamps alone, the ratio of far-red to red light (FR:R) was 1.2 at the top of the canopy and increased to 5.4 at the bottom. The supplemental LED inter-lighting decreased the FR:R ratio at the middle and low positions in the canopy and was associated with greener leaves and higher photosynthetic light use efficiency (PLUE) in the leaves in the lower canopy. The use of LED inter-lighting increased the biomass and yield by increasing the fruit weight and enhancing plant growth. The PLUE of plants receiving supplemental LED light decreased at the end of the light period, indicating that photosynthesis of the supplemented plants at the end of the day might be limited by sink capacity. The supplemental LED lighting increased the size of fruits in the middle and distal positions of the truss, resulting in a more even size for each fruit in the truss. Diurnal analysis of fruit growth showed that fruits grew more quickly during the night on the plants receiving LED light than on unsupplemented control plants. This faster fruit growth during the night was related to an increased root pressure. The LED treatment also increased the xylem levels of the phytohormone jasmonate. Supplemental LED inter-lighting increased tomato fruit weight without affecting the total soluble solid contents in fruits by increasing the total assimilates available for fruit growth and by enhancing root activity through an increase in root pressure and water supply to support fruit growth during the night.",
"Gene and genome duplications underlie the origins of evolutionary novelty in plants. Soybean, Glycine max, is considered to be a paleopolyploid species with a complex genome. We found multiple homologs of the phytochrome A gene (phyA) in the soybean genome and determined the DNA sequences of two paralogs designated GmphyA1 and GmphyA2. Analysis of the GmphyA2 gene from the lines carrying a recessive allele at a photoperiod insensitivity locus, E4, revealed that a Ty1/copia-like retrotransposon was inserted in exon 1 of the gene, which resulted in dysfunction of the gene. Mapping studies suggested that GmphyA2 is encoded by E4. The GmphyA1 gene was mapped to a region of linkage group O, which is homeologous to the region harboring E4 in linkage group I. Plants homozygous for the e4 allele were etiolated under continuous far red light, but the de-etiolation occurred partially, indicating that the mutation alone did not cause a complete loss of phyA function. The genetic redundancy suggests that the presence of duplicated copies of phyA genes accounts for the generation of photoperiod insensitivity, while protecting against the deleterious effects of mutation. Thus, this phenomenon provides a link between gene duplication and establishment of an adaptive response of plants to environments.",
"The blue light receptor NPH1 (for nonphototropic hypocotyl) has been considered to be the only UV-A/blue light receptor that induces a phototropic response by the hypocotyl and root of Arabidopsis. By analysis of root phototropism (rpt) mutants, we show, however, the involvement of another blue light receptor as well as the existence of two separate signaling pathways working downstream of these receptors in the phototropic response. A newly isolated gene, RPT2, controls one of these pathways. The RPT2 gene is light inducible; encodes a novel protein with putative phosphorylation sites, a nuclear localization signal, a BTB/POZ domain, and a coiled-coil domain; and belongs to a large gene family that includes the recently isolated NPH3 gene. From genetic, physiological, and biochemical evidence, we propose a genetic model of the signaling pathways that induce the phototropic response in Arabidopsis.",
"Light is the environmental factor that most affects plant growth and development through its impact on photomorphogenesis and photosynthesis. A quadruple photoreceptor mutant lacking four of the most important photoreceptors in plants, phytochromes A and B (phyA, phyB) and cryptochromes 1 and 2 (cry1, cry2), is severely affected in terms of growth and development. Previous studies have suggested that in addition to a photomorphogenic disorder, the phyA phyB cry1 cry2 quadruple mutant might have severe alterations in photosynthetic ability. Here, we investigated the photosynthetic processes altered in the quadruple mutant and performed a proteomic profiling approach to identify some of the proteins involved. The phyA phyB cry1 cry2 quadruple mutant showed reduced leaf area and total chlorophyll content. Photosynthetic rates at high irradiances were reduced approximately 65% compared to the wild type (WT). Light-saturated photosynthesis and the response of net CO2 exchange to low and high internal CO2 concentrations suggest that the levels or activity of the components of the Calvin cycle and electron transport might be reduced in the quadruple mutant. Most of the under-expressed proteins in the phyA phyB cry1 cry2 quadruple mutant consistently showed a chloroplastic localization, whereas components of the Calvin cycle and light reaction centers were overrepresented. Additionally, Rubisco expression was reduced threefold in the phyA phyB cry1 cry2 quadruple mutant. Together, these results highlight the importance of the phytochrome and cryptochrome families in proper autotrophy establishment in plants. They also suggest that an overall limitation in the chlorophyll levels, expression of Rubisco, and enzymes of the Calvin Cycle and electron transport that affect ribulose-1,5-biphosphate (RuBP) regeneration reduced photosynthetic capacity in the phyA phyB cry1 cry2 quadruple mutant.",
"Germination of lettuce (Lactuca sativa) 'Grand Rapids' seeds is regulated by phytochrome. The action of phytochrome includes alterations in the levels of gibberellin (GA) and abscisic acid (ABA). To determine the molecular mechanism of phytochrome regulation of ABA metabolism, we isolated four lettuce cDNAs encoding 9-cis-epoxycarotenoid dioxygenase (biosynthesis; LsNCED1-LsNCED4) and four cDNAs for ABA 8'-hydroxylase (catabolism; LsABA8ox1-LsABA8ox4). Measurements of ABA and its catabolites showed that a decrease in ABA level coincided with a slight increase in the level of the ABA catabolite phaseic acid after red light treatment. Quantitative reverse transcription-polymerase chain reaction analysis indicated that ABA levels are controlled by phytochrome through down-regulation of LsNCED2 and LsNCED4 expression and up-regulation of LsABA8ox4 expression in lettuce seeds. Furthermore, the expression levels of LsNCED4 decreased after GA(1) treatment, whereas the levels of expression of the other two genes were unaffected. The LsNCED4 expression was also down-regulated by red light in lettuce seeds in which GA biosynthesis was suppressed by AMO-1618, a specific GA biosynthesis inhibitor. These results indicate that phytochrome regulation of ABA metabolism is mediated by both GA-dependent and -independent mechanisms. Spatial analysis showed that after red light treatment, the ABA decrease on the hypocotyl side was greater than that on the cotyledon side of lettuce seeds. Moreover, phytochrome-regulated expression of ABA and GA biosynthesis genes was observed on the hypocotyl side, rather than the cotyledon side, suggesting that this regulation occurs near the photoperceptive site.",
"B-box containing proteins play an important role in light signaling in plants. Here, we identify LIGHT-REGULATED ZINC FINGER1/SALT TOLERANCE HOMOLOG3 (STH3), a B-box encoding gene that genetically interacts with two key regulators of light signaling, ELONGATED HYPOCOTYL5 (HY5) and CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1). STH3 physically interacts with HY5 in vivo and shows a COP1-dependent localization to nuclear speckles when coexpressed with COP1 in plant cells. A T-DNA insertion mutant, sth3, is hyposensitive to high fluence blue, red, and far-red light and has elongated hypocotyls under short days. Analyses of double mutants between sth3, sth2, and hy5 suggest that they have partially overlapping functions. Interestingly, functional assays in protoplasts suggest that STH3 can activate transcription both independently and together with STH2 through the G-box promoter element. Furthermore, sth3 suppresses the cop1 hypocotyl phenotype in the dark as well as the anthocyanin accumulation in the light. Finally, COP1 ubiquitinates STH3 in vitro, suggesting that STH3 is regulated by COP1. In conclusion, we have identified STH3 as a positive regulator of photomorphogenesis acting in concert with STH2 and HY5, while also being a target of COP1-mediated ubiquitination.",
"It is well known that abscisic acid (ABA) plays a central role in the regulation of seed dormancy and that transcriptional regulation of genes encoding ABA biosynthetic and degradation enzymes is responsible for determining ABA content. However, little is known about the upstream signaling pathways impinging on transcription to ultimately regulate ABA content or how environmental signals (e.g., light and cold) might direct such expression in grains. Our previous studies indicated that light is a key environmental signal inhibiting germination in dormant grains of barley (Hordeum vulgare), wheat (Triticum aestivum), and Brachypodium distachyon and that this effect attenuates as after-ripening progresses further. We found that the blue component of the light spectrum inhibits completion of germination in barley by inducing the expression of the ABA biosynthetic gene 9-cis-epoxycarotenoid dioxygenase and dampening expression of ABA 8'-hydroxylase, thus increasing ABA content in the grain. We have now created barley transgenic lines downregulating the genes encoding the blue light receptors CRYTOCHROME (CRY1) and CRY2. Our results demonstrate that CRY1 is the key receptor perceiving and transducing the blue light signal in dormant grains."
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CRISPR-based epigenome editing in plants and animals
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CRISPR-based epigenome editing uses dCas9 as a platform to recruit transcription or chromatin regulators at chosen loci. Despite recent and ongoing advances, the full potential of these approaches to studying chromatin functions in vivo remains challenging to exploit. In this review we discuss how recent progress in plants and animals provides new routes to investigate the function of chromatin regulators and address the complexity of associated regulations that are often interconnected. While efficient transcriptional engineering methodologies have been developed and can be used as tools to alter the chromatin state of a locus, examples of direct manipulation of chromatin regulators remain scarce in plants. These reports also reveal pitfalls and limitations of epigenome engineering approaches that are nevertheless informative as they are often associated with locus- and context-dependent features, which include DNA accessibility, initial chromatin and transcriptional state or cellular dynamics. Strategies implemented in different organisms to overcome and even take advantage of these limitations are highlighted, which will further improve our ability to establish the causality and hierarchy of chromatin dynamics on genome regulation.
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[
"The RNA-guided endonuclease Cas9 can be converted into a programmable transcriptional repressor, but inefficiencies in target-gene silencing have limited its utility. Here we describe an improved Cas9 repressor based on the C-terminal fusion of a rationally designed bipartite repressor domain, KRAB-MeCP2, to nuclease-dead Cas9. We demonstrate the system's superiority in silencing coding and noncoding genes, simultaneously repressing a series of target genes, improving the results of single and dual guide RNA library screens, and enabling new architectures of synthetic genetic circuits.",
"High-throughput genetic screens have become essential tools for studying a wide variety of biological processes. Here we experimentally compare systems based on clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) or its transcriptionally repressive variant, CRISPR-interference (CRISPRi), with a traditional short hairpin RNA (shRNA)-based system for performing lethality screens. We find that the CRISPR technology performed best, with low noise, minimal off-target effects and consistent activity across reagents.",
"Detailed functional analyses of many fundamentally important plant genes via conventional loss-of-function approaches are impeded by the severe pleiotropic phenotypes resulting from these losses. In particular, mutations in genes that are required for basic cellular functions and/or reproduction often interfere with the generation of homozygous mutant plants, precluding further functional studies. To overcome this limitation, we devised a clustered regularly interspaced short palindromic repeats (CRISPR)-based tissue-specific knockout system, CRISPR-TSKO, enabling the generation of somatic mutations in particular plant cell types, tissues, and organs. In Arabidopsis (Arabidopsis thaliana), CRISPR-TSKO mutations in essential genes caused well-defined, localized phenotypes in the root cap, stomatal lineage, or entire lateral roots. The modular cloning system developed in this study allows for the efficient selection, identification, and functional analysis of mutant lines directly in the first transgenic generation. The efficacy of CRISPR-TSKO opens avenues for discovering and analyzing gene functions in the spatial and temporal contexts of plant life while avoiding the pleiotropic effects of system-wide losses of gene function.",
"Several programmable transcription factors exist based on the versatile Cas9 protein, yet their relative potency and effectiveness across various cell types and species remain unexplored. Here, we compare Cas9 activator systems and examine their ability to induce robust gene expression in several human, mouse, and fly cell lines. We also explore the potential for improved activation through the combination of the most potent activator systems, and we assess the role of cooperativity in maximizing gene expression.",
"Bacterial type II CRISPR-Cas9 systems have been widely adapted for RNA-guided genome editing and transcription regulation in eukaryotic cells, yet their in vivo target specificity is poorly understood. Here we mapped genome-wide binding sites of a catalytically inactive Cas9 (dCas9) from Streptococcus pyogenes loaded with single guide RNAs (sgRNAs) in mouse embryonic stem cells (mESCs). Each of the four sgRNAs we tested targets dCas9 to between tens and thousands of genomic sites, frequently characterized by a 5-nucleotide seed region in the sgRNA and an NGG protospacer adjacent motif (PAM). Chromatin inaccessibility decreases dCas9 binding to other sites with matching seed sequences; thus 70% of off-target sites are associated with genes. Targeted sequencing of 295 dCas9 binding sites in mESCs transfected with catalytically active Cas9 identified only one site mutated above background levels. We propose a two-state model for Cas9 binding and cleavage, in which a seed match triggers binding but extensive pairing with target DNA is required for cleavage.",
"Systematic interrogation of gene function requires the ability to perturb gene expression in a robust and generalizable manner. Here we describe structure-guided engineering of a CRISPR-Cas9 complex to mediate efficient transcriptional activation at endogenous genomic loci. We used these engineered Cas9 activation complexes to investigate single-guide RNA (sgRNA) targeting rules for effective transcriptional activation, to demonstrate multiplexed activation of ten genes simultaneously, and to upregulate long intergenic non-coding RNA (lincRNA) transcripts. We also synthesized a library consisting of 70,290 guides targeting all human RefSeq coding isoforms to screen for genes that, upon activation, confer resistance to a BRAF inhibitor. The top hits included genes previously shown to be able to confer resistance, and novel candidates were validated using individual sgRNA and complementary DNA overexpression. A gene expression signature based on the top screening hits correlated with markers of BRAF inhibitor resistance in cell lines and patient-derived samples. These results collectively demonstrate the potential of Cas9-based activators as a powerful genetic perturbation technology.",
"Cancer is a major cause of death throughout the world, and there is a large need for better and more personalized approaches to combat the disease. Over the past decade, synthetic lethal approaches have been developed that are designed to exploit the aberrant molecular origins (i.e. defective genes) that underlie tumorigenesis. BLM and CHEK2 are two evolutionarily conserved genes that are somatically altered in a number of tumor types. Both proteins normally function in preserving genome stability through facilitating the accurate repair of DNA double strand breaks. Thus, uncovering synthetic lethal interactors of BLM and CHEK2 will identify novel candidate drug targets and lead chemical compounds. Here we identify an evolutionarily conserved synthetic lethal interaction between SOD1 and both BLM and CHEK2 in two distinct cell models. Using quantitative imaging microscopy, real-time cellular analyses, colony formation and tumor spheroid models we show that SOD1 silencing and inhibition (ATTM and LCS-1 treatments), or the induction of reactive oxygen species (2ME2 treatment) induces selective killing within BLM- and CHEK2-deficient cells relative to controls. We further show that increases in reactive oxygen species follow SOD1 silencing and inhibition that are associated with the persistence of DNA double strand breaks, and increases in apoptosis. Collectively, these data identify SOD1 as a novel candidate drug target in BLM and CHEK2 cancer contexts, and further suggest that 2ME2, ATTM and LCS-1 are lead therapeutic compounds warranting further pre-clinical study."
] |
Detection of Wolbachia pipientis and other bacterial symbionts in Erebidae moths
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Models estimate that up to 80% of all butterfly and moth species host vertically transmitted endosymbiotic microorganisms, which can affect the host fitness, metabolism, reproduction, population dynamics, and genetic diversity, among others. The supporting empirical data are however currently highly biased towards the generally more colourful butterflies, and include less information about moths. Additionally, studies of symbiotic partners of Lepidoptera predominantly focus on the common bacterium Wolbachia pipientis, while infections by other inherited microbial partners have more rarely been investigated. Here, we mine the whole genome sequence data of 47 species of Erebidae moths, with the aims to both inform on the diversity of symbionts potentially associated with this Lepidoptera group, and discuss the potential of metagenomic approaches to inform on host associated microbiome diversity. Based on the result of Kraken2 and MetaPhlAn2 analyses, we found clear evidence of the presence of Wolbachia in four species. Our result also suggests the presence of three other bacterial symbionts (Burkholderia spp., Sodalis spp. and Arsenophonus spp.) in three other moth species. Additionally, we recovered genomic material from bracovirus in about half of our samples. The detection of the latter, usually found in mutualistic association to braconid parasitoid wasps, may inform on host-parasite interactions that take place in the natural habitat of the Erebidae moths, suggesting either contamination with material from species of the host community network, or horizontal transfer of members of the microbiome between interacting species.
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[
"The diversity and abundance of Burkholderia species in sugarcane field soils were investigated by a 16S rRNA gene-based approach using genus-specific primers. A total of 365,721 sequences generated by the Illumina MiSeq platform were assigned to the genus Burkholderia. Nearly 58% of these sequences were placed in a previously defined cluster, including stinkbug symbionts. Quantitative PCR analysis revealed a consistent number of 16S rRNA gene copies for Burkholderia species (10(7) g(-1) soil) across the sampled fields. C/N, pH, and nitrate concentrations were important factors shaping the Burkholderia community structure; however, their impacts were not significant considering the overall genus size.",
"The genus Wolbachia is an archetype of maternally inherited intracellular bacteria that infect the germline of numerous invertebrate species worldwide. They can selfishly alter arthropod sex ratios and reproductive strategies to increase the proportion of the infected matriline in the population. The most common reproductive manipulation is cytoplasmic incompatibility, which results in embryonic lethality in crosses between infected males and uninfected females. Females infected with the same Wolbachia strain rescue this lethality. Despite more than 40 years of research and relevance to symbiont-induced speciation, as well as control of arbovirus vectors and agricultural pests, the bacterial genes underlying cytoplasmic incompatibility remain unknown. Here we use comparative and transgenic approaches to demonstrate that two differentially transcribed, co-diverging genes in the eukaryotic association module of prophage WO from Wolbachia strain wMel recapitulate and enhance cytoplasmic incompatibility. Dual expression in transgenic, uninfected males of Drosophila melanogaster crossed to uninfected females causes embryonic lethality. Each gene additively augments embryonic lethality in crosses between infected males and uninfected females. Lethality associates with embryonic defects that parallel those of wild-type cytoplasmic incompatibility and is notably rescued by wMel-infected embryos in all cases. The discovery of cytoplasmic incompatibility factor genes cifA and cifB pioneers genetic studies of prophage WO-induced reproductive manipulations and informs the continuing use of Wolbachia to control dengue and Zika virus transmission to humans.",
"Symbiotic associations can allow an organism to acquire novel traits by accessing the genetic repertoire of its partner. In the Dictyostelium discoideum farming symbiosis, certain amoebas (termed \"farmers\") stably associate with bacterial partners. Farmers can suffer a reproductive cost but also gain beneficial capabilities, such as carriage of bacterial food (proto-farming) and defense against competitors. Farming status previously has been attributed to amoeba genotype, but the role of bacterial partners in its induction has not been examined. Here, we explore the role of bacterial associates in the initiation, maintenance, and phenotypic effects of the farming symbiosis. We demonstrate that two clades of farmer-associated Burkholderia isolates colonize D. discoideum nonfarmers and infectiously endow them with farmer-like characteristics, indicating that Burkholderia symbionts are a major driver of the farming phenomenon. Under food-rich conditions, Burkholderia-colonized amoebas produce fewer spores than uncolonized counterparts, with the severity of this reduction being dependent on the Burkholderia colonizer. However, the induction of food carriage by Burkholderia colonization may be considered a conditionally adaptive trait because it can confer an advantage to the amoeba host when grown in food-limiting conditions. We observed Burkholderia inside and outside colonized D. discoideum spores after fruiting body formation; this observation, together with the ability of Burkholderia to colonize new amoebas, suggests a mixed mode of symbiont transmission. These results change our understanding of the D. discoideum farming symbiosis by establishing that the bacterial partner, Burkholderia, is an important causative agent of the farming phenomenon.",
"Despite the omnipresence of specific host-symbiont associations with acquisition of the microbial symbiont from the environment, little is known about how the specificity of the interaction evolved and is maintained. The bean bug Riptortus pedestris acquires a specific bacterial symbiont of the genus Burkholderia from environmental soil and harbors it in midgut crypts. The genus Burkholderia consists of over 100 species, showing ecologically diverse lifestyles, and including serious human pathogens, plant pathogens, and nodule-forming plant mutualists, as well as insect mutualists. Through infection tests of 34 Burkholderia species and 18 taxonomically diverse bacterial species, we demonstrate here that nonsymbiotic Burkholderia and even its outgroup Pandoraea could stably colonize the gut symbiotic organ and provide beneficial effects to the bean bug when inoculated on aposymbiotic hosts. However, coinoculation revealed that the native symbiont always outcompeted the nonnative bacteria inside the gut symbiotic organ, explaining the predominance of the native Burkholderia symbiont in natural bean bug populations. Hence, the abilities for colonization and cooperation, usually thought of as specific traits of mutualists, are not unique to the native Burkholderia symbiont but, to the contrary, competitiveness inside the gut is a derived trait of the native symbiont lineage only and was thus critical in the evolution of the insect gut symbiont.",
"Wolbachia, a maternally inherited bacterium induces reproductive alterations in its hosts such as feminization of males, male killing and parthenogenesis. It is the most diverse endosymbiont infecting more than 70% of the insects ranging from pests to pollinators. Kerria lacca-a hemipteran is a sedentary, oriental insect known to produce lac-the only resin of animal origin. The present study was conducted to screen the presence of Wolbachia and its associated phages in the two infrasubspecific forms (four insect lines) of K. lacca viz. kusmi and rengeeni differing from each other on the basis of host preference. Wolbachia and its associated phage were found to be prevalent in all the insect lines analyzed. We, hereby, report the presence of WO-phage (Wolbachia phage) for the first time in K. lacca. Further, phylogenetic data differentiated the kusmi and rengeeni infrasubspecific forms into two different groups on the basis of WO-phage sequences.",
"DNA sequencing continues to decrease in cost with the Illumina HiSeq2000 generating up to 600 Gb of paired-end 100 base reads in a ten-day run. Here we present a protocol for community amplicon sequencing on the HiSeq2000 and MiSeq Illumina platforms, and apply that protocol to sequence 24 microbial communities from host-associated and free-living environments. A critical question as more sequencing platforms become available is whether biological conclusions derived on one platform are consistent with what would be derived on a different platform. We show that the protocol developed for these instruments successfully recaptures known biological results, and additionally that biological conclusions are consistent across sequencing platforms (the HiSeq2000 versus the MiSeq) and across the sequenced regions of amplicons."
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Sentence Prosody and Sentence Recall in Cochlear Implant Users
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Speech prosody, including pitch contour, word stress, pauses, and vowel lengthening, can aid the detection of the clausal structure of a multi-clause sentence and this, in turn, can help listeners determine the meaning. However, for cochlear implant (CI) users, the reduced acoustic richness of the signal raises the question of whether CI users may have difficulty using sentence prosody to detect syntactic clause boundaries within sentences or whether this ability is rescued by the redundancy of the prosodic features that normally co-occur at clause boundaries. Twenty-two CI users, ranging in age from 19 to 77 years old, recalled three types of sentences: sentences in which the prosodic pattern was appropriate to the location of a clause boundary within the sentence (congruent prosody), sentences with reduced prosodic information, or sentences in which the location of the clause boundary and the prosodic marking of a clause boundary were placed in conflict. The results showed the presence of congruent prosody to be associated with superior sentence recall and a reduced processing effort as indexed by the pupil dilation. The individual differences in a standard test of word recognition (consonant-nucleus-consonant score) were related to the recall accuracy as well as the processing effort. The outcomes are discussed in terms of the redundancy of the prosodic features, which normally accompany a clause boundary and processing effort.
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[
"Young and older adults heard sentences in which one character was describing another character (\"The doctor said the nurse is thirsty\"), where the character being described could be determined only by the prosodic pattern in which the sentence was heard. Using computer editing, the authors generated sentences that were heard with either one (Experiment 1) or two (Experiment 2) of three ordinarily co-occurring prosodic features reduced (pitch variation, amplitude variation, timing variation). For both age groups, timing variation was the most valuable of the three prosodic features. These results add to our understanding of the effective preservation of spoken language comprehension in normal aging.",
"In the present study, a computational model of phoneme identification was applied to data from a previous study, wherein cochlear implant (CI) users' adaption to a severely shifted frequency allocation map was assessed regularly over 3 months of continual use. This map provided more input filters below 1 kHz, but at the expense of introducing a downwards frequency shift of up to one octave in relation to the CI subjects' clinical maps. At the end of the 3-month study period, it was unclear whether subjects' asymptotic speech recognition performance represented a complete or partial adaptation. To clarify the matter, the computational model was applied to the CI subjects' vowel identification data in order to estimate the degree of adaptation, and to predict performance levels with complete adaptation to the frequency shift. Two model parameters were used to quantify this adaptation; one representing the listener's ability to shift their internal representation of how vowels should sound, and the other representing the listener's uncertainty in consistently recalling these representations. Two of the three CI users could shift their internal representations towards the new stimulation pattern within 1 week, whereas one could not do so completely even after 3 months. Subjects' uncertainty for recalling these representations increased substantially with the frequency-shifted map. Although this uncertainty decreased after 3 months, it remained much larger than subjects' uncertainty with their clinically assigned maps. This result suggests that subjects could not completely remap their phoneme labels, stored in long-term memory, towards the frequency-shifted vowels. The model also predicted that even with complete adaptation, the frequency-shifted map would not have resulted in improved speech understanding. Hence, the model presented here can be used to assess adaptation, and the anticipated gains in speech perception expected from changing a given CI device parameter.",
"The present study used eight normal-hearing (NH) subjects, listening to acoustic cochlear implant (CI) simulations, to examine the effects of spectral shifting on speech recognition in noise. Speech recognition was measured using spectrally matched and shifted speech (vowels, consonants, and IEEE sentences), generated by 8-channel, sine-wave vocoder. Measurements were made in quiet and in noise (speech-shaped static noise and speech-babble at 5 dB signal-to-noise ratio). One spectral match condition and four spectral shift conditions were investigated: 2 mm, 3 mm, and 4 mm linear shift, and 3 mm shift with compression, in terms of cochlear distance. Results showed that speech recognition scores dropped because of noise and spectral shifting, and that the interactive effects of spectral shifting and background conditions depended on the degree/type of spectral shift, background conditions, and the speech test materials. There was no significant interaction between spectral shifting and two noise conditions for all speech test materials. However, significant interactions between linear spectral shifts and all background conditions were found in sentence recognition; significant interactions between spectral shift types and all background conditions were found in vowel recognition. Overall, the results suggest that tonotopic mismatch may affect performance of CI users in complex listening environments.",
"Gap detection thresholds were measured in patients with the Nucleus and Symbion cochlear implants as a function of several current waveform parameters. Detection of gaps in an electrical sinusoidal stimulus or in a train of biphasic pulses by implanted patients was similar to detection of gaps in comparable acoustic stimuli by normal listeners. Threshold gaps were 20-50 ms for low-level stimuli and improved with stimulus level to 2-5 ms for high-level stimuli. Gap detection performance was not affected by the electrode position in the cochlea or by the distance between stimulating electrodes. The data from most patients were well fitted by a trading relation between the duration of the gap and the square of stimulus intensity, indicating energy detection. The similarity of gap thresholds for normal subjects and implant patients suggests that many details of the peripheral neural activity are probably not important for this task, and that there is no retrocochlear loss of auditory temporal resolution with sensorineural hearing loss.",
"To generate an evidence-based algorithm for the use of intraoperative testing during cochlear implantation (CI). Retrospective review. Tertiary referral center. A total of 277 children (aged 6 mo to 17 yr) and adults 18 years and older with normal cochlear anatomy who underwent primary and revision cochlear implantation at a single center between 2005 and 2010 were included. Intraoperative electrophysiologic monitoring and intraoperative Stenver's view plain film radiography. Intraoperative testing included the following: 1) individual electrode impedance measurements; 2) neural response telemetry (tNRT) levels for electrodes E20, E15, E10, and E5; and 3) plain film radiograph assessment of electrode position. No patient demonstrated abnormalities on all 3 modalities. Open or short electrodes on impedance testing were found in 6% of patients; half of these normalized when remeasured. Absent tNRT responses on 1 or more electrodes occurred in 14% of patients, although complete lack of response was rare (1.4%) and did not correlate with a dysfunctional device. Spread of excitation was performed in 1 patient and was consistent with a tip rollover. Intraoperative radiography identified tip-rollover and extracochlear electrode placement in all cases (n = 5, 1.8%) and prompted the use of the backup device. Immediate intraoperative determination of device functionality and optimal electrode placement is advantageous. Of the modalities tested, including electrode impedance, tNRT, and plain radiograph, only the radiographic results impacted intraoperative surgical decision making and led to the use of the backup device.",
"Commercially available cochlear implant systems attempt to deliver frequency information going down to a few hundred Hertz, but the electrode arrays are not designed to reach the most apical regions of the cochlea, which correspond to these low frequencies. This may cause a mismatch between the frequencies presented by a cochlear implant electrode array and the frequencies represented at the corresponding location in a normal-hearing cochlea. In the following study, the mismatch between the frequency presented at a given cochlear angle and the frequency expected by an acoustic hearing ear at the corresponding angle is examined for the cochlear implant systems that are most commonly used in the United States. The angular insertion of each of the electrodes on four different electrode arrays (MED-EL Standard, MED-EL Flex28, Advanced Bionics HiFocus 1J, and Cochlear Contour Advance) was estimated from X-ray. For the angular location of each electrode on each electrode array, the predicted spiral ganglion frequency was estimated. The predicted spiral ganglion frequency was compared with the center frequency provided by the corresponding electrode using the manufacturer's default frequency-to-electrode allocation. Differences across devices were observed for the place of stimulation for frequencies below 650 Hz. Longer electrode arrays (i.e., the MED-EL Standard and Flex28) demonstrated smaller deviations from the spiral ganglion map than the other electrode arrays. For insertion angles up to approximately 270°, the frequencies presented at a given location were typically approximately an octave below what would be expected by a spiral ganglion frequency map, while the deviations were larger for angles deeper than 270°. For frequencies above 650 Hz, the frequency to angle relationship was consistent across all four electrode models. A mismatch was observed between the predicted frequency and the default frequency provided by every electrode on all electrode arrays. The mismatch can be reduced by changing the default frequency allocations, inserting electrodes deeper into the cochlea, or allowing cochlear implant users to adapt to the mismatch. Further studies are required to fully assess the clinical significance of the frequency mismatch.",
"The development of a procedure for the measurement of insertion depth angles of cochlear implant electrode arrays based on flat-panel computed tomography (FPCT) and the application of this technique to in vivo postoperative images. The knowledge of the insertion depth angle of electrode arrays is relevant for the preservation of low-frequency residual hearing and for optimizing speech coding strategies. Until now, the angular position of electrodes was derived from 2-dimensional radiographs. In the present study, 3-dimensional (3D) radiographs provided by FPCT were used to determine the insertion depth according to angular electrode positions with higher accuracy. For this purpose, a new evaluation procedure was designed and applied to radiographs of 15 cochlear implant patients. In contrast to 2-dimensional radiographs, the obtained 3D images show all 3 semicircular channels and therefore allow the determination of a clear reference, which is required for precise insertion angle measurements. Furthermore, the presented FPCT radiographs visualize distinct electrodes. Despite the constant length of the implanted electrode arrays, we have found a considerable variation of measured insertion depth angles, which is consistent with published observations on the variability and the gender dependence of the size of human cochleae. FPCT provides 3D high-resolution radiographic data that enable the determination of the insertion depth angle with high accuracy and, potentially, an angle determination of individual electrodes. Therefore, this low-dose technique is especially appropriate for postoperative investigations after cochlea implantation."
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Noninvasive index for acute hypoxemic respiratory failure in the intensive care unit
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Acute hypoxemic respiratory failure (AHRF) is a major factor for increased mortality in the intensive care unit (ICU). We hypothesized that the noninvasive index SpO
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[
"Traditional measures of ARDS severity such as Pao2/Fio2 may not reliably predict clinical outcomes. The oxygenation index (OI [Fio2 × mean airway pressure × 100)/Pao2]) may more accurately reflect ARDS severity but requires arterial blood gas measurement. We hypothesized that the oxygenation saturation index (OSI [Fio2 × mean airway pressure × 100)/oxygen saturation by pulse oximetry (Spo2)]) is a reliable noninvasive surrogate for the OI that is associated with hospital mortality and ventilator-free days (VFDs) in patients with ARDS. Critically ill patients enrolled in a prospective cohort study were eligible if they developed ARDS (Berlin criteria) during the first 4 ICU days and had mean airway pressure, Spo2/Fio2, and Pao2/Fio2 values recorded on the first day of ARDS (N = 329). The highest mean airway pressure and lowest Spo2/Fio2 and Pao2/Fio2 values were used to calculate OI and OSI. The association between OI or OSI and hospital mortality or VFD was analyzed by using logistic regression and linear regression, respectively. The area under the receiver-operating characteristic curve (AUC) for mortality was compared among OI, OSI, Spo2/Fio2, Pao2/Fio2, and Acute Physiology and Chronic Health Evaluation II scores. OI and OSI were strongly correlated (rho = 0.862; P < .001). OSI was independently associated with hospital mortality (OR per 5-point increase in OSI, 1.228 [95% CI, 1.056-1.429]; P = .008). OI and OSI were each associated with a reduction in VFD (OI, P = .023; OSI, P = .005). The AUC for mortality prediction was greatest for Acute Physiology and Chronic Health Evaluation II scores (AUC, 0.695; P < .005) and OSI (AUC, 0.602; P = .007). The AUC for OSI was substantially better in patients aged < 40 years (AUC, 0.779; P < .001). In patients with ARDS, the OSI was correlated with the OI. The OSI on the day of ARDS diagnosis was significantly associated with increased mortality and fewer VFDs. The findings suggest that OSI is a reliable surrogate for OI that can noninvasively provide prognostic information and assessment of ARDS severity.",
"Prediction of death and prolonged mechanical ventilation is important in terms of projecting resource utilization and in establishing protocols for clinical studies of acute lung injury (ALI). We aimed to identify risk factors for a combined end-point of death and/or prolonged ventilator dependence and developed an ALI-specific prediction model. In this retrospective analysis of three multicenter clinical studies, we identified predictors of death or ventilator dependence from variables prospectively recorded during the first three days of mechanical ventilation. After the prediction model was derived in an international cohort of patients with ALI, it was validated in two independent samples of patients enrolled in a clinical trial involving 17 academic centers and a North American population-based cohort. A combined end-point of death and/or ventilator dependence at 14 days or later occurred in 68% of patients in the international cohort, 60% of patients in the clinical trial, and 59% of patients in the population-based cohort. In the derivation cohort, a model based on age, oxygenation index on day 3, and cardiovascular failure on day 3 predicted death and/or ventilator dependence. The prediction model performed better in the clinical trial validation cohort (area under the receiver operating curve 0.81, 95% confidence interval 0.77 to 0.84) than in the population-based validation cohort (0.71, 95% confidence interval 0.65 to 0.76). A model based on age and cardiopulmonary function three days after the intubation is able to predict, moderately well, a combined end-point of death and/or prolonged mechanical ventilation in patients with ALI.",
"We assessed rates and predictive factors of non-invasive ventilation (NIV) failure in patients admitted to the intensive care unit (ICU) for non-hypercapnic acute hypoxemic respiratory failure (AHRF). This is an observational cohort study using data prospectively collected over a three-year period in a medical ICU of a university hospital. Among 113 patients receiving NIV for AHRF, 82 had acute respiratory distress syndrome (ARDS) and 31 had non-ARDS. Intubation rates significantly differed between ARDS and non-ARDS patients (61% versus 35%, P = 0.015) and according to clinical severity of ARDS: 31% in mild, 62% in moderate, and 84% in severe ARDS (P = 0.0016). In-ICU mortality rates were 13% in non-ARDS, and, respectively, 19%, 32% and 32% in mild, moderate and severe ARDS (P = 0.22). Among patients with moderate ARDS, NIV failure was lower among those having a PaO2/FiO2 >150 mmHg (45% vs. 74%, p = 0.04). NIV failure was associated with active cancer, shock, moderate/severe ARDS, lower Glasgow coma score and lower positive end-expiratory pressure level at NIV initiation. Among intubated patients, ICU mortality rate was 46% overall and did not differ according to the time to intubation. With intubation rates below 35% in non-ARDS and mild ARDS, NIV stands as the first-line approach; NIV may be attempted in ARDS patients with a PaO2/FiO2 > 150. By contrast, 84% of severe ARDS required intubation and NIV did not appear beneficial in this subset of patients. However, the time to intubation had no influence on mortality.",
"An American College of Chest Physicians/Society of Critical Care Medicine Consensus Conference was held in Northbrook in August 1991 with the goal of agreeing on a set of definitions that could be applied to patients with sepsis and its sequelae. New definitions were offered for some terms, while others were discarded. Broad definitions of sepsis and the systemic inflammatory response syndrome were proposed, along with detailed physiologic parameters by which a patient may be categorized. Definitions for severe sepsis, septic shock, hypotension, and multiple organ dysfunction syndrome were also offered. The use of severity scoring methods when dealing with septic patients was recommended as an adjunctive tool to assess mortality. Appropriate methods and applications for the use and testing of new therapies were recommended. The use of these terms and techniques should assist clinicians and researchers who deal with sepsis and its sequelae."
] |
Short-term versus long-term neoadjuvant aromatase inhibitor therapy for breast cancer
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Aromatase inhibitor (AI) treatment is the standard of care for postmenopausal women with primary estrogen receptor-positive breast cancer. The impact of duration of neoadjuvant endocrine therapy (NET) on molecular characteristics is still unknown. We evaluated and compared changes of gene expression profiles under short-term (2-week) versus longer-term neoadjuvant AIs.
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[
"HER2-positive breast cancer consists of four intrinsic molecular subtypes-luminal A, luminal B, HER2-enriched, and basal-like-and a normal-like subtype, with the HER2-enriched subtype having the highest activation of the EGFR-HER2 pathway. We aimed to test the hypothesis that patients with the HER2-enriched subtype benefit the most from dual HER2 blockade. PAMELA is an open-label, single-group, phase 2 trial done in 19 hospitals in Spain. We recruited female patients aged at least 18 years with previously untreated, centrally confirmed HER2-positive, stage I-IIIA invasive breast cancer regardless of hormone receptor status. Patients were given lapatinib (1000 mg per day orally) and trastuzumab (loading dose of 8 mg/kg, followed by 6 mg/kg every 3 weeks intravenously) for 18 weeks; hormone receptor-positive patients were additionally given letrozole (2·5 mg per day orally; if menopausal) or tamoxifen (20 mg per day orally; if premenopausal). Surgery was done 1-3 weeks after the last dose of study treatment. Intrinsic molecular subtypes of tumour biopsy samples taken at baseline (day 0) and day 14 were determined with the PAM50 predictor. The primary outcome was the ability of the HER2-enriched subtype to predict pathological complete response at the time of surgery. The primary outcome was assessed in the evaluable population (ie, all patients who had initial tumour biopsy samples available and who underwent definitive surgery) and safety was assessed in all patients who received at least one part of study treatment. This study is registered with ClinicalTrials.gov, number NCT01973660, and is completed. Between Oct 28, 2013, and Nov 26, 2015, we recruited 151 patients, of whom 14 (9%) discontinued treatment and 137 (91%) completed treatment as planned. At baseline, most patients had the HER2-enriched subtype (101 [67%]), followed by luminal A (22 [15%]), luminal B (16 [11%]), basal-like (nine [6%]), and normal-like (three [2%]) subtypes. At the time of surgery, 46 (30%, 95% CI 23-39) of 151 patients had pathological complete response in the breast. 41 (41%, 31-51) of 101 patients with the HER2-enriched subtype and five (10%, 4-23) of 50 patients with non-HER2-enriched subtypes achieved pathological complete response at the time of surgery (odds ratio 6·2, 95% CI 2·3-16·8; p=0·0004). The HER2-enriched subtype can identify patients with HER2-positive breast cancer who are likely to benefit from dual HER2 blockade therapies. GlaxoSmithKline, Susan Komen Foundation, CERCA Programme-Generalitat de Catalunya, Banco Bilbao Vizcaya Argentaria Foundation, Pas a Pas, and the Breast Cancer Research Foundation.",
"Background: In hormone receptor-positive (HR+)/HER2-negative breast cancer, the HER2-enriched and Basal-like intrinsic subtypes are associated with poor outcome, low response to anti-estrogen therapy and high response to chemotherapy. To date, no validated biomarker exists to identify both molecular entities other than gene expression. Methods: PAM50 subtyping and immunohistochemical data were obtained from 8 independent studies of 1,416 HR+/HER2-negative early breast tumors. A non-luminal disease score (NOLUS) from 0 to 100, based on percentage of estrogen receptor (ER), progesterone receptor (PR) and Ki67 tumor cells, was derived in a combined cohort of 5 studies (training dataset) and tested in a combined cohort of 3 studies. The performance of NOLUS was estimated using Area Under the ROC Curve (AUC). Results: In the training dataset (n = 903) and compared to luminal disease, non-luminal disease had lower percentage of ER-positive cells (median 65.2 vs. 86.2%, p < 0.01) and PR-positive cells (33.2 vs. 56.4%, p < 0.01) and higher percentage of Ki67-positive cells (18.2 vs. 13.1%, p = 0.01). A NOLUS formula was derived: -0.45*ER -0.28*PR +0.27*Ki67 + 73.02. The proportion of non-luminal tumors in NOLUS-positive (≥51.38) and NOLUS-negative (<51.38) groups was 52.6 and 8.7%, respectively. In the testing dataset (n = 514), NOLUS was found significantly associated with non-luminal disease (p < 0.01) with an AUC 0.902. The proportion of non-luminal tumors in NOLUS-positive and NOLUS-negative groups was 76.9% (56.4-91.0%) and 2.6% (1.4-4.5%), respectively. The sensitivity and specificity of the pre-specified cutoff was 59.3 and 98.7%, respectively. Conclusions: In the absence of gene expression data, NOLUS can help identify non-luminal disease within HR+/HER2-negative breast cancer.",
"Current immunohistochemical (IHC)-based definitions of luminal A and B breast cancers are imperfect when compared with multigene expression-based assays. In this study, we sought to improve the IHC subtyping by examining the pathologic and gene expression characteristics of genomically defined luminal A and B subtypes. Gene expression and pathologic features were collected from primary tumors across five independent cohorts: British Columbia Cancer Agency (BCCA) tamoxifen-treated only, Grupo Español de Investigación en Cáncer de Mama 9906 trial, BCCA no systemic treatment cohort, PAM50 microarray training data set, and a combined publicly available microarray data set. Optimal cutoffs of percentage of progesterone receptor (PR) -positive tumor cells to predict survival were derived and independently tested. Multivariable Cox models were used to test the prognostic significance. Clinicopathologic comparisons among luminal A and B subtypes consistently identified higher rates of PR positivity, human epidermal growth factor receptor 2 (HER2) negativity, and histologic grade 1 in luminal A tumors. Quantitative PR gene and protein expression were also found to be significantly higher in luminal A tumors. An empiric cutoff of more than 20% of PR-positive tumor cells was statistically chosen and proved significant for predicting survival differences within IHC-defined luminal A tumors independently of endocrine therapy administration. Finally, no additional prognostic value within hormonal receptor (HR) -positive/HER2-negative disease was observed with the use of the IHC4 score when intrinsic IHC-based subtypes were used that included the more than 20% PR-positive tumor cells and vice versa. Semiquantitative IHC expression of PR adds prognostic value within the current IHC-based luminal A definition by improving the identification of good outcome breast cancers. The new proposed IHC-based definition of luminal A tumors is HR positive/HER2 negative/Ki-67 less than 14%, and PR more than 20%.",
"PAM50/Prosigna gene expression-based assay identifies three categorical risk of relapse groups (ROR-low, ROR-intermediate and ROR-high) in post-menopausal patients with estrogen receptor estrogen receptor-positive (ER+)/ HER2-negative (HER2-) early breast cancer. Low risk patients might not need adjuvant chemotherapy since their risk of distant relapse at 10-years is below 10% with endocrine therapy only. In this study, 517 consecutive patients with ER+/HER2- and node-negative disease were evaluated for Ki67 and Prosigna. Most of Luminal A tumors (65.6%) and ROR-low tumors (70.9%) had low Ki67 values (0-10%); however, the percentage of patients with ROR-medium or ROR-high disease within the Ki67 0-10% group was 42.7% (with tumor sizes ≤2 cm) and 33.9% (with tumor sizes > 2 cm). Finally, we found that the optimal Ki67 cutoff for identifying Luminal A or ROR-low tumors was 14%. Ki67 as a surrogate biomarker in identifying Prosigna low-risk outcome patients or Luminal A disease in the clinical setting is unreliable. In the absence of a well-validated prognostic gene expression-based assay, the optimal Ki67 cutoff for identifying low-risk outcome patients or Luminal A disease remains at 14%.",
"Multiple breast cancer gene expression profiles have been developed that appear to provide similar abilities to predict outcome and may outperform clinical-pathologic criteria; however, the extent to which seemingly disparate profiles provide additive prognostic information is not known, nor do we know whether prognostic profiles perform equally across clinically defined breast cancer subtypes. We evaluated whether combining the prognostic powers of standard breast cancer clinical variables with a large set of gene expression signatures could improve on our ability to predict patient outcomes. Using clinical-pathological variables and a collection of 323 gene expression \"modules\", including 115 previously published signatures, we build multivariate Cox proportional hazards models using a dataset of 550 node-negative systemically untreated breast cancer patients. Models predictive of pathological complete response (pCR) to neoadjuvant chemotherapy were also built using this approach. We identified statistically significant prognostic models for relapse-free survival (RFS) at 7 years for the entire population, and for the subgroups of patients with ER-positive, or Luminal tumors. Furthermore, we found that combined models that included both clinical and genomic parameters improved prognostication compared with models with either clinical or genomic variables alone. Finally, we were able to build statistically significant combined models for pathological complete response (pCR) predictions for the entire population. Integration of gene expression signatures and clinical-pathological factors is an improved method over either variable type alone. Highly prognostic models could be created when using all patients, and for the subset of patients with lymph node-negative and ER-positive breast cancers. Other variables beyond gene expression and clinical-pathological variables, like gene mutation status or DNA copy number changes, will be needed to build robust prognostic models for ER-negative breast cancer patients. This combined clinical and genomics model approach can also be used to build predictors of therapy responsiveness, and could ultimately be applied to other tumor types.",
"Purpose: To identify a predictive biomarker for durvalumab, an anti-programmed death ligand 1 (PD-L1) mAb.Experimental Design: RNA sequencing of 97 advanced-stage non-small cell lung carcinoma (NSCLC) biopsies from a nonrandomized phase Ib/II clinical trial (1108/NCT01693562) were profiled to identify a predictive signature; 62 locally advanced or metastatic urothelial cancer tumors from the same study were profiled to confirm predictive utility of the signature. Thirty NSCLC patients provided pre- and posttreatment tumors for messenger RNA (mRNA) analysis. NSCLC with ≥25% tumor cells and urothelial cancer with ≥25% tumor or immune cells stained for PD-L1 at any intensity were scored PD-L1 positive (PD-L1+). Kaplan-Meier and Cox proportional hazards analyses were used to adjust for gender, age, prior therapies, histology, ECOG status, liver metastasis, and smoking. Tumor mutation burden (TMB) was calculated using data from The Cancer Genome Atlas (TCGA).Results: In the NSCLC discovery set, a four-gene IFNγ-positive (IFNγ+) signature comprising IFNγ, CD274, LAG3, and CXCL9 was associated with higher overall response rates, longer median progression-free survival, and overall survival compared with signature-low patients. IFNγ+-signature NSCLC patients had improved survival regardless of IHC PD-L1 status. These associations were replicated in a urothelial cancer cohort. The IFNγ+ signature was induced 2-fold (P = 0.003) by durvalumab after 8 weeks of therapy in patients with NSCLC, and baseline signature was associated with TMB but not survival in TCGA data.Conclusions: The IFNγ+ mRNA signature may assist in identifying patients with improved outcomes with durvalumab, independent of PD-L1 assessed by IHC. Clin Cancer Res; 24(16); 3857-66. ©2018 AACR.",
"Purpose: Cyclin-dependent kinase (CDK) 4/6 drives cell proliferation in estrogen receptor-positive (ER+) breast cancer. This single-arm phase II neoadjuvant trial (NeoPalAna) assessed the antiproliferative activity of the CDK4/6 inhibitor palbociclib in primary breast cancer as a prelude to adjuvant studies.Experimental Design: Eligible patients with clinical stage II/III ER+/HER2- breast cancer received anastrozole 1 mg daily for 4 weeks (cycle 0; with goserelin if premenopausal), followed by adding palbociclib (125 mg daily on days 1-21) on cycle 1 day 1 (C1D1) for four 28-day cycles unless C1D15 Ki67 > 10%, in which case patients went off study due to inadequate response. Anastrozole was continued until surgery, which occurred 3 to 5 weeks after palbociclib exposure. Later patients received additional 10 to 12 days of palbociclib (Cycle 5) immediately before surgery. Serial biopsies at baseline, C1D1, C1D15, and surgery were analyzed for Ki67, gene expression, and mutation profiles. The primary endpoint was complete cell cycle arrest (CCCA: central Ki67 ≤ 2.7%).Results: Fifty patients enrolled. The CCCA rate was significantly higher after adding palbociclib to anastrozole (C1D15 87% vs. C1D1 26%, P < 0.001). Palbociclib enhanced cell-cycle control over anastrozole monotherapy regardless of luminal subtype (A vs. B) and PIK3CA status with activity observed across a broad range of clinicopathologic and mutation profiles. Ki67 recovery at surgery following palbociclib washout was suppressed by cycle 5 palbociclib. Resistance was associated with nonluminal subtypes and persistent E2F-target gene expression.Conclusions: Palbociclib is an active antiproliferative agent for early-stage breast cancer resistant to anastrozole; however, prolonged administration may be necessary to maintain its effect. Clin Cancer Res; 23(15); 4055-65. ©2017 AACR."
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Targeting Ewing sarcoma via the EWS-FLI transcriptional network
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EWS/FLI is the defining mutation of Ewing sarcoma. This oncogene drives malignant transformation and progression and occurs in a genetic background characterized by few other recurrent cooperating mutations. In addition, the tumor is absolutely dependent on the continued expression of EWS/FLI to maintain the malignant phenotype. However, EWS/FLI is a transcription factor and therefore a challenging drug target. The difficulty of directly targeting EWS/FLI stems from unique features of this fusion protein as well as the network of interacting proteins required to execute the transcriptional program. This network includes interacting proteins as well as upstream and downstream effectors that together reprogram the epigenome and transcriptome. While the vast number of proteins involved in this process challenge the development of a highly specific inhibitors, they also yield numerous therapeutic opportunities. In this report, we will review how this vast EWS-FLI transcriptional network has been exploited over the last two decades to identify compounds that directly target EWS/FLI and/or associated vulnerabilities.
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[
"Among primitive neuroectodermal tumors, Ewing tumors are characterized by a gene rearrangement recombining the 5'-EWS gene portion with one of two ETS-related proto-oncogenes, FLI-1 or ERG, in roughly 90 and 10% of cases, respectively. We attempted to antagonize EWS/FLI-1 function in Ewing tumor cell lines. As a control, a cell line derived from another small round cell tumor of neuroectodermal origin, neuroblastoma, was used. This cell line was found to express almost identical patterns of ETS proteins except for EWS/FLI-1 and a novel, ELF-related gene product. Stable expression of antisense EWS/FLI-1 cDNA resulted in decreased endogenous EWS/FLI-1 RNA levels and growth reduction of ET cells but not of neuroblastoma cells. DNA-binding FLI-1 derivatives localizing to the nucleus in which the 5'-EWS regulatory domain was replaced by bacterial beta-galactosidase dominantly modulated transcriptional transactivation from a degenerate ETS-binding motif. Transient transfection of these dominant-negative recombinants resulted in a significant decrease in the relative number of mitoses in four Ewing tumor cell lines tested but not in the neuroblastoma cell line. The presented evidence for modulation of tumor cell proliferation by EWS/FLI-1 antagonists suggests a causal role for EWS/FLI-1-mediated gene activation in the malignant transformation of the enigmatic Ewing tumor-precursor cell.",
"Ewing sarcomas (ES) are highly malignant bone or soft tissue tumors. Genetically, ES are defined by balanced chromosomal EWS/ETS translocations, which give rise to chimeric proteins (EWS-ETS) that generate an oncogenic transcriptional program associated with altered epigenetic marks throughout the genome. By use of an inhibitor (JQ1) blocking BET bromodomain binding proteins (BRDs) we strikingly observed a strong down-regulation of the predominant EWS-ETS protein EWS-FLI1 in a dose dependent manner. This was further enhanced by co-treatment with an inhibitor of the PI3K pathway. Microarray analysis further revealed JQ1 treatment to block a typical ES associated expression program. The effect on this expression program was mimicked by RNA interference with BRD3 or BRD4 expression, indicating that the EWS-FLI1 mediated expression profile is at least in part mediated via such epigenetic readers. Consequently, contact dependent and independent proliferation of different ES lines was strongly inhibited. Mechanistically, treatment of ES resulted in a partial arrest of the cell cycle as well as induction of apoptosis. Tumor development was suppressed dose dependently in a xeno-transplant model in immune deficient mice, overall indicating that ES may be susceptible to treatment with epigenetic inhibitors blocking BET bromodomain activity and the associated pathognomonic EWS-ETS transcriptional program.",
"Ewing sarcoma (ES), a highly aggressive tumor of children and young adults, is characterized most commonly by an 11;22 chromosomal translocation that fuses EWSR1 located at 22q12 with FLI1, coding for a member of the ETS family of transcription factors. Although genetic changes in ES have been extensively researched, our understanding of the role of epigenetic modifications in this neoplasm is limited. In an effort to improve our knowledge in the role of epigenetic changes in ES we evaluated the in vitro antineoplastic effect of the DNA methyltransferase inhibitor 5-Aza-deoxycytidine (5-Aza-dC) and identified epigenetically silenced genes by pharmacologic unmasking of DNA methylation coupled with genome-wide expression profiling. Comparisons between untreated and 5-Aza-dC treated ES cell lines (n = 5) identified 208 probe sets with at least twofold difference in expression (P ≤ 0.05). The 208 probe sets represented 145 upregulated and 31 down-regulated genes. Of the 145 genes upregulated after 5-Aza-dC treatment, four: were further characterized. ACRC, CLU, MEST, and NNAT were found to be hypermethylated and transcriptionally down-regulated in ES cell lines. Further studies revealed that ACRC, CLU, MEST, and NNAT were often hypermethylated in primary ES tumors. Transfection-mediated reexpression of ACRC, CLU, MEST, and NNAT in ES cell lines resulted in decreased growth in culture. This study demonstrated epigenetically modified genes in ES cell lines and primary tumors and suggested that epigenetic dysregulation may contribute to disease pathogenesis in ES.",
"Ewing Sarcoma is a rare bone and soft tissue malignancy affecting children and young adults. Chromosomal translocations in this cancer produce fusion oncogenes as characteristic molecular signatures of the disease. The most common case is the translocation t (11; 22) (q24;q12) which yields the EWS-Fli1 chimeric transcription factor. Finding a way to directly target EWS-Fli1 remains a central therapeutic approach to eradicate this aggressive cancer. Here we demonstrate that treating Ewing Sarcoma cells with JQ1(+), a BET bromodomain inhibitor, represses directly EWS-Fli1 transcription as well as its transcriptional program. Moreover, the Chromatin Immuno Precipitation experiments demonstrate for the first time that these results are a consequence of the depletion of BRD4, one of the BET bromodomains protein from the EWS-Fli1 promoter. In vitro, JQ1(+) treatment reduces the cell viability, impairs the cell clonogenic and the migratory abilities, and induces a G1-phase blockage as well as a time- and a dose-dependent apoptosis. Furthermore, in our in vivo model, we observed a tumor burden delay, an inhibition of the global vascularization and an increase of the mice overall survival. Taken together, our data indicate that inhibiting the BET bromodomains interferes with EWS-FLi1 transcription and could be a promising strategy in the Ewing tumors context.",
"ETS proteins are a family of transcription factors that play important roles in the development of cancer. The Ewing's sarcoma EWS/ETS fusion oncoproteins control a number of cancer-relevant phenotypes in that disease. We recently demonstrated that EWS/FLI, the most common EWS/ETS fusion in Ewing's sarcoma, regulates a portion of its target genes, including the critical target NR0B1, via GGAA-containing microsatellites in their promoters. Given the unusual nature of microsatellites as EWS/FLI response elements, we sought to elucidate the mechanism of EWS/FLI activity at these sites. We found that the ability to bind GGAA microsatellites is shared by multiple ETS family members from distinct phylogenetic subfamilies. Importantly, however, only EWS/ETS-containing fusions are capable of mediating transcriptional activation via these elements, highlighting a neomorphic function of the Ewing's sarcoma fusion proteins. Additional analysis revealed that the GGAA microsatellite binds EWS/FLI with an affinity that is 2 to 3 orders of magnitude lower than previously identified high-affinity consensus/redundant binding sites. The stoichiometry of this interaction is 2 protein molecules for each DNA molecule, suggesting that EWS/FLI binds these elements as a homodimer. The isolated FLI ETS domain bound microsatellite sequences in a nearly identical fashion to full-length EWS/FLI, thus indicating that residues required for homodimeric binding are localized to the ETS domain. These data suggest a new paradigm for an ETS family member binding to DNA at cancer-relevant genetic loci and highlight emergent properties of EWS/FLI that are required for the development of Ewing's sarcoma.",
"Ewing sarcoma is an aggressive bone cancer of children and young adults defined by the presence of a chromosomal translocation: t(11;22)(q24;q12). The encoded protein, EWS/FLI, fuses the amino-terminal domain of EWS to the carboxyl-terminus of FLI. The EWS portion is an intrinsically disordered transcriptional regulatory domain, while the FLI portion contains an ETS DNA-binding domain and two flanking regions of unknown function. Early studies using non-Ewing sarcoma models provided conflicting information on the roles of each domain of FLI in EWS/FLI oncogenic function. We therefore sought to define the specific contributions of each FLI domain to EWS/FLI activity in a well-validated Ewing sarcoma model and, in doing so, to better understand Ewing sarcoma development mediated by the fusion protein. We analyzed a series of engineered EWS/FLI mutants with alterations in the FLI portion using a variety of assays. Fluorescence anisotropy, CUT&RUN, and ATAC-sequencing experiments revealed that the isolated ETS domain is sufficient to maintain the normal DNA-binding and chromatin accessibility function of EWS/FLI. In contrast, RNA-sequencing and soft agar colony formation assays revealed that the ETS domain alone was insufficient for transcriptional regulatory and oncogenic transformation functions of the fusion protein. We found that an additional alpha-helix immediately downstream of the ETS domain is required for full transcriptional regulation and EWS/FLI-mediated oncogenesis. These data demonstrate a previously unknown role for FLI in transcriptional regulation that is distinct from its DNA-binding activity. This activity is critical for the cancer-causing function of EWS/FLI and may lead to novel therapeutic approaches.",
"The ETS gene family is frequently involved in chromosome translocations that cause human cancer, including prostate cancer, leukemia, and sarcoma. However, the mechanisms by which oncogenic ETS proteins, which are DNA-binding transcription factors, target genes necessary for tumorigenesis is not well understood. Ewing's sarcoma serves as a paradigm for the entire class of ETS-associated tumors because nearly all cases harbor recurrent chromosomal translocations involving ETS genes. The most common translocation in Ewing's sarcoma encodes the EWS/FLI oncogenic transcription factor. We used whole genome localization (ChIP-chip) to identify target genes that are directly bound by EWS/FLI. Analysis of the promoters of these genes demonstrated a significant over-representation of highly repetitive GGAA-containing elements (microsatellites). In a parallel approach, we found that EWS/FLI uses GGAA microsatellites to regulate the expression of some of its target genes including NR0B1, a gene required for Ewing's sarcoma oncogenesis. The microsatellite in the NR0B1 promoter bound EWS/FLI in vitro and in vivo and was both necessary and sufficient to confer EWS/FLI regulation to a reporter gene. Genome wide computational studies demonstrated that GGAA microsatellites were enriched close to EWS/FLI-up-regulated genes but not down-regulated genes. Mechanistic studies demonstrated that the ability of EWS/FLI to bind DNA and modulate gene expression through these repetitive elements depended on the number of consecutive GGAA motifs. These findings illustrate an unprecedented route to specificity for ETS proteins and use of microsatellites in tumorigenesis."
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Geometric Microliths and Approximate Bayesian Computation for the Neolithic in the Iberian Peninsula
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In the present article we use geometric microliths (a specific type of arrowhead) and Approximate Bayesian Computation (ABC) in order to evaluate possible origin points and expansion routes for the Neolithic in the Iberian Peninsula. In order to do so, we divide the Iberian Peninsula in four areas (Ebro river, Catalan shores, Xúquer river and Guadalquivir river) and we sample the geometric microliths existing in the sites with the oldest radiocarbon dates for each zone. On this data, we perform a partial Mantel test with three matrices: geographic distance matrix, cultural distance matrix and chronological distance matrix. After this is done, we simulate a series of partial Mantel tests where we alter the chronological matrix by using an expansion model with randomised origin points, and using the distribution of the observed partial Mantel test's results as a summary statistic within an Approximate Bayesian Computation-Sequential Monte-Carlo (ABC-SMC) algorithm framework. Our results point clearly to a Neolithic expansion route following the Northern Mediterranean, whilst the Southern Mediterranean route could also find support and should be further discussed. The most probable origin points focus on the Xúquer river area.
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[
"Understanding the forces that influence natural variation within and among populations has been a major objective of evolutionary biologists for decades. Motivated by the growth in computational power and data complexity, modern approaches to this question make intensive use of simulation methods. Approximate Bayesian Computation (ABC) is one of these methods. Here we review the foundations of ABC, its recent algorithmic developments, and its applications in evolutionary biology and ecology. We argue that the use of ABC should incorporate all aspects of Bayesian data analysis: formulation, fitting, and improvement of a model. ABC can be a powerful tool to make inferences with complex models if these principles are carefully applied.",
"Approximate Bayesian computation (ABC) methods can be used to evaluate posterior distributions without having to calculate likelihoods. In this paper, we discuss and apply an ABC method based on sequential Monte Carlo (SMC) to estimate parameters of dynamical models. We show that ABC SMC provides information about the inferability of parameters and model sensitivity to changes in parameters, and tends to perform better than other ABC approaches. The algorithm is applied to several well-known biological systems, for which parameters and their credible intervals are inferred. Moreover, we develop ABC SMC as a tool for model selection; given a range of different mathematical descriptions, ABC SMC is able to choose the best model using the standard Bayesian model selection apparatus.",
"Mobility is one of the most important processes shaping spatiotemporal patterns of variation in genetic, morphological, and cultural traits. However, current approaches for inferring past migration episodes in the fields of archaeology and population genetics lack either temporal resolution or formal quantification of the underlying mobility, are poorly suited to spatially and temporally sparsely sampled data, and permit only limited systematic comparison between different time periods or geographic regions. Here we present an estimator of past mobility that addresses these issues by explicitly linking trait differentiation in space and time. We demonstrate the efficacy of this estimator using spatiotemporally explicit simulations and apply it to a large set of ancient genomic data from Western Eurasia. We identify a sequence of changes in human mobility from the Late Pleistocene to the Iron Age. We find that mobility among European Holocene farmers was significantly higher than among European hunter-gatherers both pre- and postdating the Last Glacial Maximum. We also infer that this Holocene rise in mobility occurred in at least three distinct stages: the first centering on the well-known population expansion at the beginning of the Neolithic, and the second and third centering on the beginning of the Bronze Age and the late Iron Age, respectively. These findings suggest a strong link between technological change and human mobility in Holocene Western Eurasia and demonstrate the utility of this framework for exploring changes in mobility through space and time.",
"Lactase persistence (LP) is common among people of European ancestry, but with the exception of some African, Middle Eastern and southern Asian groups, is rare or absent elsewhere in the world. Lactase gene haplotype conservation around a polymorphism strongly associated with LP in Europeans (-13,910 C/T) indicates that the derived allele is recent in origin and has been subject to strong positive selection. Furthermore, ancient DNA work has shown that the--13,910*T (derived) allele was very rare or absent in early Neolithic central Europeans. It is unlikely that LP would provide a selective advantage without a supply of fresh milk, and this has lead to a gene-culture coevolutionary model where lactase persistence is only favoured in cultures practicing dairying, and dairying is more favoured in lactase persistent populations. We have developed a flexible demic computer simulation model to explore the spread of lactase persistence, dairying, other subsistence practices and unlinked genetic markers in Europe and western Asia's geographic space. Using data on--13,910*T allele frequency and farming arrival dates across Europe, and approximate Bayesian computation to estimate parameters of interest, we infer that the--13,910*T allele first underwent selection among dairying farmers around 7,500 years ago in a region between the central Balkans and central Europe, possibly in association with the dissemination of the Neolithic Linearbandkeramik culture over Central Europe. Furthermore, our results suggest that natural selection favouring a lactase persistence allele was not higher in northern latitudes through an increased requirement for dietary vitamin D. Our results provide a coherent and spatially explicit picture of the coevolution of lactase persistence and dairying in Europe."
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Low-molecular weight (LMWPM) metabolites in Parkinson's disease
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Diets rich in (poly)phenols have been associated with positive effects on neurodegenerative disorders, such as Parkinson's disease (PD). Several low-molecular weight (poly)phenol metabolites (LMWPM) are found in the plasma after consumption of (poly)phenol-rich food. It is expected that LMWPM, upon reaching the brain, may have beneficial effects against both oxidative stress and neuroinflammation, and possibly attenuate cell death mechanisms relate to the loss of dopaminergic neurons in PD.
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[
"The results of seven population-based studies were examined separately and pooled to obtain age- and sex-specific estimates of the prevalence of PD. An in-person screening instrument and diagnostic clinical examination were used to detect potential PD cases. The overall prevalence (per 100 population) in persons 65 years of age and older was 1.8, with an increase from 0.6 for those age 65 to 69 years to 2.6 for those 85 to 89 years. There were no sex differences in prevalence of PD.",
"LUHMES cells are conditionally-immortalized non-transformed human fetal cells that can be differentiated to acquire a dopaminergic neuron-like phenotype under appropriate growth conditions. After differentiation by GDNF and cyclic adenosine monophosphate, LUHMES were sensitive to 1-methyl-4-phenylpyridinium (MPP(+)) toxicity at < or =5 microM, but resistant to the parental compound 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The high homogeneity and purity of the cultures allowed the detection of metabolic changes during the degeneration. Cellular ATP dropped in two phases after 24 and 48 h; cellular glutathione (GSH) decreased continuously, paralleled by an increase in lipid peroxidation. These events were accompanied by a time-dependent degeneration of neurites. Block of the dopamine transporter by GBR 12909 or mazindol completely abrogated MPP(+) toxicity. Inhibition of de novo dopamine synthesis by alpha-methyl-l-tyrosine or 3-iodo-l-tyrosine attenuated toxicity, but did not reduce the initial drop in ATP. Inhibition of mixed lineage kinases by CEP1347 completely prevented the MPP(+)-induced loss of viability and intracellular GSH, but failed to attenuate the initial drop of ATP. For the quantitative assessment of neurite degeneration, an automated imaging-based high content screening approach was applied and confirmed the findings made by pharmacological interventions in this study. Our data indicate that inhibition of mitochondrial ATP synthesis is not sufficient to trigger cell death in MPP(+)-treated LUHMES.",
"Glutathione (GSH) deficiency has been identified as an early event in the progression of Parkinson's disease. However, the role of GSH in the etiology and pathogenesis of this neurodegenerative disorder is not well established. The aim of this study is to assess the effect of transient GSH depletion in the substantia nigra pars compacta (SNpc) on neuroinflammation after the injection of a single dose of l-buthionine sulfoximine (BSO) into the SNpc of male Sprague-Dawley rats. The results showed that BSO treatment stimulates microglia (p<0.01) and astroglial response (p<0.01), c-Jun N-terminal kinase and inducible nitric oxide synthase (iNOS) (p<0.001) in the SNpc, accompanied by dopaminergic dysfunction. In addition, high levels of tumor necrosis factor α (p<0.01), interleukins IL-1β p<0.01), IL-6 p<0.001) and nitric oxide p<0.01) were found in the treated animals compared to control groups, while no significant differences were found in IL-10 levels. These results suggest that transient GSH depletion can increase the susceptibility of SNpc to degeneration by promoting an inflammatory response and nitrosative stress, reinforcing the possible role of GSH unbalance, oxygen/nitrogen reactive species and neuroinflammation as causal factors on the degeneration of the SNpc.",
"Dietary supplementation with antioxidant rich foods can decrease the level of oxidative stress in brain regions and can ameliorate age-related deficits in neuronal and behavioral functions. We examined whether short-term supplementation with blueberries might enhance the brain's ability to generate a heat shock protein 70 (HSP70) mediated neuroprotective response to stress. Hippocampal (HC) regions from young and old rats fed either a control or a supplemented diet for 10 weeks were subjected to an in vitro inflammatory challenge (LPS) and then examined for levels of HSP70 at various times post LPS (30, 90 and 240 min). While baseline levels of HSP70 did not differ among the various groups compared to young control diet rats, increases in HSP70 protein levels in response to an in vitro LPS challenge were significantly less in old as compared to young control diet rats at the 30, 90 and 240 min time points. However, it appeared that the blueberry diet completely restored the HSP70 response to LPS in the old rats at the 90 and 240 min times. This suggests that a short-term blueberry (BB) intervention may result in improved HSP70-mediated protection against a number of neurodegenerative processes in the brain. Results are discussed in terms of the multiplicity of the effects of the BB supplementation which appear to range from antioxidant/anti-inflammatory activity to signaling."
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Effects of external feedback on perceptual learning of visual speech during lipreading training
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This study investigated the effects of external feedback on perceptual learning of visual speech during lipreading training with sentence stimuli. The goal was to improve visual-only (VO) speech recognition and increase accuracy of audiovisual (AV) speech recognition in noise. The rationale was that spoken word recognition depends on the accuracy of sublexical (phonemic/phonetic) speech perception; effective feedback during training must support sublexical perceptual learning.
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[
"Auditory training has long been advocated to enhance communication but has never been time or cost-effective. This article describes the concepts underlying the development of a home-based, interactive adaptive computer program designed to engage the adult hearing-impaired listener in the hearing-aid-fitting process, provide listening strategies, build confidence, and address cognitive changes characteristic of the aging process. An investigation using a between-group, within-subject design with pre- and post-test objective and subjective measures was conducted at five clinical sites. Sixty-five subjects were randomly placed into two groups, one receiving LACE (Listening and Communication Enhancement) immediately following baseline testing and one serving as a control for one month and then receiving training as a crossover group. Results showed statistically significant improvements for the trained subjects on all but one of the outcome measures. Barriers facing the widespread implementation of home-based aural rehabilitation are discussed.",
"Auditory training (AT) helps compensate for degradation in the auditory signal. A series of three high-quality training studies are discussed, which include, (i) a randomized controlled trial (RCT) of phoneme discrimination in quiet that trained adults with mild hearing loss (n = 44), (ii) a repeated measures study that trained phoneme discrimination in noise in hearing aid (HA) users (n = 30), and (iii) a double-blind RCT that directly trained working memory (WM) in HA users (n = 57). AT resulted in generalized improvements in measures of self-reported hearing, competing speech, and complex cognitive tasks that all index executive functions. This suggests that for AT related benefits, the development of complex cognitive skills may be more important than the refinement of sensory processing. Furthermore, outcome measures should be sensitive to the functional benefits of AT. For WM training, lack of far-transfer to untrained outcomes suggests no generalized benefits to real-world listening abilities. We propose that combined auditory-cognitive training approaches, where cognitive enhancement is embedded within auditory tasks, are most likely to offer generalized benefits to the real-world listening abilities of adults with hearing loss.",
"Sensory stimuli become easier to detect or distinguish with practice. It is generally assumed that the task-relevant stimulus dimension becomes increasingly more salient as a result of attentively performing the task at a level that is neither too easy nor too difficult. However, here we show improved auditory frequency discrimination following training with physically identical tones that were impossible to discriminate. We also show that learning transfers across tone frequencies and across modalities: training on a silent visuospatial computer game improved thresholds on the auditory discrimination task. We suggest that three processes are necessary for optimal perceptual learning: sensitization through exposure to the stimulus, modality- and dimension-specific attention, and general arousal.",
"A range of tests of everyday attention is described, based on ecologically plausible activities such as searching maps, looking through telephone directories, and listening to lottery number broadcasts. An age-, sex- and IQ-stratified sample of 154 normal participants was given these tests, along with a number of existing tests of attention. The factor structure revealed by this data set matched well contemporary evidence for a set of functionally independent attentional circuits in the brain, and included factors for sustained attention, selective attention, attentional switching and auditory-verbal working memory. The Test of Everyday Attention (TEA), which was developed and standardized on the basis of these subtests, has three parallel forms, high test-retest reliability, and correlates significantly with existing measures of attention. Furthermore, selected subtests successfully discriminate among a number of brain-impaired groups, including closed head injury versus age-matched controls, minimal versus mild Alzheimer's disease, and progressive supranuclear palsy patients versus age-matched controls.",
"The aim of this research forum article was to examine accessibility, use, and adherence to computerized and online interventions for people with hearing loss. Four intervention studies of people with hearing loss were examined: 2 auditory training studies, 1 working memory training study, and 1 study of multimedia educational support. A small proportion (approximately 15%) of participants had never used a computer, which may be a barrier to the accessibility of computer and Internet-based interventions. Computer competence was not a factor in intervention use or adherence. Computer skills and Internet access influenced participant preference for the delivery method of the multimedia educational support program. It is important to be aware of current barriers to computer and Internet-delivered interventions for people with hearing loss. However, there is a clear need to develop and future-proof hearing-related applications for online delivery.",
"Viewing a speaker's articulatory movements substantially improves a listener's ability to understand spoken words, especially under noisy environmental conditions. It has been claimed that this gain is most pronounced when auditory input is weakest, an effect that has been related to a well-known principle of multisensory integration--\"inverse effectiveness.\" In keeping with the predictions of this principle, the present study showed substantial gain in multisensory speech enhancement at even the lowest signal-to-noise ratios (SNRs) used (-24 dB), but it was also evident that there was a \"special zone\" at a more intermediate SNR of -12 dB where multisensory integration was additionally enhanced beyond the predictions of this principle. As such, we show that inverse effectiveness does not strictly apply to the multisensory enhancements seen during audiovisual speech perception. Rather, the gain from viewing visual articulations is maximal at intermediate SNRs, well above the lowest auditory SNR where the recognition of whole words is significantly different from zero. We contend that the multisensory speech system is maximally tuned for SNRs between extremes, where the system relies on either the visual (speech-reading) or the auditory modality alone, forming a window of maximal integration at intermediate SNR levels. At these intermediate levels, the extent of multisensory enhancement of speech recognition is considerable, amounting to more than a 3-fold performance improvement relative to an auditory-alone condition.",
"This study was designed to address individual differences in aided speech understanding among a relatively large group of older adults. The group of older adults consisted of 98 adults (50 female and 48 male) ranging in age from 60 to 86 (mean = 69.2). Hearing loss was typical for this age group and about 90% had not worn hearing aids. All subjects completed a battery of tests, including cognitive (6 measures), psychophysical (17 measures), and speech-understanding (9 measures), as well as the Speech, Spatial, and Qualities of Hearing (SSQ) self-report scale. Most of the speech-understanding measures made use of competing speech and the non-speech psychophysical measures were designed to tap phenomena thought to be relevant for the perception of speech in competing speech (e.g., stream segregation, modulation-detection interference). All measures of speech understanding were administered with spectral shaping applied to the speech stimuli to fully restore audibility through at least 4000 Hz. The measures used were demonstrated to be reliable in older adults and, when compared to a reference group of 28 young normal-hearing adults, age-group differences were observed on many of the measures. Principal-components factor analysis was applied successfully to reduce the number of independent and dependent (speech understanding) measures for a multiple-regression analysis. Doing so yielded one global cognitive-processing factor and five non-speech psychoacoustic factors (hearing loss, dichotic signal detection, multi-burst masking, stream segregation, and modulation detection) as potential predictors. To this set of six potential predictor variables were added subject age, Environmental Sound Identification (ESI), and performance on the text-recognition-threshold (TRT) task (a visual analog of interrupted speech recognition). These variables were used to successfully predict one global aided speech-understanding factor, accounting for about 60% of the variance."
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What is the role of the REG gene in the formation of new islets?
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On 31 October 1920, Sir Frederick Banting, while preparing for a medical student lecture on diabetes, a topic that he knew little about, learned how pancreatic stones resulted in the formation of new islets of Langerhans. He then scribbled down a potential research study of tying off the ducts of the pancreas and collecting the secretions to improve diabetes. These secretions became known as insulin. A century later, 60 different oral medications and 20 different insulins are available for the treatment of diabetes, yet none stimulate new islet formation. One hundred years later, after the discovery of insulin, more than a dozen research teams from around the world have demonstrated that similar studies to Banting's pancreatic ligation studies have resulted in upregulation of the REG gene. There are now more than 200 publications on the role of Reg gene proteins and shorter Reg peptides in initiating new islet formation islet from exocrine pancreatic ducts and protecting against inflammation to islets resulting in islet death. Human data through Phase 2b in both type 1 and 2 diabetes patients with diabetes for an average of 20 years have demonstrated that the use of a shorter bioactive Reg peptide can generate new endogenous insulin production, resulting in significant reductions in hemoglobin A1C and increases in stimulated C-peptide. The observations of Frederick Banting, one century ago, may now lead to the generation of therapeutics that form new islets without the need for transplantation.
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[
"After pancreatectomy, regeneration of acinar and islet elements occurs. Recent data from a model of islet hyperplasia in the hamster suggested that induction of a local pancreatic factor stimulates islet formation. We postulate that the reg gene may be this factor. We studied reg expression during induction of islet growth by using a similar model in the rat. Rats underwent surgical wrapping of the splenic lobe of the pancreas or sham operation. At 2 days ductular proliferation and immunohistochemical evidence of insulin within ductular epithelia were evident in the wrapped lobe. By 14 and 56 days islet number per square millimeter increased 63% and 43%, respectively. Reg mRNA levels, measured by Northern blot analysis with a rat reg cDNA probe (n = 5), increased 300% at 2 days in the wrapped lobe and decreased to that of unwrapped controls by 14 days. In situ hybridization showed localization of reg to the acinar cells. In unwrapped gastric lobes of animals who underwent surgical wrapping of the splenic lobe, no change in islet number per square millimeter or reg gene expression was noted. Surgical wrapping of the pancreatic splenic lobe induces local reg gene expression that is temporally associated with duct cell hyperplasia. This is followed by islet formation within the wrapped lobe. Reg may play a role in the induction of new islets from ductular precursors and in maintenance of normal islet function.",
"The pancreatic regenerating (reg I) gene and its protein product are derived from acinar cells and are mitogenic to beta- and ductal cells. We studied the mechanism of this mitogenic response. ARIP (rat ductal) and RIN 1046-38 (rat beta-) cell lines were exposed to exogenous reg I in culture or transfected with a reg I expression vector. Mitogenesis was assessed by MTS assay (CellTiter 96; Promega, Inc., Madison, WI), and cellular mRNA was subjected to gene microarray analysis to determine signal transduction pathways. Yeast two-hybrid technology was then used to determine intracellular binding of reg I protein. Cells exposed to exogenous reg I showed a mitogenic response; cells transfected with reg I expression vector showed inhibited growth. Microarray analysis of the former showed induction of cyclin pathways and mitogen-activated protein kinase phosphatase (MKP-1); cyclins were inhibited in the latter. Northern analysis confirmed gene induction of cyclin D1 and MKP-1; JNK was phosphorylated prior to expression of both. Yeast two-hybrid analysis confirmed a protein-protein interaction with MKP-1; this was confirmed by immunoprecipitation. Pancreatic-derived cells exposed to reg I grow by activation of signal transduction pathways involving the mitogen-activated protein kinase phosphatases and cyclins, with concomitant induction of MKP-1. However, high intracellular levels of reg I lead to decreased growth, likely via a binding to and inactivation of MKP-1. Inhibition of cell growth, and possible induction of apoptosis, may lead to differentiation of these cells to other cell types.",
"Human regenerating (Reg) gene products are regionally expressed by gut-derived tissues, and are markedly up-regulated in cancer and in diseases characterized by mucosal injury. We recently identified Reg IV, a novel regenerating gene product that is uniquely expressed by the normal distal gastrointestinal mucosa. The function remains poorly understood due to the lack of significant purified Reg IV for biochemical and functional studies. Recombinant human Reg IV was efficiently expressed under the control of the AOX1 gene promoter in Pichia pastoris using the MutS strain KM71H. We describe the unique conditions that are required for efficient production of Reg IV protein in high density fermentation. Optimal protein expression was obtained by reduction of the fermentation temperature and addition of casamino acids as a supplemental nitrogen source and to minimize the activity of yeast produced proteases. Recombinant Reg IV protein was purified by tangential flow filtration and reverse phase chromatography. The purified protein was characterized by amino terminus sequence analysis and MALDI-TOFMS showing that the engineered protein had the expected sequence and molecular weight without secondary modification. Recombinant Reg IV was further characterized by specific monoclonal and polyclonal reagents that function for Western blot analysis and for immunolocalization studies.",
"Reg IV, the latest member of the regenerating gene family, has been documented in different tissues of human and rat, such as the colon, small intestine, stomach, and pancreas. Expression of Reg IV gene in distinct cell types has been correlated with its various functions in regeneration, cell growth and survival, proliferation and differentiation, cell adhesion, and resistance to apoptosis. However, there was no evidence to show whether the Reg IV protein is present in the reproductive system of normal rat. The aim of this study was to reveal the expression patterns of Reg IV in rat ovary and uterus. The expression of Reg IV was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot at mRNA and protein levels, respectively. The localization of Reg IV protein within rat ovary and uterus was investigated by immunohistochemistry (IHC). Our results showed that the expression of Reg IV in ovary was significantly higher than that in the uterus. The strong immunoreactive signals of Reg IV was observed in granulosa cells and oocytes of ovarian follicles, corpus luteum, and interstitial cells in rat ovary; only weak signals were detected in luminal and gland epithelium of rat endometrium. These findings first demonstrate the expression of Reg IV in ovary and uterus of the healthy rat at both mRNA and protein levels. It provides an evidence of Reg IV expression in rat reproductive system, which may help elucidate a potential role in cell growth and proliferation of reproductive system.",
"The regenerating islet-derived (REG) gene family encodes a group of proteins highly expressed in several human pathologies, many of which are associated with epithelial inflammation. All human family members, namely REG1A, REG1B, REG3A and REG4, are closely related in genomic sequence and all are part of the c-type lectin superfamily. REGs are highly expressed during inflammatory bowel disease (IBD)-related colonic inflammation and the in vivo expression pattern of REG1A and REG4 has been localised by using immunohistochemistry. However, the function of the encoded proteins is largely unknown and the cellular localisation of REG expression during colonic inflammation has not been described. Therefore, we have used in situ hybridisation to demonstrate the cellular localisation of REG expression in healthy and diseased colonic mucosa. Samples drawn from an IBD cohort including both inflamed and un-inflamed colonic mucosa are described, as are samples from non-IBD inflammation and healthy controls. Immunohistochemistry against known cell-type markers on serial sections has localised the expression of REGs to metaplastic Paneth cells (REG1A, REG1B and REG3A) and enteroendocrine cells (REG4), with a marked expansion of expression during inflammation. The group of REGs can, based on gene expression patterns, be divided into at least two groups; REG1A, REG1B and REG3A with their expression focused in the crypt base spreading from Paneth cells and REG4 being more highly expressed towards the luminal face. This exploration of expression pattern forms provides the background for further exploration of REG function in the intestine.",
"Pancreatic ductal adenocarcinoma (PDAC) offers an optimal model for discovering \"druggable\" molecular pathways that participate in inflammation-associated cancer development. Chronic pancreatitis, a common prolonged inflammatory disease, behaves as a well-known premalignant condition that contributes to PDAC development. Although the mechanisms underlying the pancreatitis-to-cancer transition remain to be fully elucidated, emerging evidence supports the hypothesis that the actions of proinflammatory mediators on cells harboring Kras mutations promote neoplastic transformation. Recent elegant studies demonstrated that the IL17 pathway mediates this phenomenon and can be targeted with antibodies, but the downstream mechanisms by which IL17 functions during this transition are currently unclear. In this study, we demonstrate that IL17 induces the expression of REG3β, a well-known mediator of pancreatitis, during acinar-to-ductal metaplasia and in early pancreatic intraepithelial neoplasia (PanIN) lesions. Furthermore, we found that REG3β promotes cell growth and decreases sensitivity to cell death through activation of the gp130-JAK2-STAT3-dependent pathway. Genetic inactivation of REG3β in the context of oncogenic Kras-driven PDAC resulted in reduced PanIN formation, an effect that could be rescued by administration of exogenous REG3β. Taken together, our findings provide mechanistic insight into the pathways underlying inflammation-associated pancreatic cancer, revealing a dual and contextual pathophysiologic role for REG3β during pancreatitis and PDAC initiation.",
"Regenerating islet-derived (Reg) proteins have emerged as multifunctional agents with pro-proliferative, anti-apoptotic, differentiation-inducing and bactericidal properties. Over the last 40 years since first discovered, Reg proteins have been implicated in a gamut of maladies including diabetes, various types of cancer of the digestive tract, and Alzheimer disease. Surprisingly though, a consensus is still absent on the regulation of their expression, and molecular underpinning of their function. Here, we provide a critical appraisal of recent findings in the field of Reg protein biology. Specifically, the structural characteristics are reviewed particularly in connection with established or purported functions of different members of the Reg family. Moreover, Reg expression patterns in different tissues both under normal and pathophysiological conditions are summarized. Putative receptors and cascades reported to relay Reg signaling inciting cellular responses are presented aiming at a better appreciation of the biological activities of the distinct Reg moieties. Challenges are also discussed that have hampered thus far the rapid progress in this field such as the use of non-standard nomenclature for Reg molecules among various research groups, the existence of multiple Reg members with significant degree of homology and possibly compensatory modes of action, and the need for common assays with robust readouts of Reg activity. Coordinated research is warranted going forward, given that several research groups have independently linked Reg proteins to diseased states and raised the possibility that these biomolecules can serve as therapeutic targets and biomarkers.",
"The Reg3 peptides INGAP-PP and human Reg3α/β (HIP) have been hypothesized to stimulate β-cell neogenesis in the pancreas. Administration of INGAP-PP has been shown to cause an increase in β-cell mass in multiple animal models, reverse streptozotocin (STZ) induced diabetes in mice and reduces HbA1c levels in type 2 diabetic humans. In this study, we have examined the ability of the INGAP-PP and HIP peptides to induce β-cell formation in vivo in normal mice through short-term administration of the peptides. We assessed the peptides ability to induce an increase in extra-islet insulin-positive cell clusters by looking at β-cell number by point counting morphometry on pancreata that had been randomized using the smooth fractionator principle in non-diabetic NMRI mice after short-term injections of the peptides (5 d). Five day continuous BrdU labeling was used to determine if the new β-cells were derived from replicating β-cells. Real time quantitative RT-PCR and immuno-histochemistry was used to analyze changes in pancreatic transcription factor expression. A 1.5- to 2-fold increase in the volume of small extra-islet insulin-positive clusters post 5 d treatment with INGAP-PP and HIP as compared with mice treated with a non-peptide control or scrambled peptide (p<0.05) (n = 7) was found. Five day continuous BrdU infusion during the 5 d period showed little or no incorporation in islets or small insulin clusters. Five days of treatment with INGAP-PP or HIP, showed a tendency toward increased levels of pancreatic progenitor markers such as Ngn3, Nkx6.1, Sox9 and Ins. These are the first studies to compare and indicate that the human Reg3 α/β (HIP) peptide has similar bioactivity in vivo as INGAP by causing formation of small β-cell clusters in extra-islet pancreatic tissue after only 5 d of treatment. Upregulation of pancreatic transcription factors may be part of the mechanism of action.",
"We originally isolated the HIP/PAP gene in a differential screen of a human hepatocellular carcinoma cDNA library. This gene is expressed at high levels in 25% of primary liver cancers but not in nontumorous liver. HIP/PAP belongs to the family of C-type lectins and acts as an adhesion molecule for hepatocytes. In normal adult human tissues, HIP/PAP expression is found in pancreas (exocrine and endocrine cells) and small intestine (Paneth and neuroendocrine cells). In order to gain insight into the possible role of HIP/PAP in vivo, we have investigated the pattern of HIP/PAP expression in the developing postimplantation mouse embryo by in situ hybridization. Detailed analysis of developing mouse embryos revealed that HIP/PAP gene exhibits a restricted expression pattern during development. Thus, HIP/PAP transcripts are first observed within the nervous system from day 14.5 onwards in trigeminal ganglia, dorsal root ganglia, and spinal cord where it appears to be an early specific marker of a subpopulation of motor neurons. At laster stages, HIP/PAP transcripts were detected in intestine and pancreas at day 16.5 but not in embryonic liver. This highly restricted expression pattern suggests that HIP/PAP might participate in neuronal as well as intestinal and pancreatic cell development.",
"The Wnt/beta-catenin signaling pathway is activated in many human hepatocellular carcinomas (HCC). We tried to identify the genes involved in carcinogenesis and progression of HCC with beta-catenin mutations. We used PCR-based subtractive hybridization to compare gene expression between malignant and benign components of a human HCC occurring in pre-existing adenoma activated for beta-catenin. Two of the genes identified belong to the Regenerating gene (REG) family. They encode the Regenerating islet-derived 3 alpha (REG3A/HIP/PAP/REG-III) and 1 alpha (REG1A) proteins, both involved in liver and pancreatic regeneration and proliferation. Using siRNA directed against beta-catenin, we demonstrated that REG3A is a target of beta-catenin signaling in Huh7 hepatoma cells. The upregulation of REG3A and REG1A expression is significantly correlated to the beta-catenin status in 42 HCC and 28 hepatoblastomas characterized for their beta-catenin status. Thus, we report strong evidence that both genes are downstream targets of the Wnt pathway during liver tumorigenesis."
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In situ cryo-electron microscopy of the tip link in mouse stereocilia
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Mechanosensory transduction (MT), the conversion of mechanical stimuli into electrical signals, underpins hearing and balance and is carried out within hair cells in the inner ear. Hair cells harbor actin-filled stereocilia, arranged in rows of descending heights, where the tips of stereocilia are connected to their taller neighbors by a filament composed of protocadherin 15 (PCDH15) and cadherin 23 (CDH23), deemed the 'tip link.' Tension exerted on the tip link opens an ion channel at the tip of the shorter stereocilia, thus converting mechanical force into an electrical signal. While biochemical and structural studies have provided insights into the molecular composition and structure of isolated portions of the tip link, the architecture, location, and conformational states of intact tip links, on stereocilia, remains unknown. Here, we report in situ cryo-electron microscopy imaging of the tip link in mouse stereocilia. We observe individual PCDH15 molecules at the tip and shaft of stereocilia and determine their stoichiometry, conformational heterogeneity, and their complexes with other filamentous proteins, perhaps including CDH23. The PCDH15 complexes occur in clusters, frequently with more than one copy of PCDH15 at the tip of stereocilia, suggesting that tip links might consist of more than one copy of PCDH15 complexes and, by extension, might include multiple MT complexes.
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[
"We have developed a computer software package, IMOD, as a tool for analyzing and viewing three-dimensional biological image data. IMOD is useful for studying and modeling data from tomographic, serial section, and optical section reconstructions. The software allows image data to be visualized by several different methods. Models of the image data can be visualized by volume or contour surface rendering and can yield quantitative information.",
"The vertebrate inner ear, responsible for hearing and balance, is able to sense minute mechanical stimuli originating from an extraordinarily broad range of sound frequencies and intensities or from head movements. Integral to these processes is the tip-link protein complex, which conveys force to open the inner-ear transduction channels that mediate sensory perception. Protocadherin-15 and cadherin-23, two atypically large cadherins with 11 and 27 extracellular cadherin (EC) repeats, are involved in deafness and balance disorders and assemble as parallel homodimers that interact to form the tip link. Here we report the X-ray crystal structure of a protocadherin-15 + cadherin-23 heterotetrameric complex at 2.9-Å resolution, depicting a parallel homodimer of protocadherin-15 EC1-3 molecules forming an antiparallel complex with two cadherin-23 EC1-2 molecules. In addition, we report structures for 10 protocadherin-15 fragments used to build complete high-resolution models of the monomeric protocadherin-15 ectodomain. Molecular dynamics simulations and validated crystal contacts are used to propose models for the complete extracellular protocadherin-15 parallel homodimer and the tip-link bond. Steered molecular dynamics simulations of these models suggest conditions in which a structurally diverse and multimodal protocadherin-15 ectodomain can act as a stiff or soft gating spring. These results reveal the structural determinants of tip-link-mediated inner-ear sensory perception and elucidate protocadherin-15's structural and adhesive properties relevant in disease.",
"Much is known about the mechanotransducer (MT) channels mediating transduction in hair cells of the vertrbrate inner ear. With the use of isolated preparations, it is experimentally feasible to deliver precise mechanical stimuli to individual cells and record the ensuing transducer currents. This approach has shown that small (1-100 nm) deflections of the hair-cell stereociliary bundle are transmitted via interciliary tip links to open MT channels at the tops of the stereocilia. These channels are cation-permeable with a high selectivity for Ca(2+); two channels are thought to be localized at the lower end of the tip link, each with a large single-channel conductance that increases from the low- to high-frequency end of the cochlea. Ca(2+) influx through open channels regulates their resting open probability, which may contribute to setting the hair cell resting potential in vivo. Ca(2+) also controls transducer fast adaptation and force generation by the hair bundle, the two coupled processes increasing in speed from cochlear apex to base. The molecular intricacy of the stereocilary bundle and the transduction apparatus is reflected by the large number of single-gene mutations that are linked to sensorineural deafness, especially those in Usher syndrome. Studies of such mutants have led to the discovery of many of the molecules of the transduction complex, including the tip link and its attachments to the stereociliary core. However, the MT channel protein is still not firmly identified, nor is it known whether the channel is activated by force delivered through accessory proteins or by deformation of the lipid bilayer.",
"The fibroblast growth factor FGF21 was labeled with molecularly defined gold nanoparticles (AuNPs), applied to human adipocytes, and imaged by cryo-electron tomography (cryo-ET). Most AuNPs were in pairs about 80 Å apart, on the outer cell surface. Pairs of AuNPs were also abundant inside the cells in clathrin-coated vesicles and endosomes. AuNPs were present but no longer paired in multivesicular bodies. FGF21 could thus be tracked along the endocytotic pathway. The methods developed here to visualize signaling coupled to endocytosis can be applied to a wide variety of cargo and may be extended to studies of other intracellular transactions.",
"Mouse chromosome 10 harbors several loci associated with hearing loss, including waltzer (v), modifier-of deaf waddler (mdfw) and Age-related hearing loss (Ahl). The human region that is orthologous to the mouse 'waltzer' region is located at 10q21-q22 and contains the human deafness loci DFNB12 and USH1D). Numerous mutations at the waltzer locus have been documented causing erratic circling and hearing loss. Here we report the identification of a new gene mutated in v. The 10.5-kb Cdh23 cDNA encodes a very large, single-pass transmembrane protein, that we have called otocadherin. It has an extracellular domain that contains 27 repeats; these show significant homology to the cadherin ectodomain. In v(6J), a GT transversion creates a premature stop codon. In v(Alb), a CT exchange generates an ectopic donor splice site, effecting deletion of 119 nucleotides of exonic sequence. In v(2J), a GA transition abolishes the donor splice site, leading to aberrant splice forms. All three alleles are predicted to cause loss of function. We demonstrate Cdh23 expression in the neurosensory epithelium and show that during early hair-cell differentiation, stereocilia organization is disrupted in v(2J) homozygotes. Our data indicate that otocadherin is a critical component of hair bundle formation. Mutations in human CDH23 cause Usher syndrome type 1D and thus, establish waltzer as the mouse model for USH1D.",
"Bispecific antibodies have become important formats for therapeutic discovery. They allow for potential synergy by simultaneously engaging two separate targets and enable new functions that are not possible to achieve by using a combination of two monospecific antibodies. Antagonistic antibodies dominate drug discovery today, but only a limited number of agonistic antibodies (i.e. those that activate receptor signaling) have been described. For receptors formed by two components, engaging both of these components simultaneously may be required for agonistic signaling. As such, bispecific antibodies may be particularly useful in activating multicomponent receptor complexes. Here, we describe a biparatopic (i.e. targeting two different epitopes on the same target) format that can activate the endocrine fibroblast growth factor (FGF) 21 receptor (FGFR) complex containing β-Klotho and FGFR1c. This format was constructed by grafting two different antigen-specific VH domains onto the VH and VL positions of an IgG, yielding a tetravalent binder with two potential geometries, a close and a distant, between the two paratopes. Our results revealed that the biparatopic molecule provides activities that are not observed with each paratope alone. Our approach could help address the challenges with heterogeneity inherent in other bispecific formats and could provide the means to adjust intramolecular distances of the antibody domains to drive optimal activity in a bispecific format. In conclusion, this format is versatile, is easy to construct and produce, and opens a new avenue for agonistic antibody discovery and development.",
"In mammals, environmental sounds stimulate the auditory receptor, the cochlea, via vibrations of the stapes, the innermost of the middle ear ossicles. These vibrations produce displacement waves that travel on the elongated and spirally wound basilar membrane (BM). As they travel, waves grow in amplitude, reaching a maximum and then dying out. The location of maximum BM motion is a function of stimulus frequency, with high-frequency waves being localized to the \"base\" of the cochlea (near the stapes) and low-frequency waves approaching the \"apex\" of the cochlea. Thus each cochlear site has a characteristic frequency (CF), to which it responds maximally. BM vibrations produce motion of hair cell stereocilia, which gates stereociliar transduction channels leading to the generation of hair cell receptor potentials and the excitation of afferent auditory nerve fibers. At the base of the cochlea, BM motion exhibits a CF-specific and level-dependent compressive nonlinearity such that responses to low-level, near-CF stimuli are sensitive and sharply frequency-tuned and responses to intense stimuli are insensitive and poorly tuned. The high sensitivity and sharp-frequency tuning, as well as compression and other nonlinearities (two-tone suppression and intermodulation distortion), are highly labile, indicating the presence in normal cochleae of a positive feedback from the organ of Corti, the \"cochlear amplifier.\" This mechanism involves forces generated by the outer hair cells and controlled, directly or indirectly, by their transduction currents. At the apex of the cochlea, nonlinearities appear to be less prominent than at the base, perhaps implying that the cochlear amplifier plays a lesser role in determining apical mechanical responses to sound. Whether at the base or the apex, the properties of BM vibration adequately account for most frequency-specific properties of the responses to sound of auditory nerve fibers."
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Convergence insufficiency: a review of the literature.
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Convergence insufficiency (CI) is a common binocular vision (BV) disorder characterized by difficulty in maintaining motor fusion at near, which affects approximately 7.5 percent of the population. Diagnostic criteria for the disorder are inconsistent, ranging from one to many clinical signs. Methodology for clinical tests is inconsistent in measurement technique, visual targets, required repetitions, and normative values. This manuscript demonstrates the inconsistencies amongst published studies, and highlights the importance of consistent clinical diagnostic signs, measurement techniques, visual targets, and cut-off criteria. For each clinical sign, the recommended methodology for the procedure is described. Several studies do not take age into account when diagnosing CI in their cohorts. As such, the review emphasizes changes in diagnostic signs with age. This manuscript highlights the need for consistent and clear procedures and diagnostic criteria amongst clinicians and provides the basis for future studies in terms of diagnostic testing required for CI of varying age groups.
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[
"Few studies have examined the prevalence of convergence insufficiency (CI) in a school-based population in Sudan. This study sought to determine the prevalence of CI and its related clinical characteristics among Sudanese secondary school students. A descriptive cross-sectional survey was conducted in Central Khartoum North, Sudan, in the academic year 2013/2014. A total of 4211 secondary school students, with a mean age of 15.5 ± 2.5 years, underwent complete eye examination, and were screened for symptoms associated with near work. Near and distance heterophoria was measured with the alternate cover test using a prism bar; near point of convergence (NPC) and positive fusional vergence (PFV) at near were determined. Of the 4211 students screened, 329 (7.8%) were diagnosed with CI. Of these, 173 (52.6%) students were male and 156 (47.4%) were female; there was no significant relationship between sex and CI (P > 0.05). Standard schools had a higher prevalence of CI (43%) than geographic schools (36%) and there was a significant association between CI and the type of school (P < 0.05). In most of the students (78.42%), CI was due to both remote NPC and decreased PFV; in 20.36% of the students, CI was due to remote NPC only, and in very few students (1.22%), it was due to decreased PFV only. These findings suggest that CI is prevalent in the secondary school population in Central Khartoum North, Sudan.",
"Previous reports concerning the effect of age on the AC/A ratio have been equivocal. Therefore, the present study investigated both the stimulus (AC/As) and response (AC/Ar) ratios using a subjective haploscope-optometer in a relatively large sample of subjects (n = 42) over a wide range of ages (22-65 years). The AC/As showed a small but significant decrease with age (approximately 0.04 delta/D/year). When the older subjects (> 45 years of age) were excluded, however, there was no systematic age effect. The AC/Ar exhibited a small but significant increase with age (approximately 0.08 delta/D/year) for subjects under 45 years of age. However, when the older pre-presbyopes and younger presbyopes (35-44 years of age) were excluded, there was no systematic age effect. In subjects 45 years of age and older, the AC/Ar could not be reliably assessed. This was attributed to physiological and instrumentation noise as a result of the minimal change in accommodative response. In the mid-aged subjects (35-44 years of age), the apparent increase in AC/Ar with age was speculated to be due to neural adaptation of the crosslink gain from the accommodative to the vergence system and/or slight intrusion into the upper non-linear response region of accommodation with the measurements. The finding of AC/Ar constancy with age when mid-aged pre-presbyopes and early presbyopes were excluded supports the non-linearity hypothesis, and thus there appears to be no real change in AC/Ar with increased age. The results support the Hess-Gullstrand theory of presbyopia.",
"Convergence insufficiency represents a significant problem in optometric practice. The percentage of cases in a rural area was compared with that in a previous sample from an urban area, with respect to the general incidence, and the effects of age and taking of medicines as an indicator of health. The percentage of convergence insufficiency in the rural area was about half that in the urban area, and the percentage taking medicines was about two-thirds. The incidence of convergence insufficiency increased with age in both populations. When both samples were taken together, the incidence of convergence insufficiency was three times greater in those taking medicines.",
"Although it is well known that age affects visual function, we still have much to learn about the impact of aging on binocular vision. Our aim was to establish distance heterophoria and distance fusional vergence ranges in a large non-clinical population of wide age range, to provide normal values for comparisons. A cross-sectional study was performed on 271 non-clinical subjects stratified into six age groups. Distance heterophoria measurements were obtained using the von Graefe technique and distance horizontal fusional vergence ranges using Risley rotary prisms in each subject, with best spectacle correction. The mean heterophoria value for distance fixation was exophoric except for the 71-80 year age group, which was esophoric. No effects of age were observed on distance heterophoria and distance base-in and base-out break means. However, a significant effect of age was noted on base-in recovery and base-out recovery. Between the youngest and oldest age groups, mean base-in recovery decreased by 2.5 Delta and mean base-out recovery by 3.3 Delta. Distance base-in and base-out recovery means diminished significantly with age such that recovery values fell outside the norm in subjects in the 61-70 year age group and older. This study establishes statistical normal values for distance heterophoria and fusional vergence in a non-clinical population. Mean values such as these are valuable tools for identifying subjects with far binocular anomalies.",
"The purpose of this study was to help determine the most appropriate target to be used for the assessment of the nearpoint of convergence, normative data for the break and recovery in adults, and the diagnostic value of the red-glass modification and repetition of the nearpoint of convergence. A total of 175 subjects with normal binocular vision and 38 subjects with convergence insufficiency were evaluated. The nearpoint of convergence was measured three ways, with an accommodative target, a penlight, and a penlight with red and green glasses. The nearpoint of convergence was also measured using a penlight for 10 repetitions. Results suggest a clinical cutoff value of 5 cm for the nearpoint of convergence break and 7 cm for the nearpoint of convergence recovery with either an accommodative target or a penlight with red and green glasses. This study establishes normative data for the nearpoint of convergence break and recovery in the adult population and supports the value of various test modifications when other testing is equivocal.",
"This study compared the near phoria measurement using the Bernell muscle balance card with and without prism neutralization, using both trial frame and phoropter correction, and compared with the conventional Maddox rod method. Forty young normal Chinese adults had their near phoria measured with trial frame correction using the conventional muscle balance card method (method 1). Any deviation was compensated with a prism bar as an alternative approach (method 2). The conventional Maddox rod method (method 3) was also carried out for comparison. These three methods were repeated with phoropter correction and considered as methods 4, 5 and 6. The phorias obtained from these six methods were not significantly different from each other (repeated measures anova, p > 0.05). More than half of the subjects were exophoric. Although the difference in phoria was not significant, phoria measurement using phoropter correction yielded a greater coefficient of variation. Near phoria measurement using the muscle balance card conducted with trial frame correction was less variable, and was also more natural and similar to a real reading situation. The use of prism for compensation did not affect the phoria results. Exophoria seems to be more common than esophoria in young Chinese adults.",
"Subjective measurement of the oculomotor deviation at both distance and near is a standard test in optometric practice. A number of procedures are available to the practitioner, and previous studies have demonstrated differences in the repeatability of many of the techniques. However, it is unclear whether testing the subject through a phoropter or in free space (trial frame) will alter the oculomotor deviation. Distance and near heterophoria was measured in 60 visually normal subjects between 20 and 34 years of age using the Von Graefe (VG), Maddox Rod (MR), and Modified Thorington (MT) procedures. The deviation was assessed for viewing distances of 6 m and 0.40 m using both a phoropter and trial frame. To examine the repeatability of each technique, the deviation was measured on two separate occasions for each procedure, with the two sessions being separated by at least 24 hours. The mean vertical deviations at both distance and near were extremely close to orthophoria, and only minimal variation was observed. However, the MR procedure, when used with a trial frame, gave the best repeatability for measuring vertical deviations both at distance and near. Regarding horizontal deviations, the repeatability was better for all three procedures when using a trial frame compared with the phoropter. The best repeatability at distance and near was observed when the MT and MR techniques, respectively, were used with the trial frame. Consistent with previous reports, the VG procedure had poor repeatability, especially when used with the phoropter, and the mean findings showed a greater exo deviation when compared with the other two techniques. Testing oculomotor deviations in free space provides a more repeatable response than when using a phoropter. Accordingly, we recommend that subjective measurements of heterophoria in the clinical setting can best be quantified using either the MR or MT techniques in free space."
] |
Zinc transporter ZnT2 is involved in intestinal dysbiosis and mucosal inflammation
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Loss of Paneth cell (PC) function is implicated in intestinal dysbiosis, mucosal inflammation, and numerous intestinal disorders, including necrotizing enterocolitis (NEC). Studies in mouse models show that zinc transporter ZnT2 (
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[
"Nucleotides are low molecular weight biological molecules key to biochemical processes. Sources include de novo synthesis, recovery via salvage mechanisms, and dietary intakes. Although endogenous production serves as the main nucleotide source, evidence suggests that exogenous sources are essential to immune competence, intestinal development, and recovery. Dietary nucleotides serve a marked role in rapidly proliferating cells where they are necessary for optimal function. Accordingly, dietary nucleotides are deemed conditionally essential in the presence of various physiological stresses, including growth and development, recovery from injury, infection, and certain disease states. Clinical studies that evaluated nutrition formulations of nucleotides in combination with other specific nutrient substances demonstrated improved clinical outcomes in patients characterized as critically ill, injured, immune suppressed, or with chronic gastrointestinal conditions. However, conclusions regarding specific benefits of nucleotides are limited. Scientific substantiation of nucleotide supplementation in infant formula has been reported to improve the maturation and development of the intestinal tract as well as immune function. All medical nutrition products except for one immune-modulating formulation are devoid of nucleotides. In an effort to build on this, the current review presents the data to support potential clinical applications for nucleotides in enteral nutrition that may contribute to improved outcomes in physiologically stressed patients.",
"Greater than 68% of young infants are exposed to dietary zinc (Zn) levels that are higher than the Tolerable Upper Intake Limit. However, the consequences of excess dietary Zn during early life on intestinal function and host-microbe interactions are unknown. Neonatal mice are gavaged with 100 Zn µg d-1 from postnatal day (PN) 2 through PN10 and indices of intestinal function and host-microbe interactions are compared to unsupplemented mice. Excess dietary Zn causes oxidative stress, increases goblet cell number and mucus production, and are associated with increased intestinal permeability and systemic inflammation. Over 900 genes are differentially expressed; 413 genes display a fold-change >1.60. The Gene Ontology Biological processes most significantly affected include biological adhesion, the immune system, metabolic processes, and response to stimulus. Key genes most highly and significantly upregulated include ALDH2, MT1, TMEM6, CDK20, and COX62b, while CALU, ST3GAL4, CRTC2, SLC28A2, and COMMA1 are downregulated. These changes are associated with a microbiome enriched in pathogenic taxa including Pseudomonadales and Campylobacter, and greater expression of bacterial stress response genes. Excess dietary Zn may have unforeseen influences on epithelial signaling pathways, barrier function, and luminal ecology in the intestine that may have long-term consequences on intestinal health.",
"Homology modelling has matured into an important technique in structural biology, significantly contributing to narrowing the gap between known protein sequences and experimentally determined structures. Fully automated workflows and servers simplify and streamline the homology modelling process, also allowing users without a specific computational expertise to generate reliable protein models and have easy access to modelling results, their visualization and interpretation. Here, we present an update to the SWISS-MODEL server, which pioneered the field of automated modelling 25 years ago and been continuously further developed. Recently, its functionality has been extended to the modelling of homo- and heteromeric complexes. Starting from the amino acid sequences of the interacting proteins, both the stoichiometry and the overall structure of the complex are inferred by homology modelling. Other major improvements include the implementation of a new modelling engine, ProMod3 and the introduction a new local model quality estimation method, QMEANDisCo. SWISS-MODEL is freely available at https://swissmodel.expasy.org.",
"Serine proteases are extensively known to play key roles in many physiological processes. However, their dysregulation is often associated to several diseases including inflammatory bowel diseases (IBD). Here, we used specific substrates to monitor fecal protease activities in a large cohort of healthy and IBD patients. Of interest, serine protease activity was 10-fold higher in IBD fecal samples compared to healthy controls. Moreover, functional analysis of these fecal proteolytic activities revealed that the most increased activities are trypsin-like, elastase-like and cathepsin G-like. We also show for the first time, an increase of proteinase 3-like activity in these samples compared to controls. Results presented here will guide further investigations to better understand the relevance of these peptidases in IBD.",
"SWISS-MODEL Repository (SMR) is a database of annotated 3D protein structure models generated by the automated SWISS-MODEL homology modeling pipeline. It currently holds >400 000 high quality models covering almost 20% of Swiss-Prot/UniProtKB entries. In this manuscript, we provide an update of features and functionalities which have been implemented recently. We address improvements in target coverage, model quality estimates, functional annotations and improved in-page visualization. We also introduce a new update concept which includes regular updates of an expanded set of core organism models and UniProtKB-based targets, complemented by user-driven on-demand update of individual models. With the new release of the modeling pipeline, SMR has implemented a REST-API and adopted an open licencing model for accessing model coordinates, thus enabling bulk download for groups of targets fostering re-use of models in other contexts. SMR can be accessed at https://swissmodel.expasy.org/repository.",
"The discovery of Helicobacter pylori sparked a revolution in the understanding and management of peptic ulcer disease and gastric cancer. Other Helicobacter species are recognized as important pathogenic agents in colitic diseases of rodents and primates, in particular Helicobacter bilis, Helicobacter fennelliae, Helicobacter hepaticus and Helicobacter trogontum. Helicobacter bilis and H. hepaticus are now routinely used to initiate rodent models of inflammatory bowel disease (IBD), particularly in immunocompromised hosts. Molecular evidence exists linking various non-pylori Helicobacter spp. with human IBD; however, attempts to culture organisms in this disease cohort have proved unsuccessful to date. Attributing causation has therefore proved elusive. Seven enterohepatic, non-pylori Helicobacter organisms have been successfully cultured from humans, namely Helicobacter canadensis, Helicobacter canis, Helicobacter cinaedi, H. fennelliae, Helicobacter pullorum, Helicobacter winghamensis and Helicobacter sp. flexispira taxon 8 (now classified as H. bilis). Of these, H. cinaedi and H. fennelliae are the closest to fulfilling Koch's postulates as causative agents in homosexual proctitis. The possibility that novel Helicobacter organisms have a role in the initiation of human IBD warrants further consideration and targeted investigations.",
"The prevalence of nephrocalcinosis (NC) in preterm neonates in recent reports is 7-41%. The wide range in prevalence is a consequence of different study populations and ultrasound equipment and criteria, in addition to a moderate interobserver variation. NC in preterm neonates has a multifactorial aetiology, consisting of low gestational age and birth weight, often in combination with severe respiratory disease, and occurs as a result of an imbalance between stone-promoting and stone-inhibiting factors. A limited number of histological studies suggest that calcium oxalate crystals play an important role in NC in premature neonates. In 85% of children resolution of NC occurs in the first years of life. Prematurity, per se, is associated with high blood pressure, relatively small kidneys, and (distal) tubular dysfunction. In addition, NC in preterm neonates can have long-term sequelae for glomerular and tubular function. Long-term follow-up of blood pressure and renal function of prematurely born children, especially with neonatal NC, is recommended. Prevention of NC with (low) oral doses of citrate has not resulted in a significant decrease in the prevalence of NC; a higher citrate dosage deserves further study. Future research pertaining to prevention of NC in preterm neonates is crucial."
] |
Primary EGFR T790M mutation with EGFR exon 19 del/DCTN1-ALK translocation in a patient with lung cancer brain metastases: a case report and review of the literature.
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The echinoderm microtubule associated protein-like 4 gene (EML4) encodes the predominant anaplastic lymphoma kinase (ALK) fusion partner in non-small-cell lung cancer (NSCLC); however, the dynactin subunit 1 (DCTN1)-ALK rearrangement is extremely rare. The co-occurrence of primary epidermal growth factor receptor (EGFR) T790M mutation with EGFR exon 19 deletion (del) in patients with NSCLC is uncommon. Here we report a female lung adenocarcinoma patient with brain metastases and possible coexistence of primary EGFR T790M mutation/EGFR exon 19 del/DCTN1-ALK translocation. The patient received multiline treatment including chemotherapy, antivascular, and targeted therapies. To overcome developed resistance to chemotherapy or targeted therapy to prolong overall survival, the patient's circulating tumor DNA (ctDNA) was dynamically monitored. The patient responded to successive osimertinib and alectinib treatment, and alectinib achieved a nearly complete response for lung and brain lesions after she acquired osimertinib resistance. Furthermore, we summarize 22 published cases of patients with lung adenocarcinoma with concurrent EGFR mutation and ALK rearrangement, including details of clinical characteristics, natural history, and pertinent therapy of this uncommon tumor subtype. This literature review shows that EGFR inhibition was an indispensable aspect of the treatment of patients with EGFR/ALK co-alterations in the pre-alectinib era and that ALK inhibition with crizotinib did not show more eye-catching therapeutic results. Considering the effectiveness achieved by alectinib, this case study provides a new perspective for the treatment of lung cancer brain metastasis patients with concurrent EGFR/ALK mutations.
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[
"Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKI), including crizotinib, are effective treatments in preclinical models and in cancer patients with ALK-translocated cancers. However, their efficacy will ultimately be limited by the development of acquired drug resistance. Here we report two mechanisms of ALK TKI resistance identified from a crizotinib-treated non-small cell lung cancer (NSCLC) patient and in a cell line generated from the resistant tumor (DFCI076) as well as from studying a resistant version of the ALK TKI (TAE684)-sensitive H3122 cell line. The crizotinib-resistant DFCI076 cell line harbored a unique L1152R ALK secondary mutation and was also resistant to the structurally unrelated ALK TKI TAE684. Although the DFCI076 cell line was still partially dependent on ALK for survival, it also contained concurrent coactivation of epidermal growth factor receptor (EGFR) signaling. In contrast, the TAE684-resistant (TR3) H3122 cell line did not contain an ALK secondary mutation but instead harbored coactivation of EGFR signaling. Dual inhibition of both ALK and EGFR was the most effective therapeutic strategy for the DFCI076 and H3122 TR3 cell lines. We further identified a subset (3/50; 6%) of treatment naive NSCLC patients with ALK rearrangements that also had concurrent EGFR activating mutations. Our studies identify resistance mechanisms to ALK TKIs mediated by both ALK and by a bypass signaling pathway mediated by EGFR. These mechanisms can occur independently, or in the same cancer, suggesting that the combination of both ALK and EGFR inhibitors may represent an effective therapy for these subsets of NSCLC patients.",
"Lung cancers harboring common EGFR mutations respond to EGFR tyrosine kinase inhibitors (TKI). We previously reported that tumors with exon 18 mutations are particularly sensitive to irreversible second-generation (2G) afatinib compared with first-generation TKIs (1G-TKI). However, data on the mechanisms of acquired resistance to afatinib are limited. We established afatinib-resistant cells by transfecting Ba/F3 cells with common or exon 18 (G719A and Del18) mutations and subjecting them to chronic exposure to increasing concentrations of afatinib. Afatinib-resistant clones were separately established through N-ethyl-N-nitrosourea (ENU) mutagenesis and exposure to fixed concentrations of afatinib. Rebiopsy samples from patients whose tumors acquired resistance to afatinib were analyzed. Afatinib-resistant cells with Del19, L858R, or G719A developed T790M, whereas those with Del18 acquired novel L792F mutation. ENU mutagenesis screening established 84 afatinib-resistant clones. All Del19 clones and most of the other clones acquired only T790M. However, C797S occurred in subsets of L858R, G719A, and Del18 clones. In addition, subsets of Del18 clones acquired L792F. C797S-acquired cells were sensitive to 1G erlotinib. L792F demonstrated intermediate resistance between T790M and C797S to both 1G- and 3G-TKIs, whereas L792F was the least resistant to 2G-TKIs, particularly dacomitinib. Chronic exposure of Del18 + L792F cells to dacomitinib induced additional T790M. T790M was detected in one of four clinical samples. In conclusion, L792F and C797S, in addition to the major T790M, can develop in afatinib-resistant cells particularly using a low dose of afatinib, and these minor mutations appear to exhibit sensitivity to dacomitinib and erlotinib, respectively. These secondary mutations should be tested in clinical practice. Mol Cancer Ther; 16(2); 357-64. ©2016 AACRSee related article by Talbert et al., p. 344.",
"For practical and robust de novo identification of genomic fusions and breakpoints from targeted paired-end DNA sequencing data, we developed Fusion And Chromosomal Translocation Enumeration and Recovery Algorithm (FACTERA). Our method has minimal external dependencies, works directly on a preexisting Binary Alignment/Map file and produces easily interpretable output. We demonstrate FACTERA's ability to rapidly identify breakpoint-resolution fusion events with high sensitivity and specificity in patients with non-small cell lung cancer, including novel rearrangements. We anticipate that FACTERA will be broadly applicable to the discovery and analysis of clinically relevant fusions from both targeted and genome-wide sequencing datasets. http://factera.stanford.edu.",
"Purpose: EGFR and anaplastic lymphoma kinase (ALK) alterations have been regarded as oncogenic drivers and incorporated into clinical practices to manage nonsmall cell lung cancer (NSCLC). Alterations of these two genes were traditionally considered to be mutually exclusive, but recent studies have suggested that they can occur concomitantly. Here, we investigated the prevalence, clinical features and outcomes in response to the treatment of NSCLC patients who harbor EGFR and ALK co-alterations. Methods: We reviewed the genomic profiles of 419 ALK-rearranged NSCLC patients with the intent of investigating the EGFR kinase domain (exon 18-21) and ALK co-alterations. The genomes of these patients were sequenced in a Clinical Laboratory Improvement Amendments-certified laboratory. Results: The overall frequency of concomitant EGFR (exon 18-21) and ALK alterations was 5.01% (21/419) in ALK-rearranged NSCLC patients. The concomitant rate of EGFR alterations in patients with EML4-ALK co-alterations (3.06%, 11/359) was dramatically lower than that in patients with non-EML4-ALK co-alterations (16.67%, 10/60, p<0.01). EML4-ALK/EGF R co-alterations were more prone to occur in females than in males, and non-EML4-ALK/EGFR co-alterations were more common in males than in females (p=0.02). Before the detection of EGFR-ALK co-alterations, some patients were treated with EGFR-TKIs (n=16) according to previously detected EGFR alterations; Kaplan-Meier analysis revealed that EML4-ALK/EGFR co-altered patients (n=7) had a significantly shorter progression-free survival (PFS) after EGFR-TKI treatment than that of non-EML4-ALK/EGFR co-altered patients (n=8; mPFS, 6.0 vs 15.0 months, p=0.046). In addition, we demonstrated the subsequent clinical outcomes of co-altered patients after previous EGFR-TKI treatment. Five EGFR/ALK co-altered patients treated with single TKIs (EGFR-TKIs or ALK-TKIs) displayed diverse clinical outcomes. Three patients who received dual-TKI treatment (EGFR-TKI plus ALK-TKI) all achieved a PFS of more than 5 months (8.4 months, 8.6 months, >5.2 months). Conclusion: EML4-ALK/EGFR and non-EML4-ALK/EGFR co-alterations displayed distinct clinical features and responses to EGFR-TKIs, suggesting that non-EML4-ALK co-alterations are likely to occur as a resistance mechanism to EGFR-TKI. In addition, dual-TKI therapy might be a better choice than single-TKI treatments for these co-altered patients. To the best of our knowledge, this is the largest dual-positive EGFR/ALK cohort study in People's Republic of China.",
"We report a man with advanced adenocarcinoma who harboring exon 19 (E746-A750del) epidermal growth factor receptor (EGFR) deletion and echinoderm microtubule-associated protein like 4-anaplastic lymphoma kinase (EML4-ALK) gene translocation in the re-biospy specimen. The patient was treated with erlotinib with a stable disease but progressed slowly, while crizotinib showed a complete response."
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How can I get a list of all dayflowers in the world?
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Asiatic dayflower (
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[
"Atrazine, a broad-leaf herbicide, has been used widely to control weeds in corn and other crops for several decades and its extensive used has led to widespread contamination of soils and water bodies. Phytoremediation with switchgrass and other native prairie grasses is one strategy that has been suggested to lessen the impact of atrazine in the environment. The goal of this study is to characterize: (1) the uptake of atrazine into above-ground switchgrass biomass; and (2) the degradation and transformation of atrazine over time. A fate study was performed using mature switchgrass columns treated with an artificially-created agricultural runoff containing 16 ppm atrazine. Soil samples and above-ground biomass samples were taken from each column and analyzed for the presence of atrazine and its chlorinated metabolites. Levels of atrazine in both soil and plant material were detectable through the first 2 weeks of the experiment but were below the limit of detection by Day 21. Levels of deethylatrazine (DEA) and didealkylatrazine (DDA) were detected in soil and plant tissue intermittently over the course of the study, deisopropylatrazine (DIA) was not detected at any time point. A radiolabel study using [(14)C]atrazine was undertaken to observe uptake and degradation of atrazine with more sensitivity. Switchgrass columns were treated with a 4 ppm atrazine solution, and above-ground biomass samples were collected and analyzed using HPLC and liquid scintillation counting. Atrazine, DEA, and DIA were detected as soon as 1d following treatment. Two other metabolites, DDA and cyanuric acid, were detected at later time points, while hydroxyatrazine was not detected at all. The percentage of atrazine was observed to decrease over the course of the study while the percentages of the metabolites increased. Switchgrass plants appeared to exhibit a threshold in regard to the amount of atrazine taken up by the plants; levels of atrazine in leaf material peaked between Days 3 and 4 in both studies.",
"The present study has evaluated effect of fluroxypyr concentrations 0-0.8 mg l(-1) (a widely-used herbicide for controlling annual or perennial weeds growth) on selected metabolic and stress-related parameters in Oryza sativa plants after 6 days of exposure. Increasing concentrations decreased shoot growth and accumulation of chlorophylls but had no effect on root biomass. Increasing doses led also to increase in superoxide radical, hydrogen peroxide and proline accumulation, while malondialdehyde, an indicator of lipid peroxidation, was constitutively elevated. Histochemical staining with nitroblue tetrazolium and 3, 3-diaminobenzidine were positively correlated with the generation of superoxide radical and H(2)O(2). The fluroxypyr-induced oxidative stress triggered significant changes in activities of superoxide dismutase, catalase, ascorbate peroxidase and peroxidase (POD). Activities of the antioxidant enzymes show a general increase at low fluroxypyr concentrations and a decrease at high fluroxypyr levels (except for POD). Analysis of naturing polyacrylamide gel electrophoresis confirmed these results. These data support the observation that fluroxypyr-triggered oxidative stress was responsible for the disturbance of the growth in the rice plants.",
"Pesticides sprayed on farmlands can end up in rivers and be transported into estuaries, where they could affect aquatic organisms in freshwater and marine habitats. A series of experiments were conducted using the amphipod Corophium volutator Pallas (Amphipoda, Corophiidae) and single pesticides, namely atrazine (AT), azinphos-methyl (AZ), carbofuran (CA) and endosulfan (EN) that were added to sediments and covered with seawater. Our goal was to compare the concentrations affecting the survival of the animals relative to potential attractant or repellent properties of sediment-spiked pesticides. The avoidance/preference of contaminated/reference sediments by amphipods was examined after 48 and 96 h of exposure using sediments with different organic carbon content. The octanol-water partition coefficients (log K(ow)) ranked the pesticides binding to sediments as EN > AZ > AT > CA. LC(50) and LC(20) covered a wide range of nominal concentrations and ranked toxicity as CA-AZ > EN > AT. Under the experimental set up, only EN initiated an avoidance response and the organic carbon normalised concentration provided consistent results. Using the present data with wide confidence limits, >20% of a population of C. volutator could perish due to the presence of EN before relocation or detecting CA or AZ in sediments by chemical analysis.",
"Anatomical, histochemical and biochemical approaches were used to study mercury uptake and phytotoxicity as well as anti-oxidative responses in two species of ferns [Chinese brake fern (Pteris vittata) and Boston fern (Nephrolepis exaltata)], grown in a hydroponic system. The roots of both cultivars accumulated large amounts of mercury, but exhibited limited mercury translocation to shoots. Mercury exposure led to more pronounced phytotoxicity accompanied by stronger oxidative stress in the shoots of P. vittata than in N. exaltata. N. exaltata established a more effective anti-oxidative system against mercury-induced oxidative stress than did P. vittata. The activity of anti-oxidative enzymes (superoxide dismutase, catalase and glutathione reductase) increased. The reduced ascorbate (ASA) and oxidized ascorbate (DHA) are regulated. Mercury exposure led to an increase in the concentration of glutathione (GSH) in both fern species. The present study suggests that N. exaltata is more tolerant to mercury exposure than P. vittata, which has been also reported to be more tolerant to arsenic exposure. N. exaltata may thus have potential for phytostabilization of soils or phytofiltration of waste water contaminated with mercury."
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The Cambridge Study in Delinquent Development: Health and Offending Across Generations
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Research suggests that convicted persons are more likely than non-convicted persons to suffer poor health. However, few longitudinal studies have investigated associations between health and offending across generations. Using the Cambridge Study in Delinquent Development, this article prospectively investigates the relationship between health and offending across generations and between genders. At the average age of 25, third generation convicted males and females reported a higher incidence of serious drug use than non-convicted persons. Convicted males reported a higher incidence of mental illness and self-harm, whereas convicted females reported a lower incidence of physical illness, mental illness, self-harm and hospitalizations when compared to non-convicted females. Convicted males reported a higher incidence of industrial accidents, sports injuries and fight injuries, but a lower incidence of road accidents, whereas convicted females were more likely to report road accidents. Like their fathers, convicted males show worse health compared to non-convicted individuals.
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[
"The traditional approach to 'exact' small-sample interval estimation of the odds ratio for binomial, Poisson, or multinomial samples uses the conditional distribution to eliminate nuisance parameters. This approach can be very conservative. For two independent binomial samples, we study an unconditional approach with overall confidence level guaranteed to equal at least the nominal level. With small samples this interval tends to be shorter and have coverage probabilities nearer the nominal level.",
"We examined self-reported health among formerly incarcerated mothers. We used data from the Fragile Families and Child Wellbeing Study (n = 4096), a longitudinal survey of mostly unmarried parents in urban areas, to estimate the association between recent incarceration (measured as any incarceration in the past 4 years) and 5 self-reported health conditions (depression, illicit drug use, heavy drinking, fair or poor health, and health limitations), net of covariates including health before incarceration. In adjusted logistic regression models, recently incarcerated mothers, compared with their counterparts, have an increased likelihood of depression (odds ratio [OR] = 1.60; 95% confidence interval [CI] = 1.18, 2.17), heavy drinking (OR = 1.79; 95% CI = 1.19, 2.68), fair or poor health (OR = 1.49; 95% CI = 1.08, 2.06), and health limitations (OR = 1.78; 95% CI = 1.27, 2.50). This association is similar across racial/ethnic subgroups and is larger among mothers who share children with fathers who have not been recently incarcerated. Recently incarcerated mothers struggle with even more health conditions than expected given the disadvantages they experience before incarceration. Furthermore, because incarceration is concentrated among those who are most disadvantaged, incarceration may increase inequalities in population health.",
"This article examines the relationship between incarceration and health functioning. Using data from the National Longitudinal Survey of Youth, the relationship between incarceration and more than 20 different measures of health are tested. Using multiple analytic procedures, a distinctive pattern of association emerges. Individuals with a history of incarceration appear consistently more likely to be afflicted with infectious disease and other illnesses associated with stress. In contrast, no consistent relationships were observed between incarceration status and ailments unrelated to stress or infectious disease. The results suggest that exposure to infectious disease and stress are important to understanding the lasting impact of incarceration on health.",
"Mass incarceration has profoundly restructured the life courses of not only marginalized adult men for whom this event is now so prevalent but also their families. We examined research published from 2000 to 2017 on the consequences of parental incarceration for child health in the United States. In addition to focusing on specific health outcomes, we also considered broader indicators of child well-being because there has been little research on the association between parental incarceration and objectively measured child health outcomes. Our findings support 4 conclusions. First, paternal incarceration is negatively associated-possibly causally so-with a range of child health and well-being indicators. Second, although some research has suggested a negative association between maternal incarceration and child health, the evidence on this front is mixed. Third, although the evidence for average effects of paternal incarceration on child health and well-being is strong, research has also suggested that some key factors moderate the association between paternal incarceration and child health and well-being. Finally, because of the unequal concentration of parental incarceration and the negative consequences this event has for children, mass incarceration has increased both intracountry inequality in child health in the United States and intercountry inequality in child health between the United States and other developed democracies. In light of these important findings, investment in data infrastructure-with emphasis on data sets that include reliable measures of parental incarceration and child health and data sets that facilitate causal inferences-is needed to understand the child health effects of parental incarceration.",
"Parental incarceration is now prevalent in community samples (e.g., with 11% of children reporting paternal imprisonment and 3% reporting maternal imprisonment in a national sample), pointing to a potentially important childhood trauma that should be included in work on contemporary childhood stressors in this era of mass incarceration. This paper investigates the influences of maternal and paternal imprisonment on changes in young adult mental health using a nationally representative sample. We assess four perspectives-gendered loss, same-sex role model, intergenerational stress, and maternal salience - on the joint influences of maternal and paternal incarceration within the broader stress process paradigm. The results generalize support for a gendered loss perspective developed in work on parental death and an early small study of parental incarceration. This pattern reveals maternal incarceration increases depressive symptoms while paternal incarceration increases substance role problems. Chronicity of parental imprisonment and its timing are also influential. Analyses further specify a vulnerability of male and minority young adults to high levels of mental health problems following maternal and paternal incarceration in adolescence."
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A simple and effective mixing carbonization-activation process to prepare rice hull-derived porous Si-carbon materials
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A simple and effective mixing carbonization-activation process was developed to prepare rice hull-derived porous Si-carbon materials. The morphologies and pore structures of the materials were controlled effectively without any loading or additions at various carbonization temperatures. The structures of the samples changed from large pores and thick walls after 800 ∘C carbonization to small pores and thin walls after 1000 ∘C carbonization. An additional alkali activation-carbonization process led to coral reef-like structures surrounded by squama in the sample that underwent 900 ∘C carbonization (Act-RH-900). This optimal material (Act-RH-900) had a large specific surface area (768 m
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[
"We successfully prepared two different classes of hypercrosslinked porous organic polymers (HPPs)-the tetraphenylethene (TPE) and (4-(5,6-Diphenyl-1H-Benzimidazol-2-yl)-triphenylamine (DPT) HPPs-through the Friedel-Crafts polymerization of tetraphenylethene and 4-(5,6-diphenyl-1H-benzimidazol-2-yl)-triphenylamine, respectively, with 1,4-bis(chloromethyl)benzene (Ph-2Cl) in the presence of anhydrous FeCl3 as a catalyst. Our porous materials exhibited high BET surface areas (up to 1000 m2 g-1) and good thermal stabilities. According to electrochemical and dyes adsorption applications, the as-prepared DPT-HPP exhibited a high specific capacitance of 110 F g-1 at a current density of 0.5 A g-1, with an excellent cycling stability of over 2000 times at 10 A g-1. In addition, DPT-HPP showed a high adsorption capacity up to 256.40 mg g-1 for the removal of RhB dye from water.",
"Covalent organic frameworks (COFs) are a family of crystalline porous networks having applications in various fields, including gas and energy storage. Despite respectable progress in the synthesis of such crystalline materials, examples of the use of template-free methods to construct COFs having hollow nano- and microstructures are rare. Furthermore, all reported methods for synthesizing these hollow structural COFs have involved [4 + 2] and [3 + 2] condensations. Herein, we report the synthesis of hollow microspherical and microtubular carbazole-based COFs through template-free, one-pot, [3 + 3] condensations of the novel triamine 9-(4-aminophenyl)-carbazole-3,6-diamine (Car-3NH2) and triformyl linkers with various degrees of planarity. Depending upon the monomer's planarity, a unique morphological variety was observed. A time-dependent study revealed that each COF formed through an individual mechanism depended on the degree of planarity of the triformyl linker; it also confirmed that the hollow structures of these COFs formed through inside-out Ostwald ripening. Our COFs exhibited high Brunauer-Emmett-Teller surface areas (up to ca. 1400 m2 g-1), excellent crystallinity, and high thermal stability. Moreover, the CO2 uptake capacities of these COFs were excellent: up to 61 and 123 mg g-1 at 298 and 273 K, respectively. The high surface areas facilitated greater numbers of strong interactions with CO2 molecules, leading to high CO2 uptake capacities. Moreover, the prepared COFs exhibited redox activity because of their redox-active triphenylamine and pyridine groups, which can be utilized in electrochemical energy storages. Accordingly, such hollow COFs having high surface areas appear to be useful materials for industrial and biological applications.",
"Desiccant driven dehumidification for maintaining the proper humidity levels and atmospheric water capture with minimum energy penalty are important aspects in heat pumps, refrigeration, gas and liquid purifications, gas sensing, and clean water production for improved human health and comfort. Water adsorption by using nanoporous materials has emerged as a viable alternative to energy-intensive industrial processes, thus understanding the significance of their porosity, high surface areas, vast pore volumes, chemical and structural features relative to the water adsorption is quite important. In this review article, important features of nanoporous materials are presented, including zeolites, porous carbons, as well as crystalline and amorphous porous organic polymers (POPs) to define the interactions between the water molecules and the polar/non-polar functional groups on the surface of these nanoporous materials. In particular, focus is placed on the recent developments in POPs in the context of water capture as a result of their remarkable stability towards water and wide range of available synthetic routes and building blocks for their synthesis. We also highlighted recent approaches to increase the water sorption capacity of POPs by modifying their structure, morphology, porosity, and chemical functionality while emphasizing their promising future in this emerging area."
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Untargeted LC-Ion Mobility Spectrometry-Mass Spectrometry-Based Analysis of Per- and Polyfluoroalkyl Substances in Houston Ship Channel/Galveston Bay
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Per- and polyfluoroalkyl substances (PFAS) are persistent organic pollutants of concern because of their ubiquitous presence in surface and ground water; analytical methods that can be used for rapid comprehensive exposure assessment and fingerprinting of PFAS are needed. Following the fires at the Intercontinental Terminals Company (ITC) in Deer Park, TX in 2019, large quantities of PFAS-containing firefighting foams were deployed. The release of these substances into the Houston Ship Channel/Galveston Bay (HSC/GB) prompted concerns over the extent and level of PFAS contamination. A targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based study of temporal and spatial patterns of PFAS associated with this incident revealed presence of 7 species; their levels gradually decreased over a 6-month period. Because the targeted LC-MS/MS analysis was focused on about 30 PFAS molecules, it may have missed other PFAS compounds present in firefighting foams. Therefore, we utilized untargeted LC-ion mobility spectrometry-mass spectrometry (LC-IMS-MS)-based analytical approach for a more comprehensive characterization of PFAS in these water samples. We analyzed 31 samples from 9 sites in the HSC/GB that were collected over 5 months after the incident. Our data showed that additional 19 PFAS were detected in surface water of HSC/GB, most of them decreased gradually after the incident. PFAS features detected by LC-MS/MS correlated well in abundance with LC-IMS-MS data; however, LC-IMS-MS identified a number of additional PFAS, many known to be components of firefighting foams. These findings therefore illustrate that untargeted LC-IMS-MS improved our understanding of PFAS presence in complex environmental samples.
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[
"Firefighting foams contain per- and polyfluoroalkyl substances (PFAS) - a class of compounds widely used as surfactants. PFAS are persistent organic pollutants that have been reported in waterways and drinking water systems across the United States. These substances are of interest to both regulatory agencies and the general public because of their persistence in the environment and association with adverse health effects. PFAS can be released in large quantities during industrial incidents because they are present in most firefighting foams used to suppress chemical fires; however, little is known about persistence of PFAS in public waterways after such events. In response to large-scale fires at Intercontinental Terminal Company (ITC) in Houston, Texas in March 2019, almost 5 million liters of class B firefighting foams were used. Much of this material flowed into the Houston Ship Channel and Galveston Bay (HSC/GB) and concerns were raised about the levels of PFAS in these water bodies that have commercial and recreational uses. To evaluate the impact of the ITC incident response on PFAS levels in HSC/GB, we collected 52 surface water samples from 12 locations over a 6-month period after the incident. Samples were analyzed using liquid chromatography-mass spectrometry to evaluate 27 PFAS, including perfluorocarboxylic acids, perfluorosulfonates and fluorotelomers. Among PFAS that were evaluated, 6:2 FTS and PFOS were detected at highest concentrations. Temporal and spatial profiles of PFAS were established; we found a major peak in the level of many PFAS in the days and weeks after the incident and a gradual decline over several months with patterns consistent with the tide- and wave-associated water movements. This work documents the impact of a large-scale industrial fire, on the environmental levels of PFAS, establishes a baseline concentration of PFAS in HSC/GB, and highlights the critical need for development of PFAS water quality standards.",
"Global profiling of xenobiotics in human matrices in an untargeted mode is gaining attention for studying the environmental chemical space of the human exposome. Defined as the study of a comprehensive inclusion of environmental influences and associated biological responses, human exposome science is currently evolving out of the metabolomics science. In analogy to the latter, the development and applications of high resolution mass spectrometry (HRMS) has shown potential and promise to greatly expand our ability to capture the broad spectrum of environmental chemicals in exposome studies. HRMS can perform both untargeted and targeted analysis because of its capability of full- and/or tandem-mass spectrum acquisition at high mass accuracy with good sensitivity. The collected data from target, suspect and non-target screening can be used not only for the identification of environmental chemical contaminants in human matrices prospectively but also retrospectively. This review covers recent trends and advances in this field. We focus on advances and applications of HRMS in human biomonitoring studies, and data acquisition and mining. The acquired insights provide stepping stones to improve understanding of the human exposome by applying HRMS, and the challenges and prospects for future research.",
"The term \"exposome\" was coined in 2005 to underscore the importance of the environment to human health and to bring research efforts in line with those on the human genome. The ability to characterize environmental exposures through biomonitoring is key to exposome research efforts. Our objectives were to describe why traditional and nontraditional (exposomic) biomonitoring are both critical in studies aiming to capture the exposome and to make recommendations on how to transition exposure research toward exposomic approaches. We describe the biomonitoring needs of exposome research and approaches and recommendations that will help fill the gaps in the current science. Traditional and exposomic biomonitoring approaches have key advantages and disadvantages for assessing exposure. Exposomic approaches differ from traditional biomonitoring methods in that they can include all exposures of potential health significance, whether from endogenous or exogenous sources. Issues of sample availability and quality, identification of unknown analytes, capture of nonpersistent chemicals, integration of methods, and statistical assessment of increasingly complex data sets remain challenges that must continue to be addressed. To understand the complexity of exposures faced throughout the lifespan, both traditional and nontraditional biomonitoring methods should be used. Through hybrid approaches and the integration of emerging techniques, biomonitoring strategies can be maximized in research to define the exposome.",
"Per- and polyfluoroalkyl substances (PFAS) are an ensemble of persistent organic pollutants of global interest because of their associations with adverse health outcomes. Currently, environmental PFAS pollution is prolific as a result of the widespread manufacturing of these compounds and their chemical persistence. In this work, we demonstrate the advantages of adding ion mobility spectrometry (IMS) separation to existing LC-MS workflows for PFAS analysis. Using a commercially available drift tube IMS-MS, we characterized PFAS species and isomeric content in both analytical standards and environmental water samples. Molecular trendlines based on intrinsic mass and structural relationships were also explored for individual PFAS subclasses (e.g. PFSA, PFCA, etc.). Results from rapid IMS-MS analyses provided a link between mass and collision cross sections (CCS) for specific PFAS families and are linked to compositional differences in molecular structure. In addition, CCS values provide additional confidence of annotating prioritized features in untargeted screening studies for potential environmental pollutants. Results from this study show that the IMS separation provides novel information to support traditional LC-MS PFAS analyses and will greatly benefit the evaluation of unknown pollutants in future environmental studies.",
"We report a new method for fast and sensitive analyses of biologically relevant fatty acids (FAs) in red blood cells (RBC) by liquid chromatography mass spectrometry (LC-MS). A new chemical derivatization approach was developed forming picolylamides from FAs in a quantitative reaction. Fourteen derivatized FA standards, including saturated and unsaturated FAs from C14 to C22, were efficiently separated within 15 min. In addition, the use of a recently introduced benchtop orbitrap mass spectrometer under positive electrospray ionization (ESI) full scan mode showed a 2-10-fold improvement in sensitivity compared with a conventional tandem MS method, with a limit of detection in the low femtomole range for saturated and unsaturated FAs. The developed method was applied to determine FA concentrations in RBC with intra- and interday coefficients of variation below 10%.",
"The production, use, environmental fate, occurrence, and toxicity of perfluoroalkylated substances have been reviewed. Although only a limited number of essential physicochemical data are available, thus hampering a complete assessment of the environmental fate of PFAS, it has become clear that PFAS behave differently from other nonpolar organic micropollutants. PFAS are present in environmental media in urbanized areas both with and without fluorochemicals production sites. The presence of PFOS at levels above the limit of detection has been demonstrated in almost all organisms sampled in a global survey as well as in both nonexposed and exposed human populations. The acute and chronic ecotoxicity of PFOS, PFOA, and 8:2 FTOH to aquatic organisms is moderate to low. Acute toxicity to rodents is also low. PFOS concentrations in effluents have been reported that approach indicative target values derived from available aquatic toxicity data. PFOA has been found to be weakly carcinogenic. This review shows the importance of the perfluoroalkylated substances for the environment and the necessity to fill the current gaps in knowledge of their environmental fate and effects.",
"EXPOsOMICS is a European Union funded project that aims to develop a novel approach to the assessment of exposure to high priority environmental pollutants, by characterizing the external and the internal components of the exposome. It focuses on air and water contaminants during critical periods of life. To this end, the project centres on 1) exposure assessment at the personal and population levels within existing European short and long-term population studies, exploiting available tools and methods which have been developed for personal exposure monitoring (PEM); and 2) multiple \"omic\" technologies for the analysis of biological samples (internal markers of external exposures). The search for the relationships between external exposures and global profiles of molecular features in the same individuals constitutes a novel advancement towards the development of \"next generation exposure assessment\" for environmental chemicals and their mixtures. The linkage with disease risks opens the way to what are defined here as 'exposome-wide association studies' (EWAS)."
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Development of a novel poly(methyl methacrylate)-based resin for stereolithography additive manufacturing
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Poly(methyl methacrylate) (PMMA) is widely used in dental applications. However, PMMA specialized for stereolithography (SLA) additive manufacturing (3D-printing) has not been developed yet. This study aims to develop a novel PMMA-based resin for SLA 3D-printing by mixing methyl methacrylate (MMA), ethylene glycol dimethacrylate (EGDMA), and PMMA powder in various mixing ratios. The printability and the viscosity of the PMMA-based resins were examined to determine their suitability for 3D-printing. The mechanical properties (flexural strength and Vickers hardness), shear bond strength, degree of conversion, physicochemical properties (water sorption and solubility), and cytotoxicity for L929 cells of the resulting resins were compared with those of three commercial resins: one self-cured resin and two 3D-print resins. EGDMA and PMMA were found to be essential components for SLA 3D-printing. The viscosity increased with PMMA content, while the mechanical properties improved as EGDMA content increased. The shear bond strength tended to decrease as EGDMA increased. Based on these characteristics, the optimal composition was determined to be 30% PMMA, 56% EGDMA, 14% MMA with flexural strength (84.6 ± 7.1 MPa), Vickers hardness (21.6 ± 1.9), and shear bond strength (10.5 ± 1.8 MPa) which were comparable to or higher than those of commercial resins. The resin's degree of conversion (71.5 ± 0.7%), water sorption (19.7 ± 0.6 μg/mm
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[
"To optimize the 3D printing of a dental material for provisional crown and bridge restorations using a low-cost stereolithography 3D printer; and compare its mechanical properties against conventionally cured provisional dental materials. Samples were 3D printed (25×2×2mm) using a commercial printable resin (NextDent C&B Vertex Dental) in a FormLabs1+ stereolithography 3D printer. The printing accuracy of printed bars was determined by comparing the width, length and thickness of samples for different printer settings (printing orientation and resin color) versus the set dimensions of CAD designs. The degree of conversion of the resin was measured with FTIR, and both the elastic modulus and peak stress of 3D printed bars was determined using a 3-point being test for different printing layer thicknesses. The results were compared to those for two conventionally cured provisional materials (Integrity®, Dentsply; and Jet®, Lang Dental Inc.). Samples printed at 90° orientation and in a white resin color setting was chosen as the most optimal combination of printing parameters, due to the comparatively higher printing accuracy (up to 22% error), reproducibility and material usage. There was no direct correlation between printing layer thickness and elastic modulus or peak stress. 3D printed samples had comparable modulus to Jet®, but significantly lower than Integrity®. Peak stress for 3D printed samples was comparable to Integrity®, and significantly higher than Jet®. The degree of conversion of 3D printed samples also appeared higher than that of Integrity® or Jet®. Our results suggest that a 3D printable provisional restorative material allows for sufficient mechanical properties for intraoral use, despite the limited 3D printing accuracy of the printing system of choice.",
"Major changes are taking place in dental laboratories as a result of new digital technologies. Our aim is to provide an overview of these changes. In this article the reader will be introduced to the range of layered fabrication technologies and suggestions are made how these might be used in dentistry. Key publications in English from the past two decades are surveyed. The first digital revolution took place many years ago now with the production of dental restorations such as veneers, inlays, crowns and bridges using dental CAD-CAM systems and new improved systems appear on the market with great rapidity. The reducing cost of processing power will ensure that these developments will continue as exemplified by the recent introduction of a new range of digital intra-oral scanners. With regard to the manufacture of prostheses this is currently dominated by subtractive machining technology but it is inevitable that the additive processing routes of layered fabrication, such as FDM, SLA, SLM and inkjet printing, will start to have an impact. In principle there is no reason why the technology cannot be extended to all aspects of production of dental prostheses and include customized implants, full denture construction and orthodontic appliances. In fact anything that you might expect a dental laboratory to produce can be done digitally and potentially more consistently, quicker and at a reduced cost. Dental device manufacturing will experience a second revolution when layered fabrication techniques reach the point of being able to produce high quality dental prostheses. The challenge for the dental materials research community is to marry the technology with materials that are suitable for use in dentistry. This can potentially take dental materials research in a totally different direction.",
"The application of 3-dimensional printing technology is emerging in dentistry and is being increasingly used to fabricate dental restorations. To date, scientific evidence is lacking regarding the effect of different factors on the mechanical properties of the printed restorations with the additive manufacturing technique. The purpose of this in vitro study was to evaluate the effect of build direction (layer orientation) on the mechanical properties of a novel 3-dimensionally (3D)-printed dental restorative material. Based on the printing direction, 2 groups were tested. In the first group (n=20), the specimens were vertically printed with the layers oriented perpendicular to the load direction. In the second group (n=20), the specimens were horizontally printed with the layers oriented parallel to the load direction. All specimens were fabricated using the DW028D 3D-printer. The specimens were loaded with a universal testing machine at a crosshead speed of 1 mm/min with a 10-kN load cell. The test was performed at room temperature (22°C) under dry testing conditions. The compressive strength was calculated for both groups, and the results were compared using the unpaired t test (α=.05). The mean ±SD compressive strength for the vertically printed specimens was 297 MPa (±34) compared with 257 MPa (±41) for the horizontally printed specimens (P=.002). Within the limitations of this study, the layer orientation was found to influence the compressive strength of the material. Vertically printed specimens with the layers oriented perpendicular to load direction have improved mechanical properties more than horizontally printed specimens with the layers oriented parallel to load direction."
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Wnt signaling pathway in liver fibrosis
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Liver fibrosis is a life-threatening disease, with challenging morbidity and mortality for healthcare systems worldwide. It imparts an enormous economic burden to societies, making continuous research and informational updates about its pathogenesis and treatment crucial. This review's focus is on the current knowledge about the Wnt signaling pathway, serving as an important pathway in liver fibrosis development and activation of hepatic stellate cells (HSCs). Two types of Wnt pathways are distinguished, namely the ß-catenin-dependent canonical and non-canonical Ca
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[
"Stellate cells, either hepatic (HSCs) or pancreatic (PSCs), are a type of interstitial cells characterized by their ability to store retinoids in lipid vesicles. In pathological conditions both HSCs and PSCs lose their retinoid content and transform into fibroblast-like cells, contributing to the fibrogenic response. HSCs also participate in other functions including vasoregulation, drug detoxification, immunotolerance, and maintenance of the hepatocyte population. PSCs maintain pancreatic tissue architecture and regulate pancreatic exocrine function. Recently, PSCs have attracted the attention of researchers due to their interactions with pancreatic ductal adenocarcinoma cells. PSCs promote tumour growth and angiogenesis, and their fibrotic activity increases the resistance of pancreatic cancer to chemotherapy and radiation. We are reviewing the current literature concerning the role played by retinoids in the physiology and pathophysiology of the stellate cells, paying attention to their developmental aspects as well as the function of stellate cells in tissue repair and organ regeneration.",
"The canonical Wnt-β-catenin pathway is a complex, evolutionarily conserved signalling mechanism that regulates fundamental physiological and pathological processes. Wnt-β-catenin signalling tightly controls embryogenesis, including hepatobiliary development, maturation and zonation. In the mature healthy liver, the Wnt-β-catenin pathway is mostly inactive but can become re-activated during cell renewal and/or regenerative processes, as well as in certain pathological conditions, diseases, pre-malignant conditions and cancer. In hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA), the two most prevalent primary liver tumours in adults, Wnt-β-catenin signalling is frequently hyperactivated and promotes tumour growth and dissemination. A substantial proportion of liver tumours (mainly HCC and, to a lesser extent, CCA) have mutations in genes encoding key components of the Wnt-β-catenin signalling pathway. Likewise, hepatoblastoma, the most common paediatric liver cancer, is characterized by Wnt-β-catenin activation, mostly as a result of β-catenin mutations. In this Review, we discuss the most relevant molecular mechanisms of action and regulation of Wnt-β-catenin signalling in liver development and pathophysiology. Moreover, we highlight important preclinical and clinical studies and future directions in basic and clinical research.",
"Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy with a dismal survival rate. Persistent activation of pancreatic stellate cells (PSCs) can perturb the biomechanical homoeostasis of the tumour microenvironment to favour cancer cell invasion. Here we report that ATRA, an active metabolite of vitamin A, restores mechanical quiescence in PSCs via a mechanism involving a retinoic acid receptor beta (RAR-β)-dependent downregulation of actomyosin (MLC-2) contractility. We show that ATRA reduces the ability of PSCs to generate high traction forces and adapt to extracellular mechanical cues (mechanosensing), as well as suppresses force-mediated extracellular matrix remodelling to inhibit local cancer cell invasion in 3D organotypic models. Our findings implicate a RAR-β/MLC-2 pathway in peritumoural stromal remodelling and mechanosensory-driven activation of PSCs, and further suggest that mechanical reprogramming of PSCs with retinoic acid derivatives might be a viable alternative to stromal ablation strategies for the treatment of PDAC.",
"In mouse fetal liver, hepatoblasts, sinusoidal endothelial cells and macrophages (or erythroblastic islands) promote differentiation and proliferation of hematopoietic cells through cell-cell interactions and secretion of cytokines and extracellular matrix factors. Until now, we have had little knowledge of the hematopoietic cytokines or extracellular matrix mRNAs expressed in hepatic stellate cells. Using p75 neurotrophin receptor (p75NTR) to mark this cell population, we sorted 12.5, 14.5 and 16.5 dpc hepatic stellate cells and analyzed expression of cytokines and extracellular matrix mRNAs. Among cytokines, insulin-like growth factor 2 (Igf2) was highly expressed at all three stages analyzed. The extracellular matrix molecule fibronectin (Fn1) was highly expressed in 12.5 dpc cells, whereas vitronectin (Vtn) was highly expressed in 14.5 and 16.5 dpc hepatic stellate cells. Among liver cells, Igf2 was predominantly expressed in hepatoblast-like cells at all three stages examined, suggesting that hepatoblast-like cells are an essential part of the niche that maintains homeostasis of hematopoietic cells in embryonic mouse liver. Defining these expression patterns could facilitate our understanding of cross talk between cytokine and extracellular matrix molecules in hepatic stellate cells and benefit research in developmental hematopoiesis as well as the study of liver biology.",
"In the development of liver fibrosis, activated hepatic stellate cells (HSCs) contribute to the synthesis and deposition of extracellular matrix (ECM) proteins. HSC activation is considered as a central driver of liver fibrosis. Recently, microRNAs (miRNAs) have been reported to act as key regulators in HSC activation. Pinostilbene hydrate (PSH), a methylated derivative of resveratrol, has demonstrated anti-inflammatory, antioxidant and anti-tumour activities. However, the effects of PSH on HSC activation remain unclear. The effects of PSH on HSC activation were examined. Moreover, the roles of WNT inhibitory factor 1 (WIF1) and miR-17-5p in the effects of PSH on HSC activation were examined. PSH induced a significant reduction in HSC proliferation. PSH also effectively inhibited HSC activation, with reduced α-SMA and collagen expression. Notably, it was found that Wnt/β-catenin signalling was involved in the effects of PSH on HSC activation. PSH resulted in Wnt/β-catenin signalling inactivation, with a reduction in TCF activity as well as β-catenin nuclear translocation. Further studies showed that PSH inhibited Wnt/β-catenin signalling via regulation of WIF1 and miR-17-5p. Reduced HSC activation caused by PSH could be restored by loss of WIF1 or miR-17-5p mimics. Luciferase reporter assays further confirmed that WIF1 was a target of miR-17-5p. PSH has a significant protective effect against HSC activation. In addition, we demonstrate that PSH enhances WIF1 expression and inhibits Wnt/β-catenin signalling via miR-17-5p, contributing to the suppression of HSC activation.",
"There is an urgent need to develop antifibrotic therapies for chronic liver disease, and clarify which endpoints in antifibrotic trials will be acceptable to regulatory agencies. The American Association for the Study of Liver Diseases sponsored an endpoints conference to help accelerate the efficient testing of antifibrotic agents and develop recommendations on clinical trial design for liver fibrosis. In this review, we summarize the salient and novel elements of this conference and provide directions for future clinical trial design. The article follows the structure of the conference and is organized into five areas: (1) antifibrotic trial design; (2) preclinical proof-of-concept studies; (3) pharmacological targets, including rationale and lessons to learn; (4) rational drug design and development; and (5) consensus and recommendations on design of clinical trials in liver fibrosis. Expert overviews and collaborative discussions helped to summarize the key unmet needs and directions for the future, including: (1) greater clarification of at-risk populations and study groups; (2) standardization of all elements of drug discovery and testing; (3) standardization of clinical trial approaches; (4) accelerated development of improved noninvasive markers; and (5) need for exploration of potential off-target toxicities of future antifibrotic drugs.",
"Niclosamide has preclinical activity against a wide range of cancers. In prostate cancer, it inhibits androgen receptor variant 7 and synergizes with abiraterone. The approved niclosamide formulation has poor oral bioavailability. The primary objective of this phase Ib trial was to identify a maximum tolerated dose (MTD) and recommended phase 2 dose (RP2D) of a novel reformulated orally-bioavailable niclosamide/PDMX1001 in combination with abiraterone and prednisone in men with castration-resistant prostate cancer (CRPC). Eligible patients had progressing CRPC, adequate end-organ function, and no prior treatment with abiraterone or ketoconazole. Patients were treated with escalating doses of niclosamide/PDMX1001 and standard doses of abiraterone and prednisone. Peak and trough niclosamide plasma levels were measured. Common Terminology Criteria for Adverse Events (CTCAE) v4.0 and Prostate Cancer Working Group 2 criteria were used to evaluate toxicities and responses. Nine patients with metastatic CRPC were accrued, with no dose-limiting toxicities observed at all dose levels. The recommended Phase II dose of niclosamide/PDMX1001 was 1200 mg orally (PO) three times daily plus abiraterone 1000 mg PO once daily and prednisone 5 mg PO twice daily. Trough and peak niclosamide concentrations exceeded the therapeutic threshold of > 0.2 µM. The combination was well tolerated with most frequent adverse effects of diarrhea. Five out of eight evaluable patients achieved a PSA response; two achieved undetectable PSA and radiographic response. A novel niclosamide/PDMX1001 reformulation achieved targeted plasma levels when combined with abiraterone and prednisone, and was well tolerated. Further study of niclosamide/PDMX1001 with this combination is warranted."
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-helix/-crystallin domain architecture of Candida glabrata cell wall adhesins.
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Candida glabrata is an opportunistic pathogenic yeast frequently causing infections in humans. Though it lacks typical virulence factors such as hyphal development, C. glabrata contains a remarkably large and diverse set of putative wall adhesins that is crucial for its success as pathogen. Here, we present an analysis of putative adhesins from the homology clusters V and VI. First, sequence similarity network analysis revealed relationships between cluster V and VI adhesins and S. cerevisiae haze protective factors (Hpf). Crystal structures of A-regions from cluster VI adhesins Awp1 and Awp3b reveal a parallel right-handed β-helix domain that is linked to a C-terminal β-sandwich. Structure solution of the A-region of Awp3b via single wavelength anomalous diffraction phasing revealed the largest known lanthanide cluster with 21 Gd3+ ions. Awp1-A and Awp3b-A show structural similarity to pectate lyases but binding to neither carbohydrates nor Ca2+ was observed. Phenotypic analysis of awp1Δ, awp3Δ, and awp1,3Δ double mutants did also not confirm their role as adhesins. In contrast, deletion mutants of the cluster V adhesin Awp2 in the hyperadhesive clinical isolate PEU382 demonstrated its importance for adhesion to polystyrene or glass, biofilm formation, cell aggregation and other cell surface-related phenotypes. Together with cluster III and VII adhesins our study shows that C. glabrata CBS138 can rely on a set of 42 Awp1-related adhesins with β-helix/α-crystallin domain architecture for modifying the surface characteristics of its cell wall.
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"Candida glabrata is among the most prevalent causes of candidiasis. Unlike Candida albicans, it is not capable of changing morphology between yeast and hyphal forms but instead has developed other virulence factors. An important feature is its unprecedented large repertoire of predicted cell wall adhesins, which are thought to enable adherence to a variety of surfaces under different conditions. Here, we analyzed the wall proteome of PEU1221, a high biofilm-forming clinical strain isolated from an infected central venous catheter, under biofilm-forming conditions. This isolate shows increased incorporation of putative adhesins, including eight proteins that were not detected in walls of reference strain ATCC 2001, and of which Epa22, Awp14, and Awp2e were identified for the first time. The proteomics data suggest that cluster III adhesin Awp14 is relatively abundant in PEU1221. Phenotypic studies with awp14Δ deletion mutants showed that Awp14 is not responsible for the high biofilm formation of PEU1221 onto polystyrene. However, awp14Δ mutant cells in PEU1221 background showed a slightly diminished binding to chitin and seemed to sediment slightly slower than the parental strain suggesting implication in fungal cell-cell interactions. By structural modeling, we further demonstrate similarity between the ligand-binding domains of cluster III adhesin Awp14 and those of cluster V and VI adhesins. In conclusion, our work confirms the increased incorporation of putative adhesins, such as Awp14, in high biofilm-forming isolates, and contributes to decipher the precise role of these proteins in the establishment of C. glabrata infections.",
"Fungal adhesins (Als) or flocculins are family of cell surface proteins that mediate adhesion to diverse biotic and abiotic surfaces. A striking characteristic of Als proteins originally identified in the pathogenic Candida albicans is to form functional amyloids that mediate cis-interaction leading to the formation of adhesin nanodomains and trans-interaction between amyloid sequences of opposing cells. In this report, we show that flocculins encoded by FLO11 in Saccharomyces cerevisiae behave like adhesins in C. albicans. To do so, we show that the formation of nanodomains under an external physical force requires a threshold number of amyloid-forming sequences in the Flo11 protein. Then, using a genome editing approach, we constructed strains expressing variants of the Flo11 protein under the endogenous FLO11 promoter, leading to the demonstration that the loss of amyloid-forming sequences strongly reduces cell-cell interaction but has no effect on either plastic adherence or invasive growth in agar, both phenotypes being dependent on the N- and C-terminal ends of Flo11p. Finally, we show that the location of Flo11 is not altered either by the absence of amyloid-forming sequences or by the removal of the N- or C-terminus of the protein.",
"The prediction of interresidue contacts and distances from coevolutionary data using deep learning has considerably advanced protein structure prediction. Here, we build on these advances by developing a deep residual network for predicting interresidue orientations, in addition to distances, and a Rosetta-constrained energy-minimization protocol for rapidly and accurately generating structure models guided by these restraints. In benchmark tests on 13th Community-Wide Experiment on the Critical Assessment of Techniques for Protein Structure Prediction (CASP13)- and Continuous Automated Model Evaluation (CAMEO)-derived sets, the method outperforms all previously described structure-prediction methods. Although trained entirely on native proteins, the network consistently assigns higher probability to de novo-designed proteins, identifying the key fold-determining residues and providing an independent quantitative measure of the \"ideality\" of a protein structure. The method promises to be useful for a broad range of protein structure prediction and design problems.",
"The pectate lyase (Bsp165PelA) from Bacillus sp. N16-5 has great potential in industrial applications because it shows high specific activity under extremely alkaline conditions. Besides, activity measurement of Bsp165PelA does not require addition of calcium, in a way different from the other pectate lyases. Here we report crystal structures of Bsp165PelA in apo-form and in complex with trigalacturonate. The parallel β-helix, active site residues and substrate binding cleft are similar to those in the other pectate lyases from Polysaccharide Lyase family 1. However, some of the highly conserved Ca(2+) binding residues and secondary structures are altered in Bsp165PelA, making it difficult to coordinate with Ca(2+) as in the other pectate lyases. We found Bsp165PelA forms some direct enzyme-substrate interactions instead of using Ca(2+) ions bridging in the extremely alkaline environment.",
"For host colonization, the human fungal pathogen Candida glabrata is known to utilize a large family of highly related surface-exposed cell wall proteins, the lectin-like epithelial adhesins (Epas). To reveal the structure-function relationships within the entire Epa family, we have performed a large scale functional analysis of the adhesion (A) domains of 17 Epa paralogs in combination with three-dimensional structural studies of selected members with cognate ligands. Our study shows that most EpaA domains exert lectin-like functions and together recognize a wide variety of glycans with terminal galactosides for conferring epithelial cell adhesion. We further identify several conserved and variable structural features within the diverse Epa ligand binding pockets, which affect affinity and specificity. These features rationalize why mere phylogenetic relationships within the Epa family are weak indicators for functional classification and explain how Epa-like adhesins have evolved in C. glabrata and related fungal species.",
"The present paper describes the SSM algorithm of protein structure comparison in three dimensions, which includes an original procedure of matching graphs built on the protein's secondary-structure elements, followed by an iterative three-dimensional alignment of protein backbone Calpha atoms. The SSM results are compared with those obtained from other protein comparison servers, and the advantages and disadvantages of different scores that are used for structure recognition are discussed. A new score, balancing the r.m.s.d. and alignment length Nalign, is proposed. It is found that different servers agree reasonably well on the new score, while showing considerable differences in r.m.s.d. and Nalign.",
"Glycosylphosphatidylinositol (GPI)-anchored proteins are an important class of cell wall proteins in Candida albicans because of their localization and their function, even if more than half of them have no characterized homolog in the databases. In this study, we focused on the IFF protein family, investigating their exposure on the cell surface and the sequences that determine their subcellular localization. Protein localization and surface exposure were monitored by the addition of a V5 tag on all members of the family. The data obtained using the complete proteins showed for Iff3 (or -9), Iff5, Iff6, and Iff8 a covalent linkage to the β-1,6-glucan network but, remarkably, showed that Iff2/Hyr3 was linked through disulfide bridges or NaOH-labile bonds. However, since some proteins of the Iff family were undetectable, we designed chimeric constructions using the last 60 amino acids of these proteins to test the localization signal. These constructions showed a β-1,6-glucan linkage for Iff1/Rbr3, Iff2/Hyr3, Iff4 and Iff7/Hyr4 C-terminal-Iff5 fusion proteins, and a membrane localization for the Iff10/Flo9 C terminus-Iff5 fusion protein. Immunofluorescence analyses coupled to these cell fraction data confirmed the importance of the length of the central serine/threonine-rich region for cell surface exposure. Further analysis of the Iff2/Hyr3 linkage to the cell surface showed for the first time that a serine/threonine central region of a GPI-anchored protein may be responsible for the disulfide and the NaOH bonds to the glucan and glycoproteins network and may also override the signal of the proximal ω site region."
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COVID-19 Vaccination Acceptance among Chinese Citizens: A Cross-Sectional Study
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The outbreak of coronavirus disease 2019 (COVID-19) has led to numerous tragic deaths all over the world. Great efforts have been made by worldwide nations for COVID-19 targeted vaccine development since the disease outbreak. In January 2021, the Chinese government started to provide free vaccination among nationwide communities, which was optional for citizens. As no evidence has been provided so far regarding COVID-19 vaccination acceptance since the initiation of nationwide vaccination, this study aims to investigate COVID-19 vaccination acceptance among Chinese citizens as well as its associated factors as an attempt to bridge such gap embedded in the current literature. An anonymous cross-sectional study was conducted online in March and April 2021 among adults, with the survey questionnaire designed based on the framework of the health belief model (HBM). Information on socio-demographics, risk perception, past pandemic-related experience, awareness of vaccination as well as acceptance of COVID-19 vaccination were collected. Chi-squared test and multi-level regression were performed to distinguish the acceptance between different groups as well as to identify the significant predictors. A total of 3940 participants completed the survey, with 90.6% of the participants reporting strong willingness to get vaccinated. A list of factors were found to be significantly associated with individuals' acceptance of vaccination, including the region of residence, ethnicity, annual income, whether or not they had experienced a major pandemic event in the past, risk perception of the COVID-19 as well as the awareness of receiving vaccination. Safety concerns about the vaccine (27.7%), concerns about receiving vaccination immediately after newly developed vaccines were released into the market (22.4%) as well as concerns about the potential side effects induced by vaccination (22.1%) were identified as the primary reasons of residents' resistance against vaccination. Overall, residents demonstrated strong willingness to receive vaccination against COVID-19 in China. However, the improvement of vaccination-related knowledge among Chinese residents should be highlighted as a critical strategy to facilitate the penetration of nationwide vaccination in order to ultimately achieve the establishment of herd immunity in China.
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"This study attempts to understand coronavirus disease 2019 (COVID-19) vaccine demand and hesitancy by assessing the public's vaccination intention and willingness-to-pay (WTP). Confidence in COVID-19 vaccines produced in China and preference for domestically-made or foreign-made vaccines was also investigated. A nationwide cross-sectional, self-administered online survey was conducted on 1-19 May 2020. The health belief model (HBM) was used as a theoretical framework for understanding COVID-19 vaccination intent and WTP. A total of 3,541 complete responses were received. The majority reported a probably yes intent (54.6%), followed by a definite yes intent (28.7%). The perception that vaccination decreases the chances of getting COVID-19 under the perceived benefit construct (OR = 3.14, 95% CI 2.05-4.83) and not being concerned about the efficacy of new COVID-19 vaccines under the perceived barriers construct (OR = 1.65, 95% CI 1.31-2.09) were found to have the highest significant odds of a definite intention to take the COVID-19 vaccine. The median (interquartile range [IQR]) of WTP for COVID-19 vaccine was CNY¥200/US$28 (IQR CNY¥100-500/USD$14-72). The highest marginal WTP for the vaccine was influenced by socio-economic factors. The majority were confident (48.7%) and completely confident (46.1%) in domestically-made COVID-19 vaccine. 64.2% reported a preference for a domestically-made over foreign-made COVID-19 vaccine. The findings demonstrate the utility of HBM constructs in understanding COVID-19 vaccination intent and WTP. It is important to improve health promotion and reduce the barriers to COVID-19 vaccination.",
"The COVID-19 pandemic continues to adversely affect the U.S., which leads globally in total cases and deaths. As COVID-19 vaccines are under development, public health officials and policymakers need to create strategic vaccine-acceptance messaging to effectively control the pandemic and prevent thousands of additional deaths. Using an online platform, we surveyed the U.S. adult population in May 2020 to understand risk perceptions about the COVID-19 pandemic, acceptance of a COVID-19 vaccine, and trust in sources of information. These factors were compared across basic demographics. Of the 672 participants surveyed, 450 (67%) said they would accept a COVID-19 vaccine if it is recommended for them. Males (72%) compared to females, older adults (≥55 years; 78%) compared to younger adults, Asians (81%) compared to other racial and ethnic groups, and college and/or graduate degree holders (75%) compared to people with less than a college degree were more likely to accept the vaccine. When comparing reported influenza vaccine uptake to reported acceptance of the COVID-19 vaccine: 1) participants who did not complete high school had a very low influenza vaccine uptake (10%), while 60% of the same group said they would accept the COVID-19 vaccine; 2) unemployed participants reported lower influenza uptake and lower COVID-19 vaccine acceptance when compared to those employed or retired; and, 3) Black Americans reported lower influenza vaccine uptake and lower COVID-19 vaccine acceptance than all other racial groups reported in our study. Lastly, we identified geographic differences with Department of Health and Human Services (DHHS) regions 2 (New York) and 5 (Chicago) reporting less than 50 percent COVID-19 vaccine acceptance. Although our study found a 67% acceptance of a COVID-19 vaccine, there were noticeable demographic and geographical disparities in vaccine acceptance. Before a COVID-19 vaccine is introduced to the U.S., public health officials and policymakers must prioritize effective COVID-19 vaccine-acceptance messaging for all Americans, especially those who are most vulnerable.",
"In December 2019, a cluster of patients with pneumonia of unknown cause was linked to a seafood wholesale market in Wuhan, China. A previously unknown betacoronavirus was discovered through the use of unbiased sequencing in samples from patients with pneumonia. Human airway epithelial cells were used to isolate a novel coronavirus, named 2019-nCoV, which formed a clade within the subgenus sarbecovirus, Orthocoronavirinae subfamily. Different from both MERS-CoV and SARS-CoV, 2019-nCoV is the seventh member of the family of coronaviruses that infect humans. Enhanced surveillance and further investigation are ongoing. (Funded by the National Key Research and Development Program of China and the National Major Project for Control and Prevention of Infectious Disease in China.).",
"Vaccine regulation in China meets World Health Organization standards, but China's vaccine industry and immunization program have some characteristics that differ from other countries. We described the history, classification, supply and prices of vaccines available and used in China, compared with high-and middle-incomes countries to illustrate the development of Chinese vaccine industry and immunization program. Immunization policy documents were obtained from the State Council and the National Health and Family Planning Commission (NHFPC). Numbers of doses of vaccines released in China were obtained from the Biologicals Lot Release Program of the National Institutes for Food and Drug Control (NIFDC). Vaccine prices were obtained from Chinese Central Government Procurement (CCGP). International data were collected from US CDC, Public Health England, European CDC, WHO, and UNICEF. Between 2007 and 2015, the annual supply of vaccines in China ranged between 666 million and 1,190 million doses, with most doses produced domestically. The government's Expanded Program on Immunization (EPI) prevents 12 vaccine preventable diseases (VPD) through routine immunization. China produces vaccines that are in common use globally; however, the number of routinely-prevented diseases is fewer than in high- and middle-income countries. Contract prices for program (EPI) vaccines ranged from 0.1 to 5.7 US dollars per dose - similar to UNICEF prices. Contract prices for private-market vaccines ranged from 2.4 to 102.9 US dollars per dose - often higher than prices for comparable US, European, and UNICEF vaccines. China is a well-regulated producer of vaccines, but some vaccines that are important globally are not included in China's EPI system in China. Sustained and coordinated effort will be required to bring Chinese vaccine industry and EPI into an era of global leadership.",
"The ongoing outbreak of the novel coronavirus disease (COVID-19) that occurred in China is rapidly spreading globally. China's bond and strict containment measures have been proved (in practice) to significantly reduce the spread of the epidemic. This was obtained through the use of emergency control measures in the epidemic areas and the integration of resources from multiple systems, including business, community, technology, education, and transportation, across the country. In order to better understand how China has managed to reduce the public health and economic impacts of the COVID-19 epidemic, this editorial systematically reviews the specific measures for infection prevention and control of the disease. The best practices for COVID-19 eradication in China provide evidence-based strategies that could be replicated in other countries."
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Assessing COVID-19 Vaccine Booster Hesitancy in a National Sample
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Given the emergence of breakthrough infections, new variants, and concerns of waning immunity from the primary COVID-19 vaccines, booster shots emerged as a viable option to shore-up protection against COVID-19. Following the recent authorization of vaccine boosters among vulnerable Americans, this study aims to assess COVID-19 vaccine booster hesitancy and its associated factors in a nationally representative sample. A web-based 48-item psychometric valid survey was used to measure vaccine literacy, vaccine confidence, trust, and general attitudes towards vaccines. Data were analyzed through Chi-square (with a post hoc contingency table analysis) and independent-sample
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[
"There can be little doubt about the ethical imperative to ensure adequate vaccination uptake against certain infectious diseases. In the face of vaccine refusal, health authorities and providers instinctively appeal to coercive approaches or increased education as methods to ensure adequate vaccine uptake. Recently, some have argued that public fear around Ebola should be used as an opportunity for such approaches, should an Ebola vaccine become available. In this article, the author describes the difficulties associated with coercion and education when addressing vaccine opposition. Both coercion and education can cause opposite effects than intended in certain circumstances. The correct area of focus is to address the breakdown in trust within clinical relationships. The author presents suggestions for an approach towards vaccine refusal that may be more promising.",
"Two US Food and Drug Administration (FDA)-authorized coronavirus disease 2019 (COVID-19) mRNA vaccines, BNT162b2 (Pfizer/BioNTech) and mRNA-1273 (Moderna), have demonstrated high efficacy in large phase 3 randomized clinical trials. It is important to assess their effectiveness in a real-world setting. This is a retrospective analysis of 136,532 individuals in the Mayo Clinic health system (Arizona, Florida, Iowa, Minnesota, and Wisconsin) with PCR testing data between December 1, 2020 and April 20, 2021. We compared clinical outcomes for a vaccinated cohort of 68,266 individuals who received at least one dose of either vaccine (nBNT162b2 = 51,795; nmRNA-1273 = 16,471) and an unvaccinated control cohort of 68,266 individuals propensity matched based on relevant demographic, clinical, and geographic features. We estimated real-world vaccine effectiveness by comparing incidence rates of positive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PCR testing and COVID-19-associated hospitalization and intensive care unit (ICU) admission starting 7 days after the second vaccine dose. The real-world vaccine effectiveness of preventing SARS-CoV-2 infection was 86.1% (95% confidence interval [CI]: 82.4%-89.1%) for BNT162b2 and 93.3% (95% CI: 85.7%-97.4%) for mRNA-1273. BNT162b2 and mRNA-1273 were 88.8% (95% CI: 75.5%-95.7%) and 86.0% (95% CI: 71.6%-93.9%) effective in preventing COVID-19-associated hospitalization. Both vaccines were 100% effective (95% CIBNT162b2: 51.4%-100%; 95% CImRNA-1273: 43.3%-100%) in preventing COVID-19-associated ICU admission. BNT162b2 and mRNA-1273 are effective in a real-world setting and are associated with reduced rates of SARS-CoV-2 infection and decreased burden of COVID-19 on the healthcare system. This study was funded by nference.",
"Several reports claim that the purported similarity between syncytin-1 and the SARS-CoV-2 spike protein may induce immune cross-reactivity resulting in female sterility. We used frozen embryo transfer as a model for comparing the implantation rates between SARS-CoV-2 vaccine seropositive, infection seropositive, and seronegative women. No difference was found in serum hCG documented implantation rates or sustained implantation rates between the three groups. Reports claiming that COVID-19 vaccines or illness cause female sterility are unfounded.",
"Throughout the COVID-19 pandemic, health care personnel (HCP) have been at high risk for exposure to SARS-CoV-2, the virus that causes COVID-19, through patient interactions and community exposure (1). The Advisory Committee on Immunization Practices recommended prioritization of HCP for COVID-19 vaccination to maintain provision of critical services and reduce spread of infection in health care settings (2). Early distribution of two mRNA COVID-19 vaccines (Pfizer-BioNTech and Moderna) to HCP allowed assessment of the effectiveness of these vaccines in a real-world setting. A test-negative case-control study is underway to evaluate mRNA COVID-19 vaccine effectiveness (VE) against symptomatic illness among HCP at 33 U.S. sites across 25 U.S. states. Interim analyses indicated that the VE of a single dose (measured 14 days after the first dose through 6 days after the second dose) was 82% (95% confidence interval [CI] = 74%-87%), adjusted for age, race/ethnicity, and underlying medical conditions. The adjusted VE of 2 doses (measured ≥7 days after the second dose) was 94% (95% CI = 87%-97%). VE of partial (1-dose) and complete (2-dose) vaccination in this population is comparable to that reported from clinical trials and recent observational studies, supporting the effectiveness of mRNA COVID-19 vaccines against symptomatic disease in adults, with strong 2-dose protection.",
"Data from randomized clinical trials and real-world observational studies show that all three COVID-19 vaccines currently authorized for emergency use by the Food and Drug Administration* are safe and highly effective for preventing COVID-19-related serious illness, hospitalization, and death (1,2). Studies of vaccine effectiveness (VE) for preventing new infections and hospitalizations attributable to SARS-CoV-2, the virus that causes COVID-19), particularly as the B.1.617.2 (Delta) variant has become predominant, are limited in the United States (3). In this study, the New York State Department of Health linked statewide immunization, laboratory testing, and hospitalization databases for New York to estimate rates of new laboratory-confirmed COVID-19 cases and hospitalizations by vaccination status among adults, as well as corresponding VE for full vaccination in the population, across all three authorized vaccine products. During May 3-July 25, 2021, the overall age-adjusted VE against new COVID-19 cases for all adults declined from 91.8% to 75.0%. During the same period, the overall age-adjusted VE against hospitalization was relatively stable, ranging from 89.5% to 95.1%. Currently authorized vaccines have high effectiveness against COVID-19 hospitalization, but effectiveness against new cases appears to have declined in recent months, coinciding with the Delta variant's increase from <2% to >80% in the U.S. region that includes New York and relaxation of masking and physical distancing recommendations. To reduce new COVID-19 cases and hospitalizations, these findings support the implementation of a layered approach centered on vaccination, as well as other prevention strategies such as masking and physical distancing.",
"In pharmacokinetic data analysis, it is frequently necessary to select the number of exponential terms in a polyexponential expression used to describe the concentration-time relationship. The performance characteristics of several selection criteria, the Akaike Information Criterion (AIC), and the Schwarz Criterion (SC), and the F test (alpha = 0.05), were examined using Monte Carlo simulations. In particular, the ability of these criteria to select the correct model, to select a model allowing estimation of pharmacokinetic parameters with small bias and good precision, and to select a model allowing precise predictions of concentration was evaluated. To some extent interrelationships among these procedures is explainable. Results indicate that the F test tends to choose the simpler model more often than does either the AIC or SC, even when the more complex model is correct. Also, the F test is more sensitive to deficient sampling designs. Clearance estimates are generally very robust to the choice of the wrong model. Other pharmacokinetic parameters are more sensitive to model choice, particularly the apparent elimination rate constant. Prediction of concentrations is generally more precise when the correct model is chosen. The tendency for the F test (alpha = 0.05) to choose the simpler model must be considered relative to the objectives of the study."
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How do I find the number of Russian words in a sentence?
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9 Russian
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[
"Croatian viticulture was most extensive at the beginning of the 20th century, when about 400 varieties were in use. Autochthonous varieties are the result of spontaneous hybridization from the pre-phylloxera era and are still cultivated today on about 35 % of vineyard area, while some exist only in repositories. We present what is the most comprehensive genetic analysis of all major Croatian national repositories, with a large number of microsatellite, or simple sequence repeat (SSR) markers, and it is also the first study to apply single nucleotide polymorphism (SNP) markers. After 212 accessions were fingerprinted, 95 were classified as unique to Croatian germplasm. Genetic diversity of Croatian germplasm is rather high considering its size. SNP markers proved useful for fingerprinting but less informative and practical than SSRs. Analysis of the genetic structure showed that Croatian germplasm is predominantly part of the Balkan grape gene pool. A high number of admixed varieties and synonyms is a consequence of complex pedigrees and migrations. Parentage analysis confirmed 24 full parentages, as well as 113 half-kinships. Unexpectedly, several key genitors could not be detected within the present Croatian germplasm. The low number of reconstructed parentages (19%) points to severe genetic erosion and stresses the importance of germplasm repositories.",
"The decreasing cost along with rapid progress in next-generation sequencing and related bioinformatics computing resources has facilitated large-scale discovery of SNPs in various model and nonmodel plant species. Large numbers and genome-wide availability of SNPs make them the marker of choice in partially or completely sequenced genomes. Although excellent reviews have been published on next-generation sequencing, its associated bioinformatics challenges, and the applications of SNPs in genetic studies, a comprehensive review connecting these three intertwined research areas is needed. This paper touches upon various aspects of SNP discovery, highlighting key points in availability and selection of appropriate sequencing platforms, bioinformatics pipelines, SNP filtering criteria, and applications of SNPs in genetic analyses. The use of next-generation sequencing methodologies in many non-model crops leading to discovery and implementation of SNPs in various genetic studies is discussed. Development and improvement of bioinformatics software that are open source and freely available have accelerated the SNP discovery while reducing the associated cost. Key considerations for SNP filtering and associated pipelines are discussed in specific topics. A list of commonly used software and their sources is compiled for easy access and reference."
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what is black rot disease?
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Black rot disease, caused by
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"ABSTRACT Two hundred and seventy-six accessions of mainly Brassica spp. were screened for resistance to Xanthomonas campestris pv. campestris races. In Brassica oleracea (C genome), the majority of accessions were susceptible to all races, but 43% showed resistance to one or more of the rare races (2, 3, 5, and 6) and a single accession showed partial resistance to races 1, 3, 5, and 6. Further searches for resistance to races 1 and 4, currently the most important races worldwide, and race 6, the race with the widest host range, were made in accessions representing the A and B genomes. Strong resistance to race 4 was frequent in B. rapa (A genome) and B. napus (AC genome), indicating an A genome origin. Resistance to races 1 and 4 was present in a high proportion of B. nigra (B genome) and B. carinata (BC genome) accessions, indicating a B genome origin. B. juncea (AB genome) was the most resistant species, showing either strong resistance to races 1 and 4 or quantitative resistance to all races. Potentially race-nonspecific resistance was also found, but at a lower frequency, in B. rapa, B. nigra, and B. carinata. The combination of race-specific and race-nonspecific resistance could provide durable control of black rot of crucifers.",
"Black rot, caused by Xanthomonas campestris pv. campestris (Xcc), is a seed borne disease of Brassicaceae. Eleven pathogenic races have been identified based on the phenotype interaction pattern of differential brassica cultivars inoculated with different strains. Race 1 and 4 are the two most frequent races found in Brassica oleracea crops. In this study, a PCR molecular diagnostic tool was developed for the identification of Xcc races 1 and 4 of this pathogen. Whole genomic sequences of races 1, 3, 4 and 9 and sequences of three other Xanthomonas pathovars/species (X. campestris pv. incanae (Xci), X. campestris pv. raphani (Xcr) and X.euvesicatoria (Xev) were aligned to identify variable regions among races. To develop specific markers for races 1 and 4, primers were developed from a region where sequences were dissimilar in other races. Sequence-characterized amplified regions (SCAR) and insertion or deletion of bases (InDel) were used to develop each specific set of primers. The specificity of the selected primers was confirmed by PCR tests using genomic DNA of seven different Xcc races, two strains of X. campestris pathovars and other species of bacteria. Bacterial samples of the races 1 and 4 isolates were collected from artificially inoculated cabbage leaves to conduct bio-PCR. Bio-PCR successfully detected the two Xcc isolates. By using our race-specific markers, a potential race 1 strain from the existing Korean Xcc collection was identified. The Xcc race 1 and 4-specific markers developed in this study are novel and can potentially be used for rapid detection of Xcc races through PCR.",
"Xanthomonas campestris pv. campestris is the causal agent of black rot on Brassicaceae. The draft genome sequences of strains CFBP 1869 and CFBP 5817 have been determined and are the first ones corresponding to race 1 and race 4 strains, which have a predominant agronomic and economic impact on cabbage cultures worldwide."
] |
Temporal changes in the transcriptome of implant-associated monocytes and macrophages in the foreign body response
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Poly(ethylene glycol) (PEG) hydrogels hold promise for in vivo applications but induce a foreign body response (FBR). While macrophages are key in the FBR, many questions remain. This study investigates temporal changes in the transcriptome of implant-associated monocytes and macrophages. Proinflammatory pathways are upregulated in monocytes compared to control monocytes but subside by day 28. Macrophages are initially proinflammatory but shift to a profibrotic state by day 14, coinciding with fibrous capsule emergence. Next, this study assesses the origin of macrophages responsible for fibrous encapsulation using wildtype, C-C Motif Chemokine Receptor 2 (CCR2)
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"The recruitment of monocyte-macrophages into the artery wall is one of the earliest events in the pathogenesis of atherosclerosis. Monocyte chemoattractant protein 1 (MCP-1) is a potent monocyte chemoattractant secreted by many cells in vitro, including vascular smooth muscle and endothelial cells. To test whether it is expressed in the artery in vivo, we used Northern blot analysis, in situ hybridization, and immunocytochemistry to study the expression of MCP-1 in normal and atherosclerotic human and rabbit arteries. Northern blot analysis showed that MCP-1 mRNA could be isolated from rabbit atherosclerotic lesions but not from the intima media of normal animals. Furthermore, MCP-1 mRNA was extracted from macrophage-derived foam cells isolated from arterial lesions of ballooned cholesterol-fed rabbits, whereas alveolar macrophages isolated simultaneously from the same rabbits did not express MCP-1 mRNA. MCP-1 mRNA was detected by in situ hybridization in macrophage-rich regions of both human and rabbit atherosclerotic lesions. No MCP-1 mRNA was found in sublesional medial smooth muscle cells or in normal arteries. By using immunocytochemistry, MCP-1 protein was demonstrated in human lesions, again only in macrophage-rich regions. Immunostaining of the serial sections with an antiserum against malondialdehyde-modified low density lipoprotein indicated the presence of oxidized low density lipoprotein indicated the presence of oxidized low density lipoprotein and/or other oxidation-specific lipid-protein adducts in the same areas that contained macrophages and MCP-1. We conclude that (i) MCP-1 is strongly expressed in a small subset of cells in macrophage-rich regions of human and rabbit atherosclerotic lesions and (ii) MCP-1 may, therefore, play an important role in the ongoing recruitment of monocyte-macrophages into developing lesions in vivo.",
"Invasive tumor dissemination in vitro and in vivo involves the proteolytic degradation of ECM barriers. This process, however, is only incompletely attenuated by protease inhibitor-based treatment, suggesting the existence of migratory compensation strategies. In three-dimensional collagen matrices, spindle-shaped proteolytically potent HT-1080 fibrosarcoma and MDA-MB-231 carcinoma cells exhibited a constitutive mesenchymal-type movement including the coclustering of beta 1 integrins and MT1-matrix metalloproteinase (MMP) at fiber bindings sites and the generation of tube-like proteolytic degradation tracks. Near-total inhibition of MMPs, serine proteases, cathepsins, and other proteases, however, induced a conversion toward spherical morphology at near undiminished migration rates. Sustained protease-independent migration resulted from a flexible amoeba-like shape change, i.e., propulsive squeezing through preexisting matrix gaps and formation of constriction rings in the absence of matrix degradation, concomitant loss of clustered beta 1 integrins and MT1-MMP from fiber binding sites, and a diffuse cortical distribution of the actin cytoskeleton. Acquisition of protease-independent amoeboid dissemination was confirmed for HT-1080 cells injected into the mouse dermis monitored by intravital multiphoton microscopy. In conclusion, the transition from proteolytic mesenchymal toward nonproteolytic amoeboid movement highlights a supramolecular plasticity mechanism in cell migration and further represents a putative escape mechanism in tumor cell dissemination after abrogation of pericellular proteolysis.",
"The chemokines are a large family of cytokines that control the recruitment of leukocytes in immune and inflammatory responses. We describe the isolation of a novel murine CC chemokine that, based on its biological and structural features, we have named monocyte chemoattractant protein (MCP)-5. MCP-5 mapped to the CC chemokine cluster on mouse chromosome 11 and was most closely related to human MCP-1 in structure (66% amino acid identity). Purified recombinant MCP-5 protein was a potent chemoattractant for peripheral blood monocytes, was only weakly active on eosinophils at high doses, and was inactive on neutrophils. MCP-5 induced a calcium flux in peripheral blood mononuclear cells, but not in purified murine eosinophils or neutrophils. Consistent with these results, MCP-5 induced a calcium flux in human embryonic kidney (HEK)-293 cells transfected with human and murine CCR2, a CC chemokine receptor expressed on monocytes. MCP-5 did not induce a calcium flux in HEK-293 cells transfected with CCR1, CCR3, or CCR5. Constitutive expression of MCP-5 mRNA was detected predominantly in lymph nodes, and its expression was markedly induced in macrophages activated in vitro and in vivo. Moreover, MCP-5 expression was up-regulated in the lungs of mice following aerosolized antigen challenge of sensitized mice, and during the host response to infection with Nippostrongylus brasiliensis. These data indicate that MCP-5 is a novel and potent monocyte active chemokine that is involved in allergic inflammation and the host response to pathogens.",
"Monocyte chemoattractant protein-1 (MCP-1) is a potent agonist for mononuclear leukocytes and has been implicated in the pathogenesis of atherosclerosis and granulomatous lung disease. To determine the role of MCP-1 and related family members in vivo, we used homologous recombination in embryonic stem cells to generate mice with a targeted disruption of C-C chemokine receptor 2 (CCR2), the receptor for MCP-1. CCR2-/- mice were born at the expected Mendelian ratios and developed normally. In response to thioglycollate, the recruitment of peritoneal macrophages decreased selectively. In in vitro chemotaxis assays, CCR2-/- leukocytes failed to migrate in response to MCP-1. Granulomatous lung disease was induced in presensitized mice by embolization with beads coupled to purified protein derivative (PPD) of Mycobacterium bovis. As compared with wild-type littermates, CCR2-/- mice had a decrease in granuloma size accompanied by a dramatic decrease in the level of interferon gamma in the draining lymph nodes. Production of interferon gamma was also decreased in PPD-sensitized splenocytes from CCR2-/- mice and in naive splenocytes activated by concanavalin A. We conclude that CCR2-/- mice have significant defects in both delayed-type hypersensitivity responses and production of Th1-type cytokines. These data suggest an important and unexpected role for CCR2 activation in modulating the immune response, as well as in recruiting monocytes/macrophages to sites of inflammation.",
"Intracerebral hemorrhage (ICH) is a devastating type of stroke that lacks a specific treatment. An intense immune response develops after ICH, which contributes to neuronal injury, disability, and death. However, the specific mediators of inflammation-induced injury remain unclear. The objective of the present study was to determine whether blood-derived CCR2+ Ly6C(hi) inflammatory monocytes contribute to disability. ICH was induced in mice and the resulting inflammatory response was quantified using flow cytometry, confocal microscopy, and neurobehavioral testing. Importantly, blood-derived monocytes were distinguished from resident microglia by differential CD45 staining and by using bone marrow chimeras with fluorescent leukocytes. After ICH, blood-derived CCR2+ Ly6C(hi) inflammatory monocytes trafficked into the brain, outnumbered other leukocytes, and produced tumor necrosis factor. Ccr2(-/-) mice, which have few circulating inflammatory monocytes, exhibited better motor function following ICH than control mice. Chimeric mice with wild-type CNS cells and Ccr2(-/-) hematopoietic cells also exhibited early improvement in motor function, as did wild-type mice after inflammatory monocyte depletion. These findings suggest that blood-derived inflammatory monocytes contribute to acute neurological disability. To determine the translational relevance of our experimental findings, we examined CCL2, the principle ligand for the CCR2 receptor, in ICH patients. Serum samples from 85 patients were collected prospectively at two hospitals. In patients, higher CCL2 levels at 24 h were independently associated with poor functional outcome at day 7 after adjusting for potential confounding variables. Together, these findings suggest that inflammatory monocytes worsen early disability after murine ICH and may represent a therapeutic target for patients.",
"Dense fibrosis and a robust macrophage infiltrate are key therapeutic barriers in pancreatic ductal adenocarcinoma (PDAC). CD40 activation can circumvent these barriers by inducing macrophages, originating from peripheral blood monocytes, to deplete fibrosis. The precise mechanism and therapeutic implications of this antifibrotic activity, though, remain unclear. Here, we report that IFNγ and CCL2 released systemically in response to a CD40 agonist cooperate to redirect a subset of Ly6C(+)CCR2(+)monocytes/macrophages to infiltrate tumors and deplete fibrosis. Whereas CCL2 is required for Ly6C(+)monocyte/macrophage infiltration, IFNγ is necessary for tumor-infiltrating monocytes/macrophages to shift the profile of matrix metalloproteinases (MMP) in tumors, leading to MMP-dependent fibrosis degradation. In addition, MMP13-dependent loss of extracellular matrix components induced by a CD40 agonist increased PDAC sensitivity to chemotherapy. Our findings demonstrate that fibrosis in PDAC is a bidirectional process that can be rapidly altered by manipulating a subset of tumor-infiltrating monocytes, leading to enhanced chemotherapy efficacy. We report that CD40 agonists improve chemotherapy efficacy in pancreatic carcinoma by redirecting tumor-infiltrating monocytes/macrophages to induce fibrosis degradation that is dependent on MMPs. These findings provide novel insight into the plasticity of monocytes/macrophages in cancer and their capacity to regulate fibrosis and modulate chemotherapy efficacy in pancreatic carcinoma.",
"Here, we aimed to investigate migration of a model tumor cell line (HT-1080 fibrosarcoma cells, HT-1080s) using synthetic biomaterials to systematically vary peptide ligand density and substrate stiffness. A range of substrate elastic moduli were investigated by using poly(ethylene glycol) (PEG) hydrogel arrays (0.34 - 17 kPa) and self-assembled monolayer (SAM) arrays (~0.1-1 GPa), while cell adhesion was tuned by varying the presentation of Arg-Gly-Asp (RGD)-containing peptides. HT-1080 motility was insensitive to cell adhesion ligand density on RGD-SAMs, as they migrated with similar speed and directionality for a wide range of RGD densities (0.2-5% mol fraction RGD). Similarly, HT-1080 migration speed was weakly dependent on adhesion on 0.34 kPa PEG surfaces. On 13 kPa surfaces, a sharp initial increase in cell speed was observed at low RGD concentration, with no further changes observed as RGD concentration was increased further. An increase in cell speed ~ two-fold for the 13 kPa relative to the 0.34 kPa PEG surface suggested an important role for substrate stiffness in mediating motility, which was confirmed for HT-1080s migrating on variable modulus PEG hydrogels with constant RGD concentration. Notably, despite ~ two-fold changes in cell speed over a wide range of moduli, HT-1080s adopted rounded morphologies on all surfaces investigated, which contrasted with well spread primary human mesenchymal stem cells (hMSCs). Taken together, our results demonstrate that HT-1080s are morphologically distinct from primary mesenchymal cells (hMSCs) and migrate with minimal dependence on cell adhesion for surfaces within a wide range of moduli, whereas motility is strongly influenced by matrix mechanical properties.",
"Cancer cells can invade three-dimensional matrices by distinct mechanisms, recently defined by their dependence on extracellular proteases, including matrix metalloproteinases. Upon treatment with protease inhibitors, some tumour cells undergo a 'mesenchymal to amoeboid' transition that allows invasion in the absence of pericellular proteolysis and matrix degradation. We show here that in HT1080 cells, this transition is associated with weakened integrin-dependent adhesion, consistently reduced cell surface expression of the alpha2beta1 integrin collagen receptor and impaired signalling downstream, as judged by reduced autophosphorylation of focal adhesion kinase (FAK). On examining cancer cells that use defined invasion strategies, we show that distinct from mesenchymal invasion, amoeboid invasion is independent of intracellular calpain 2 proteolytic activity that is usually needed for turnover of integrin-linked adhesions during two-dimensional planar migration. Moreover, an inhibitor of Rho/ROCK signalling, which specifically impairs amoeboid-like invasion, restores cell surface expression of alpha2beta1 integrin, downstream FAK autophosphorylation and calpain 2 sensitivity--features of mesenchymal invasion. These findings link weakened integrin function to a lack of requirement for calpain 2-mediated integrin adhesion turnover during amoeboid invasion. In keeping with the need for integrin adhesion turnover, mesenchymal invasion is uniquely sensitive to Src inhibitors. Thus, the need for a major pathway that controls integrin adhesion turnover defines and distinguishes cancer cell invasion strategies.",
"Cell migration on 2D surfaces is governed by a balance between counteracting tractile and adhesion forces. Although biochemical factors such as adhesion receptor and ligand concentration and binding, signaling through cell adhesion complexes, and cytoskeletal structure assembly/disassembly have been studied in detail in a 2D context, the critical biochemical and biophysical parameters that affect cell migration in 3D matrices have not been quantitatively investigated. We demonstrate that, in addition to adhesion and tractile forces, matrix stiffness is a key factor that influences cell movement in 3D. Cell migration assays in which Matrigel density, fibronectin concentration, and beta1 integrin binding are systematically varied show that at a specific Matrigel density the migration speed of DU-145 human prostate carcinoma cells is a balance between tractile and adhesion forces. However, when biochemical parameters such as matrix ligand and cell integrin receptor levels are held constant, maximal cell movement shifts to matrices exhibiting lesser stiffness. This behavior contradicts current 2D models but is predicted by a recent force-based computational model of cell movement in a 3D matrix. As expected, this 3D motility through an extracellular environment of pore size much smaller than cellular dimensions does depend on proteolytic activity as broad-spectrum matrix metalloproteinase (MMP) inhibitors limit the migration of DU-145 cells and also HT-1080 fibrosarcoma cells. Our experimental findings here represent, to our knowledge, discovery of a previously undescribed set of balances of cell and matrix properties that govern the ability of tumor cells to migration in 3D environments.",
"A mathematical model, based on thermodynamics, was developed to demonstrate how substrate rigidity influences cell morphology and migration. The mechanisms by which substrate rigidity are translated into cell-morphological changes and cell movement are described. The model takes into account the competition between the elastic energies in the cell-substrate system and work of adhesion at the cell periphery. The cell morphology and migration are dictated by the minimum of the total free energy of the cell-substrate system. By using this model, reported experimental observations on cell morphological changes and migration can be better understood with a theoretical basis. In addition, these observations can be more accurately correlated with the variation of substrate rigidity. This study indicates that the activity of the adherent cell is dependent not only on the substrate rigidity but also is correlated with the relative rigidity between the cell and substrate. Moreover, the study suggests that the cell stiffness can be estimated based on the substrate stiffness corresponding to the change of trend in morphological stability."
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Vaccine hesitancy in visible minorities: A scoping review
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Vaccine hesitancy is one of the top ten greatest threats to global health. During the COVID-19 era, vaccine hesitancy poses substantial risks, especially in visible minorities, who are disproportionately affected by the pandemic. Although evidence of vaccine hesitancy exists, there is minimal focus on visible minorities and the reasons for hesitancy in this group are unclear. Identifying these populations and their reasons for vaccine hesitancy is crucial in improving vaccine uptake and curbing the spread of COVID-19. This scoping review follows a modified version of the Arksey and O'Malley strategy. Using comprehensive search strategies, advanced searches were conducted on Medline, CINAHL, and PubMed databases to acquire relevant articles. Full-text reviews using inclusion and exclusion criteria were performed to extract themes of vaccine hesitancy. Themes were grouped into factors using thematic qualitative analysis and were objectively confirmed by principal component analysis (PCA). To complement both analyses, a word cloud of titles and abstracts for the final articles was generated. This study included 71 articles. Themes were grouped into 8 factors and the top 3 recurring factors were safety and effectiveness of the vaccine, mistrust, and socioeconomic characteristics. Shedding light on these factors could help mitigate health inequities and increase overall vaccine uptake worldwide through interventions and policies targeted at these factors. Ultimately, this would help achieve global herd immunity.
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"We investigate how the anticipation of COVID-19 vaccines affects voluntary social distancing. In a large-scale preregistered survey experiment with a representative sample, we study whether providing information about the safety, effectiveness, and availability of COVID-19 vaccines affects the willingness to comply with public health guidelines. We find that vaccine information reduces peoples' voluntary social distancing, adherence to hygiene guidelines, and their willingness to stay at home. Getting positive information on COVID-19 vaccines induces people to believe in a swifter return to normal life. The results indicate an important behavioral drawback of successful vaccine development: An increased focus on vaccines can lower compliance with public health guidelines and accelerate the spread of infectious disease. The results imply that, as vaccinations roll out and the end of a pandemic feels closer, policies aimed at increasing social distancing will be less effective, and stricter policies might be required.",
"Given the results from early trials, COVID-19 vaccines will be available by 2021. However, little is known about what Americans think of getting immunized with a COVID-19 vaccine. Thus, the purpose of this study was to conduct a comprehensive and systematic national assessment of COVID-19 vaccine hesitancy in a community-based sample of the American adult population. A multi-item valid and reliable questionnaire was deployed online via mTurk and social media sites to recruit U.S. adults from the general population. A total of 1878 individuals participated in the study where the majority were: females (52%), Whites (74%), non-Hispanic (81%), married (56%), employed full time (68%), and with a bachelor's degree or higher (77%). The likelihood of getting a COVID-19 immunization in the study population was: very likely (52%), somewhat likely (27%), not likely (15%), definitely not (7%), with individuals who had lower education, income, or perceived threat of getting infected being more likely to report that they were not likely/definitely not going to get COVID-19 vaccine (i.e., vaccine hesitancy). In unadjusted group comparisons, compared to their counterparts, vaccine hesitancy was higher among African-Americans (34%), Hispanics (29%), those who had children at home (25%), rural dwellers (29%), people in the northeastern U.S. (25%), and those who identified as Republicans (29%). In multiple regression analyses, vaccine hesitancy was predicted significantly by sex, education, employment, income, having children at home, political affiliation, and the perceived threat of getting infected with COVID-19 in the next 1 year. Given the high prevalence of COVID-19 vaccine hesitancy, evidence-based communication, mass media strategies, and policy measures will have to be implemented across the U.S. to convert vaccines into vaccinations and mass immunization with special attention to the groups identified in this study.",
"Availability of reliable and valid race/ethnicity data is essential for monitoring and improving quality of care for minority groups. We explore the limitations and challenges posed by existing means of data collection and discuss issues that need to be considered as the data are analyzed and used.",
"Coronavirus disease 2019 (COVID-19) was declared a pandemic in March 2020. Several prophylactic vaccines against COVID-19 are currently in development, yet little is known about people's acceptability of a COVID-19 vaccine. We conducted an online survey of adults ages 18 and older in the United States (n = 2,006) in May 2020. Multivariable relative risk regression identified correlates of participants' willingness to get a COVID-19 vaccine (i.e., vaccine acceptability). Overall, 69% of participants were willing to get a COVID-19 vaccine. Participants were more likely to be willing to get vaccinated if they thought their healthcare provider would recommend vaccination (RR = 1.73, 95% CI: 1.49-2.02) or if they were moderate (RR = 1.09, 95% CI: 1.02-1.16) or liberal (RR = 1.14, 95% CI: 1.07-1.22) in their political leaning. Participants were also more likely to be willing to get vaccinated if they reported higher levels of perceived likelihood getting a COVID-19 infection in the future (RR = 1.05, 95% CI: 1.01-1.09), perceived severity of COVID-19 infection (RR = 1.08, 95% CI: 1.04-1.11), or perceived effectiveness of a COVID-19 vaccine (RR = 1.46, 95% CI: 1.40-1.52). Participants were less likely to be willing to get vaccinated if they were non-Latinx black (RR = 0.81, 95% CI: 0.74-0.90) or reported a higher level of perceived potential vaccine harms (RR = 0.95, 95% CI: 0.92-0.98). Many adults are willing to get a COVID-19 vaccine, though acceptability should be monitored as vaccine development continues. Our findings can help guide future efforts to increase COVID-19 vaccine acceptability (and uptake if a vaccine becomes available).",
"The COVID-19 pandemic continues to adversely affect the U.S., which leads globally in total cases and deaths. As COVID-19 vaccines are under development, public health officials and policymakers need to create strategic vaccine-acceptance messaging to effectively control the pandemic and prevent thousands of additional deaths. Using an online platform, we surveyed the U.S. adult population in May 2020 to understand risk perceptions about the COVID-19 pandemic, acceptance of a COVID-19 vaccine, and trust in sources of information. These factors were compared across basic demographics. Of the 672 participants surveyed, 450 (67%) said they would accept a COVID-19 vaccine if it is recommended for them. Males (72%) compared to females, older adults (≥55 years; 78%) compared to younger adults, Asians (81%) compared to other racial and ethnic groups, and college and/or graduate degree holders (75%) compared to people with less than a college degree were more likely to accept the vaccine. When comparing reported influenza vaccine uptake to reported acceptance of the COVID-19 vaccine: 1) participants who did not complete high school had a very low influenza vaccine uptake (10%), while 60% of the same group said they would accept the COVID-19 vaccine; 2) unemployed participants reported lower influenza uptake and lower COVID-19 vaccine acceptance when compared to those employed or retired; and, 3) Black Americans reported lower influenza vaccine uptake and lower COVID-19 vaccine acceptance than all other racial groups reported in our study. Lastly, we identified geographic differences with Department of Health and Human Services (DHHS) regions 2 (New York) and 5 (Chicago) reporting less than 50 percent COVID-19 vaccine acceptance. Although our study found a 67% acceptance of a COVID-19 vaccine, there were noticeable demographic and geographical disparities in vaccine acceptance. Before a COVID-19 vaccine is introduced to the U.S., public health officials and policymakers must prioritize effective COVID-19 vaccine-acceptance messaging for all Americans, especially those who are most vulnerable.",
"The success of vaccination efforts to curb the COVID-19 pandemic will require broad public uptake of immunization and highlights the importance of understanding factors associated with willingness to receive a vaccine. U.S. adults aged 65 and older enrolled in the HeartlineTM clinical study were invited to complete a COVID-19 vaccine assessment through the HeartlineTM mobile application between November 6-20, 2020. Factors associated with willingness to receive a COVID-19 vaccine were evaluated using an ordered logistic regression as well as a Random Forest classification algorithm. Among 9,106 study participants, 81.3% (n = 7402) responded and had available demographic data. The majority (91.3%) reported a willingness to be vaccinated. Factors most strongly associated with vaccine willingness were beliefs about the safety and efficacy of COVID-19 vaccines and vaccines in general. Women and Black or African American respondents reported lower willingness to vaccinate. Among those less willing to get vaccinated, 66.2% said that they would talk with their health provider before making a decision. During the study, positive results from the first COVID-19 vaccine outcome study were released; vaccine willingness increased after this report. Even among older adults at high-risk for COVID-19 complications who are participating in a longitudinal clinical study, 1 in 11 reported lack of willingness to receive COVID-19 vaccine in November 2020. Variability in vaccine willingness by gender, race, education, and income suggests the potential for uneven vaccine uptake. Education by health providers directed toward assuaging concerns about vaccine safety and efficacy can help improve vaccine acceptance among those less willing. Clinicaltrials.gov NCT04276441.",
"Amongst the most robust consensus related to the COVID-19 disease is that the elderly are by far the most vulnerable population group. Hence, public authorities target older people in order to convince them to comply with preventive measures. However, we still know little about older people's attitudes and compliance toward these measures. In this research, I aim to improve our understanding of elderly people's responses to the pandemic using data from 27 countries. Results are surprising and quite troubling. Elderly people's response is substantially similar to their fellow citizens in their 50's and 60's. This research (i) provides the first thorough description of the most vulnerable population's attitudes and compliance in a comparative perspective (ii) suggest that governments' strategies toward elderly people are far from successful and (iii) shows that methodologically, we should be more cautious in treating age as having a linear effect on COVID-19 related outcomes."
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Influence of underlying diseases and immunosuppressive regimen on seroconversion rates in liver transplant recipients
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As COVID-19 remains an issue in transplantation medicine, a successful vaccination can prevent infections and life-threatening courses. The probability of poor immune response in liver transplant recipients gained attention and insecurity among those patients, leading us to investigate the humoral immune response alongside the influence of underlying diseases and immunosuppressive regimen on seroconversion rates. We included 118 patients undergoing anti-spike-protein-IgG testing at least 21 days after completed SARS-CoV-2 vaccination. Ninety-seven patients also underwent anti-spike-protein-IgA testing. The influence of baseline demographics, immunosuppressive regimen and underlying disease on seroconversion was analyzed, and 92 of 118 patients (78.0%) developed anti-spike-protein-IgG antibodies. Patients with a history of alcoholic liver disease before transplantation showed significantly lower seroconversion rates (
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"This study quantifies antispike protein antibody responses to first-dose messenger RNA (mRNA) COVID-19 vaccines in solid organ transplant recipients to better understand the immunogenicity of the vaccines in immunocompromised individuals.",
"Knowledge on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in liver transplant recipients is lacking, particularly in terms of severity of the disease. The aim of this study was to describe the demographic, baseline clinical characteristics and early outcomes of a European cohort of liver transplant recipients with SARS-CoV-2 infection. We conducted an international prospective study across Europe on liver transplant recipients with SARS-CoV-2 infection confirmed by microbiological assay during the first outbreak of COVID-19 pandemic. Baseline characteristics, clinical presentation, management of immunosuppressive therapy and outcomes were collected. 57 patients were included (70% male, median (IQR) age at diagnosis 65 (57-70) years). 21 (37%), 32 (56%) and 21 (37%) patients had one cardiovascular disease, arterial hypertension and diabetes mellitus, respectively. The most common symptoms were fever (79%), cough (55%), dyspnoea (46%), fatigue or myalgia (56%) and GI symptoms (33%). Immunosuppression was reduced in 22 recipients (37%) and discontinued in 4 (7%). With this regard, no impact on outcome was observed. Forty-one (72%) subjects were hospitalised and 11 (19%) developed acute respiratory distress syndrome. Overall, we estimated a case fatality rate of 12% (95% CI 5% to 24%), which increased to 17% (95% CI 7% to 32%) among hospitalised patients. Five out of the seven patients who died had a history of cancer. In this European multicentre prospective study of liver transplant recipients, COVID-19 was associated with an overall and in-hospital fatality rate of 12% (95% CI 5% to 24%) and 17% (95% CI 7% to 32%), respectively. A history of cancer was more frequent in patients with poorer outcome.",
"The BNT162b2 vaccine showed high efficacy against COVID-19 in a phase III randomized clinical trial. A vaccine effectiveness evaluation in a real-world setting is needed. To assess the short-term effectiveness of the first dose of the BNT162b2-vaccine against SARS-CoV-2 infection 13 to 24 days after immunization in a real-world setting. This comparative effectiveness study used data from a 2.6 million-member state-mandated health care system in Israel. Participants included all individuals aged 16 years and older who received 1 dose of the BNT162b2 vaccine between December 19, 2020, and January 15, 2021. Data were analyzed in March 2021. Receipt of 1 dose of the BNT162b2 vaccine. Information was collected regarding medical history and positive SARS-CoV-2 polymerase chain reaction test and COVID-19 symptoms from 1 day after first vaccine to January 17, 2021. Daily and cumulative infection rates in days 13 to 24 were compared with days 1 to 12 after the first dose using Kaplan-Meier survival analysis and generalized linear models. Data for 503 875 individuals (mean [SD] age, 59.7 [14.7] years; 263 228 [52.4%] women) were analyzed, of whom 351 897 had follow-up data for days 13 to 24. The cumulative incidence of SARS-CoV-2 infection was 2484 individuals (0.57%) during days 1 through 12 and 614 individuals (0.27%) in days 13 through 24. The weighted mean (SE) daily incidence of SARS-CoV-2 infection in days 1 through 12 was 43.41 (12.07) infections per 100 000 population and 21.08 (6.16) infections per 100 000 population in days 13 through 24, a relative risk reduction (RRR) of 51.4% (95% CI, 16.3%-71.8%). The decrease in incidence was evident from day 18 after the first dose. Similar RRRs were calculated in individuals aged 60 years or older (44.5%; 95% CI, 4.1%-67.9%), those younger than 60 years (50.2%; 95% CI, 14.1%-71.2%), women (50.0%; 95% CI, 13.5%-71.0%), and men (52.1%; 95% CI, 17.3%-72.2%). Findings were similar in subpopulations (eg, ultraorthodox Jewish: RRR, 53.5% [95% CI, 19.2%-73.2%]) and patients with various comorbidities (eg, cardiovascular diseases: RRR, 47.2% [95% CI, 7.8%-69.8%]). Vaccine effectiveness against symptomatic COVID-19 was 54.4% (95% CI, 21.4%-73.6%). In this comparative effectiveness study of a single dose of the BNT162b2 vaccine, results were comparable to that of the phase III randomized clinical trial.",
"The rapid vaccination campaign against COVID-19 in Israel relied on the BNT162b2 vaccine. We performed a longitudinal analysis of multiple cohorts, using individual data, to evaluate the effectiveness of the vaccine against new and breakthrough cases. We estimated vaccine effectiveness (VE) for 27 consecutive cohorts, each comprised of individuals vaccinated on specific days. VE against new COVID-19 cases was evaluated for five SARS-CoV-2-related outcomes: infection, symptomatic disease, hospitalisation, severe/critical disease and death. For breakthrough cases, rate reduction was evaluated for hospitalisation, severe/critical disease and death. Outcomes were evaluated at predetermined time-periods after vaccination, the last one dedicated to individuals who became SARS-CoV-2-positive 22-28 days after the second dose. The highest VE estimates against new cases in ≥16 year old individuals, for all outcomes, were reached at the 15-21 day period after the second dose, ranging between 97.7% (95% CI: 95.9-98.7%) for deaths and 98.6% (95% CI: 97.8-99.1%) for severe/critical disease. VE estimates of the 14-20 day period after the first dose ranged between 54.3% (95% CI: 50.6-57.8%) for infection and 77.3% (95% CI: 71.2-82.1%) for severe/critical disease. VE rose more slowly among ≥80 year old individuals. Rate reductions of breakthrough complications were highest at the 22-28 day period after the second dose, ranging between 47.4% (95% CI: 4.3-71.2%) for death and 66.2% (95% CI: 44.2-79.6%) for severe/critical disease. The BNT162 vaccine is highly effective in preventing new SARS-CoV-2 cases. Among ≥80 year old individuals, high effectiveness develops more slowly. In breakthrough cases, vaccination reduces complications and death. None."
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Impact of the COVID-19 pandemic and income supplements provided by the Australian government on diet cost and affordability for low-income households in Greater Brisbane, Queensland, Australia
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The COVID-19 pandemic has increased food insecurity worldwide, yet there has been limited assessment of shifts in the cost and affordability of healthy, equitable and sustainable diets. This study explores the impact of the COVID-19 pandemic and income supplements provided by the Australian government on diet cost and affordability for low-income households in an Australian urban area. The Healthy Diets ASAP method protocol was applied to assess the cost and cost differential of current and recommended diets before (in 2019) and during the COVID-19 pandemic (late 2020) for households with a minimum-wage and welfare-only disposable household income, by area of socioeconomic disadvantage, in Greater Brisbane, Queensland, Australia. Data were collected between August and October, 2020, from 78 food outlets and compared with data collected in the same locations between May and October, 2019, in an earlier study. The price of most healthy food groups increased significantly during the pandemic-with the exception of vegetables and legumes, which decreased. Conversely, the price of discretionary foods and drinks did not increase during the pandemic. The cost of the current and recommended diets significantly increased throughout this period, but the latter continued to be less expensive than the former. Due to income supplements provided between May and September 2020, the affordability of the recommended diet improved greatly, by 27% and 42%, for households with minimum-wage and welfare-only disposable household income, respectively. This improvement in the affordability of the recommended diet highlights the need to permanently increase welfare support for low-income families to ensure food security.
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"Many historical, environmental, socioeconomic, political, commercial, and geographic factors underscore the food insecurity and poor diet-related health experienced by Aboriginal people in Australia. Yet, there has been little exploration of Aboriginal food practices or perspectives on food choice recently. This study, with 13 households in remote communities on the Anangu Pitjantjatjara Yankunytjatjara (APY) Lands, fills this gap using ethnographic and Indigenist methods. Results highlight Anangu resourcefulness, securing food despite poverty and adversity, and provide unique insights into factors influencing the three major types and range of dietary patterns identified. These factors include household economic cycles and budgeting challenges; overcrowding and family structures, mobility and 'organization'; available food storage, preparation and cooking infrastructure; and familiarity and convenience. Structural and systemic reform, respecting Aboriginal leadership, is required to improve food security.",
"Poor diet is the leading preventable risk factor contributing to the burden of disease globally and in Australia, and is inequitably distributed. As the price of healthy foods is a perceived barrier to improved diets, evidence on the cost and affordability of current (unhealthy) and recommended (healthy, more equitable and sustainable) diets is required to support policy action. This study applied the Healthy Diets ASAP (Australian Standardised Affordability and Pricing) methods protocol to measure the cost, cost differential and affordability of current and recommended diets for a reference household in Queensland, Australia. Food prices were collected in 18 randomly selected locations stratified by area of socioeconomic disadvantage and remoteness. Diet affordability was calculated for three income categories. Surprisingly, recommended diets would cost 20% less than the current diet in Queensland as a whole. Households spent around 60% of their food budget on discretionary choices (that is, those not required for health that are high in saturated fat, added sugar, salt and/or alcohol). Queensland families would need to spend around 23% of their income on recommended diets. However, recommended diets would not be affordable in low socioeconomic or very remote areas, costing 30 and 35% of median household income respectively. The government supplements due to the SARS-CoV-2 pandemic would improve affordability of recommended diets by 29%. Study findings highlight that while price is one factor affecting consumer food choice, other drivers such as taste, convenience, advertising and availability are important. Nevertheless, the study found that recommended diets would be unaffordable in very remote areas, and that low-income families are likely experiencing food stress, irrespective of where they live in Queensland. Policy actions, such as increasing to 20% the current 10% tax differential between basic healthy, and unhealthy foods in Australia, and supplementing incomes of vulnerable households, especially in remote areas, are recommended to help improve diet equity and sustainability, and health and wellbeing for all.",
"Prior to the 2020 outbreak of COVID-19, 70% of Australians' food purchases were from supermarkets. Rural communities experience challenges accessing healthy food, which drives health inequalities. This study explores the impact of COVID-19 on food supply and purchasing behaviour in a rural supermarket. Group model building workshops explored food supply experiences during COVID-19 in a rural Australian community with one supermarket. We asked three supermarket retailers \"What are the current drivers of food supply into this supermarket environment?\" and, separately, 33 customers: \"What are the current drivers of purchases in this supermarket environment?\" Causal loop diagrams were co-created with participants in real time with themes drawn afterwards from coded transcripts. Retailers' experience of COVID-19 included 'empty shelves' attributed to media and government messaging, product unavailability, and community fear. Customers reported fear of contracting COVID-19, unavailability of food, and government restrictions resulting in cooking more meals at home, as influences on purchasing behaviour. Supermarket management and customers demonstrated adaptability and resilience to normalise demand and combat reduced supply.",
"EAT-Lancet Commission's Planetary Health Diet proposed a diet that integrates nutrition and sustainability considerations, however its affordability is unknown in many country-specific contexts, including Australia. The aim of this study is to develop a healthy and sustainable food basket modelled on the Planetary Health Diet to determine the affordability of the Planetary Health Diet basket across various socio-economic groups, and compare this affordability with a food basket modelled on the typical current diet, in an Australian setting. An Australian-specific Planetary Health Diet basket was developed for a reference household (2 adults and 2 children) modelled on the Planetary Health Diet reference diet, and compared to a previously-developed Typical Australian Diet basket. The cost of each food basket was determined by online supermarket shopping surveys in low, medium and high socio-economic areas in each Australian state. Basket affordability was determined for the reference household by comparing the basket cost to disposable income in each socio-economic group in each state. Mann-Whitney U tests then determined if there were significant differences between the median costs and the median affordability of both baskets. The Planetary Health Diet basket was shown to be less expensive and more affordable than the Typical Australian Diet basket nationally, in all metropolitan areas, in all socio-economic groups across Australia (median cost: Planetary Health Diet = AUD$188.21, Typical Australian Diet = AUD$224.36; median affordability: Planetary Health Diet = 13%, Typical Australian Diet = 16%; p = < 0.05). This study showed the Planetary Health Diet to be more affordable than the Typical Australian Diet for metropolitan-dwelling Australians. These results can help to inform public health and food policy aimed at achieving a healthy and sustainable future for all Australians, including reductions in overweight/obesity rates and increased food security.",
"The public health literature suggests that the cheapness of energy-dense foods is driving the obesity epidemic. We examined food purchases in low-income families and its relationship to the price of food and availability of funds. In-depth interviews were conducted with 22 parents with children less than 15 years of age whose major source of income was a government pension. A photo taxonomy, where participants sorted 50 photos of commonly purchased foods, was used to explore food choice. The most common food groupings used by the participants were: basic, emergency, treat and comfort. The process of food purchase was described by participants as weighing up the attributes of a food in relation to price and money available. Shoppers nominated the basic unit of measurement as quantity per unit price and the heuristic for food choice when shopping as determining \"value for money\" in a process of triage relating to food purchase decisions. Participants stated satiation of hunger to be the most common \"value\" relative to price. Given that the foods nominated as filling tended to be carbohydrate-rich staples, we suggest that public health initiatives need to acknowledge this triage process and shape interventions to promote nutrition over satiation.",
"Few Australians consume a healthy, equitable and more sustainable diet consistent with the Australian Dietary Guidelines (ADGs). Low socioeconomic groups (SEGs) suffer particularly poor diet-related health problems. However, granular information on dietary intakes and affordability of recommended diets was lacking for low SEGs. The Healthy Diets Australian Standardised Affordability and Pricing protocol was modified for low SEGs to align with relevant dietary intakes reported in the National Nutrition Survey 2011-2012(which included less healthy and more discretionary options than the broader population), household structures, food purchasing habits, and incomes. Cost and affordability of habitual and recommended diets of low SEGs were calculated using prices of 'standard brands' and 'cheapest options'. With 'standard brands', recommended diets cost less than habitual diets, but were unaffordable for low SEGs. With 'cheapest options', both diets were more affordable, but recommended diets cost more than habitual diets for some low SEGs, potentially contributing to perceptions that healthy food is unaffordable. The study confirms the need for an equity lens to better target dietary guidelines for low SEGs. It also highlights urgent policy action is needed to help improve affordability of recommended diets."
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Carbazole derivatives as potential inhibitors of porcine epidemic diarrhea virus
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Porcine epidemic diarrhea virus (PEDV), an enteric coronavirus, causes neonatal pig acute gastrointestinal infection with a characterization of severe diarrhea, vomiting, high morbidity, and high mortality, resulting in tremendous damages to the swine industry. Neither specific antiviral drugs nor effective vaccines are available, posing a high priority to screen antiviral drugs. The aim of this study is to investigate anti-PEDV effects of carbazole alkaloid derivatives. Eighteen carbazole derivatives (No.1 to No.18) were synthesized, and No.5, No.7, and No.18 were identified to markedly reduce the replication of enhanced green fluorescent protein (EGFP) inserted-PEDV, and the mRNA level of PEDV N. Flow cytometry assay, coupled with CCK8 assay, confirmed No.7 and No.18 carbazole derivatives displayed high inhibition effects with low cell toxicity. Furthermore, time course analysis indicated No.7 and No.18 carbazole derivatives exerted inhibition at the early stage of the viral life cycle. Collectively, the analysis underlines the benefit of carbazole derivatives as potential inhibitors of PEDV, and provides candidates for the development of novel therapeutic agents.
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"Coronavirus-like particles were detected by electron microscopy in the intestinal contents of pigs during a diarrhea outbreak on 4 swine breeding farms. Diarrhea was reproduced in experimental pigs with one of the isolates, designated CV777, which was found to be distinct from the 2 known porcine coronaviruses, transmissible gastroenteritis virus and hemagglutinating encephalomyelitis virus.",
"Porcine epidemic diarrhea virus (PEDV), which re-emerged in China in October 2010, has spread rapidly worldwide. Detailed analyses of the complete genomes of different PEDV strains are essential to understand the relationships among re-emerging and historic strains worldwide. Here, we analysed the complete genomes of 409 strains from different countries, which were classified into five subgroup strains (i.e., GI-a, GI-b, GII-a, GII-b, and GII-c). Phylogenetic study of different genes in the PEDV strains revealed that the newly discovered subgroup GII-c exhibited inconsistent topologies between the spike gene and other genes. Furthermore, recombination analysis indicated that GII-c viruses evolved from a recombinant virus that acquired the 5' part of the spike gene from the GI-a subgroup and the remaining genomic regions from the GII-a subgroup. Molecular clock analysis showed that divergence of the GII-c subgroup spike gene occurred in April 2010, suggesting that the subgroup originated from recombination events before the PEDV re-emergence outbreaks. Interestingly, Ascaris suum, a large roundworm occurring in pigs, was found to be an unusual PEDV host, providing potential support for cross-host transmission. This study has significant implications for understanding ongoing global PEDV outbreaks and will guide future efforts to develop effective preventative measures against PEDV.",
"Griffithsin is a lectin with potent antiviral activity against enveloped viruses. The objective of this study was to assess Griffithsin's inhibitory effect on porcine epidemic diarrhea virus (PEDV). The results showed that Griffithsin reduced PEDV infection of Vero cells by approximately 82.8%. Moreover, using time-of-addition assays and RT-qPCR, we found that delayed addition of Griffithsin had a weaker inhibitory effect on PEDV than earlier treatment. The mechanism of Griffithsin's action against PEDV involved both preventing viral attachment to host cells and disrupting cell-to-cell transmission; its dual mode of action distinguished Griffithsin from most other antiviral drugs. In conclusion, Griffithsin was identified as a potent PEDV inhibitor and may represent a candidate drug for preventing PEDV infection.",
"Pleomorphic fringed particles bearing some resemblance to orthomyxoviruses and coronaviruses were seen in 90 percent of stools from south Indian children and adults but not in stools from neonates. This finding may be related to the abnormalities of intestinal structure and function common in this region of India."
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Effect of calcium and vitamin D supplementation on the risk for kidney stone formation
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Kidney stone disease is a multifactorial condition influenced by both genetic predisposition and environmental factors such as lifestyle and dietary habits. Although different monogenic polymorphisms have been proposed as playing a causal role for calcium nephrolithiasis, the prevalence of these mutations in the general population and their complete pathogenetic pathway is yet to be determined. General dietary advice for kidney stone formers includes elevated fluid intake, dietary restriction of sodium and animal proteins, avoidance of a low calcium diet, maintenance of a normal body mass index, and elevated intake of vegetables and fibers. Thus, balanced calcium consumption protects against the risk for kidney stones by reducing intestinal oxalate availability and its urinary excretion. However, calcium supplementation given between meals might increase urinary calcium excretion without the beneficial effect on oxalate. In kidney stone formers, circulating active vitamin D has been found to be increased, whereas higher plasma 25-hydroxycholecalciferol seems to be present only in hypercalciuric patients. The association between nutritional vitamin D supplements and the risk for stone formation is currently not completely understood. However, taken together, available evidence might suggest that vitamin D administration worsens the risk for stone formation in patients predisposed to hypercalciuria. In this review, we analyzed and discussed available literature on the effect of calcium and vitamin D supplementation on the risk for kidney stone formation.
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"A low-calcium diet is recommended to prevent recurrent stones in patients with idiopathic hypercalciuria, yet long-term data on the efficacy of a low-calcium diet are lacking. Recently, the efficacy of a low-calcium diet has been questioned, and greater emphasis has been placed on reducing the intake of animal protein and salt, but again, long-term data are unavailable. We conducted a five-year randomized trial comparing the effect of two diets in 120 men with recurrent calcium oxalate stones and hypercalciuria. Sixty men were assigned to a diet containing a normal amount of calcium (30 mmol per day) but reduced amounts of animal protein (52 g per day) and salt (50 mmol of sodium chloride per day); the other 60 men were assigned to the traditional low-calcium diet, which contained 10 mmol of calcium per day. At five years, 12 of the 60 men on the normal-calcium, low-animal-protein, low-salt diet and 23 of the 60 men on the low-calcium diet had had relapses. The unadjusted relative risk of a recurrence for the group on the first diet, as compared with the group on the second diet, was 0.49 (95 percent confidence interval, 0.24 to 0.98; P=0.04). During follow-up, urinary calcium levels dropped significantly in both groups by approximately 170 mg per day (4.2 mmol per day). However, urinary oxalate excretion increased in the men on the low-calcium diet (by an average of 5.4 mg per day [60 micromol per day]) but decreased in those on the normal-calcium, low-animal-protein, low-salt diet (by an average of 7.2 mg per day [80 micromol per day]). In men with recurrent calcium oxalate stones and hypercalciuria, restricted intake of animal protein and salt, combined with a normal calcium intake, provides greater protection than the traditional low-calcium diet.",
"In humans, familial or idiopathic hypercalciuria (IH) is a common cause of hypercalciuria and predisposes to calcium oxalate nephrolithiasis. Intestinal calcium hyperabsorption is a constant feature of IH and may be due to either a vitamin D-independent process in the intestine, a primary overproduction of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], or a defect in renal tubular calcium reabsorption. Selective breeding of spontaneously hypercalciuric male and female Sprague-Dawley rats resulted in offspring with hypercalciuria, increased intestinal calcium absorption, and normal serum 1,25(OH)2D3 levels. The role of the vitamin D receptor (VDR) in the regulation of intestinal calcium absorption was explored in 10th generation male genetic IH rats and normocalciuric controls. Urine calcium excretion was greater in IH rats than controls (2.9 +/- 0.3 vs. 0.7 +/- 0.2 mg/24 h, P < 0.001). IH rat intestine contained twice the abundance of VDR compared with normocalciuric controls (536 +/- 73 vs. 243 +/- 42 nmol/mg protein, P < 0.001), with no difference in the affinity of the receptor for its ligand. Comparable migration of IH and normal intestinal VDR on Western blots and of intestinal VDR mRNA by Northern analysis suggests that the VDR in IH rat intestine is not due to large deletion or addition mutations of the wild-type VDR. IH rat intestine contained greater concentrations of vitamin D-dependent calbindin 9-kD protein. The present studies strongly suggest that increased intestinal VDR number and normal levels of circulating 1,25(OH)2D3 result in increased functional VDR-1,25(OH)2D3 complexes, which exert biological actions in enterocytes to increase intestinal calcium transport. Intestinal calcium hyperabsorption in the IH rat may be the first example of a genetic disorder resulting from a pathologic increase in VDR.",
"This study aims to prospectively assess the incidence of hypercalciuria and hypercalcemia with different doses of vitamin D and with a calcium intake of approximately 1,200 mg/day. This was a 1-year randomized placebo-controlled study of vitamin D (400-4,800 IU/d) in 163 white women aged 57 to 90 years. Calcium citrate tablets (200 mg) were added to the diet to achieve a total calcium intake of approximately 1,200 mg/day in all groups. All women had vitamin D insufficiency at baseline, with serum 25-hydroxyvitaminD levels lower than 20 ng/mL (50 nmol/L). Serum and 24-hour urine calcium were collected every 3 months on supplementation, any test result above the upper reference range represented an episode of hypercalcemia or hypercalciuria. Mixed-effects models and multivariate logistic regression were used in the analysis. Hypercalcemia (>10.2 mg/dL [2.55 mmol/L]) occurred in 8.8% of white women. Hypercalciuria (>300 mg/d [7.5 mmol]) occurred in 30.6% of white women. Episodes of hypercalciuria were transient in half of the group and recurrent in the other half. No relationship between hypercalcemia or hypercalciuria and vitamin D dose was found, and hypercalciuria was equally common in the placebo group. Hypercalciuria and hypercalcemia commonly occur with vitamin D and calcium supplements. Whether hypercalciuria and hypercalcemia are caused by calcium, vitamin D, or both is unclear. These findings may have relevance to the reported increase in kidney stones in the Women's Health Initiative trial. Because calcium 1,200 mg and vitamin D 800 IU/day are widely recommended in postmenopausal women, systematic evaluation of the safety of supplements is warranted in clinical management and in future studies.",
"There are striking inequities in calcium intake between rich and poor populations. Appropriate calcium intake has shown many health benefits, such as reduction of hypertensive disorders of pregnancy, lower blood pressure particularly among young people, prevention of osteoporosis and colorectal adenomas, lower cholesterol values, and lower blood pressure in the progeny of mothers taking sufficient calcium during pregnancy. Studies have refuted some calcium supplementation side effects like damage to the iron status, formation of renal stones and myocardial infarction in older people. Attention should be given to bone resorption in post-partum women after calcium supplementation withdrawal. Mechanisms linking low calcium intake and blood pressure are mediated by parathyroid hormone raise that increases intracellular calcium in vascular smooth muscle cells leading to vasoconstriction. At the population level, an increase of around 400-500 mg/day could reduce the differences in calcium intake between high- and middle-low-income countries. The fortification of food and water seems a possible strategy to reach this goal.",
"Nephrolithiasis is a common health problem in autosomal dominant polycystic kidney disease (ADPKD) and significantly contributes to patient morbidity. Recently, Tolvaptan has been introduced for the treatment of ADPKD, but whether it is associated with alterations of the urinary lithogenic risk profile remains unknown. We conducted an analysis of participants enrolled in the Bern ADPKD registry, a prospective observational cohort study. Twenty-four-hour urine analyses were performed at baseline and then at yearly follow-ups. Relative supersaturation ratios for calcium oxalate, brushite, and uric acid were calculated with the program EQUIL2. Unadjusted and multivariable mixed-effects linear regression models, adjusted for age, sex, body mass index, eGFR, net acid excretion, and height-adjusted total kidney volume, were used to assess the association of Tolvaptan with urinary parameters relevant for kidney stone formation. The maximum individual follow-up time was 3 years, median follow-up time 1.9 years, and cumulative follow-up time 169 years. In total, 125 participants (38 with and 87 without Tolvaptan treatment) were included in the analysis. In multivariable analysis, Tolvaptan treatment was associated [adjusted estimate of the difference between Tolvaptan and no Tolvaptan; 95% confidence interval (CI)] with lower urine relative supersaturation ratios for calcium oxalate (-0.56; 95% CI, -0.82 to -0.3; P<0.001), brushite (-0.33; 95% CI, -0.54 to -0.11; P=0.004), and uric acid (-0.62; 95% CI, -0.88 to -0.37; P<0.001), and with higher urine citrate in mmol/mmol creatinine per day (0.25; 95% CI, 0.05 to 0.46; P=0.02) and calcium in mmol/mmol creatinine per day (0.31; 95% CI, 0.09 to 0.53; P=0.006) excretion. In addition, Tolvaptan treatment was associated with lower net acid excretion in mEq/mmol creatinine per day (-0.54; 95% CI, -0.90 to -0.17; P=0.004) and higher net gastrointestinal alkali absorption in mEq/mmol creatinine per day (0.57; 95% CI, 0.26 to 0.88; P<0.001). Tolvaptan treatment is associated with a significantly improved urinary lithogenic risk profile in patients with ADPKD.",
"Mutations of the CYP24A1 gene, which encodes the 1,25-dihydroxyvitamin D-24-hydroxylase cytochrome P450, Cyp24A1, are predicted to result in elevated 1,25-dihydroxyvitamin D concentrations, hypercalcemia, hypercalciuria, nephrolithiasis, and bone disease. Treatment of hypercalcemia associated with CYP24A1 gene mutations has not been described. The genetic basis of a syndrome in a 44-yr-old Caucasian male characterized by intermittent hypercalcemia, hypercalciuria, elevated serum 1,25-dihydroxyvitamin D, undetectable serum 24,25-dihydroxyvitamin D, metabolically active nephrolithiasis, and reduced bone mineral density of the lumbar spine was examined. Sequencing of the CYP24A1 gene and biochemical and genetic analysis of seven family members in three generations was carried out. Because of hypercalcemia, hypercalciuria, and metabolically active nephrolithiasis, the patient was treated with a cytochrome 3A inhibitor, ketoconazole, 200 mg orally every 8 h, for 2 months. The sequence of the CYP24A1 gene showed two canonical splice junction mutations in the proband. Analysis of family members showed a phenotype associated one or both mutations, suggesting autosomal dominant transmission with partial penetrance of the trait. After therapy with ketoconazole, statistically significant reductions in previously elevated urinary calcium into the normal range were noted. Previously elevated serum 1,25-dihydroxyvitamin D and calcium concentrations decreased, and previously decreased PTH concentrations increased into the normal range, but the differences were not statistically significant. In a syndrome characterized by intermittent hypercalcemia, hypercalciuria, elevated 1,25-dihydroxyvitamin D, undetectable 24,25-dihydroxyvitamin D concentrations, splice junction mutations of the CYP24A1 gene, and autosomal dominant transmission of the trait, treatment with ketoconazole is useful in reducing urinary calcium.",
"Idiopathic infantile hypercalcaemia (IIH) is an autosomal recessively inherited disease, presented in the first year of life with hypercalcaemia, precipitated by normal amounts of vitamin D supplementation. Recently loss-of-function mutations in the CYP24A1 gene, which encodes the vitamin D-metabolizing enzyme 24-hydroxylase, have been found in these patients. We describe a young man homozygous for a novel missense mutation (c.628T>C) of the CYP24A1 gene. He had suffered from severe hypercalcaemia in early childhood. At age 29 he presented with medullary nephrocalcinosis, chronic kidney disease (CKD) stage 2, microalbuminuria, mild hypertension and nephrogenic diabetes insipidus. He had mild hypercalcaemia and moderate hypercalciuria. As a novel finding, fibroblast growth factor 23 (FGF23) was elevated."
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Hepatitis C virus screening in primary care and a private gastroenterology practice.
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(1) Background: Low rates of hepatitis C virus (HCV) diagnosis and sub-optimal linkage to care constitute barriers toward eliminating the infection. In 2012/2013, we showed that HCV screening in primary care detects unknown cases. However, hepatitis C patients may not receive further diagnostics and therapy because they drop out during the referral pathway to secondary care. Thus, we used an existing network of primary care physicians and a practice of gastroenterology to investigate the pathway from screening to therapy. (2) Methods: HCV screening was prospectively included in a routine check-up of primary care physicians who cooperated regularly with a private gastroenterology practice. Anti-HCV-positive patients were referred for further specialized diagnostics and treatment if indicated. (3) Results: Seventeen primary care practices screened 1875 patients. Twelve individuals were anti-HCV-positive (0.6%), six of them reported previous antiviral HCV therapy, and one untreated patient was HCV-RNA-positive (0.05% of the population). None of the 12 anti-HCV-positive cases showed up at the private gastroenterology practice. Further clinical details of the pathway from screening to therapy could not be analyzed. (4) Conclusions: The linkage between primary and secondary care appears to be problematic in the HCV setting even among cooperating partners, but robust conclusions require larger datasets.
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"Management of hepatitis C virus (HCV)-infected individuals requires referral to specialist care. To determine whether patients newly diagnosed as anti-HCV positive are appropriately referred for further investigation and management, and if not, to determine why not. We studied patients tested for antibodies to HCV by Nottingham Public Health Laboratory in a 2-year period (2000-2002). The progress of newly diagnosed anti-HCV positive patients into specialist clinics for further management was documented. For patients not referred for specialist care, a questionnaire was sent to the clinician requesting the initial anti-HCV test, to identify reasons for nonreferral. Eleven thousand one hundred and seventy-seven patients were tested for anti-HCV. Two hundred and fifty-six (2.3%) were newly diagnosed as being anti-HCV positive. Two per cent of samples sent from primary care were anti-HCV positive, compared to 18.8, 18.9 and 1.3% sent from prison, drug and alcohol units, and secondary care, respectively. About 64.3% of positive patients diagnosed in primary care were referred to specialist care, compared to 18.4, 42.4 and 62.6% of patients diagnosed in the other three settings. One hundred and twenty-five (49%) newly diagnosed patients were referred appropriately for further management. 68 of these attended clinic, 45 underwent liver biopsy and 26 (10%) began treatment. One hundred and thirty-one patients (51%) were not referred. In 54 cases, there was no evidence that the anti-HCV positive result reached the patient. In 15, referral was considered but rejected, and 20 patients were referred to non-HCV-specialists (their general practitioners or to genito-urinary medicine). Hence less than 50% of newly diagnosed anti-HCV positive patients are referred to an appropriate clinic for further investigation and management. Reasons for this are multifarious and complex, reflecting both systems failure and patient choice. Unless these are understood and addressed, the Department of Health Hepatitis C Strategy (2002) and Action Plan for England (2004) will fail to achieve their intended objectives.",
"Hepatitis C virus (HCV) infection remains a pressing public health issue. Our aim is to assess the linkage to care of patients with HCV diagnosis and to support the proactive case-finding of new HCV-infected patients in an Italian primary care setting. This was a retrospective cohort study of 44 general practitioners (GPs) who managed 63,955 inhabitants in the Campania region. Adults with already known HCV diagnosis or those with HCV high-risk profile at June 2019 were identified and reviewed by GPs to identify newly diagnosed of HCV and to assess the linkage to care and treatment for the HCV patients. Overall, 698 HCV patients were identified, 596 with already known HCV diagnosis and 102 identified by testing the high-risk group (2614 subjects). The 38.8% were already treated with direct-acting antivirals, 18.9% were referred to the specialist center and 42.3% were not sent to specialist care for treatment. Similar proportions were found for patients with an already known HCV diagnosis and those newly diagnosed. Given that the HCV infection is often silent, case-finding needs to be proactive and based on risk information. Our findings suggested that there needs to be greater outreach, awareness and education among GPs in order to enhance HCV testing, linkage to care and treatment.",
"To define the process of outpatient consultation, the authors conducted a prospective study of 716 consecutive outpatient consultations in a university-based primary care internal medicine practice. The overall consultation rate was 11.9 per 100 patient visits, with 78% of the referrals to other physicians and 22% to non-physician specialists. Consultation rates and patterns of referral varied little between physicians with different levels of experience. Eighteen per cent of the consultations resulted in a no-show by the patient to the consultant. Referring physicians received communications from the consultants 80.5% of the time when appointments were kept. By multivariate regression two variables were shown to be most important in determining the internist's overall satisfaction: 1) how well the consultant aided the internist in his ongoing management of the patient's problem, and 2) how well specific questions were addressed by the specialist. Other statistically significant variables were the clarity and promptness of the consultant's reply, the educational value of the consultation, and specific management recommendations made by the consultant. To improve the consultation process no-shows must be minimized, communication from the consultant maximized, and the interaction between the internist and the consultant bolstered.",
"The Chronic Hepatitis Cohort Study (CHeCS), a dynamic prospective, longitudinal, observational cohort study, was created to assess the clinical impact of chronic viral hepatitis in the United States. This report describes the cohort selection process, baseline demographics, and insurance, biopsy, hospitalization, and mortality rates. Electronic health records of >1.6 million adult patients seen from January 2006 through December 2010 at 4 integrated healthcare systems in Detroit, Michigan; Danville, Pennsylvania; Portland, Oregon; and Honolulu, Hawaii were collected and analyzed. Of 2202 patients with chronic hepatitis B virus (HBV) infection, 50% were aged 44-63 years, 57% male, 58% Asian/Pacific Islander, and 13% black; and 5.1% had Medicaid, 16.5% Medicare, and 76.3% private insurance. During 2001-2010, 22.3% had a liver biopsy and 37.9% were hospitalized. For the 8810 patients with chronic hepatitis C virus (HCV) infection, 75% were aged 44-63 years, 60% male, 23% black; and 12% had Medicaid, 23% Medicare, and 62% private insurance. During 2001-2010, 38.4% had a liver biopsy and 44.3% were hospitalized. Among persons in care, 9% of persons with HBV and 14% of persons with HCV infection, mainly those born during 1945-1964, died during the 2006-2010 five-year period. Baseline demographic, hospitalization, and mortality data from CHeCS highlight the substantial US health burden from chronic viral hepatitis, particularly among persons born during 1945-1964.",
"Globally, hepatitis C virus (HCV) infects ∼3% of the population. The objective of this study was to review published work and determine the direct medical costs for diseases associated with HCV infection globally, with the exception of the US. A systematic literature search was conducted to identify studies reporting the costs of hepatitis C sequelae between January 1990 and January 2011. Over 400 references were identified, of which 45 were pertinent. The costs were compiled, converted to US dollars, and adjusted to 2010 costs using the medical component of the consumer price index. The median cost of liver transplants was estimated at $139,070 ($15,430-$443,700), refractory ascites at $16,740 ($8990-$35,940), hepatocellular carcinoma (HCC) at $15,310 ($3370-$84,710), decompensated cirrhosis at $14,660 ($3810-$48,360), variceal hemorrhage at $12,190 ($3550-$46,120), hepatic encephalopathy at $9180 ($5370-$50,120), diuretic sensitive ascites at $3400 ($1320-$7470), compensated cirrhosis at $820 ($50-$2890), and chronic hepatitis C at $280 ($90-$1860). The variation among studies was mainly due to the methodology used to assess cost, local cost and government reimbursement, and country-specific treatment protocols. All costs were adjusted to 2010 US dollars using the US medical component of the consumer price index (CPI) which may not reflect the change in medical costs in other countries. In addition, the costs, in the local currency were converted to US dollars in the year of the study. However, medical expenses may not vary with exchange rate, leading to artificial variations. Finally, there was no assessment of the quality of individual studies, which resulted in the same weighting to all studies. Hepatitis C imposes a high economic burden globally. Knowing the burden of HCV sequelae is useful for policy decisions as well as serving as a basis for determining the value of HCV screening and treatment."
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Quality assessment of oocytes from in vitro-developed antral follicles.
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In vitro follicle development (IVFD) is an adequate model to obtain basic knowledge of folliculogenesis and provides a tool for ovarian toxicity screening. IVFD yielding competent oocytes may also offer an option for fertility and species preservation. To promote follicle growth and oocyte maturation in vitro, various culture systems are utilized for IVFD in rodents, domestic animals, wild animals, nonhuman primates, and humans. Follicle culture conditions have been improved by optimizing gonadotropin levels, regulatory factors, nutrient supplements, oxygen concentration, and culture matrices. This review summarizes quality assessment of oocytes generated from in vitro-developed antral follicles from the preantral stage, including oocyte epigenetic and genetic profile, cytoplasmic and nuclear maturation, preimplantation embryonic development following in vitro fertilization, as well as pregnancy and live offspring after embryo transfer. The limitations of oocyte quality evaluation following IVFD and the gaps in our knowledge of IVFD to support proper oocyte development are also discussed. The information may advance our understanding of the requirements for IVFD, with a goal of producing competent oocytes with genetic integrity to sustain embryonic development resulting in healthy offspring.
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"The removal and cryostorage of ovarian cortical biopsies is now offered as a fertility preservation option for young women. The only available option to restore fertility using this tissue is by transplantation, which may not be possible for all patients. The full potential of this tissue to restore fertility could be achieved by the development of in vitro systems that support oocyte development from the most immature stages to maturation. The techniques of in vitro growth (IVG) combined with in vitro maturation (IVM) are being developed with human tissue, but comparing different systems has been difficult because of the scarcity of tissue so nonhuman primates are being used as model systems. There are many challenges to developing a complete culture system that would support human oocyte development, and this review outlines the approaches being taken by several groups using tissue from women and nonhuman primate models to support each of the stages of oocyte development.",
"This study aimed to evaluate the role of anethole during the in vitro culture of caprine early antral follicles. Early antral follicles were isolated from caprine ovaries and cultured for 18 days without (control) or with anethole (300 µg/ml). After culture, the cumulus-oocyte complexes were subjected to in vitro maturation, followed by parthenogenetic activation or in vitro fertilization (IVF) and embryo culture. Follicular walls were used for the quantification of messenger RNA (mRNA) of CYP19A1, CYP17, MMP-9, TIMP-2, Bax, and Bcl-2 genes, and culture medium was used for evaluation of ferric reducing antioxidant power (FRAP) and estradiol levels. After in vitro follicle culture (IVFC), anethole induced higher total antioxidant capacity, that is, it produced higher FRAP levels, reduced the Bax/Bcl-2 ratio, and increased the levels of mRNA for CYP19A1 and CYP17, which was associated with a greater estradiol production (p < .05). Also, anethole improved the ability of oocytes to resume meiosis and reach metaphase II stage, as well as yielded higher (p < .05) embryo production (e.g., morulas and blastocysts) in both parthenogenetic activation and IVF techniques. One pregnancy (Day 30) was obtained from IVFC with anethole. In conclusion, anethole promoted in vitro growth and maturation of goat early antral follicles and oocytes and enabled embryo production. Furthermore, this study reports, for the first time in goats, a pregnancy after IVF using oocytes originated from early antral follicles grown in vitro.",
"The availability of viable oocytes is the limiting factor in the development of new reproductive techniques. Many attempts have been made to grow immature oocytes in vitro during recent decades. Recently, a modified alginate-based three-dimensional culture system was designed to support the growth and maturation of multilayered secondary follicles. This system was able to produce oocytes that successfully completed meiosis, fertilization, and development to the blastocyst stage. Subsequent attempts to culture two-layered secondary follicles were unsuccessful under the original conditions. Herein, we investigated the effect of alginate consistency on two-layered follicle growth and oocyte developmental competence by encapsulating follicles into alginate scaffolds of various concentrations. Although there were no significant differences in survival rates, 0.25% and 0.5% alginate supported more rapid growth of follicles and antrum formation compared with 1.5% and 1.0% alginate after 8 days of culture. Alginate scaffold concentration also affected the proliferation and differentiation of somatic cells (theca and granulosa cells), measured in terms of morphological changes, steroid profiles (androstenedione, estradiol, and progesterone), and specific molecular markers (Fshr, Lhcgr, and Gja1). Theca cell proliferation and steroid production were hindered in follicles cultured in 1.5% alginate. In vitro fertilization and embryo culture revealed that oocytes obtained from 0.25% alginate retained the highest developmental competence. Overall, the present study showed that the alginate scaffold consistency affects folliculogenesis and oocyte development in vitro and that the alginate culture system can and should be tailored to maximally support follicle growth depending on the size and stage of the follicles selected for culture.",
"The present study examines the use of buffalo preantral follicles as a source of oocytes for in vitro embryo production. Preantral follicles were isolated from abattoir-derived buffalo ovaries and were grown for 100 days in five different culture systems: (1) minimum essential medium (MEM); (2) coconut water; (3) MEM + ovarian mesenchymal cell (OMC) co-culture; (4) MEM + granulosa cell (GC) co-culture; or (5) MEM + cumulus cell (CC) co-culture. Low growth rates for the preantral follicles were observed when follicles were cultured in MEM or coconut water medium. Moderate growth rates were seen for OMC and GC co-cultures, and high rates of growth were observed when follicles were grown in CC co-culture. The survival of preantral follicles was low in the MEM culture (<25%), but was over 75% in the other culture systems. Oocytes were not recovered from the MEM group, while an oocyte recovery rate of 80-100% was observed when the follicles were cultured with coconut water/somatic cells. Transferable embryos could be produced only with the oocytes obtained from preantral follicles grown in the OMC and CC co-culture systems. This study demonstrates, for the first time, that it is possible to produce buffalo embryos by in vitro fertilization of oocytes derived from in vitro grown preantral follicles.",
"In recent years several follicle culture systems have been pioneered in different mammalian species for studying ovarian folliculogenesis and culturing immature oocytes. Applications of these in vitro techniques include fertility preservation for humans, conservation of rare animals and development of oocyte banks for research purposes. Immature female gametes in the ovarian cortex can be cryopreserved for later use if culture techniques are available afterwards to promote growth and maturation. This review focuses on biochemical and biophysical factors related to oocyte culture in mice, the only animal in which live offspring have been produced after folliculogenesis in vitro. The advantage of using mice for these studies is that, in parallel to development of follicle culture systems, essential knowledge on folliculogenesis can be obtained from knockout mouse models. Recent experiments in mice stressed the principal role of the oocyte in follicle development and the strict timing of the biological processes underlying oogenesis in vitro. In large domestic animals and humans, study of oocyte culture is confounded by the constitutively prolonged nature of ovarian follicle development. In humans, only some aspects of follicle development have been studied because of the limited availability of suitable material for experimentation, technical difficulties related to manipulation of very small structures and lack of knowledge on physiological regulation of the early stages of follicle growth. Only a few reports describe ovarian follicular growth in vitro. In this review, relevant information on hormonal and growth factor regulation of the earliest stages of follicle growth in mammals is reviewed. Techniques are becoming available for the precise isolation of distinct classes of follicle and powerful molecular biology techniques can be used in studies of ovarian tissue culture.",
"The two principal functions of ovarian follicles are developmental and endocrine. The cumulus cells surrounding the oocyte are specialized to serve the development of the oocyte and steroidogenesis is a principal role of mural granulosa cells that line the follicle wall. The findings in this report demonstrate that oocytectomy or treatment with an inhibitor of SMAD2/3 activation results in decreased cumulus marker mRNA transcript levels and allows FSH to induce mural marker transcripts in cumulus cells. In addition, SMAD2/3 signaling is involved in enabling cumulus expansion and EGF-induced increases in Ptx3, Ptgs2 and Has2 mRNA levels. By contrast, follicle-stimulating hormone (FSH) stimulated expression of mural transcripts, but suppressed levels of cumulus transcripts. Thus, FSH and oocyte-stimulated SMAD2/3 signaling establish opposing gradients of influence in the follicle. These specify the mural and cumulus granulosa cell phenotypes that are pivotal for appropriate endocrine function and oocyte development.",
"We describe a novel 30-kDa secretory protein, Acrp30 (adipocyte complement-related protein of 30 kDa), that is made exclusively in adipocytes and whose mRNA is induced over 100-fold during adipocyte differentiation. Acrp30 is structurally similar to complement factor C1q and to a hibernation-specific protein isolated from the plasma of Siberian chipmunks; it forms large homo-oligomers that undergo a series of post-translational modifications. Like adipsin, secretion of Acrp30 is enhanced by insulin, and Acrp30 is an abundant serum protein. Acrp30 may be a factor that participates in the delicately balanced system of energy homeostasis involving food intake and carbohydrate and lipid catabolism. Our experiments also further corroborate the existence of an insulin-regulated secretory pathway in adipocytes."
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Epigenetics and Huntington's disease
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In light of our aging population, neurodegenerative disorders are becoming a tremendous challenge, that modern societies have to face. They represent incurable, progressive conditions with diverse and complex pathological features, followed by catastrophic occurrences of massive neuronal loss at the later stages of the diseases. Some of these disorders, like Huntington's disease (HD), rely on defined genetic factors. HD, as an incurable, fatal hereditary neurodegenerative disorder characterized by its mid-life onset, is caused by the expansion of CAG trinucleotide repeats coding for glutamine (Q) in exon 1 of the huntingtin gene. Apart from the genetic defect, environmental factors are thought to influence the risk, onset and progression of HD. As epigenetic mechanisms are known to readily respond to environmental stimuli, they are proposed to play a key role in HD pathogenesis. Indeed, dynamic epigenomic remodeling is observed in HD patients and in brains of HD animal models. Epigenetic signatures, such as DNA methylation, histone variants and modifications, are known to influence gene expression and to orchestrate various aspects of neuronal physiology. Hence, deciphering their implication in HD pathogenesis might open up new paths for novel therapeutic concepts, which are discussed in this review.
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[
"Alzheimer's disease and other neurodegenerative disorders of aging are characterized by clinical and pathological features that are relatively specific to humans. To obtain greater insight into how brain aging has evolved, we compared age-related gene expression changes in the cortex of humans, rhesus macaques, and mice on a genome-wide scale. A small subset of gene expression changes are conserved in all three species, including robust age-dependent upregulation of the neuroprotective gene apolipoprotein D (APOD) and downregulation of the synaptic cAMP signaling gene calcium/calmodulin-dependent protein kinase IV (CAMK4). However, analysis of gene ontology and cell type localization shows that humans and rhesus macaques have diverged from mice due to a dramatic increase in age-dependent repression of neuronal genes. Many of these age-regulated neuronal genes are associated with synaptic function. Notably, genes associated with GABA-ergic inhibitory function are robustly age-downregulated in humans but not in mice at the level of both mRNA and protein. Gene downregulation was not associated with overall neuronal or synaptic loss. Thus, repression of neuronal gene expression is a prominent and recently evolved feature of brain aging in humans and rhesus macaques that may alter neural networks and contribute to age-related cognitive changes.",
"Cytosine guanine dinucleotide (CpG) island methylation is a known mechanism of epigenetic inheritance in postmeiotic cells. Through associated chromatin changes and silencing, such epigenetic states can influence cellular physiology and affect disease risk and severity. Our studies of CpG island methylation in normal colorectal mucosa revealed progressive age-related increases at multiple gene loci, suggesting genome-wide molecular alterations with potential to silence gene expression. However, there was considerable variation in the degree of methylation among individuals of comparable ages. Such variation could be related to genetic factors, lifestyle, or environmental exposures. Studies in ulcerative colitis and hepatocellular cirrhosis and neoplasia revealed that chronic inflammatory states are accompanied by marked increases in CpG island methylation in normal-appearing tissues, confirming the hypothesis that proinflammatory exposures could account for part of the epigenetic variation in human populations. Preliminary data also suggest potential influences of lifestyle and exposure factors on CpG island methylation. It is suggested that epigenetic variation related to aging, lifestyle, exposures and possibly genetic factors, is one of the modulators of acquired, age-related human diseases, including neoplasia.",
"Progression of brain ageing is influenced by a complex interaction of genetic and environmental factors. Analysis of genetically modified animals with uniform genetic backgrounds in a standardised, controlled environment enables the dissection of critical determinants of brain ageing on a molecular level. Human and animal studies suggest that increased load of damaged macromolecules, efficacy of DNA maintenance, mitochondrial activity, and cellular stress defences are critical determinants of brain ageing. Surprisingly, mouse lines with genetic impairment of anti-oxidative capacity generally did not show enhanced cognitive ageing but rather an increased sensitivity to oxidative challenge. Mouse lines with impaired mitochondrial activity had critically short life spans or severe and rapidly progressing neurodegeneration. Strains with impaired clearance in damaged macromolecules or defects in the regulation of cellular stress defences showed alterations in the onset and progression of cognitive decline. Importantly, reduced insulin/insulin-like growth factor signalling generally increased life span but impaired cognitive functions revealing a complex interaction between ageing of the brain and of the body. Brain ageing is accompanied by an increased risk of developing Alzheimer's disease. Transgenic mouse models expressing high levels of mutant human amyloid precursor protein showed a number of symptoms and pathophysiological processes typical for early phase of Alzheimer's disease. Generally, therapeutic strategies effective against Alzheimer's disease in humans were also active in the Tg2576, APP23, APP/PS1 and 5xFAD lines, but a large number of false positive findings were also reported. The 3xtg AD model likely has the highest face and construct validity but further studies are needed.",
"Mammalian genomes encode tens of thousands of long-noncoding RNAs (lncRNAs), which are capable of interactions with DNA, RNA and protein molecules, thereby enabling a variety of transcriptional and post-transcriptional regulatory activities. Strikingly, about 40% of lncRNAs are expressed specifically in the brain with precisely regulated temporal and spatial expression patterns. In stark contrast to the highly conserved repertoire of protein-coding genes, thousands of lncRNAs have newly appeared during primate nervous system evolution with hundreds of human-specific lncRNAs. Their evolvable nature and the myriad of potential functions make lncRNAs ideal candidates for drivers of human brain evolution. The human brain displays the largest relative volume of any animal species and the most remarkable cognitive abilities. In addition to brain size, structural reorganization and adaptive changes represent crucial hallmarks of human brain evolution. lncRNAs are increasingly reported to be involved in neurodevelopmental processes suggested to underlie human brain evolution, including proliferation, neurite outgrowth and synaptogenesis, as well as in neuroplasticity. Hence, evolutionary human brain adaptations are proposed to be essentially driven by lncRNAs, which will be discussed in this review.",
"Unbalanced epigenetic regulation is thought to contribute to the progression of several neurodegenerative diseases, including Huntington's disease (HD), a genetic disorder considered as a paradigm of epigenetic dysregulation. In this review, we attempt to address open questions regarding the role of epigenetic changes in HD, in the light of recent advances in neuroepigenetics. We particularly discuss studies using genome-wide scale approaches that provide insights into the relationship between epigenetic regulations, gene expression and neuronal activity in normal and diseased neurons, including HD neurons. We propose that cell-type specific techniques and 3D-based methods will advance knowledge of epigenome in the context of brain region vulnerability in neurodegenerative diseases. A better understanding of the mechanisms underlying epigenetic changes and of their consequences in neurodegenerative diseases is required to design therapeutic strategies more effective than current strategies based on histone deacetylase (HDAC) inhibitors. Researches in HD may play a driving role in this process.",
"Molecules used for communication in mature nervous systems also play important roles in development, maintenance and plasticity of individual neurons. This paper reviews the evidence that neurotransmitters, in addition to their mediation of trans-synaptic information coding, can induce a spectrum of effects on neuronal cytoarchitecture, ranging from neurite sprouting to dendritic pruning and even cell death. Such profound alterations may well constitute a part of the normal functioning and structuring of the nervous system as well as contribute to severe pathological processes.",
"Huntington's Disease (HD) is caused by an expansion of a polyglutamine tract within the huntingtin (htt) protein. Pathogenesis in HD appears to include the cytoplasmic cleavage of htt and release of an amino-terminal fragment capable of nuclear localization. We have investigated potential consequences to nuclear function of a pathogenic amino-terminal region of htt (httex1p) including aggregation, protein-protein interactions, and transcription. httex1p was found to coaggregate with p53 in inclusions generated in cell culture and to interact with p53 in vitro and in cell culture. Expanded httex1p represses transcription of the p53-regulated promoters, p21(WAF1/CIP1) and MDR-1. httex1p was also found to interact in vitro with CREB-binding protein (CBP) and mSin3a, and CBP to localize to neuronal intranuclear inclusions in a transgenic mouse model of HD. These results raise the possibility that expanded repeat htt causes aberrant transcriptional regulation through its interaction with cellular transcription factors which may result in neuronal dysfunction and cell death in HD.",
"Huntington disease (HD) is caused by a CAG repeat expansion that is translated into an abnormally long polyglutamine (polyQ) tract in the huntingtin protein. The precise mechanisms leading to neurodegeneration in HD have not been fully elucidated, but alterations in gene transcription could well be involved because the activities of several nuclear proteins are compromised by the polyQ mutation. Recent microarray studies also show relevant changes in gene expression profiles in HD models, providing useful information on the potential consequences of disrupted transcriptional pathways in HD.",
"A better understanding of the molecular effects of aging in the brain may help to reveal important aspects of organismal aging, as well as processes that lead to age-related brain dysfunction. In this study, we have examined differences in gene expression in the hypothalamus and cortex of young and aged mice by using high-density oligonucleotide arrays. A number of key genes involved in neuronal structure and signaling are differentially expressed in both the aged hypothalamus and cortex, including synaptotagmin I, cAMP-dependent protein kinase C beta, apolipoprotein E, protein phosphatase 2A, and prostaglandin D. Misregulation of these proteins may contribute to age-related memory deficits and neurodegenerative diseases. In addition, many proteases that play essential roles in regulating neuropeptide metabolism, amyloid precursor protein processing, and neuronal apoptosis are up-regulated in the aged brain and likely contribute significantly to brain aging. Finally, a subset of these genes whose expression is affected by aging are oppositely affected by exposure of mice to an enriched environment, suggesting that these genes may play important roles in learning and memory.",
"The protein composition of the neuronal plasma membrane is regulated by clathrin-mediated endocytosis and changes drastically over the neuronal lifespan. Here, we utilize the transition out of the period of early postnatal growth as a model system to study age-related changes in endocytosis. Previously, we have found that the dynamic behavior of endocytic clathrin coats in dendrites changes during this period, and that clathrin coat lifetime increases in older neurons. In this study, we examine endocytosis in neuronal dendrites by measuring transferrin (Tf) uptake, and find that it is markedly reduced in older neurons in culture. This decrease was not due to a reduction in transferrin receptor protein levels, nor to a decrease in the expression of endocytic proteins. However, imaging of endocytosis in living dendrites demonstrated that cargo transport through clathrin-coated pits was slower during internalization. Thus, endocytic function in dendrites is altered in older neurons, suggesting that as neurons age, protein trafficking mechanisms are controlled to complement maturational requirements."
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Prevalence of depression and anxiety in mothers of young children during the COVID-19 pandemic
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Parents have experienced considerable challenges and stress during the COVID-19 pandemic, which may impact their well-being. This meta-analysis sought to identify: (1) the prevalence of depression and anxiety in parents of young children (<age 5) during the COVID-19 pandemic, and (2) sociodemographic (e.g., parent age, being racially minoritized) and methodological moderators (e.g., study quality) that explain heterogeneity among studies. A systematic search was conducted across four databases from January 1, 2020 to March 3, 2021. A total of 18 non-overlapping studies (8981 participants), all focused on maternal mental health, met inclusion criteria. Random-effect meta-analyses were conducted. Pooled prevalence estimates for clinically significant depression and anxiety symptoms for mothers of young children during the COVID-19 pandemic were 26.9% (95% CI: 21.3-33.4) and 41.9% (95% CI: 26.7-58.8), respectively. Prevalence of clinically elevated depression and anxiety symptoms were higher in Europe and North America and among older mothers. Clinically elevated depressive symptoms were lower in studies with a higher percentage of individuals who were racially minoritized. In comparison, clinically elevated anxiety symptoms were higher among studies of low study quality and in samples with highly educated mothers. Policies and resources targeting improvements in maternal mental health are essential.
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[
"Childbearing age has progressively increased in industrialized countries. The impact of this delay on motherhood, however, requires further research. The study sample included a prospective cohort of healthy nulliparous pregnant women aged between 18 and 40 years (n=148) assessed at 38 weeks gestation (Time#1, T1), 48h after birth (Time#2, T2), and 3 months after birth (Time#3, T3). The effect of age on psychological, biological, and social variables was evaluated. Maternal psychological symptoms in terms of depression and anxiety were assessed at T1-T3; and parenting stress at T3. Stress biomarkers (cortisol, α-amylase) were determined in mothers at T1-T3. Questionnaires addressing social functioning (i.e., family functioning, maternal attitudes, and social support) were conducted at T3. Bayesian additive models were used to analyze the data. Depressive symptoms showed a steep increase starting from 35 years of age at T1 and an U-shaped relationship with a minimum around 30 years old at T3. The same results were observed for parenting stress. Cortisol levels increased sharply from 30 years of age at T3. Family functioning, maternal attitudes, and social support improved moderately from 30 years of age. Prenatal depressive symptoms were higher in older women, but postpartum depressive symptoms and parenting stress increased in both younger and older women. Nevertheless, cortisol levels just increased in older ages at postpartum. In contrast, social functioning (family functioning, maternal attitudes, and social support) improved with age. We conclude that these social advantages may compensate for other disadvantages of delayed childbearing (i.e., depressive symptoms, parenting stress, and high cortisol level).",
"The COVID-19 pandemic has resulted in an unprecedented situation for new parents, with public health orders greatly affecting daily life as well as various aspects of parenting and new parent wellbeing. To understand the impact of the COVID-19 pandemic on mothers/parents across Nova Scotia who are caring for a child 0-12 months of age. This study utilized an online qualitative survey to collect data. Feminist poststructuralism and discourse analysis guided the analysis and discussion. Nova Scotia, Canada PARTICIPANTS: : 68 participants were recruited from across the province of Nova Scotia. Mental health and socialization were both major concerns for new mothers/parents, as many expressed feelings of worry, anxiety, loneliness, isolation, and stress. Online support was sought by many new mothers/parents as a way of supporting their own mental health. Some found ways to make it meaningful for them, while others believed that it could not replace or offer the same benefits as in-person interaction and support. Informal and formal support systems are both essential for new mothers. As public health systems and health care services learn to adapt to COVID-19, further research is required to examine how health services may best meet the needs of new mothers/parents.",
"Stay-at-home orders and the removal of care and domestic supports during the early days of the COVID-19 pandemic substantially disrupted US parents' work and family lives. Although much is known about changes in US parents' paid labor arrangements, the evidence regarding changes in unpaid domestic labor has been largely anecdotal. This study uses novel data from 1,025 US parents in different-sex partnerships to provide a descriptive overview of changes in mothers' and fathers' participation in, and division of, housework and childcare from March 2020 to the early days of the pandemic (late April 2020). Findings show an overall increase in domestic responsibilities for mothers who were already doing most of the household labor. Still, both mothers and fathers report a general shift toward more egalitarian divisions of household labor, driven by increases in fathers' contributions. The shift toward more egalitarian sharing of domestic labor is observed across demographic groups and across types of domestic tasks. Consistent with findings from other countries, egalitarian divisions of domestic labor increased among U.S. parents during the early days of the COVID-19 pandemic. Mothers, nonetheless, report retaining primary responsibility for domestic labor in the majority of families.",
"Objectives: Older adults' greater susceptibility to mortality from COVID-19 may have meaningful psychological implications not only for them, but also for their children. In this study, we focused on daughters of older women and examined the intergenerational relationships as a correlate of daughters' anxiety, depressive symptoms, and psychosomatic complaints.Method: Data were collected from 456 daughters of older mothers (M(age) = 40.82) during the first wave of the COVID-19 outbreak in Israel, when a relatively strict lockdown was enforced, separating mothers and daughters.Results: Findings suggest that while mothers' objective risk factors (age and morbidity) were mostly not associated with their daughters' distress, the daughters' concern about their mothers, and their perceived ambivalence in the relationship with the mother, as well as structural and affectual solidarity, were.Conclusion: We conclude that the mother-daughter relationship is an important correlate of daughters' reactions to this health crisis. Practically, it suggests that some daughters to aging mothers could be at a greater risk for emotional distress following the COVID-19 outbreak.",
"Although many young mothers (aged <21 years) are exposed to multiple adversities that increase their risk for mental illness, prevalence data are largely limited self-report questionnaires estimating only the prevalence of postpartum depression. Gaining a greater understanding of the burden of a broader range of common mental illnesses affecting these young women has the potential to improve their health as well as the development and functioning of their children. The Young Mothers Health Study recruited 450 mothers aged <21 years and 100 comparison mothers (aged >20 years old at first delivery) living in urban and rural central-west Ontario. Age-matched young mothers were also compared with 15- to 17-year-old women without children (N = 630) from the 2014 Ontario Child Health Study. The prevalence of current mental disorders was assessed using the Mini-International Neuropsychiatric Interview for Children and Adolescents. Nearly 2 of 3 young mothers reported at least one mental health problem, and almost 40% had more than one. Young mothers were 2 to 4 times as likely to have an anxiety disorder (generalized anxiety disorder, separation anxiety disorder, social phobia, and specific phobia), attention-deficit/hyperactivity disorder, oppositional defiant disorder, or conduct disorder and were 2 to 4 times more likely to have more than one psychiatric problem than older comparison mothers or women aged 15-17 years. Given the high rates of mental health problems and complex needs of young mothers in Canada and the possible adverse effects of maternal psychopathology on their children, further efforts should be directed at engaging and treating this high-risk group.",
"The results of 46 observational studies were analyzed to assess the strength of the association between depression and parenting behavior and to identify variables that moderated the effects. The association between depression and parenting was manifest most strongly for negative maternal behavior and was evident to a somewhat lesser degree in disengagement from the child. The association between depression and positive maternal behavior was relatively weak, albeit significant. Effects for negative maternal behavior were moderated by timing of the depression: Current depression was associated with the largest effects. However, residual effects of prior depression were apparent for all behaviors. Socioeconomic status, child age, and methodological variables moderated the effects for positive behavior: Effects were strongest for studies of disadvantaged women and mothers of infants. Studies using diagnostic interviews and self-report measures yielded similar effects, suggesting that deficits are not specific to depressive disorder. Research is needed to identify factors that affect the magnitude of parenting deficits among women who are experiencing depression and other psychological difficulties.",
"Using data from 747 rural African American mothers, this study incorporated role accumulation theory to test direct and indirect effects of stressors, coping behaviors, and role responsibilities on health functioning. Results indicated that demands emerging from financial strain were related to compromised mental health and decreases in mothers' use of effective coping strategies and role responsibility engagement. Conversely, mothers who effectively responded to stressors and fulfilled responsibilities to their children and communities experienced enhanced mental health, which in turn promoted optimal physical health. The results can inform research and intervention with African American women."
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Involvement of photosynthesis-related genes in resistance to soybean mosaic virus
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Increasing lines of evidence indicate that chloroplast-related genes are involved in plant-virus interactions. However, the involvement of photosynthesis-related genes in plant immunity is largely unexplored. Analysis of RNA-Seq data from the soybean cultivar L29, which carries the Rsv3 resistance gene, showed that several chloroplast-related genes were strongly induced in response to infection with an avirulent strain of soybean mosaic virus (SMV), G5H, but were weakly induced in response to a virulent strain, G7H. For further analysis, we selected the PSaC gene from the photosystem I and the ATP-synthase α-subunit (ATPsyn-α) gene whose encoded protein is part of the ATP-synthase complex. Overexpression of either gene within the G7H genome reduced virus levels in the susceptible cultivar Lee74 (rsv3-null). This result was confirmed by transiently expressing both genes in Nicotiana benthamiana followed by G7H infection. Both proteins localized in the chloroplast envelope as well as in the nucleus and cytoplasm. Because the chloroplast is the initial biosynthesis site of defence-related hormones, we determined whether hormone-related genes are involved in the ATPsyn-α- and PSaC-mediated defence. Interestingly, genes involved in the biosynthesis of several hormones were up-regulated in plants infected with SMV-G7H expressing ATPsyn-α. However, only jasmonic and salicylic acid biosynthesis genes were up-regulated following infection with the SMV-G7H expressing PSaC. Both chimeras induced the expression of several antiviral RNA silencing genes, which indicate that such resistance may be partially achieved through the RNA silencing pathway. These findings highlight the role of photosynthesis-related genes in regulating resistance to viruses.
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[
"Effector-triggered immunity (ETI) is an active immune response triggered by interactions between host resistance proteins and their cognate effectors. Although ETI is often associated with the hypersensitive response (HR), various R genes mediate an HR-independent process known as extreme resistance (ER). In the soybean-Soybean mosaic virus (SMV) pathosystem, the strain-specific CI protein of SMV functions as an effector of Rsv3-mediated ER. In this study, we used the soybean (Rsv3)-SMV (CI) pathosystem to gain insight into the molecular signaling pathway involved in ER. We used genome-wide transcriptome analysis to identify a subset of the type 2C protein phophatase (PP2C) genes that are specifically up-regulated in Rsv3-mediated ER. Gain-of-function analysis of the most significantly expressed soybean PP2C gene, GmPP2C3a, showed that ABA-induced GmPP2C3a functions as a key regulator of Rsv3-mediated ER. Our results further suggest that the primary mechanism of ER against viruses is the inhibition of viral cell-to-cell movement by callose deposition in an ABA signaling-dependent manner.",
"The cylindrical inclusion (CI) protein of potyviruses is involved in virus replication and cell-to-cell movement. These two processes should rely on multiple plant-virus interactions; however, little is known about the host factors that are involved in, or that may interfere with, CI functions. By using a yeast two-hybrid system, the CI protein from Plum pox virus (PPV) was found to interact with the photosystem I PSI-K protein, the product of the gene psaK, of Nicotiana benthamiana. Coexpression of PPV CI was shown to cause a decrease in the accumulation level of PSI-K transiently expressed in N. benthamiana leaves. To test the biological relevance of this interaction, we have analyzed the infection of PPV in N. benthamiana plants in which psaK gene expression has been silenced by RNA interference, as well as in Arabidopsis thaliana psaK knockout plants. Our results show that downregulation of the psaK gene leads to higher PPV accumulation, suggesting a role for the CI-PSI-K interaction in PPV infection.",
"Photosystem II (PSII) is a light-driven protein, involved in the primary reactions of photosynthesis. In plant photosynthetic membranes PSII forms large multisubunit supercomplexes, containing a dimeric core and up to four light-harvesting complexes (LHCs), which act as antenna proteins. Here we solved a three-dimensional (3D) structure of the C2S2M2 supercomplex from Arabidopsis thaliana using cryo-transmission electron microscopy (cryo-EM) and single-particle analysis at an overall resolution of 5.3 Å. Using a combination of homology modelling and restrained refinement against the cryo-EM map, it was possible to model atomic structures for all antenna complexes and almost all core subunits. We located all 35 chlorophylls of the core region based on the cyanobacterial PSII structure, whose positioning is highly conserved, as well as all the chlorophylls of the LHCII S and M trimers. A total of 13 and 9 chlorophylls were identified in CP26 and CP24, respectively. Energy flow from LHC complexes to the PSII reaction centre is proposed to follow preferential pathways: CP26 and CP29 directly transfer to the core using several routes for efficient transfer; the S trimer is directly connected to CP43 and the M trimer can efficiently transfer energy to the core through CP29 and the S trimer.",
"Redox-controlled phosphorylation of thylakoid membrane proteins represents a unique system for the regulation of light energy utilization in photosynthesis. The molecular mechanisms for this process remain unknown, but current views suggest that the plastoquinone pool directly controls the activation of the kinase. On the basis of enzyme activation by a pH shift in the darkness combined with flash photolysis, EPR, and optical spectroscopy we propose that activation occurs when plastoquinol occupies the quinol-oxidation (Qo) site of the cytochrome bf complex, having its high-potential path components in a reduced state. A linear correlation between kinase activation and accessibility of the Qo site to plastoquinol was established by quantification of the shift in the g(y) EPR signal of the Rieske Fe-S center resulting from displacement of the Qo-site plastoquinol by a quinone analog. Activity persists as long as one plastoquinol per cytochrome bf is still available. Withdrawal of one electron from this plastoquinol after a single-turnover flash exciting photosystem I leads to deactivation of the kinase parallel with a decrease in the g(z) EPR signal of the reduced Rieske Fe-S center. Cytochrome f, plastocyanin, and P(700) are rereduced after the flash, indicating that the plastoquinol at the Qo site is limiting in maintaining the kinase activity. These results give direct evidence for a functional cytochrome bf-kinase interaction, analogous to a signal transduction system where the cytochrome bf is the receptor and the ligand is the plastoquinol at the Qo site.",
"The diffusion of proteins in chloroplast thylakoid membranes is believed to be important for processes including the photosystem-II repair cycle and the regulation of light harvesting. However, to date there is very little direct information on the mobility of thylakoid proteins. We have used fluorescence recovery after photobleaching in a laser-scanning confocal microscope to visualize in real time the exchange of chlorophyll proteins between grana in intact spinach (Spinacia oleracea L.) and Arabidopsis chloroplasts. Most chlorophyll proteins in the grana appear immobile on the 10-min timescale of our measurements. However, a limited population of chlorophyll proteins (accounting for around 15% of chlorophyll fluorescence) can exchange between grana on this timescale. In intact, wild-type chloroplasts this mobile population increases significantly after photoinhibition, consistent with a role for protein diffusion in the photosystem-II repair cycle. No such increase in mobility is seen in isolated grana membranes, or in the Arabidopsis stn8 and stn7 stn8 mutants, which lack the protein kinases required for phosphorylation of photosystem II core proteins and light-harvesting complexes. Furthermore, mobility under low-light conditions is significantly lower in stn8 and stn7 stn8 plants than in wild-type Arabidopsis. The changes in protein mobility correlate with changes in the packing density and size of thylakoid protein complexes, as observed by freeze-fracture electron microscopy. We conclude that protein phosphorylation switches the membrane system to a more fluid state, thus facilitating the photosystem-II repair cycle.",
"The photosystems (PS), catalyzing the photosynthetic reactions of higher plants, are unevenly distributed in the thylakoid membrane: PSII, together with its light harvesting complex (LHC)II, is enriched in the appressed grana stacks, while PSI-LHCI resides in the non-appressed stroma thylakoids, which wind around the grana stacks. The two photosystems interact in a third membrane domain, the grana margins, which connect the grana and stroma thylakoids and allow the loosely bound LHCII to serve as an additional antenna for PSI. The light harvesting is balanced by reversible phosphorylation of LHCII proteins. Nevertheless, light energy also damages PSII and the repair process is regulated by reversible phosphorylation of PSII core proteins. Here, we discuss the detailed composition and organization of PSII-LHCII and PSI-LHCI (super)complexes in the thylakoid membrane of angiosperm chloroplasts and address the role of thylakoid protein phosphorylation in dynamics of the entire protein complex network of the photosynthetic membrane. Finally, we scrutinize the phosphorylation-dependent dynamics of the protein complexes in context of thylakoid ultrastructure and present a model on the reorganization of the entire thylakoid network in response to changes in thylakoid protein phosphorylation.",
"Extreme resistance (ER) is a type of R-gene-mediated resistance that rapidly induces a symptomless resistance phenotype, which is different from the phenotypical R-resistance manifested by the programmed cell death, accumulation of reactive oxygen species, and hypersensitive response. The Rsv3 gene in soybean cultivar L29 is responsible for ER against the avirulent strain G5H of soybean mosaic virus (SMV), but is ineffective against the virulent strain G7H. Rsv3-mediated ER is achieved through the rapid accumulation of callose, which arrests SMV-G5H at the point of infection. Callose accumulation, however, may not be the lone mechanism of this ER. Analyses of RNA-seq data obtained from infected soybean plants revealed a rapid induction of the abscisic acid pathway at 8 h post infection (hpi) in response to G5H but not to G7H, which resulted in the down-regulation of transcripts encoding β-1,3 glucanases that degrade callose in G5H-infected but not G7H-infected plants. In addition, parts of the autophagy and the small interfering (si) RNA pathways were temporally up-regulated at 24 hpi in response to G5H but not in response to G7H. The jasmonic acid (JA) pathway and many WRKY factors were clearly up-regulated only in G7H-infected plants. These results suggest that ER against SMV-G5H is achieved through the quick and temporary induction of ABA, autophagy, and the siRNA pathways, which rapidly eliminate G5H. The results also suggest that suppression of the JA pathway in the case of G5H is important for the Rsv3-mediated ER.",
"Callose synthesis occurs at specific stages of plant cell wall development in all cell types, and in response to pathogen attack, wounding and physiological stresses. We determined the expression pattern of \"upstream regulatory sequence\" of 12 Arabidopsis callose synthase genes (CalS1-12) genes and demonstrated that different callose synthases are expressed specifically in different tissues during plant development. That multiple CalS genes are expressed in the same cell type suggests the possibility that CalS complex may be constituted by heteromeric subunits. Five CalS genes were induced by pathogen (Hyaloperonospora arabidopsis, previously known as Peronospora parasitica, the causal agent of downy mildew) or salicylic acid (SA), while the other seven CalS genes were not affected by these treatments. Among the genes that are induced, CalS1 and CalS12 showed the highest responses. In Arabidopsis npr1 mutant, impaired in response of pathogenesis related (PR) genes to SA, the induction of CalS1 and CalS12 genes by the SA or pathogen treatments was significantly reduced. The patterns of expression of the other three CalS genes were not changed significantly in the npr1 mutant. These results suggest that the high induction observed of CalS1 and CalS12 is Npr1 dependent while the weak induction of five CalS genes is Npr1 independent. In a T-DNA knockout mutant of CalS12, callose encasement around the haustoria on the infected leaves was reduced and the mutant was found to be more resistant to downy mildew as compared to the wild type plants.",
"Plants regulate photosynthetic light harvesting to maintain balanced energy flux into photosystems I and II (PSI and PSII). Under light conditions favoring PSII excitation, the PSII antenna, light-harvesting complex II (LHCII), is phosphorylated and forms a supercomplex with PSI core and the PSI antenna, light-harvesting complex I (LHCI). Both LHCI and LHCII then transfer excitation energy to the PSI core. We report the structure of maize PSI-LHCI-LHCII solved by cryo-electron microscopy, revealing the recognition site between LHCII and PSI. The PSI subunits PsaN and PsaO are observed at the PSI-LHCI interface and the PSI-LHCII interface, respectively. Each subunit relays excitation to PSI core through a pair of chlorophyll molecules, thus revealing previously unseen paths for energy transfer between the antennas and the PSI core.",
"PsaI represents one of three low molecular weight peptides of PSI. Targeted inactivation of the plastid PsaI gene in Nicotiana tabacum has no measurable effect on photosynthetic electron transport around PSI or on accumulation of proteins involved in photosynthesis. Instead, the lack of PsaI destabilizes the association of PsaL and PsaH to PSI, both forming the light-harvesting complex (LHC)II docking site of PSI. These alterations at the LHCII binding site surprisingly did not prevent state transition but led to an increased incidence of PSI-LHCII complexes, coinciding with an elevated phosphorylation level of the LHCII under normal growth light conditions. Remarkably, LHCII was rapidly phosphorylated in ΔpsaI in darkness even after illumination with far-red light. We found that this dark phosphorylation also occurs in previously described mutants impaired in PSI function or state transition. A prompt shift of the plastoquinone (PQ) pool into a more reduced redox state in the dark caused an enhanced LHCII phosphorylation in ΔpsaI Since the redox status of the PQ pool is functionally connected to a series of physiological, biochemical, and gene expression reactions, we propose that the shift of mutant plants into state 2 in darkness represents a compensatory and/or protective metabolic mechanism. This involves an increased reduction and/or reduced oxidation of the PQ pool, presumably to sustain a balanced excitation of both photosystems upon the onset of light."
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Preliminary study of 3D printed spacers containing antibiotics for joint replacement surgery
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In recent years, the use of 3D printing technologies in orthopedic surgery has markedly increased, as they offer the possibility of printing personalized prostheses. The work presented in this article is a preliminary study of a research project which aims to manufacture customized spacers containing antibiotics for use in joint replacement surgery. The objective of this work was to design and print different 3D constructs to evaluate the use of different materials, their properties after the process of 3D printing, such as resistance, and the release kinetics of drugs from the constructs. Different designs and different materials were analyzed to obtain a 3D construct with suitable properties. Our design takes advantage of the micropores created between the layers of the 3D printed filaments to release the contained drug. Using polylactic acid (PLA) we were able to print cylindrical structures with interconnected micropores and a hollow chamber capable of releasing methylene blue, which was selected as a model drug. The final PLA 3D construct was printed with a 10% infill. The physical and technological characteristics, morphological changes at body temperature and interaction with water were considered to be acceptable. The PLA 3D printed constructs were found to have sufficient strength to withstand a force of 500 kg. The results obtained allow to continue research in this project, with the aim of manufacturing prostheses containing a reservoir of antibiotics or other drugs in their interior for their subsequent controlled release.
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[
"Bacterial infection remains a significant complication following total joint replacement. If infection is suspected when revision surgery is being performed, a large dose of antibiotic, usually gentamicin sulphate, is often blended with the acrylic bone cement powder in an attempt to reduce the risk of recurrent infection. In this in-vitro study the effect of small and large doses of gentamicin sulphate on the handling and mechanical properties of the cement, gentamicin release from the cement, and in-vitro biofilm formation by clinical Staphylococcus spp. isolates on the cement was determined. An increase in gentamicin loading of 1, 2, 3, or 4 g, in a cement powder mass of 40 g, resulted in a significant decrease in the compressive and four-point bending strength, but a significant increase in the amount of gentamicin released over a 72h period. When overt infection was modelled, using Staphylococcus spp. clinical isolates at an inoculum of 1 x 10(7) colony-forming units/ml, an increase in the amount of gentamicin (1, 2, 3, or 4 g) added to 40 g of poly(methyl methacrylate) cement resulted in an initial decrease in bacterial colonization but this beneficial effect was no longer apparent by 72 h, with the bacterial strains forming biofilms on the cements despite the release of high levels of gentamicin. The findings suggest that orthopaedic surgeons should carefully consider the clinical consequences of blending large doses (1 g or more per 40 g of poly(methyl methacrylate)) of gentamicin into Palacos R bone cement for use in revision surgery as the increased gentamicin loading does not prevent bacterial biofilm formation and the effect on the mechanical properties could be important to the longevity of the prosthetic joint.",
"High-dose antibiotic-loaded bone cement (ALBC) spacers are commonly used to treat prosthetic joint infections following total hip and knee arthroplasties. This methodology can provide high local antibiotic concentrations while minimizing systemic exposure and toxicity. The occurrence of acute kidney injury (AKI) is rarely reported. Available literature suggests that the rate may be higher than previously thought. We report a case of significant systemic tobramycin absorption with concomitant acute renal failure in a 69-year-old female following the implantation of a high-dose ALBC spacer containing both tobramycin and vancomycin. The tobramycin level 24 h post-surgery was 5.8 mcg/mL. Due to concomitant renal failure, antibiotic clearance was poor and resulted in prolonged exposure to elevated aminoglycoside levels. Recovery of renal function occurred, but clinicians should be vigilant in considering the potential impact ALBC spacers can have on post-operative renal function if antibiotic elution is higher than expected.",
"The use of 3D printing in Orthopedics is set to transform the way surgeries are planned and executed. The development of X rays and later the CT scan and MRI enabled surgeons to understand the anatomy and condition better and helped plan surgeries on images obtained. 3DGraphy a term used for 3D printed orthopedic patient models and Jigs has gone a step further by providing surgeons with a physical copy of the patient's affected part that can not only be seen but also felt and moved around spatially. Similarly 3D printed Jigs are patient specific devices that are used to ensure optimal screw trajectory and implant placement with minimal exposure. While the use of 3D printed models and Jigs have now become routine, a similar revolution is happening in the field of designing and printing patient specific implants. Metal printing along with enhanced capability to print other biocompatible materials like PEEK and PLA is likely to improve the current implant manufacturing process. On the horizon is another interesting development related to this field - 3D Bio printing. Printing human tissues and organs is considered the final frontier and impressive strides have been made in printing bone graft substitutes and cartilage like material. This paper is an overview of all the current developments and the road ahead in this invigorating field.",
"To investigate the feasibility of the use of 3D-printed guiding templates for accurate placement of plates and screws for internal fixation of acetabular fractures. 3D models of the pelvises of 14 adult cadavers were reconstructed using computed tomography (CT). Twenty-eight acetabular fractures were simulated and placement positions for plates and screw trajectories were designed. Bending module was obtained by 3D cutting; guiding template was manufactured using 3D printing, and the plate was pre-bent according to the bending module. Plates and screws were placed in cadaveric pelvises using the guiding template, and 3D model was reconstructed using CT. The designed and real trajectories were matched using 3D registration including the coordinates (entry and exit points) of designed trajectory. The number of qualified points with different accuracy levels was compared using Chi-squared test. Sixty-four plates and 339 screws were placed with no cortical breach. The absolute difference of the X, Y, and Z coordinates between the designed and real entry points were 0.52±0.45, 0.43±0.36, and 0.53±0.44mm, respectively. The corresponding values for the exit points were 0.83±0.67, 1.22±0.87, and 1.26±0.83mm, respectively. With an accuracy degree ≥1.9mm for the entry points and ≥3.8mm for the exit points, there was no significant difference between the designed and the real trajectories. The 3D-printed guiding template helped achieve accurate placement of plates and screws in the pelvis of adult cadavers."
] |
UCSF Immunoprofiler Initiative: Understanding Cancer Immunity as a Collection of Dominant Patterns of Immune Organization
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Cancers display significant heterogeneity with respect to tissue of origin, driver mutations, and other features of the surrounding tissue. It is likely that individual tumors engage common patterns of the immune system-here "archetypes"-creating prototypical non-destructive tumor immune microenvironments (TMEs) and modulating tumor-targeting. To discover the dominant immune system archetypes, the University of California, San Francisco (UCSF) Immunoprofiler Initiative (IPI) processed 364 individual tumors across 12 cancer types using standardized protocols. Computational clustering of flow cytometry and transcriptomic data obtained from cell sub-compartments uncovered dominant patterns of immune composition across cancers. These archetypes were profound insofar as they also differentiated tumors based upon unique immune and tumor gene-expression patterns. They also partitioned well-established classifications of tumor biology. The IPI resource provides a template for understanding cancer immunity as a collection of dominant patterns of immune organization and provides a rational path forward to learn how to modulate these to improve therapy.
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[
"Knowledge of immune cell phenotypes in the tumor microenvironment is essential for understanding mechanisms of cancer progression and immunotherapy response. We profiled 45,000 immune cells from eight breast carcinomas, as well as matched normal breast tissue, blood, and lymph nodes, using single-cell RNA-seq. We developed a preprocessing pipeline, SEQC, and a Bayesian clustering and normalization method, Biscuit, to address computational challenges inherent to single-cell data. Despite significant similarity between normal and tumor tissue-resident immune cells, we observed continuous phenotypic expansions specific to the tumor microenvironment. Analysis of paired single-cell RNA and T cell receptor (TCR) sequencing data from 27,000 additional T cells revealed the combinatorial impact of TCR utilization on phenotypic diversity. Our results support a model of continuous activation in T cells and do not comport with the macrophage polarization model in cancer. Our results have important implications for characterizing tumor-infiltrating immune cells.",
"Tumor infiltrating leukocytes (TILs) are an integral component of the tumor microenvironment and have been found to correlate with prognosis and response to therapy. Methods to enumerate immune subsets such as immunohistochemistry or flow cytometry suffer from limitations in phenotypic markers and can be challenging to practically implement and standardize. An alternative approach is to acquire aggregative high dimensional data from cellular mixtures and to subsequently infer the cellular components computationally. We recently described CIBERSORT, a versatile computational method for quantifying cell fractions from bulk tissue gene expression profiles (GEPs). Combining support vector regression with prior knowledge of expression profiles from purified leukocyte subsets, CIBERSORT can accurately estimate the immune composition of a tumor biopsy. In this chapter, we provide a primer on the CIBERSORT method and illustrate its use for characterizing TILs in tumor samples profiled by microarray or RNA-Seq.",
"Tumour-specific CD8 T cell dysfunction is a differentiation state that is distinct from the functional effector or memory T cell states1-6. Here we identify the nuclear factor TOX as a crucial regulator of the differentiation of tumour-specific T (TST) cells. We show that TOX is highly expressed in dysfunctional TST cells from tumours and in exhausted T cells during chronic viral infection. Expression of TOX is driven by chronic T cell receptor stimulation and NFAT activation. Ectopic expression of TOX in effector T cells in vitro induced a transcriptional program associated with T cell exhaustion. Conversely, deletion of Tox in TST cells in tumours abrogated the exhaustion program: Tox-deleted TST cells did not upregulate genes for inhibitory receptors (such as Pdcd1, Entpd1, Havcr2, Cd244 and Tigit), the chromatin of which remained largely inaccessible, and retained high expression of transcription factors such as TCF-1. Despite their normal, 'non-exhausted' immunophenotype, Tox-deleted TST cells remained dysfunctional, which suggests that the regulation of expression of inhibitory receptors is uncoupled from the loss of effector function. Notably, although Tox-deleted CD8 T cells differentiated normally to effector and memory states in response to acute infection, Tox-deleted TST cells failed to persist in tumours. We hypothesize that the TOX-induced exhaustion program serves to prevent the overstimulation of T cells and activation-induced cell death in settings of chronic antigen stimulation such as cancer.",
"Cancer therapy is challenged by the diversity of molecular implementations of oncogenic processes and by the resulting variation in therapeutic responses. Projects such as The Cancer Genome Atlas (TCGA) provide molecular tumor maps in unprecedented detail. The interpretation of these maps remains a major challenge. Here we distilled thousands of genetic and epigenetic features altered in cancers to ∼500 selected functional events (SFEs). Using this simplified description, we derived a hierarchical classification of 3,299 TCGA tumors from 12 cancer types. The top classes are dominated by either mutations (M class) or copy number changes (C class). This distinction is clearest at the extremes of genomic instability, indicating the presence of different oncogenic processes. The full hierarchy shows functional event patterns characteristic of multiple cross-tissue groups of tumors, termed oncogenic signature classes. Targetable functional events in a tumor class are suggestive of class-specific combination therapy. These results may assist in the definition of clinical trials to match actionable oncogenic signatures with personalized therapies.",
"An improved understanding of the anti-tumor CD8+ T cell response after checkpoint blockade would enable more informed and effective therapeutic strategies. Here we examined the dynamics of the effector response of CD8+ tumor-infiltrating lymphocytes (TILs) after checkpoint blockade therapy. Bulk and single-cell RNA profiles of CD8+ TILs after combined Tim-3+PD-1 blockade in preclinical models revealed significant changes in the transcriptional profile of PD-1- TILs. These cells could be divided into subsets bearing characterstics of naive-, effector-, and memory-precursor-like cells. Effector- and memory-precursor-like TILs contained tumor-antigen-specific cells, exhibited proliferative and effector capacity, and expanded in response to different checkpoint blockade therapies across different tumor models. The memory-precursor-like subset shared features with CD8+ T cells associated with response to checkpoint blockade in patients and was compromised in the absence of Tcf7. Expression of Tcf7/Tcf1 was requisite for the efficacy of diverse immunotherapies, highlighting the importance of this transcriptional regulator in the development of effective CD8+ T cell responses upon immunotherapy.",
"The role of the adaptive immune response in controlling the growth and recurrence of human tumors has been controversial. We characterized the tumor-infiltrating immune cells in large cohorts of human colorectal cancers by gene expression profiling and in situ immunohistochemical staining. Collectively, the immunological data (the type, density, and location of immune cells within the tumor samples) were found to be a better predictor of patient survival than the histopathological methods currently used to stage colorectal cancer. The results were validated in two additional patient populations. These data support the hypothesis that the adaptive immune response influences the behavior of human tumors. In situ analysis of tumor-infiltrating immune cells may therefore be a valuable prognostic tool in the treatment of colorectal cancer and possibly other malignancies.",
"Wounding, apoptosis, or infection can trigger a proliferative response in neighboring cells to replace damaged tissue. Studies in Drosophila have implicated c-Jun amino-terminal kinase (JNK)-dependent activation of Yorkie (Yki) as essential to regeneration-associated growth, as well as growth associated with neoplastic tumors. Yki is a transcriptional coactivator that is inhibited by Hippo signaling, a conserved pathway that regulates growth. We identified a conserved mechanism by which JNK regulated Hippo signaling. Genetic studies in Drosophila identified Jub (also known as Ajuba LIM protein) as required for JNK-mediated activation of Yki and showed that Jub contributed to wing regeneration after wounding and to tumor growth. Biochemical studies revealed that JNK promoted the phosphorylation of Ajuba family proteins in both Drosophila and mammalian cells. Binding studies in mammalian cells indicated that JNK increased binding between the Ajuba family proteins LIMD1 or WTIP and LATS1, a kinase within the Hippo pathway that inhibits the Yki homolog YAP. Moreover, JNK promoted binding of LIMD1 and LATS1 through direct phosphorylation of LIMD1. These results identify Ajuba family proteins as a conserved link between JNK and Hippo signaling, and imply that JNK increases Yki and YAP activity by promoting the binding of Ajuba family proteins to Warts and LATS."
] |
are unsaturated fatty acids allowed in canada?
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The 2019 Canada's Food Guide (CFG) recommends that foods containing mostly unsaturated fatty acid (UFA) should replace foods that contain mostly SFA to reduce SFA intakes.
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[
"The 2019 revised version of Canada's Food Guide (CFG) recommends limiting the consumption of processed foods that are high in saturated fatty acids (SFA). Yet, the contributions of each CFG group to the total SFA intake of Canadians are not specifically known. The objectives of this study were to quantify the total SFA intake of Canadians, determine the sources of SFA consumed by Canadian adults, and identify potential differences in these sources. A nation representative sample from the Canadian Community Health Survey (CCHS - Nutrition 2015) was used for these analyses. Dietary intakes were measured using a single 24-h recall. Food sources of SFA were classified according to the revised 2019 CFG categories. We have also examined the contribution of foods not included in these three categories to total SFA intake. Among Canadian adults, total SFA contributed to 10.4 ± 0.1% (SE) of total energy intake (E). The \"Protein foods\" (47.7 ± 0.5% with 23.2 ± 0.4% from milk and alternatives and 24.5 ± 0.4% from meats and alternatives) and \"All other foods\" (44.2 ± 0.5%) categories were the main sources of total SFA intake. Few differences in SFA sources were identified between sexes, age groups, education levels, and body mass index (BMI) categories. These data show that the mean SFA consumption is greater than the 10% E cut-off previously proposed in Canada. Future studies should examine which food substitution is most likely to contribute to a greater reduction in SFA intake at the population level.",
"Estimating the population distribution of the usual intake of a nutrient relative to that of another nutrient requires determination of individual-level ratios. If intake data are available on a per-day basis, as with 24-h dietary recalls, those ratios can be determined in 1 of 2 ways: as the usual ratio of intakes or the ratio of usual intakes. Each of these ratios has its own meaning and determination; the ratio of usual intakes is conceptually consistent with determinations obtained from FFQ data. We present a method for estimating the ratio of usual intakes that uses bivariate modeling of the 2 nutrient intakes in question. Application of the method to the NHANES data for the years 2001-2004 yielded estimated distributions for percent of usual energy intake from total fat, percent of usual energy intake from saturated fat, and usual sodium intake per 1000 kcal (4184 kJ) of usual energy intake. Distributions for both the total population and for age-gender subgroups were estimated. Approximately 60% of adults (>19 y) had a usual total fat intake that was within the recommended range of 20-35% of total energy, but only approximately 34% had a usual saturated fat intake <10% of total energy. The results changed only minimally when the other definition of usual intake, the usual ratio of intakes, was adopted.",
"Dietary factors are major contributors to morbidity and mortality, and significant attention is being paid to interventions to support healthy eating, including through the creation of a healthier food supply. The objective of this study was to inform interventions to support healthy eating by examining the top dietary sources of calories, sodium, sugars, and saturated fats among Canadians in relation to sex, age, and income. We drew upon data from the 2015 Canadian Community Health Survey, which collected interviewer-administered 24-h dietary recalls from Canadians who were 1 year of age and older (n = 20 176), residing in the 10 provinces. Foods and beverages were grouped into 91 mutually exclusive categories (e.g., 100% fruit juice, fruit drinks). On the basis of the average proportion contributed, the top 20 sources of each dietary component were identified for all individuals and by sex-age and income groups. The mean amount of each dietary component contributed by each category (per capita and per consumer) and the proportions of persons consuming items in each category were also examined. Top sources included commonly consumed items (e.g., breads and flatbreads as sources of sodium), as well as those high in a given dietary component (e.g., soda as a source of sugars). Several food and beverage categories were top contributors to more than one dietary component examined, suggesting possible priorities for intervention and future analyses. The identification of major sources of calories and nutrients of concern can inform population health efforts, such as reformulation, to improve the health of Canadians."
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How do I find the genus of a plant?
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The genus
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[
"Ribosomal RNA genes have become the standard molecular markers for microbial community analysis for good reasons, including universal occurrence in cellular organisms, availability of large databases, and ease of rRNA gene region amplification and analysis. As markers, however, rRNA genes have some significant limitations. The rRNA genes are often present in multiple copies, unlike most protein-coding genes. The slow rate of change in rRNA genes means that multiple species sometimes share identical 16S rRNA gene sequences, while many more species share identical sequences in the short 16S rRNA regions commonly analyzed. In addition, the genes involved in many important processes are not distributed in a phylogenetically coherent manner, potentially due to gene loss or horizontal gene transfer. While rRNA genes remain the most commonly used markers, key genes in ecologically important pathways, e.g., those involved in carbon and nitrogen cycling, can provide important insights into community composition and function not obtainable through rRNA analysis. However, working with ecofunctional gene data requires some tools beyond those required for rRNA analysis. To address this, our Functional Gene Pipeline and Repository (FunGene; http://fungene.cme.msu.edu/) offers databases of many common ecofunctional genes and proteins, as well as integrated tools that allow researchers to browse these collections and choose subsets for further analysis, build phylogenetic trees, test primers and probes for coverage, and download aligned sequences. Additional FunGene tools are specialized to process coding gene amplicon data. For example, FrameBot produces frameshift-corrected protein and DNA sequences from raw reads while finding the most closely related protein reference sequence. These tools can help provide better insight into microbial communities by directly studying key genes involved in important ecological processes.",
"Genome sequencing enhances our understanding of the biological world by providing blueprints for the evolutionary and functional diversity that shapes the biosphere. However, microbial genomes that are currently available are of limited phylogenetic breadth, owing to our historical inability to cultivate most microorganisms in the laboratory. We apply single-cell genomics to target and sequence 201 uncultivated archaeal and bacterial cells from nine diverse habitats belonging to 29 major mostly uncharted branches of the tree of life, so-called 'microbial dark matter'. With this additional genomic information, we are able to resolve many intra- and inter-phylum-level relationships and to propose two new superphyla. We uncover unexpected metabolic features that extend our understanding of biology and challenge established boundaries between the three domains of life. These include a novel amino acid use for the opal stop codon, an archaeal-type purine synthesis in Bacteria and complete sigma factors in Archaea similar to those in Bacteria. The single-cell genomes also served to phylogenetically anchor up to 20% of metagenomic reads in some habitats, facilitating organism-level interpretation of ecosystem function. This study greatly expands the genomic representation of the tree of life and provides a systematic step towards a better understanding of biological evolution on our planet.",
"Utricularia are rootless aquatic carnivorous plants which have recently attracted the attention of researchers due to the peculiarities of their miniaturized genomes. Here, we focus on a novel aspect of Utricularia ecophysiology-the interactions with and within the complex communities of microorganisms colonizing their traps and external surfaces. Bacteria, fungi, algae, and protozoa inhabit the miniature ecosystem of the Utricularia trap lumen and are involved in the regeneration of nutrients from complex organic matter. By combining molecular methods, microscopy, and other approaches to assess the trap-associated microbial community structure, diversity, function, as well as the nutrient turn-over potential of bacterivory, we gained insight into the nutrient acquisition strategies of the Utricularia hosts. We conclude that Utricularia traps can, in terms of their ecophysiological function, be compared to microbial cultivators or farms, which center around complex microbial consortia acting synergistically to convert complex organic matter, often of algal origin, into a source of utilizable nutrients for the plants.",
"The alignment of sequencing reads against a protein reference database is a major computational bottleneck in metagenomics and data-intensive evolutionary projects. Although recent tools offer improved performance over the gold standard BLASTX, they exhibit only a modest speedup or low sensitivity. We introduce DIAMOND, an open-source algorithm based on double indexing that is 20,000 times faster than BLASTX on short reads and has a similar degree of sensitivity."
] |
Mitochondrial and Cholinergic Dysfunction in TgF344-AD Rats
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Glucose hypometabolism, mitochondrial dysfunction, and cholinergic deficits have been reported in early stages of Alzheimer's disease (AD). Here, we examine these parameters in TgF344-AD rats, an Alzheimer model that carries amyloid precursor protein and presenilin-1 mutations, and of wild type F344 rats. In mitochondria isolated from rat hippocampi, we found reductions of complex I and oxidative phosphorylation in transgenic rats. Further impairments, also of complex II, were observed in aged (wild-type and transgenic) rats. Treatment with a "cocktail" containing magnesium orotate, benfotiamine, folic acid, cyanocobalamin, and cholecalciferol did not affect mitochondrial activities in wild-type rats but restored diminished activities in transgenic rats to wild-type levels. Glucose, lactate, and pyruvate levels were unchanged by age, genetic background, or treatment. Using microdialysis, we also investigated extracellular concentrations of acetylcholine that were strongly reduced in transgenic animals. Again, ACh levels in wild-type rats did not change upon treatment with nutrients, whereas the cocktail increased hippocampal acetylcholine levels under physiological stimulation. We conclude that TgF344-AD rats display a distinct mitochondrial and cholinergic dysfunction not unlike the findings in patients suffering from AD. This dysfunction can be partially corrected by the application of the "cocktail" which is particularly active in aged rats. We suggest that the TgF344-AD rat is a promising model to further investigate mitochondrial and cholinergic dysfunction and potential treatment approaches for AD.
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[
"Tau becomes characteristically altered both functionally and structurally in several neurodegenerative diseases now collectively called tauopathies. Although increasing evidence supports that alterations of tau may directly cause neuronal degeneration and cell death, the mechanisms, which render tau to become a toxic agent are still unclear. In addition, it is obscure, whether neurodegeneration in tauopathies occurs via a common mechanism or specific differences exist. The aim of this review is to provide an overview about the different experimental models that currently exist, how they are used to determine the role of tau during degeneration and what has been learnt from them concerning the mechanistic role of tau in the disease process. The review begins with a discussion about similarities and differences in tau alteration in paradigmatic tauopathies such as frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) and Alzheimer's disease (AD). The second part concentrates on major experimental models that have been used to address the mechanistic role of tau during degeneration. This will include a discussion of cell-free assays, culture models using cell lines or dissociated neurons, and animal models. How these models aid to understand (i) alterations in the function of tau as a microtubule-associated protein (MAP), (ii) direct cytotoxicity of altered tau protein, and (iii) the potential role of tau aggregation in neurodegenerative processes will be the central theme of this part. The review ends with concluding remarks about a general mechanistic model of the role of tau alteration and neuronal degeneration in tauopathies and future perspectives.",
"For nonhumans, it has been shown that the hippocampus (HPC) is critical for spatial memory. We tested patients with unilateral HPC resections on a virtual analogue of a classic spatial task to assess HPC functioning in nonhumans: the Morris water task. We found that when humans are required to use spatial cues to navigate to a hidden escape platform in a pool, patients with HPC resections display severe impairments in spatial navigation relative to age-matched controls and age-matched patients who have had extra-HPC resections. This effect occurred for every patient tested and was evident regardless of side of surgery. Hence, it is apparent across species and irrespective of which hemisphere is damaged that the human HPC is critical for spatial/relational memory.",
"The purpose of our study was to understand the toxic effects of hippocampal phosphorylated tau in tau mice. Using rotarod and Morris water maze (MWM) tests, immunoblotting and immunofluorescence, Golgi-Cox staining and transmission electron microscopy, we assessed cognitive behavior, measured protein levels of mitochondrial dynamics, MAP2, total and phosphorylated tau, and quantified dendritic spines and mitochondrial number and length in 12-month-old tau mice with P301L mutation. Mitochondrial function was assessed by measuring the levels of H2O2, lipid peroxidation, cytochrome oxidase activity and mitochondrial ATP. MWM and rotarod tests revealed that hippocampal learning and memory and motor learning and coordination were impaired in tau mice relative to wild-type (WT) mice. Increased levels of mitochondrial fission proteins, Drp1 and Fis1 and decreased levels of mitochondrial fusion proteins, Mfn1, Mfn2 and Opa1 were found in 12-month-old tau mice relative to age-matched WT mice, indicating that the presence of abnormal mitochondrial dynamics in tau mice. Decreased levels of dendritic protein, MAP2 and increased levels of total and phosphorylated tau proteins were found in tau mice relative to WT mice. Mitochondrial function was defective. Golgi-Cox staining analysis revealed that dendritic spines are significantly reduced. Transmission electron microscopy revealed significantly increased mitochondrial numbers and reduced mitochondrial length in tau mice. These findings suggest that hippocampal accumulation of phosphorylated tau is responsible for abnormal mitochondrial dynamics and reducing dendritic protein MAP2 and dendritic spines and hippocampal based learning and memory impairments, and mitochondrial structural and functional changes in tau mice. Based on these observations, we propose that reduced hippocampal phosphorylated tau is an important therapeutic strategy for AD and other tauopathies.",
"Alzheimer's disease (AD) is a chronic progressive neurodegenerative disease, but the pathogenesis is unclear. Damaged mitochondrial biogenesis has been observed in AD. Increasing evidence suggests that mitochondrial biogenesis is involved in the pathogenesis of AD, but the exact mechanism is unclear. In this study, we used the amyloid precursor protein Swedish mutations K594N/M595L (APPswe)/presenilin 1 with the exon-9 deletion (PS1dE9) transgenic mouse model of AD, which was successfully established by the expression of amyloid β precursor protein and presenilin 1 (PS1). Then, we compared APPswe/PS1dE9 transgenic mice with and without melatonin (MT) in drinking water for 4 months (estimated 0.5 mg/day) and control C57BL/6J mice without MT for expression of mitochondrial biogenesis factors (mitochondrial transcription factor A, nuclear respiratory factor 1 and 2, peroxisome proliferator-activated receptor γ coactivator 1-α), mitochondrial structure, mitochondrial DNA to nuclear DNA ratio, behavioral changes, and amyloid β (Aβ) deposition and soluble Aβ levels in the cerebral cortex and hippocampus. Compared with controls, APPswe/PS1dE9 mice with long-term MT intake showed increased levels of mitochondrial biogenesis factors, alleviated mitochondrial impairment, enhanced mitochondrial DNA copy number, improved spatial learning and memory deficits, and reduced Aβ deposition and soluble Aβ levels. Defective mitochondrial biogenesis may contribute toward the damaged mitochondrial structure and function in AD. MT may alleviate AD by promoting mitochondrial biogenesis.",
"Causes of initiation and progression of sporadic Alzheimer's disease (sAD) are likely multiple and include impairment of mitochondrial bioenergetics. We analyzed RNA expression levels of multiple mitochondrial oxidative phosphorylation (OXPHOS) and biogenesis (mitobiogenesis) genes in unfixed hippocampal (WH) frozen sections (10 sAD; 9 CTL) and laser-captured hippocampal pyramidal neurons (PyNs, ~1000 neurons from each case) from 8 sAD and 7 CTL cases. Nuclear-encoded OXPHOS genes in WH were significantly increased in sAD, whereas in isolated sAD PyNs, these same genes were significantly decreased. Mitochondrial DNA-encoded genes were increased in sAD PyNs but showed a non-significant downward trend in sAD WH. Relationships among WH and PyN gene expression levels in sAD distributed in a different population compared to CTL. Principal component analysis (PCA) revealed clustering of CTL but widespread heterogeneity of sAD samples. In sAD, mitochondrial bioenergetics at the gene expression level are depressed in vulnerable PyNs. PCA revealed that CTL samples clustered together, whereas sAD samples varied widely. From the perspective of OXPHOS bioenergetics, sAD is a heterogeneous syndrome and not likely due to a single abnormality. Increased stimulation of nuclear-encoded OXPHOS gene expression in PyNs is a rational therapeutic approach for most but not all cases of sAD."
] |
Molecular mechanisms of waning immunity in the case of mRNA COVID-19 vaccines
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mRNA COVID-19 vaccines have hegemonized the world market, and their administration to the population promises to stop the pandemic. However, the waning of the humoral immune response, which does not seem to last so many months after the completion of the vaccination program, has led us to study the molecular immunological mechanisms of waning immunity in the case of mRNA COVID-19 vaccines. We consulted the published scientific literature and from the few articles we found, we were convinced that there is an immunological memory problem after vaccination. Although mRNA vaccines have been demonstrated to induce antigen-specific memory B cells (MBCs) in the human population, there is no evidence that these vaccines induce the production of long-lived plasma cells (LLPCs), in a SARS-CoV-2 virus naïve population. This obstacle, in our point of view, is caused by the presence, in almost all subjects, of a cellular T and B cross-reactive memory produced during past exposures to the common cold coronaviruses. Due to this interference, it is difficult for a vaccination with the Spike protein alone, without adjuvants capable of prolonging the late phase of the generation of the immunological memory, to be able to determine the production of protective LLPCs. This would explain the possibility of previously and completely vaccinated subjects to become infected, already 4-6 months after the completion of the vaccination cycle.
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[
"Common laboratory mice such as BALB/c and C57BL/6 mice are not permissive to SARS-CoV2 infection. Sensitization of laboratory mice with Adenovirus expressing human ACE2 (Ad5-hACE2) provides a rapid model for testing viral intervention in vivo. Despite the lack of lethal outcome, Ad5-hACE2-sensitized mice show 20% weight loss on average upon viral challenge with infectious virus being detected at the site of sensitization. This protocol describes the sensitization and subsequent infection of common laboratory mice for use in testing anti-viral interventions. For complete details on the use and execution of this protocol, please refer to Sun et al. (2020).",
"Two-dose messenger RNA vaccines against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are highly effective in preventing symptomatic COVID-19 infection. However, the durability of protection is not known, nor is the effectiveness against emerging viral variants. Additionally, vaccine responses may differ based on prior SARS-CoV-2 exposure history. To investigate protection against SARS-CoV-2 variants we measured binding and neutralizing antibody responses following both vaccine doses. We document significant declines in antibody levels three months post-vaccination, and reduced neutralization of emerging variants, highlighting the need to identify correlates of clinical protection to inform the timing of and indications for booster vaccination.",
"Current COVID-19 vaccines and many clinical diagnostics are based on the structure and function of the SARS-CoV-2 spike ectodomain. Using hydrogen-deuterium exchange monitored by mass spectrometry, we have uncovered that, in addition to the prefusion structure determined by cryo-electron microscopy, this protein adopts an alternative conformation that interconverts slowly with the canonical prefusion structure. This new conformation-an open trimer-contains easily accessible receptor-binding domains. It exposes the conserved trimer interface buried in the prefusion conformation, thus exposing potential epitopes for pan-coronavirus antibody and ligand recognition. The population of this state and kinetics of interconversion are modulated by temperature, receptor binding, antibody binding, and sequence variants observed in the natural population. Knowledge of the structure and populations of this conformation will help improve existing diagnostics, therapeutics, and vaccines.",
"The emergence of SARS-CoV-2 variants with evidence of antibody escape highlight the importance of addressing whether the total CD4+ and CD8+ T cell recognition is also affected. Here, we compare SARS-CoV-2-specific CD4+ and CD8+ T cells against the B.1.1.7, B.1.351, P.1, and CAL.20C lineages in COVID-19 convalescents and in recipients of the Moderna (mRNA-1273) or Pfizer/BioNTech (BNT162b2) COVID-19 vaccines. The total reactivity against SARS-CoV-2 variants is similar in terms of magnitude and frequency of response, with decreases in the 10%-22% range observed in some assay/VOC combinations. A total of 7% and 3% of previously identified CD4+ and CD8+ T cell epitopes, respectively, are affected by mutations in the various VOCs. Thus, the SARS-CoV-2 variants analyzed here do not significantly disrupt the total SARS-CoV-2 T cell reactivity; however, the decreases observed highlight the importance for active monitoring of T cell reactivity in the context of SARS-CoV-2 evolution.",
"In March 2020, the World Health Organization (WHO) declared coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)1, a pandemic. With rapidly accumulating numbers of cases and deaths reported globally2, a vaccine is urgently needed. Here we report the available safety, tolerability and immunogenicity data from an ongoing placebo-controlled, observer-blinded dose-escalation study (ClinicalTrials.gov identifier NCT04368728) among 45 healthy adults (18-55 years of age), who were randomized to receive 2 doses-separated by 21 days-of 10 μg, 30 μg or 100 μg of BNT162b1. BNT162b1 is a lipid-nanoparticle-formulated, nucleoside-modified mRNA vaccine that encodes the trimerized receptor-binding domain (RBD) of the spike glycoprotein of SARS-CoV-2. Local reactions and systemic events were dose-dependent, generally mild to moderate, and transient. A second vaccination with 100 μg was not administered because of the increased reactogenicity and a lack of meaningfully increased immunogenicity after a single dose compared with the 30-μg dose. RBD-binding IgG concentrations and SARS-CoV-2 neutralizing titres in sera increased with dose level and after a second dose. Geometric mean neutralizing titres reached 1.9-4.6-fold that of a panel of COVID-19 convalescent human sera, which were obtained at least 14 days after a positive SARS-CoV-2 PCR. These results support further evaluation of this mRNA vaccine candidate."
] |
Effects of fruits and vegetables on the host whole blood cell transcriptome and the composition and function of the intestinal microbiome in pigs.
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A study was conducted to determine the effects of a diet supplemented with fruits and vegetables (FV) on the host whole blood cell (WBC) transcriptome and the composition and function of the intestinal microbiome. Nine six-week-old pigs were fed a pig grower diet alone or supplemented with lyophilized FV equivalent to half the daily recommended amount prescribed for humans by the Dietary Guideline for Americans (DGA) for two weeks. Host transcriptome changes in the WBC were evaluated by RNA sequencing. Isolated DNA from the fecal microbiome was used for 16S rDNA taxonomic analysis and prediction of metabolomic function. Feeding an FV-supplemented diet to pigs induced differential expression of several genes associated with an increase in B-cell development and differentiation and the regulation of cellular movement, inflammatory response, and cell-to-cell signaling. Linear discriminant analysis effect size (LEfSe) in fecal microbiome samples showed differential increases in genera from
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"Genes in metabolic and nutrient signaling pathways play important roles in lifespan in model organisms and human longevity. The aim of this study was to examine the relation of a quantitative measure of healthy diet to gene expression in a community-based cohort. We used the 2015 Dietary Guidelines for Americans Adherence Index (DGAI) score to quantify key dietary recommendations of an overall healthy diet. Our current analyses included 2220 Offspring participants (mean age 66 ± 9 y, 55.4% women) and 2941 Third-Generation participants (mean age 46 ± 9 y, 54.5% women) from the Framingham Heart Study. Gene expression was profiled in blood through the use of the Affymetrix Human Exon 1.0 ST Array. We conducted a transcriptome-wide association study of DGAI adjusting for age, sex, smoking, cell counts, and technical covariates. We also constructed a combined gene score from genes significantly associated with DGAI. The DGAI was significantly associated with the expression of 19 genes (false discovery rate <0.05). The most significant gene, ARRDC3, is a member of the arrestin family of proteins, and evidence in animal models and human data suggests that this gene is a regulator of obesity and energy expenditure. The DGAI gene score was associated with body mass index (P = 1.4 × 10-50), fasting glucose concentration (P = 2.5 × 10-11), type 2 diabetes (P = 1.1 × 10-5), and metabolic syndrome (P = 1.8 × 10-32). Healthier diet was associated with genes involved in metabolic function. Further work is needed to replicate our findings and investigate the relation of a healthy diet to altered gene regulation.",
"A high-capacity system was developed to monitor the expression of many genes in parallel. Microarrays prepared by high-speed robotic printing of complementary DNAs on glass were used for quantitative expression measurements of the corresponding genes. Because of the small format and high density of the arrays, hybridization volumes of 2 microliters could be used that enabled detection of rare transcripts in probe mixtures derived from 2 micrograms of total cellular messenger RNA. Differential expression measurements of 45 Arabidopsis genes were made by means of simultaneous, two-color fluorescence hybridization.",
"Comprehensive identification of all functional elements encoded in the human genome is a fundamental need in biomedical research. Here, we present a comparative analysis of the human, mouse, rat and dog genomes to create a systematic catalogue of common regulatory motifs in promoters and 3' untranslated regions (3' UTRs). The promoter analysis yields 174 candidate motifs, including most previously known transcription-factor binding sites and 105 new motifs. The 3'-UTR analysis yields 106 motifs likely to be involved in post-transcriptional regulation. Nearly one-half are associated with microRNAs (miRNAs), leading to the discovery of many new miRNA genes and their likely target genes. Our results suggest that previous estimates of the number of human miRNA genes were low, and that miRNAs regulate at least 20% of human genes. The overall results provide a systematic view of gene regulation in the human, which will be refined as additional mammalian genomes become available.",
"Although genomewide RNA expression analysis has become a routine tool in biomedical research, extracting biological insight from such information remains a major challenge. Here, we describe a powerful analytical method called Gene Set Enrichment Analysis (GSEA) for interpreting gene expression data. The method derives its power by focusing on gene sets, that is, groups of genes that share common biological function, chromosomal location, or regulation. We demonstrate how GSEA yields insights into several cancer-related data sets, including leukemia and lung cancer. Notably, where single-gene analysis finds little similarity between two independent studies of patient survival in lung cancer, GSEA reveals many biological pathways in common. The GSEA method is embodied in a freely available software package, together with an initial database of 1,325 biologically defined gene sets.",
"The development of B cells from haematopoietic stem cells proceeds along a highly ordered, yet flexible, pathway. At multiple steps along this pathway, cells are instructed by transcription factors on how to further differentiate, and several check-points have been identified. These check-points are initial commitment to lymphocytic progenitors, specification of pre-B cells, entry to the peripheral B-cell pool, maturation of B cells and differentiation into plasma cells. At each of these regulatory nodes, there are transcriptional networks that control the outcome, and much progress has recently been made in dissecting these networks. This article reviews our current understanding of this exciting field.",
"The gut microbiome is shaped by diet and influences host metabolism; however, these links are complex and can be unique to each individual. We performed deep metagenomic sequencing of 1,203 gut microbiomes from 1,098 individuals enrolled in the Personalised Responses to Dietary Composition Trial (PREDICT 1) study, whose detailed long-term diet information, as well as hundreds of fasting and same-meal postprandial cardiometabolic blood marker measurements were available. We found many significant associations between microbes and specific nutrients, foods, food groups and general dietary indices, which were driven especially by the presence and diversity of healthy and plant-based foods. Microbial biomarkers of obesity were reproducible across external publicly available cohorts and in agreement with circulating blood metabolites that are indicators of cardiovascular disease risk. While some microbes, such as Prevotella copri and Blastocystis spp., were indicators of favorable postprandial glucose metabolism, overall microbiome composition was predictive for a large panel of cardiometabolic blood markers including fasting and postprandial glycemic, lipemic and inflammatory indices. The panel of intestinal species associated with healthy dietary habits overlapped with those associated with favorable cardiometabolic and postprandial markers, indicating that our large-scale resource can potentially stratify the gut microbiome into generalizable health levels in individuals without clinically manifest disease.",
"Acute lymphoblastic leukemias carrying a chromosomal translocation involving the mixed-lineage leukemia gene (MLL, ALL1, HRX) have a particularly poor prognosis. Here we show that they have a characteristic, highly distinct gene expression profile that is consistent with an early hematopoietic progenitor expressing select multilineage markers and individual HOX genes. Clustering algorithms reveal that lymphoblastic leukemias with MLL translocations can clearly be separated from conventional acute lymphoblastic and acute myelogenous leukemias. We propose that they constitute a distinct disease, denoted here as MLL, and show that the differences in gene expression are robust enough to classify leukemias correctly as MLL, acute lymphoblastic leukemia or acute myelogenous leukemia. Establishing that MLL is a unique entity is critical, as it mandates the examination of selectively expressed genes for urgently needed molecular targets."
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