bluuebunny/crossref_metadata_2025 huggingface dataset:
Breastmilk contains primitive stem cells from the lactating breast with multi‐lineage differentiation potential
Authors: Hassiotou Foteini , Filgueira Luis , Trengove Naomi , Lai Ching Tat , Hartmann Peter
Published: June 2021

The remodelling of the mammary gland during pregnancy/lactation is fuelled by mammary stem cells. Because most of our knowledge on these cells comes from the resting gland, recent efforts have focused on the lactating gland in which these cells are activated. We non‐invasively accessed stem cells from the human lactating gland via freshly expressed breastmilk. We performed ex vivo analysis of the cell subpopulations present in breastmilk using numerous phenotypic and functional stem cell and differentiation markers for various lineages. A population displaying the phenotypic profile of mammary stem cells previously characterised in the resting gland (CD49f+/CD29+) was identified, able to self‐renew and differentiate in vitro into the two main mammary epithelial subtypes. In addition, a population expressing the pluripotency markers Oct‐4, Sox‐2, Nanog, SSEA‐4, Tra‐1–60 and Tra‐1–81 was identified. Culture of these cells in stem cell and differentiation media revealed the potential both for self‐renewal and differentiation into cells originating from the three germ layers. The identified subpopulations were also localised in paraffin‐embedded human lactating tissues. Based on these results, a new cellular hierarchy is proposed for the human mammary gland, at the top of which sits a cell population with potential pluripotency, which can be easily accessed via breastmilk.

Grant Funding Source : Women and Infants Research Foundation, Medela AG
Identification of neural crest‐like synovial stem cells
Authors: Huang Fang , Tang Zhenyu , Li Song
Published: June 2021

Recent studies suggest that bioactive scaffolds can be used to regenerate cartilage by local cell homing. However, the specific cell types participating in regeneration are not fully understood. In this study, we identified a novel type of neural crest like synovial stem cell (NCL‐SSC) residing within the synovial membrane, which has several important characteristics distinct from previously identified stem cells. Protein marker screening revealed that NCL‐SSCs homogeneously expressed neural crest stem cell markers such as Sox1, Sox10, Snail and Vimentin. Furthermore, differentiation assays NCL‐SSCs can be induced into mesenchymal lineages (osteoblasts, adipocytes, and chondrocytes), peripheral neurons and Schwann cells. In addition, we found that the combination of chick embryo extract and basic fibroblast growth factor can maintain the multipotency and maker expression of NCL‐SSCs. Cloning assay showed that the average plating efficiency of NCL‐SSCs is about 10% and the cloned colonies retained the marker expression and multipotency. In summary, our study indentified a new type of local cells in synovial knee joint, which could be an important cell source for in situ tissue engineering.
Initial Characterization of the Cellular Basis for Bladder Regeneration in a Rodent Model
Authors: Peyton Charles , Burmeister David , Andersson Karl-Erik , Christ George
Published: June 2021

Purpose

A better understanding of intrinsic mammalian organ regeneration would benefit the field regenerative medicine. Our prior work shows functional bladder recovery in rats after removal of ~70% of the bladder (subtotal cystectomy/STC). Our objective is to begin identifying the spatiotemporal characteristics of cellular response to STC.

Methods

Trigone‐sparing STC was performed on 12‐week‐old female Fischer F344 rats. At 1, 3, 5, and 7 days post‐STC, bladders were harvested 2 hours after IP 50mg/kg 5‐Bromo‐2’‐deoxyuridine (BrdU). Fluorescent BrdU labeling was used to quantify and localize S‐phase cells within the bladder wall. Immunohistochemistry for Shh, Gli‐1 and BMP‐4 was also performed. Normal rat bladders served as controls.

Results

Sparse BrdU labeling was observed in native bladders (Fig. 1). The total number of BrdU‐positive cells/400x HPF was significantly higher post‐STC at all time points. A time‐depended location change in the percentage of BrdU‐positive cells was seen. Increased positive staining for Shh, Gli‐1 and BMP‐4 was also noted post‐STC.

Conclusions

STC induced a robust proliferative response with quantifiable spatiotemporal characteristics and up‐regulation of key targets in the Shh developmental pathway. Future studies will identify the cell source(s) and details of the molecular pathways that modulate functional bladder regeneration.
Glycans of Trypanosoma cruzi virulence factors are effective targets for vaccine development
Authors: Almeida Igor C. , Veas Esteban C. , Ganiko Luciane , Nohara Lilian L. , Garza Kristine , Silva Luiz S. , Travassos Luiz R. , Nakayasu Ernesto S. , Marques Alexandre F.
Published: June 2021

Trypanosoma cruzi causes Chagas disease (CD), which affects millions of people. There is no human vaccine despite numerous experimental studies based on T‐cell epitopes. In contrast, B‐cell epitopes remain poorly explored, despite the fact that very high levels of protective anti‐α‐Gal Abs elicited against highly abundant α‐Gal epitopes on the parasite surface are ubiquitous in CD patients. We hypothesize that terminal α‐Gal‐containing epitopes are potential vaccine targets. Here, we evaluate the Galα1,3LacNAc‐BSA in the α1,3‐galactosyltransferase‐KO (αGalT‐KO) mouse model, which closely mimics the human humoral immune response against T. cruzi. Following immunization, αGalT‐KO mice showed high titers of protective anti‐α‐Gal Abs, very low parasitemia, and high levels of proinflammatory cytokines. All immunized animals survived up to one year (experiment endpoint) following parasite challenges. Histological and qRT‐PCR analyses of the heart of protected animals indicated extremely low parasite levels. Depletion of CD4+ and/or CD8+ T cells before or after immunization resulted in much lower survival rates, corroborating the key role of T cells in controlling disease progression. In sum, our data show that protective anti‐α‐Gal Abs and proinflammatory immune responses can effectively control experimental CD.
Characterization of a Novel Overexpression System in Haloferax volcanii
Authors: Huff Brian , Hilty Jenna , Hemmingsen Jens , Ginn-Pease Margaret E.
Published: June 2021

A novel inducible protein expression system has been developed in the halophilic archaea, Haloferax volcanii (Hvo.) The expression system is based on the histidine utilization (Hut) operon and can be induced by histidine or urocanate, a histidine metabolite. This study characterizes the 88 base promoter region between the gene that encodes a newly discovered Hut regulatory protein (Hut R) and the first gene in the Hut operon, Hut U. Plasmids have been constructed that contain the gene for Hut R, the promoter region, the first three codons of Hut U and the gene for L7ae, an RNA binding protein, used as an expression reporter (see figure). The plasmid containing the wildtype promoter yields a high level of inducible gene expression when transformed into Hvo. Two specific mutations have been constructed in the promoter near the Hut U start sequence, deleting either 5 base (Del 1) or 20 bases (Del 2) immediately upstream of the Hut U start codon. These mutants yield decreased gene expression as measured by Northern and Western analysis. The new system appears to be generally useful for overexpression of halophilic proteins and has the potential for protein overexpression from a variety of halophilic organisms. This work was supported by Capital University.
Intersectin 1 is required for Neuroblastoma tumorigenesis
Authors: Russo Angela , O'Bryan John
Published: June 2021

Neuroblastoma (NB) is a malignant tumor deriving from the neural crest that is responsible for 13% of cancer‐related deaths in children. Phosphoinositide 3‐kinases (PI3Ks) play a central role in NB tumorigenesis. PI3Ks activate AKT, which regulates survival and stabilizes the MYCN oncogene. Both MYCN and AKT are markers for poor prognosis in NBs. Inhibiting PI3Ks with pan‐inhibitors blocks malignant progression in vivo. These inhibitors have high toxicity that limits their use in NB treatment. Though there are three classes of PI3Ks, only Class I has been implicated in NB. Class 2 beta PI3K (PI3KC2β) is overexpressed in cancer and involved in adherent and anchorage‐independent growth of cancer cells. We found that the scaffold protein intersectin 1 (ITSN1) interacts with PI3KC2β and regulates its activation and Akt activation in mouse NB lines. Thus, we examined the role of the ITSN1‐PI3KC2β pathway in human NB tumorigenesis. We found that stable silencing of ITSN1 in human NB cells attenuates their growth both in soft agar assays in vitro and in xenograft tumor assays in vivo. Furthermore, PI3KC2β overexpression restores anchorage‐independent growth of ITSN1‐depleted NB cells suggesting that PI3KC2β mediates ITSN1's tumorigenic phenotype. This study reveals a new pathway, i.e., ITSN1‐ PI3KC2β is involved in regulating the pathogenesis of NBs.
Chondrocyte Phenotype of Mice Expressing Reduced Levels of Perlecan/HSPG2, an Essential Component of Cartilage
Authors: Lowe Dylan Andrew , Srinivasan Padma Pradeepa , Kirn-Safran Catherine
Published: June 2021

Perlecan/Hspg2 (PLN) is a heparan sulfate proteoglycan functioning as a modulator of growth factor bioavailability and scaffold in the extracellular matrix (ECM). PLN gene loss‐of‐function mutations result in severe skeletal defects and embryonic death. In this study, we use a viable PLN mutant mouse model in which PLN expression is reduced. Adult PLN mutant mice display a short stature, early osteoarthritis, and altered bone properties. Immunostaining of developing growth plates revealed an absence of columnar organization and severe reduction of PLN secretion in the cartilage matrix of PLN mutant versus control bones. Furthermore, the majority of PLN in the hypomorphs co‐localized with a strong signal for BiP, an endoplasmic reticulum (ER) chaperone involved in the unfolded protein response (UPR). Our data suggest that mutant PLN retained within the ER elevates ER stress, which contributes to the disease phenotype. Furthermore, chondrocyte proliferation is altered in the developing growth plates of PLN mutant mice. Ongoing work further investigates how elevated ER stress and altered fibroblast growth factor signaling in the developing growth plate contribute to the abnormal phenotype in PLN mutant mice. Supported by NIH P20 RR016458 COBRE and HHMI and UD Science and Engineering Scholarship.
Brain‐derived neurotrophic factor receptor TrkB exists as a preformed dimer in living cells
Authors: Maruyama Ichiro , Shen Jianying
Published: June 2021

Background

Neurotrophins and their receptors play crucial roles in the development and functioning of nervous systems. It is widely believed that neurotrophin‐induced dimerization of the receptors initiates the transmembrane signalling. However, it is still controversial whether the receptor molecules have a monomeric or dimeric structure on the cell surface before its ligand binding.

Findings

Using chemical cross‐linking, bimolecular fluorescence complementation assay and luciferase fragment complementation assay, in this study, we show the brain‐derived neurotrophic factor (BDNF) receptor TrkB exists as a homo‐dimer before ligand binding. We also report that the receptor fs intracellular domain is critical for the formation of the dimeric structure, since mutant receptors lacking its intracellular domain failed to have the homodimeric structure, and that the dimer is formed in the endoplasmic reticulum before the newly synthesized receptors reach the cell surface.

Conclusions

Most, if not all, of the TrkB receptor has a preformed, yet inactive, homodimeric structure before BDNF binding. The intracellular domain of TrkB plays a crucial role in the spontaneous dimerization of the newly synthesized receptors, which occurs in ER. These findings provide new insight into an understanding of a molecular mechanism underlying transmembrane signaling mediated by neurotrophin receptors.
Bacterial GTPases may be regulators of cell wall assembly
Authors: Winters Jenifer Nichole , Rizzo Nicholas , March Paul E
Published: June 2021

Scrutiny of bacterial genomes has led to the identification of a core set of genes that are essential for cell growth: the minimal genome. Many of the genes represented in the minimal genome have well defined and unambiguous functions. The function of some of the highly conserved bacterial GTPases, in particular the Era‐subfamily (Der/EngA and Era), is not completely understood. These GTPases are the topic of this investigation. Recently we have reported that over‐expression of Der caused a defect in cell wall structure and that Der could bind specifically to membrane protein. The binding reaction was reconstituted in vitro and we developed an electrophoresis based co‐migration binding assay to detect the Der‐membrane interaction. We have used that assay to assess that OmpC, and not OmpF is necessary for Der to form a high molecular weight complex when combined with membrane proteins. Based on these results, and crystal structures of OmpC and Der, we propose that domain 3 of Der binds to loop 4 on OmpC, and that this interaction is necessary for the proper regulation of cell wall assembly in Escherichia coli.
Distribution and localization of Rho GTPases in rat brain
Authors: Martínez-Alcantar Lorena , Meléndez-Herrera Esperanza , Clemente-Guerrero Mónica , Saavedra-Molina Alfredo , Manzo-Avalos Salvador
Published: June 2021

Rho GTPases proteins are essential for cytoskeletal reorganization. Several studies have implicated members of the Rho GTPase family in the neuronal polarization and the formation of axons and dendrites. Only few studies have investigated the cellular distribution of RhoA, RhoB and Rac1. Therefore the aim of this study was to investigate the cellular distribution of these proteins using immunohistochemical technique. Rac1, RhoA and RhoB proteins show a differential distribution and the pattern is different in rat brain. RhoA was localized in the soma and dendrites of neurons in most brain regions, hypothalamus. In brainstem nucleus RhoA showed increased expression. RhoB showed a more restricted distribution, and was located in the brain stem and in the plasma membrane in the pyramidal cell layer of the cortex. Rac1 was localized in neuronal cell bodies and distributed in all brain regions. The differential localization of the Rho GTPases in rat brain may reflect different roles of these proteins within these structures.
The cpSRP54‐cpFtsY Interaction in the Chloroplast SRP Pathway
Authors: Lu Zeyu Mike , Chandrasekar Sowmya , Shan Shu-ou
Published: June 2021

The universally conserved signal recognition particle (SRP) and SRP receptor (SR) mediate the cotranslational targeting of proteins to cellular membranes. In contrast, a recently discovered unique chloroplast SRP in green plants is primarily dedicated to the post‐translational targeting of light harvesting chlorophyll a/b binding (LHC) proteins to the thylakoid membranes. The pathway involves cpSRP54 and its receptor cpFtsY, two GTPases that are similar to the cytosolic SRP54 and SR GTPases. Intriguingly, the otherwise universally conserved SRP RNA which accelerates complex formation between the two GTPases has not been found in the chloroplast pathway. Instead, a new 43‐kDa protein, cpSRP43, forms the chloroplast SRP together with cpSRP54. Kinetic analyses have shown that cpSRP43 does not significantly affect the rate of the cpSRP54·cpFtsY complex formation or the GTP hydrolysis rate of the complex. Instead, cpSRP43 facilitates the substrate recognition through interactions with the cargo proteins, LHCPs. This project seeks to understand how cpSRP54 and cpFtsY are able to associate more efficiently than their bacterial homologues in the absence of the SRP RNA by studying their complex structure using crystallography and lysine footprinting mass spectrometry. To date, 2‐dimensional crystal plates have been obtained, and progress continues toward the determination of a cpSRP54·cpFtsY structure.
“Role of GPER in Estrogen‐Mediated Inhibition of Cardiomyocyte Hypertrophy.”
Authors: Huffman Sarah Elizabeth , Kennedy Timothy Ryan , Baute Audrey Joy , Waikel Rebekah L.
Published: June 2021

Estrogen is known to attenuate cardiac hypertrophy and heart failure in females. To understand how estrogen mediates its effects on the heart, it is important to understand how the cell receives the estrogen signal. Estrogen activates two classes of receptors: Nuclear Hormone Receptors (Estrogen Receptor Alpha (ERa) and Estrogen Receptor Beta (ERβ) and a membrane bound G‐Protein Coupled Receptor for Estrogen (GPER or GPR30). It has not been established whether GPER plays a role in estrogen's ability to prevent cardiac hypertrophy or what that role may be. To determine if GPER plays a role in the antihypertrophic effects of estrogen, we have performed a series of in‐vitro experiments using primary rat cardiomyocyte cultures that have been serum starved for 24 hours and then exposed to the hypertrophic agent, Phenylephrine, in the presence or absence of a GPER specific inhibitor (G15) plus estrogen or the presence or absence of a GPER specific activator (G1) and compared to estrogen alone. The degree of hypertrophy was determined between the groups. Expression of known estrogen target genes were also assessed in cells exposed to the GPER specific activator by relative RT‐PCR. Taken together our results suggest that GPER does play a role in estrogen's antihypertrophic effects.
Study Of Molecular Events Associated With Single Wall Carbon Nanotubes Mediated Cellular Cytotoxicity
Authors: Goornavar Virupaxi , Ravichandran Prabakaran , Biradar Santoshkumar , Ramesh Govindarajan T
Published: June 2021

Carbon nanotubes, due to their unique and outstanding properties, have generated extraordinary interest and expectations for industrial and biomedical applications. The increasing mass production of SWCNT requires their biocompatibility studies to prevent the possible health hazards among workers involved in the research, manufacturing of these materials and in the general public. Lung epithelial cell lines were used to investigate the mechanism underlying for apoptosis induced by single wall carbon nanotubes. Our results showed a significant amount of incorporation of dUTPs in the nucleus confirming the induction of apoptosis, which was also validated by DNA fragmentation assay. The activity of caspases‐3 and caspase‐8 increased in a dose and time dependent manner in parallel to the morphological apoptosis. The levels of cytochrome c, cellular ATP, Ap‐1 factor, p53, p21, bax, and bcl‐2 also changed reflecting the activation of the apoptosis pathway. Together, in this study we show that apoptosis in lung epithelial cells exposed to single wall carbon nanotubes occurred through the activation of caspase cascade, which in turn triggers nuclear transcription factor AP‐1 and the release of cytochrome c from the mitochondria into the cytosol where it binds the apoptotic protease activating factor resulting in cell death.
Alternate day fasting (ADF) with a high fat background diet produces similar weight loss and cardio‐protection when compared to ADF with a low fat background diet
Authors: Klempel Monica Catherine , Kroeger Cynthia M , Varady Krista A
Published: June 2021

Background

Alternate day fasting (ADF) with a low‐fat (LF) background diet facilitates weight loss and is cardio‐protective in obese individuals. Whether these beneficial effects can be reproduced by a high‐fat (HF) ADF diet is not known.

Objective

This study examined the effects of an ADF‐HF versus ADF‐LF diet on body weight, body composition and cardiovascular disease (CVD) risk in obese humans.

Methods

Thirty‐three obese subjects were randomized to an ADF‐HF or ADF‐LF diet for 10 weeks. All food was provided during the study.

Results

Body weight was reduced (P < 0.0001) by 4.9%, and 4.2% in the ADF‐HF and ADFLF group, respectively. Waist circumference decreased (P < 0.0001) by 7.6%, and 7.5% in the ADF‐HF and ADF‐LF groups, respectively. LDL cholesterol was reduced (P < 0.0001) by 19.1%, and 24.8%, whereas triglycerides were lowered (P = .0001) by 13.3% and 14.3%, in the ADF‐HF and ADF‐LF group, respectively. There were no changes in blood pressure or heart rate.

Conclusion

These findings suggest that an ADF‐HF diet is equally as effective as an ADF‐LF diet in helping obese subjects lose weight and improve CVD risk factors.
Knowledge, Use and Perceptions of Probiotics and Prebiotics in Medical Center Healthcare Providers
Authors: Rasmussen Heather , Oliver Laura , Gregoire Mary , Chen Yimin
Published: June 2021

Dietary modulation of the gut microbiota through probiotics and prebiotics may be beneficial for health. Despite their potential importance, no studies have determined knowledge, use and perceptions of probiotics and prebiotics in healthcare providers. In the present study, an online questionnaire was completed to assess these parameters in healthcare providers at a large, urban medical center (n=256). Most of the participants were White (77%), female (74%), and nurses (49%) or physicians (39%). Participants were familiar with probiotics (97%), but not familiar with prebiotics (28%). Most of the participants had not read any recent peer‐reviewed publications on probiotics (77%) or prebiotics (79%). If substantiated by peer‐reviewed literature, healthcare providers would be “quite a bit” to “very much” willing to recommend probiotics (53%) and prebiotics (78%). Despite believing that probiotics and prebiotics are beneficial to overall health (63% and 55%, respectively), the majority of healthcare providers had never recommended probiotics (57%) or prebiotics (75%) to patients. A total of 62% of healthcare providers personally consume probiotics and 31% consume prebiotics, with a significant difference in probiotic and prebiotic recommendations for patient use based on personal consumption (P = 0.001; P = 0.001, respectively). In conclusion, healthcare providers are familiar with probiotics, but not prebiotics, and believe they are beneficial to overall health; however, they do not follow the current literature or frequently recommend them to patients.
Interleukin 10 (IL‐10) inhibits GABAergic transmission in rat hippocampal neurons by a mechanism involving phosphatidylinositol 3‐kinase
Authors: Suryanarayanan Asha , McCrary Ellen J , Morrow A Leslie , Spigelman Igor
Published: June 2021

We previously demonstrated that a single intoxicating dose of ethanol significantly increases hippocampal interleukin 10 (IL‐10). Additionally, in whole‐cell patch clamp recordings from rat brain dentate granule cells, we observed that acute application of IL‐10 (10–50 ng/ml) decreased GABAergic miniature inhibitory postsynaptic current (mIPSC) peak amplitude, area and frequency. A significant decrease in tonic current was also observed. Since IL‐10 causes rapid activation of phosphatidylinositol (PI) 3‐kinase, we hypothesized that PI3‐kinase is involved in mediating IL‐10 induced inhibition of GABAergic transmission. To test this hypothesis, we carried out recordings including either 20 μM LY294002 (a PI3‐kinase inhibitor) or 20 μM LY303511 (a negative control compound which does not inhibit PI3‐kinase) in the recording pipette. We found that in presence of LY294002, IL‐10 no longer caused a significant decrease in mIPSC peak amplitude, area and tonic current. Also, in the presence of LY294002, a significant decrease in mIPSC frequency was observed only with 50 ng/ml IL‐10. Results with the negative control compound LY303511 showed that the inhibitory effects of IL‐10 mIPSC area, peak amplitude, frequency and tonic current were maintained. These data demonstrate, for the first time, the involvement of PI3‐kinase in IL‐10 mediated modulation of GABAergic transmission.
Novel positive allosteric modulators of the human α7 nicotinic acetylcholine receptor
Authors: Miller Marc A. , Gu Ruo-Xu , Feuerbach Dominik , Ye Yong , Wei* Dong-Qing , Arias Hugo R.
Published: June 2021

The pharmacological activity of a series of novel amide derivatives was characterized on several nicotinic acetylcholine receptors (AChRs). Ca2+ influx results indicate that these compounds are not agonists of the human (h) α4β2, α3β4, α7, and α1β1γδ AChRs; but are specific positive allosteric modulators (PAMs) of hα7 AChRs. Radioligand binding results indicate that PAMs do not inhibit binding of [3H]methyllycaconitine but enhance binding of [3H]epibatidine to hα7 AChRs, indicating that these compounds do not directly, but allosterically, interact with the hα7 agonist sites. Hydrogen bond interactions between the amide group of the PAMs and the hα7 AChR binding site are found to be critical for their activity. The dual PAM and antagonistic activities elicited by these compounds might be therapeutically important in treating diseases such as depression and drug addiction.

We thank the National Institute on Drug Abuse (NIDA) for the gift of phencyclidine.
Modulation of heart glycogen by isoproterenol‐induced transient tachycardia in newly born rats
Authors: Fewell James Edwin , Zhang Chunfen
Published: June 2021

Newly born animals of a number of species have an increased cardiac glycogen content compared to older animals which is purported to support cardiac function and promote survival during hypoxia. As a first step in investigating the role of cardiac glycogen in influencing protective responses of newly born rat pups to hypoxia, our experiments were carried out to determine if cardiac glycogen would be altered following isoproterenol(ISO)‐induced transient tachycardia. In preliminary dose–response experiments testing doses of 0 to 10 mg/kg of the nonselective beta adrenergic receptor agonist ISO, we determined 1.25 mg/kg to be the smallest dose that elicited a maximal heart rate response (i.e., the EC100) in 1 day‐old pups and in which the heart rate returned back to control levels by 120 minutes. One day old rat pups received vehicle (VEH) or ISO (1.25 mg/kg) ip and heart rate was recorded for 120 min before decapitation and removal of the heart for determination of glycogen content. Whole heart glycogen content in untouched pups was 4.1±0.5 mg/g of tissue (n=4). Whole heart glycogen content decreased significantly (p<0.05) following ISO‐induced transient tachycardia compared to VEH: 1.5±0.5 mg/g (n=8) versus 3.5±0.3 mg/g (n=5), respectively. Thus, our data provide evidence that whole heart glycogen is significantly decreased in newly born rat pups following ISOinduced transient tachycardia.
Central arterial stiffness is positively associated with parent‐reported inactivity and systolic blood pressure in children
Authors: Price Paula Holland , Lee Joshua F. , Bartholomew John B. , Brothers R. Matthew
Published: June 2021

Carotid‐femoral arterial pulse wave velocity (central PWV), “an index of central arterial stiffness”, has become a recognized physiologic measure of vascular pathology. Central PWV is gaining acceptance as an indicator of overall atherosclerotic disease and shows potential as a screening tool for atherosclerotic disease in hypertensive adults. There is evidence that atherosclerotic disease begins in childhood, however, the mechanism is poorly understood. An association between fitness and central PWV has been reported in children. This pilot study explores the association between inactivity and central PWV in children. Central arterial stiffness of thirteen subjects, ages 8–13 years, was measured by carotid‐femoral PWV. Parents of the subjects reported the child's inactivity behavior on a typical day. Central PWV was found to correlate with inactivity (correlation coefficient=.656, p<0.05) and was also found to correlate with systolic blood pressure (correlation coefficient=0.667, p<0.02). These data indicate that inactivity may be a strong predictor of central arterial stiffness in children.

Funding Source: University of Texas at Austin
The role of CaMKII activation in the regulation of carnitine palmitoyl transferase and lipid oxidation in rats muscle
Authors: Mukwevho Emmanuel
Published: June 2021

The mismatch between high fatty acids supply and mitochondrial fatty acid oxidation in skeletal muscle has been associated with insulin resistance, the hallmark of type II diabetes (Holloway et al., 2009). In this study we wanted to investigate the role of CaMKII activation in the reduction of lipids and whether carnitine palmitoyl transferase (CPT)‐1, the rate limiting step oxidation was involved. Improved mitochondrial fatty acid oxidation is seen as a crucial step in the reduction of lipids that interferes with insulin signaling pathway.

In this study, male Wistar rats were used. The exercise regime was designed for 5 days, where each session per day was 5 × 12 min with 3 minutes rests, after which gastrocnemius muscle was excised after 18h of the 5th day. Using GC‐MS (Gas chromatography mass spectroscopy) with flame ionization detector (FID), we found that CaMKII activation resulted in significant reduction in total lipid content. When KN93 (CaMKII inhibitor) was injected in the muscle prior exercise, the content of lipid was significantly higher than of muscle of the exercised rat without KN93, however similar to that of non‐exercised rat muscle. Exercise also resulted in increased CPT‐1 protein content whereas inclusion of KN93 prior exercise significantly altered the observed CPT‐1 upregulation. These observations indicate that CaMKII plays a significant role in lipid oxidation metabolism.
Keratinocyte Growth Factor and Dexamethasone Treatments Protect Lung Epithelial Cells from Death by Oxidative Stress
Authors: Benipal Bavneet , Lien Yu-Chin , Fisher Aron , Feinstein Sheldon
Published: June 2021

The goal of this study was to determine whether keratinocyte growth factor (KGF) and dexamethasone (Dex) treatments, which induce peroxiredoxin 6 (Prdx6) production, can protect lung epithelial cells against oxidative stress. Prdx6 is an enzyme that has both phospholipase A2 and peroxidase activities and can reduce peroxides, including lipid hydroperoxides, by a unique mechanism employing a single conserved cysteine. It is highly expressed in lung, both protecting against oxidative stress and regulating pulmonary surfactant phospholipid metabolism. A549 cells and primary cultures of rat lung alveolar type II cells were treated with 20 ng/ml KGF or 1 μM Dex or both and exposed to tert‐butyl hydroperoxide (tBOOH) or hydrogen peroxide (H2O2) for 24 hours at several concentrations. Cell survival was assessed using the MTT or neutral red assays. For A549 cells and rat type II cells treated with tBOOH, the LD50 was about 150 μM while for A549 cells treated with H2O2 it was 250 μM. Treatment with KGF or Dex alone protected cells substantially, while treatment with both KGF and Dex had an additive effect. Our studies show that KGF and Dex, which increase Prdx6 expression, can protect cells from oxidative stress and the combination is more protective than either treatment alone. Further study will be needed to determine whether Prdx6 is responsible for the observed protection in lung epithelial cells. Supported by HL P01‐75587.
Thirteen days of normobaric intermittent hypoxia increases SpO2 in a 71‐year‐old male
Authors: Kambis Kenneth Wicker , Yasukawa Mihio , Moran Thomas , Sobel Bart
Published: June 2021

PURPOSE

Normobaric Intermittent Hypoxic Exposure (NIHE) is thought to reduce risk of Acute Mountain Sickness (AMS) during subsequent exposure to higher altitudes by increasing arterial oxygen saturation of hemoglobin (SpO2). The purpose of this study was to evaluate the effect of 13d of NIHE on older adult SpO2.

METHODS

A 71 y.o., 185 cm tall, 84 kg, healthy male informed volunteer, participated in this IRB approved study. A former marathoner, BR has a history of mitral valve replacement (1995) and hemorrhagic stroke (2001). After a thorough physical examination and stress test by a cardiologist, he was pronounced in superior fitness and approved for the study. BR was exposed to normobaric hypoxic simulated altitudes of 2438 m to 5486 m in ~300 m increments for 3–4h/d for 13d. Every 15 minutes of exposure, SpO2 and heart rate (HR) were measured. On d0 and on d13, BR was exposed to 6 normobaric hypoxic simulated altitudes (1830 m – 4663 m).

RESULTS

During 13d of NIHE, SpO2 was reduced from d1 (2486 m) to d13 (5486 m; r= −0.947, P< 0.001). When the d0 SpO2 values at the 6 simulated altitudes were compared to d13 values, significant increases (P<0.05) were found.

CONCLUSIONS

NIHE appears to be a safe and effective method of increasing SpO2 in older adults preparing for strenuous high altitude exertion. Supported by: The Foundation for Aging Studies and Exercise Science Research and, The Borgenicht Program at William & Mary
Validation of Epigenetic Markers in Endometriosis
Authors: Colon Mariano , Colon Maricarmen , Monteiro Janice , Baez Perla , Ruiz Abigail , Flores Idhaliz
Published: June 2021

Endometriosis is a gynecological disease that affects 1 in 10 women during their reproductive years, cause incapacitating pelvic pain and infertility. Endometriosis is characterized by the growth of endometrial tissue outside the uterine cavity. Endometriosis has no cure, its diagnosis can only be done via surgery, there are no non‐invasive markers and its etiology is unknown. Epigenetic mechanisms may play an important role in the etiology of this disease. The modification of histones by acetylation and methylation of lysine residues has been shown to regulate gene expression by changing chromatin structure. We have shown by ELISA that endometriotic lesions have high levels of acetylated histone 3 at lysine residue 9 (H3K9ac).

Aim

To measure H3K9ac levels using immunohistochemistry (IHC).

Methods

Using a Tissue Microarray (TMA) composed of 163 biopses (endometriotic lesions and control tissues) we will assess levels of H3K9ac by IHC. Troubleshooting experiments identified the 1:100 as the appropriate dilution of the antibody.

Results

We expect that endometriotic lesions will express lower levels of H3K9ac than controls. Also, we expect that endometrium of patients express higher levels of H3K9ac than endometrioctic lesions. IHC will be able to show the cell specificity (i.e., glands vs. stroma) and will confirm the nuclear localization of this histone mark. In addition, we will be able to determine possible differences in this mark by lesion type (ovarian vs. peritoneal). These results will help understand how genes are regulated in endometriosis, and may support the use of novel treatments for this incurable disease. R25GM096955
DNA oxidation in regulatory D‐loop region of the mitochondrial genome is important for hypoxia‐induced mitochondrial biogenesis in rat pulmonary artery endothelial cells (PAECs)
Authors: Ruchko Mykhaylo V. , Gorodnya Olena M. , Gillespie Mark N. , Pastukh Viktor M.
Published: June 2021

Emerging evidence suggests that hypoxia‐induced transcription of nuclear genes requires controlled DNA damage and repair (Al‐ Mehdi et al, Sci Signaling, 2012). Whether such a mechanism operates in mitochondria is unknown. Here we explored the hypothesis that hypoxia‐induced mitochondrial (mt) DNA replication requires sequence‐specific oxidative DNA modifications in the mitochondrial genome. Using slot blot and quantitative Southern blot analyses, respectively, we found that increased mtDNA copy number in hypoxic PAECs was accompanied by oxidative modifications in the D‐loop region of the mitochondrial genome. Transfection of rat PAECs with a mitochondria‐targeted DNA repair enzyme OGG1 suppressed hypoxia‐induced DNA oxidation in the D‐loop region and reduced binding of mitochondrial transcription factor TFAM to both regulatory and coding regions of the mitochondrial genome, as assessed by ChIP analysis. OGG1 over‐expression also attenuated hypoxia‐induced mtDNA replication. Interestingly, hypoxia increased number of hyperpolarized mitochondria in wild‐type PAECs, as evidenced by JC‐1 fluorescence. Mitochondria in OGG1‐transfected cells were hyperpolarized under normal conditions and less affected by hypoxia. These observations support the concept that oxidative DNA modifications in the D‐loop region are important for hypoxia‐induced mitochondrial biogenesis. Supported by NIH.
Vitamin E and nonalcoholic fatty liver disease (NAFLD)
Authors: Thakur Varsha , Ulatowski Lynn , Parker Robert , Zhang Renliang , Liu Xiuli , Manor Danny
Published: June 2021

Nonalcoholic fatty liver disease consists of a continuum of hepatic injuries, from simple steatosis to nonalcoholic steatohepatitis (NASH), and accounts for the majority of chronic liver diseases in the Western world. The penetrance of NAFLD is especially high amongst obese individuals. Presently, no effective treatment option is available for this disease, and it sometimes progresses to cirrhosis and liver cancer. Multiple lines of evidence indicate that oxidative stress may be an important factor in the pathological progression of NAFLD. Moreover, dietary supplementation with the antioxidant vitamin E was shown to be efficacious in attenuating pathologic progression in human NAFLD patients (Sanyal et al NEJM 2010).

To gain mechanistic insights into the effects of vitamin E on NAFLD, we examined the presence and progression of liver injury in a genetic model of vitamin E deficiency, namely mice lacking the alpha‐tocopherol transfer protein (αTTP). When placed on a vitamin E deficient diet, TTP‐null mice displayed many hallmarks of NAFLD, namely accumulation of macro‐ and micro‐vesicular lipid droplets, hepatic injury evidenced by elevation of circulating alanine aminotransferase, increased hepatic collagen deposition, and inflammation of the liver, evidenced by marked increase in TNFα. Importantly, many of these parameters normalized upon dietary supplementation with vitamin E. Interestingly, dietary supplementation with vitamin E did not prevent lipid deposition in the TTP‐null livers, indicating that TTP is essential for mediating the full complement of benefits associated with vitamin E supplementation. (This work was supported by NIH grant ROIDK067494).
The SUMO Ligase PIAS1 Promotes the Progression and Survival of Solid Tumors of Different Histopathological Origins
Authors: Constanzo Jerfiz D , Rabellino Andrea , Konstantinidou Georgia , Schuster Katja , Scaglioni Pier
Published: June 2021

Protein inhibitor of activated STAT‐1 (PIAS1) regulates cell signaling via posttranslational modifications of proteins. PIAS1 conjugates small ubiquitin like modifiers (SUMO‐chains) onto target proteins. Our lab discovered that PIAS1 SUMOylates the promyelocytic leukemia (PML) tumor suppressor. This event promotes degradation of the tumor suppressor (PML) aiding tumor progression.

Thus, we hypothesized that PIAS1 will promote cell survival and progression in human cancers where it is highly expressed, representing an unexplored vulnerability in these cancers. We performed gain and loss of function experiments to test PIAS1 effect on tumorigenesis with transformation assays, and studied cell survival after PIAS1 depletion using RNAi. Ectopic overexpression of Pias1 protein in NIH3T3 and mouse embryonic fibroblasts (MEFs) promotes colony formation and loss of contact inhibition in culture. PIAS1 depletion using shRNA in non‐small cell lung cancer (NSCLC) and breast cancer cell lines leads to increased PML levels, following susceptibility to apoptosis and impaired cell proliferation. Moreover, siRNA depletion of PIAS1 in NSCLC cell line H460 reduces cell spreading and impairs migration in wound healing assays. Our findings show for the first time a novel role for the SUMO ligase PIAS1 in tumor progression and survival. Our findings lay the groundwork for the study of PIAS1 in human cancers.
Detection of Aniline resistant and Streptomycin sensitive Yeast R11 in processed Biosolid
Authors: Kannan Subburaj , Montoya Hector , Barrera Jorge , Longoria Elena , Castillion Ariana
Published: June 2021

It is known that five nitroaniline and dinitroaniline isomers including 2‐nitroaniline, 3‐nitroaniline, 4‐nitroaniline, 2,4‐dinitroaniline and 2, 6‐dinitroaniline were detected in the acidic wastewater and in printing and dyeing wastewater. In this study, Yeast R112 was inoculated in the biosolid(biosludge) to determine the growth pattern and tolerance level to aniline isomer derived toxicants. Biosolid was obtained from the City of Eagle Pass Waste Water processing plant. The biosolid represent organic and inorganic remains of raw sewage subjected to oxygen treatment in the aeration chamber. Yeast R112 showed substantial growth in the undiluted biosludge, but not in the higher dilutions. Virtually no Yeast R112 was detected in the 10–2, 10–3, 10–4, and 10–5 dilutions. Also, antibiotic sensitivity was performed by Kirby‐Bauer disk diffusion antibiotic sensitivity testing. Yeast R112 strain resistant to Aniline isomers showed selective growth inhibition by streptomycin 50μg/disc. Whereas a normal growth pattern was observed in the presence of Methicillin, Chloramphenicol, and Kanamycin. Taken together, Yeast R112 strain showed resistance to Aniline isomers present in the sewage water. Also, Yeast R112 strain showed sensitive to Streptomycin.
Role of the GRK2 extreme amino terminus and active site tether in forming G protein‐coupled receptor docking site
Authors: Sterne‐Marr Rachel , Michalski Kevin , Beautrait Alexandre , Lopez Thomas , Mannix Katelynn , McDonald Devin , Cutter Amber , Francis Charnelle , Medina Christopher , Bouvier Michel , Tesmer John
Published: June 2021

G protein‐coupled receptor (GPCR) kinases (GRKs) phosphorylate agonist‐activated GPCRs to initiate receptor desensitization, but the mechanism of kinase domain activation is unclear. Because agonist‐activated receptors dramatically stimulate the catalytic activity of GRKs, we sought to identify GRK2 residues located outside the active site that play a role in receptor interaction. Previous work suggested that active site tether (AST) residues of the kinase carboxyl‐tail extension are required for GRK activation, and that N‐terminal helix and AST interaction is important for receptor phosphorylation. We therefore carried out systematic mutagenesis of residues 3–18 and the AST of GRK2, and characterized these mutants using in vitro rhodopsin and peptide phosphorylation, GRK activation, intact cell β2‐adrenergic receptor phosphorylation, and cell based α2‐adrenergic receptor recruitment assays. Our results suggest that the N‐terminus and AST, both intrinsically disordered in the inactive kinase, play important roles in formation of the interaction site with activated receptors. Funding: NSF (RSM), NIH (JT), Canadian Institute for Health Research (MB), and FRSQ & Groupe de Recherche Universitaire sur le Médicament (AB).
The relationship between sex hormones and osteoarthritis
Authors: Feresin Rafaela G , Elam Marcus L , Zhao Yitong , Hooshmand Shirin , Arjmandi Bahram H
Published: June 2021

Osteoarthritis (OA) is the most common joint disorder affecting more than 27 million Americans. Sex hormones have been shown to play an important role in the pathogenesis of OA. For instance, up‐regulation of locally synthesized (by cartilage) estrogens and androgens deleteriously influence cartilage and jeopardize joint integrity in women and men, respectively. Thus, the purpose of this study is to provide some evidence for the etiology of OA by investigating the influence of systemic versus local estrogen and testosterone levels in serum and synovial fluid (knee joint) of men and women with varying degrees of OA. In addition, this study will compare the abundance of ERα, ERβ, androgen receptors, aromatase, and 5α‐reductase protein and mRNA levels in the cartilage to the severity of OA. Cartilage, synovial fluid, and serum samples were obtained from 30 men and 30 women, ages of 44 to 85 years with knee OA who underwent arthroscopic surgery. Estradiol and testosterone levels in serum and synovial fluid will be measured using ELISA. Protein and mRNA levels of ERα, ERβ, androgen receptors, aromatase, and 5α‐reductase will be assessed by western blot and RT‐PCR, respectively. The long‐term goal of this study is to make progress towards the development of OA treatments that could alter the progression of this degenerative disease by blocking the deleterious effects of local estrogen and testosterone on cartilage.
The changing influence of wealth, education and urbanization on overweight and obesity in Guatemalan women between 1995 and 2008
Authors: Kanter Rebecca , Fort Meredith P. , Ramirez Manuel , Martorell Reynaldo
Published: June 2021

Overweight and obesity (BMI≥25.0) prevalence in Guatemalan women (15–49y) was examined in the 1995, 1998, 2002, and 2008 National Maternal and Infant Health Surveys (ENSMI). Associations with socio‐demographic factors (age, education, ethnicity, urban‐rural, parity, and wealth) were analyzed using STATA SE 12.0. Survey weighted logistic regression models adjusted for age and parity assessed associations between BMI≥25.0 and socio‐demographic factors. Prevalence of BMI≥25.0 was 34.1% in 1995 and 59.4% in 2008; and BMI≥30.0 rose from 7.9% to 22.4%. Tests for trend (p<0.0001) confirmed BMI≥25.0 increased in most socio‐demographic groups over time. In 1995, high wealth (OR = 5.97, CI: 4.23, 8.43, p<0.001) and urban areas (OR = 1.32, CI: 1.08, 1.62, p<0.01) were associated with a BMI≥25.0. In 1998, 2002, and 2008, education and urban areas were significantly positively associated with BMI≥25.0. In 1995, education (OR = 1.81, CI: 1.19, 2.75, p<0.01) and high wealth (OR = 7.07, CI: 3.99, 12.52, p<0.001) were associated with BMI≥30.0; in 1998 wealth was not significant. From 2002 and 2008 the association with BMI≥30.0 and education decreased (OR = 1.97, CI: 1.55, 2.50, p<0.001; OR = 1.52, CI: 1.30, 1.78, p<0.001), while urban areas increased (OR = 1.34, CI: 1.05, 1.72, p<0.05; OR = 1.67, CI: 1.42, 1.95, p<0.001). Education, and urban areas explain more of the increase in BMI≥25.0 from 1995 to 2008. This research was not funded.

Grant Funding Source: None
Factors influencing the 25‐hydroxyvitamin D level of pregnant women and their newborns in Giessen, Germany
Authors: Wuertz Catrin , Gilbert Peter , Baier Wolfgang , Kunz Clemens
Published: June 2021

There is mounting evidence of an association between a low maternal 25‐hydroxyvitamin D (25(OH)D) level and an increased risk of adverse pregnancy outcomes. In a cross‐sectional study blood samples of 261 women and 328 newborns were collected for analysis of 25(OH)D (CLIA, DiaSorin), parathyroid hormone, alkaline phosphatase and calcium. Demographical, habitual and clinical data were obtained from a questionnaire and from the medical file. 77% of the women and 69% of the newborns had a vitamin D deficiency (25(OH)D < 50 nmol/l). The median maternal and newborn 25(OH)D levels were 24.9 (25th, 75th percentile 12.6, 45.4) nmol/l and 34.1 (17.7, 58.5) nmol/l. Logistic regression analysis revealed that significant risk factors for maternal vitamin D deficiency were physical inactivity (aOR 2.7, 95% CI 1.1, 6.8) and a non‐European country of origin (aOR 3.2, 95% CI 1.0, 10.3) after controlling for season and independent risk factors. Maternal vitamin D deficiency was a significant predictor of newborn vitamin D deficiency after controlling for season (aOR 70.2, 95% CI 17.9, 275.8). These are the first 25(OH)D data for pregnant women and their newborns in Germany. They very clearly indicate the great need for urgent action to prevent low vitamin D levels. Because of the marginal dietary sources and the lacking endogenous synthesis of vitamin D during the winter a daily supplementation is strongly recommended. DRKS0000324