bluuebunny/crossref_metadata_2025 huggingface dataset:
Interspecific effects of body size on grasshopper jumping performance
Authors: Kirkton Scott D , Harrison Jon F
Published: April 2020

The scaling of jump performance has been relatively well studied in vertebrates; however, less is known about how interspecific body size differences affect insect jumping. In this study, we compare the morphology, jump performance, lactate production, and relative cost of drag in sixteen species of grasshoppers using both standard regression and phylogenetic generalized least squares analyses. In contrast to the situation observed across species in vertebrates, and during ontogeny in Schistocerca americana grasshoppers, small grasshopper species had similar jump distances as larger species, despite losing more energy to drag. This surprising result could be at least partially explained by the fact that smaller grasshopper species had relatively more jumping muscle mass, relatively longer femurs, and produced more lactate during repeated jumping. These data suggest that selection has acted strongly to improve jump performance in smaller grasshoppers via a variety of morphological and biochemical adaptations.
Dimethyl fumarate regulates the IL‐17‐ACT1‐TBK1 axis‐mediated IκB‐ζ expression by influencing the phosphorylation of Regnase‐1
Authors: Matsuda Tadashi , Ohagakiuchi Yui , Saino Yuka , Komori Yuki , Sato Ami , Hirashima Koki , KItai Yuichi , Kashiwakura Jun-ichi , Oritani Kenji , Muromoto Ryuta
Published: April 2020

The signaling elicited by the cytokine interleukin‐17A (IL‐17) is important for antimicrobial defense responses, whereas excessive IL‐17 production leads to autoimmune diseases such as psoriasis and multiple sclerosis. IL‐17–induced stabilization of mRNAs has been recognized as a unique and important feature of IL‐17 signaling. We previously demonstrated that an IL‐17 signaling protein, ACT1 is required to counteract constitutive inhibitor of nuclear factor kappa B zeta (IκB‐ζ) mRNA degradation by the ribonuclease Regnase‐1. In the present study, we tried to clarify this mechanism in more detail and identify an agent that can inhibit IL‐17–induced mRNA stabilization. Experiments using siRNA and an inhibitor of TANK‐binding kinase 1 (TBK1) revealed that TBK1 was also required for IκB‐ζ mRNA stabilization through Regnase‐1 phosphorylation. Interestingly, this TBK1‐mediated phosphorylation of Regnase‐1 was suppressed by the addition of an electrophilic small molecule, dimethyl fumarate (DMF) that has been used to treat IL‐17–related autoimmune diseases. Furthermore, confocal microscopic observation of the cellular localization of ACT1 revealed that DMF treatment resulted in the disappearance of ACT1 nuclear dots and perinuclear accumulation of ACT1. These results suggested that DMF is a small molecule that compromises IL‐17–induced activation of the ACT1‐TBK1 pathway, thereby inhibiting IL‐17–induced mRNA stabilization.

Support or Funding Information

This work was supported in part by the Japan Society for the Promotion of Science (JSPS) KAKENHI grant (numbers 254600560, 17K08263, and 19H03364).
TgMIF regulates inflammatory response by mediating TLR signaling
Authors: Kim Donggyu
Published: April 2020

Toxoplasma gondii (Tg) is an obligate intracellular parasite that can invade and replicate in almost all nucleated cells of warm‐blooded animals. It has a world‐wide geographic distribution and is known to infect many species of birds and mammals, including approximately one third of humans. Macrophage migration inhibitory factor (MIF) is a proinflammatory molecule in mammals that, unusually for a cytokine, exhibits tautomerase and oxidoreductase enzymatic activities. MIF has been demonstrated to be involved in innate and adaptive immune responses by stimulating the production of pro‐inflammatory mediators, such as TNF‐α and IL‐8. In this study, we measured that Toxoplasma gondii MIF (rTgMIF)‐induced levels of TNF‐α, IL‐1β, and IL‐6 by ELISA and protein‐protein interaction involved in TLR signal transduction.

We investigated the function of rTgMIF in the inflammatory response. We showed that rTgMIF affects the secretion of pro‐ and anti‐inflammatory cytokines. We showed that rTgMIF also regulates MAPK signaling in BMDMs. Taken together, these results demonstrated that TgMIF is related to pathogenesis of Toxoplasma gondii through regulation of MAPK pathway in TLR signaling.

Support or Funding Information

ACKNOWLEDGEMENTS

This work was supported by the NRF grant funded by the Korea government (MSIP) 2019R1I1A2A01064237.
Inhibitory effects of ginsenosides on S100 protein‐mediated rheumatic diseases
Authors: Cho Eun-i
Published: April 2020

Rheumatoid arthritis(RA) is a chronic autoimmune disorder that primarily affects joints. It causes severe long‐term damage to various organ systems and results in progressive disability, systemic complications. The cause of rheumatoid arthritis is not well understood but involves a complex interplay of exogenous and endogenous inflammatory factors. Especially, numerous evidence show that S100 family also known as the pro‐inflammatory calcium‐binding protein, plays a pivotal role in the pathogenesis of rheumatic diseases. Some S100 proteins are released at the site of inflammation and participate in inflammatory signalling pathways via a variety of receptors. Therefore, these S100 proteins could be useful biomarkers for clinical joint damage in RA. Ginsenosides are a class of natural product steroid glycosides and triterpene saponins, which has a long history of use in traditional medicine. It has also been used in clinical practice for resolution of inflammation. In this study, we measured that RAW264.7 murine macrophage cell line and murine BMDMs are transfected or treated with S100 in presence of ginsenosides. The levels of cytokines were measured by ELISA. The results show that secretion of S100 proteins was inhibited by ginsenosides and pro‐inflammatory cytokines was decreased at the site of inflammation. Taken together, we suggested that pro‐inflammatory signaling of S100 proteins is inhibited by ginsenoside and this result indicated that effect of ginsenoside in S100‐mediated inflammation might be potential therapeutic target for resolving rheumatoid disease.

Support or Funding Information

This work was supported by the NRF grant funded by the Korea government (MSIP) 2019R1I1A2A01064237
Development of substitute of R‐spondin 1 for production of prostate organoid
Authors: Cha Ji Hyeon , Heo Hee Sun , Lee Seung Hwan , Lee Jong Hun
Published: April 2020

Organoid that is organ‐like cell aggregates has both the advantages of 'in vitro' and three‐dimensional advantages of animal experiment. It is very useful for modeling disease, drug development research, and testing customized medicine.

R‐spondin 1 stimulates Wnt / β‐catenin signaling to induce proliferation of organoids. Thus, high concentrations of R‐spondin 1 are essential for organoid culture. However, production of R‐spondin 1 is limited and, due to the high cost of R‐spondin 1, there are financial difficulties in the practical application of organoids. We have found an alternative to the R‐spondin 1 protein from natural compounds. We also evaluated how this natural component affects prostate organoid.

We confirmed that prostate organoids were established from selected natural products. Cultures were divided into Group A with R‐spondin 1, Group B with natural products and Group C with R‐spondin 1 and natural products.

In order to confirm whether organoids actually have prostate tissue characteristics, we confirmed expression of basal specific markers and, lumen specific markers including prostate specific antigen (PSA). Cyrokeratin 8 and PSA were used as luminal specific markers, and p63 and cytokeratin 5 were used as basal specific markers.

This study showed that R‐spondin 1, Lgr5 antagonist, could be replaced by natural compound in organoid culture. We also could observe that natural‐derived components helped R‐spondin 1 to promote growth of prostate organoid and inhibit apoptosis. This research is expected to contribute to the research of organoids that will be developed in the future.
RNA‐Binding Protein QKI is a critical pre‐RNA splicing regulator for cardiac development and function
Authors: Liu Ying , Chen Xinyun , Xu Chen , Sun Ning , Shou Weinian
Published: April 2020

RNA‐binding protein QKI (also known as Quaking) belongs to hnRNP K‐homology (KH)‐domain protein family and is implicated a novel regulator in pre‐RNA splicing. As previously shown, the QKI homolog how potentially regulates heart lumen formation and function in fruit fly. However, QKI’s function in mammalian heart development is largely unknown. We found that Qki specifically expressed in mouse developing hearts as early as E7.0 and this high‐level expression was maintained in the hearts to adulthood. Consistent with this finding, by using human embryonic stem cells (hESCs)‐cardiomyocyte differentiation system, QKI expression was found up‐regulated at Day 6, a critical time frame that cardiogenic progenitor transition into early cardiomyocytes. To determine the biological function of QKI in cardiogenesis, we generated hESCsQKIdef by using CRISPR/Cas9 gene editing technology and analyzed the physiological impact of QKI‐deficiency to cardiomyocyte differentiation and maturation. As expected, we did not find defect in pluripotency in hESCsQKIdef, but the contractile function was dramatically reduced in hESCsQKIdef‐derived cardiomyocytes. Based on scRNA‐seq and clustering analyses, there was no significant difference in the distribution of the clusters when compared hESCsQKIdef‐cardiomyocyte group to normal control group, suggesting that the cardiomyocyte differentiation was largely unaffected in hESCsQKIdef. However, scRNA‐seq demonstrated a handful of genes encoding sarcomere proteins were significantly altered in cardiomyocyte clusters, which included ACTN2, ACTA1, MYH7, and MYL2, etc., indicating a significant defect in cardiomyocyte maturation. Bulk RNA‐seq data not only further confirmed this notion, but also provided additional depth for detailed analysis on the QKI‐mediated pre‐RNA splicing events. In conjunction with QKI‐binding motif search and RNA immunoprecipitation (RIP) analysis, we were able to demonstrate that QKI was a key pre‐RNA splicing regulator for sets of genes involved in cardiac myofibrillogenesis and cardiac excitation‐contraction (E‐C) coupling, which ultimately impacted on cardiomyocyte maturation and function. Additional analysis of cardiac developmental defects in Qkibgeo mutant mice further validated that QKI was an indispensable splicing regulator in cardiovascular development. Our results not only revealed a novel mechanism by which QKI regulates pre‐mRNA splicing in cardiogenesis, but also implied that QKI was likely involved in the pathogenesis of certain forms of congenital heart defects and cardiomyopathies.

Support or Funding Information

This work was supported in part by National Institute of Health P01HL134599
Differential ontogeny of adipose tissue development and the impact of maternal dietary supplementation on gene expression pathways and profiles.
Authors: Alagal Reham , Fainberg Hernan , Budge Helen , Symonds Michael
Published: April 2020

To understand the developmental origins of the molecular changes within different adipose depots in early life and how maternal diet supplementation with canola oil alters the same, a microarray based gene expression study was conducted on three fat depots (epicardial, perirenal and sternal) of 28 day old sheep. Multi‐region bioinformatics analysis demonstrated that each depot exhibited a distinct gene expression profile and contained a small number of unique modules of co‐expressed genes associated with distinct biological processes. Perirenal adipose tissue therefore exhibited a greater tendency to differentiate. In this fat depot, genes involve in energy metabolism (DECR1, GHR and PPARG), angiogenesis (HOXA5 and HIF1α) and pre‐adipocyte precursor genes (HOXB6, HOXB8 and HOXB5) were co‐expressed. In contrast, sternal and epicardial adipose tissues showed lower plasticity. Interestingly, genes involved in energy metabolism were actively repressed in the sternal depot. Following maternal dietary supplementation, the perirenal adipose tissue depot was the most responsive. It showed an upregulation of genes involved in inflammatory responses, lipid metabolism, cell differentiation and modulation of thermogenesis in brown adipose tissue. Namely, the following genes were modulated i.e. NR3C1, CYB5B, GLTSCR1L and FATR. We also have shown the unique response of different adipose depots following maternal supplementation of diet with canola oil. In conclusion, we highlight the potential effect of anatomical location on adipose tissues development. In particular, the perirenal adipose depot shows more plasticity in changes in gene expression during development at early life and is the most responsive following maternal dietary manipulation with canola oil, as compared to sternal and epicardial adipose depots.
Antifungal and cytotoxic activities of acetone and aqueous extracts of Morinda lucida
Authors: Olawuwo Stephen Olasunkanmi , Aro Abimbola Obemisola , Kabongo-Kayoka Prudence Ngagule , Eloff Jacob Nicholas , McGaw Lyndy Joy
Published: April 2020

Aspergillosis is an opportunistic airborne fungal infection caused by ubiquitous saprophytic moulds of the Aspergillus genus. Infection has been documented in humans and animals, and it usually occurs through inhalation of unicellular spores known as conidia or, more rarely, after ingestion or wound contamination. Oral candidiasis is also an important sign for clinical diagnosis and an indicator of the evolution of immunodeficiency among HIV carriers. Mycotic stomatitis in animals is caused by Candida spp. in most cases. Cryptococcosis is a fungal disease of man and domestic animals, caused by the Cryptococcus neoformans complex. In animals it is often characterized by upper respiratory symptoms, subcutaneous nodules, pneumonia, central nervous system or ocular disorders, lymphadenopathy, or subcutaneous nodules. Antifungal drugs are often unaffordable to rural farmers and their efficacy is hampered as a result of antimicrobial resistance. Therefore, the need to seek alternative measures from phytogenic sources is urgent. The aim of this study was to investigate the in vitro antifungal activity of acetone and aqueous extracts of Morinda lucida (Rubiaceae) against ATCC strains of Aspergillus fumigatus, A. flavus, Candida albicans and Cryptococcus neoformans as well as clinical isolates of A. fumigatus and C. albicans using a serial microdilution assay. The cytotoxicity against human colon cancer (Caco‐2) cells and total activity of the acetone and aqueous extracts were also determined. The minimum inhibitory concentration (MIC) of both extracts against tested organisms ranged from 0.11 to 2.50 mg/ml and 0.03 to 2.50 mg/ml after 48 and 72 hours respectively. The LC50 values of the acetone and aqueous extracts against the Caco‐2 cells were 0.46 and 0.58 mg/ml respectively. The range of selectivity index (SI) values of both extracts was 0.23 to 5.27 and 0.23 to 19.33 after 48 and 72 hours respectively. The range of total activity of the extracts was 56 to 2 454 and 56 to 9 000 ml/g after 48 and 72 hours respectively. The potential of this plant species as an alternative for treatment of human and livestock mycoses is supported by these results. However, in vivo data is necessary to further investigate this claim
Mid‐gestation low‐dose LPS administration results in excessive weight gain upon a Western Style Diet in female mouse offspring
Authors: Plösch Torsten , Dijkstra Dorieke J. , Verkaik-Schakel Rikst Nynke , Stojanovska Violeta , Scherjon Sicco A.
Published: April 2020

Exposure to gestational complications can have a life‐long influence on the offspring’s health. Inflammation is observed in many complications, including preeclampsia and gestational diabetes mellitus, and is likely mediating the long‐term consequences for the offspring. Here we use a mouse model of LPS‐induced maternal inflammation to examine the consequences of gestational inflammation on offspring’s metabolic health. 25μg/kg LPS was administered to pregnant C57Bl6/J mice on gestational day 10. After weaning, all offspring was fed a semi‐synthetic control diet. At 12 weeks of age, half of the animals were switched to a Western‐Style Diet (WSD). Body composition (MRI) and glucose‐ and insulin tolerance were measured at 12 and 24 weeks of age. At 24 weeks of age, blood pressure was measured, and plasma, fat pads, liver and brain were collected and analyzed at the molecular level. Males and females were analyzed separately. LPS‐exposed female offspring showed an increase in body fat mass at 12 weeks of age, and the combination of LPS‐exposure and WSD markedly increased food intake, body weight and body fat mass in the females at 24 weeks of age. Observed changes in glucose‐and insulin tolerance, leptin and insulin levels and gene expression in liver, adipose tissue and hypothalamus were associated with body weight, not with in utero LPS exposure. In utero LPS exposure leads to increased intake of WSD in female offspring, which is likely causative of the observed increased body weight and fat mass and metabolic and molecular changes. A metabolic driver for increased food intake was not identified. Thus, in this model, gestational inflammation might lead to increased food intake through programmed behavioral traits rather that fetal programming of metabolism.

Support or Funding Information

This work was supported by ZonMw (91211053).
Calciprotein Particle Inhibition Explains Magnesium‐mediated Protection against Vascular Calcification
Authors: de Baaij Jeroen H.F. , ter Braake Anique , Eelderink Coby , Pasch Andreas , de Borst Martin , Hoenderop Joost
Published: April 2020

Phosphate toxicity is a strong determinant of vascular calcification development in chronic kidney disease (CKD). Phosphate induces the formation of calciprotein particles (CPP), which were identified to drive the calcification process. Epidemiological data of suggests that magnesium may improve cardiovascular risk by inhibiting vascular calcification. However, the mechanism by which magnesium counteracts vascular calcification is poorly understood. Here, we investigated the effects of magnesium on phosphate and secondary crystalline calciprotein particles (CPP2) induced calcification and crystal maturation.

Vascular smooth muscle cells were treated with high phosphate or CPP2 and supplemented with magnesium to study cellular calcification. The effect of magnesium on CPP maturation, morphology and composition was studied by medium absorbance, electron microscopy and energy dispersive spectroscopy. Effects of magnesium intake on aortic calcification was assessed in Klotho knock‐out mice. To translate our findings to CKD patients, the effects of magnesium on calcification propensity (T50) was measured in sera from CKD patients and healthy controls.

Magnesium supplementation prevented phosphate‐induced calcification in vascular smooth muscle cells. In contrast, magnesium failed to inhibit CPP2‐induced calcification, indicating it acts before the formation of CPP2. Increased expression of osteogenic genes osteopontin and alkaline phosphatase were unchanged after magnesium supplementation. Furthermore, in a Klotho knock‐out mouse model for vascular calcification, high dietary magnesium intake prevented aortic calcification effectively. Showing the potential of magnesium in patients, magnesium dose‐dependently delayed the maturation of CPP2 by several days. In human serum, addition of 0.2 mmol/L magnesium increased T50 from healthy controls by 51 ± 5 minutes (P < 0.05) and CKD patients by 44 ± 4 minutes (P < 0.05). Each further 0.2 mmol/L addition of magnesium led to further increases in both groups.

Our results demonstrate crystalline CPP2 mediate phosphate‐induced calcification. Magnesium prevents crystalline CPP2 formation and thereby prevents phosphate toxicity leading to calcification. Magnesium supplementation, even at low dosages, is a potential therapeutic strategy to reduce vascular calcification in CKD.

Support or Funding Information

This research was funded by grants from the Netherlands Organization for Scientific Research (NWO Veni 016.186.012 and VICI 016.130.668) and the Dutch Kidney Foundation (Kolff 14OKG17 and 15OP02). This work was furthermore supported by the NIGRAM2+ consortium funded by Health Holland (LSHM17034) and the Dutch Kidney Foundation (16TKI02).
Aqueous lyophilized extract of Moringa oleifera ameliorates streptozotocin‐induced diabetes in a fashion similar to pioglitazone
Authors: Ogunmiluyi Iyanuoluwa Omolola , Falayi Olufunke Olubunmi , Ogunpolu Blessing Seun , Oyagbemi Ademola Adetokunbo , Omobowale Temidayo Olutayo , Adedapo Adeolu Alex
Published: April 2020

Abnormally high glucose levels in the blood, a major characteristic of diabetes, has been associated with a high level of oxidative stress, believe to play a major role in the development of diabetes and diabetic complications through various mechanisms that result in cellular damage. This study evaluates the role of lyophilized Moringa oleifera in ameliorating streptozotocin‐induced diabetes and the possible signaling pathway involved. Diabetes was induced by a single intraperitoneal (I.P.) injection of freshly prepared solution of 25 mg/kg of Streptozotocin (STZ); and after 72 hours, rats with glucose level of 250 mg/dL and above were taken to be diabetic and were used in the study. Twenty‐five animals were divided into five equal groups. Group A was the control that received water alone throughout, B was the untreated diabetic group, C was the non‐diabetic but treated with Moringa oleifera (100 mg/kg), D was the diabetic and treated with Moringa oleifera (100 mg/kg) and E was the diabetic group treated with 25 mg/kg dose of Pioglitazone. Blood glucose values were taken weekly. Serum, liver, kidney and heart were taken for markers of organ damage, inflammation and oxidative stress. Expression of GLUT 4 in the pancreas and liver was done using immunohistochemistry technique. Treatment with aqueous lyophilized Moringa extract resulted in significant decrease in blood glucose levels, decrease in the level of indicators of renal and hepatic damage (ALT, AST, urea and creatinine), as well as decrease in the levels of the markers of renal, hepatic, and cardiac oxidative stress (MDA, hydrogen peroxide, and PC). Enzymatic and non‐enzymatic anti‐oxidants (SOD, GST, GPx, GSH) were also improved in the renal, cardiac and hepatic homogenate. Expression of GLUT 4 was down‐regulated in the untreated group while treatment with Moringa and Pioglitazone caused increase in the expression of this protein. It can thus be concluded that aqueous lyophilized Moringa extract has an anti‐diabetic effect in a fashion similar to pioglitazone, as evidenced by the decrease in the level of glucose, markers of oxidative stress; increase in the levels of both enzymatic and non‐enzymatic antioxidants and the up‐regulation of GLUT 4 which is relevant in reversing resistance to insulin in the same manner as the standard drug used in this study.
Understanding the Structure of Trabecular Excrescences Found in Osteoarthritic and Non‐Osteoarthritic Joint Samples
Authors: Taylor Adam M. , Allison Kathryn , Rhodes Bryan , Kerns Jemma G. , Shepherd Rebecca F.
Published: April 2020

The presence of novel bone structures, termed trabecular excrescences, have previously been found in surgical waste tissue samples from patients with osteoarthritis, alkaptonuria and from aged bone samples. A form of trabecular excrescence is found associated with the presence of bone marrow adipocytes and the absence of bone lining cells. We hypothesize that the adipocytes are contributing to the formation of bone like structures. This is inconsistent with the current paradigm of osteoclast‐osteoblast coupled remodelling. This study aimed to determine the topographical distribution of trabecular excrescences throughout the trabecular bone in human joint samples and to further characterise their internal structure.

Joint samples were obtained as surgical waste, with informed patient consent, from 20 patients with OA (mean age = 75.3) and 20 control patients without OA (mean age = 78.5). North of Scotland NHS Research Ethics Committee (UK) approved this research. Joint samples obtained included femoral heads, humeral heads, ankles, knees and shaft of femur. Each joint sample was sliced into sagittal sections. The sections were decalcified and processed for histology. Unstained sections were viewed under fluorescence microscopy. Sections were stained with haematoxylin and eosin and with immunohistochemical staining for collagen I, collagen VI and osteocalcin.

Excrescences were found in both control and osteoarthritic samples. When found, the excrescences were often clustered, with several protrusions shown along a region of trabeculae. These results demonstrate that trabecular excrescences are globally distributed throughout the femoral head, and not specific to areas of osteoarthritic change. This suggests the formation of these structures may be linked to the ageing process rather than disease progression. Immunohistochemical staining demonstrated the presence of collagen VI within the extracellular matrix of the newly formed excrescences. However, we also noted the presence of osteocalcin in ‘smoothed over’ versions of excrescences.

This study has shown that trabecular excrescences are initially formed by the deposition of collagen VI by adipocytes. However, the presence of osteocalcin suggested that excrescences have the potential to be remodelled through typical osteoblast‐osteoclast remodelling. Future treatments for skeletal disorders could look to exploit the scaffold like properties of structures formed by adipocytes.

Support or Funding Information

Anatomical Society PhD Studentship
Controlling the Bioluminescence Output from Shewanella woodyi using Soluble Electron Acceptors
Authors: Alsabia Sahar , Mortensen Chris , Theberge Allison , Blair Anna , Wendel Nina , Biffinger Justin
Published: April 2020

We will present bioluminescence data from Shewanella woodyi cultures in order to correlate bioluminescence intensity with changes in the electrochemical potential in a saltwater medium using soluble electron acceptors. A relationship between the concentration of NaNO3, CoCl2, or ZnCl2 to bioluminescence intensity was confirmed using aerobic cultures of S. woodyi at 20°C with glucose as the sole carbon source. In general, increasing the concentration of nitrate, Co(II), or Zn(II) reduced the bioluminescence per cell, with complete luminescence being repressed at ≥5 mM nitrate and ≥0.5 mM Co(II). Results from cell viability fluorescent staining concluded that increasing the concentration of Co(II) or nitrate did not affect the overall viability of the cells when compared to cultures lacking Co(II) or nitrate. Our presentation will show that electrochemical potentials of < 0.2 V vs NHE repress the luminescence from the cells, but the exact mechanism is unclear. Our results indicate that the luminescence intensity from S. woodyi could be systematically reduced using these three soluble electron acceptors making S. woodyi a potential model bacterium for whole‐cell luminescence bioelectrochemical sensor applications.

Support or Funding Information

University of Dayton Graduate Student Summer Fellowship and Dean's Summer Fellowship programs
The Problems with the Concepts of Osmole and Osmotically Active Particles
Authors: Kuang Serena Y. , Walter Stefan , Yang Xiaoqi , Li Xiaonan
Published: April 2020

Introduction

In this presentation, we address and resolve the problems with the current definitions of osmole and the widely used term “osmotically active particles”. This presentation precedes and connects to our second abstract titled “The Concept of Osmolarity: Problems and Resolutions” in EB2020.

Method

Logical reasoning.

Results

1) A simple osmosis system is first defined as S‐m‐H2O: a solution (S) compartment and a water compartment separated by a selectively permeable membrane (m) that can be either water‐permeable (only permeable to water) or SP‐permeable (permeable to water and some species of solute particles (SP)). This is a missing step to define osmole and true osmolarity. 2) The problems mentioned above can be identified in three sample definitions of osmole: “One osmole (Osm) is equal to 1 mole (mol) (6.02 × 1023) of solute particles”; “One mole of osmotically active particles is called one osmole”; and “the osmole (Osm or osmol) is a non‐SI unit of measurement that defines the number of moles of a chemical compound that contribute to a solution’s osmotic pressure”. These definitions are consistent if the m in S‐m‐H2O is water‐permeable but have different meanings when the m is SP‐permeable: the first includes osmotically ineffective, permeant SP, while the third excludes them. The second definition has a problem with the ambiguous term “osmotically active particles”: are these particles osmotically active when facing a water‐permeable membrane or facing a SP‐permeable membrane or both? Only the third definition still works when the membrane is SP‐permeable, since the moles of particles “that contribute to a solution’s osmotic pressure” are the moles of the impermeant SP regardless of whether m is water‐permeable or SP‐permeable. 3) To fix the problem with the term “osmotically active particles”, we replace it with a new term “osmotically effective SP,” defined as the physical quantity representing the amount of the impermeant (i.e., osmotic pressure (π)‐contributable) SP in the S compartment of S‐m‐H2O. 4) Osmosis does not take place in an isolated solution but in S‐m‐H2O. When the m is given, the total solute particles (TSP) of S are differentiated to be permeant SP measured using millimole (mmol) and impermeant SP measured using milliosmole (mOsm). Since the unit mOsm defined above excludes any permeant SP to a given m, the unit of osmolarity (i.e., mOsm/L) defines true osmolarity: the molar concentration of the impermeant SP in the S compartment of S‐m‐H2O.

Conclusion

Our definitions of osmole, osmotically effective SP, and true osmolarity are all given membrane‐dependent, system parameters of S‐m‐H2O, not owned by an isolated S. These definitions are essential but not enough to thoroughly resolve the many unaddressed problems with the current definitions of osmolarity. Our second abstract addresses these problems and offers resolutions.
Effect of Mesenchymal Stromal Cells (MSC) and Platelet‐Rich Plasma (PRP) on Rat Achilles Tendon Repair
Authors: Fagan Diana , Austin Brittany , Drew Stuart , Etty Thywill , Juzbasich Dragan , Pham Errek , Marie Hazel
Published: April 2020

Achilles tendon rupture requires long recovery times and has the possibility of re‐rupture. The goal of this project is to improve healing in surgically repaired Achilles tendons in a Lewis rat model. Full thickness surgical transections were made 6 mm proximal to the calcaneus bone. Following suture repair, a 20 x 30 mm piece of CollatapeTM was wrapped around the tendon in groups receiving collagen and the skin closed. MSC or PRP or both (100 ml) was injected subcutaneously adjacent to the incision. Both test Achilles and control Achilles from the same animal were harvested and the biomechanical properties determined using a standard force‐extension analysis of tensile properties after 1 week or 2 weeks. A significant improvement in Strain at UTS and Strain at Failure (calculated as percent of control tendon) was seen at 1 week in rats receiving CollatapeTM, plus PRP and MSC. This project was approved by the Northeast Ohio Medical School IACUC.

Support or Funding Information

Supported by Mercy Health, Youngstown and Youngstown State University.
Chronic Intermittent Hypoxia Alters the Chloride Gradient in Median Preoptic Nucleus (MnPO) Neurons of Rats
Authors: Farmer George E , Little Joel T , Bachelor Martha E. , Rybalchenko Nataliya , Yuan Joseph , Cunningham J. Thomas
Published: April 2020

Chronic intermittent hypoxia (CIH) is an animal model that simulates the hypoxemia associated with obstructive sleep apnea (OSA). Rats exposed to CIH exhibit increased sympathetic tone and reactivity and a persistent elevation in blood pressure during periods of normoxia, similar to patients with OSA. In MnPO neurons, angiotensin II type 1 receptor knock down (AT1aR KD) mediated reductions in GABAa inhibition are exacerbated to the point where GABAa activation is excitatory following CIH. Here, we use the genetically encoded ratiometric Cl‐sensor, ClopHensorN, to monitor the chloride flux of PVN projecting MnPO neurons in normoxic (Norm) and CIH treated rats in response to GABAa receptor activation.

Using isoflurane (2–3%) anesthesia, male Sprague‐Dawley rats (250–350g) received microinfusions (0.4 μL) of a retrograde AAV9‐Cre in the PVN and a DIO‐ClopHensorN in the MnPO. After recovery, rats were subjected to 7 days of CIH (0800–1600 hrs) or Normoxia. The CIH protocol consisted of 6 min cycles (3 min 21% O2, 3 min 10% O2) repeated 10x/hr for 8 hours (during the normal inactive/sleep phase) on 7 consecutive days. For ClopHensorN measurements, rats were anesthetized with isoflurane (2–3%) and coronal slices (300 μm) containing the MnPO were cut using standard in vitro slice procedures. Image sets were captured every 3 sec. Cl‐flux was determined from the ratio of GFP (ex 488 nm, em 500–550 nm) to tdTomato (ex 594 nm, em 650–700 nm) fluorescence in response to 10 s focal application of muscimol (100 uM).

Twelve rats (6 Norm, 6 CIH) were used for ClopHensorN studies. In MnPO neurons from CIH treated rats (n = 632), 20.1% showed a decrease in the ratio of GFP to tdTomato fluorescence (n = 127) while 0.3% showed an increase in this ratio (n = 2) indicative of Cl‐efflux. In MnPO neurons from Norm rats (n = 482), 41.9% showed a muscimol dependent decrease in the ratiometric Cl‐signal (n = 202) with 0 cells showing an increase. Furthermore, the magnitude if muscimol dependent decreases in the Cl‐signal were reduced in the CIH treated rats suggestive of reduced GABAa mediated inhibition. In a separate groups of rats, Western blot analysis showed that CIH increased the expression of pKCC2 (n = 6) in punches containing the MnPO compared to MnPO punches from Norm controls (n = 5) injected with Scr. CIH/AT1a KD rats showed decreased pKCC2 (n = 6) compared to CIH/Scr but increased pKCC2 in Norm/AT1a KD rats (n = 6) compared to Norm/Scr rats.

The results demonstrate that CIH alters the Cl‐flux of PVN projecting MnPO neurons and these changes are likely due to altered function of KCC2. These changes may contribute to the increased sympathetic tone and reactivity, as well as the persistent hypertension associated with CIH.

Support or Funding Information

Supported by P01 HL088052
Role of Thioredoxin‐interacting Protein in Diabetic Myocardial Ischemia‐Reperfusion Injury
Authors: Ye Xiaodong , Cai Yin , Zhang Dengwen , Wang Sheng , Liu Danyong , Irwin Michael Garnet , Xia Zhengyuan
Published: April 2020

Background

Myocardial ischemia‐reperfusion injury is a major threat to human health, causing great cardiac morbidity and mortality worldwide. Yet, it’s even more severe among diabetic patients. Thioredoxin‐interacting protein (TXNIP), a critical regulator with its inhibitory effect against antioxidation, is a novel target during myocardial ischemia‐reperfusion injury. TXNIP is also found to be highly expressed in hyperglycemic conditions, but its role in diabetic myocardial ischemia‐reperfusion injury is unclear. The current research aimed to study the role of TXNIP in myocardial ischemia‐reperfusion injury in diabetes mellitus.

Methods

Diabetes was induced by streptozotocin injection via tail vein (65mg/kg) in Sprague‐Dawley rats. In vivo model of myocardial ischemia‐reperfusion injury was established as 30 minutes occlusion of the left anterior descending coronary artery and then 2 hours of reperfusion. In vitro hypoxia/re‐oxygenation (H/R) model was established as 6 hours of hypoxia and 12 hours of re‐oxygenation of H9C2 cells. Also, glucose was added to the medium at a concentration of 33 mmol/L to mimic the condition of diabetes. TXNIP small interference RNA (siRNA) was introduced to knock down the expression of TXNIP.

Results

The expression of TXNIP significantly increased in the diabetic rat group, together with much greater infarct size and higher release of CK‐MB after myocardial ischemia‐reperfusion injury, compared to non‐diabetic group. In vitro study showed that TXNIP expression was significantly higher in high glucose (HG) group with greater cell injury and much higher LDH release after H/R treatment, compared to normal glucose (NG) group. However, TXNIP knock‐down via siRNA could alleviate this cellular injury with lower LDH release and higher cell viability. The expression of cleaved caspase‐3 significantly decreased even after H/R treatment in the TXNIP knock‐down group. Also, down‐regulation of NF‐kB and less phosphorylation of IkBα were detected after H/R in the TXNIP knock‐down group. Further study found higher conversion of LC3II from LC3I and lower expression of p62, indicating the activation of autophagy via TXNIP knock‐down. Finally, quantitative polymerase chain reaction test revealed significantly lower gene expression of TNF‐α and IL‐6.

Conclusion

TXNIP is highly expressed in diabetes after myocardial ischemia‐reperfusion injury. TXNIP knock‐down can alleviate myocardial ischemia‐reperfusion injury, possibly via activating autophagy and also via inhibiting inflammation. It is reasonable to believe that TXNIP would be a potential target for the treatment of diabetic myocardial ischemia‐reperfusion injury.

Support or Funding Information

Supported by RGC (17118619) and NSFC (81970247)
H2S Photosystems in the Treatment of Glioblastoma
Authors: Esposito Emanuela , Campolo Michela , Paterniti Irene , Lanza Marika , Filippone Alessia , Casili Giovanna , Conoci Sabrina , Cuzzocrea Salvatore
Published: April 2020

Glioblastoma multiforme (GBM) is the most frequent and deadly primary malignant central nervous system (CNS) tumour in adults. GBM is usually treated through the reduction of tumour mass and cerebral decompression, together with radiotherapy and chemotherapy, which improve the quality of life of the subject and prolong its survival. Treatment remains challenging, as GBM inevitably re‐emerges despite surgical debulking, radiotherapy and chemotherapy. Therefore, the discovery of effective therapy for GBM is an area of interest. Photothermal therapy (PTT) is a newly developed and encouraging therapeutic strategy, in treating cancer. Gold nanorods (AuNRs) are ideal nanomaterials for PTT applications both in vitro and in vivo. There is a reasonable body of evidence suggesting that endogenous H2S production is important for the growth and proliferation of colon and ovarian cancer. A limited number of studies are available in CNS cancer. Photoresponsive systems that delivers H2S on demand were used to treat glioblastoma to design and generate compounds based on photo‐thermal release of H2S that act as anti‐cancer therapy for glioblastoma. AuNRDs were prepared by a modified procedure based on the seed assisted method. These systems were tailored with a thiolate spacer appendages to permit the covalently link to the AuNRDs surface. The dual H2S photoactive‐systems were characterized by electronic microscopy methods, optical absorption, and emission techniques. Photocontrolled H2S‐based therapies were tested on GBM cell lines U87, A172, U138, LN‐18, T98G, U118‐MG. Measurement of H2S relase, Cell Cycle Analysis, Comet assay, determination of intracellular ROS, and apoptotic pathways were performed. Moreover, xenograft and orthotopic model were used to demonstrated the effect of dual H2S photoactive‐systems on tumour volume and weight, neoangiogenesis, tissue invasion. Animals were also subjected to magnetic resonance imaging. Photocontrolled H2S‐based therapies could emerge as new unconventional treatment modalities that complement better‐known photodynamic and photothermal therapies. Nanosystems carrying chemotherapeutic drugs or as mediators of alternative therapies offer several advantages over the standard dosing forms in the treatment of GBM.

Support or Funding Information

no funding
Exercise Decreases Estrogen Receptor β and Inflammatory Cytokines in the Mesenteric Fat of an Endometriosis Rat Model
Authors: Del Mar Méndez-Casillas María , Cruz Myrella L. , Velazquez-Cruz Johnathan , Chompre Gladys , Appleyard Caroline B.
Published: April 2020

Endometriosis, a gynecological disorder, is characterized by increased levels of the estrogen receptor β (ERβ) in endometrial tissue. Estrogen receptors (ER) are also present in visceral fat and dysregulation of their function is known to correlate with increased white adipose tissue. Preliminary data in our lab found increased mesenteric fat and serum leptin levels in endometriosis animals, which are decreased by exercise. In rodents, morphology of the mesenteric fat can be regulated by exercise, however it is still unclear what role exercise plays in this change and how it might impact inflammatory conditions such as endometriosis.

Hypothesis

Voluntary exercise can decrease endometriosis through modulation of ER in the mesenteric fat and a decrease in inflammatory cytokines.

Methods

Endometriosis (ENDO) was induced in female Sprague Dawley rats by the implantation of uterine tissue next to the intestinal mesentery while control group (SHAM) received only sutures. The exercised endometriosis group (ENDO‐Ex) had access to a running wheel before and after surgery. After sixty days, rats were sacrificed and mesenteric fat was collected for measurement of ERβ, ERα, interleukin‐1β (IL‐1β), and interleukin‐6 (IL‐6) by RT‐PCR. After fixation and staining with hematoxylin & eosin micro‐sections of mesenteric fat were analyzed to ascertain adipocyte morphology and their number and size.

Results

The Endo animals had significantly higher amounts of mesenteric fat in contrast with the SHAM (p<0.05) and the ENDO‐Ex animals (p<0.01; n=9–10/group). The serum leptin hormone levels were significantly decreased in ENDO‐Ex animals when compared with the SHAM (p<0.001) and ENDO (p<0.001) animals. Across all groups a significant correlation was found between percentage of fat/body weight and ER‐β (p=0.011; n=12). Increased mesenteric fat also correlated with increased levels of IL‐1β (p=0.024) and IL‐6 (p=0.030) in the endometriosis animals (n=4/group). ENDO‐Ex animals developed less endometriotic lesions and these were smaller in size. Exercise significantly decreased ERβ levels (p=0.045) and IL‐1β (p=0.008, n=4/group) compared with ENDO. No significant differences were found in ERα levels between groups.

Conclusion

Our results demonstrate that exercise can reverse the increased levels of mesenteric fat, ERβ and inflammatory cytokines found in an animal model of endometriosis.

Support or Funding Information

P20GM103475‐16, R15AT009915 & R25GM096955
Features of Stable Plaque Phenotype Are Increased by Macrophage Specific Insulin‐Like Growth Factor 1
Authors: Snarski Patricia , Danchuk Svitlana , Sukhanov Sergiy , Yoshida Tadashi , Higashi Yusuke , Shai Shaw-Yung , Chandrasekar Bysani , Delafontaine Patrice
Published: April 2020

Insulin‐like growth factor I (IGF‐1) reduces atherosclerosis in Apoe‐null mice and promotes stable plaque phenotype. We reported that deficiency for macrophage (MF)‐specific IGF‐1 receptor upregulated matrix metalloproteinases (MMPs) and promoted atherogenesis. To test whether MF‐specific IGF‐1 will suppress MMPs, decrease atherosclerosis, and change plaque stability, we generated Apoe‐null mice with IGF‐1 overexpression driven by MF‐specific promoter (scavenger receptor A) (SRA12 and SRA17 mice). IGF‐1 level was increased in peritoneal MF isolated from SRA12 (21±4% increase) and from SRA17 mice (52±9% increase) compared to control Apoe‐null mice (both P<0.05). SRA12 and SRA17 mice were fed with a high fat diet for 12 weeks. MF‐specific IGF‐1 overexpression significantly decreased atherosclerotic burden (SRA12, 23±3% decrease, n=20, and SRA17, 28±6% decrease, n=16, vs. control, n=29, en face analysis) and increased plaque collagen levels (11.1±6%, SRA12 and 66.5±11%, n=11, SRA17, Trichrome). Atherosclerotic plaques in both SRA12 and SRA17 mice had reduced necrotic cores suggesting elevated plaque stability. Increased levels of plaque collagen in SRA17 mice correlated with a significant decrease of MMP1, MMP2, MMP8, MMP12, MMP13, and MMP14 protein levels in peritoneal MF (immunoblotting, 30–72% decrease vs. control). To assess monocyte recruitment into the plaque, macrophage‐cleavable fluorescent red beads were injected (IV). Plaques in both SRA12 and SRA17 mice have reduced number of red beads compared to Apoe‐null mice (77±2% decrease, p=0.027, 74±3% decrease, p=0.0592, respectively) suggesting decreased monocyte recruitment. Cutlured human THP‐1 MFs were incubated with IGF‐1 (0, 20, 100ng/mL) for 48 hrs. IGF‐1 (20ng/ml) decreased expression of MMP8 (34±13% decrease, P<0.05) and MMP14 protein (35±17% decrease, P<0.05) (ELISA). To demonstrate that downregulation of MMPs was mediated via IGF1R‐dependent mechanism, we used Picropodophyllin (PPP), a specific IGF1R inhibitor. MF pre‐treatment with PPP (1ug/ml, 2hrs) abolished IGF‐1‐induced MMP8 downregulation in MF. In summary, we show that IGF‐1 reduced MMPs in vitro and MF IGF‐1 decreased MMPs, upregulated collagen and suppressed atherosclerotic burden in vivo. We identified the novel MMP‐dependent mechanism potentially mediating MF IGF‐1 atheroprotective effect by increasing features of stable plaque phenotype.

Support or Funding Information

NIH/NHLBI 2R01HL070241‐13A1
Effect of a Medical Device containing Xyloglucan and Pea Protein on a Murine Model of Atopical Dermatitis.
Authors: Campolo Michela , Paterniti Irene , Casili Giovanna , Filippone Alessia , lanza Marika , Ardizzone Alessio , Cuzzocrea Salvatore , Esposito Emanuela
Published: April 2020

Atopic dermatitis (AD) is a chronic inflammatory disease of the skin, characterized by intense itching and dry skin. The etiology of AD is complex and has not been fully clarified, involving genetic susceptibility, immunological dysfunction, epidermal barrier and environmental factors. Xyloglucan (XG) and pea protein (PP) are two compounds of natural origin characterized respectively by a physical barrier creation that protects mucosal tissues reducing inflammation. The aim of the present study was to evaluate the potential beneficial effects of XG + PP in a mouse models of AD and S. aureus infection‐ associated AD.

Mice were topically treated with 200 μl of 0.5% oxazolone on the dorsal skin three times a week for AD induction. Mice received XG and PP by topical administration, 1 hour before oxazolone treatment. In experiment of S. aureus infection‐ associated AD, to induce superficial superinfection of the skin, mice were also treated with 5 μl of 108 of a culture of S. aureus for 2 weeks; mice superinfected received XG and PP by topical administration 1 hour before oxazolone + S. aureus. Following 4 weeks, the skin was removed for histological and biochemical analysis. Our results clearly demonstrated the protective barrier effect of XG and PP characterized by a reduction in histological tissue changes induced‐AD, mastocyte degranulation and tight junction permeability in the skin following oxazolone treatment. Moreover, XG+PP was able to reduce filaggrin expression, a hallmark of AD. Our data also support the effectiveness of XG+PP to reduce the damage by superinfection post AD induced by s. aureus. In conclusion, the medical device containing XG and PP could be considered an ideal approach for the treatment of AD.