bluuebunny/crossref_metadata_2025_split huggingface dataset:
Scleraxis over‐expression promotes the cardiac myofibroblast phenotype (1152.13)
Authors: Roche Patricia , Bagchi Rushita , Aroutiounova Nina , Czubryt Michael 
Published: June 2021

Myofibroblasts are the primary mediators of cardiac remodeling, and their persistence in the myocardium is implicated in the development of fibrosis. Thus, inhibiting myofibroblast differentiation represents a novel approach for treating cardiac fibrosis. Scleraxis is a transcription factor required for development of collagen rich tissues. Interestingly, the same stimuli that induce differentiation also cause increased scleraxis levels in vitro and in vivo. Thus, we hypothesized that scleraxis promotes the myofibroblast phenotype. To test this, we over‐expressed scleraxis via adenovirus in primary cardiac fibroblasts (CFs) and measured changes in key myofibroblast markers. We observed that scleraxis over‐expression in CFs caused increased mRNA and protein levels of these markers. Since myofibroblasts are also distinguished from fibroblasts by their contractility, we used a contraction assay to determine the functional implications of increased marker levels. We found that over‐expressing scleraxis in CFs caused a significant increase in contraction. However, over‐expression of a dominant negative DNA binding‐deficient scleraxis mutant did not induce contraction, indicating a requirement for intact scleraxis. These results demonstrate that scleraxis promotes the cardiac myofibroblast phenotype, implicating scleraxis as a potential target for the intervention of cardiac fibrosis.Grant Funding Source: Supported by Canadian Institute of Healthy Research, St. Boniface Hospital Foundation
Hyperglycemia suppresses cardiomyocyte bioenergetic reserve independent of O‐GlcNAcylation (1155.5)
Authors: Dassanayaka Sujith , Readnower Ryan , Long Bethany , Aird Allison , Salabei Joshua , Hill Bradford , Jones Steven 
Published: June 2021

Hyperglycemia, a hallmark of diabetes, can augment flux through accessory glucose metabolism pathways, such as the hexosamine biosynthetic pathway, which produces the sugar donor for the β‐O‐linked‐N‐acetylglucosamine (O‐GlcNAc) post‐translational modification of proteins. We examined whether hyperglycemia‐induced elevations in O‐GlcNAcylation of cultured neonatal rat cardiomyocytes (NRCMs) affected mitochondrial function using extracellular flux analysis. Hyperglycemia (33 mmol/L) augmented O‐GlcNAcylation and suppressed mitochondrial reserve capacity (p<0.05); however, an osmotic control recapitulated the effect of hyperglycemia‐suppressed reserve capacity (p<0.05) without increasing O‐GlcNAcylation. Furthermore, neither adenoviral overexpression nor pharmacologic inhibition of either OGA or OGT altered mitochondrial reserve capacity. Specific interrogation of individual electron transport Complex II function showed that hyperglycemia reduced Complex II mediated oxygen consumption rate (OCR), which, again was recapitulated by an osmotic control. Interestingly, pharmacologic augmentation of O‐GlcNAcylation increased Complex II OCR (p<0.05). Thus, hyperglycemia may perturb mitochondrial function independent of O‐GlcNAcylation, thereby questioning the purported causal relationship between O‐GlcNAcylation and diabetic cardiomyocyte dysfunction.Grant Funding Source: Supported by NIH and AHA
Limb specific comparison of flow‐mediated dilation in overweight, pre‐menopausal women (1158.1)
Authors: Barlow Matthew , Restaino Robert 
Published: June 2021

The dilatory responses of the brachial artery as compared to the popliteal artery have been found to be equal in healthy young women and reduced in normal aging in healthy, post‐menopausal women (Parker et al. 2006). However, limb‐specific differences have not been investigated among young overweight, premenopausal women who are at a greater propensity for advanced cardiovascular aging. The hypothesis is that overweight women will exhibit limb specific differences in artery dilation with reduced dilation in the popliteal artery as compared to the brachial artery. Artery dilatory responses were assessed using flow‐mediated dilation (FMD) in 18 women (9 controls, 9 overweight) aged 24.7+1.29, non‐smokers, and sedentary. Significant difference were found between groups including % body fat, BMI, weight, waist to hip ratio, fasting blood glucose and oxLDL. A between group comparison of percent dilation revealed that the popliteal artery in overweight participants were significantly lower than control (p<0.05). We conclude that young, metabolically compromised women exhibit a limb specific difference in dilation of the upper and lower extremities as compared to healthy controls, showing possible complications of oxidative stress within the vasculature.Grant Funding Source: HSI STEM – Engaging Students in STEM Careers Award Number: P031C110114
The impact of aging on mitochondrial proton leaks in human skeletal muscle (1159.9)
Authors: Porter Craig , Hurren Nicholas , Sidossis Labros , Børsheim Elisabet 
Published: June 2021

Proton leaks are thought to account for 20‐25% of mitochondrial oxygen consumption. It has been postulated that mitochondrial proton cycling in senescent tissue may reduce free radical production. Here, we determined mitochondrial respiratory control in skeletal muscle biopsies obtained from young and older individuals. 6 young (31±3 yrs) and 10 older (58±7 yrs) subjects participated in this study. Mitochondrial respiration was determined in permeabilized myofibers after the addition of substrates, ADP and oligomycin. Thereafter, coupling control ratios were calculated. Coupled mitochondrial respiration tended to be higher in muscle from young vs. older subjects (38.0±3.1 vs. 31.7±2.8 pmol/mg/sec, P=0.16). ADP sensitivity was similar between groups, whereas oligomycin sensitivity was significantly lower in the older vs. young group (P<0.05). Mitochondrial ADP sensitivity is preserved in skeletal muscle of older individuals. However, mitochondria of older individuals are less sensitive to the ATP synthase inhibitor oligomycin. These data suggest that inner membrane proton conductance is elevated in skeletal muscle mitochondria with aging.Grant Funding Source: Supported by NIH (1R01 AG033761, P30 AG024832, 1 U54RR026141) and NIDRR (H133P110012) grants.
GQ‐16, a novel partial PPARγ ligand, decreases visceral adiposity and induces the expression of thermogenesis‐related genes in mice with obesity and insulin resistance induced by diet (1160.8)
Authors: Coelho Michella , Lima Caroline , Royer Carine , Pitta Ivan , Amato Angélica , Neves Francisco 
Published: June 2021

Thiazolidinediones are used for the treatment of type 2 diabetes (T2DM) and their therapeutic actions are mediated via activation of PPARγ. Despite their efficacy, their use is limited by side effects such as weight gain. We have previously identified a novel PPARγ ligand with similar anti‐diabetic efficacy as rosiglitazone (RSG), but without weight gain.Objective: To evaluate the effects of different doses of GQ‐16 on adiposity and gene expression in mice with insulin resistance and obesity induced by diet.Methods: Mice were fed normal‐fat (NFD) or high‐fat (HFD) diet since weaning. At the age of 16 wk, they received GQ‐16 (5, 10 or 20mg/kg/d), RSG (4mg/kg/d) or vehicle by gavage. White adipose tissue (WAT) pads were excised for determination of adiposity and gene expression by qPCR.Results (P<0.05, 4 mice/group): In HFD‐treated mice, weight gain was reduced by treatment with 5, 10 and 20 mg/kg/d of GQ‐16, but increased by RSG. GQ‐16 (10 and 20 mg/kg/d) decreased blood glucose similarly to RSG. In addition, GQ‐16 treatment (20 mg/kg/d) reduced WAT pads, whereas RSG treatment increased them. Brown fat‐selective genes were upregulated in WAT in response to GQ‐16.Conclusions: Lower doses of GQ‐16 decreased blood glucose and visceral adiposity. This latter finding might possibly reflect increased thermogenesis, since GQ‐16 induced the expression of thermogenesis‐related genes, and reinforces the potential of GQ‐16 as a new strategy to treat obese patients with T2DM.Grant Funding Source: CNPq
Regulation of leucyl‐tRNA synthetase and RagB expression in human skeletal muscle by essential amino acids (1161.4)
Authors: Carlin Matthew , Tanner Ruth , Agergaard Jakob , Jalili Thunder , Askew E , McClain Donald , Supiano Mark , Marcus Robin , LaStayo Paul , Drummond Micah 
Published: June 2021

Essential amino acids (EAA) are a potent stimulator of mammalian target of rapamycin (mTORC1) signaling and protein synthesis. However, the molecular regulators upstream of mTORC1 signaling that are stimulated by EAA are not well described especially in human skeletal muscle. Leucyl tRNA Synthetase (LRS) and RagB have recently been shown to be important in mTORC1 activation. The purpose of this study was to determine if the LRS‐ and RagB‐mTORC1 complexes and LRS and RagB gene and protein expression are altered following acute EAA ingestion in healthy human skeletal muscle. Muscle biopsies (vastus lateralis) were obtained from 8 young adults (4M, 4F) in the fasted state (basal) and 1 and 3h after EAA (13g; 2.4g Leu) ingestion. Co‐immunoprecipates were used to determine LRS‐ and RagB‐mTORC1 protein complexes and whole muscle homogenates were analyzed for LRS and RagB gene and protein expression and mTOR phosphorylation. We found that EAA increased RagB protein abundance and mTOR phosphorylation 1h following EAA ingestion (P<0.05). However EAA ingestion did not alter LRS‐ or RagB‐mTORC1 complexes, LRS protein expression or change LRS and RagB gene expression 1 or 3h‐post EAA. We conclude that LRS‐ and RagB‐mTORC1 complexes are not altered 1 and 3h‐post EAA. However, RagB protein increased after EAA ingestion in healthy human skeletal muscle, which may be a mechanism to promote protein anabolism.Grant Funding Source: Supported by K01 AG038556 and UL1 TR000105
PPARβ/δ is not required by PGC‐1α to enhance skeletal muscle oxidative metabolism (1164.3)
Authors: Pérez‐Schindler Joaquín , Vargas‐Fernández Elyzabeth , Santos Gesa , Svensson Kristoffer , Wahli Walter , Handschin Christoph 
Published: June 2021

Impaired skeletal muscle metabolism is a hallmark of numerous, chronic metabolic diseases. Exercise training efficiently improves skeletal muscle metabolic fitness, increases mitochondrial biogenesis and exerts beneficial therapeutic effects. Aberrant expression of metabolic‐related genes in skeletal muscle is thought to contribute to the development and progression of such pathologies. The peroxisome proliferator‐activated receptor γ (PPARγ) coactivator‐1α (PGC‐1α) and the nuclear receptor PPARβ/δ have both been independently shown to increase skeletal muscle oxidative capacity and exercise performance. Therefore, current understanding is that the PGC‐1α‐PPARβ/δ axis plays a pivotal role in the regulation of skeletal muscle metabolism. However, the interplay and hierarchical regulation of these two proteins remains unclear, with limited research conducted in vivo. In the present study, we demonstrate that the disruption of the PGC‐1α‐PPARβ/δ axis in mouse skeletal muscle does not affect body composition, whole body metabolism and glucose homeostasis under basal conditions. Importantly, PPARβ/δ seems dispensable for most of the effects of PGC‐1α on VO2 and lactate metabolism during exercise as well as the enhancement of skeletal muscle oxidative metabolism. Therefore, these results indicate that PPARβ/δ is not an essential partner of PGC‐1α in the control of skeletal muscle remodeling.Grant Funding Source: Supported by the Swiss National Science Foundation
Aging alters muscle reflex control of autonomic cardiovascular responses to rhythmic contractions in humans (1165.8)
Authors: Sidhu Simranjit , Weavil Joshua , Venturelli Massimo , Rossman Matthew , Gmelch Benjamin , Bledsoe Amber , Jessop Jacob , Richardson Russell , Amann Markus 
Published: June 2021

We investigated the influence of aging on the role of group III/IV muscle afferents in determining circulatory responses to physical activity. Six old (OLD; 70 ± 2 years) and 8 young (YNG; 24 ± 2 years) males performed single‐leg knee extensor exercise (14 W / 30 W / 80% max) under control conditions and with lumbar intrathecal fentanyl impairing feedback from µ‐opioid receptor‐sensitive lower limb muscle afferents. Cardiac output (Q), leg perfusion pressure (PP), leg vascular conductance (LVC), and femoral blood flow (QL) were collected continuously. There were no hemodynamic differences between conditions at rest. Fentanyl blockade during exercise did not affect Q in OLD (P=0.2); however, secondary to significant decreases in heart rate and stroke volume, Q was attenuated by ~20% in YNG (P<0.05). Furthermore, in the face of a similar 5‐7% reduction in PP (P<0.05) during exercise with fentanyl blockade in both groups, LVC increased ~13% in OLD, but decreased ~10% in YNG (both P<0.05). The net effect of these blockade‐induced changes was an unaltered QL response to exercise in OLD, contrasting the significant 15‐17% decrease in YNG. In conclusion, healthy aging alters muscle reflex control of autonomic circulatory responses to exercise. While muscle afferent feedback remains critical in regulating PP, their contribution to central (Q) and peripheral (QL) hemodynamic responses might diminish with age.Grant Funding Source: HL_103786; HL‐116579
The C‐terminal tail of aquaporin 1 modulates β‐catenin expression in pulmonary arterial smooth muscle cells (1175.2)
Authors: Yun Xin , Jiang Haiyang , Lai Ning , Shimoda Larissa 
Published: June 2021

PASMC proliferation and migration are important contributors to the vascular remodeling that occurs during hypoxic pulmonary hypertension. Hypoxia increases the expression of AQP1, a member of the water channel family of proteins, in rat PASMCs and we previously showed that AQP1 controls proliferation and migration in PASMCs via a mechanism that includes the C‐terminal tail but is independent of water transport. β‐catenin, part of the Wnt signaling pathway, activates the transcription of crucial target genes controlling proliferation. In melanoma and endothelial cells, Lin‐7, a cell junction protein, interacts with AQP1 to regulate β‐catenin levels. Thus, we explored whether β‐catenin, Lin‐7 and AQP1 interact in PASMCs and, if so, whether the C‐terminal tail of AQP1 was required. Rat PASMCs were harvested, plated onto culture dishes and infected with adenoviral constructs expressing green fluorescent protein (AdGFP), the control for infection, wild‐type AQP1 (AdAQP1) or AQP1 lacking the cytoplasmic C‐terminal tail (AdAQP1CT). Compared to AdGFP cells, infection with AdAQP1, but not AdAQP1CT, increased PASMC migration and proliferation and β‐catenin protein expression. Interestingly, immunoblot analysis revealed that Lin‐7 was not expressed in rat PASMCs. Our results suggest that increased AQP1 levels upregulate β‐catenin protein levels via a mechanism requiring the AQP1 C‐terminal tail but independent of Lin‐7.Grant Funding Source: AHA0755479
Mitochondrial bioenergetics and permeability response in rat pulmonary microvascular endothelial cells in culture (1176.1)
Authors: Merker Marilyn , Townsley Mary , Bongard Robert 
Published: June 2021

Hyperoxia causes lung endothelial injury and depresses mitochondrial complex I (CI) activity. In isolated rat lung perfused with a physiological saline solution (5.5 mM glucose), CI inhibitors decrease lung tissue ATP and increase endothelial permeability (measured as Kf), effects that are overcome using a quinone (CoQ1) (Free Radic Biol Med. doi: 10.1016/j.freeradbiomed.2013.07.040). To unmask the mitochondrial contribution to pulmonary endothelial cell culture permeability responses, PMVEC were treated with the CI inhibitor rotenone (0.5 µM) in 0.2 mM glucose without or with CoQ1 (10 µM). In PMVEC monolayers, rotenone decreased ATP from 18.4 ± 1.6 (mean ± SE) to 4.6 ± 0.8 nmol/mg protein, decreased O2 consumption from 2.6 ± 0.1 to 0.3 ± 0.1 nmol/min/mg protein and increased permeability to FITC‐dextran from 8.3 ± 0.7 to 14.3 ± 0.4 nl/min/cm2. When CoQ1 was present with rotenone, ATP (17.4 ± 1.4 nmol/mg protein), O2 consumption (1.6 ± 0.2 nmol/min/mg protein) and permeability (10.1 ± 1.2 nl /min/cm2) were maintained. CoQ1 also prevented rotenone‐induced mitochondrial depolarization. Protective effects of CoQ1 on ATP, O2 consumption and mitochondrial membrane potential were blocked by inhibitors of the quinone mediated cytosol‐mitochondria electron shuttle. We conclude that, as in the lung, mitochondrial bioenergetics can be a determinant of the PMVEC permeability response.Grant Funding Source: Supported by Dept of Veterans Affairs
Whole body plethysmography measurement of respiratory function of mice in vivo (1178.9)
Authors: Quindry John , Ballmann Christopher , Selsby Joshua 
Published: June 2021

Applied physiology research commonly incorporates mouse models, but in vivo physiologic measures in conscious rodents can be difficult to collect. Since recommendations for standardized whole body plethysmography (WBP) use do not currently exist, the current study employed WBP to quantify pulmonary function in conscious mice. Two month old Male C57BL6 and MDX mice (n=8/group) were housed on a reverse light cycle and examined on consecutive days using a calibrated WBP (Fine Pointe unrestrained WBP, Buxco Electronics). Multiple measurements such as time of day (8 am, 12 pm, 3 pm, 6:30 pm), environment lighting influence, and day‐to‐day reproducibility as well as chamber habituation time required for stable data collection plateau were recorded. Fine Pointe software was used to determine respiratory rate, tidal volume, minute ventilation, peak inspiratory flow, peak expiratory flow, inspiratory time, expiratory time, and relaxation time between breaths. Statistical observance of respiratory chamber habituation occurred between 5‐10 mins from the start of collection for all variables measured. Reverse light cycle housed mice exhibited significant time of day pulmonary function differences, performing more forceful and less frequent breaths during the middle of the day (12 pm & 3 pm). Significant differences were observed for WBP performed in lighted vs. darkened conditions, with the latter eliciting more forceful breathing. Day‐to‐day reproducibility was significant (p<0.05) with a range of from r2=0.58 to r2=0.62 for measures collected. Findings indicate good reproducibility of WBP is possible provided investigators collect data after at least a 10 minute habituation period and standardize time of day and room lighting used for data collection. This work supported by the Duchenne Alliance and its member foundations.Grant Funding Source: Duchenne Alliance and member foundations
Increased differentiating kidney epithelial cell density induced by changes in fluid flow velocity (1180.12)
Authors: Seetharaman Vikram , Resnick Andrew 
Published: June 2021

Mouse cortical collecting duct cells were allowed to differentiate under conditions of no fluid flow (vf = 0; negative control) or constant rapid flow (vf = max; positive control) on orbital shakers. Cells were subjected to Δvf at 3, 5, 10, 20 days post‐differentiation, from vf = 0 to vf = max, or vice versa. At various times post‐Δvf, cells were harvested, nuclei were DAPI stained for cell counting, and RNA/protein extracted. Some cells were fixed and immunostained for the presence of the markers of MCCD cell differentiation E‐cadherin (E‐cad), important in adherens junction formation, and polycystin 1 (PC1), important in cellular mechanosensation. If MCCD’s exposed to Δvf do indeed de‐differentiate, then the relative expression of these 2 proteins should decrease. Although PC1 was absent from all cells sampled before 10 days post‐differentiation and unchanged in cells exposed to 5 days of vf = max followed by 5 days of vf = 0 compared to cell exposed to 10 days of vf = max only, E‐cad expression appeared slightly lower in cells exposed to 5 days of vf = max followed by 4 hours of vf = 0 compared to cells exposed to both 5 days of vf = 0 only and 5 days of vf = max only. Quantitative real time‐polymerase chain reaction and Western blot analyses will subsequently be used to determine concentrations of several mRNA transcripts [cyclin dependent kinase 1 (cdk1)/c‐Myc/cyclin D] and phosphorylated proteins [phospho‐Protein Kinase B (PKB, also known as Akt)/phospho‐mammalian Target of Rapamycin (mTOR)/phospho‐Mitogen Activated Protein Kinase (MAPK, also known as ERK1&amp;2)], respectively, considered markers of cell proliferation. If MCCD’s exposed to Δvf do indeed divide, then all of these mRNA transcripts and phospho‐proteins should increase in concentration except for phospho‐PKB, which should decrease in concentration.Grant Funding Source: Supported by the National Institute of Diabetes and Digestive and Kidney Diseases 1R15DK092716
Hydrogen sulfide activates heterologously expressed human CFTR in Xenopus oocytes (1181.16)
Authors: Althaus Mike , Perniss Alexander , Beuter Liesa 
Published: June 2021

Hydrogen sulfide (H2S) has recently been recognized as a cellular signaling molecule. Physiological responses to H2S are often attributed to its impact on various sodium, calcium and potassium channels. However, little is known about the influence of H2S on chloride channels. This study investigated the putative regulation of the chloride channel CFTR (cystic fibrosis transmembrane conductance regulator) by H2S.Human CFTR was heterologously expressed in Xenopus oocytes and its activity was measured by microelectrode recordings using the two‐electrode voltage‐clamp technique. CFTR expression was verified by the application of forskolin/IBMX which elicited a transmembrane current that was sensitive to the CFTR inhibitor CFTRinh‐172. The H2S‐forming salt Na2S (5‐300 µM) significantly increased transmembrane currents of CFTR‐expressing oocytes but had no effect on native oocytes. The effect of Na2S was dose dependent and blocked by CFTRinh‐172. The slow H2S‐releasing molecule GYY4137 also activated CFTR, although to a smaller extent compared to Na2S. The adenylate cyclase inhibitor MDL‐12330A significantly decreased the activation of CFTR by Na2S. These data indicate that H2S activates CFTR in Xenopus oocytes by the cAMP/adenylate cyclase pathway. In epithelia, an increased CFTR activity may correspond to a secretory response to H2S which may be endogenously produced by the epithelium or a H2S‐generating microflora.Grant Funding Source: Supported by the German Research Foundation (AL‐1453/1‐2).
An intrinsically disordered region of the transcription factor, NFAT5, becomes more ordered with an increase in osmolality (1182.8)
Authors: DuMond Jenna , Kumar Raj , Ramkissoon Kevin , Izumi Yuichiro , Thompson Brad , Burg Maurice , Ferraris Joan 
Published: June 2021

Hypertonicity stimulates NFAT5 nuclear localization and transactivating activity. Many transcription factors are known to contain intrinsically disordered regions (IDRs) which become more structured with local environmental changes such as osmolality, temperature and tonicity. The N‐terminal region of NFAT5 is predicted to contain an IDR. Using far‐UV CD and fluorescence we found that addition of the organic osmolyte trehalose to the purified N‐terminal IDR of NFAT5 caused an increase in the structure of the IDR, consistent with a 2‐state transition, ID to native. To investigate how this increased structure affects cellular protein‐protein interactions, we looked for a correlation between change in structure and association with other proteins. Mass spectrometry identified NFAT5 IDR protein binding partners. We thus have demonstrated that a change in osmolality gives rise to an increase in IDR structure and affects NFAT5 protein‐protein interactions. These data will aid in understanding transcriptional upregulation of NFAT5 under cellular osmotic stress.Grant Funding Source: Supported by SBC, NHLBI, National Institutes of Health, Intramural Research Program