bluuebunny/crossref_metadata_2025_split huggingface dataset:
Renin Overexpression Leads to Increased Titin‐Based Stiffness Contributing to Diastolic Dysfunction in Hypertensive mRen2 Rats
Authors: Csipo Tamas , Kovacs Arpad , Fulop Gabor Aron , Kovacs Andrea , Bodi Beata , Juhasz Bela , Priksz Daniel , Granzier Henk , Edes Istvan , Papp Zoltan , Barta Judit , Toth Attila 
Published: November 2023

Hypertension is a major risk factor for heart failure (HF). We investigated the influence of hypertension on cardiac contraction and relaxation in transgenic renin overexpressing rats (carrying mouse Ren‐2 renin gene, mRen2, n=6). Blood pressure (BP) was measured by tail‐cuff method, echocardiography was used to characterize cardiac contractility, cellular force measurements and biochemical assays were applied to reveal molecular mechanisms. Sprague‐Dawley (SD) rats (n=6) were used as control. Systolic BP was elevated in mRen2 rats (235.11±5.32 vs. 127.03±7.56 mmHg in SD, P<0.05), resulting in increased left ventricular (LV) weight/body weight ratio (4.05±0.09 vs. 2.77±0.08 mg/g in SD, P<0.05). No effects of transgenic renin expression on systolic parameters, such as LV ejection fraction (EF, 68.50±2.09 vs. 70.17±1.45 % in SD), maximal Ca2+‐activated force (25.89±0.99 vs. 29.05±2.80 kN/m2 in SD) and Ca2+‐sensitivity of force production (pCa50, 5.83±0.01 vs. 5.87±0.05 in SD) were found. In contrast, pronounced diastolic dysfunction (DD) was observed in mRen2 rats: ratio of peak early and late LV diastolic filling (E/A) was lower (1.14±0.04 vs. 1.87±0.08, P<0.05), LV isovolumetric relaxation time was longer (43.85±0.89 vs. 28.55±1.33 ms, P<0.05) and cardiomyocyte passive stiffness was higher (1.74±0.06 vs. 1.28±0.18 kN/m2, P<0.05), compared to SD, respectively. Finally, hyperphosphorylation of titin at Ser‐12742 within the PEVK domain (P<0.05) proposed a molecular mechanism for DD. Our data suggest a link between the activation of renin‐angiotensin‐aldosterone system and increased titin‐based stiffness through phosphorylation of titin's PEVK element, contributing to DD. Accordingly, mRen2 rat might provide an in vivo model for HF with preserved EF (HFpEF).Support or Funding InformationThis work was supported by a European Union FP7 grant (MEDIA: metabolic road to diastolic heart failure) and by the Hungarian Scientific Research Fund (K116940 and K109083).
Transient Receptor Potential Melastatin 4 Channel Regulation by Inositol Trisphosphate Receptor in Human Urinary Bladder Smooth Muscle
Authors: Provence Aaron , Hristov Kiril L. , Rovner Eric S. , Petkov Georgi V. 
Published: November 2023

Overactive bladder (OAB) is a highly prevalent and debilitating condition that lacks an effective therapeutic treatment. The contraction and relaxation of urinary bladder smooth muscle (UBSM) facilitates the voiding and storage of urine, the two main functions of the urinary bladder. Some forms of OAB have been directly linked to UBSM dysfunction. Thus, efforts to uncover unknown regulatory mechanisms in UBSM function are urgently needed. Among the potential novel targets for OAB, emerging evidence points to key roles for transient receptor potential melastatin 4 (TRPM4) channels in UBSM excitability and contractility. However, the cellular mechanisms regulating TRPM4 channel activity in human UBSM has been virtually unexplored. As a potential cellular mechanism regulating TRPM4 channel activity in UBSM, we have directed our attention to the inositol trisphosphate (IP3) receptors (IP3Rs), which facilitate the release of Ca2+ from the sarcoplasmic reticulum (SR). Since the TRPM4 channels are activated by Ca2+, IP3R‐mediated Ca2+ release of the SR represents a potential Ca2+ source for TRPM4 channel activation. In accordance with the approved IRB Pro00045232 of the Medical University of South Carolina, we used clinically‐characterized human UBSM tissues from 6 donor patients to investigate the molecular and functional interactions of the TRPM4 channels and IP3Rs in human UBSM. Using in situ proximity ligation assay (PLA) and amphotericin‐B perforated patch‐clamp electrophysiology, we sought to test the hypothesis that TRPM4 channels are tightly associated with the IP3Rs and are activated by IP3R‐mediated Ca2+ release of the SR in human UBSM. As determined by in situ PLA, we demonstrated colocalization of the TRPM4 channels and IP3Rs in human UBSM cells. As the TRPM4 channels and IP3Rs must be located within close apposition to functionally interact, these findings support the concept of a TRPM4‐IP3R regulatory mechanism in human UBSM function. To investigate IP3R regulation of TRPM4 channel activity, we sought to determine the consequences of IP3R pharmacological inhibition on TRPM4 channel‐mediated transient inward cation currents (TICCs). In freshly‐isolated human UBSM cells, the selective IP3R inhibitor xestospongin C significantly decreased TICCs. These findings suggest that the SR IP3Rs have a key role in mediating the Ca2+‐dependent activation of TRPM4 channels in human UBSM. In conclusion, this study provides novel insight into the cellular mechanisms regulating TRPM4 channel activity in human UBSM function. Our study reveals that the TRPM4 channels and IP3Rs are spatially and functionally coupled in human UBSM, information that is critical for further evaluating the potential role of the TRPM4 channels as novel therapeutic targets for OAB.Support or Funding InformationSupported by NIH grant R01‐DK106964 to Georgi V. Petkov and F31‐DK104528 to Aaron Provence.
Chronic Binge Alcohol Administration (CBA) Impairs Skeletal Muscle Mitochondrial Gene Expression in Simian Immunodeficiency Virus (SIV)‐Infected Rhesus Macaques
Authors: Duplanty Anthony A , Song Kejing , Simon Liz , Molina Patricia E 
Published: November 2023

Previous studies from our laboratory have shown that CBA accentuates muscle wasting at end‐stage SIV infection and this is associated with pro‐oxidative and pro‐inflammatory skeletal muscle milieu as well as dysregulation of myoblast myogenic gene expression and myotube formation. Additionally, muscle of CBA/SIV macaques showed a greater reduction in the number and oxidative capacity (succinate dehydrogenase activity) of type 1 muscle fibers compared to that of sucrose administered SIV‐infected (SUC/SIV) and uninfected controls, Chronic alcohol consumption has been reported to result in mitochondrial DNA damage and impaired biogenesis; and this is considered an important mechanism in the pathogenesis of muscle myopathy. We hypothesized that CBA would impair the expression of genes involved in mitochondrial function in SIV‐infected macaques.PurposeThe purpose of this study was to investigate the effects of CBA on expression of selected mitochondrial genes in the skeletal muscle of SIV‐infected rhesus macaques at end‐stage disease.MethodsMale rhesus macaques were administered daily CBA (blood alcohol levels ~50 mM) or sucrose (SUC) intragastrically 3 months prior to intravenous SIVmac251 inoculation and continued throughout the study period. Skeletal muscle (gastrocnemius) samples were obtained at necropsy when animals met criteria for euthanasia and total RNA was extracted and mitochondrial gene expression was analyzed by qPCR.ResultsThe relative expression of peroxisome proliferator‐activated receptor gamma, coactivator 1 beta (PGC‐1 beta) and mitofusin‐1 (MFN1) was significantly decreased in the skeletal muscle of CBA/SIV macaques (75% and 39% respectively; P<0.05), without altering the gene expression of MFN2, TFAM, NRF1, NRF2, or PGC‐1 alpha.ConclusionThese results indicate that CBA administration decreases expression of genes essential for mitochondrial biogenesis in the skeletal muscle of SIV‐infected rhesus macaques and are in agreement with our previous observations of decreased oxidative capacity of type 1 muscle fibers. We speculate that impaired mitochondrial biogenesis may contribute to the underlying pathophysiology of alcoholic and HIV/AIDS associated myopathy. Developing therapeutic or life style interventions aimed at attenuating mitochondrial dysregulation in persons living with HIV/AIDS with alcohol use disorders should lead to improved muscle function and strength.Support or Funding InformationThis study was supported by grants from the National Institute on Alcohol Abuse and Alcoholism (NIAAA): T32 Postdoctoral Fellowship (AA07577) and the Comprehensive Alcohol Research Center P60 (AA09803) in the department of Physiology, LSU Heath Science Center, New Orleans, LA.
ErbB3 negatively regulates homeostatic Paneth cell numbers
Authors: Almohazey Dana , Hsieh Jonathan J , Vossler Claire V , Frey Mark R 
Published: November 2023

Paneth cells (PCs) are located at the base of the intestinal crypt, where they support the intestinal stem cells (ISC) with growth factors and participate in innate immunity by secreting antimicrobial peptides (AMPs). Loss of or functional defects in PCs are seen in Crohn's disease, linking abnormalities in these cells to inflammatory bowel disease (IBD). We recently showed that activation of the neuregulin receptor ErbB4 protects PCs against injury. However, the role of ErbB3, the other neuregulin receptor, in PCs is unknown. In this study we tested the effects of ErbB3 activation or deletion on PC numbers and AMP production.METHODSIleal tissue and enteroid cultures derived from C57BL/6 (WT), ErbB4flox/flox, ErbB4flox/flox; Villin‐Cre (E4KOIE), ErbB3flox/flox, and ErbB3flox/flox; Villin‐Cre (E3KOIE) mice were used. Enteroids were given NRG1β (ErbB3/4 ligand; 10ng/ml) or NRG4 (ErbB4 ligand; 100ng/ml) for 24 h and RNA was extracted. Fixed tissue for immunofluorescence analysis, RNA for qPCR, and protein lysates for western blotting were collected from mice.RESULTSNRG1β treatment of WT enteroids for 24 h resulted in reduced expression of Lyz1 (by 29.47%, p=0.03) and Defa5 (by 46.60%, p= 0.001), and fewer lysozyme‐positive cells (by 18.03%, p=0.03). This effect of NRG1β was maintained in E4KOIE enteroids but was not recapitulated in NRG4‐treated WT enteroids, suggesting a specific role for ErbB3. Consistently, E3KOIE intestinal crypts displayed an increased number of lysozyme‐positive cells (by 25.36%, p=0.001) and MMP7/MUC2 double positive intermediate/precursor cells (by 16.24%, p=0.011). E3KOIE crypts also had increased Lyz1 (by 67.53%, p<0.0001) and Lgr5 (by 46.65%, p=0.012) RNA expression. This increase was not accompanied by differences in expression of other ErbB family members (Egfr, ErbB2, and ErbB4) or other secretory cell markers (Muc2 and Chga). Interestingly, E3KOIE mice had a significant increase in expression of the secretory regulator Atoh1 (by 59.20%, p=0.0009), which is required for PC development. Since MAPK signaling has been implicated in PC regulation, we performed western blot analysis for P‐Erk1/2 and P‐p38 on tissue lysates, but there was no difference between WT and E3KOIE mice. In contrast, E3KOIE mice had a lower level of P‐Akt (by 53.1%, p=0.02), suggesting potential involvement of the PI3K pathway.CONCLUSIONSThese results suggest that ErbB3 restricts PC numbers by a mechanism that may involve PI3K/Akt and Atoh1. Understanding the role of ErbB3 receptor could identify new therapeutic targets for regulating PCs and the ISC niche.Support or Funding InformationThis work was supported by a Senior Research Award from the Crohn's and Colitis Foundation of America and by NIH award R01DK095004.
Metformin enhances intestinal epithelial health through activation of AMPK
Authors: Xue Yansong , Sun Xiaofei , Zhu Mei‐Jun 
Published: November 2023

The intestinal epithelium is a vital barrier that is consistently renewed. The balance among proliferation, migration, and differentiation are tightly regulated, which is critical for intestinal epithelial homeostasis. The objective of this study was to evaluate the beneficial role of metformin, a widely used antidiabetic drug, on epithelial health using interleukin‐10 deficient (IL10KO) mice. Metformin supplementation increased the abundance of E‐cadherin, a core component of the adhesion junctions, in both WT and IL10KO mice. In addition, metformin supplementation suppressed macrophage pro‐inflammatory activity in IL10KO mice as indicated by reduced M1 macrophage abundance as well as decreased pro‐inflammatory cytokine IL‐1b, TNF‐a and INF‐g expression. As a target of metformin, AMP‐activated protein kinase (AMPK) phosphorylation was enhanced, which might explain the suppression of inflammation in metformin group. Bone morphogenetic protein (BMP) signaling is one of the key pathways regulating gastrointestinal development and homeostasis. Consistently, metformin supplementation enhanced BMP signaling in IL10KO mice. Furthermore, the expression of two key transcription factors promoting epithelial differentiation, Hes1 and Math, was promoted by metformin. To confirm the mediator role of AMPK, we further over‐expressed AMPK in Caco‐2 cells and found that the expression of BMP4 and the phosphorylation of Smad, which is a down‐stream effector of BMP signaling, was enhanced in AMPK over‐expressed Caco‐2 cells. In addition, AMPK activity was correlated with the mRNA expression and/or protein content of Notch1, Math, Hes1, which are key transcription factors regulating epithelial differentiation. In conclusion, our data demonstrated that metformin suppresses inflammation and enhanced intestinal epithelial differentiation via enhancing AMPK signaling.Support or Funding InformationR15HD073864
Suppressive effect of nicotine on expression of MAdCAM‐1 in colitis
Authors: Maruta Koji , Kurihara Chie , Furuhashi Hirotaka , Takajo Takeshi , Yasutake Yuichi , Okada Yoshikiyo , Yoshikawa Kenichi , Higashiyama Masaaki , Watanabe Chikako , Komoto Shunsuke , Tomita Kengo , Nagao Shigeaki , Miura Soichiro , Hokari Ryota 
Published: November 2023

IntroductionAdhesion molecules play important roles in inflammatory bowel disease (IBD), and modulation of gut‐specific adhesion molecules like MAdCAM‐1 and α4β7‐integrin could be a new therapeutic target. Epidemically, nicotine absorbed by smoking is reported to work protectively in some ulcerative colitis cases. We investigated the effect of nicotine on dextran sulfate sodium (DSS)‐induced murine colitis in point of adhesion molecules.AimWe investigated the effect of nicotine on the following things; 1) symptoms of murine colitis; 2) leukocyte adhesion to colonic microvascular endothelium; 3) expression of α4β7‐integrin on lymphocytes; and 4) expression of adhesion molecules on endothelium.Method1) C57BL/6J mice were treated with 3% DSS (40kDa) in drinking water for 7 days. Some mice were treated with 0.1 mg/ml nicotine in drinking water simultaneously. Disease activities of colitis were assessed by DAI score, which consists of body weight loss, stool consistency and bleeding. 2) C57BL/6J mice were treated with 0.1mg/ml nicotine in drinking water 1 day after 3% DSS treatment for 7 days. Mice were injected FITC‐labeld splenocytes (3×107 dissoleved in 0.3ml) obtained from another mouse, via cervical vein. The number of splenocytes adhering to colonic submucosal vasculature were counted with an intravital fluorescence microscope. 3) C57BL/6J mice were treated with 0.1mg/ml nicotine in drinking water 1 day or for 7 days. The expression of α4β7‐integrin on splenocytes was analyzed with a flow cytometer. 4) The bEnd.3 cells, a cell line derived from murine endothelioma, were treated with 5 ng/ml TNF‐α for 12 hours. Some groups were treated with 1 mM nicotine simultaneously. The mRNA expression of MAdCAM‐1 was measured by quantitative RT‐PCR.ResultNicotine treatment significantly decreased DAI score induced by DSS. In intravital microscopic study, the number of splenocytes adhering to vascular endothelium was increased by DSS treatment, and the increase was significantly attenuated by additional nicotine treatment. Expression of α4β7‐integrin on splenocytes was not affected by nicotine. In bEnd.3 cells, nicotine treatment significantly attenuated the increased expression of MAdCAM‐1 induced by TNF‐α.ConclusionEnhanced expression of adhesion molecules, especially MAdCAM‐1, on the microvascular endothelium in inflamed condition, was significantly inhibited by nicotine‐treatment. This study showed that nicotine might have a protective effect on inflammation of colonic mucosa of IBD via modulation of expression of MAdCAM‐1.Support or Funding InformationNone
Mechanism of copper uptake from ceruloplasmin by cells and the involvement of an additional copper uptake transporter
Authors: Dinh Lauren , Juarez Jacqueline , Vargas Rebecca , Ramos Danny , Mar David , Ishida Michael , Linder Maria C 
Published: November 2023

Ceruloplasmin (Cp) is the most abundant copper‐containing protein in the blood plasma. Though well known for its ferroxidase activity, it also has other functions. We have now demonstrated it is a circulating source of copper for cells, using 64Cu‐labeled Cp purified from mouse plasma, and mouse embryonic fibroblasts (MEFs) that do and do not express the only as yet identified copper uptake transporter in mammalian cells, CTR1. During copper delivery, Cp was converted from the holo (Cu‐containing) to the apo form, as demonstrated by immunoblotting of native PAGE gels, and 64Cu‐uptake was not reduced by inhibitors of endocytosis. Uptake of Cp‐Cu was significantly greater in cells expressing vs not expressing Ctr1, indicating that Ctr1 can take up copper from Cp. However, most of the uptake was through an as yet unidentified transporter, that like Ctr1 preferred Cu(I) and was inhibited by Ag(I). Attempts to cross‐link Cp to this transporter in the Ctr1−/− MEFs did not result in the identification of plausible transporter proteins. To improve chances of cross‐linking sufficient protein for identification by MS‐MS, studies with plasma membrane fractions from brains and hearts of rats incubated at 4ºC with purified 67Cu‐labeled rat plasma Cp were initiated (a method used previously to identify Cp “receptors”; BBRC 139: 822, 1986) and Cp‐binding to the membranes was confirmed. We conclude that uptake of copper from Cp occurs at the cell surface, with formation of apoCp and delivery of the copper to Ctr1 and an additional transporter.Support or Funding InformationR15 GM100464 from the National Institutes of Health
MicroRNA Profiling and Regulation in Hypertensive Kidney
Authors: Weber Gregory , Pushpakumar Sathnur , Mukhopadhyay Manas , Sen Utpal 
Published: November 2023

Hypertension is a major risk factor for chronic kidney disease and renal inflammation is an integral part in this pathology. In recent years, hydrogen sulfide (H2S) has emerged as an important regulator of blood pressure. Studies in spontaneously hypertensive rats have shown that endogenous hydrogen sulfide (H2S) production is impaired due to decreased CBS/CSE expression, and exogenous H2S supplementation decreased blood pressure and vascular remodeling. MicroRNAs are short, single‐stranded RNA genes that regulate post‐transcriptional gene expression by targeting messenger RNAs (mRNAs), and are associated with a broad spectrum of physiological processes. While there are several studies aimed at investigating the role of non‐coding RNA in hypertension, the roles and implications of these RNAs are not well defined. In this study, we sought to identify microRNAs that are dysregulated in response to angiotensin‐induced hypertension in the kidney and whether hydrogen sulfide donor, GYY 4137, could reverse the microRNA alteration. Wild‐type mice were treated without or with Ang‐II (1000ng/Kg/Min) and without or with GYY4137 for 4 weeks. Blood pressure, renal blood flow, renal resistive index (RI) measurements were performed. microRNA microarrays were conducted and subsequent target prediction revealed genes associated with proinflammatory response. Ang‐II treated mice showed a significant increase in blood pressure accompanied by a decrease in blood flow in the renal cortex and an increase in the resistive index. These effects were attenuated in mice treated with GYY4137. Microarrays of the kidneys revealed downregulation of miR‐129 and miR‐299b in Ang‐II treated mice and upregulation following GYY4137 treatment. In contrast, miR‐369 was upregulated in Ang‐II treatment and suppressed in Ang‐II+GYY4137 mice. Quantitation of genes involved in proinflammatory response revealed upregulation of Tnfα, Il1β, Mcp1 and Mip2 mRNA. Our data suggests that downregulation of miR‐129 and ‐299b plays a significant role in Ang‐II induced renal inflammation, and GYY4137 reduces inflammation and is associated with reversal of miR expression.Support or Funding InformationNIH: HL104103, AHA: 15SDG25840013
The Otolithic Membrane Mediates Otolith Organization
Authors: Kramer Kenneth L. , Thiessen Kevin D. 
Published: November 2023

Otoconia are small biocrystals that link mechanical forces to the sensory hair cells in the mammalian utricle and saccule, a process essential for sensing linear acceleration and gravity as well as maintaining bodily balance. In fish, structurally similar otoliths mediate both balance and hearing. While research across several vertebrates has identified at least seven homologous proteins that are found in both otoliths and otoconia, little is known about how and when these proteins termed otoconins are organized. The goals of this project were to begin to characterize otoconin organization in the zebrafish otolith, determine when otoconin organization occurs during development, and identify some of the cellular and molecular mediators of otoconin organization. Using antibodies that we generated to the otoconins Otoconin‐90 and Starmaker, we found that these otoconins initial colocalize in early otoliths. Remarkably, Otoconin‐90 and Starmaker expression gradually changed after attachment of the otolith to early tether cells to become complementary by 30 hours post fertilization. When tether cells were ablated, otoconin expression fails to organize; yet otoconin organization eventually occurred in some embryos when tether cells appeared later in development. We further demonstrated that expression of Otogelin and Otogelin‐like in the otolithic membrane are necessary for otolith attachment and otoconin organization since unattached otoconins aggregate in both morphant and mutant embryos. We conclude that zebrafish otolith organization during early development is mediated by Otogelin and Otogelin‐like expression on the tether cell surface.Support or Funding InformationThis project was supported by grants from the National Center for Research Resources (5P20RR018788‐09) and the National Institute of General Medical Sciences (8 P20 GM103471‐09) from the National Institutes of Health.