bluuebunny/crossref_metadata_2025_split huggingface dataset:
Iron controls lung fibroblast activation and invasion via SH3RF1/HIPK2 pathway
Authors: Huang Chaoqun , Dang Quanjin , Liang Yurong , Hewawasam Sankha , Liu Lin 
Published: May 2023

Iron is a metal element indispensable for nearly all living organisms, as it participates in a variety of biological processes. The homeostasis of iron is essential to human health. Iron-responsive element-binding proteins (IRP1 and IRP2) bind the iron-responsive element (IRE) of mRNAs to control mRNA stability or protein translation. Abnormal iron homeostasis can lead to a broad spectrum of human diseases. The objective of this study was to investigate the role and mechanism of iron in lung fibroblast activation. We hypothesize that iron overload enhances lung fibroblast activation via the SH3 domain containing ring finger 1 (SH3RF1)/homeodomain interacting protein kinase 2 (HIPK2) pathway. Search for iron-responsive elements (SIREs) software was employed to predict IREs in human and mouse genomes. Lung fibroblast function studies were performed by determining the expression of myofibroblast marker α-smooth actin (α-SMA) using real-time PCR, Western blotting, and immunostaining and analyzing fibroblast invasion. Iron supplementation with ferric ammonium citrate (FAC) enhanced the mRNA and protein expression of α-SMA and fibroblast invasion. SH3RF1 was found to contain a novel IRE in its 3’-UTR and was down-regulated by iron treatment. Overexpression of SH3RF1 suppressed the iron-induced lung fibroblast activation. Moreover, iron treatment enhanced the expression of HIPK2 protein in lung fibroblasts, which was reduced by SH3RF1 overexpression. Knockdown of HIPK2 protein inhibited α-SMA mRNA expression. Furthermore, treatment of the lung fibroblasts with MG132 increased HIPK2 protein level, which was suppressed by SH3RF1 overexpression. In summary, iron controls lung fibroblast activation and invasion via the SH3RF1/HIPK2 regulation system and may be involved in the pathogenesis of idiopathic pulmonary fibrosis. R01 HL157450, R01HL135152 and P20GM103648 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Cell-free hemoglobin-mediated oxidation of low-density lipoprotein (oxLDL) contributes to lung microvascular endothelial dysfunction during sepsis
Authors: Meegan Jamie , Ware Lorraine , Bastarache Julie 
Published: May 2023

Introduction: Disruption of the microvascular endothelial barrier is a critical pathological feature of sepsis-induced acute lung injury. Plasma cell-free hemoglobin (CFH) is elevated in approximately 80% of patients with sepsis and is independently associated with development of acute respiratory distress syndrome (ARDS) and mortality. Oxidized CFH (ferric, 3+) can oxidize low-density lipoprotein (oxLDL), which signals through its major endothelial receptor, the lectin-like oxidized LDL receptor 1 (LOX-1), to cause endothelial dysfunction through several pro-inflammatory pathways including activation of mitogen-activated protein kinases (MAPKs). However, little is known regarding whether LOX-1 receptor signaling leads to microvascular endothelial hyperpermeability or acute lung injury, especially in the context of sepsis. We hypothesized that oxidation of LDL by CFH contributes to lung microvascular endothelial barrier dysfunction and worse outcomes during sepsis through LOX-1 and downstream p38 MAPK. Methods: To test whether generation of oxLDL by CFH is associated with endothelial injury in clinical sepsis, circulating levels of CFH, oxLDL, and vascular injury marker sVE-cadherin were measured in 24 sepsis patients via ELISA and tested for association with development of ARDS. LDL was oxidized by combining LDL with CFH in a test tube overnight at 37°C and oxLDL was quantified by TBARS assay. In primary human lung microvascular endothelial cells (HLMVEC) transendothelial electrical resistance (TER), a measure of barrier dysfunction, was assessed by Electric Cell-substrate Impedance Sensing (ECIS). LOX-1 receptor was blocked using BI-0115 (Boehringer Ingelheim, 20 μM) and p38 MAPK was inhibited using NiPp (100 μM). Results: In sepsis patients, plasma oxLDL levels correlated with CFH (r=0.686, p=0.016) and sVE-cadherin (r=0.603, p=0.012), and tended to be higher in those who developed ARDS (38 U/L [IQR 27, 45] vs. 27 U/L [IQR 19, 35], p=0.1). Oxidation of LDL by CFH exacerbated HLMVEC barrier dysfunction compared to control, LDL, or CFH. Barrier dysfunction induced by CFH or oxLDL was attenuated by blocking the LOX-1 receptor or p38 MAPK. Conclusions: Increased plasma CFH and oxLDL are associated with vascular injury during clinical sepsis; one mechanism by which CFH may cause vascular hyperpermeability and sepsis-mediated lung injury is through oxidation of LDL which can drive signaling through the endothelial LOX-1 receptor and activation of p38 MAPK. NIH R35HL150783, R21GM144915, R01HL158906, R01HL164937, T32HL094296; Parker B. Francis Fellowship This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Fatty acid-binding proteins promote endothelial glycolysis and arterial programming in pulmonary arterial hypertension
Authors: Dai Zhiyu , Liu Bin , Yi Dan , Hamid Syed , Kala Mrinalini , Glembotski Chris , Fallon Michael 
Published: May 2023

Introduction: Pulmonary arterial hypertension (PAH) is a disaster disease characterized by obliterative vascular remodeling and persistent increase of vascular resistance, leading to right heart failure and premature death. Understanding the cellular and molecular mechanisms will help develop novel therapeutic approaches for PAH patients. Hypothesis: We hypothesis that endothelial fatty acid metabolism is critical for obstructive vascular remodeling in the pathogenesis of PAH. Methods: A severe mouse model of PH Egln1Tie2Cre mice were bred with Fabp45-/- mice to generate Egln1Tie2Cre/ Fabp45-/- mice. Single-cell RNA sequencing (scRNA-seq) analysis and metabolomic analysis were used to profile the pulmonary cells in Egln1Tie2Cre mice and Egln1Tie2Cre/ Fabp45-/- mice. Human hPAEC from idiopathic PAH patients and healthy donors, monocrotaline (MCT)-induced and Sugen5416/hypoxia (SuHx)-induced PH rats were used to measure fatty acid-binding protein 4 and 5 (FABP4 and FABP5) expression. siRNA mediated knockdown of FABP4 and FABP5 and lentivirus mediated FABP4 and 5 overexpression were performed to study cell proliferation, apoptosis, glycolysis, fatty acid oxidation. Echocardiography, hemodynamics, histological and immunostaining assay were performed to evaluate the PH phenotypes. Results: Both FABP4 and 5 were highly induced in the ECs of Egln1Tie2Cre mice and PAECs from IPAH patients, as well as the whole lungs of MCT and SuHx-induced PH rats. Knockdown or overexpression of FABP4-5 reduced or enhanced EC proliferation, starvation-induced Caspase 3/7 activity, glycolysis and fatty acid oxidation. Genetic deletion of Fabp4 and 5 in Egln1Tie2Cre mice exhibited a reduction of right ventricular systolic pressure (RVSP), RV hypertrophy, and attenuation of pulmonary vascular remodeling, prevention of right heart failure. Fabp4-5 deletion also normalized EC glycolysis and arterial gene programming, reduced HIF-2a expression and endothelial proliferation in Egln1Tie2Cre mice. Conclusions: FABP4 and 5 control EC glycolysis and arterial programming and contribute to the development of severe PH. This work was supported in part by NIH grants R00HL138278, R01HL158596, American Heart Association Career Development Award 20CDA35310084, American Thoracic Society Foundation and Pulmonary Hypertension Association Research Fellowship, The Cardiovascular Medical and Education Fund, Arizona Biomedical Research Centre funding (ADHS18-198871), and University of Arizona Startup funding to Z. Dai This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Morphology of neuromuscular junction in patients with peripheral artery disease
Authors: Tu Huiyin , Kim Julian , Hakim Ali , Zhu Zhen , Pipinos Iraklis , Li Yulong 
Published: May 2023

Peripheral artery disease (PAD) is primarily caused by occlusion of arteries that supply blood to the lower limbs and presents significantly pathological changes of calf muscles, include degenerating and regenerating fibers, reduction of the skeletal muscle area, connective tissue proliferation and fibrosis, fat accumulation, and abnormal mitochondria, all of which are predictors of muscle functional decline. Additionally, skeletal muscles also require the motor nerve innervation for the signal transmission through the neuromuscular junction (NMJ) to induce skeletal muscle contraction. To observe whether the NMJ is changed in the PAD patients, gastrocnemius muscles were collected from 2 health subjects and 6 PAD stage III/IV patients. The motor nerve terminals were labeled with anti-neurofilament 200 and anti-synaptophysin antibodies, the nicotinic acetylcholine receptors (nAChRs) were labeled with α-Bungarotoxin. Labeled NMJs were imaged using a laser scanning confocal microscope with Z-stack. The image was captured in 65 NMJs from health subjects and 140 NMJs from PAD patients, respectively. The total nAChR area and the number of discrete fragments of each nAChR cluster, the area per fragment, the percentage of fragmented nAChR clusters, the nerve innervation and the nerve terminal occupancy were calculated. The results were represented with mean ± standard error (SE). The nAChR area was 354.9 ± 12.7 and 209.4 ± 8.2, the average number of fragment/ nAChR cluster was 2.6 ± 0.2 and 3.1 ± 0.2, 6.2% and 19.3% of the nAChR clusters were fragmented, and the nerve terminal occupancy was 61.5 ± 1.4 % and 52.0 61.5 ± 1.8 in the health subjects and PAD patients, respectively. All nAChR clusters were innervated by the motor nerve terminal in the health subjects, whereas 8.6% of nAChR clusters were denervated in PAD Patients. These data indicate that the total nAChR area per nAChR cluster and the area of each discrete fragment are reduced and fragmented nAChR clusters are increased in PAD patients. Additionally, the denervation of nAChR clusters is increased in PAD patients. These results suggest that the neuromuscular transmission is compromised in PAD patients, which limits patients’ physical activities. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Role of Mfge8 in the reduced contractility of diabetic gastroparesis
Authors: Dutcher Dustin , Perrino Brian 
Published: May 2023

Coordinated gastric contractility is imperative for ideal digestion and absorption of nutrients. This is impaired during diabetic gastroparesis, a disorder characterized by abnormal gastric smooth muscle contractions leading to delayed gastric emptying. The objective of this research was to further the understanding of the gastric smooth muscle contraction regulatory mechanisms and how these cascades are affected by diabetes. In addition to its anti-inflammatory properties, it has been shown that milk fat globule-epidermal growth factor 8 (Mfge8) binding to ɑ8β1 integrin decreases murine gastric contractile activity by reducing the inhibitory phosphorylation of MLCP, resulting in MLCP activation and reduced myosin phosphorylation. Additionally, Mfge8 is elevated in human serum from type 2 diabetic individuals compared to non-diabetic individuals. It is hypothesized that elevated levels of Mfge8 in smooth muscle tissue will lead to decreased contractility strength, a phenomenon that would resemble gastroparesis secondary to diabetes. We analyzed the contractility and Mfge8 expression of human gastric antrum smooth muscles from obese diabetic and obese non-diabetic individuals who elected to undergo sleeve gastrectomy surgery for weight loss. Short strips of gastric muscle were stimulated to contract with serial concentrations of carbachol and contractile activity was measured with AcqKnowledge software. Mfge8 protein expression was then analyzed using a Wes Simple western instrument. We report that human gastric antrum smooth muscle tissue levels of Mfge8 are elevated in diabetic obese individuals (n=22) compared to non-diabetic obese individuals (n=37, P=0.004). However, we show that the contractile activity of antrum smooth muscle from diabetic obese individuals (n=38) is not dependent on Mfge8 levels and is not significantly different from the contractility of non-diabetic obese individuals (n=38). The rate of contraction, rate of relaxation, and overall tension ( P=0.40) produced by the muscle tissues from the two groups did not significantly differ from each other. It is possible that Mfge8 was not elevated enough to cause reduced contractility, or that those with elevated Mfge8 had not yet developed symptoms of gastroparesis. Mfge8 may serve as a biomarker for diabetic gastroparesis in the future, and additional research is required to determine the pathologic significance of elevated Mfge8 levels in obese diabetic human gastric antrum smooth muscle. This research was supported by a National Institute of Diabetes and Digestive and Kidney Diseases Diabetic Complications Consortium (DiaComp, http://www.diacomp.org ) Grant DK076169, a Takeda Pharmaceuticals Innovation Center Grant to BP, and a UNRSOM Office of Medical Research Grant FD110-PG08935 to DD. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Bacillus subtilis 67989 improves endothelial-mediated dilation in male mice fed a western diet
Authors: Risk Briana , Graham Elliot , Brown Gabriele , Stark Grace , Zhang Mingye , Hinkle Kayla , Wei Yuren , Weir Tiffany , Gentile Christopher 
Published: May 2023

The gut microbiota has emerged as a novel regulator of vascular function. We have shown that targeting the microbiota can improve endothelial dysfunction in the settings of nutrient excess and obesity. In a previous clinical trial, intake of the spore-forming bacteria Bacillus subtilis was associated with improved markers of endothelial function in healthy adults. Here we examined whether consumption of Bacillus subtilis 67989 reverses endothelial dysfunction in a diet-induced obesogenic mouse model. Male C57/BJ mice were fed a Western diet (WD; n=24) or standard diet (SD; n=24) for 10 weeks, then further subdivided for an additional 9-12 weeks such that half (n=12) of all mice received B. subtilis 67989 (PB) formulated into the diets, thus yielding 4 experimental groups (WD, SD, WD+PB, SD+PB). After the dietary intervention, both WD-fed cohorts exhibited significant increases in body weight, glucose intolerance, markers of intestinal inflammation and alterations in gut microbiome composition, with no effect of PB. Endothelium-dependent dilation was impaired in WD vs SD (60.6±6.5% vs 85.3±3.8%, respectively p<0.05), whereas this impairment was mitigated in WD+PB (77.5±3.2%) such that dilation was not significantly different than SD-fed mice. In conclusion, the probiotic Bacillus subtilis 76989 attenuates obesity-related endothelial dysfunction in male mice independent of changes in body weight or glucose intolerance. Current analyses are examining putative mechanism(s) that mediate this improvement. National Heart, Lung, and Blood Institute Grant R01-HL-144611-01 (to C. L. Gentile and T. L. Weir) This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Acute hyperglycemia lowers endothelial TRPV4 channel activity
Authors: Kulshrestha Astha , Chen Yen-Lin , Sonkusare Swapnil 
Published: May 2023

Calcium signaling mechanisms regulate endothelium-dependent dilation of small arteries. Endothelial dysfunction has been linked to the clinical symptoms observed in diabetic patients, although the underlying mechanisms behind compromised endothelial health are unclear. The clinical symptoms of hyperglycemia are exacerbated in diabetic patients due to their higher initial spike in blood glucose concentration after food intake. However, the effects of acute increases in glucose levels on endothelial calcium signals have not been studied. Recent studies indicate that transient receptor potential vanilloid 4 (TRPV4) channels, a calcium entry pathway in endothelial cells, promote the dilation of small arteries. Therefore, we hypothesized that hyperglycemia lowers endothelial TRPV4 channel activity to impair the dilation of small arteries. The calcium signal activity of endothelial cells (ECs) was compared in small mesenteric arteries (MA, ~ 100 μm) from C57BL6J mice under normal (5 mM Glucose) and hyperglycemic (25 mM Glucose) conditions using spinning disk confocal microscopy. En face MAs were loaded with fluo-4 AM (10 μM) and images were acquired at 30 frames per second to capture real-time changes in calcium signals. The baseline activity of endothelial calcium signals was lower under hyperglycemic conditions compared to normal and osmotic control (5 mM Glucose/20 mM Mannitol) conditions. In the presence of TRPV4 channel inhibitor GSK2193874 (GSK219, 100 nM), hyperglycemia-induced decrease in endothelial calcium signals was abolished, suggesting that hyperglycemic conditions target endothelial TRPV4 channels. To verify this possibility, we recorded the activity of endothelial TRPV4 channels as unitary calcium influx signals, called TRPV4 sparklets, in the presence of cyclopiazonic acid to eliminate the interference from intracellular calcium release signals. TRPV4 channels were activated using GSK1016790A (GSK101, 3 nM), a selective channel activator. The number of TRPV4 sparklet sites and activity per site were both reduced under hyperglycemic conditions compared to normal conditions. Overall, these data suggest that acute hyperglycemia reduces the activity of endothelial TRPV4 channels, which may attenuate endothelial function. Future work will determine the acute contribution of hyperglycemia-induced decrease of TRPV4 channel activity to impaired endothelium-dependent vasodilation. This work was funded by grants from the NIH (HL142808 and HL147555) to Dr. Sonkusare. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
Renal NHE1 is required for regulation of intracellular pH in principal cells and type B intercalated cells
Authors: Xue Jianxiang , Thomas Linto , Rieg Jessica Dominguez , Tomilin Viktor , Pochynyuk Oleh , Rieg Timo 
Published: May 2023

The sodium-hydrogen exchanger isoform 1 (NHE1) is a plasma membrane transporter with roles in transepithelial Na+/H+ transport, intracellular pH (pHi) and cell volume regulation. Within the kidney, NHE1 is previously documented to be expressed in all nephron segments. However, recent transcriptomic and proteomic analyses, and our staining results in mice, consistently find no expression of NHE1 in the proximal tubule or thick ascending limb, but in the distal nephron, including distal convoluted tubule, connecting tubule, and collecting duct (principal and intercalated cells, PC and IC, respectively). We hypothesized that basolateral NHE1 in IC (co-labelling with H+-ATPase B1 subunit, labeling all 3 types of IC: type A, B, and non-A-non-B) plays an important role in acid-base homeostasis. Due to lack of a suitable animal model, we generated kidney-specific NHE1 knockout (NHE1KS-KO) mice. NHE1KS-KO mice completely lack renal NHE1 protein expression compared to their control littermates (n=6 male mice/genotype). No differences were observed in body weight, food and water intake (determined in their home cages) or concentrations of blood Na+, K+, Ca2+, pH or HCO3-. Urine analysis showed no significant differences in electrolytes, minerals of pH. To mechanistically determine the role of NHE1 for pHi regulation in IC and PC we used confocal microscopy of acutely-split open cortical collecting ducts. This method allows for the analysis of a vast number of PC/IC simultaneously. To study the role of NHE1 we employed a NH4Cl pre-pulse (40 mM) and measured pHi recovery after switching back to a physiological solution. Intracellular pH recovery in PC (n=171) was significantly slower (~50%) in NHE1KS-KO mice compared to controls (n=85). Similar to PC, pHi recovery in type B IC of NHE1KS-KO mice (n=38) was significantly slower (~35%) compared to controls (n=40). No differences in type A IC were observed between NHE1KS-KO mice (n=35) and controls (n=29). Inhibition of ClC-K2 via NPPB (Cl-channel blocker) was used to confirm cell types: PC did not respond with a change in pHi but NPPB increased pHi in type A and decreased pHi in type B ICs. In summary, our results identify for the first time an important role of NHE1 for pHi regulation particularly in PCs and type B ICs. Funding to Dr. Rieg was provided by the NIDDK grant 1R01DK110621, VA Merit Review Award IBX004968A, AHA Transformational Research Award 19TPA34850116, and NIDDK Diabetic Complications Consortium grants DK076169 and DK115255. DK095029 and DK117865 (both to O. Pochynyuk). Dr. Thomas was supported by an AHA postdoctoral grant (828731), V. Tomilin by an AHA Career Development Award 19CDA34660148. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.