bluuebunny/crossref_metadata_2025 huggingface dataset:
Functional properties of mouse ferroportin, a cellular iron‐export protein
Authors: Azucenas Corbin P , Bonamer John P , Ruwe T Alex , Qiao Bo , Ganz Tomas , Nemeth Elizabeta , Mackenzie Bryan
Published: June 2021

Iron homeostasis is a topic of great public‐health importance because of the high prevalence of iron‐deficiency anemia and of hereditary disorders that cause iron overload. Ferroportin (Fpn)—the only known mammalian cellular iron exporter—is expressed on the plasma membrane of macrophages, enterocytes, and hepatocytes. Fpn, under the control of the iron‐regulatory hormone hepcidin, serves a critical role in the regulation of systemic iron. Whereas we have previously characterized the functional properties of human Fpn, a great deal of research in iron homeostasis utilizes mouse models, highlighting the need to study mouse Fpn in vitro. For example, the flatiron mouse model of classical ferroportin disease bears the mutation H32R in mFpn and is characterized by systemic iron deficiency and macrophage iron loading. The flatiron mouse also appears to exhibit a manganese phenotype [Seo YA and Wessling‐Resnick M (2015) FASEB J 28, 2726–2733], raising the possibility that mouse Fpn (mFpn) serves a role in Mn metabolism, although Mn is not a substrate of human Fpn. We therefore tested the hypothesis that mouse but not human Fpn can transport Mn. We characterized the functional properties of mFpn by using radiotracer assays in RNA‐injected Xenopus oocytes expressing mouse or human Fpn. We found that mFpn can export several transition metals, with selectivity Fe > Co > Ni, but not Cd. Whereas the H32R mutant abolished mFpn‐mediated Fe efflux relative to wildtype, neither wildtype nor mutant mFpn could transport Mn. mFpn‐mediated Fe efflux was pH‐dependent with estimated optimal pH of 8.2 ± (SEM) 0.2 when [Ca2+]o = 2 mM. mFpn‐mediated Fe efflux was activated by extracellular Ca2+, proceeded independently of extracellular Na+, and was inhibited 51% ± 1% by 10 μM hepcidin (30‐min preincubation). Our data (i) reveal that human and mouse Fpn share similar functional properties and (ii) do not support the notion that Fpn can transport Mn.

Support or Funding Information

NIH grants R01 DK107309, P30 DK078392, and R25 HL115473, and the American Physiological Society

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Identification of novel, potent and selective inhibitor of VRAC
Authors: Jeon Dong‐Kyu , Namkung Wan
Published: June 2021

Tight regulation of cell volume is essential for normal cell function. Sudden changes in the osmolarity of the extracellular fluid (ECF) cause cell swelling or shrinkage, and leads to regulatory volume decrease (RVD) and regulatory volume increase (RVI). In the cell volume regulation, RVD involves activation of ion transport which mediates the efflux of K+ and Clfrom the cells. The volume‐regulated anion channel (VRAC) is involved in Cl− efflux and plays a pivotal role in the regulation of cell volume. In addition, it is known that various processes such as cell division, migration, necrotic and apoptotic cell death are directly related to VRAC. Short circuit current measurement in FRT cells expressing ANO1 chloride channel revealed that the most potent and selective VRAC inhibitor, DCPIB, strongly inhibited ANO1 chloride current. Several cell lines express both ANO1 and VRAC, and there is cross talk between these chloride channels. Thus, the lack of selective inhibitors of VRAC limits VRAC and ANO1 research. In this study, we performed a cell‐based high‐throughput screening to identify potent and selective VRAC inhibitors. Screening of 55,000 compounds and electrophysiological studies revealed a novel, potent and selective VRAC inhibitor, VRACi‐A01 (IC50 = 2.7 μM). It did not affect the ANO1 chloride channel activity at the concentration showing complete inhibition of VRAC. To the best of our knowledge, VRACi‐A01 is the most potent and selective VRAC inhibitor reported to date and a useful research tool for pharmacological dissection of VRAC in the cells expressing VRAC and ANO1.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MicroRNA‐103a plays a role in regulating human sodium‐dependent vitamin C transporter‐1 (hSVCT1) in intestinal epithelial cells
Authors: SUBRAMANIAN VEEDAMALI , Sabui Subrata , Marchant Jonathan , Said Hamid
Published: June 2021

Ascorbic acid (AA; vitamin C) is essential for normal human health and well‐being. Vitamin C acts as a cofactor for many important enzymatic reactions and is a potent antioxidant. Humans cannot synthesize vitamin C endogenously and obtain the vitamin from dietary sources via intestinal absorption, a process that involves the human sodium‐dependent vitamin C transporters‐1 &‐2 (hSVCT1 and hSVCT2; products of the SLC23A1 and SLC23A2 genes, respectively). Previous studies have shown that hSVCT1 is predominantly expressed in the intestine and localized at the apical membrane of polarized enterocytes where it mediates AA absorption. Thus, hSVCT1 expression levels are an important determinant of physiological vitamin C homeostasis. To date, little is known about post‐transcriptional mechanisms that regulate hSVCT1 expression in intestinal epithelial cells. Here, we investigated posttranscriptional regulation of hSVCT1 by microRNA (miRNA) in intestinal epithelial cells. A pmirGLO‐SLC23A1‐3′‐UTR construct transfected into human intestinal epithelial cells (Caco‐2 and NCM460 cells) showed significantly reduced luminescence. Bioinformatic analysis of the SLC23A1‐3′‐UTR predicted five putative miRNA binding sites (miR‐103a, miR‐107, miR‐328, miR‐384, and miR‐499‐5p) in the 3′‐UTR. Expression of mature miR‐103a was significantly higher than four other miRNA regulators in both intestinal cell lines as well as mouse jejunal mucosa. Addition of a miR‐103a mimic, but not a miR‐103a mutant construct, significantly reduced luciferase reporter activity of pmirGLO‐SLC23A1‐3′‐UTR. In contrast, addition of a miR‐103a inhibitor markedly increased luminescence. Further, addition of the miR‐103a mimic led to a marked inhibition in AA uptake, associated with decreased hSVCT1 mRNA and protein expression in intestinal epithelial cells. Reciprocally, the miR‐103a inhibitor significantly increased AA uptake, associated with increased levels of hSVCT1 mRNA and protein expression in intestinal epithelial cells. These findings demonstrate for the first time that hSVCT1 is post‐transcriptionally regulated by miR‐103a in human intestinal epithelial cells.

Support or Funding Information

Supported by NIH grants DK 107474, DK 58057, DK 56057 and a grant from the DVA.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Increased HDAC9 Expression is Associated with Decreased Estrogen in Female Patients with Intracranial Aneurysm
Authors: Lansdell Theresa A , Fisher Courtney , Demel Stacie , Dorrance Anne
Published: June 2021

Background and purpose

Subarachnoid hemorrhage is a devastating consequence of intracranial aneurysm (IA) rupture. Therefore early identification of patients with IA is important. Identification of biomarkers for IA will help improve diagnosis, the understanding of IA pathogenesis, and patient management. A risk allele (rs10230207), located nearby the histone deacetylase 9 (HDAC9) gene on chromosome 7, has been associated with IA incidence. HDAC9 can repress the transcription of estrogen receptors α and β (ESR1 and ESR2). We tested the hypothesis that lymphocytes from patients with IA will have increased HDAC9 and decreased estrogen receptor gene transcription compared to controls.

Methods

Immortalized B‐lymphocytes from male and female patients with un‐ruptured IA (N=109) and population controls (N=85) were obtained from the NINDS repository and cultured in RPMI‐1640 medium supplemented with 10% fetal calf serum. Subject genomic DNA was genotyped for presence of the risk allele using TaqMan SNP genotyping assays directed toward rs10230207. HDAC9 and ESR gene expression were analyzed using TaqMan expression assays. In a separate study using subject case (N=23) and control (N=10) blood samples, HDAC9 and ESR2 transcriptional differences were confirmed in peripheral blood mononuclear cells. Plasma estrogen levels were also measured in these subjects.

Results

HDAC9 expression was increased in IA patient lymphoblasts compared to population controls (P= 0.0311) and was associated with the presence of the rs10230207 risk allele (P=0. 0235). Patients harboring the risk allele had an increased risk for IA (OR 9.1, 95% CI 3.6–23.1). While the risk allele puts both males and females at risk for IA, the risk of IA is larger for females (OR 23.9, 95% CI 5.4–105.4) compared to males (OR 5.9, 95% CI 2.7–13.2). Lymphoblasts from female subjects between 19–50 years (premenopausal) were further analyzed (N= 17 cases and 22 population controls). HDAC9 gene expression was increased (P=0.0243) and ESR2 expression was decreased (P= 0.0351) in premenopausal females with IA compared to premenopausal female population controls. Decreased plasma estrogen levels were also detected in the premenopausal female cases (N=7) compared to control (N=5) subjects (P =0.0015).

Conclusions

The presence of the rs10230207 risk allele was associated with increased expression of HDAC9 and decreased expression of ESR2 in case and control immortalized lymphoblasts. Furthermore, we found that female IA patients that had not yet undergone menopause had decreased plasma estrogen levels. Future studies are aimed at defining the positive and negative predictive values of estrogen levels and IA in female patients.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Extracellular vesicles from Wistar Kyoto and spontaneously hypertensive rats have differential vasodilatory responses in resistance arteries
Authors: Good Miranda Elizabeth , Musante Luca , LaSalvia Sabrina , Carey Robert M , Howell Nancy L , Le Thu H , Erdbruegger Uta , Isakson Brant E
Published: June 2021

Extracellular vesicles (EVs) have been described as novel bio‐markers and bio‐activators in vascular dysfunction in hypertension (HTN). However, the mechanism by which EVs affect vascular function is unknown. To examine the effects of EVs on acetylcholine (ACh)‐mediated vasodilation, we isolated 3rd order mesenteric arteries and EVs from 12‐wk old normotensive Wistar‐Kyoto (WKY) and spontaneously hypertensive (SHR) rats. Arteries were cannulated on a pressure myograph and EVs (~6×107EV/mL) were added to the vessel lumen and circulating bath. No significant difference in ACh vasodilation was observed in control WKY and SHR arteries (no EVs). Interestingly, WKY arteries treated with WKY EVs reduced vasodilation compared to control arteries, an effect lost when treated with SHR EVs. ACh‐mediated vasodilation was not altered in SHR arteries when treated with either WKY or SHR EVs. WKY arteries treated with EVs isolated from 6‐wk old WKY and SHR, an age where SHR rats have not yet developed HTN, or with delipidated EVs from 12‐wk old WKY and SHR rats resulted in reduced vasodilation. Together, these data suggest that normotensive EVs reduce ACh‐mediated vasodilation and upon development of hypertension fail to reduced vasodilation. However, reduced vasodilation can be restored by delipidation of the SHR EVs or SHR EVs obtained prior to development of hypertension. Analysis of isolated EVs showed a larger number of CD45‐positive EVs in 12 week‐old SHR rats, indicating an increase in EVs from inflammatory cells following the development of hypertension. Using EVs isolated from human blood samples, a similar reduction in vasodilation was observed in arteries exposed to EVs isolated from normotensive humans. This effect was lost when arteries were treated with EVs isolated from hypertensive patients. This data supports the functional role of EVs in vascular regulation that is altered in HTN.

Support or Funding Information

K99HL143165, K23HL12610, HL088554

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
AdipoRon, adiponectin receptor agonist, improves vascular function in the mesenteric arteries of type 2 diabetic mice
Authors: Choi Soo‐Kyoung , Kwon Youngin , Byeon Seonhee , Lee Young‐Ho
Published: June 2021

Background

Adiponectin is one of the most abundant adipokines secreted from adipose tissue. An orally active synthetic adiponectin receptor agonist, adipoRon has been suggested to ameliorate insulin resistance, myocardial apoptosis, and pancreatic tumor. It has been reported that adiponectin directly induces vascular relaxation however, the chronic effect of adipoRon in the vascular dysfunction in type 2 diabetes has not been studied yet. Thus, in this study, we examined whether adipoRon improves vascular function in type 2 diabetes and what mechanism is involved.

Methods

Ten to 12‐week old male type 2 diabetic (db−/db−) mice were treated with adiponectin receptor agonist (adipoRon, 10 mg/kg/everyday, by oral gavage) for 2 weeks. Isolated mesenteric arteries were mounted in the arteriography and arterial diameter was measured. And western blot analysis was assessed.

Results

Pressure‐induced myogenic response was significantly increased, whereas endothelium‐dependent relaxation was significantly reduced in the mesenteric arteries from type 2 diabetic mice. Interestingly, treatment of adipoRon normalized potentiated myogenic response. However, endothelium‐dependent relaxation was not affected by treatment of adipoRon. The expression levels of adiponectin receptor 1, 2 and adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1, 2 (APPL 1, 2) were increased in the mesenteric arteries from Type 2 diabetic mice and treatment of adipoRon did not affect them. Interestingly, adipoRon treatment increased the phosphorylation level of AMP‐activated protein kinase (AMPK) and decreased phosphorylation of myosin phosphatase target subunit 1 (MYPT1) in the type 2 diabetic mice while there was no change in the level of eNOS phosphorylation.

Conclusion

The treatment of adipoRon improves vascular function in the mesenteric arteries from type 2 diabetic mice through endothelium‐independent mechanism. We suggest that myosin light chain phosphatase (MLCP) activation through reduced phosphorylation of MYPT1 might be the dominant mechanism in the adipoRon‐induced vascular effect.

Support or Funding Information

This work was supported by the Basic Science Research Program of the National Research Foundation (NRF) of Korea funded by the Ministry of Education, Science and Technology (NRF‐2018R1D1A1B07041820).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Differential diagnosis of cardiac tumors on a cadaveric specimen
Authors: Raja Dewan S.
Published: June 2021

Objectives

Primary cardiac tumors are a relatively rare occurrence in autopsy and surgery reports of United States. Notably, around 80% of all cardiac tumors are benign. The most common cardiac tumor is myxoma, which typically hangs inside the left atrium. On a female cadaveric specimen of about 70 years of age—we recently discovered nodular masses on the heart surface in the gross anatomy laboratory. We found two rounded cardiac tumors of 2 cm and 3 cm in diameter, both on the right ventricle encroaching on the pulmonary trunk. We explored her lungs and liver, finding no tumors in or on the cadaver's other viscera. We examined the tumors macroscopically and, on palpating, found it hard. Then we incised the nodule, finding a semi‐solid blood clot and some cheesy materials. I took photographs before and after the incision.

Methods

We searched several pathology textbooks and relevant scientific literature the cause of this swelling.

Result and Conclusion

The differential diagnosis includes aneurysms of the right atrium with clotted blood, lipomas, epicardial cysts and Chagas cardiomyopathy. Our discovery has subsequently drawn keen attention from faculty and students.

Support or Funding Information

None

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Evidence of Endothelin‐B Receptor Dysfunction in Obesity
Authors: Derella Cassandra C , Looney Jacob , Bieck Maire Rose , Jeong Jin hee , Rodriguez‐Miguelez Paula , Tucker Matthew A , Thomas Jeffrey , Stanheqicz Anna E , Alexander Lacy M , Pollock David M , Harris Ryan A
Published: June 2021

Background

The endothelin system has been identified as a key regulator of vascular function and is comprised of two opposing receptors; a constricting A‐receptor (ETAR) and dilating B‐receptor (ETBR). Dysregulation of the ETBR and obesity both independently contribute to vascular endothelial dysfunction. However, whether obese individuals exhibit ETBR dysfunction has yet to be determined. Thus, this study sought to test the hypothesis that microvascular ETBR dysfunction is present in obesity compared to normal weight individuals.

Methods

Intradermal microdialysis assessments were completed on 13 individuals (9 men; 4 women; age range: 19–38 years old). Participants were recruited according to BMI: lean (BMI ≤ 24.9 kg/m2; n = 6) or overweight/obese (BMI ≥ 25.0 kg/m2; n = 7). Two microdiaylsis fibers were inserted into the dermal layer of the right forearm where cutaneous blood flow was measured using laser speckle contrast imaging. ETBR function was determined two ways: 1) by a seven dose incremental perfusion of ET‐1 (10−20–10−8 M) combined with the selective ETAR antagonist, BQ‐123 (750 nM), and 2) by a seven dose incremental perfusion of ET‐3 (10−17–10−11 M), the isoform that has ETBR selectivity at low doses. The dose response area under the curve (AUC) for cutaneous vascular conductance was calculated for each individual.

Results

The AUC for the ET‐1 + BQ‐123 dose response was significantly (p=0.02 reduced in the overweight/obese group (AUC = 157.0 ± 11.0 AU) when compared to the lean group (AUC = 221.8 ± 41.1 AU). The AUC for the ET‐3 dose response tended (p=0.21) to be lower in the overweight/obese group (AUC = 155.2 ± 8.5 AU) compared to the lean group (AUC = 191.3 ± 46.4 AU).

Conclusion

These data suggest that ETBR function is impaired in obesity compared to normal weight individuals. The reduced ETBR function may contribute to the observed prevalence of vascular endothelial dysfunction in obesity; however, this hypothesis warrants further investigation.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Angiotensin‐(1‐7) Prevents Thyroid Hormone‐induced Cardiomyocyte Hypertrophy by FOXO3 Signaling Pathway
Authors: Senger Nathalia , Santos Robson Augusto , Barreto‐Chaves Maria Luiza Morais
Published: June 2021

It is well‐known that high levels of thyroid hormone promote cardiomyocyte hypertrophy, which in part is mediated by activation of Renin‐Angiotensin System (RAS). Recent study of our group demonstrated that the contralateral RAS axis (ACE2/Angiotensin‐(1–7)/Mas receptor) is also involved in this cardiac hypertrophy model and that transgenic rats overexpressing circulating levels of Angiotensin‐(1‐7) (Ang‐(1‐7)) present an improvement of cardiac function and attenuation of cardiac hypertrophy. However, the molecular mechanisms involved in these Ang‐(1‐7) actions have not been fully elucidated. The aim of this study was to investigate if the anti‐hypertrophic effect of Ang‐(1‐7) also occurs in isolated cardiomyocytes, as well as the involvement of the Foxo3 signaling pathway for these effects. All experiments were approved by the Ethics Committee on Animal Experimentation of ICB‐USP (n030/12/CEUA). Primary cardiomyocytes cultures obtained from neonatal rats were treated with triiodothyronine (T3) (15nM) and/or Angiotensin‐(1‐7) (Ang‐(1‐7)) (100nM). All data were analyzed by ANOVA followed by Tukey's post hoc test, and values of P<0.05 were considered statistically significant. The Ang‐(1‐7) treatment prevented the cardiomyocyte hypertrophy induced by T3, which was confirmed by reduction cell surface area, L‐[3,4,5‐3H] leucine incorporation and BNP gene expression. Combined treatment with the MAS receptor antagonist (A779, 100nM) impaired the anti‐hypertrophic effect of Ang‐(1‐7). Regarding molecular mechanisms, although Ang‐(1‐7) treatment did not impair the rapid activation of Akt promoted by T3, Ang‐(1‐7) treatment reduced the phosphorylation of Foxo3 (inactive form) and promoted the nuclear accumulation of Foxo3 in cardiomyocytes, indicating the activation of this protein. Collectively, these findings indicate an important cardioprotective effect of Ang‐(1‐7) in isolated cardiomycoytes treated with T3, occurring via MAS receptor and with possible participation of Foxo3.

Support or Funding Information

Foundation for the Support of Research in the State of São Paulo (FAPESP) and National Council of Technological and Scientific Development (CNPq).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Antiarrhythmic Effects of Chronic Melatonin Treatment Are Not Associated with Its Antioxidative Action in Rat Myocardial Ischemia/Reperfusion Model
Authors: Azarov Jan E , Efremova Yulia , Bernikova Olesya , Pliss Michael , Lopatina Ekaterina , Vaykshnorayte Marina , Sedova Ksenia
Published: June 2021

Introduction

Melatonin was proposed as a potential antiarrhythmic agent, and its antioxidative properties are considered to be the basis for the antiarrhythmic effect. The present study was aimed at the evaluation of association between ventricular tachycardia and/or ventricular fibrillation (VT/VF) incidence, oxidative stress parameters and electrophysiological changes in myocardial ischemia/reperfusion setting.

Methods

18 rats were treated by melatonin (10 mg/kg/day, orally) for 7 days, while 13 control animals were given placebo. The left anterior descending artery was reversibly ligated for 5 min to induce ischemia and subsequent reperfusion in the anesthetized animals. Unipolar electrograms were recorded from ventricular epicardium with a 64‐lead array connected to a custom‐made high‐resolution (4 kHz) mapping system. Local activation and repolarization times were measured as instants of dV/dt min during QRS complexes and dV/dt max during T waves in unipolar electrograms, respectively. Local repolarization durations and dispersion of repolarization were calculated. After experiments, the hearts were excised, and parameters of the oxidative stress were determined in ventricular myocardium.

Results

VT/VF incidence was less in the animals given melatonin as compared to controls (6/18 vs 9/13, p=0.047, respectively). Superoxide dismutase activity was higher in the melatonin group as compared to the control group (69.6±0.5 vs 60.2±2.0 inhibition rate %, p=0.001, respectively), while total antioxidant capacity, 4‐HNE adduct and total glutathione content did not differ in the studied groups. No parameters of oxidative stress were associated with arrhythmia susceptibility. Among the studied ventricular epicardial electrophysiological parameters, local activation times at the baseline (preischemic) state were both reduced by melatonin treatment (9.39±0.26 vs 10.58±0.33 ms, p=0.012) and associated with VT/VF incidence (B=1.872, 95% CI 1.106–3.169, p=0.020) in logistic regression analysis. No association between parameters of ventricular electrical excitation and oxidative stress were found.

Conclusion

Though chronic melatonin treatment enhanced enzymatic antioxidant activity, its antiarrhythmic action in ischemia/reperfusion model was associated with its direct effect on ventricular activation pattern.

Support or Funding Information

The study was supported by Russian Science Foundation (RSF 18‐15‐00309).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Exogenous MG53 Protects Adult Mouse Cardiomyocytes by Preventing Mitochondria Damage in Response to Oxidative Stress
Authors: Gumpper Kristyn Nicole , Ko Jae‐Kyun , Xu Li , Han Renzhi , Zhou Hua , Lin Pei‐Hui , Ma Jianjie
Published: June 2021

Ischemic heart disease is a major cause of mortality and morbidity world‐wide and its incidence is continually increasing as other health factors, such as diabetes type II and obesity, climb. Ischemic injury of any tissue causes a loss of healthy mitochondria, due to an increase in reactive oxygen species leading to oxidative stress. MG53, also known as TRIM72, is highly expressed in skeletal muscle and modestly in other tissues and is essential to repair damage to plasma membrane by acting as a myokine. We have previously shown that MG53's membrane repair function is associated with changes in oxidative state inside the cell. Moreover, we have shown that mg53−/− mice are more susceptible to ischemia‐reperfusion injury, whereas treatment with exogenous recombinant human MG53 (rhMG53) reduces both infarct damage and fibrosis and restores cardiac function. We hypothesize that in addition to cell membrane repair, exogenous MG53 enters cardiomyocytes and can act as a myokine to protect cardiomyocytes by maintaining mitochondrial function through control of oxidative‐stress induced mitophagy. In this study, MG53 protection of cardiomyocyte mitochondria was assessed by subjecting HL‐1 cells (an immortalized mouse atrial cardiomyocyte cell line) to oxidative stress followed by analysis for mitochondrial structure and function. Overall, we show that treatment with rhMG53 allowed cells to maintain a healthy mitochondrial membrane potential to protect mitochondrial function. We are currently dissecting the mechanism that facilitates the endocytic uptake of MG53 from the circulation into the cardiomyocytes as well as downstream signaling pathways that mediate the protection of the mitochondria function. The novelty of this study is that it shows exogenous MG53 can protect mitochondria in cardiomyocytes upon oxidative stress. This provides a mechanism behind how rhMG53 treatment may be a clinically relevant strategy to reduce cardiomyocyte injury and maintain cardiac function in patients after ischemic injuries.

Support or Funding Information

Research Support: R01 grants (DK106394 and AR070752) from NIH to J. M and AHA predoctoral fellowship (18PRE34030430) to K.G.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Regulation of Brown Adipose Tissue Function by HuR
Authors: Anthony Sarah R. , Lanzillotta Lindsey , Guarnieri Adrienne , Green Lisa , Slone Sam , Helsley Robert N. , Brown J. Mark , Civelek Mete , Owens A. Phillip , Tranter Michael
Published: June 2021

Stimulating the metabolic function of brown adipose tissue (BAT) and/or inducing the beiging of white adipocytes (WAT) to BAT‐like cells represent potential therapeutic strategies for increasing energy expenditure and reducing obesity. HuR (ELAVL1) is an RNA binding protein that mediates gene expression through stabilization of mRNA targets and is highly expressed in both WAT and BAT. We identified an inverse correlation between HuR expression and obesity in humans via the METabolic Syndrome In Men study of Finnish men. Thus, we hypothesized that HuR expression in adipocytes plays a critical role in maintaining metabolic homeostasis.

Our results show that, compared to control littermates, adipocyte‐specific HuR‐null (adipo‐HuR−/−) mice display a lean phenotype with decreased fat mass and increased energy expenditure, independent of diet. Interestingly, adipo‐HuR−/− mice also show a disruption of BAT structure and function, with increased lipid droplet size, decreased BAT density, and an inability to thermoregulate when subjected to cold. Examination of skeletal muscle suggests that the lean phenotype in adipo‐HuR−/− mice is due to compensatory shivering to thermoregulate in the absence of functional BAT, a result consistent with UCP1−/− mice. Surprisingly, we found no difference in basal expression of UCP1 in BAT from control and adipo‐HuR−/− mice. However, HuR is activated downstream of β3 adrenergic stimulation and appears to play a role in cold/β3 induced UCP1 expression. Additionally, adipo−HuR−/− mice show a decrease in mitochondrial density in both BAT and subcutaneous (sc) WAT, as well as a decrease in brown/beige adipocyte marker genes (PRDM16, Elovl3) in scWAT.

In conclusion, our results demonstrate that HuR plays a central role in the structure and function of BAT. Work is ongoing to identify the transcriptome wide changes in HuR‐dependent gene expression as well as the potential role of HuR in beiging of WAT.

Support or Funding Information

This work was partially funded by a University of Cincinnati Heart, Lung, and Vascular Institute Pilot Grant (MT, APO).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Genistein and Exercise Reverse Hyperglycemia and Hyperinsulinemia Induced by High Fat/High Sucrose Diet via Influences on Hepatic Gluconeogenesis in Mice
Authors: St Aubin Chaheyla , Fisher Amy , Broderick Thomas , Al‐Nakkash Layla
Published: June 2021

Consumption of a high fat/high sugar (HFHS) diet has not only been identified as risk factor for obesity, but it also predisposes towards type 2 diabetes mellitus (T2DM), by resulting in hyperglycemia and impaired insulin signaling. A targeted metabolic pathway for treatment of T2DM is hepatic gluconeogenesis. Gluconeogenesis is responsible for maintaining blood glucose levels during times of exercise, fasting, and stress. Insulin is responsible for suppressing gluconeogenic activity. However, as the liver becomes insulin resistant, hepatic gluconeogenesis becomes over activated exacerbating T2DM pathology. We aimed to assess whether genistein and/or exercise regulate hepatic gluconeogenesis. C57BL/6J male and female mice aged 5–6 weeks were randomly assigned to one of the following groups (n=8–10/group): lean control (Ln), high fat/high sucrose diet (HFHS), HFHS+Gen (genistein), HFHS+Ex (exercise), and HFHS+Gen+Ex. The HFD consisted of 60% saturated fat, 20% carbohydrate, 20% protein and included sucrose and fructose (42 g/L) in the drinking water. Exercise consisted of daily moderate treadmill running for a total duration of 150 minutes/week for 12 weeks. Genistein dose was 600 mg genistein/kg HF diet. C57BL/6J mice fed HFHS exhibited phenotypes seen in T2DM pathology; including rapid weight gain, hyperinsulinemia and hyperglycemia. Our results demonstrate that serum glucose and insulin levels in males were rescued by Gen+Ex treatment combined compared to controls. Corticosterone levels were significantly elevated in females fed HFHS diet (2755±375 pg/mL, n=6) compared to Ln controls (1449 ± 275 pg/mL, n=7), with significant recovery by Gen+Ex (964±234 pg/mL, n=7). Measures of total protein expression of renal 11βHSD2 (responsible for the conversion of active cortisol to inactive cortisone) however, showed no significant difference indicating that 11βHSD2 may not be playing a role in corticosterone deactivation. We are currently determining total protein expression levels of hepatic 11βHSD1 (responsible for the conversion of corticosterone to its active form). Current studies are also evaluating the roles of two rate limiting enzymes; phosphoenolpyruvate carboxykinase (PEPCK) and glucose‐6‐phosphotase (G6Pase). These studies provide evidence of the protective role that genistein and exercise may provide to ameliorate diabetic‐obesity induced by HFHS diet.

Support or Funding Information

Support: Layla Al‐Nakkash, Tom Broderick were supported by Midwestern‐Arizona Alzheimer's Consortium and Midwestern University Intramural Funds. Chaheyla St Aubin and Amy Fisher were supported by the Department of Biomedical Sciences.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Gene and protein expression of exercise pressor reflex‐related molecular receptors in dorsal root ganglion of spontaneous‐hypertensive and normotensive rats
Authors: Weavil Joshua C , Kwon Oh Sung , Hughen Ronald W , Zhang Jie , Richardson Russell S , Light Alan R , Amann Markus
Published: June 2021

BACKGROUND

The exercise pressor reflex (EPR), mediated by mechano‐ and metabosensitive skeletal muscle afferent feedback, is exaggerated in spontaneously hypertensive rats. This cardiovascular reflex dysfunction could be the result of disease‐related abnormalities in the expression and/or sensitivity of molecular receptors located on the terminal end of these muscle afferents, altered central integration of muscle afferent feedback, and/or altered end‐organ responsiveness to a given sympathetic activity.

PURPOSE

To examine dorsal root ganglion (DRG) gene and protein expression of molecular receptors recognized to be significant determinants of the EPR in control (CTRL) and spontaneous hypertensive rats (SHR).

METHODS

Twelve lumbar DRGs (6 left, 6 right) were harvested from each of the ten CTRL (mean arterial blood pressure (MAP): 99 ± 2 mmHg) and the ten SHR (MAP: 146 ± 3 mmHg) male rats. DRGs from the right side were quickly frozen for later extraction and RNA sequencing. All mRNA values were normalized to transcripts per million (TPM). DRGs from the left were quickly frozen and later assessed for protein expression by Western Blot. All protein was normalized to GAPDH. Analyses were focused on the transient receptor potential cation channel 1 (TRPV1), acid‐sensing ion channel 3 (ASIC3), purinergic receptor 2X3 (P2X3), and prostaglandin E2 receptor 4 (EP4).

RESULTS

mRNA content was similar between SHR and CTRL animals (TRPV1: 113 ± 9 TPM vs 110 ± 13 TPM; ASIC3: 39 ± 9 vs 44 ± 7; P2X3: 246 ± 16 vs 246 ± 26; EP4: 10 ± 1 vs 9 ± 1, respectively; all P > 0.18). In contrast, SHR animals demonstrated a greater protein expression for all receptors measured compared to CTRL animals (TRPV1: 1.47 ± 0.20% vs 0.86 ± 0.06%; ASIC3: 1.36 ± 0.09 vs 0.77 ± 0.07; P2X3: 1.49 ± 0.11 vs 1.18 ± 0.11; EP4: 1.36 ± 0.21 vs 1.11 ± 0.13; all P < 0.01).

CONCLUSION

In the face of an equivalent mRNA content in the SHR and CRTL DRG, the substantially greater receptor protein content in the SHR supports the potential for a dysregulated expression of muscle receptor protein in the hypertensive rats. Furthermore, these findings suggest an abnormally increased content of EPR‐related molecular receptors as a potential mechanism contributing to the exaggerated EPR which characterizes these animals.

Support or Funding Information

HL‐116579

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Central Administration of Hydrogen Sulfide Alleviates Rodent Angiotensin II Hypertension
Authors: Donertas Basak , Malphurs Wendi L. , Baekey David M. , Julian David , Sirmagul Basar , Zubcevic Jasenka
Published: June 2021

Introduction

Activation of autonomic neural pathways plays a significant role in pathogenesis of hypertension (HTN). Hydrogen sulfide (H2S) is an endogenous neuro‐ and immunomodulatory molecule that is reportedly reduced in both rodent and human HTN. However, central role of H2S in HTN is not well understood. We hypothesized that chronic intracerebroventricular (ICV) infusion of sodium hydrosulfide (NaHS), an H2S donor, will alleviate rodent angiotensin II (Ang II) HTN.

Methods

Adult male Sprague Dawley rats were implanted with radiotelemetry transmitters (DSI) in the descending aorta. Following baseline BP measurements, rats were randomly divided into three groups: (i) ICV NaHS; (ii) HTN; and (iii) ICV NaHS‐treated HTN rats. HTN was induced by chronic infusion of Ang II (200 ng/kg/min s.c.) for four weeks using mini‐osmotic pumps. ICV NaHS (30 nmol/h) or ICV saline was administered simultaneously with s.c. Ang II or saline infusion. At endpoint, H2S levels were measured in plasma samples of all rats.

Results

At week four, mean arterial pressure (MAP) was significantly reduced in the ICV NaHS‐treated HTN group when compared with HTN group (104±7 vs. 166±5 mmHg, respectively; P< 0.01). This was associated with a lower MAP at night in the ICV NaHS‐treated HTN vs. HTN group (Delta MAP from day to night: 15.29±4 vs. 31.37±17 mmHg, respectively), suggesting reduced sympathetic drive following treatment with NaHS. Interestingly, this also reflected in significantly higher plasma H2S levels in the ICV NaHS‐treated HTN vs. HTN group (0.070±0.000 AU vs. 0.067±0.000 AU, respectively; P<0.05).

Conclusion

These results support our hypothesis that alterations in central H2S signaling are associated with HTN, as centrally administered NaHS alleviated rodent Ang II HTN via reduction of sympathetic tone at night.

Support or Funding Information

TUBITAK 2214‐A and UF CVM.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Orexin Function in DOCA‐Salt Rat Model
Authors: Bigalke Jeremy A , Chen Qing‐Hui , Shan Zhiying
Published: June 2021

Orexin is a neuropeptide with a large range of roles, including sleep/wake regulation, hunger, and more recently blood pressure (BP) regulation. Although the role of orexin in the paraventricular nucleus (PVN) has been investigated in several animal models such as spontaneously hypertensive rats, it remains unclear whether or not it is active in the Deoxycorticosterone‐acetate (DOCA) salt rat model, a model of salt sensitive hypertension. In this study, we investigated the role of PVN orexin signaling on metabolism and fluid homeostasis in the DOCA‐salt rat. Seven to eight week old male Sprague Dawley (SD) rats were split into three groups including DOCA‐salt (n=7), high salt (HS, n=2), or untreated control (n=6) groups. For 3 weeks, the DOCA group was treated with a subcutaneous implantation of a 75mg DOCA pellet, and fed a 1% NaCl/ 0.2% KCl water solution, the HS group was given 1% NaCl drinking water without any implantation, and the control group was left untreated. Following 3 weeks, the HS alone and DOCA‐high salt were put into metabolic cages to measure food and water intake, as well as fecal and urine output for 72 hours. Results show a significant increase in saline intake in the DOCA‐salt group when compared to the HS alone (HS: 46.41±0.52, vs DOCA‐salt: 176.69±29.24 ml, P<0.05), as well as an increased urine output (HS: 25.63±4.46, vs DOCA‐salt: 126.06±5.35 ml, P<0.001), while food intake was not significantly different. Following metabolic cages, all rats were sacrificed and subjected to either real‐time PCR or immunostaining, as well as heart weight measurements. The heart weight to body weight ratio was also measured to be larger in the DOCA‐salt group when compared to the control group (Control: 0.34±0.005%, vs DOCA‐salt: 0.43±0.025%, P<0.005), although there were no significant differences in the HS alone group compared to the others. PCR data showed the mRNA levels of orexin‐1‐receptor (OX1R) was increased by 23%, while vasopressin showed a 2.9 fold increase in the PVN of DOCA‐salt rats compared to controls. Immunostaining data also showed an increase in OX1R as well as vasopressin expression within the PVN. The combination of these results indicate a potential role for PVN orexin system hyperactivity in DOCA‐salt rat pathology, as well as vasopressin expression, salt appetite, and fluid homeostasis.

Support or Funding Information

Songer Award (Bigalke)

NIH R15HL129213 (Shan)

Michigan Technological University and Portage Health Foundation REF (Shan)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Resolving Thrombo‐inflammation following ischemia reperfusion injury
Authors: Gavins Felicity N. E. , Becker Felix , Senchenkova Elena , Ansari Junaid , Vital Shantel
Published: June 2021

Background

The intertwined processes of thrombosis and inflammation resulting in inadequate resolution are major pathophysiological contributors of ischemia reperfusion injury (I/RI). Accumulating data supports the hypothesis that anti‐inflammatory therapies may limit thrombosis and that anti‐thrombotic therapies may reduce vascular inflammation. One such target is the anti‐inflammatory and pro‐resolving endogenous mediator Annexin A1 (AnxA1). Whilst much work has focused on the effect of AnxA1 on leukocyte accumulation during I/RI, less is known regarding its effect on platelets. Here we tested for the first time the therapeutic potential AnxA1 in resolving thrombo‐inflammation after I/RI.

Material and Methods

We used a well‐characterized model of transient middle cerebral artery occlusion with reperfusion (MCAo/R), coupled with pharmacological and genetic approaches. Fluorescence intravital microscopy (IVM) was used to monitor thrombo‐inflammatory responses, along with flow cytometry and ELISAs. (All animal experiments were approved by LSUHSC‐S Animal Care and Use Committee and were conducted in accordance with the American Association for Accreditation of Laboratory Animal Care guidelines and complied with ARRIVE [Animal Research: Reporting In Vivo Experiments]).

Results

IVM revealed heightened platelet adherence and aggregate formation within cerebral microvessels post I/RI. Using AnxA1 knock‐out mice, we found heightened cellular (neutrophil and platelet) activation as well as neutrophil‐platelet aggregate formation within the cerebral microcirculation. Administration of AnxA1 reduced platelet adherence to the inflamed cerebral endothelium following I/RI, along with a reduction in thrombus formation, thereby facilitating the resolution of thrombo‐inflammation post cerebral I/RI.

Conclusion

Herein we found a previously unknown phenomenon that the endogenous anti‐inflammatory and pro‐resolving mediator AnxA1 is able to exert protection through the alteration of platelet responses by orchestrating the delicate platelet phenotype in I/RI from pro‐pathogenic to regulatory. Thus, AnxA1 is a possible therapeutic strategy for effecting the resolution of cerebral thrombo‐inflammation and may therefore provide a novel anti‐thrombo‐inflammatory therapeutic target following ischemic insults.

Support or Funding Information

Dr. Gavins acknowledges the support of the National Institutes of Health/National Heart, Lung, and blood Institute (NIH/NHLBI): HL125572‐01A1.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
CD8 T‐cells regulate macrophage recruitment leading to exacerbated cardiac remodeling
Authors: Pitts Cooper , Clayton Joshua , DeLeon‐Pennell Kristine Y
Published: June 2021

Elevated CD8 T‐cell counts (>1065 cells/mm3) have been linked with increased mortality rate (2‐fold) in patients after myocardial infarction (MI). However, the exact mechanisms by which CD8 T‐cells impair cardiac wound healing post‐MI are not totally understood. We hypothesized that CD8 T‐cells regulate the inflammatory response leading to decreased survival and cardiac function post‐MI. Permanent occlusion of the left anterior descending artery was performed on C57BL/6J (WT; 4.5±0.1 months of age; n=6 males and females/day) and CD8atm1Mak mice (CD8KO; 4.1±0.2 months of age; n=6 males and females/day). CD8KO mice had improved day 7 survival (73%) compared to WT mice (50%; p<0.05). Interestingly, 100% of the CD8KO mice who died did so due to cardiac rupture of the left ventricle (LV; rupture/total deaths; 3/3) whereas 50% of WT animals (5/10; p<0.05) died of cardiac rupture. Strain analysis showed that as early as post‐MI day 3, global longitudinal strain was improved in CD8KO animals (−5.3±0.1%) compared to WT (−2.8±0.1%; p<0.05) suggesting WT mice have increased myocardial wall stress. Echocardiography measured at day 7 post‐MI showed improved cardiac physiology, as indicated by fractional shortening in CD8KO mice (11±2%) compared to WT mice (5±1%; p<0.05) implicating CD8 T‐cells as a negative regulator of wound healing during the later phase post‐MI. Proteomic plasma analysis of 111 proteins indicated CD8KO had elevated immune response at both post‐MI day 3 and day 7. This was supported by increased macrophage numbers at day 3 post‐MI in CD8KO animals compared to WT mice (p<0.05) as shown by Mac3 staining of the infarct. Collagen within the infarct was also elevated at day 3 in CD8KO compared to WT (p<0.05) as shown by picosirius red staining implicating CD8 T‐cells in regulation of myocardial fibrosis. In conclusion, CD8 T‐cells regulate macrophage recruitment and impair collagen deposition during the early phase post‐MI leading to adverse cardiac remodeling and decreased survival.

Support or Funding Information

This work was supported by the Biomedical Laboratory Research and Development Service of the Veterans Affairs Office of Research and Development Award IK2BX003922 to KYD‐P. The content is solely the responsibility of the authors and does not necessarily represent the official views of the Veterans Administration.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
In Utero Exposure to Electronic Cigarettes Causes Cerebrovascular Impairment with Aging
Authors: Burrage Emily Nicole , Aboaziza Eiman A. , Marshall Kent L. , Parsley Matthew C. , Moore Josh , Johnson Abigail , O'Reilly Juliana , Hare Lance , Dangott Sydney , Tice Abigail L. , Chantler Paul D , Olfert IM
Published: June 2021

Electronic cigarettes (Ecigs) are being promoted as safer than traditional tobacco cigarettes, leading to increased use even during pregnancy. However, the risks associated with Ecig usage during pregnancy on offspring has not been examined.

Rat dams were either exposed to nicotine‐free Ecig vapor (Ecig+0) or ambient air. Rat pups were sacrificed at either one month (Air n=6, Ecig+0 n=5) or three months (Air n=4, Ecig+0 n=3). Maternal Ecig exposure (Joyetech eGrip OLED using 5‐sec puff @ 17.5 W) consisted of 30 puffs over 1‐hour each day, 5 days/week, resulting in an average daily TPM of 120mg/m3. Maternal exposure began on gestational day 2 and continued until pups were weaned. Pups themselves were never directly exposed to Ecig vapors. To examine cerebrovascular function, the middle cerebral artery (MCA) was isolated positioned in a pressurized myobath and exposed to increasing concentrations of acetylcholine (Ach; 10−9M to 10−4M), serotonin (5‐HT; 10−9M to 10−4M) and sodium nitroprusside (SNP; 10−9M to 10−4M).

Significant impairment in MCA dilation to the highest dose of Ach was observed in Ecig+0 compared to air exposed pups at one month of age (13±2% vs. 30±1%, ANOVA p<0.05, respectively) as well as at three months of age (8±1% vs 23±2%, ANOVA p<0.05, respectively). The magnitude of decline in vascular reactivity for Ecig+0 exposed pups at one month and three months was different, although not statistically significant (ANOVA p<0.08). MCA constriction to 5‐HT was similar across all groups and only slight nonsignificant differences were noted in endothelial‐independent dilation in response to SNP.

These data suggest offspring born to mothers who vape during pregnancy have impaired cerebrovascular function, with this impairment worsening with age, suggesting there are risks to the offspring associated with maternal Ecig use.

Support or Funding Information

WVU Cancer Institute Philip R Dino Innovative Research Grant (IMO); APS STRIDE Fellowship (JO); NIHGMS 5U54GM104942‐03 (PDC)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.