bluuebunny/crossref_metadata_2025_split huggingface dataset:
VEGF/eNOS‐mediated Cardiomyocyte Proliferation: a Novel Mechanism to Regulate Cardiovascular Regeneration in Mice and Humans
Authors: GENTILE CARMINE , Chuang Christine Y , Drake Christopher J , Davies Michael J 
Published: November 2023

An absence of cardiomyocyte proliferation in adult hearts limits their regeneration potential following tissue damage arising from adverse cardiac events such as a myocardial infarction or heart failure. There is considerable interest, and therapeutic potential, in unveiling mechanisms that enhance heart regeneration using stem cells and progenitor cells. Our studies have previously demonstrated that vascular endothelial growth factor (VEGF) regulates proliferation of cardiomyocyte progenitor cells via endothelial nitric oxide synthase (eNOS) in murine embryos. We have now investigated the role played by the VEGF/eNOS signalling pathway by evaluating cardiomyocyte proliferation in both WT and eNOS KO mouse embryos, and in human heart samples. Confocal analysis of WT and KO embryonic hearts showed no difference at embryonic stage E8.0 by using antibodies against markers of progenitor cells in the heart (VEGFR2/Flk1), cardiomyocytes (smooth muscle α‐actin), endothelial cells (CD31), and cell proliferation (phospho‐histone H3). In contrast, cardiomyocyte proliferation was reduced in E8.5 and E9.5 NOS3 KO embryos when compared to WT controls. Heart tube formation was also impaired. Consequently, E17 NOS3 KO embryos had smaller hearts (approximately 20% decrease of the volume) compared to WT controls. To translate our findings to humans, we analysed human heart biopsies from young and older donors using similar antibodies. Confocal analysis showed, for the first time, a correlation between eNOS expression and cardiomyocyte proliferation in a young human heart. To further investigate the role played by the VEGF/eNOS signaling pathway in human hearts, we developed an in vitro 3D co‐culture model of human endothelial cells, cardiomyocytes and fibroblasts, the human cardiac spheroid (hCS). A higher level of extracellular matrix (ECM) synthesis was observed in hCSs generated by young cells compared to hCSs generated by older cells. Interestingly, VEGF‐treatment of older hCSs increased ECM synthesis compared to untreated hCSs controls, demonstrating therapeutic potential in enhancing the activity of the VEGF/eNOS signaling pathway in hCSs. In conclusion, our data suggest that a similar role is played by the VEGF/eNOS signaling pathway in mice and human cardiomyocyte proliferation. Current studies are focusing on identifying molecular targets of the VEGF/eNOS signaling pathway in the regulation of human cardiomyocyte proliferation as novel therapeutics to induce tissue regeneration in diseased hearts.Support or Funding InformationThis study was supported by AHA 11PRE7530048 and Marcus Blackmore Fellowship to CG
Inhibition of the Sodium‐Glucose Co‐Transporter Protein Type 2 Decreases Oxidative Stress and Improves Cardiac Remodeling in Rats with Diabetes Mellitus
Authors: Rosa Camila Moreno , Campos Dijon Henrique Salome , Reyes David Rafael , Pagan Luana Urbano , Gomes Mariana Janini , Damatto Felipe , Fernandes Ana Angélica Henrique , Okoshi Marina Politi , Okoshi Katashi 
Published: November 2023

PurposeA new class of antidiabetic drugs aims to inhibit the sodium‐glucose co‐transporter type 2 (SGLT2). Thus, SGLT2 inhibition results in a reduction of hyperglycemia by decreasing reabsorption of glucose and increasing urinary excretion. Therefore, this study aimed to analyze the effects of SGLT2 inhibition by dapagliflozin (DAPA) on cardiac remodeling in rats with diabetes mellitus (DM).MethodsMale Wistar rats with 450 g were divided into four groups: control (CTL, n=15); CTL+DAPA (n=15); DM (n=15); DM+DAPA (n=15). Diabetes was induced by streptozotocin (40 mg/kg, i.p., single dose). CTL+DAPA and DM+DAPA groups received DAPA (5 mg/kg/day, added to the rat chow) for 8 weeks. In vivo cardiac structures and functions were assessed by echocardiogram. Oxidative stress was evaluated in left ventricular myocardial tissue by measuring the activity of superoxide dismutase (SOD), glutathione peroxidase (GSH‐Px), catalase, and lipid hydroperoxide (LPO).StatisticsComparisons were performed by ANOVA ‐ 2×2 factorial (p<0.05).ResultsCompared to the DM group, DM+DAPA group presented higher body weight (DM: 343±52; DM+DAPA: 389±46 g; p<0.05), lower blood glucose (DM: 487±80; DM+DAPA: 172±89 mg/dL; p<0.05), and lower systolic blood pressure (DM: 140±7; DM+DAPA: 133±7 mmHg; p<0.05). Echocardiography showed lower left ventricular (LV) and left atrial diameter, and improved LV systolic and diastolic function in DM+DAPA compared to the DM group. Activities of SOD (DM: 13.7±3.90; DM+DAPA: 23.5±4.12 nmol/mg protein; p<0.05), GSH‐Px (DM: 19.7±5.07; DM+DAPA: 33.5±6.84 nmol/mg tissue; p<0.05), and catalase (DM: 85.4±23.0; DM+DAPA: 117±20.1 μmol/g tissue; p<0.05) were increased, and LPO levels (DM: 385±53.8; DM+DAPA: 304±39.8 nmol/g tissue; p<0.05) decreased in the DM+DAPA, compared to the DM group.ConclusionInhibition of the sodium‐glucose co‐transporter type 2 by dapagliflozin decreases oxidative stress and improves cardiac remodeling in rats with diabetes mellitus.Support or Funding InformationFAPESP, and CNPq.
Acute GPER1 Activation During Reperfusion Protects the Myocardium Against Ischemia/Reperfusion Injury by inhibiting of the mPTP Opening and Mitophagy
Authors: Feng Yansheng , Kabir E. Mohammad , Madungwe Ngonidzashe Benson , Bopassa Jean C. 
Published: November 2023

IntroductionWe recently found that acute pre‐ischemic estrogen (17 b‐estradiol) treatment induces cardioprotection against ischemia/reperfusion (I/R) injury mainly via G protein‐coupled estrogen receptor1 (GPER1) activation. We, furthermore, showed that pre‐ischemic estrogen‐GPER1 effects preserve mitochondrial integrity and function via MEK/ERK/GSK‐3b pathway.HypothesisWe investigated whether acute post‐ischemic estrogen (PI‐E2) treatment can also induce cardio‐protective effects via GPER1 activation and determine whether PI‐E2 action involves the reduction of mitophagy.MethodsMale and ovarectomized female Sprague‐Dawley rats were anesthetized with ketamine (80 mg/kg i.p.) and xylazine (8 mg/kg i.p.). Hearts were subjected to 35 min of the left anterior descending (LAD) artery occlusion, followed by 180 min reperfusion. An estrogen bolus (50 mg/kg body weight) or PBS (same volume) was applied via the femoral vein 5 min before reperfusion, and GPER1 antagonist, G15, was given 10 min before estrogen. Area at risk (AAR) was identified using Evans Blue dye and myocardial infarct size assessed by TTC staining method. Mitochondrial calcium retention capacity (CRC) required to induce mitochondrial permeability transition pore (mPTP) opening was assessed after 20 min reperfusion. Expression levels of LC3I LC3II; p62; PINK1; and pERK were detected by western blot in whole cell lysates. Parkin and ubiquitinated protein were measured by Western blot in cytosolic and mitochondrial fractions.ResultsWe found that PI‐E2 treatment reduced myocardial infarct size normalized to the AAR or to the whole LV and increased mitochondrial CRC compared to untreated. We also found that PI‐E2 treatment decreased the level of LC3II, increased the level of p62 and pERK1/2, inhibited the translocation of Parkin to mitochondria and decreased ubiquitinated proteins in mitochondria compared to control. Interestingly, all these PI‐E2 effects were abolished by addition of G15.ConclusionAcute post‐ischemic GPER1 activation by estrogen treatment induces cardioprotective effects against I/R injury. PI‐E2 effects through GPER1 involve the inhibition of mitophagy and reduction of mitochondrial proteins ubiquitination. Together, these post‐ischemic estrogen effects via GPER1 lead to inhibition of the mPTP opening, and subsequently cardioprotection.Support or Funding InformationVoelcker Fund
Impact of Static and Pulsatile Blood Pressure on Cerebral Blood Flow Regulation
Authors: Tarumi Takashi , Zhang Li , Liu Jie , Xing Changyang , Turner Marcel , Repshas Justin , Lewis Tammy , Zhang Rong 
Published: November 2023

BackgroundElevated blood pressure (BP) is a major risk factor for stroke and cognitive impairment in older adults. BP consists of static and pulsatile components that may have differential impacts on cerebral hemodynamics. The objective of this study is to investigate the main and interaction effects of static and pulsatile BP components on cerebral blood flow (CBF), perfusion, and pressure‐flow relations.MethodsWe tested 126 subjects aged 55–80 years (mean age = 65±6 SD). Static and pulsatile BP components were defined as diastolic BP and pulse pressure respectively, as measured by arterial tonometry from the carotid artery. Global CBF, regional brain perfusion, and vascular resistance were measured by phase‐contrast MRI and arterial spin labeling. Transfer function metrics of cerebral dynamic pressure‐flow relations were calculated from the cardiac and lower frequencies to quantify cerebrovascular impedance and autoregulation.ResultsCBF pulsatility increased dose‐dependently with pulse pressure while global CBF and regional perfusion remained at the similar levels. Figure shows the main and interaction effects of static and pulsatile BP components on cerebral pressure‐flow relations. Cerebrovascular resistance increased with diastolic BP whereas the modulus of impedance increased with both diastolic BP and pulse pressure. There were significant main and interaction effects of diastolic BP and pulse pressure on transfer function phase of cerebral autoregulation. These findings remained statistically significant after adjusting for covariates (e.g. age).ConclusionsElevations of static and pulsatile BP components increase cerebrovascular resistance/impedance and alter dynamic cerebral autoregulation in middle‐aged and older adults, which potentially may have impact on brain structure and function.Support or Funding InformationThis study was supported by the National Institute of Health (R01AG033106 and R01HL102457) and the American Heart Association (14POST20140013).
Induction of Heme Oxygenase with Hemin Improves Impaired Insulin Signaling and Compromised Cardiac Structure and Function in Obese Zucker Rats
Authors: Tiwari Shuchita , Ndisang Joseph Fomusi 
Published: November 2023

BackgroundDiabetic cardiomyopathy is characterized by cardiac dysfunction and impaired insulin signaling. The mechanisms that drive obesity and insulin‐resistant mediated cardiomyopathy are not completely elucidated. Accordingly, the aim of this study was to examine the effects of upregulating the heme‐oxygenase (HO) on obesity related cardiac dysfunction and impaired insulin signaling in obese Zucker rats, a model characterized by insulin resistance and cardiomyopathy.MethodsImmunohistochemistry, spectrophotometry, enzyme‐linked immunosorbent assay (ELISA) and Western blotting were used. In addition non‐invasive echocardiography and invasive left ventricular (LV) cannulation was employed to evaluate the hemodynamic, echocardiographic and electrophysiological parameters of the myocardium. HO was enhanced with hemin or blocked with stannous mesoporphyrin.ResultsOur data demonstrated that administering hemin to obese Zucker rats; (i) enhanced important proteins of insulin signaling (IRS‐1, PI3K and GLUT4), (ii) reduced glucose intolerance, (iii) suppressed HOMA‐index, but (iv) increased adiponectin and atrial‐natriuretic‐peptide (ANP). Interestingly, the improvement of insulin sensitivity and glucose metabolism was accompanied by attenuation of cardiomyocytes longitudinal muscle‐fiber thickness, the reduction of diastolic/systolic free‐wall thickness and the suppression of LV hypertrophy. Furthermore, hemin attenuated inflammation/oxidative stress, suppressed fibrosis, extracellular matrix/pro‐fibrotic proteins (collagen‐IV, fibronectin, TGF‐β1) as well as proteins of heart failure such as osteopontin and osteoprotegerin. These were associated with improved cardiac function as evidenced by improved hemodynamics such as mean‐arterial pressure, arterial‐systolic pressure, +dP/dt, arterial‐diastolic pressure and improved electrocardiographic parameters associated with cardiac conduction system including PR‐interval, QRS‐duration, ST‐segment and QT interval. However, the HO blocker, stannous‐mesoporphyrin reversed the cardio‐protective effects of hemin.ConclusionOur findings demonstrates that hemin improved compromised cardiac structure and function in obese Zucker rats. Interestingly, attenuation of inflammation, oxidative stress, extracellular‐matrix/profibrotic proteins, cardiac hypertrophy and fibrosis alongside the potentiation of adiponectin, ANP and proteins of insulin signaling are among the multifaceted mechanisms through which an upregulated HO‐system improves insulin sensitivity, glucose metabolism and diabetic cardiomyopathy. Thus, HO‐inducers may be explored as a novel therapy for diabetic cardiomyopathy.Support or Funding InformationHeart & Stroke Foundation of Saskatchewan, Canada
Long QT‐syndrome Associated Caveolin‐3 Mutations F97C and S141R Alter the Cardiac Transient Outward Kv Current and the L‐type Ca2+ Currents
Authors: Balijepalli Ravi , Tyan Leonid , Keefe Alexis , Foell Jason D , Kamp Timothy J 
Published: November 2023

Caveolae are discrete microdomains in cardiomyocytes that localize many ion channels and signaling proteins. Mutations in the CAV3 gene, encoding caveolin‐3 (Cav‐3), a muscle specific scaffolding protein integral to caveolae in the cardiomyocytes, have been associated with a congenital Long QT syndrome (LQTS9). Previously we have shown that the LQTS associated Cav‐3 mutations, F97C and S141R; increase late sodium channel current in cardiomyocytes contributing to the prolonged action potential duration (APD), but co‐immunoprecipitation using lysates from mouse ventricular myocytes and western blot analysis revealed that Kv4.2, Kv4.3 and Cav1.2 channels also co‐IP with Cav‐3. Thus we evaluated whether disease the LQT associated mutations impacted heterologously expressed Kv4.2, Kv4.3, and Cav1.2 channels in HEK293 cells using whole cell patch clamp electrophysiology. Co‐expression of F97C (40.7 ± 5.9 pA/pF) or S141R (58.8 ± 7.7 pA/pF) significantly (p<0.001) reduced the peak IKv4.2 densities compared to cells co‐expressing either Cav‐3 WT (80 ± 9.1 pA/pF) or pcDNA3.1 (88.6 ± 12.7 pA/pF). The co‐expression of F97C significantly reduced IKv4.3 (213.8 ± 35 pA/pF) compared to Cav‐3 WT (336.7 ± 48.2 pA/pF). F97C also caused a reduction in the activation time constants of Ikv4.2 and Ikv4.3, but did not impact the inactivation. Co‐expression of F97C and S141R significantly (p<0.05) slowed the recovery from inactivation of Kv4.2. On the other hand, co‐expression of S141R significantly increased the peak ICa,L (‐14.8 ± 2.5 pA/pF, p<0.05) compared to Cav‐3 WT (−9.5 ± 2.4 pA/pF). Co‐expression of F97C (−7.6 ± 1.2 pA/pF) did not alter the peak ICa,L density but significantly slowed the calcium dependent inactivation (CDI) of the ICa,L. We conclude that the LQTS linked Cav‐3 mutations F97C and S141R, impart loss‐of‐function effects on Ito but cause a gain‐of function effect on the ICa,L and together these may contribute to the delay in ventricular repolarization and arrhythmogenesis in a setting of LQTS.Support or Funding InformationR01 HL078878
Cytotoxic CD8+ Interleukin (IL)‐17+ T‐cells Play a Role in Hypertension‐Specific Inflammatory Responses Associated with Ovariectomy in Dahl Rats
Authors: Pai Amrita V , West Crystal A , Souza Aline A , Ji Hong , Wu Xie A , Sandberg Kathryn L 
Published: November 2023

Background and ObjectiveOophorectomy is associated with increased body weight (BW) gain and incidence of hypertension (HT) compared with age‐matched women. Increased BW gain and HT coincide with increased inflammatory biomarkers. To separate immune responses due to HT from those due to BW gain, we compared T‐cell subpopulations in normotensive Dahl (DR) and hypertensive Dahl (DS) rats after ovariectomy.MethodsSix week old DS and DR rats were ovariectomized (Ovx) or had sham surgery (Sham). Mean arterial pressure (MAP) was measured in anesthetized rats after 4 and 10 months (mo) acutely by indwelling vascular catheter. Splenic T‐cells were assessed by flow cytometry at 10 mo.ResultsOvariectomy increased the MAP in DS but not DR rats at 4 mo [MAP (mmHg): DS‐Ovx, 143±1.0 vs DS‐Sham, 129±2.9; p=0.0002 & DR‐Ovx, 123±4.4 vs DR‐Sham, 126±1.3; ns n=10–12/group]. At 10 mo, the MAP remained higher in the ovariectomized DS compared to DR rats [MAP (mmHg): DS‐Ovx, 190±12 vs DR‐Ovx, 114±1.2; n=5–8/group; p=0.002]. At 10 mo, ovariectomy increased BW to a similar extent in DS and DR rats [BW gain (g): DS, 179±13 vs DR, 202±14; ns]. The frequency of CD8+IL17a+ T‐cells increased in the ovariectomized DS but not DR rats [Frequency (%) at 10 mo: DS‐Ovx, 9.6±2 vs DR‐Ovx, 1.3±0.4; n=5–8/group; p<0.0001].ConclusionCytotoxic CD8+ IL‐17a+ T‐cells may play a role in the HT‐specific inflammatory response in ovariectomized DS rats.Support or Funding InformationGeorgetown University Medical Center, and Translational Biomedical Science Predoctoral Training Program Georgetown‐Howard Universities Center for Clinical and Translational Science
Pioglitazone Inhibits Pyruvate Metabolism in Hepatocytes
Authors: Shannon Christopher , Daniele Giuseppe , Galindo Cynthia , DeFronzo Ralph , Abdul‐Ghani Mohammed , Norton Luke 
Published: November 2023

The thiazoldinedione compound pioglitazone (PIO) is an effective hypoglycemic agent commonly prescribed for the treatment of type 2 diabetes mellitus (T2DM), the actions of which have typically been assigned to transcriptional regulation through the peroxisome proliferator‐activated receptor gamma (PPAR‐γ). Growing evidence supports the presence of non‐PPAR‐γ related targets of PIO. For example, modulation of glucose metabolism has been demonstrated in human skeletal muscle myotubes through PIO‐mediated inhibition of the newly identified mitochondrial pyruvate carriers MPC1 and MPC2 (Divakuruni et al,. 2013). However, the existence of PPAR‐γ sparing mechanisms of PIO in hepatocytes has not been established. Therefore, we explored the effects of pharmacologically‐relevant concentrations of PIO in cultured hepatocytes (H4IIE) on whole‐cell substrate oxidation (14CO2 capture), mitochondrial ATP production (luciferin‐luciferase assay) and pyruvate/lactate‐driven glucose production, as well as any impact upon the expression of key regulatory gluconeogenic genes. We also assessed the dependence of these effects, if any, on MPC1 and MPC2 using siRNA transfection by electroporation.Four hour PIO treatment (10 μM) suppressed pyruvate oxidation by 29% (P<0.01), but had no impact upon palmitate oxidation. Consistent with this, a dose‐dependent inhibition of mitochondrial ATP production from 500 μM pyruvate/2 mM malate was observed with PIO at 1 μM (27%; p=0.07), 5 μM (36%; P<0.05) and 10 μM (61%; P<0.01) concentrations, but did not influence ATP production from glutamate, succinate or palmitoylcarnitine substrates. Glucose production from pyruvate/lactate was 53% lower following overnight treatment with 10 μM PIO (P<0.01) and this occurred in the absence of any changes in the expression of phosphoenolpyruvate carboxykinase, fructose 1,6‐bisphosphatase, glucose 6‐phosphatase or pyruvate carboxylase. Treatment with the specific inhibitor of mitochondrial pyruvate import UK50995 (5 – 10 μM) similarly inhibited pyruvate oxidation (−85%), pyruvate‐driven mitochondrial ATP production (−66%) and glucose production (−71%). Silencing of MPC1 or MPC2 led to a ~20% reduction in pyruvate oxidation, but only MPC1 silencing significantly impaired glucose production (−48%). The inhibition of pyruvate oxidation by PIO was preserved in cells silenced for MPC1 (P<0.001) and MPC2 (P<0.05), as was the suppression of glucose production in cells silenced for MPC1 (P<0.05).Together, these data demonstrate a role for PIO in the downregulation of liver mitochondrial metabolism that is specific to pyruvate and not dependent on the expression of MPC1 or MPC2. Moreover, these experiments indicate the existence of a novel PPAR‐γ‐independent mechanism through which PIO may suppress hepatic glucose production and hence ameliorate hyperglycemia in T2D patients.
CHANGE IN ENVIRONMENTAL GLUCOSE CONCENTRATION ALTERS ASTROCYTIC INSULIN AND INSULIN RECEPTOR LEVELS
Authors: Cohen Susan , Liu Qiuli , Ryan Robert , Gebremedhin Debebe , Harder David , Rarick Kevin 
Published: November 2023

BACKGROUNDThe actions of insulin in the brain include effects on glucose homeostasis, cell proliferation, cell differentiation, cognition, and memory. Cellular constituents of the neurovascular unit, including the astrocyte, contribute to insulin responses in the brain. Astrocytes are an ideal target cell type for investigation because of their dynamic character, multitude of functions, and their important role in brain homeostasis. We hypothesize that astrocytes respond to environmental glucose changes and could be a source of insulin supply to the microenvironment.OBJECTIVETo investigate astrocyte insulin and insulin receptor response to environmental glucose changes and to determine insulin secretion levels.METHODSNeonatal rat astrocytes in culture were exposed to changes in environmental glucose levels (LOW‐ 0mM, NORMAL‐5.5mM, and HIGH‐25mM) over three passages. Astrocyte whole cell lysates were used to determine message (qPCR) and protein levels (Western blot) for insulin and insulin receptor. Insulin Ultrasensitive ELISA was used to measure NORMAL astrocytic insulin secretion in response to glucose (25mM) and potassium (30mM) stimuli.RESULTSAstrocytes grown in LOW, NORMAL, and HIGH glucose levels resulted in equal cell counts and morphological appearances. Increasing glucose level from LOW, to NORMAL, or to HIGH in astrocytic culture media significantly increased insulin‐1 (exon 1–2) and insulin‐2 (exon 1–2) mRNA levels (One‐way ANOVA, n=4, p<0.05). Increased glucose level also resulted in a positive correlation with increased protein levels of insulin and insulin receptor in astrocytic cell lysate (Pearson Correlation, r=>0.85, p<0.01, n=3, R2=>0.73). Astrocytes secreted measurable levels of insulin (0.02–0.05μg/L) but did not increase significantly with glucose or potassium stimuli.CONCLUSIONSAstrocytes respond to changes in environmental glucose levels with increased insulin and insulin receptor message and protein levels. Astrocytes secrete measurable amounts of insulin but do not respond to stimuli as other insulin secreting cells. We speculate that such insulin responses to glucose levels from the astrocyte may be adaptive for its own homeostasis rather than to affect the microenvironment.Support or Funding InformationFunding from NIH (NHLBI) R01 HL033833 and HL105997 and the Veterans Administration Research Career Scientist Award