Hypertrophy In Rats Research Articles

Evaluation of Anti-hypertrophic Potential of Enicostemma littorale Blume on Isoproterenol Induced Cardiac Hypertrophy.

The main objective of this study is to evaluate the anti-hypertrophic potential of the aqueous extract of Enicostemma littorale (E. littorale) against isoproterenol induced cardiac hypertrophic rat models (male albino Wistar rats) through biochemical investigations. Aqueous extract of E. littorale known for various beneficial properties was administered (100mg/kg, 12days, oral) to isoproterenol (ISO) induced cardiac hypertrophic rats (low ISO-60mg/kg, 12days and high ISO-100mg/kg, 12days, subcutaneous) and were compared with group that was treated with the reference drug, Losartan (10mgkg, administered for 12days, oral). The anti-hypertrophic effect of E. littorale was evaluated by analysing the morphometric indices of the heart, ECG tracings, changes in blood biochemical parameters viz., serum glucose, serum total protein, serum albumin, lipid profile, cardiac specific enzymes (SGOT, SGPT and LDH) and histopathological examination of the heart tissue. The results fundamentally revealed that the plant extract efficiently ameliorated cardiac hypertrophy induced by ISO injected in experimental rats. The outcomes of biochemical investigations of this study highlighted the association between the hypertrophic β-adrenergic receptor signalling (β-AR) and the 5' AMP-activated protein kinase (AMPK)-peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) axis in the metabolism of cardiac fibrosis and hypertrophy. This β-AR/AMPK-PGC1α signalling stem can serve as a key target in ameliorating cardiac hypertrophy through focus on its principal regulators. To add, we also propose that the glycoside, swertiamarin present in this plant with the reported anti-fibrotic potential in liver can be further isolated and evaluated for its anti-hypertrophic potential to treat cardiac hypertrophy.

PKA phosphorylation underlies functional recruitment of sarcolemmal SK2 channels in ventricular myocytes from hypertrophic hearts.

Key points Small‐conductance Ca2+‐activated K+ (SK) channels expressed in ventricular myocytes are dormant in health, yet become functional in cardiac disease.SK channels are voltage independent and their gating is controlled by intracellular [Ca2+] in a biphasic manner. Submicromolar [Ca2+] activates the channel via constitutively‐bound calmodulin, whereas higher [Ca2+] exerts inhibitory effect during depolarization.Using a rat model of cardiac hypertrophy induced by thoracic aortic banding, we found that functional upregulation of SK2 channels in hypertrophic rat ventricular cardiomyocytes is driven by protein kinase A (PKA) phosphorylation. Using site‐directed mutagenesis, we identified serine‐465 as the site conferring PKA‐dependent effects on SK2 channel function.PKA phosphorylation attenuates I SK rectification by reducing the Ca2+/voltage‐dependent inhibition of SK channels without changing their sensitivity to activating submicromolar [Ca2+]i.This mechanism underlies the functional recruitment of SK channels not only in cardiac disease, but also in normal physiology, contributing to repolarization under conditions of enhanced adrenergic drive. Small‐conductance Ca2+‐activated K+ (SK) channels expressed in ventricular myocytes (VMs) are dormant in health, yet become functional in cardiac disease. We aimed to test the hypothesis that post‐translational modification of SK channels under conditions accompanied by enhanced adrenergic drive plays a central role in disease‐related activation of the channels. We investigated this phenomenon using a rat model of hypertrophy induced by thoracic aortic banding (TAB). Western blot analysis using anti‐pan‐serine/threonine antibodies demonstrated enhanced phosphorylation of immunoprecipitated SK2 channels in VMs from TAB rats vs. Shams, which was reversible by incubation of the VMs with PKA inhibitor H89 (1 μmol L–1). Patch clamped VMs under basal conditions from TABs but not Shams exhibited outward current sensitive to the specific SK inhibitor apamin (100 nmol L–1), which was eliminated by inhibition of PKA (1 μmol L–1). Beta‐adrenergic stimulation (isoproterenol, 100 nmol L–1) evoked I SK in VMs from Shams, resulting in shortening of action potentials in VMs and ex vivo optically mapped Sham hearts. Using adenoviral gene transfer, wild‐type and mutant SK2 channels were overexpressed in adult rat VMs, revealing serine‐465 as the site that elicits PKA‐dependent phosphorylation effects on SK2 channel function. Concurrent confocal Ca2+ imaging experiments established that PKA phosphorylation lessens rectification of I SK via reduction Ca2+/voltage‐dependent inhibition of the channels at high [Ca2+] without affecting their sensitivity to activation by Ca2+ in the submicromolar range. In conclusion, upregulation of SK channels in diseased VMs is mediated by hyperadrenergic drive in cardiac hypertrophy, with functional effects on the channel conferred by PKA‐dependent phosphorylation at serine‐465.

Curcumin alleviates isoproterenol-induced cardiac hypertrophy and fibrosis through inhibition of autophagy and activation of mTOR.

Curcumin has been reported to possess cardioprotective effects. However, the potential molecular mechanism of curcumin is still not clear. The aim of the present study was to investigate the role of curcumin in regulating autophagy and mammalian target of rapamycin (mTOR) signaling in isoproterenol-induced cardiac hypertrophy and fibrosis in the rat. Rats model of cardiac hypertrophy and fibrosis was induced by isoprenaline (5 mg/kg/day, subcutaneous injection), which were treated with or without curcumin (200 mg/kg/day, intragastric administration). Masson's trichrome staining was performed to investigate the effect of curcumin on fibrosis of cardiac hypertrophy rat. The expression of hypertrophic and fibrosis markers was determined by RT-qPCR. The protein expression of autophagic markers, mTOR, and phosphorylated-mTOR (p-mTOR) was performed by Western blotting. Isoprenaline treatment significantly up-regulated the mRNA expression of hypertrophic (ANP and MYH7) and fibrotic (procollagen I and III) markers in the hearts from rats. All of these markers were reversed by curcumin treatment in isoproterenol-treated rats. Histological analysis showed that curcumin attenuated the interstitial fibrosis of heart triggered by isoproterenol. Moreover, isoproterenol significantly reduced the mRNA levels of mTOR and the protein expression of p-mTOR. However, isoprenaline caused a significant induction of the mRNA levels of LC3 and Beclin-1 and the protein expression of LC3-II and Beclin-1, as well as LC3-II/I ratio. Curcumin abolished these isoprenaline-mediated changes in mTOR/autophagy signaling pathway. Our data demonstrated that curcumin targeted mTOR/autophagy axis could attenuate cardiac hypertrophy and fibrosis in a rat model.

Apigenin-induced HIF-1α inhibitory effect improves abnormal glucolipid metabolism in AngⅡ/hypoxia-stimulated or HIF-1α-overexpressed H9c2 cells

BackgroundApigenin, a natural flavonoid compound, can improve the myocardial abnormal glucolipid metabolism and down-regulate the myocardial hypoxia inducible factor-1α (HIF-1α) in hypertensive cardiac hypertrophic rats. However, whether or not the ameliorative effect of glucolipid metabolism is from the reduction of HIF-1α expression remains uncertain. PurposeThis study aimed to investigate the exact relationship between them in angiotensin Ⅱ (Ang Ⅱ)/hypoxia-stimulated or HIF-1α overexpressed H9c2 cells. MethodsTwo cell models with Ang Ⅱ/hypoxia-induced hypertrophy and HIF-1α overexpression were established. After treatment of the cells with different concentrations of apigenin, the levels of total protein, free fatty acids (FFA), and glucose were detected by the colorimetric method, the level of atrial natriuretic peptide (ANP) was detected by the ELISA method, and the expressions of HIF-1α, peroxisome proliferator-activated receptor α/γ (PPARα/γ), carnitine palmitoyltmnsferase-1 (CPT-1), pyruvate dehydrogenase kinase-4 (PDK-4), glycerol-3-phosphate acyltransferase genes (GPAT), and glucose transporter-4 (GLUT-4) proteins were detected by the Western blot assay. ResultsFollowing treatment of the both model cells with apigenin 1–10 μM for 24 h, the levels of intracellular total protein, ANP, and FFA were decreased, while the level of cultured supernatant glucose was increased. Importantly, apigenin treatment could inhibit the expressions of HIF-1α, PPARγ, GPAT, and GLUT-4 proteins, and increase the expressions of PPARα, CPT-1, and PDK-4 proteins. ConclusionApigenin could exert an ameliorative effect on abnormal glucolipid metabolism in AngⅡ/hypoxia-stimulated or HIF-1α-overexpressed H9c2 cells, and its mechanisms were associated with the inhibition of HIF-1α expression and subsequent upregulation of PPARα-mediated CPT-1 and PDK-4 expressions and downregulation of PPARγ-mediated GPAT and GLUT-4 expressions.

A4989 Induction of endoplasmic reticulum stress and variation in expression levels of Zip14 in hypertrophic rat heart

A4989 Induction of endoplasmic reticulum stress and variation in expression levels of Zip14 in hypertrophic rat heart

Effect of Calcium on Slow Force Responses in Isolated Right Ventricle Preparations of Healthy and Hypertrophied Myocardium in Male and Female Rats.

Effect of different Ca2+ concentrations in the bathing solution [Ca2+]o on the parameters of single isometric contraction and slow force response to stretching was studied in isolated preparations of healthy and hypertrophied myocardium of male and female Wistar rats. In all groups of experimental animals, the increase in calcium concentration was followed by a decrease in the myocardium slow response intensity. We revealed a complementary relationship between the current and medium-term systems of myocardial contractility regulation by the length of the myocardium aimed at the maintenance of the constant level during adaptation to the load. Slow responses of the hypertrophied rat heart myocardium were suppressed in comparison with those in the healthy myocardium and their intensity did not depend on animal sex.

NaHS Induced ANP Secretion via PI3K/Akt/NO/cGMP and KATP Channel Pathway

The third gasotransmitter, hydrogen sulfide (H2S), is related to the pathogenesis of cardiovascular diseases. However, it's still not clear about the relation between H2S and cardiac hormone, atrial natriuretic peptide (ANP). The purpose of this study is to investigate the effects of H2S donor on ANP secretion and seek its mechanism using normal and isoproterenol (ISP)‐treated hypertrophied rat atria. Different doses of H2S donors were perfused into isolated beating rat atria, and atrial pressure and ANP secretion were measured. NaHS (1, 10, 50, 100 μM) augmented high stretch‐induced ANP secretion and decreased systolic atrial pressure (SAP) dose‐dependently. Pretreatment with an inhibitor or antagonist for phosphatidylinositol 3‐kinase (PI3K; wortmannin), protein kinase B (Akt; API‐2), nitric oxide synthase (NOS; L‐NAME), soluble guanylyl cyclase (sGC; ODQ), and KATP channel (glibenclamide) blocked the augmentation of ANP secretion induced by NaHS. The high stretch‐induced ANP secretion was stimulated by Na2S but was not changed by GYY4137 and sodium thiosulfate. H2S synthesis enzyme inhibitor (DL‐propargylglycine, PAG) did not show any significant changes in atrial parameters. However, the response of ANP secretion to NaHS markedly attenuated and PAG suppressed ANP secretion in ISP‐treated rat atria. The expression of eNOS protein was decreased but the expression of cardiomyocyte‐specific H2S producing enzyme, cystathione γ‐lyase, was not changed in ISP‐treated rat ventricles. These findings clarify that NaHS stimulates ANP secretion through PI3K/Akt/NO/cGMP and KATP channel pathway. The modification of ANP secretion by NaHS and H2S synthesis enzyme inhibitor suggests the possible role of endogenous H2S in the pathogenesis of cardiac hypertrophy.Support or Funding InformationSupported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIP) (NO 2017‐R1A2B‐4002214).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Protective effect of hydrogen-rich saline on pressure overload-induced cardiac hypertrophyin rats: possible role of JAK-STAT signaling

BackgroundMolecular hydrogen has been shown to have antioxidant effect and have been used to prevent oxidative stress-related diseases. The goal of this study was to explore if hydrogen-rich saline (HRS) plays a cardioprotective effect on abdominal aortic constriction (AAC) induced cardiac hypertrophy in rats. 60adult Sprague–Dawley rats received surgically the AAC for 6-week. After the surgery, the rats were randomly divided into 4 groups (15 for each):1: sham-operated (sham); 2: AAC-model; 3: AAC + Low HRS (LHRS); and 4: AAC + High HRS (HHRS). The rats in sham and AAC-model groups were treated with normal saline intraperitoneally, while rats in LHRS and HHRS groups were intraperitoneally treated with 3 or 6 mL/kg HRS daily, respectively, for 6-week.ResultsThe ratios of HW/BW and LVW/BW were shown in an order of Model > LHRS > HHRS > SHAM groups. The cardiac hypertrophy was also manifested with increased expressions of atrial natriuretic peptide (ANP), brain natriuretic peptides (BNP) and fibrosis of cardiac tissues in AAC-model group, which could likewise be restrained in LHRS and HHRS groups. Moreover, the JAK-STAT (Janus Kinase-Signal transducers and activators of transcription) signaling molecule expressions were decreased with HRS treatment.ConclusionsOur results showed a protective effect of HRS on pressure overload-induced cardiac hypertrophy in rats, which may be associated to a decreasing in JAK-STAT signaling pathway.

Protective effect of Dendrobium candidum on isoproterenol induced cardiac hypertrophy in rats

To study the effect and mechanism of Dendrobium candidum on isoproterenol-induced myocardial hypertrophy in rats, 60 healthy SD rats(30 males and 30 females) were randomly divided into 5 groups(12 in each group): normal group, model group, three D. candidum preventive administration groups(0.09, 0.18, 1.1 g·kg⁻¹). Except for the normal group, rats of other groups were injected back subcutaneously with ISO(5 mg·kg⁻¹) for 10 consecutive days. At the same time, preventive administration groups began to give different doses of the sample for 30 days and model group began to give normal saline. Left ventricular systolic pressure(LVSP) was measured in each group by common carotid artery cannulation, and the left ventricle(LW)/tibia length, heart weight index(HWI) and myocardial hydroxyproline(Hydro) content were calculated. Myocardial tissue HE staining and Masson staining were used to observe the myocardial structure and the degree of myocardial fibrosis respectively. Atrial natriuretic peptide(ANP), brain natriuretic peptide(BNP), and cardiac troponin I(cTN-I) concentration were measured by enzyme-linked immunosorbent assay(ELISA). The results showed that as compared with the normal group, the levels of ANP, BNP and cTN-I in plasma were significantly increased in ISO-induced hypertrophic rats; as compared with the model group, D. candidumcan inhibit ISO-induced ventricular pressure and ventricular hypertrophy, reduce myocardial collagen synthesis, improve myocardial fibrosis and ventricular remodeling, and significantly down-regulate ANP, BNP and cTN-I levels in plasma. This study shows that D. candidum has a protective effect on isoproterenol-induced cardiac hypertrophy.

The Role and Determinants of Biphasic Regulation of SK Channels by Ca2+ in Hypertrophic Rat Ventricular Cardiomyocytes

The Role and Determinants of Biphasic Regulation of SK Channels by Ca2+ in Hypertrophic Rat Ventricular Cardiomyocytes

Enhanced Redox State and Efficiency of Glucose Oxidation With miR Based Suppression of Maladaptive NADPH-Dependent Malic Enzyme 1 Expression in Hypertrophied Hearts.

Metabolic remodeling in hypertrophic hearts includes inefficient glucose oxidation via increased anaplerosis fueled by pyruvate carboxylation. Pyruvate carboxylation to malate through elevated ME1 (malic enzyme 1) consumes NADPH necessary for reduction of glutathione and maintenance of intracellular redox state. To elucidate upregulated ME1 as a potential maladaptive mechanism for inefficient glucose oxidation and compromised redox state in hypertrophied hearts. ME1 expression was selectively inhibited, in vivo, via non-native miR-ME1 (miRNA specific to ME1) in pressure-overloaded rat hearts. Rats subjected to transverse aortic constriction (TAC) or Sham surgery received either miR-ME1 or PBS. Effects of ME1 suppression on anaplerosis and reduced glutathione (GSH) content were studied in isolated hearts supplied 13C-enriched substrate: palmitate, glucose, and lactate. Human myocardium collected from failing and nonfailing hearts during surgery enabled RT-qPCR confirmation of elevated ME1 gene expression in clinical heart failure versus nonfailing human hearts (P<0.04). TAC induced elevated ME1 content, but ME1 was lowered in hearts infused with miR-ME1 versus PBS. Although Sham miR-ME1 hearts showed no further reduction of inherently low anaplerosis in normal heart, miR-ME1 reduced anaplerosis in TAC to baseline: TAC miR-ME1=0.034±0.004; TAC PBS=0.081±0.005 (P<0.01). Countering elevated anaplerosis in TAC shifted pyruvate toward oxidation in the tricarboxylic acid cycle. Importantly, via the link to NADPH consumption by pyruvate carboxylation, ME1 suppression in TAC restored GSH content, reduced lactate production, and ultimately improved contractility. A maladaptive increase in anaplerosis via ME1 in TAC is associated with reduced GSH content. Suppressing increased ME1 expression in hypertrophied rat hearts, which is also elevated in failing human hearts, reduced pyruvate carboxylation thereby normalizing anaplerosis, restoring GSH content, and reducing lactate accumulation. Reducing ME1 induced favorable metabolic shifts for carbohydrate oxidation, improving intracellular redox state and enhanced cardiac performance in pathological hypertrophy.

Effect of Combined Intervention of Electroacupuncture and Astragaloside IV on Myocardial Hypertrophy and TGF-β 1/Smad Signaling in Rats with Myocardial Fibrosis

To observe the effect of combined intervention of electroacupuncture (EA) and astragaloside IV(ASIV) on cardiac hypertrophy and transforming growth factor β 1 (TGF-β 1)/Smad signaling in isoproterenol (ISO) induced cardiac hypertrophy rats, so as to investigate its underlying mechanisms in improving myocardial fibrosis. A total of 50 SD rats were randomly divided into 5 groups: normal control, model (ISO), Propranolol (PRO)，ASIV and EA＋ASIV groups (n＝10 in each group). The myocardial fibrosis model was established by intraperitoneal injection (i.p.) of ISO (10 mg·kg－1·d－1), once daily for 30 days. Rats of the control group were given normal saline (i.p.), those of the PRO group given with PRO (40 mg·kg－1·d－1, gavage), and those of the ASIV and EA＋ASIV groups were treated by gavage of ASIV (40 mg·kg－1·d－1), once daily for 30 days. EA (20 Hz, 6 V) was applied to bilateral "Neiguan" (PC 6) for 10 min, once every day for 30 d. The heart mass index (HMI, whole heart weight/body weight) and left ventricular (LV) mass index (LVMI, weight of the LV/body weight) were calculated to assess the state of cardiac hypertrophy. The enzyme linked immunosorbent assay (ELISA) was used to determine the levels of procollagen I carboxy-terminal propeptide (PICP，a marker of extracellular matrix remodeling) and carboxyterminal telopeptide of type I collagen (ICTP, a metabolite of type I collagen) in serum, and Western blot was used to test protein contents of TGF- β 1, Smad 2 / 3, Smad 4, Smad 7 in the left ventricle tissue of the heart. After modeling, the HMI and LVMI, serum PICP and ICTP contents and the expression levels of myocardial TGF-β 1, Smad 2/3 and Smad 4 proteins were significantly increased in the model (ISO) group (P<0.05), suggesting a deposition of collagen and cardiac hypertrophy, and were considerably decreased in PRO, ASIV and EA＋ASIV groups after the intervention (P<0.05). The expression level of myocardial Smad 7 protein was significantly lower in the model group than in the normal control group (P<0.05), and significantly up-regulated in PRO, ASIV and EA＋ASIV groups (P<0.05). Sirius Red staining of the left ventricular myocardium showed a dense deposition of collagen and a severer myocardial fibrosis in the model group, and a relatively lighter fibrosis in the PRO, ASIV and EA＋ASIV groups. The therapeutic effects of EA＋ASIV were comparable to those of PRO, and were significantly superior to those of ASIV in down-regulating HMI, serum ICTP, and myocardial Smad 2/3 and Smad 4 expression and up-regulating Smad 7 protein (P<0.05). There were no significant differences among the PRO, ASIV and EA＋ASIV groups in LVMI, PICP and TGF-β 1 levels, and between the PRO and EA＋ ASIV groups in HMI, ICTP, Smad 2/3, Smad 4 and Smad 7 levels (P> 0.05). EA stimulation of PC 6 combined with ASIV can relieve cardiac hypertrophy and myocardial fibrosis in rats, which may be associated with its effects in regulating myocardial TGF-β 1/Smad signaling pathway.

Apelin‑13 promotes cell proliferation in the H9c2 cardiomyoblast cell line by triggering extracellular signal‑regulated kinase 1/2 and protein kinase B phosphorylation.

Apelin‑13 (APL‑13), a peptide hormone that serves as a ligand for G‑protein coupled receptors, has been demonstrated to be highly expressed in left ventricular hypertrophy rat models. It has been implicated in cardio‑protection under pathological states. The present study aimed to assess the physiological proliferation effect of APL‑13 in cultured H9c2 cardiomyoblast cells, and to elucidate the underlying mechanisms. Cell proliferation was determined by MTT assay. The extracellular signal‑regulated kinase (ERK) 1/2 and protein kinase B (Akt) signaling pathway was identified, and protein expression levels were detected using western blot analysis. The results demonstrated that APL‑13 markedly increased cell proliferation. Western blotting results suggested that APL‑13 significantly enhanced the expression of phosphoinositide ERK1/2 and Akt activation in a dose‑dependent manner. U0126 (10µM; ERK1/2 inhibitor) and/or 10µM LY294002 (Akt inhibitor) were used to help to determine the APL‑signaling mechanism. As a result, LY294002 and U0126 partially blocked the APL‑13 induced H9c2 proliferation. In conclusion, these data suggested that APL‑13 has a proliferative effect on myocardium cells via the Akt and ERK1/2 signaling pathways, and provide potential novel pharmaceutical targets for cardiovascular disease.

Upregulation of α-enolase protects cardiomyocytes from phenylephrine-induced hypertrophy.

Cardiac hypertrophy often refers to the abnormal growth of heart muscle through a variety of factors. The mechanisms of cardiomyocyte hypertrophy have been extensively investigated using neonatal rat cardiomyocytes treated with phenylephrine. α-Enolase is a glycolytic enzyme with "multifunctional jobs" beyond its catalytic activity. Its possible contribution to cardiac dysfunction remains to be determined. The present study aimed to investigate the change of α-enolase during cardiac hypertrophy and explore its role in this pathological process. We revealed that mRNA and protein levels of α-enolase were significantly upregulated in hypertrophic rat heart induced by abdominal aortic constriction and in phenylephrine-treated neonatal rat cardiomyocytes. Furthermore, knockdown of α-enolase by RNA interference in cardiomyocytes mimicked the hypertrophic responses and aggravated phenylephrine-induced hypertrophy without reducing the total glycolytic activity of enolase. In addition, knockdown of α-enolase led to an increase of GATA4 expression in the normal and phenylephrine-treated cardiomyocytes. Our results suggest that the elevation of α-enolase during cardiac hypertrophy is compensatory. It exerts a catalytic independent role in protecting cardiomyocytes against pathological hypertrophy.

Induction of endoplasmic reticulum stress and changes in expression levels of Zn2+-transporters in hypertrophic rat heart.

Clinical and experimental studies have shown an association between intracellular free Zn2+ ([Zn2+]i)-dyshomeostasis and cardiac dysfunction besides [Ca2+]i-dyshomeostasis. Since [Zn2+]i-homeostasis is regulated through Zn2+-transporters depending on their subcellular distributions, one can hypothesize that any imbalance in Zn2+-homeostasis via alteration in Zn2+-transporters may be associated with the induction of ER stress and apoptosis in hypertrophic heart. We used a transverse aortic constriction (TAC) model to induce hypertrophy in young male rat heart. We confirmed the development of hypertrophy with a high ratio of heart to body weight and cardiomyocyte capacitance. The expression levels of ER stress markers GRP78, CHOP/Gadd153, and calnexin are significantly high in TAC-group in comparison to those of controls (SHAM-group). Additionally, we detected high expression levels of apoptotic status marker proteins such as the serine kinase GSK-3β, Bax-to-Bcl-2 ratio, and PUMA in TAC-group in comparison to SHAM-group. The ratios of phospho-Akt to Akt and phospho-NFκB to the NFκB are significantly higher in TAC-group than in SHAM-group. Furthermore, we observed markedly increased phospho-PKCα and PKCα levels in TAC-group. We, also for the first time, determined significantly increased ZIP7, ZIP14, and ZnT8 expressions along with decreased ZIP8 and ZnT7 levels in the heart tissue from TAC-group in comparison to SHAM-group. Furthermore, a roughly calculated total expression level of ZIPs responsible for Zn2+-influx into the cytosol (increased about twofold) can be also responsible for the markedly increased [Zn2+]i detected in hypertrophic cardiomyocytes. Taking into consideration the role of increased [Zn2+]i via decreased ER-[Zn2+] in the induction of ER stress in cardiomyocytes, our present data suggest that differential changes in the expression levels of Zn2+-transporters can underlie mechanical dysfunction, in part due to the induction of ER stress and apoptosis in hypertrophic heart via increased [Zn2+]i- besides [Ca2+]i-dyshomeostasis.

Change of acetylcholine-sensitive K+ channel current in atrial myocytes of myocardial hypertrophy rat model

Objective To explore the change of acetylcholine-sensitive K+ channel(Kach)current in artial myocytes of myocardial hypertrophy rat model,to provide partial theoretical basis of hypertrophic cardiomyopathy(HCM)complicated with atrial fibrillation(AF). Methods Based on the rat′s hypertrophic model made by clipping both kidneys,we acutely isolated the atrial muscle cell,and used the whole-cell patch clamp technique to test the current of Kach from the atrial myocytes. Results The Kach of experimental group fell by(54.4±5)% compared to the control group,the difference had statistical significance(P<0.05). Conclusion The IKach signifiantly decreased in the rat hypertrophic model,which was similar with past AF model investigative result,supposing the connection of Kach and cardiomyopathy complicated AF. Key words: Myocardial hypertrophy; Hypertrophic cardiomyopathy; Atrial fibrillation; Acetylcholine-sensitive K+ channel

Abstract 286: ATPase Inhibitory Factor 1 Regulates Glycolysis in Cardiomyocytes

It is known that pathological cardiac hypertrophy is associated with decreased fatty acid oxidation (FAO) and increased reliance on glycolysis. This metabolic remodeling is generally recognized and considered ultimately maladaptive for sustaining myocardial energetics and function. The mechanisms responsible for the switch are poorly understood but appear to be coupled with impaired mitochondrial function. We found that mitochondrial ATPase inhibitor factor 1 (ATPIF1) was up-regulated (by 4-fold, p=0.01) in the failing heart induced by TAC surgery as well as in hypertrophied adult rat cardiomyocytes (CMs) induced by 10 uM phenylephrine (PE, by 3-fold, p=0.01), but not in the physiological hypertrophied heart (p=0.9). ATPIF1 is an inhibitor of ATPase in the mitochondrial ATP synthase, or Complex V. It was shown that upregulation of ATPIF1 increased glycolysis in non-cardiomyocytes. Here we wish to test the hypothesis that ATPIF1 stimulates glycolysis in the heart undergoing pathological hypertrophy. Overexpress ATPIF1 (OE) by adenovirus vector in CMs, which mimics the up-regulation of ATPIF1 induced by PE, increased glycolysis in CMs (control: 8.54±0.61 mpH/min, OE: 11.46±0.48 mpH/min; p=0.006), while ATPIF1 knockdown (KD) by shRNA in PE treated CMs abolished the upregulation of glycolytic capacity (control: 29.27±1.31 mpH/min, KD: 28.61±1.81 mpH/min, control-PE: 40.12±1.33 mpH/min, KD-PE: 32.51±2.09 mpH/min; p=0.003). ATPIF1OE or pathological hypertrophy induced by PE increased the expression of glycolytic enzymes, e.g. GAPDH, GLUT1, LDHA and PKM2, which were abolished by ATPIF1KD. We also found that elevation of ATPIF1 decreased mitochondrial oxygen consumption rate (OCR) and promoted mitochondrial reactive oxygen species (mtROS) production, while the generation of mtROS caused by PE was abrogated in the ATPIF1 deficient CMs. Blockade of mtROS production by expressing mitochondria localized catalase suppressed the increased glycolysis in ATPIF1-OE CMs. In conclusion, our results identify ATPIF1 as a regulator of glycolysis in pathological cardiac hypertrophy, which may provide a mechanism for the regulation of glycolysis through mitochondrial bioenergetics.

Abstract 245: Heart Failure With Preserved Ejection Fraction- a New Animal Model and a Novel Mechanistic Perspective

Diagnosis of heart failure with preserved ejection fraction (HFpEF) is becoming the prevalent form of disease type. Established treatments for heart failure with reduced ejection fraction (HFrEF) have proven minimally effective for HFpEF. This may reflect differences in underlying cardiomyocyte pathophysiology. In HFrEF cardiomyocyte phenotype is characterized by impaired contractile response and diminished systolic Ca 2+ levels. Studies of intact HFpEF-derived cardiomyocytes are lacking. Progress in understanding the etiology of HFpEF has been impeded by limited availability of appropriate pre-clinical models. Our goal was to validate and characterize a new rodent model of HFpEF, the ‘Hypertrophic Heart Rat’ (HHR), undertaking longitudinal investigations to delineate the associated cardiac and cardiomyocyte pathophysiology. The selectively inbred HHR strain exhibits adult cardiac enlargement (without hypertension) and premature mortality (40% at age 50 weeks) compared to the control ‘Normal Heart Rat’ (NHR). Echo analyses established that cardiac hypertrophy was characterized in vivo by maintained systolic parameters (i.e. ejection fraction at 85-90% control) with marked diastolic dysfunction (i.e. increased E/E’). Diastolic dysfunction was detectable in young adult HHR, as an early disease marker. Histological examination identified regions of focal reparative fibrosis in HHR hearts, most prominent in the transverse midwall area of the left ventricle adjacent to the interventricular septum. Evaluation of cardiomyocyte function using left ventricular myocytes isolated from hearts of 30 week (prefailing ) HHR revealed a hypercontractile phenotype with high Ca 2+ operational levels and arrhythmogenic vulnerability. HHR cardiomyocytes exhibited dramatically increased L-type Ca 2+ channel current density (almost 2-fold), and molecular analyses identified hyperphosphorylation of key sarcoplasmic reticulum Ca 2+ regulatory proteins, without change in total phospho-titin. These findings strongly support the contention that HFpEF and HFrEF can have different underlying cardiomyocyte phenotypes. New directions for HFpEF therapies are indicated, and the HHR provides a new model for preclinical HFpEF investigations.

Abstract 431: Upregulation of Small Conductance Ca 2+ Activated K + Channels Promotes Formation of Mitochondrial Supercomplexes and Increases Respiration Reserve

Introduction: Small conductance calcium (Ca 2+ )-activated K + (SK) channels are present in the inner mitochondria membrane of ventricular cardiomyocytes. We recently reported that pharmacological enhancement of mito-SKs protects from Ca 2+ dependent arrhythmia by reducing production of reactive oxygen species (ROS) by mitochondria thereby normalizing oxidation state and activity of Ca 2+ release channels ryanodine receptors in hypertrophic rat hearts. Recent advances demonstrate that respiratory components of the mitochondrial electron transport chain can organize into supercomplexes (SCs) resulting in more efficient electron transfer and reduced generation of ROS. Hypothesis: Functional upregulation of mito-SK channels protects from Ca 2+ -dependent arrhythmia via promotion of mitochondrial SC formation and improved respiration. Methods: H9C2 myoblasts were infected with adenoviral vectors to overexpress SK2 and Cox7rp. After 48 hours in culture, cell respiration was measured using Seahorse XFe Analyzer; ROS was measured using general ROS indicator CFDA; and mitochondria were isolated to assess SC formation using Blue-Native electrophoresis (BN-PAGE). The effects of SK enhancer NS309 (2 μM, 10 min.) on SC formation were studied in Langendorff perfused rat hearts with hypertrophy induced by thoracic aortic banding (TAB). Results: rSK2 overexpression in H9C2s reduced intracellular ROS ~ 30%, enhanced maximum respiration rate vs. controls (0.74±0.04 and 0.58±0.03 pM/min/μg respectively, p&lt;0.05); and caused a robust shift (&gt;2 fold) of complex IV towards large mitochondrial SCs (~1.2 mDa). As a control, similar results were obtained with overexpression of Cox7rp protein that promotes SC formation. In addition, BN-PAGE experiments in isolated rat hearts demonstrated that robust decrease in high MW Complexes I and IV in hypertrophy was fully restored to control levels upon treatment with SK enhancer NS309. Conclusion: Upregulation of SK channels reduces mito-ROS by an increase in respiration reserve which is linked to enhanced mitochondrial SC formation. Our data suggest that targeting mito-SK channels has therapeutic potential to treat cardiac arrhythmia in conditions accompanied by oxidative stress.

Effects of a Proteasome Inhibitor on Cardiomyocytes in a Pressure-Overload Hypertrophy Rat Model: An Animal Study.

BackgroundThe ubiquitin-proteasome system (UPS) is an important pathway of proteolysis in pathologic hypertrophic cardiomyocytes. We hypothesize that MG132, a proteasome inhibitor, might prevent hypertrophic cardiomyopathy (CMP) by blocking the UPS. Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and androgen receptor (AR) have been reported to be mediators of CMP and heart failure. This study drew upon pathophysiologic studies and the analysis of NF-κB and AR to assess the cardioprotective effects of MG132 in a left ventricular hypertrophy (LVH) rat model.MethodsWe constructed a transverse aortic constriction (TAC)-induced LVH rat model with 3 groups: sham (TAC-sham, n=10), control (TAC-cont, n=10), and MG132 administration (TAC-MG132, n=10). MG-132 (0.1 mg/kg) was injected for 4 weeks in the TAC-MG132 group. Pathophysiologic evaluations were performed and the expression of AR and NF-κB was measured in the left ventricle.ResultsFibrosis was prevalent in the pathologic examination of the TAC-cont model, and it was reduced in the TAC-MG132 group, although not significantly. Less expression of AR, but not NF-κB, was found in the TAC-MG132 group than in the TAC-cont group (p<0.05).ConclusionMG-132 was found to suppress AR in the TAC-CMP model by blocking the UPS, which reduced fibrosis. However, NF-κB expression levels were not related to UPS function.