scholarly journals Antiapoptotic Actions of Methyl Gallate on Neonatal Rat Cardiac Myocytes Exposed to H2O2

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Sandhya Khurana ◽  
Amanda Hollingsworth ◽  
Matthew Piche ◽  
Krishnan Venkataraman ◽  
Aseem Kumar ◽  
...  
Keyword(s):  
Epigallocatechin Gallate ◽  
Annexin V ◽  
Neonatal Rat ◽  
Methyl Gallate ◽  
Rat Cardiomyocytes ◽  
Therapeutic Value ◽  
Intracellular Ros ◽  
Neonatal Rat Cardiac Myocytes ◽  
Cardiac Disorders ◽  
Polyphenol Intake

Reactive oxygen species trigger cardiomyocyte cell death via increased oxidative stress and have been implicated in the pathogenesis of cardiovascular diseases. The prevention of cardiomyocyte apoptosis is a putative therapeutic target in cardioprotection. Polyphenol intake has been associated with reduced incidences of cardiovascular disease and better overall health. Polyphenols like epigallocatechin gallate (EGCG) can reduce apoptosis of cardiomyocytes, resulting in better health outcomes in animal models of cardiac disorders. Here, we analyzed whether the antioxidant N-acetyl cysteine (NAC) or polyphenols EGCG, gallic acid (GA) or methyl gallate (MG) can protect cardiomyocytes from cobalt or H2O2-induced stress. We demonstrate that MG can uphold viability of neonatal rat cardiomyocytes exposed to H2O2by diminishing intracellular ROS, maintaining mitochondrial membrane potential, augmenting endogenous glutathione, and reducing apoptosis as evidenced by impaired Annexin V/PI staining, prevention of DNA fragmentation, and cleaved caspase-9 accumulation. These findings suggest a therapeutic value for MG in cardioprotection.

Diazoxide triggers cardioprotection against apoptosis induced by oxidative stress

2003 ◽  
Vol 284 (6) ◽  
pp. H2235-H2241 ◽  
Author(s):  
Masashi Ichinose ◽  
Hidetoshi Yonemochi ◽  
Toshiaki Sato ◽  
Tetsunori Saikawa
Keyword(s):  
Oxidative Stress ◽  
Flow Cytometry ◽  
Time Course ◽  
Channel Blocker ◽  
Annexin V ◽  
Neonatal Rat ◽  
Nuclear Staining ◽  
Protective Effects ◽  
Rat Cardiomyocytes ◽  
Induced Apoptosis

Although mitochondrial ATP-sensitive potassium (mitoKATP) channels have been reported to reduce the extent of apoptosis, the critical timing of mitoKATP channel opening required to protect myocytes against apoptosis remains unclear. In the present study, we examined whether the mitoKATP channel serves as a trigger of cardioprotection against apoptosis induced by oxidative stress. Apoptosis of cultured neonatal rat cardiomyocytes was determined by flow cytometry (light scatter and propidium iodide/annexin V-FITC fluorescence) and by nuclear staining with Hoechst 33342. Mitochondrial membrane potential (ΔΨ) was measured by flow cytometry of cells stained with rhodamine-123 (Rh-123). Exposure to H2O2 (500 μM) induced apoptosis, and the percentage of apoptotic cells increased progressively and peaked at 2 h. This H2O2-induced apoptosis was associated with the loss of ΔΨ, and the time course of decrease in Rh-123 fluorescence paralleled that of apoptosis. Pretreatment of cardiomyocytes with diazoxide (100 μM), a putative mitoKATP channel opener, for 30 min before exposure to H2O2 elicited transient and mild depolarization of ΔΨ and consequently suppressed both apoptosis and ΔΨ loss after 2-h exposure to H2O2. These protective effects of diazoxide were abrogated by the mitoKATP channel blocker 5-hydroxydecanoate (500 μM) but not by the sarcolemmal KATP channel blocker HMR-1098 (30 μM). Our results suggest for the first time that diazoxide-induced opening of mitoKATP channels triggers cardioprotection against apoptosis induced by oxidative stress in rat cardiomyocytes.

Antagonism of endothelin-1 inhibits hypoxia-induced apoptosis in cardiomyocytesThis article is one of a selection of papers published in the special issue (part 2 of 2) on Forefronts in Endothelin.

2008 ◽  
Vol 86 (8) ◽  
pp. 536-540 ◽  
Author(s):  
Anjing Ren ◽  
Xiaohong Yan ◽  
Hong Lu ◽  
Jingsong Shi ◽  
Yuanjun Yin ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Annexin V ◽  
Neonatal Rat ◽  
Receptor Subtypes ◽  
Rat Cardiomyocytes ◽  
Endothelin 1 ◽  
Apoptosis Rate ◽  
Hypoxic Conditions ◽  
Induced Apoptosis

Apoptosis is well documented to be a common feature of many pathological processes of the heart. Exogenous endothelin-1 (ET-1) has been shown to be proapoptotic or antiapoptotic, depending on ET-1 concentration, cell type, and the ratio of ETA/ETB receptor subtypes. The role of endogenous ET-1 in cardiomyocyte apoptosis, however, is not clarified. This study observed the effects of the ETA-receptor antagonists BQ610 and BQ123 and the ETB-receptor antagonist BQ788 on hypoxia-induced apoptosis in primary cultured neonatal rat cardiomyocytes. Hypoxic apoptosis was induced by incubating cardiomyocytes in serum-free medium under 3% O2 and 5% CO2 for 24 h and evaluated by TUNEL analysis and flow cytometry. TUNEL analysis showed that the apoptotic cardiomyocytes constituted 24.2% ± 2.2% of the total cells under hypoxic conditions. Treatment with BQ610 (5 μmol/L) significantly reduced the apoptosis rate to 13.2% ± 3.7% (data from 4 independent experiments, p < 0.01 vs. hypoxia). Flow cytometry showed that the percentage of apoptotic cells positively stained with annexin V and propidium iodide was 42.76% ± 4.44% (n = 12) in cultures subjected to hypoxia. BQ123 at 0.04, 0.2, and 1.0 μmol/L dose-dependently reduced the apoptosis rate to 34.00% ± 10.35% (n = 6, p < 0.05), 30.38% ± 8.28% (n = 6, p < 0.01), and 22.89% ± 4.19% (n = 6, p < 0.01), respectively. In contrast, BQ788 did not affect hypoxic apoptosis. These findings suggested that endogenous ET-1 contributed to hypoxia-induced apoptosis in cultured cardiomyocytes, which was mediated by ETA receptors, but not by ETB receptors.

Abstract 1778: Estrogen Protects Cardiomyocytes from Apoptosis by Suppressing p38alpha-mediated p53 activation and by Downregulating a Negative Feedback from p53 on p38beta

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Han Liu ◽  
Ali Pedram ◽  
Ellis R Levin ◽  
Jin K Kim
Keyword(s):  
Molecular Mechanisms ◽  
Ischemia Reperfusion ◽  
Fluorescent Microscopy ◽  
Annexin V ◽  
Neonatal Rat ◽  
Rat Cardiomyocytes ◽  
Downstream Target ◽  
Whole Cell Lysate ◽  
P53 Activation

To investigate the molecular mechanisms of estrogen-mediated protection of cardiomyocytes in hypoxic stress. Estrogen is anti-apoptotic and promotes survival of cardiomyocytes under ischemia-related stress. We have previously shown that estrogen protects cardiomyocytes exposed to simulated ischemia-reperfusion (I/R) by differentially regulating pro-apoptotic p38alpha and pro-survival p38beta. However, little is known about how E2 modulation of the kinases alters the apoptotic signaling in cardiomyocytes. An attractive downstream target is p53, a well-known mediator of apoptosis and a substrate of p38alpha. We propose that the cytoprotective actions of estrogen involve regulation of p53 via p38. Cultured neonatal rat cardiomyocytes underwent hypoxia followed by reoxygenation (H/R) to simulate I/R. The extent of apoptosis was determined by examining annexin V-positive cells under fluorescent microscopy. The whole cell lysate was collected after H/R, and the protein of interest was immunoprecipitated and immunoblotted. The p38 activity was determined by immunoprecipitating the protein and performing in-vitro kinase assays on the substrate, ATF2. In our model, p53 played a significant role in H/R-induced myocyte death: Inhibition of p53 (by pifithrin-alpha or by siRNA) significantly reduced the number of apoptotic cells by half. Phosphorylation of p53 (p-p53) at serine 15 increased after H/R, while E2 effectively inhibited this form of p53 activation. Use of a specific agonist for each estrogen receptor isoform (ERalpha or ERbeta) demonstrated that both isoforms participate in inhibition of p-p53. E2 attenuated transcription of apoptosis-specific targets of p53, puma and noxa , as determined by RT PCR. In addition, inhibition of p53 augmented the p38beta activity, suggesting a baseline repression of this pro-survival kinase by p53 that may be reversed by E2. Our results demonstrate that during H/R stress, cardiomyocytes undergo p53-dependent apoptosis following phosphorylation of p53 by p38alpha. E2 protects cardiomyocytes by inhibiting p38alpha-p53 signaling, leading to pro-survival p38beta activation.

Necrostatin-1 Analog DIMO Exerts Cardioprotective Effect against Ischemia Reperfusion Injury by Suppressing Necroptosis via Autophagic Pathway in Rats

Pharmacology ◽  
2021 ◽  
Vol 106 (3-4) ◽  
pp. 189-201
Author(s):  
Shigang Qiao ◽  
Wen-jie Zhao ◽  
Huan-qiu Li ◽  
Gui-zhen Ao ◽  
Jian-zhong An ◽  
...  
Keyword(s):  
Cell Death ◽  
Ischemia Reperfusion Injury ◽  
Myocardial Infarct ◽  
Ischemia Reperfusion ◽  
Glucose Deprivation ◽  
Neonatal Rat ◽  
Ligand Docking ◽  
Autophagic Flux ◽  
Myocardial Infarct Size ◽  
Rat Cardiomyocytes

Aim: It has been reported that necrostatin-1 (Nec-1) is a specific necroptosis inhibitor that could attenuate programmed cell death induced by myocardial ischemia/reperfusion (I/R) injury. This study aimed to observe the effect and mechanism of novel Nec-1 analog (Z)-5-(3,5-dimethoxybenzyl)-2-imine-1-methylimidazolin-4-1 (DIMO) on myocardial I/R injury. Methods: Male SD rats underwent I/R injury with or without different doses of DIMO (1, 2, or 4 mg/kg) treatment. Isolated neonatal rat cardiomyocytes were subjected to oxygen-glucose deprivation/reoxygenation (OGD/R) treatment with or without DIMO (0.1, 1, 10, or 100 μM). Myocardial infarction was measured by TTC staining. Cardiomyocyte injury was assessed by lactate dehydrogenase assay (LDH) and flow cytometry. Receptor-interacting protein 1 kinase (RIP1K) and autophagic markers were detected by co-immunoprecipitation and Western blotting analysis. Molecular docking of DIMO into the ATP binding site of RIP1K was performed using GLIDE. Results: DIMO at doses of 1 or 2 mg/kg improved myocardial infarct size. However, the DIMO 4 mg/kg dose was ineffective. DIMO at the dose of 0.1 μM decreased LDH leakage and the ratio of PI-positive cells followed by OGD/R treatment. I/R or OGD/R increased RIP1K expression and in its interaction with RIP3K, as well as impaired myocardial autophagic flux evidenced by an increase in LC3-II/I ratio, upregulated P62 and Beclin-1, and activated cathepsin B and L. In contrast, DIMO treatment reduced myocardial cell death and reversed the above mentioned changes in RIP1K and autophagic flux caused by I/R and OGD/R. DIMO binds to RIP1K and inhibits RIP1K expression in a homology modeling and ligand docking. Conclusion: DIMO exerts cardioprotection against I/R- or OGD/R-induced injury, and its mechanisms may be associated with the reduction in RIP1K activation and restoration impaired autophagic flux.

scholarly journals Id2 Represses Aldosterone-Stimulated Cardiac T-Type Calcium Channels Expression

2021 ◽  
Vol 22 (7) ◽  
pp. 3561
Author(s):  
Jumpei Ito ◽  
Tomomi Minemura ◽  
Sébastien Wälchli ◽  
Tomoaki Niimi ◽  
Yoshitaka Fujihara ◽  
...  
Keyword(s):  
Transcriptional Repressor ◽  
Neonatal Rat ◽  
Protective Mechanism ◽  
Pathological State ◽  
Rat Cardiomyocytes ◽  
Adult Mice ◽  
Spontaneous Action ◽  
Sirna Knockdown ◽  
Spontaneous Action Potentials ◽  
Ca2 Channels

Aldosterone excess is a cardiovascular risk factor. Aldosterone can directly stimulate an electrical remodeling of cardiomyocytes leading to cardiac arrhythmia and hypertrophy. L-type and T-type voltage-gated calcium (Ca2+) channels expression are increased by aldosterone in cardiomyocytes. To further understand the regulation of these channels expression, we studied the role of a transcriptional repressor, the inhibitor of differentiation/DNA binding protein 2 (Id2). We found that aldosterone inhibited the expression of Id2 in neonatal rat cardiomyocytes and in the heart of adult mice. When Id2 was overexpressed in cardiomyocytes, we observed a reduction in the spontaneous action potentials rate and an arrest in aldosterone-stimulated rate increase. Accordingly, Id2 siRNA knockdown increased this rate. We also observed that CaV1.2 (L-type Ca2+ channel) or CaV3.1, and CaV3.2 (T-type Ca2+ channels) mRNA expression levels and Ca2+ currents were affected by Id2 presence. These observations were further corroborated in a heart specific Id2- transgenic mice. Taken together, our results suggest that Id2 functions as a transcriptional repressor for L- and T-type Ca2+ channels, particularly CaV3.1, in cardiomyocytes and its expression is controlled by aldosterone. We propose that Id2 might contributes to a protective mechanism in cardiomyocytes preventing the presence of channels associated with a pathological state.

Mechanical activity in heart regulates translation of α-myosin heavy chain mRNA but not its localization

1999 ◽  
Vol 276 (6) ◽  
pp. H2013-H2019 ◽  
Author(s):  
Gordana Nikcevic ◽  
Maria C. Heidkamp ◽  
Merja Perhonen ◽  
Brenda Russell
Keyword(s):  
Myosin Heavy Chain ◽  
Heavy Chain ◽  
Mrna Translation ◽  
Neonatal Rat ◽  
Mechanical Activity ◽  
Mrna Levels ◽  
Rat Cardiomyocytes ◽  
Significant Difference ◽  
Polysomal Fraction ◽  
Mechanical Transduction

Mechanical inactivity depresses protein expression in cardiac muscle tissue and results in atrophy. We explore the mechanical transduction mechanism in spontaneously beating neonatal rat cardiomyocytes expressing the α-myosin heavy chain (α-MyHC) isoform by interfering with cross-bridge function [2,3-butanedione monoxime (BDM), 7.5 mM] without affecting cell calcium. The polysome content and α-MyHC mRNA levels in fractions from a sucrose gradient were analyzed. BDM treatment blocked translation at initiation (162 ± 12% in the nonpolysomal RNA fraction and 43 ± 6% in the polysomal fraction, relative to control as 100%; P < 0.05). There was an increase in α-MyHC mRNA from the nonpolysomal fraction (120.5 ± 7.7%; P < 0.05 compared with control) with no significant change in the heavy polysomes. In situ hybridization of α-MyHC mRNA was used to estimate message abundance as a function of the distance from the nucleus. The mRNA was dispersed through the cytoplasm in spontaneously beating cells as well as in BDM-treated cells (no significant difference). We conclude that direct inhibition of contractile machinery, but not calcium, regulates initiation of α-MyHC mRNA translation. However, calcium, not pure mechanical signals, appears to be important for message localization.

Cross-bridge cycling gives rise to spatiotemporal heterogeneity of dynamic subcellular mechanics in cardiac myocytes probed with atomic force microscopy

2010 ◽  
Vol 298 (3) ◽  
pp. H853-H860 ◽  
Author(s):  
Evren U. Azeloglu ◽  
Kevin D. Costa
Keyword(s):  
Contractile Activity ◽  
Major Axis ◽  
Neonatal Rat ◽  
Pharmacological Treatments ◽  
Rat Cardiomyocytes ◽  
Force Microscopy ◽  
Cross Bridge ◽  
Apparent Modulus ◽  
Atomic Force ◽  
And Function

To study how the dynamic subcellular mechanical properties of the heart relate to the fundamental underlying process of actin-myosin cross-bridge cycling, we developed a novel atomic force microscope elastography technique for mapping spatiotemporal stiffness of isolated, spontaneously beating neonatal rat cardiomyocytes. Cells were indented repeatedly at a rate close but unequal to their contractile frequency. The resultant changes in pointwise apparent elastic modulus cycled at a predictable envelope frequency between a systolic value of 26.2 ± 5.1 kPa and a diastolic value of 7.8 ± 4.1 kPa at a representative depth of 400 nm. In cells probed along their major axis, spatiotemporal changes in systolic stiffness displayed a heterogeneous pattern, reflecting the banded sarcomeric structure of underlying myofibrils. Treatment with blebbistatin eliminated contractile activity and resulted in a uniform apparent modulus of 6.5 ± 4.8 kPa. This study represents the first quantitative dynamic mechanical mapping of beating cardiomyocytes. The technique provides a means of probing the micromechanical effects of disease processes and pharmacological treatments on beating cardiomyocytes, providing new insights and relating subcellular cardiac structure and function.

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