P4463The pineal hormone melatonin inhibits doxorubicin-induced mitochondrial dysfunction and apoptosis in cardiomyocytes

2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
T Kasi Ganeshan ◽  
J D Bell ◽  
N W Chong
Keyword(s):  
Mitochondrial Dysfunction ◽  
Cardiac Cells ◽  
H9c2 Cell ◽  
H9c2 Cells ◽  
Doxorubicin Treatment ◽  
Cell Energy ◽  
Student’S T ◽  
Induced Apoptosis ◽  
Measure Oxygen Consumption ◽  
Student’S T Test

Abstract Background Heart failure (HF) is a major end-point of cardiovascular diseases (CVD). The pathogenesis of HF is mostly unresolved but involves metabolic alterations. Treatment of animals and cardiomyocytes with β-adrenergic receptor agonists induces HF. Mitochondrial dysfunction and HF are common complications of anticancer drugs such as doxorubicin (DOX). Melatonin synthesis dramatically decreases with age and in patients with CVD. Purpose The aim of this study was to investigate whether DOX-induced cardiac dysfunction can be attenuated by melatonin. Methods The Seahorse XF analyser was utilised (with the XFp Cell Energy Phenotype kit) to measure oxygen consumption rate [OCR; oxidative phosphorylation (OXPHOS)] and extracellular acidification rate (ECAR; glycolysis) in living rat cardiomyocyte-derived H9c2 cell line. Mono-layers of cells were treated with cardiotoxic drugs [isoproterenol (ISO, 100μM) or DOX (0.1μM)] for 24hr with and without melatonin co-treatment (MEL, 1μM). Cyan ADP flow cytometry was used to examine the anti-apoptotic properties of MEL (1μM) on DOX-treatment (0.5μM, 24hr). Data are given as mean±SEM (n=separate experiments) and analysis was performed using ANOVA and two-tail unpaired Student's T-test, as applicable. Results Isoproterenol-treatment increased peak OCR of H9c2 cells by ∼30% which was inhibited by MEL [CON, 384±17; ISO, 496±33; ISO+MEL, 412±31pmol/min; n=3 (six replicates); CON vs. ISO, p<0.05; ISO vs. ISO+MEL, p<0.05; CON vs. ISO+MEL, p>0.05]. Doxorubicin-treatment decreased OCR by ∼40% which was reversed by MEL [CON, 934±69; DOX, 554±52; DOX+MEL, 858±97pmol/min; n=3 (six replicates); CON vs. DOX, p<0.05; DOX vs. DOX+MEL, p<0.05; CON vs. DOX+MEL, p>0.05]. ISO and DOX significantly increased (∼30%) and decreased (∼25%) ECAR respectively (n=3, p<0.05) which was not inhibited by MEL. Melatonin alone had no significant effect on OCR and ECAR. Melatonin inhibited DOX-induced apoptosis in H9c2 cells [CON, 6.3±0.8%; DOX, 22±1.8%; DOX+MEL, 11±1.7%, n=4 (two replicates); CON vs. DOX, p<0.001; DOX vs. DOX+MEL, p<0.004; CON vs. DOX+MEL, p>0.05]. Conclusions ISO and DOX-treatment induced mitochondrial dysfunction in H9c2 cells by alteration of OXPHOS and glycolysis; changes in OXPHOS were prevented by MEL. These data indicate that DOX-induced apoptosis in cardiac cells may be mediated, at least in part, by OXPHOS dysfunction which was attenuated by MEL treatment.

scholarly journals Sheng Mai San protects H9C2 cells against hyperglycemia-induced apoptosis

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Bing Pang ◽  
Li-Wei Shi ◽  
Li-juan Du ◽  
Yun-Chu Li ◽  
Mei-Zhen Zhang ◽  
...  
Keyword(s):  
Cell Viability ◽  
Molecular Mechanisms ◽  
Annexin V ◽  
Signalling Pathway ◽  
H9c2 Cell ◽  
Protective Effects ◽  
H9c2 Cells ◽  
P53 Expression ◽  
Fasl Gene ◽  
Induced Apoptosis

Abstract Background Sheng Mai San (SMS) has been proven to exhibit cardio-protective effects. This study aimed to explore the molecular mechanisms of SMS on hyperglycaemia (HG)-induced apoptosis in H9C2 cells. Methods HG-induced H9C2 cells were established as the experimental model, and then treated with SMS at 25, 50, and 100 μg/mL. H9C2 cell viability and apoptosis were quantified using MTT and Annexin V-FITC assays, respectively. Furthermore, Bcl-2/Bax signalling pathway protein expression and Fas and FasL gene expression levels were quantified using western blotting and RT-PCR, respectively. Results SMS treatments at 25, 50, 100 μg/mL significantly improved H9C2 cell viability and inhibited H9C2 cell apoptosis (p < 0.05). Compared to the HG group, SMS treatment at 25, 50, and 100 μg/mL significantly downregulated p53 and Bax expression and upregulated Bcl-2 expression (p < 0.05). Moreover, SMS treatment at 100 μg/mL significantly downregulated Fas and FasL expression level (p < 0.05) when compared to the HG group. Conclusion SMS protects H9C2 cells from HG-induced apoptosis probably by downregulating p53 expression and upregulating the Bcl-2/Bax ratio. It may also be associated with the inhibition of the Fas/FasL signalling pathway.

scholarly journals Lingonberry anthocyanins protect cardiac cells from oxidative-stress-induced apoptosis

2017 ◽  
Vol 95 (8) ◽  
pp. 904-910 ◽  
Author(s):  
Cara K. Isaak ◽  
Jay C. Petkau ◽  
Heather Blewett ◽  
Karmin O ◽  
Yaw L. Siow
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Cardiac Cells ◽  
Control Group ◽  
Protective Effects ◽  
H9c2 Cells ◽  
Induced Apoptosis ◽  
Hoechst Staining

Lingonberry grown in northern Manitoba, Canada, contains exceptionally high levels of anthocyanins and other polyphenols. Previous studies from our lab have shown that lingonberry anthocyanins can protect H9c2 cells from ischemia–reperfusion injury and anthocyanin-rich diets have been shown to be associated with decreased cardiovascular disease and mortality. Oxidative stress can impair function and trigger apoptosis in cardiomyocytes. This study investigated the protective effects of physiologically relevant doses of lingonberry extracts and pure anthocyanins against hydrogen-peroxide-induced cell death. Apoptosis and necrosis were detected in H9c2 cells after hydrogen peroxide treatment via flow cytometry using FLICA 660 caspase 3/7 combined with YO-PRO-1 and then confirmed with Hoechst staining and fluorescence microscopy. Each of the 3 major anthocyanins found in lingonberry (cyanidin-3-galactoside, cyanidin-3-glucoside, and cyanidin-3-arabinoside) was protective against hydrogen-peroxide-induced apoptosis in H9c2 cells at 10 ng·mL−1 (20 nmol·L−1) and restored the number of viable cells to match the control group. A combination of the 3 anthocyanins was also protective and a lingonberry extract tested at 3 concentrations produced a dose-dependent protective effect. Lingonberry anthocyanins protected cardiac cells from oxidative-stress-induced apoptosis and may have cardioprotective effects as a dietary modification.

scholarly journals L-Carnitine: An Antioxidant Remedy for the Survival of Cardiomyocytes under Hyperglycemic Condition

2018 ◽  
Vol 2018 ◽  
pp. 1-12 ◽  
Author(s):  
Fernanda Vacante ◽  
Pamela Senesi ◽  
Anna Montesano ◽  
Alice Frigerio ◽  
Livio Luzi ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Mitochondrial Dysfunction ◽  
Signaling Pathways ◽  
Stress Condition ◽  
Cardiac Cells ◽  
Ros Production ◽  
Protective Effects ◽  
H9c2 Cells ◽  
Rat Cardiomyocytes ◽  
Oxidative Stress Condition

Background. Metabolic alterations as hyperglycemia and inflammation induce myocardial molecular events enhancing oxidative stress and mitochondrial dysfunction. Those alterations are responsible for a progressive loss of cardiomyocytes, cardiac stem cells, and consequent cardiovascular complications. Currently, there are no effective pharmacological measures to protect the heart from these metabolic modifications, and the development of new therapeutic approaches, focused on improvement of the oxidative stress condition, is pivotal. The protective effects of levocarnitine (LC) in patients with ischemic heart disease are related to the attenuation of oxidative stress, but LC mechanisms have yet to be fully understood. Objective. The aim of this work was to investigate LC’s role in oxidative stress condition, on ROS production and mitochondrial detoxifying function in H9c2 rat cardiomyocytes during hyperglycemia. Methods. H9c2 cells in the hyperglycemic state (25 mmol/L glucose) were exposed to 0.5 or 5 mM LC for 48 and 72 h: LC effects on signaling pathways involved in oxidative stress condition were studied by Western blot and immunofluorescence analysis. To evaluate ROS production, H9c2 cells were exposed to H2O2 after LC pretreatment. Results. Our in vitro study indicates how LC supplementation might protect cardiomyocytes from oxidative stress-related damage, preventing ROS formation and activating antioxidant signaling pathways in hyperglycemic conditions. In particular, LC promotes STAT3 activation and significantly increases the expression of antioxidant protein SOD2. Hyperglycemic cardiac cells are characterized by impairment in mitochondrial dysfunction and the CaMKII signal: LC promotes CaMKII expression and activation and enhancement of AMPK protein synthesis. Our results suggest that LC might ameliorate metabolic aspects of hyperglycemic cardiac cells. Finally, LC doses herein used did not modify H9c2 growth rate and viability. Conclusions. Our novel study demonstrates that LC improves the microenvironment damaged by oxidative stress (induced by hyperglycemia), thus proposing this nutraceutical compound as an adjuvant in diabetic cardiac regenerative medicine.

Doxorubicin-induced cardiotoxicity: direct correlation of cardiac fibroblast and H9c2 cell survival and aconitase activity with heat shock protein 27

2007 ◽  
Vol 293 (5) ◽  
pp. H3111-H3121 ◽  
Author(s):  
Samir Turakhia ◽  
C. D. Venkatakrishnan ◽  
Kathy Dunsmore ◽  
Hector Wong ◽  
Periannan Kuppusamy ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Heat Shock Protein 27 ◽  
Cardiac Cells ◽  
H9c2 Cell ◽  
Heat Shock Factor 1 ◽  
H9c2 Cells ◽  
Sod Activity ◽  
Patients With Cancer ◽  
Si Rna

The use of doxorubicin (Dox) and its derivatives as chemotherapeutic drugs to treat patients with cancer causes dilated cardiomyopathy and congestive heart failure due to Dox-induced cardiotoxicity. In this work, using heat shock factor-1 wild-type (HSF-1+/+) and HSF-1 knockout (HSF-1−/−) mouse fibroblasts and embryonic rat heart-derived cardiac H9c2 cells, we show that the magnitude of protection from Dox-induced toxicity directly correlates with the level of the heat shock protein 27 (HSP27). Western blot analysis of normal and heat-shocked cells showed the maximum expression of HSP27 in heat-shocked cardiac H9c2 cells and no HSP27 in HSF-1−/− cells (normal or heat-shocked). Correspondingly, the cell viability, measured [with (3,4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay] after treatment with various concentrations of Dox, was the highest in heat-shocked H9c2 cells and the lowest in HSF-1−/− cells. Depleting HSP27 in cardiac H9c2 cells by small interfering (si)RNA also reduced the viability against Dox, confirming that HSP27 does protect cardiac cells against the Dox-induced toxicity. The cells that have lower HSP27 levels such as HSF-1−/−, were found to be more susceptible for aconitase inactivation. Based on these results we propose a novel mechanism that HSP27 plays an important role in protecting aconitase from Dox-generated O2•−, by increasing SOD activity. Such a protection of aconitase by HSP27 eliminates the catalytic recycling of aconitase released Fe(II) and its deleterious effects in cardiac cells.

scholarly journals Heart-on-a-Chip: An Investigation of the Influence of Static and Perfusion Conditions on Cardiac (H9C2) Cell Proliferation, Morphology, and Alignment

2017 ◽  
Vol 22 (5) ◽  
pp. 536-546 ◽  
Author(s):  
Anna Kobuszewska ◽  
Ewelina Tomecka ◽  
Kamil Zukowski ◽  
Elzbieta Jastrzebska ◽  
Michal Chudy ◽  
...  
Keyword(s):  
Lab On A Chip ◽  
Heart Tissue ◽  
Cardiac Cells ◽  
H9c2 Cell ◽  
Cardiac Diseases ◽  
H9c2 Cells ◽  
Parallel Microchannels ◽  
Static Conditions ◽  
Longitudinal Channel ◽  
Number Of Cells

Lab-on-a-chip systems are increasingly used as tools for cultures and investigation of cardiac cells. In this article, we present how the geometry of microsystems and microenvironmental conditions (static and perfusion) influence the proliferation, morphology, and alignment of cardiac cells (rat cardiomyoblasts—H9C2). Additionally, studies of cell growth after incubation with verapamil hydrochloride were performed. For this purpose, poly(dimethylsiloxane) (PDMS)/glass microfluidic systems with three different geometries of microchambers (a circular chamber, a longitudinal channel, and three parallel microchannels separated by two rows of micropillars) were prepared. It was found that static conditions did not enhance the growth of H9C2 cells in the microsystems. On the contrary, perfusion conditions had an influence on division, morphology, and the arrangement of the cells. The highest number of cells, their parallel orientation, and their elongated morphology were obtained in the longitudinal microchannel. It showed that this kind of microsystem can be used to understand processes in heart tissue in detail and to test newly developed compounds applied in the treatment of cardiac diseases.

scholarly journals Protective Effect of Two Alkaloids from Hippophae rhamnoides Linn. against Doxorubicin-Induced Toxicity in H9c2 Cardiomyoblasts

Molecules ◽  
2021 ◽  
Vol 26 (7) ◽  
pp. 1946
Author(s):  
Wenna Zhou ◽  
Jian Ouyang ◽  
Na Hu ◽  
Gang Li ◽  
Honglun Wang
Keyword(s):  
Protein Expression ◽  
Protective Effect ◽  
Caspase 3 ◽  
Cardiac Cells ◽  
Potential Candidate ◽  
H9c2 Cells ◽  
Cardiac Muscle Cells ◽  
Anti Cancer ◽  
H9c2 Cardiomyoblasts ◽  
Induced Apoptosis

Background: Doxorubicin (Dox) is one of the most frequently prescribed anti-cancer drugs. However, clinical application with Dox is limited due to its potentially fatal cumulative cardiotoxicity. N-p-coumaroyl-4-aminobutan-1-ol (alk-A), an organic amide alkaloid and hippophamide (alk-B), a rare pyridoindole alkaloid were successfully obtained by purification and separation of seabuckthorn seed residue in our previous research. This study was undertaken to investigate the protective effect of alk-A and alk-B against Dox-induced embryonic rat cardiac cells (H9c2 cells) apoptosis. Methods: H9c2 cells were treated with Dox (2.5 µM) in the presence of alk-A and alk-B (10, 20, and 40 µM) and incubated for 24 h. Results: It was shown that pretreatment of the H9c2 cells with alk-A and alk-B significantly reduced Dox-induced apoptosis. Alk-A and alk-B both inhibited reactive oxygen species (ROS) production and suppressed cleaved-caspase-3 protein expression and the activation of JNK (Jun N-terminal kinases), as well as increasing ATP levels, favoring mitochondrial mitofusin protein expression, and relieving damage to mitochondrial DNA. Conclusions: These results suggest that alk-A and alk-B can inhibit Dox-induced apoptosis in H9C2 cardiac muscle cells via inhibition of cell apoptosis and improvement of mitochondrial function, while alk-B showed more protection. Alk-B could be a potential candidate agent for protecting against cardiotoxicity in Dox-exposed patients.

scholarly journals MicroRNA 379 Regulates Klotho Deficiency–Induced Cardiomyocyte Apoptosis Via Repression of Smurf1

Hypertension ◽  
2021 ◽  
Author(s):  
Kai Chen ◽  
Bo Zhang ◽  
Zhongjie Sun
Keyword(s):  
Heart Failure ◽  
Apoptotic Cell ◽  
Heart Function ◽  
Underlying Mechanism ◽  
Cardiomyocyte Apoptosis ◽  
H9c2 Cell ◽  
Heart Cells ◽  
Sequencing Analysis ◽  
H9c2 Cells ◽  
Induced Apoptosis

Klotho is an aging-suppressor gene. Klotho gene deficiency impairs heart function leading to heart failure, but the underlying mechanism remains poorly understood. MicroRNAs are increasingly recognized to play important roles in the pathogenesis of cardiomyopathy. The objective of this study is to investigate whether microRNA 379 (Mir379) regulates Klotho deficiency-associated cardiomyocyte apoptosis. Using inducible Cre-Loxp recombination technology, we first found that kidney-specific deletion of the Klotho gene caused heart failure. Using microRNA sequencing analysis, we found that Mir379 may be a target of Klotho. In cultured H9c2 heart cells, we found that treatment with Klotho-free medium increased Mir379 levels and induced apoptosis. To test whether Mir379 mediates Klotho deficiency–induced apoptosis, H9c2 cells were transfected with Mir379 inhibitor. Interestingly, Mir379 inhibitor (anti-Mir379) prevented Klotho deficiency–induced H9c2 cell apoptosis. On the contrary, Mir379 mimic itself caused apoptosis in H9c2 cells. These findings suggest that Mir379 may mediate Klotho deficiency–induced apoptosis in H9C2 cells. Using the mRNA-miRNA target interaction assay, we found that Smurf1(SMAD specific E3 ubiquitin protein ligase 1) mRNA contained the 3-UTR binding site for Mir379. Importantly, Mir379 mimic suppressed Smurf1 expression, and the Mir379 mimic–induced apoptosis can be rescued by treatment with exogenous Smurf1 protein. Therefore, Smurf1 repression may be involved in Mir379–induced H9c2 cells apoptosis. In conclusion, Mir379 may mediate Klotho deficiency-associated cardiomyocyte apoptosis through repression of Smurf1 which is required for Mir379-induced apoptotic cell death. Mir379 may be a potential therapeutic target for cardiomyocyte apoptosis-associated heart failure due to Klotho deficiency.

scholarly journals Polyphenol‑rich extract of Aronia melanocarpa inhibits TNF‑α induced apoptosis in H9c2 cells

2016 ◽  
Vol 85 (3) ◽  
pp. 185 ◽  
Author(s):  
Krzysztof Borecki ◽  
Marta Żuchowski ◽  
Aldona Siennicka ◽  
Grażyna Adler ◽  
Maria Jastrzębska
Keyword(s):  
Significant Inhibition ◽  
Cardiac Cells ◽  
H9c2 Cells ◽  
Cytoprotective Effect ◽  
Aronia Melanocarpa ◽  
Myocardial Oxidative Stress ◽  
Commercial Extract ◽  
Tnf Α ◽  
H9c2 Cardiomyoblasts ◽  
Induced Apoptosis

Introduction. In certain pathological states within cardiovascular system, cardiomyocytes may exhibit overexpression of TNF‑α that results in induction of myocardial oxidative stress and cardiac cells apoptosis. Thus, they may participate in development and progression of post‑infarct congestive heart failure. The aim of this study was to evaluate the effect of polyphenol‑rich Aronia melanocarpa extract (AME) on TNF‑α induced apoptosis in cardiomyoblast H9c2 cells. Material and Methods. Apoptosis was measured in H9c2 cells preincubated with increasing concentrations of commercial extract of aronia – Aronox (10–50 μg/mL) for 24h and then treated with TNF‑α: 100 ng/mL for 24h as well. The MTT assay was used to determine cardiomyoblasts viability. Alteration of the mitochondrial membrane potential, specific for apoptotic cells, was evaluated with caspase-3 activity assay kit. Results. Our results showed that AME significantly inhibited TNF‑α induced apoptosis (IC50 = 55.84 µg/mL) and cytotoxicity in H9c2 cells. Significant inhibition of apoptosis was observed in all tested concentrations of AME. The highest anti‑apoptotic and cytoprotective effect was observed at the highest concentration (50 μg/mL), while in lower the concentrations cytoprotective effect was statistically insignificant. Conclusions. Polyphenol‑rich AME exhibits anti‑apoptotic and cytoprotective effect in H9c2 cardiomyoblasts treated with TNF‑α. Further studies are required in context of its possible application in prevention and/or therapy of cardiovascular diseases.

Metformin attenuates ischemia/reperfusion-induced apoptosis of cardiac cells by downregulation of p53/microRNA-34a via activation SIRT1

2021 ◽  
Author(s):  
Weiwei Li ◽  
Sheng Jin ◽  
Jie Hao ◽  
Yun Shi ◽  
Wenjie Li ◽  
...  
Keyword(s):  
Ischemia Reperfusion ◽  
Coronary Intervention ◽  
Cardiac Cells ◽  
H9c2 Cells ◽  
St Segment Elevation ◽  
St Segment ◽  
Associated Proteins ◽  
Myocardial Ischemia Reperfusion ◽  
Induced Apoptosis

Metformin has been demonstrated to be beneficial for the treatment of an impaired myocardium as a result of ischemia reperfusion (I/R) injury and miR-34a may be involved in this process. The aim of the present study was to determine the mechanisms by which metformin attenuated myocardial I/R injury induced apoptosis. In the in vivo I/R rat model, metformin reduced the area of damaged myocardium and CKMB activity for protection of the myocardium. Metformin also reduced apoptosis, the expression of apoptosis associated proteins and miR-34a, which resulted in corresponding changes of Bcl-2 expression. To further examine the role of miR-34a, H9C2 cells were transfected by miR-34a mimic and inhibitor. Overexpression of miR-34a increased apoptosis in H9C2 cells induced by OGD/R and knockdown of miR-34a reduced apoptosis. Metformin decreased the deacetylation activity of SIRT 1 resulting in reduced Ac-p53 levels, which reduced the levels of pri-miR-34a. To confirm these results clinically, 90 patients with ST-segment elevation myocardial infarction following percutaneous coronary intervention were recruited. Patients who took metformin regularly before infarction had lower miR-34a levels and lower serum CKMB activity. Metformin also improved sum ST-segment recovery following I/R injury. In conclusion, metformin may be helpful in the treatment of myocardial ischemia reperfusion.

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