Suppression of isoproterenol induced apoptosis in H9c2 cardiomyoblasts cells by aescin

Paper Details

Research Paper 21/11/2022
Views (435) Download (47)

Suppression of isoproterenol induced apoptosis in H9c2 cardiomyoblasts cells by aescin

Kanimozhi Kaliyamoorthi, Tani Carmel Raj TG, Nivedha Jayaseelan, Sindhu Ganapathi, Vennila Lakshmanan
Int. J. Biosci.21( 5), 261-272, November 2022.
Certificate: IJB 2022 [Generate Certificate]


Medicinal herbs have been utilized for centuries to alleviate a spectrum of drugs. Horse chestnut (Aesculus hippocastanum) is used in phytomedicine to prevent and treat a wide range of conditions, and its seed extracts contain phytocompounds such as flavonoids, polyphenols, and triterpenoids (aescin). This study aims to investigate the potential mechanisms associated with the cardioprotective effect of aescin on isoproterenol (ISO) induced cardiotoxicity in H9c2 cell lines. The effect of the drug on cell morphology was studied by using a phase contrast microscope, and cell viability was studied by using 3-(4,5-dimethyl-thiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) staining, and reactive oxygen species (ROS) production was estimated by 20-7’dichlorofluorescein diacetate staining. H9c2 cells were treated with ISO to cause cell damage and the effect of the aescin on cell morphology, mitochondrial membrane potential, intracellular ROS generation, cell viability, and apoptosis was studied. The results of this study showed that pre-administration of aescin significantly reduced the ISO-induced toxic effects on cell morphology and enhanced the number of viable cells in a dose-dependent manner. This study also demonstrates that ROS generation was significantly increased in ISO-administered cells and ISO-induced ROS production was significantly reduced in the aescin-pre-administered H9c2 cells. ISO-induced changes in the mitochondrial membrane potential of H9c2 cells were remarkably improved with aescin pretreatment. These results clearly suggest that pretreatment of aescin protects the cells against ISO-induced damage by resuming mitochondrial function and regulating apoptosis.


Branco AF, Pereira SP, Gonzalez S, Gusev O, Rizvanov AA, Oliveira PJ. 2015. Gene Expression Profiling of H9c2 Myoblast Differentiation towards a Cardiac-Like Phenotype. PloS one 10, e0129303.

Costa VM, Carvalho F, Bastos ML, Carvalho RA, Carvalho M, Remião F. 2011. Contribution of catecholamine reactive intermediates and oxidative stress to the pathologic features of heart diseases. Current medicinal chemistry 18, 2272–2314.

Dallons M, Schepkens C, Dupuis A, Tagliatti V, Colet JM. 2020. New Insights about Doxorubicin-Induced Toxicity to Cardiomyoblast-Derived H9c2 Cells and Dexrazoxane Cytoprotective Effect: Contribution of In Vitro1H-NMR Metabonomics. Frontiers in pharmacology 11, 79.

D’Oria R, Schipani R, Leonardini A, Natalicchio A, Perrini S, Cignarelli A, Laviola L, Giorgino F. 2020. The Role of Oxidative Stress in Cardiac Disease: From Physiological Response to Injury Factor. Oxidative medicine and cellular longevity 2020, 5732956.

Gallelli L. 2019. Escin: a review of its anti-edematous, anti-inflammatory, and venotonic properties. Drug design, development and therapy 13, 3425-3437.

Hausenloy DJ, Yellon DM. 2013. Myocardial ischemia-reperfusion injury: a neglected therapeutic target. The Journal of clinical investigation 123, 92-100.

Johannsen DL, Ravussin E. 2009. The role of mitochondria in health and disease. Current opinion in pharmacology 9, 780–786.

Kasibhatla S, Amarante-Mendes GP, Finucane D, Brunner T, Bossy-Wetzel E, Green DR. 2006. Acridine Orange/Ethidium Bromide (AO/EB) Staining to Detect Apoptosis. CSH protocols 2006, pdb. Prot 4493.

Khan MA, Hashim MJ, Mustafa H, Baniyas MY, Al Suwaidi SKBM, AlKatheeri R, Alblooshi FMK, Almatrooshi MEAH, Alzaabi MEH, Al Darmaki RS, Lootah SNAH. 2020. Global Epidemiology of Ischemic Heart Disease: Results from the Global Burden of Disease Study. Cureus 12, e9349.

Kim H, Xue X. 2020. Detection of Total Reactive Oxygen Species in Adherent Cells by 2′,7′-Dichlorodihydrofluorescein Diacetate Staining. Journal of visualized experiments: JoVE 10, 3791/60682.

Ling T, Boyd L, Rivas F. 2022. Triterpenoids as Reactive Oxygen Species Modulators of Cell Fate. Chemical research in toxicology 35, 569–584.

Liu K, Liu PC, Liu R, Wu X. 2015. Dual AO/EB staining to detect apoptosis in osteosarcoma cells compared with flow cytometry. Medical science monitor basic research 21, 15-20.

Lu LY, Ou N, Lu QB. 2013. Antioxidant induces DNA damage, cell death and mutagenicity in human lung and skin normal cells. Scientific reports 3, 3169.

Marchi S, Giorgi C, Suski JM, Agnoletto C, Bononi A, Bonora M, De Marchi E, Missiroli S, Patergnani S, Poletti F, Rimessi A, Duszynski J, Wieckowski MR, Pinton P. 2012. Mitochondria-ros crosstalk in the control of cell death and aging. Journal of signal transduction, 2012, 329635.

Pizzino G, Irrera N, Cucinotta M, Pallio G, Mannino F, Arcoraci V, Squadrito F, Altavilla D, Bitto A. 2017. Oxidative Stress: Harms and Benefits for Human Health. Oxidative medicine and cellular longevity 2017, 8416763.

Redza-Dutordoir M, Averill-Bates DA. 2016. Activation of apoptosis signalling pathways by reactive oxygen species. Biochimica et biophysica acta 1863, 2977–2992.

Sakamuru S, Attene-Ramos MS, Xia M. 2016. Mitochondrial Membrane Potential Assay. Methods in molecular biology (Clifton, N.J.) 1473, 17–22.

Sedlic F, Sepac A, Pravdic D, Camara AK, Bienengraeber M, Brzezinska AK, Wakatsuki T, Bosnjak ZJ. 2010. Mitochondrial depolarization underlies delay in permeability transition by preconditioning with isoflurane: roles of ROS and Ca2+. American Jornal of Physiol Cell Physiol 299, C506-15.

Severino P, D’Amato A, Pucci M, Infusino F, Birtolo LI, Mariani MV, Lavalle, C, Maestrini V, Mancone M, Fedele F. 2020. Ischemic Heart Disease and Heart Failure: Role of Coronary Ion Channels. International journal of molecular sciences 21, 3167.

Singh NP, McCoy MT, Tice RR, Schneider EL. 1988. A simple technique for quantitation of low levels of DNA damage in individual cells. Experimental cell research 175, 184-191.

Sirtori CR. 2001. Aescin: pharmacology, pharmacokinetics and therapeutic profile. Pharmacological research 44, 183-193.

Soetikno V, Sari FR, Lakshmanan AP, Arumugam S, Harima M, Suzuki K, Kawachi H, Watanabe K. 2013. Curcumin alleviates oxidative stress, inflammation, and renal fibrosis in remnant kidney through the Nrf2-keap1 pathway. Molecular nutrition & food research 57, 1649-1659.

Suhaeri M, Subbiah R, Van SY, Du P, Kim IG, Lee K, Park K. 2015. Cardiomyoblast (h9c2) differentiation on tunable extracellular matrix microenvironment. Tissue Engineering Part A 21, 1940-51.

Tungmunnithum D, Thongboonyou A, Pholboon A, Yangsabai A. 2018. Flavonoids and Other Phenolic Compounds from Medicinal Plants for Pharmaceutical and Medical Aspects: An Overview. Medicines (Basel, Switzerland) 5, 93.

Vishnu KV, Ajeesh Kumar KK, Chatterjee NS, Lekshmi RGK, Sreerekha PR, Mathew S, Ravishankar CN. 2018. Sardine oil-loaded vanillic acid grafted chitosan microparticles, a new functional food ingredient: attenuates myocardial oxidative stress and apoptosis in cardiomyoblast cell lines (H9c2). Cell stress & chaperones 23, 213-222.

Wang C, Youle RJ. 2009. The role of mitochondria in apoptosis*. Annual review of genetics 43, 95–118.

Watkins SJ, Borthwick GM, Arthur HM. 2011 The H9c2 cell line and primary neonatal cardiomyocyte cells show similar hypertrophic responses in vitro. In Vitro Cellular & Developmental Biology – Animal 47, 125-131.

WHO- World Health Organization. 2021. sheets/detail/cardiovascular-diseases-(CVD)

Wilson DW, Nash P, Buttar HS, Griffiths K, Singh R, De Meester F, Horiuchi R, Takahashi T. 2017. The Role of Food Antioxidants, Benefits of Functional Foods, and Influence of Feeding Habits on the Health of the Older Person: An Overview. Antioxidants (Basel, Switzerland) 6, 81.

Witaicenis A, Seito LN, da Silveira Chagas A, de Almeida LD, Jr Luchini AC, Rodrigues-Orsi P, Cestari SH, Di Stasi LC. 2014. Antioxidant and intestinal anti-inflammatory effects of plant-derived coumarin derivatives. Phytomedicine: international journal of phytotherapy and phytopharmacology 21, 240-246.

Ye B, Hou N, Xiao L, Xu Y, Xu H, Li F. 2016. Dynamic monitoring of oxidative DNA double-strand break and repair in cardiomyocytes. Cardiovascular pathology: the official journal of the Society for Cardiovascular Pathology 25, 93-100.

Zorov DB, Juhaszova M, Sollott SJ. 2014. Mitochondrial reactive oxygen species (ROS) and ROS-induced ROS release. Physiological reviews 94, 909-950.