Protective activity of Malus doumeri leaf extract on H2O2-induced oxidative injury in H9C2 rat cardiomyocytes

Abstract

In this study, <i>Malus doumeri</i> leaf extract (MDLE) was used to test its anti-oxidation capacity <i>in vitro</i>, it has been preliminarily analyzed for H₂O₂-induced oxidative damage in H9C2 cells and its main active components. The antioxidant capacity through DPPH (1, 1-Diphenyl-2-Picrylhydrazyl), ABTS⁺• [2,2,2'-azino-BIS-(3-ethylbenzo-thiazoline-6-sulfonic acid)] radical ion, •OH (hydroxyl radical), and <mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML"><mml:mo>•</mml:mo> <mml:msup> <mml:mrow> <mml:msub><mml:mrow><mml:mtext>O</mml:mtext></mml:mrow> <mml:mrow><mml:mn>2</mml:mn></mml:mrow> </mml:msub> </mml:mrow> <mml:mrow><mml:mo>-</mml:mo></mml:mrow> </mml:msup> </mml:math> (superoxide anion) were determined <i>in vitro</i>. The proliferation of H9C2 cells was examined by MTT [3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-Tetrazolium bromide]. MDA (malondialdehyde), SOD (superoxide dismutase), CAT (catalase), GSH (glutathione), and GSH-Px (glutathione peroxidase) were determined by colorimetry. Apoptosis induced by oxidative damage was detected by flow cytometry. The mRNA expression of antioxidant related genes of SOD, CAT, GSH, and GSH-Px were checked by qRT-PCR (quantitative real-time polymerase chain reaction). The MDLE main active components were analyzed by HPLC (high-performance liquid chromatography). MDLE had significant scavenging effects on DPPH, ABTS⁺•, •OH, and superoxide anion radicals in a concentration-dependent manner. H₂O₂ treatment could significantly lead to oxidative stress injury of H9C2 cells, and MDLE treatment significantly improved the degree of H9C2 cell damage, and showed a positive correlation with concentration. MDLE can also reduce apoptosis caused by oxidative damage. MDLE treatment could significantly reduce MDA content and increase CAT, SOD, GSH, and GSH-Px contents and expression. In addition, by HPLC analysis, the following six bioactive components were detected from MDLE: chlorogenic acid, isoquercitrin, quercetin, baicalin, and phloretin. Therefore, MDLE has a good protective effect on myocardial cells.