Protective Mechanism of Quercetin on Oxidative Injury in H9C2 Cardiomyocytes

HUO Kui-yuan; CHANG Hong; WANG Yong; LI Chun; TANG Bing-hua

doi:10.13422/j.cnki.syfjx.2016150122

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Protective Mechanism of Quercetin on Oxidative Injury in H9C2 Cardiomyocytes

更新时间：2024-09-23

- Protective Mechanism of Quercetin on Oxidative Injury in H9C2 Cardiomyocytes
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 22, Issue 15, Pages: 122-127(2016)
- 作者机构：
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.2016150122
  CLC： R285
- Published：2016
- 稿件说明：
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HUO Kui-yuan, CHANG Hong, WANG Yong, et al. Protective Mechanism of Quercetin on Oxidative Injury in H9C2 Cardiomyocytes[J]. Chinese journal of experimental traditional medical formulae, 2016, 22(15): 122-127.
DOI：

HUO Kui-yuan, CHANG Hong, WANG Yong, et al. Protective Mechanism of Quercetin on Oxidative Injury in H9C2 Cardiomyocytes[J]. Chinese journal of experimental traditional medical formulae, 2016, 22(15): 122-127. DOI： 10.13422/j.cnki.syfjx.2016150122.

摘要

目的：探讨在H2O2诱导的氧化应激损伤中槲皮素对H9C2心肌细胞存活率、活性氧分子（ROS）、还原型辅酶Ⅱ（NADPH）氧化酶、细胞凋亡等关键因子的影响，初步阐明槲皮素对心肌细胞的保护机制。方法： H2O2刺激H9C2心肌细胞建立氧化应激损伤模型，分为正常组、模型组、槲皮素组，采用噻唑蓝（MTT）法检测心肌细胞活力，活性氧检测试剂盒检测槲皮素处理后ROS变化，流式细胞术检测各组细胞凋亡率，实时荧光定量PCR（Real-time PCR）与蛋白免疫印迹法（Western blot）分别检测细胞氧化应激反应、细胞凋亡机制相关分子[NADPH氧化酶-4（NOX-4），NADPH氧化酶亚单位p22phox，B淋巴细胞瘤-2原瘤基因（Bcl-2），Bcl-2相关X基因（Bax），半胱胺酸蛋白酶-8（Caspase-8）]基因和蛋白表达水平。结果： 10 μmol·L-1槲皮素具有显著的抗氧化、抗凋亡作用。与模型组比较，槲皮素组细胞存活率明显提高（P<0.05），ROS水平与细胞凋亡率明显降低（P<0.05，P<0.01）。此外，槲皮素组中Bcl-2，NOX-4基因表达上调（P<0.05）；p22phox，Bax，Caspase-8基因表达明显下调（P<0.05）。同时槲皮素能够抑制Bax，p22phox蛋白的表达（P<0.05），上调磷脂酰肌醇-3激酶（PI3K），Bcl-2蛋白的表达（P<0.05）。结论： 10 μmol·L-1槲皮素能有效减少H2O2介导的心肌细胞氧化损伤，进而抑制心肌细胞凋亡。该机制可能是通过调节NADPH氧化酶亚型NOX-4与p22phox，减少ROS生成，激活PI3K表达，升高Bcl-2实现。

Abstract

Objective: To preliminarily investigate the protective effects of quercetin on the survival rate of H2O2-induced oxidative injury in H9C2 cardiomyocytes

and such key factors as reactive oxygen species (ROS)

nicotinamide adenosine denucleotide hydro-phosphoric acid(NADPH) oxidase and apoptosis

in order to clarify the underlying mechanism. Method: A model of hydrogen peroxide H2O2-induced H9C2 cells oxidative injury was established in vitro

and divided into the normal group

the model group and the quercetin group. Cell viability was examined with an methylthiazolyldiphenyl-tetrazolium bromide(MTT) assay to determine the available concentrations of H2O2 and quercetin. The effects of quercetin on ROS level and apoptosis rate were detected by 2'

7'-Dichloroflurescin diacetate (DCFH-DA) and flow-cytometry

respectively. We also used Real-time PCR and Western blot to examine the effect of quercetin on oxidative stress and apoptosis mechanism-related molecules (NOX-4

p22phox

Bcl-2

Bax and Caspase-8)genes and protein expression level. Result: Quercetin of 10 μmol·L-1 has significant anti-oxidative and anti-apoptotic effect. Cell survival rate increased obviously in quercetin group compared with H2O2-model group (P<0.05)

ROS level and apoptosis rate significantly decreased (P<0.05

P<0.01). Moreover

quercetin had an up-regulation on PI3K

Bcl-2 and NOX4 (P<0.05) and down-regulation of p22phox

Bax and Caspase-8 (P<0.05). Meanwhile

quercetin can inhibit Bax

p22phox protein expressions (P<0.05)

and up-regulate PI3K

Bcl-2 protein expressions (P<0.05). Conclusion: Quercetin protects H9C2 cells from H2O2-induced oxidative injury and inhibit myocardial cell apoptosis by regulating NOX-4 p22phox

reducing intracellular ROS level

activating PI3K and increasing Bcl-2.

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