高铜诱导大鼠心肌细胞线粒体自噬

doi:10.11843/j.issn.0366-6964.2020.02.019

摘要/Abstract

摘要： 旨在探讨高铜对大鼠心肌细胞线粒体自噬的影响。将32只8月龄健康大鼠随机分为4组，每组8只，饲喂不同铜含量日粮（对照组：15 mg·kg^-1；高铜Ⅰ组：30 mg·kg^-1；高铜Ⅱ组：60 mg·kg^-1；高铜Ⅲ组：120 mg·kg^-1），连续饲喂6个月后采集心。检测心组织铜含量及其病理变化；荧光定量检测线粒体自噬相关基因LC3A、LC3B、Parkin、PINK1、p62的mRNA水平；蛋白印迹检测Parkin、PINK1、LC3B/LC3A的蛋白表达水平；免疫组化和免疫荧光检测LC3B的定位及表达。结果显示，高铜各组大鼠心组织内的铜含量均高于对照组。高铜各组出现心肌纤维排列疏松、无规则的现象。与对照组相比，高铜各组LC3A、LC3B mRNA表达量增加；高铜Ⅱ组、高铜Ⅲ组p62的mRNA表达量极显著降低；高铜各组PINK1 mRNA表达量显著升高，而Parkin mRNA的表达量有上升趋势但不显著。随着日粮中铜含量的增加，PINK1、Parkin、LC3B/LC3A的蛋白的表达量显著上调。LC3B定位于细胞质中，且高铜Ⅰ、Ⅱ、Ⅲ组LC3B表达量均显著增加。试验结果表明，高铜可诱导大鼠心肌细胞线粒体自噬的发生。

关键词: 铜, 大鼠, 心肌细胞, 线粒体自噬

Abstract: The aim of this study was to investigate the effects of the high level of copper on mitophagy in cardiomyocytes of rats. Thirty-two eight-month-old healthy rats were randomly divided into 4 groups with 8 rats in each group. The rats were fed diet with different content of copper (control group:15 mg·kg^-1, high-copper group Ⅰ:30 mg·kg^-1, high-copper group Ⅱ:60 mg·kg^-1, high-copper group Ⅲ:120 mg·kg^-1). After feeding for 6 months, heart tissues were collected from anesthetized rats. The content of copper and the pathological change of myocardial tissues were measured. The mRNA levels of LC3A, LC3B, Parkin, PINK1 and p62 were detected by real-time quantitative PCR. Additionally, the protein expression of Parkin, PINK1 and LC3B/LC3A were measured by Western blot and the expression and localization of LC3B was detected by immunohistochemistry and immunofluorescence. The results showed that content of copper in the heart of the high-copper groups was higher than that of the control group, the myocardial fibers were loose and irregular in the high-copper groups. Additionally, compared with the control group, the mRNA expression of LC3A and LC3B in the high-copper groups increased. The p62 mRNA expression in the high-copper group Ⅱ and Ⅲ was lower than that of control group. Furthermore, compared with the control group, the expression of PINK1 mRNA in the high-copper group was significantly increased, and the expression of Parkin mRNA was increased but not significantly. Meanwhile, with the increase of copper content in the diet, the protein expressions of PINK1, Parkin and LC3B/LC3A were significantly up-regulated. Moreover, LC3B was located in the cytoplasm, and the LC3B expression in high-coppergroups was significantly increased compared with the control group. These results suggested that high level of copper could induce mitophagy of rat cardiomyocytes.

Key words: copper, rat, cardiomyocyte, mitophagy

中图分类号:

S856.9

杨艳阳, 余文兰, 廖建昭, 杨帆, 裴若男, 常晓月, 邓纪昌, 黄坤玄, 唐兆新. 高铜诱导大鼠心肌细胞线粒体自噬[J]. 畜牧兽医学报, 2020, 51(2): 374-381.

YANG Yanyang, YU Wenlan, LIAO Jianzhao, YANG Fan, PEI Ruonan, CHANG Xiaoyue, DENG Jichang, HUANG Kunxuan, TANG Zhaoxin. High Level of Copper Induced Mitophagy in Cardiomyocytes of Rats[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(2): 374-381.

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