miR-499通过Drp1介导线粒体自噬保护缺氧/复氧心肌细胞

doi:10.3969/j.issn.1006-5725.2023.17.008

Abstract

Abstract:

Objective To explore the protective effect of miR?499 on myocardial ischemia?reperfusion injury and its possible mechanism from the aspect of mitochondrial autophagy. Methods Myocardial cell H9c2（2?1） was induced by ischemia/reperfusion （I/R）， and hypoxia/reoxygenation （H/R） myocardial cell model was established. The cells were treated with miR?499 mimics and/or P110. The cells were divided into BC group， I/R group， I/R + mi group， I/R + NC group， I/R + mi + P110 group and I/R + NC + P110 group. Cell proliferation was detected by CCK8 method. ROS， mitochondrial membrane potential and apoptosis were detected by flow cytometry. MDA， SOD and ATP contents were detected by their kit. The mRNA and protein expression levels of mitochondrial fusion， division and autophagy related genes Fis1， Mfn1， Parkin， LC3?Ⅱ， p62 and Drp1 were detected by qRT?PCR and Western blot. Mitochondrial autophagy was observed by transmission electron microscope. Results miR?499 mimics and/or P110 can reduce the proliferation inhibition rate and apoptosis rate of myocardial cells induced by I/R. The mitochondrial membrane potential decreased. The mitochondrial autophagy decreased and the ATP content increased. The gene and protein expression levels of Fis1， Parkin， LC3?Ⅱ and Drp1 decreased， while those of Mfn1 and p62 increased. The content of ROS and MDA in cells decreased， but the content of SOD increased. Conclusions miR?499 can reduce the occurrence of oxidative stress by reducing mitochondrial autophagy mediated by Drp1， thus protecting I/R?induced cardiomyocytes。

Key words: microRNA?499, Drp1, mitochondrial autophagy, oxidative stress, myocardial ischemia/reperfusion

CLC Number:

Jing WU,Zuqiong NIE,Wanling. YIN. MiR⁃499 protects hypoxia/reoxygenation （H/R） cardiomyocytes through Drp1⁃mediated mitochondrial autophagy[J]. The Journal of Practical Medicine, 2023, 39(17): 2196-2203.

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References 23

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引物名称	序列	扩增片段大小（bp）
GAPDH-F	CAAGTTCAACGGCACAG	138
GAPDH-R	CCAGTAGACTCCACGACAT
U6-F	CTCGCTTCGGCAGCACATATACT	93
U6-R	ACGCTTCACGAATTTGCGTGTC
miR-499-F	CTGGTGTCGTGGAGTCGG	56
miR-499-R	GGGGTTAAGACTTGCAGTG
Fis1-F	TTTGAATACGCCTGGTGC	116
Fis1-R	ACATAATCCCGCTGCTCC
Mfn1-F	GAATAATCGTTGGGATGCTT	241
Mfn1-R	AACTCCTGTAATCTTGCCTGA
Drp1-F	ATTGACATCTTGACCGCC	191
Drp1-R	TCCTGTGTGCTGTAGTTGCT
Parkin-F	CGCTCAACTTGGCTACTCC	129
Parkin-R	CACTCCTCGGCACCATACT
LC3-II-F	CTTCGCCGACCGCTGTA	194
LC3-II-R	GCGTGGGGTTGAGTTGC
p62-F	GGGATGACGACTGGACG	141
p62-R	TCTGTAGGAGCCTGGTGAG

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MiR⁃499 protects hypoxia/reoxygenation （H/R） cardiomyocytes through Drp1⁃mediated mitochondrial autophagy

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