人胚胎干细胞外泌体中miRNAs的鉴定及其对缺氧心肌细胞作用机制的探讨  

Identification of exosomal miRNAs in human embryonic stem cells and their protective effect on hypoxic cardiomyocytes

作　　者：黄冰鑫 陈景 吴文涛 周建荣 田苗 涂贾子超 刘湘 李晓红 陈寄梅 HUANG Bing-xin;CHEN Jing;WU Wen-tao;ZHOU Jian-rong;TIAN Miao;TU Jia-zi-chao;LIU Xiang;LI Xiao-hong;CHEN Ji-mei(Department of Cardiology,School of Medicine,South China University of Technology,Guangzhou 510006,China;Guangdong Provincial Key Laboratory of South China Structural Heart Disease,Guangdong Provincial People's Hospital,Guangdong Academy of Medical Sciences,Guangzhou 510080,China)

机构地区：[1]华南理工大学医学院,广东广州510006 [2]广东省人民医院(广东省医学科学院),广东省心血管病研究所,广东省华南结构性心脏病重点实验室,广东广州510080

出　　处：《中国病理生理杂志》2022年第4期592-598,共7页Chinese Journal of Pathophysiology

基　　金：国家重点研发计划(No.2018YFC1002600);国家自然科学基金资助项目(No.82170259);广东省登峰计划项目(No.DFJH2019;No.DFJH201802);广东省科技计划项目(No.2019B020230003)。

摘　　要：目的:胚胎干细胞来源的外泌体对于缺氧心肌细胞的作用机制未明,本研究将对其中微小RNA(miRNA)的角色及作用进行探讨。方法:提取并鉴定胚胎干细胞H9分泌的外泌体,将外泌体处理缺氧条件下培养的AC16心肌细胞,通过Western blot、细胞活力实验、流式细胞术、线粒体膜电位检测等方法分析外泌体对缺氧心肌细胞的作用;对H9细胞来源的外泌体进行转录组测序,分析其中高丰度的miRNAs对缺氧心肌细胞的可能作用机制。结果:成功获得H9细胞来源的外泌体,发现其可以促进缺氧情况下AC16细胞的存活,抑制线粒体膜电位降低及细胞凋亡。对外泌体中富集的miRNAs进行测序后发现miR-302a/b/d-3p在外泌体中含量丰富,且与对照组相比,经外泌体处理后AC16细胞内miR-302a/b/d-3p含量也显著增加,其与凋亡通路密切相关。结论:胚胎干细胞来源外泌体可以减少缺氧情况下心肌细胞AC16的凋亡,外泌体携带的miR-302a/b/d-3p可能是其发挥保护作用的关键因素之一。AIM:The mechanism of exosomes derived from embryonic stem cells on hypoxic cardiomyocytes is still unknown.This study explores the role of exosomal microRNAs(miRNAs)in embryonic stem cells.METHODS:Exosomes secreted by embryonic stem cell line H9 were isolated and identified.The AC16 cardiomyocytes were treated with exosomes under hypoxic conditions.The effects of exosomes on hypoxic cardiomyocytes were analyzed by Western blot,cell viability experiment,flow cytometry and mitochondrial membrane potential detection.Moreover,transcriptome sequencing of H9 cell-derived exosomes was performed to analyze the possible mechanism by which high-abundance miRNAs protected hypoxic cardiomyocytes.RESULTS:The H9 cell-derived exosomes were successfully isolated and proved to promote the survival of AC16 cells under hypoxia.The H9 cell-derived exosomes rescued the reduction of mitochondrial membrane potential,and reduced cell apoptosis.After transcriptome sequencing,we found miR-302a/b/d-3p was abundantly expressed in exosomes.Moreover,compared with control group,miR-302a/b/d-3p in AC16 cells was also significantly increased after treatment with exosomes.CONCLUSION:The H9 cell-derived exosomes reduce the apoptosis of AC16 cells under hypoxia,and miR-302a/b/d-3p in exosomes may serve as a critical role.

关 键 词：胚胎干细胞 外泌体 心肌细胞 缺氧 微小RNA 

分 类 号：R363.2[医药卫生—病理学] R541[医药卫生—基础医学]