抑制miR-29a-3p表达对小鼠巨噬细胞凋亡的影响及机制  

Mechanism and influence of down-regulation of miR-29a-3p expression on apoptosis in mouse macrophage

作　　者：李木楠[1,2] 李双双 孟凡爽[1] 韩鹏 张文慧[1,2] 张林波[1,2] LI Munan;LI Shuangshuang;MENG fanshuang;HAN Peng;ZHANG Wenhui;ZHANG Linbo(College of Life Science, Jilin Agricultural University, Changchun 130118, China;Engineering Research Center for Bioreactor and Pharmaceutical Development, Ministry of Education, Changchun 130118, China)

机构地区：[1]吉林农业大学生命科学学院,吉林长春130118 [2]生物反应器与药物开发教育部工程研究中心,吉林长春130118

出　　处：《华南农业大学学报》2018年第1期64-69,共6页Journal of South China Agricultural University

基　　金：吉林省科技发展计划项目(20130101105JC;20140204018YY);吉林省教育厅"十三五"科学技术项目(2016-177JJKH20170304KJ);吉林农业大学博士科研启动基金(2015015)

摘　　要：【目的】研究miR-29a-3p下调表达对小鼠巨噬细胞凋亡及相关基因表达的影响,为探讨结核病发病机制、研发新的诊断与治疗方法提供依据。【方法】将重组抑制载体p EZX-AM02-miR-29a-3p转染RAW264.7细胞,24、36和48 h荧光显微镜观察转染效率,Real time-PCR法检测miR-29a-3p及凋亡相关基因caspase3、caspase7、caspase8、Bcl-2、Mcl-1和Bax的表达水平,流式细胞术检测RAW264.7细胞的凋亡率。【结果】重组抑制载体转染后,细胞内miR-29a-3p表达水平明显下降;而caspase7、caspase8、Bcl-2、Mcl-1和Bax等基因的表达出现不同程度的上调;流式细胞术分析发现随着转染时间延长,细胞凋亡率增加,且36 h凋亡率最高。【结论】p EZX-AM02-miR-29a-3p重组载体可抑制小鼠巨噬细胞中miR-29a-3p的表达,通过靶向上调caspase7、caspase8、Bcl-2和Mcl-1等基因的表达促进巨噬细胞的凋亡。【Objective】To study the influence of down-regulation of miR-29a-3p expression on apoptosis in mouse macrophage and expression of apoptosis-associated genes,and provide references for investigating the pathogenesis mechanism of mycobacterium tuberculosis(MTB)and developing new diagnosis and treatment methods.【Method】The recombinant inhibitor vector pEZX-AM02-miR-29a-3p was transfected into RAW264.7cells.Transfection efficiency was observed through fluorescence microscopy,the expression levels of miR-29a-3p and apoptosis-associated genes caspase3,caspase7,caspase8,Bcl-2,Mcl-1and Bax were detected by RT-PCR,and the apoptosis rates of RAW264.7cells were detected by flow cytometry at24,36,48h after transfection.【Result】After transfected with pEZX-AM02-miR-29a-3p recombinant inhibitor vector,miR-29a-3p expression in RAW264.7cells decreased significantly,while the target genes caspase7,caspase8,Bcl-2,Mcl-1and Bax were up-regulated at different levels.The results of flow cytometry indicated that the apoptosis rates increased as the transfection time progressed and reached the peak at36h.【Conclusion】The expression of miR-29a-3p in mouse macrophage are inhibited by transfecting with pEZX-AM02-miR-29a-3p,which promotes the apoptosis of macrophage by up-regulating the expression of target genes,suah as caspase7,caspase8,Bcl-2and Mcl-1.

关 键 词：miR-29a-3p 抑制载体 巨噬细胞 细胞凋亡 靶基因 

分 类 号：S854.4[农业科学—临床兽医学]