中文摘要：

目的：研究miR-142-3p对自噬相关基因ATG4c的靶向调控作用,探究miR-142-3p影响RAW264.7细胞自噬途径的作用机制。方法：生物信息学软件分析miR-142-3p的靶基因为ATG4c,构建pMIR-Report-ATG4c和pMIR-Report-ATG4c mut重组质粒,双荧光素酶报告系统、qRT-PCR、Western blot验证miR-142-3p与ATG4c的靶向作用;将做不同处理的RAW264.7细胞分为4组：正常细胞作为对照、50ng/ml雷帕霉素作用2h、EBSS饥饿作用12 h、10 nmol/L的3-甲基腺嘌呤（3-MA）作用12h后,实时荧光定量PCR（qRT-PCR）检测miR-142-3p不同干预组中的相对表达情况;将miR-142-3p mimics、miR-142-3p inhibitor及miR-142-3p control分别转染到RAW264.7细胞中,检测miR-142-3p和LC3Ⅱ的相对表达。结果：双荧光素酶报告系统、qRT-PCR、Western blot验证miR-142-3p通过靶向作用于ATG4c的3＇-UTR抑制其表达;与对照组相比,雷帕霉素和饥饿处理的RAW264.7细胞miR-142-3p明显上调,而3-MA处理组miR-142-3p明显下调;与miR-142-3p control组相比,转染miR-142-3p mimics组中LC3Ⅱ蛋白表达显著下调,而miR-142-3p inhibitor组中表达显著上调。结论：miR-142-3p通过靶向调控自噬相关基因ATG4c,参与RAW264.7小鼠巨噬细胞自噬的调控。

英文摘要：

Objective： To study the mechanism of miR-142-3p regulated Autophagy Association Gene（ ATG4c）,to explore autophagy pathway in RAW264. 7 macrophages. Methods： To predict the target genes of miR-142-3p by Bioinformatics Software—ATG4c. Then to establish pMIR-Report-ATG4 c and pMIR-Report-ATG4 c mut recombinant plasmid,in order to confirm the regulatory relation between miR-142-3p and ATG4 c through dual luciferase assay,qRT-PCR,Western blot. Four groups was assigned by different treatment,they were normal cells control group,2 h 50 ng/ml rapamycin treated group,12 h EBSS hunger treated group and 12 h 10 nmol/L 3-methyl adenine（ 3-MA） treated group,to detect miR-142-3p relative expression in RAW264. 7 cell line by quantitative RealTime PCR（ qRT-PCR）. Mi R-142-3p mimics,miR-142-3p inhibitor and miR-142-3p control was transfected in RAW264. 7 cells respectively,the relative expression of miR-142-3p and LC3Ⅱ expression was examined by qRT-PCR and Western blot. Results： It revealed the target gene（ ATG4c） of miR-142-3p by Dual luciferase assay,qRT-PCR and Western blot; Compared with the control group,the expression of miR-142-3p was obviously up-reguated from the rapamycin and hunger treated groups in RAW264. 7cell line,on the contrary,miR-142-3p was obviously down regulated from the 3-MA treated group in RAW264. 7 cell line; meanwhile,compared with the control group,LC3Ⅱ expression was significantly lower from miR-142-3p mimics group; however,LC3 Ⅱ expression was significantly higher from miR-142-3p inhibitor group. Conclusion： Mi R-142-3p can target associated genes ATG4 c to regulate autophagy in RAW264. 7 cell line.