摘要
目的:研究miR-199a-5p对幼年类风湿关节炎(JRA)软骨细胞增殖、凋亡的影响和潜在的分子机制。方法:qRT-PCR检测软骨细胞中miR-199a-5p和PDCD5 RNA的表达,Western blot检测PDCD5、增殖相关蛋白CyclinD1、p21和p27以及凋亡相关蛋白Bcl-2、Bax和cleaved-caspase3的表达,MTT法检测软骨细胞增殖,流式细胞术测定软骨细胞凋亡率,双荧光素酶报告系统验证miR-199a-5p和PDCD5的调控关系。结果:与正常软骨细胞相比,在类风湿关节炎(RA)软骨细胞中miR-199a-5p表达量显著降低(P<0.05),而PDCD5 mRNA和蛋白表达量则显著升高(P<0.05);过表达miR-199a-5p和抑制PDCD5表达均可促进RA软骨细胞增殖,抑制RA软骨细胞凋亡;双荧光素酶报告系统结果表明,miR-199a-5p靶向负调控PDCD5的表达;过表达PDCD5逆转了miR-199a-5p过表达对软骨细胞增殖和凋亡的作用。结论:miR-199a-5p通过靶向调控PDCD5表达调控RA软骨细胞增殖和凋亡,miR-199a-5p是RA的潜在分子靶点。
Objective:To investigate the efffects of miR-199a-5p on proliferation and apoptosis of chondrocytes in jvenile rheumatoid arthritis(JRA)and the underlying molecular mechanism.Methods:RNA expression levels of miR-199a-5p and PDCD5 were detected by quantitative real-time polymerase chain reaction(qRT-PCR).The protein expression levels of PDCD5,proliferation-related proteins CyclinD1,p21 and p27,and apoptosis-related proteins Bcl-2,Bax,and cleaved-caspase3 were detected by Western blot.Cell proliferation of chondrocyte was measured by MTT assay.Apoptotic rate of chondrocyte was detected by flow cytometry.And the relationship between miR-199a-5p and PDCD5 was verified by dual-luciferase reporter assay system.Results:Compared with normal chondrocytes,the expression level of miR-199a-5p in RA chondrocytes was significantly decreased(P<0.05),while the expression levels of PDCD5 mRNA and protein were significantly increased(P<0.05).Overexpression of miR-199a-5p and inhibition of PDCD5 expression both promoted the proliferation and inhibited the apoptosis of RA chondrocytes.The results of the dual-luciferase reporting system suggested that miR-199a-5p negatively regulated the expression of PDCD5.Overexpression of PDCD5 reversed the effect of miR-199a-5p overexpression on the proliferation and apoptosis of RA chondrocytes.Conclusion:MiR-199a-5p regulates the proliferation and apoptosis of RA chondrocytes by targeting PDCD5.MiR-199a-5p is a potential molecular target for rheumatoid arthritis.
作者
符艺影
许志有
韩萍
董战玲
郑诗华
王小花
FU Yi-Ying;XU Zhi-You;HAN Ping;DONG Zhan-Ling;ZHENG Shi-Hua;WANG Xiao-Hua(Department of Pediatrics,the Third People′s Hospital of Haikou,Haikou 571100,China)
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2020年第11期1374-1379,共6页
Chinese Journal of Immunology
