miR-30a-5p靶向作用蛋白激酶B基因促进骨关节炎患者软骨细胞的凋亡  

miR-30a-5p promotes the apoptosis of chondrocytes in patients with osteoarthritis by targeting protein kinase B

作　　者：沈鹏飞[1] 瞿玉兴[1] 王斌[1] 徐建达[1] 魏康[1] 谢子康[1] 马勇[2] 

机构地区：[1]南京中医药大学附属常州中医医院骨科,江苏常州213004 [2]南京中医药大学骨伤科教研室

出　　处：《中华医学杂志》2017年第39期3079-3084,共6页National Medical Journal of China

基　　金：常州市卫生计生委青年科技项目（QN201501）

摘　　要：目的观察miR-30a-5p在骨关节炎（OA）患者软骨组织中的表达及其对软骨细胞凋亡的作用机制。方法2015年5月至2016年12月，收集南京中医药大学附属常州中医医院骨科289例OA软骨组织标本。聚合酶链反应（qPCR）检测不同患者软骨组织中miR．30a-5p以及蛋白激酶B（Akt）mRNA的表达情况，Tunel法检测软骨细胞凋亡情况，免疫印迹法组织和细胞中相关蛋白的表达，流式细胞仪检测不同细胞凋亡情况和细胞周期。结果OA患者软骨组织miR．30a-5p相对表达量显著高于非OA[（3．64±0．95）比（1．03±0．31），P〈0．05]，Akt基因表达较非OA显著下调[（0．41±0．17）比（1．08±0．26），P〈0．05]；miR-30a-5p在OA患者软骨组织中的表达与AktmRNA表达呈负相关（r=0．7293，P〈0．001），与细胞凋亡率呈正相关（r=0．8475，P〈0．001）；在SW1353细胞内miR-30a-5p靶向负调控Akt基因表达，高表达的miR-30a-5p显著下调p-Akt，IkB-α，p-IkB-α，p65，p—p65和mTOR基因，p-mTOR蛋白表达（P〈0．05）；与正常SW1353细胞相比，转入miR-30a-5p—mimics的SW1353细胞凋亡率增加9．65倍，G0／G1期细胞增加1．37倍，S期细胞降低了60．94％，G2／M期细胞降低了19．53％。结论miR-30a-5p在OA患者软骨组织中呈现高表达，并且其高表达可以通过靶向作用Akt基因而将软骨细胞阻滞在GO／G1期，进而诱导软骨细胞的凋亡。Objective To investigate the expression of miR-30a-Sp in cartilage of osteoarthritis patients, and to explore its mechanism of chondrocyte apoptosis. Methods From May 2015 to December 2016, tissue specimen of 289 patients with osteoarthritis was collected in Department of Orthopedics, Changzhou traditional Chinese Medicine Hospital Affiliated to Nanjing University of Chinese Medicine. The expression of miR-30a-Sp and protein kinase B （Akt） mRNA in cartilage of different patients was detected by qPCR. The apoptosis of chondrocytes was detected by Tunel method. The expression of related proteins in tissues and ceils was detected by immunoblotting, and apoptosis and cell cycle were detected by flow cytometry. Results The expression of miR-30a-Sp in OA patients was significantly higher than control patients （ P 〈 0. 05 ）, but Aktwas positively related [ （ 3.64 ± 0. 95 ） vs （ 1.03 ± 0. 31 ）, P 〈 0. 05 ]. The expression of miR-30a-Sp in cartilage of OA patients was negatively correlated with Akt mRNA expression （r =0. 729 3, P 〈0. 001 ）, but it had a positive correlation with the apoptotic rate （r = 0. 847 5, P 〈 0. 001 ）. miR-30a-Sp targets negative regulation of Akt gene expression in SW1353 cells, and the expression of p-Akt, IkB-α, p-IkB-α, p65, p-p65 and mTOR and p-mTOR were significantly down-regulated by miR- 30a-Sp （P〈0. 05）. Compared with normal SW1353 ceils, the apoptosis rate of SW1353 cells which was transfected with miR-30a-Sp-mimics increased by 9.65 times, G0/G1 phase cells increased by 1.37 times, S phase ceils decreased by 60. 94%, G2/M phase cells decreased 19.53%. Conclusion miR-30a-Sp is highly expressed in cartilage of osteoarthritis patients, and its high expression can block chondrocytes in G0/ G1 phase by targeting Akt gene, and induce apoptosis of chondrocytes.

关 键 词：软骨细胞 骨关节炎 软骨组织 miR-30a-5p 蛋白激酶B 

分 类 号：R684.3[医药卫生—骨科学]