miR-140对早期骨关节炎软骨细胞衰老的调控作用及机制  

Regulatory effect and mechanism of miR-140 on chondrocyte senescence in early-stage osteoarthritis

作　　者：李兰 梁明玮 陆燕蓉 斯海波 沈彬 LI Lan;LIANG Ming-wei;LU Yan-rong;SI Hai-bo;SHEN Bin(West China Hospital,Sichuan University,Chengdu 610041,China)

机构地区：[1]四川大学华西医院

出　　处：《中国矫形外科杂志》2020年第3期252-259,共8页Orthopedic Journal of China

基　　金：国家自然科学基金项目(编号：81802210、81672219);四川省科技厅重点项目(编号：2018SZ0223、2018SZ0250)

摘　　要：[目的]探索miR-140对早期骨关节炎(OA)软骨细胞衰老的调控作用及机制。[方法]收集人膝关节正常、早期OA(E-OA)和中晚期OA(ML-OA)软骨组织,酶消化法获得相应组别软骨细胞。首先,PCR检测各组miR-140水平,流式细胞仪分析细胞周期,衰老相关β-半乳糖苷酶(SA-βGal)试剂盒分析SA-βGal活性,PCR和免疫细胞化学法检测衰老分子p16^INK4a、p21和p53的表达。然后,取E-OA软骨细胞设空白、IL-1β、miR-Scr和miR-140 mimic组并给予相应干预,检测并比较各组miR-140水平。最后,取E-OA软骨细胞设空白、IL-1β、miR-Scr+IL-1β和miR-140 mimic+IL-1β组并给予相应干预,分析和比较各组细胞周期、SA-βGal活性及衰老分子表达情况。[结果]正常、E-OA和ML-OA三组间miR-140水平、G0/G1期细胞比例(G0/G1%)、SA-βGal活性及衰老分子水平比较差异均有统计学意义(P<0.05),miR-140水平与OA分级呈显著负相关(P<0.05),而G0/G1%、SA-βGal活性及衰老分子阳性率与OA分级呈显著正相关(P<0.05)。在E-OA软骨细胞中,与空白组相比,IL-1β组miR-140水平显著降低(P<0.05),miR-Scr组无明显改变(P>0.05),miR-140 mimic组显著升高(P<0.05)。在E-OA软骨细胞中,IL-1β组G0/G1%、SA-βGal活性及衰老分子表达均显著高于空白组(P<0.05)、与miR-Scr+IL-1β组相比差异无统计学意义(P>0.05),miR-140+IL-1β组G0/G1%、SA-βGal活性及衰老分子表达均显著低于miR-Scr+IL-1β组(P<0.05)。[结论]软骨细胞衰老与OA发生发展密切相关,上调miR-140表达可有效抑制E-OA软骨细胞衰老,是延缓E-OA进展的潜在治疗策略。[Objective]To explore the regulatory effect and mechanism of miR-140 on chondrocyte senescence in early-stage osteoarthritis(OA).[Methods]Human articular cartilage from normal,early-stage OA(E-OA)and middle-late-stage OA(ML-OA)knee joint were collected,and the chondrocytes were obtained by enzymatic digestion.First,the levels of miR-140 in each group were detected by PCR,the cell cycle was analyzed by flow cytometer,the activity of SA-βGal was analyzed by senescence-associated β-galactosidase(SA-βGal)kit,and the expression of senescence-associated molecules,including p16^INK4a,p21 and p53,were detected by PCR and immunocytochemical staining.Then,E-OA chondrocytes were divided into blank,IL-1β,miR-Scr and miR-140 mimic groups and corresponding interventions were performed,and miR-140 levels in each group were detected and compared.Finally,E-OA chondrocytes were divided into four groups:blank,IL-1β,miR-Scr+IL-1β and miR-140 mimic+IL-1β,and the corresponding interventions were performed.The cell cycle,SA-β Gal activity and the expression of senescence-associated molecules in each group were analyzed and compared.[Results]There were significant differences in miR-140 levels,ratio of cells in G0/G1 phase(G0/G1%),SA-β Gal activity and levels of senescence-associated molecules between the normal,E-OA and ML-OA groups(P<0.05),meanwhile,there was a significant negative correlation between miR-140 level and OA severity(P<0.05),while G0/G1%,SA-β Gal activity and senescence-associated molecules levels were significantly positively correlated with OA severity(P<0.05).In E-OA chondrocytes,compared with the blank group,miR-140 levels were significantly decreased in the IL-1β group(P<0.05),not significantly changed in the miR-Scr group(P>0.05),and significantly increased in the miR-140 mimic group(P<0.05).Furthermore,in E-OA chondrocytes,G0/G1%,SA-β Gal activity and senescence-associated molecules levels were significantly higher in the IL-1βgroup than in the blank group(P<0.05),were similar between IL-1β and miR-S

关 键 词：早期骨关节炎 microRNA-140 软骨细胞 衰老 

分 类 号：R684.3[医药卫生—骨科学]