微小RNA-29b调控表达在人骨关节炎软骨细胞中的生物学功能及其机制  

Expression of microRNA-29b in human osteoarthritic chondrocytes and its effect on biological function

作　　者：刘阳[1] 周颖 方舟 Liu Yang;Zhou Ying;Fang Zhou(Department of Clinical Medicine,Guizhou Medical University,Guiyang 550004,China;Department of Orthopedics,Affiliated Hospital of Guizhou Medical University,Guiyang 550004,China)

机构地区：[1]贵州医科大学临床医学系,贵阳550004 [2]贵州医科大学附属医院骨科,贵阳550004

出　　处：《中华实验外科杂志》2022年第7期1236-1239,共4页Chinese Journal of Experimental Surgery

摘　　要：目的探讨微小RNA-29b(miR-29b)在人骨关节炎软骨组织细胞中的表达和对骨关节炎软骨细胞生物学功能影响及其机制。方法利用实时荧光定量聚合酶链反应(qPCR)检测miR-29b在人骨关节炎软骨组织细胞和正常骨软骨组织细胞(2018年1月至2019年12月贵州医科大学附属医院骨科手术取样)中表达。培养人骨关节炎软骨细胞,并分成miR-29b inhibitor组、miR-29b mimics组、Negative Control组和Blank Control组。细胞计数法(CCK-8)和流式细胞术分别检测细胞增殖、凋亡;蛋白印迹法(Western blot)和qPCR分别检测各组细胞中Ⅱ型胶原蛋白(ColⅡ)、Ⅹ型胶原蛋白(ColⅩ)、基质金属蛋白酶-13(MMP-13)蛋白及基因mRNA的表达。组间比较采用单因素方差分析和t检验。结果人骨关节炎软骨组织细胞中miR-29b表达量显著高于正常骨组织(3.21±0.26比1.04±0.06,t=8.355,P<0.05)。48~96 h时,miR-29b inhibitor组骨关节炎软骨细胞吸光度(A)值显著高于Negative Control组(0.59±0.07、0.81±0.07、1.06±0.15比0.43±0.08、0.68±0.07、0.82±0.11,F=5.133、7.155、8.242,P<0.05);miR-29b inhibitor组细胞凋亡率显著低于Negative Control组[(5.23±0.89)%比(10.16±1.55)%,F=11.244,P<0.05];miR-29b inhibitor组细胞中ColⅡ蛋白和mRNA表达均显著高于Negative Control组(0.86±0.06比0.46±0.05和1.85±0.05比0.97±0.04,F=4.284、9.545,P<0.05),而ColⅩ、MMP-13蛋白和mRNA表达均显著低于Negative Control组(0.19±0.06、0.35±0.04比0.76±0.04、0.93±0.06)和(0.36±0.04、0.51±0.05比0.96±0.07、0.99±0.06,F=5.299、6.024、6.973、7.111,P<0.05)。结论miR-29b在人骨关节炎软骨细胞中表达升高,其促进ColⅩ、MMP-13并抑制ColⅡ基因mRNA和蛋白表达,进而发挥抑制人骨关节炎软骨细胞增殖和促进细胞凋亡的作用。Objective To study the effect of microRNA-29b(miR-29b)expression on biological function and mechanism in human osteoarthritic chondrocytes.Methods Real-time fluorescence quantitative polymerase chain reaction(qPCR)was used to detect the expression of miR-29b in human osteoarthritic cartilage tissue cells and normal osteochondral tissue cells(From January 2018 to December 2019,orthopaedic surgically-resected samples were taken from the Affiliated Hospital of Guizhou Medical University).Human osteoarthritis chondrocytes were cultured and divided into miR-29b inhibitor group,miR-29b mimics group,negative control group and blank control group.Cell proliferation and apoptosis were detected by cell counting kit-8(CCK-8)assay and flow cytometry respectively.Western blotting and qPCR were used to detect the expression of CollagenⅡ(ColⅡ),CollagenⅩ(ColⅩ),matrix metalloproteinase-13(MMP-13)protein and mRNA respectively.Data were analyzed by one-way ANOVA and t-test.Results The expression of miR-29b in chondrocytes of human osteoarthritis was significantly higher than that in normal bone(3.21±0.26 vs.1.04±0.06,t=8.355,P<0.05].At 48-96 h,the absorbance value of osteoarthritis chondrocytes in miR-29b inhibitor group was significantly higher than megative control group(0.59±0.07,0.81±0.07,1.06±0.15 vs.0.43±0.08,0.68±0.07,0.82±0.11,F=5.133,7.155,8.242,all P<0.05).The apoptosis rate of miR-29b inhibitor group was significantly lower than negative control group[(5.23±0.89)%vs.(10.16±1.55)%,F=11.244,P<0.05].The expression of ColⅡprotein and mRNA in miR-29b inhibitor group was significantly higher than negative control group(0.86±0.06 vs.0.46±0.05 and 1.85±0.05 vs.0.97±0.04,F=4.284,9.545,all P<0.05),and the expression of ColⅩ,MMP-13 protein and mRNA was significantly lower than negative control group(0.19±0.06,0.35±0.04 vs.0.76±0.04,0.93±0.06 and 0.36±0.04,0.51±0.05 vs.0.96±0.07,0.99±0.06,F=5.299,6.024 and 6.973,7.111,all P<0.05).Conclusion MiR-29b is highly expressed in human osteoarthritic chondro

关 键 词：骨关节炎 软骨细胞 微小RNA 增殖 凋亡 

分 类 号：R684.3[医药卫生—骨科学]