miR-140在不同阶段骨关节炎软骨细胞中的表达规律及功能

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摘要目的观察人膝关节不同退变阶段骨关节炎(osteoarthritis，OA)软骨细胞中微小RNA-140(micro RNA-140，miR-140)表达规律及转染双链miR-140模拟物/抑制物(double strands-miR-140 mimic/inhibitor，ds-miR-140 mimic/inhibitor)对OA软骨细胞功能的影响。方法根据Kellgren and Lawrence X线分级，膝关节软骨分为正常组(O级)、中期OA组(Ⅲ级)和晚期OA组(Ⅳ级)。各组收集软骨标本6～8例，体外分离及培养软骨细胞，采用第2代软骨细胞进行检测。运用实时荧光定量聚合酶链式反应(polymerase chain reaction，PCR)检测各组miR-140相对表达量及Ⅱ型胶原(collagentype Ⅱa，COL₂A1)及Ⅹ型胶原(Collagentype Xa，COL10A1)的表达情况。向中期OA组软骨细胞转染ds-miR-140 mimic／inhibitor后，观察软骨形成因子SRY基因(SRYrelated high mobility group box gene 9，SOX9)，COL₂A1及COL10A1表达情况。结果 miR-140在OA软骨细胞中表达显著下降，中、晚期OA组miR-140和正常组相比分别减少78.9％和94.5％(P＜0.05)，晚期OA组miR-140较中期OA组减少73.9％(P＜0.05)。中、晚期OA组SOX9的mRNA较正常组分别减少34.1％和49.3％(P＜0.05)，COL₂A1的mRNA较正常组分别减少36.7％和78.7％(P＜0.05)，中、晚期OA组COL10A1的mRNA较正常组分别增加1.66倍和2.10倍(P＜0.05)。向中期OA组转染ds-miR-140 mimic／inhibitor后，SOX9的mRNA分别增加1.95倍和减少59.1％，COL₂A1的mRNA分别增加1.87倍和减少68.8％(P＜0.05)，COL10A1的mRNA分别减少73.0％和增加1.96倍(P＜0.05)。结论 miR-140在OA软骨细胞中表达显著下降，且随病情加重而进一步减少；转染ds-miR-140 mimic可增加SOX9和COL₂A1的表达，同时降低COL10A1的表达。

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关键词 ：骨关节炎, 软骨细胞, 微小RNA140, Ⅱ型胶原, X型胶原

Abstract：Objective To investigate the expression miR-140 in different stage of osteoarthritis and its function after transfecting ds-miR-140 mimic/inhibitor into chondrocytes. Methods Cartilage was classified into three groups，normal group，mid-staged group and advanced-staged group，according to the Kellgren and Lawrence criterion.6-8 cases of normal or OA cartilage were collected and chondrocytes were isolated.The second generation of chondrocytes were used to test.The levels of miR-140，COL₂A1 and COL10A1 mRNA were measured with real-time fluorescent quantitative PCR.The ds-miR-140 mimic/inhibitorwere transfected into the mid-staged group，and the expression of COL₂A1，COL10A1 and SOX9 were measured.Results Compared with the normal group，miR-140 expression in the mid- and advanced-staged group was decreased by 34.1％ and 49.3％ respectively，and its expression in advanced-staged group was decreased by 73.9％.Beside，SOX9 expression in the mid- and advanced-staged group was decreased by 75％ and 82.7％respectively，COL₂A1 expression decreased by 36.7％ and 78.7％ respectively，while COL10A1 expression increased to 1.66 times and 2.10 times respectively.After transfecting ds-miR-140 mimic/inhibitor into the mid-staged group，SOX9 expression was increased by 1.95 times or decreased by 59.1％，COL₂A1 expression was increased by 1.87 times or decreased by 68.8％，and COL10A1 expression was decreased by 73.0％ or increased by 1.96 times.Conclusion The expression level of miR-140 decreased in osteoarthritic chondrocytes and was negatively related to the severity of OA.Overexpression its level by transfecting ds-miR-140 mimic into chondrocytes could up-regulate SOX9 and COL-2A1 expression while down-regulate COL10A1 expression.

Key words： osteoarthritis chondrocyte microRNA-140 collagen type Ⅱ collagen type X

基金资助:国家自然科学基金项目(81672219、81601936)
四川省科技厅科技支撑计划(2014SZ0023-2)

通讯作者: 沈彬

作者简介: 李灿锋(1987－)，男，研究生在读，四川大学华西医院骨科，610041。

引用本文:

李灿锋 1，陈卓 2，杨静 1，周宗科 1，康鹏德 1，裴福兴 1，沈彬 1*. miR-140在不同阶段骨关节炎软骨细胞中的表达规律及功能[J]. 实用骨科杂志, 2017, 23(11): 995-999.
Li Canfeng 1，Chen Zhuo 2，Yang Jing 1，et al. Expression and Function of miR-140 in Different Stage Chondrocytes of Patients with Osteoarthritis. sygkzz, 2017, 23(11): 995-999.

链接本文:

http://www.sygkzz.com/CN/ 或 http://www.sygkzz.com/CN/Y2017/V23/I11/995

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