MicroRNA-145在原发性膝关节骨关节炎软骨中的表达及意义

Expression of microRNA-145 in cartilage of primary knee OA and its biological effects

目的明确microRNA-145(miR-145)在骨关节炎(OA)组织中的表达水平及miR-145对软骨细胞胶原蛋白表达的作用。方法收集20例膝关节OA患者的膝关节软骨和10例正常患者的膝关节软骨,根据改良的Mankin评分标准对OA关节软骨进行分组。使用RT-PCR方法检测miR-145在关节炎组织及TNF-α处理的软骨细胞中的表达,软骨细胞中X型胶原、RANK、MMP13、ALP的蛋白表达用Western blot检测。结果 miR-145在原发性OA和正常膝关节软骨中均能检测到表达,RT-PCR结果表明,miR-145的表达在不同退变程度的OA软骨组织及软骨细胞中有表达差异,而且随着OA退变程度的提高,miR-145在软骨细胞及软骨组织中的表达呈下降趋势,在重度退变的OA软骨中,上述差异与正常及轻度退变软骨有统计学意义(P<0.05)。将miR-145转染至培养的软骨细胞,Western blot对软骨细胞进行检测分析,MMP13、X型胶原纤维蛋白受到抑制。结论原发性膝关节OA的软骨组织及细胞中miR-145的表达较正常软骨的表达明显减低,而且,随着软骨退变程度的加重,miR-145表达呈下降趋势;miR-145可以抑制软骨细胞MMP13、X型胶原、RANK的表达。

Objective： To identify the expression and activity of miR-145 in cartilage osteoarthritis patients,and the effect of miR-145 on expression of in chondrocytes. Methods： The samples of cartilage were obtained from twenty patients with osteoarthritis and ten patients with normal knee. The samples of osteoarthritic cartilage were classified based on Man Kin score standard. The real-time PCR was used to identify the activity of miR-145 in arthritis tissue and the chondrocytes treated by TNF-α. Western Blot technique were used to evaluate the expressions of matrix metalloproteinase（MMP13）,collagen X,receptor Activator for Nuclear Factor-κ B RANK（RANK） and alkaline phosphatase（ALP）. The statistical analysis was performed using the Statistical Package for the Social Sciences（SPSS） version 19. 0（SPSS,Chicago,IL,USA）. Results：The expression of miR-145 was detected in both primary OA and normal knee cartilage. The results of RT-PCR showed that the expression of miR-145 was differentially expressed in OA cartilage and cartilage with different degrees of degeneration. The expressions of miR-145 in chondrocytes and cartilage decreased with the degree of OA degeneration. In the severely degenerated OA cartilage,the difference was statistically significant compared with normal and mild degenerative cartilage（P〈0. 05）. MiR-145 was transfected into cultured chondrocytes,and chondrocytes were analyzed by western blot. The activity and expression of MMP13,collagen X and RANK were disturbed by miR-145. Conclusion： Expression and activity of miR-145 in normal cartilage are significantly higher than those in degenerative cartilage,and in corralation with degenerative severity of cartilage positively. miR-145 can inhibit the expression of MMP13,type X collagen and RANK in chondrocytes.