来源：风湿病与关节炎,2017,6（9）:10-14.

补肾活血方调节miRNA-140-5p表达干预人原代软骨细胞退变的

研究（基础研究）

姚　楠¹，陈　能^2,3，刘文刚²，许学猛²，黄丹娥¹，蔡大可¹，赵传喜²

　　【摘　要】目的：研究补肾活血方含药血清对白细胞介素-1β（IL-1β）刺激的人原代软骨细胞miRNA-140及其靶基因表达的影响。方法：通过采用生理盐水和补肾活血方分别灌胃雌性SD大鼠制备空白血清和含药血清。将人软骨细胞种植于6孔板并且分为3组。空白对照组和模型对照组细胞培养的血清采用大鼠空白血清，含药血清组细胞培养的血清采用大鼠含药血清。另外，模型对照组和含药血清组细胞加入10 ng·mL^-1 IL-1β。以上各组细胞设3个复孔，继续培养细胞至IL-1β刺激后24 h收集各组细胞提取总RNA。采用RT-qPCR检测软骨细胞miRNA-140-5p以及聚蛋白样金属蛋白酶-5（ADAMTS-5）、胰岛素样生长因子结合蛋白5（IGFBP5）和基质金属蛋白酶-13（MMP-13）mRNA表达。结果：与空白对照组比较，模型对照组软骨细胞miRNA-140-5p表达和IGFBP5 mRNA表达明显降低（P < 0.05），ADAMTS-5和MMP-13mRNA表达明显升高（P < 0.05）；与模型对照组比较，补肾活血方含药血清显著升高了IL-1β刺激的软骨细胞miRNA-140-5p表达和IGFBP5 mRNA表达（P < 0.05），并且显著降低ADAMTS-5和MMP-13 mRNA表达（P < 0.05）。结论：补肾活血方含药血清可以抑制IL-1β导致的人原代软骨细胞退变，其作用机制与提高miRNA-140-5p和IGFBP5 mRNA表达，降低ADAMTS-5和MMP-13 mRNA表达有关。

　　【关键词】　骨关节炎，膝；补肾活血方；miRNA-140-5p；软骨细胞

Study on the Effects of Bushen Huoxue Fang（补肾活血方） on Regulating the

Expression of miRNA-140-5p to Intervene Human Primary Chondrocyte Degeneration

YAO Nan,CHEN Neng,LIU Wen-gang,XU Xue-meng,HUANG Dan-e,CAI Da-ke,ZHAO Chuan-xi

　　【ABSTRACT】Objective:To study the effects of drug serum on the expression of miRNA-140 and its target genes in human primary chondrocytes stimulated by IL-1β.Methods:The blank serum and drug serum were prepared by intragastric administration of normal saline and Bushen Huoxue Fang（补肾活血方）respectively to female SD rats.Human chondrocytes were seeded on six orifice plates and divided into three groups.The blank control group and the model control group were cultured with blank serum,while the drug serum group was cultured with drug serum.In addition,10 ng·mL^-1 IL-1β was added to the model control group and the drug serum group.The cells in each group were cultured in three double holes and then collected after the stimulation of IL-1β for 24 hours to extract the total RNA.RT-qPCR was used to detect the expression of miRNA-140-5p,ADAMTS-5,IGFBP5 and MMP-13 mRNA.Results:Compared with the blank control group,the expressions of miRNA-140-5p and IGFBP5 mRNA in the model control group were significantly reduced （P < 0.05）,while the expressions of ADAMTS-5 and MMP-13 mRNA were significantly increased（P < 0.05）.Compared with the model group,the serum containing Bushen Huoxue Fang significantly increased the expressions of chondrocyte miRNA-140-5p and IGFBP5 mRNA stimulated by IL-1β（P < 0.05）,and significantly decreased the expressions of ADAMTS-5 and MMP-13 mRNA（P < 0.05）.Conclusion:The serum containing Bushen Huoxue Fang can inhibit the degeneration of human primary chondrocytes induced by IL-1β,and its mechanism is related to the increase of the expressions of miRNA-140-5p and IGFBP5 mRNA and the decrease of the expressions of ADAMTS-5 and MMP-13 mRNA.

　　【Keywords】 osteoarthritis,knee;Bushen Huoxue Fang（补肾活血方）;miRNA-140-5p;chondrocytes