牛蒡子苷元对体外培养软骨细胞Ⅱ型胶原表达的影响  

Expression of Collagen Ⅱ mRNA for Arctigenin on Chondrocytes In Vitro

作　　者：陈世宣 冯伟[1,3] 卢远坚 李安琪[1] 张增乔 刘益杰[1] 曹彦俊[1] 

机构地区：[1]上海中医药大学,上海201203 [2]温州市中西医结合医院,温州325000 [3]上海市第七人民医院,上海200137

出　　处：《中国中医骨伤科杂志》2017年第10期6-10,共5页Chinese Journal of Traditional Medical Traumatology & Orthopedics

基　　金：上海市自然基金资助项目(15ZR1441000);上海市卫生和计划生育委员会中医药科研课题(2016LP037);上海市浦东新区卫计委卫生科技资助项目(PW2014A-20)

摘　　要：目的:探讨牛蒡子苷元(Arctigenin,ARC-G)对软骨细胞的增殖,及对Ⅱ型胶原(Collagen Ⅱ,Col Ⅱ)、Y染色体性别决定相关高泳动蛋白盒基因9(SRY-related high mobility group-box gene 9,SOX9)和基质金属蛋白酶-13(Matrix Metalloproteinase-13,MMP-13)表达的影响。方法:体外分离培养软骨细胞,取第1代细胞进行实验干预。实验分为空白组(ARC-0组)、低浓度牛蒡子苷元组(ARC-L组)和高浓度牛蒡子苷元组(ARC-H组)。各组分别予相应干预措施处理48h后,以SABC免疫细胞化学染色法观察Col Ⅱ的表达情况;以CCK-8法检测软骨细胞的增殖情况;以Real-time quantitative PCR检测Col Ⅱ,SOX9及MMP-13 mRNA的表达情况;以酶联免疫吸附法(ELISA)检测细胞培养上清液中Col Ⅱ的含量。结果:1)ARC-L组和ARC-H组的Col Ⅱ阳性染色强度均强于ARC-0组,其中ARC-H组染色最强;2)与ARC-0组相比,在不同时间点ARC-L组和ARC-H组均可促进软骨细胞的增殖,且ARC-H组促增殖作用更显著,除24h和96h的ARC-L组外其余各时间点差异均有统计学意义(P<0.05);3)与ARC-0组相比,ARC-L组和ARC-H组均可上调ColⅡ和SOX9 mRNA的表达,下调MMP-13 mRNA的表达,且ARC-H组作用更为显著,除ARC-L组Col Ⅱ及MMP-13 mRNA外,差异均有统计学意义(P<0.05);4)与ARC-0组相比,ARC-L组和ARC-H组均可促进Col Ⅱ蛋白的表达,且ARC-H组作用更为显著,除ARC-0组和ARC-L组外,差异均有统计学意义(P<0.05)。结论:牛蒡子苷元可通过上调SOX9和Col Ⅱ mRNA的表达,下调MMP-13 mRNA的表达以促进Col Ⅱ蛋白的合成。Objective：To detect the effect of arctigenin on chondrocytes proliferation, and on expression of Collagen Ⅱ , SRY-related highmobility group-box gene 9 （SOX 9）, and matrix metallopr-oteinase-13 （MMP-13）mRNA. Methods： Chondrocytes were isolated and then cultured in vitro, the first generation of chondrocytes were used for experimental intervention. The chondrocytes were divided into blank group （ARC-0 group） ,low concentration of arctigenin group（ARC- L group）, and high concentration of arctigenin group （ARC- H group）. The chondrocytes in each group were respectively treated for 48 h by the corresponding intervention methods, then the expression of Col Ⅱ was observed by immunocytochemisty SABC method. The chondrocytes proliferation was detected by Cell counting kit-8 （CCKS） method. The expression of Col Ⅱ ,SOX 9,and MMP-13 mRNA were detected by real-time quantitative PCR. And the collagen Ⅱ content of supernatants was detected by enzyme-linked immunosorbent assay（ELISA）. Results： 1）The positive staining potency of ARC-L group and ARC-H group were stronger than ARC-0 group,and ARC-H group was the strongest one. 2）The ARC- L group and the ARC-H group could promote chondrocytes proliferation at different point of time compared with ARC-0 group, and the ARC-H group was more significantly, the optical density were significantly different at other all-time points except the 24 h and 96 h point（P〈 0.05）. 3）Compared with ARC-0 group, both of the ARC-L group and the ARC-H group were able to up-regulate the expression of Col ]l and SOX9 mRNA, and down-regulate the expression MMP-13 mRNA,and the ARC-H group was more significantly. There was significantly different（P〈0.05）expected expression of Col Ⅱ and MMP-13 mRNA in ARC-L group. 4）Compared with ARC-0 group,the content of collagen Ⅱ in both ARC-L group and ARC-H group were upregulated,and ARC-H group was more significantly. Expect for ARC-0 group and ARC- L group,the difference among other group was significa

关 键 词：膝骨性关节炎 软骨细胞 牛蒡子苷元 Ⅱ型胶原 基质金属蛋白酶-13 

分 类 号：R-33[医药卫生]