转化生长因子-β1对人软骨细胞基质金属蛋白酶-1mRNA 及其阻滞剂mRNA 表达的影响

Regulating effects of transforming growth factor-β1 on gene expressions of MMP-1 and TIMP-1 mRNA in human hyaline chondrocytes

目的观察转化生长因子-β1(TGF-β1)对人透明软骨细胞基质金属蛋白酶-1(MMP-1)及其阻滞剂T(IMP-1)mRNA表达的作用及相关意义,更好地理解骨关节炎软骨损伤的相关机制。方法TGF-β1作用于传代培养的人软骨细胞12h,浓度分别为1ng/mL、10ngm/L、100ng/mL;上述不同浓度TGF-β1与白介素-1β(IL-1β)10ng/mL组成联合作用组,继续培养12h。应用逆转录PCRR(TP-CR)方法及实时荧光定量方法(FQP-CR),检测TGF-β1及其与IL-1β联合作用于传代培养的人透明软骨细胞MMP-1、TIMP-1mRNA表达的含量。结果正常对照组透明软骨细胞可见MMP-1、TIMP-1mRNA扩增产物,而实验组TGF-β11ng/mL、10ng/mL、100ngm/L作用于透明软骨细胞12h后,MMP-1mRNA表达逐渐降低,TIMP-1mRNA表达逐渐增高;TGF-β1与IL-1β联合作用后,随着TGF-β1浓度的升高,MMP-1mRNA表达逐渐降低,TIMP-1mRNA表达逐渐增高,各组之间差异有显著性意义(P<0.05)。提示TGF-β1与MMP-1、TIMP-1mRNA表达之间存在剂量依赖关系。结论不同浓度TGF-β1按照剂量依赖方式抑制人软骨细胞MMP-1mRNA基因的表达,刺激人软骨细胞TIMP-1mRNA基因表达;TGF-β1具有对抗IL-1β对人软骨细胞MMP-1、TIMP-1mRNA表达的作用,也呈现剂量依赖关系。研究结果对揭示骨关节炎的发病机理,指导其治疗具有积极意义。

Objective To investigate the role of transforming growth factor - β1 （TGF-β1） in regulating gene expression of the matrix metalloproteinase-1 （MMP-1） and its inhibitor, the tissue inhibitor of metalloproteinase -1 （TIMP-1） in human hyaline chondrocytes so as to have a better understanding of the mechanism of cartilage injury in osteoarthritis. Methods In the experiment, human hyaline chondrocytes were harvested enzymatically and cultured in DMEM and given respectively TGF-β1 of 1 ng/mL, 10 ng/mL and 100 ng/mL, 1 ng/mL TGF-β1 ＋ 10 ng/mL IL-1β, 10 ng/mL TGF-β1 ＋ 10 ng/mL IL-1β, 100 ng/mL TGF-β1 ＋ 10 ng/mL IL-1β before they were cultured for another 12 hours. The MMP-1 and TIMP-1 mRNA levels of passaged monolayer cell cultures of human hyaline chondrocyte were assessed by reverse transcriptase polymerase chain reaction （RT-PCR） and fluorescent quantitative PCR （FQ-PCR）. Results Normal human chondrocytes expressed MMP-1 and TIMP-1 mRNA. In the groups treated with TGF-β1 and IL-1β, TGF-β1 could decrease the MMP-1 mRNA level, but increase TIMP-1 mRNA expression level. The difference was significant （ P 〈 0.05） and the levels of MMP-1 and TIMP-1 mRNA expression had significant association with the dosages of TGF-β1. Conclusion Our results show that TGF-β1 may have a crucial effect on chondrocytes by regulating the balance between the expressions of MMP-1 mRNA and TIMP-1 mRNA, which are known to be important in osteoarthritis development, and have an antagonistic effect on IL-1β in osteoarthritis.