摘要目的:探讨骨桥蛋白(OPN)对人Tenon's囊成纤维细胞(HTFs)的调控作用.方法:实验研究.采用不同浓度(0、0.05、0.5、5μtmol/L)的OPN对HTFs进行处理,采用MTT法检测HTFs的增殖速度,采用细胞划痕实验检测HTFs的移行速度.实时荧光定量PCR(Real-time PCR)检测不同浓度的OPN处理后HTFs的血管内皮生长因子(VEGF)和转化生长因子β(TGF-β)的mRNA水平变化.采用Western blot检测不同浓度的OPN处理后HTFs的Ⅰ型胶原蛋白的表达.采用单因素方差分析对数据进行比较.结果:不同浓度OPN处理组HTFs增殖能力和移行速度差异有统计学意义(F=174.5、297.1,P＜ 0.01).OPN处理组的HTFs增殖能力增强,移行速度加快,而且随着OPN的浓度增加而增加.给予OPN单克隆抗体1A12(5 μtmol/L OPN+20 μtg/ml 1A12)处理后HTFs增殖和移行速度变慢.不同浓度的OPN处理后HTFs中VEGF和TGF-β的mRNA含量变化差异无统计学意义.OPN处理后HTFs中Ⅰ型胶原蛋白的蛋白表达增加,呈浓度依赖性(F=30.2,P＜0.01).结论:OPN能促进HTFs增殖和移行,呈浓度依赖性,该效应可被OPN抗体1A12抑制.上调OPN可以诱导HTFs中Ⅰ型胶原蛋白的表达,其单克隆抗体1A12能抑制这种效应.

abstractsObjective:To assess the modulating function of osteopontin (OPN) on human Tenon's fibroblasts (HTFs).Methods:With the intervention of OPN in different concentrations (0,0.05 μtmol/L,0.5 μmol/L,and 5 μmol/L),the proliferation rate of HTFs was measured by 3-(4,5-dimethylthiazol-2-Y1)-2,5-diphenyltetrazolium bromide (MTT),and the motility was surveyed by the cell scratch method.Real-time polymerase chain reaction (PCR) was applied to measure the mRNA of vascular endothelial growth factor (VEGF) and transforming growth factor β (TGF-β) in HTFs in the presence of different concentration of OPN.Western blot was used to measure the collagen-I levels.Data were analyzed using ANOVA.Results:OPN induced the proliferation of HTFs as indicated by MTT and enhanced the motility of HTFs as shown by the cell scratch method.The effects were concentration dependent (F=174.5,297.1,P＜0.01).In the presence of anti-OPN 1 A12 (5 μmol/L OPN+20 μtg/ml 1A12),the effects of OPN on HTFs were inhibited.None of the concentrations of OPN affected mRNA levels of VEGF and TGF-β mRNA.OPN augmented HTF expression of collagen-I protein in a concentration-dependent manner (F=30.2,P＜0.01).Conclusions:The enhancement of HTF proliferation,motility,and collagen-I expression OPN was concentration dependent.Incubation with the monoclonal OPN antibody 1A12 inhibited the effects of OPN on the HTFs.