摘要目的：观察热休克蛋白(HSP)47在增生性玻璃体视网膜病变(PVR)增生膜中的表达及转化生长因子(TGF)-β2对视网膜色素上皮(RPE)细胞 HSP47表达的影响。方法收集行玻璃体切割手术的PVR 患者增生膜病理标本,苏木精-伊红、Masson 及免疫组织化学染色,观察其组织病理特征及 HSP47的表达。加入浓度分别为0、1、5、10 ng/ml 的 TGF-β2刺激培养人 RPE(ARPE-19)细胞,并根据此浓度此分为4组；浓度为5 ng/ml TGF-β2刺激细胞0、12、24、48 h。荧光定量聚合酶链反应(RT-PCR)、蛋白免疫印迹法(Western blot)检测细胞内 HSP47、Ⅰ型胶原蛋白(Col-Ⅰ)的 mRNA 和蛋白表达情况。结果光学显微镜观察发现,PVR 增生膜中可见类圆形上皮样细胞及色素颗粒,大量胶原蛋白成分；HSP47在多数上皮样细胞胞浆及间质内呈阳性表达。RT-PCR 检测结果显示,1、5、10 ng/ml 组细胞内 HSP47、Col-ⅠmRNA表达较0 ng/ml 组分别上调了1.32、2.35、1.85倍和1.29、1.52、2.11倍,差异均有统计学意义(HSP47：F =27.21,P <0.05；Col-Ⅰ：F =23.45,P <0.05)；TGF-β2作用0、12、24、48 h 后,细胞内 HSP47、Col-ⅠmRNA 表达较0 h 分别上调了1.56、1.84、2.86倍和1.57、1.86、2.78倍,差异均有统计学意义(HSP47：F =31.56,P <0.05；Col-Ⅰ：F =54.43,P <0.001)。Western blot 检测结果显示,1、5、10 ng/ml 组细胞内HSP47、Col-Ⅰ蛋白表达较0 ng/ml 组分别上调了2.33、2.89、2.60倍和1.18、1.49、2.11倍,差异均有统计学意义(HSP47：F =39.78,P <0.05；Col-Ⅰ：F =29.10,P <0.05)。TGF-β2作用12、24、48 h 后,细胞内HSP47、Col-Ⅰ蛋白表达量较0 h 分别上调了2.08、2.37、2.80倍和1.38、1.59、2.16倍,差异均有统计学意义(HSP47：F =49.18,P <0.05；Col-Ⅰ：F =42.52,P <0.05)。结论 HSP47在 PVR 增生膜中呈阳性表达,TGF-β2可能通过促进 ARPE-19细胞 HSP47的表达,增加 Col-Ⅰ的合成与沉积。

abstractsObjective To observe the expression of hot shock protein 47 (HSP47 )in pre-retinal membrane of proliferative vitreoretinopathy (PVR)and the influence of transforming growth factor-β2 (TGF-β2 )on the expression of HSP47 in retinal pigment epithelial (RPE)cell.Methods Pre-retinal membranes were collected and observed by hematoxylin-eosin,Masson and immunohistochemical staining. Cultured ARPE-1 9 cells were treated with TGF-β2 at serial concentration (0,1,5,10 ng/ml)and time (0, 12,24,48 hours),respectively.And then the mRNA and protein expressions of HSP47 and Col-Ⅰ were measured by fluorescence quantitative reverse transcription polymerase chain reaction and Western blot at the same time.Results A lot of epithelial cells with pigmental particles were observed in pre-retinal membranes of PVR,much accumulated collagen protein was observed in the specimens,and HSP47 positive expression was bserved in cytoplasm and stroma of most of the epithelioid cells.Compared with 0 ng/ml group,the expressions of HSP47 mRNA in ARPE-19 were up-regulated by 1.32,2.35,1.85 fold, significant differences were observed in all groups (F =27.21,P <0.05);the expressions of protein were up-regulated by 2.33,2.89,2.60 fold,significant differences were observed in all groups (F =39.78,P < 0.05).The expressions of Col-ⅠmRNA were up-regulated by 1.29,1.52,2.11 fold,significant differences were observed in all groups (F =23.45,P <0.05);the expressions of protein were up-regulated by 1.18, 1.49,2.11 fold and significant differences were observed in all groups (F =29.10,P <0.05).Compared with 0 hour group,the expressions of HSP47 mRNA were up-regulated by 1.56,1.84,2.86 fold in ARPE-19 cells stimulated by 5 ng/ml TGF-β2 for 12,24 and 48 hours,and the differences were all significant (F =31.56,P <0.05);the expressions of protein were up-regulated by 2.08,2.37,2.80 fold, and the differences were all significant (F = 49.18,P < 0.05).The expressions of Col-Ⅰ mRNA were up-regulated by 1.57,1.86,2.78 fold and the differences were all significant (F = 54.43,P <0.05),the expressions of protein were up-regulated by 1.38,1.59,2.16 fold and the differences were all significant (F =42.52,P <0.05).Conclusion TGF-β2 may play a role in the pathologic process of PVR by promoting the expression of HSP47 and then increasing the synthesis and accumulation of Col-Ⅰ.