核心蛋白多糖对兔晶状体上皮细胞增生的抑制作用

背景 研究发现,白内障囊外摘出术后组织修复反应可诱导房水中活性转化生长因子β(TGF-β)的增加,残留的晶状体上皮细胞(LECs)移行、分化,细胞外基质沉积,引起上皮-间质转化,导致后囊膜混浊(PCO)的发生.寻求有效的抑制LECs增生的药物对于临床上防治PCO的发生具有重要意义. 目的 探讨核心蛋白多糖( decorin)对兔LECs增生的抑制作用及其剂量-效应与时间-效应的关系. 方法 兔LECs细胞株进行培养和传代,将处于指数生长期的细胞以8×106个/L密度接种于96孔板,将0.1、1.0、10.0 mg/Ldecorin分别加入培养基中培养24、48、72 h,加入体积分数0.1％ DMSO培养的细胞为阳性对照组,常规培养基培养的细胞为空白对照组.MTT比色法分别测定不同质量浓度的decorin作用于LECs不同时间后的细胞生长抑制率；采用流式细胞技术测定各组细胞的细胞周期,用ELISA法检测各组培养液上清中TGF-β的水平,逆转录聚合酶链反应(RT-PCR)法测定TGF-βmRNA在LECs中的表达,免疫细胞化学法观察α-平滑肌肌动蛋白(α-SMA)的表达.结果 ELISA检测结果表明,各组培养基上清液中TGF-β表达量的差异有统计学意义(F=39.24,P=0.03),不同质量浓度组TGF-β水平均明显低于空白对照组,差异有统计学意义(P＜0.01),1.0 mg/L、10.0 mg/L decorin组TGF-β水平均明显低于0.1 mg/L decorin组(P＜0.05).MTT比色法结果显示,≥1.0 mg/L decorin组LECs增生的抑制率明显高于空白对照组,各质量浓度decorin组药物作用48 h和72 h后LECs增生的抑制率明显高于24 h的值,药物作用72 h后LECs增生的抑制率明显高于48 h的值,差异均有统计学意义(P＜0.05),各质量浓度decorin组G0/G1期LECs所占比例均较空白对照组明显增加,差异均有统计学意义(P＜0.05).RT-PCR检测显示,TGF-β mRNA的表达随着decorin质量浓度的升高而降低.免疫组织化学染色表明,10.0 mg/L decorin组α-SMA在LECs中的表达明显弱于空白对照组. 结论 Decorin可以抑制LECs的增生,也可以诱导LECs凋亡,其效应呈明显的剂量和时间依赖性,有望成为最具潜力的防治PCO的药物之一.

The inhibitory effect of decorin on proliferation of rabbit epithelial cell

Background Researches found that the posterior capsular opacification (PCO) after lensextraction is associated with the elevation of the transforming growth factor-β(TGF-β).To seek the drug for inhibitingproliferation of lens epithelial cells (LECs) is crucial in the treatment and prevention of PCO. Objective Thisstudy was to investigate the preventing effects of decorin on the proliferation of LECs. Methods Rabbit LECs wascultured and passaged.The LECs in growth phase were incubated in 96 well plate at the density of 8×106/L.Decorinwith the concentrations 0.1,1.0,10.0 mg/L was added into the medium for 24,48 and 72 hours respectively.0.1％DMSO was used at the same way as positive control group,and the regular cultured cells worked as blank controlgroup.The inhibitory rates of different concentrations of decorin on the growth of LECs were detected by MTT at 24,48and 72 hours after addition of decorin.The percentage of LECs in different cell cycles in various groups was assayedusing flow cytometry.TGF-β level in medium suspension was detected using ELISA.The expression of TGF-β mRNA in LECs was checked by RT-PCR,and α-SMA expression in LECs was determined using immunochemistry. Results ELISA assay showed a statistical difference in the TGF-β levels of different groups (F=39.24,P=0.03 ).The TGF-β levels in 1.0,10.0 mg/L decorin groups were significantly decreased in comparison with blank control group (P＜0.01) and 0.1 mg/L decorin group (P＜0.05 ).The inhibitory rates of decorin in the concentrations of ≥ 1.0 mg/L on the growth of LECs were higher than the blank control group,and those in various concentrations of decorin groups were considerably lower in 24 hours compared with 48 and 72 hours ( P＜0.05 ) and so was the 48 hours compared with 72 hours (P＜0.05 ).The percentages of LECs in G0/G1 phase were ascent in 0.1,1.0 and 10.0 mg/L decorin groups in comparison with G2/M and S phase (P＜0.05).Immunochemistry revealed the weak expression of α-SMA in various decorin groups in comparison with control group. Conclusions Decorin can effectively inhibit LECs growth and induce LECs apoptosis in concentration- and time-dependent manner.It is suggested that decorin can be used in the prevention and treatment of after cataract.