阿托品对人视网膜色素上皮细胞D407株表达及分泌TGF-β_2的调控  

Expression and secretion of TGF-β_2 in human retinal pigment epithelium cell line D407 regulated by atropine

作　　者：谭佳[1] 邓志宏[1] 刘双珍[1] 

机构地区：[1]中南大学湘雅医院眼科,长沙410008

出　　处：《中南大学学报（医学版）》2010年第5期518-523,共6页Journal of Central South University ：Medical Science

基　　金：湖南省自然科学基金(08JJ3047)~~

摘　　要：目的:通过观察不同浓度阿托品联合10-5mol/L卡巴胆碱干预下D407细胞表达及分泌TGF-β2的变化,探讨阿托品对RPE细胞表达及分泌TGF-β2的调控作用。方法:常规培养D407细胞,药物干预前换用无血清培养基培养,分为4组。(1)实验组(A组):A1～A5组依次加入10-4～10-8mol/L阿托品,孵育30 min后每组加入10-5mol/L卡巴胆碱;(2)阴性对照组(B组):B1～B5组依次加入10-4～10-8mol/L阿托品;(3)阳性对照组(C组):加入10-5mol/L卡巴胆碱;(4)空白对照组(D组):不加药物。干预24 h后采用RT-PCR,Western印迹及ELISA法检测细胞胞浆中TGF-β2mRNA及蛋白质的表达水平及上清液中TGF-β2的含量。统计学方法采用单因素方差分析。结果:实验组D407细胞胞浆TGF-β2mRNA和蛋白质表达水平及上清中TGF-β2蛋白质含量均较阳性对照组低,10-4mol/L阿托品可完全阻断10-5mol/L卡巴胆碱上调TGF-β2表达及分泌的作用,其效应具有浓度依赖性(F=1 056.897,1 320.170,475.657;P<0.001)。阴性对照组D407细胞胞浆TGF-β2mRNA和蛋白质表达水平及上清中TGF-β2蛋白质含量与空白对照组比较差异无统计学意义(P>0.05)。结论:阿托品可有效抑制卡巴胆碱促进人RPE细胞表达及分泌TGF-β2的功能,提示M受体参与介导此过程。Objective To investigate the regulation of atropine to the expression and secretion of TGF-β2 in retinal pigment epithelium（RPE） cells by observing the changes of those under different treatments of atropine and carbachol.Methods D407 cells were cultured conventionally and divided into 4 groups as follows：（1） An experimental group（Group A）,cells were pretreated with 10^-4-10^-8 mol/L atropine for 30 min,and then treated with 10^-5 mol/L carbachol;（2） a negative control group（Group B）,cells were treated with 10^-4-10^-8 mol/L atropine;（3） a positive control group（Group C）,cells were treated with 10^-5 mol/L carbachol;（4） a blank control group（Group D）.The concentration of TGF-β2 in the supernate,and the level of TGF-β2 mRNA and protein were measured by ELISA,RT-PCR,and Western blot after the 24-hour treatment.The data were analyzed by analysis of variance.Results The levels of TGF-β2 mRNA and protein in the cytoplasm and the concentration of TGF-β2 in the supernate in the experimental groups were lower than those of the positive control group.Atropine at 10^-4 mol/L could completely inhibit the effect of carbachol at 10^-5 mol/L.The effect of atropine was concentration-dependent（F=1 056.897,1 320.170,and 475.657;P〈0.001）.There was no change of TGF-β2 level in the cytoplasm and supernate with the treatment of atropine alone（P〉0.05）.Conclusion Carbachol can promote the expression and secretion of TGF-β2 in human RPE cells and atropine could reverse it effectively,suggesting that M receptor may be involved.

关 键 词：视网膜色素上皮 转化生长因子Β2 卡巴胆碱 阿托品 

分 类 号：R774.1[医药卫生—眼科]