卡巴胆碱对人视网膜色素上皮细胞D407自分泌TGF-β2的调控
谭佳，刘双珍
作者单位：（410008）中国湖南省长沙市，中南大学湘雅医院眼科
作者简介：谭佳，在读博士，研究方向：屈光。
通讯作者：刘双珍. [email protected]
收稿日期：2007-10-30修回日期：2007-12-03
The regulation of carbachol on TGF-β2 autocrine of D407 human retinal pigment epithelial cells
Jia Tan, Shuang-Zhen Liu
Department of Ophthalmology, Xiangya Hospital of Central South University, Changsha 410008, Hunan Province, China
Correspondence to: Shuang-Zhen Liu. Department of Ophthalmology, Xiangya Hospital of Central South University, Changsha 410008, Hunan Province, China. [email protected]
Received:2007-10-30Accepted:2007-12-03
Abstract
AIM: To investigate the regulation of carbachol on the expression and secretion of transforming growth factor β2 (TGF-β2) of D407 human retinal pigment epithelial cells.
METHODS: D407 cells were cultured conventionally and divided into two groups. One group was incubated with carbachol (10-8-10-4mol/L) as experimental group, the other was untreated as blank group. The level of TGF-β2 secretion induced by carbachol was measured by enzyme-linked immunosorbent assay (ELISA) at 2, 4, 8, 16, 24 and 48 hours after D407 cells were treated, and the time- and dose-effect relations were explored. The level of TGF-β2 protein expression was detected by immunocyto- chemistry after D407 cells were incubated with carbachol of different concentrations for 24 hours, and the semi-quantitative analysis was carried out by using Image-Pro Plus 6.0. The data were analyzed by means of analysis of variance (ANOVA).
RESULTS: In blank group, the level of TGF-β2 in the supernate was increased after 4 hours of incubation, and reached 586.9±28.5ng/L at 24 hours. There was a significant difference in the level of TGF-β2 at various time points (F=86.59, P<0.01). In experimental group, the level of TGF-β2 in the supernate was higher than that in the blank group, and this effect was more significant when the incubation time was longer or the concentration of carbachol was higher. There were significant differences among the groups of different concentrations (10-8-10-4mol/L) (F=65.21,183.52,237.07, 250.08, 344.40; P<0.01) and among the groups at different time points (at 2,4,8,16,24,48 hours) (F=191.19,293.32,44426,47321,315.83,329.08; P<0.01). The expression of TGF-β2 protein of D407 cells in experimental group was higher than that in blank group (F=26.11; P<0.01).
CONCLUSION: Carbachol might be involved in the regulation of the cytokine network in the retina-retinal pigment epithelium-choroid complex by promoting the autocrine function of human retinal pigment epithelial cells.
KEYWORDS: carbachol; retinal pigment epithelium; transforming growth factor β2; cytokine
Tan J, Liu SZ. The regulation of carbachol on TGF-β2 autocrine of D407 human retinal pigment epithelial cells. Int J Ophthalmol
(Guoji Yanke Zazhi)2008;8(1):56-59
摘要
目的：研究卡巴胆碱对人视网膜色素上皮细胞株D407表达及分泌转化生长因子β2（transforming growth factor β2 , TGF-β2）的影响。
方法：常规培养D407细胞，分为实验组，加入卡巴胆碱10-8～10-4mol/L；空白组：不加药物。处理后2，4，8，16，24，48h采用ELISA法检测上清液中TGF-β2的含量，分析时效与量效关系；采用细胞免疫化学法检测卡巴胆碱孵育24h后D407细胞TGF-β2蛋白质的表达，运用图像分析软件Image-Pro Plus 6.0行半定量分析。统计学方法采用单因素方差分析。
结果：空白组细胞上清液中TGF-β2含量于培养4h后开始上升，24h达586.9±28.5ng/L，各时间点比较有统计学意义（F=86.59,P<0.01)。实验组细胞上清液中TGF-β2含量较空白组增加，随孵育时间的延长该效应逐渐明显，各浓度(10-8～10-4mol/L)组内比较均有统计学意义（F=65.21, 183.52,237.07,250.08,344.40; P<0.01)；各干预时间点（2，4，8，16，24，48h）实验组细胞上清液中TGF-β2含量随卡巴胆碱浓度升高逐渐增加，组内比较均有统计学意义（F=191.19,293.32,444.26,473.21,315.83,329.08; P<0.01)。孵育24h后实验组D407细胞TGF-β2蛋白质的表达较空白组增高（F=26.11，P<0.01）。
结论：卡巴胆碱可促进人视网膜色素上皮细胞分泌TGF-β2。
关键词：卡巴胆碱；视网膜色素上皮细胞；转化生长因子β2；细胞因子
谭佳,刘双珍.卡巴胆碱对人视网膜色素上皮细胞D407自分泌TGF-β2的调控.国际眼科杂志 2008;8(1):56-59
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