人源促红细胞生成素在Mller细胞中拮抗谷氨酸毒性作用的研究  

Erythropoietin protects Mller cells through maintaining its expression of glutamate-aspartate transporter

作　　者：雇利敏[1,2] 张敬法[2] 徐华[1,2] 吕立夏[2] 李维业 徐国彤[2] 

机构地区：[1]同济大学生命科学与技术学院,上海200092 [2]同济大学附属第十人民医院眼科,上海200072 [3]美国Drexel大学医学院眼科,费城19129

出　　处：《同济大学学报（医学版）》2013年第1期1-7,共7页Journal of Tongji University(Medical Science)

基　　金：国家自然科学基金(81000383)

摘　　要：目的观察高浓度谷氨酸刺激条件下Muller细胞的变化以及促红细胞生成素(erythropoietin,EPO)的保护作用是否与维持Muller细胞的谷氨酸转运功能相关。方法用MTT法检测不同浓度谷氨酸处理大鼠原代视网膜Muller细胞和大鼠视网膜Muller细胞系rMC-1所引起的细胞活力变化。选择能显著降低细胞活力的最低谷氨酸剂量建立细胞损伤模型,并研究不同浓度EPO干预后细胞活力的变化。TUNEL(terminal deoxynucleotidyltransferase-mediated dUTP nick-end labeling)法检测各组细胞凋亡情况,Western印迹法检测谷氨酸转运体GLSAT在蛋白水平的变化。结果无论是在原代大鼠Muller细胞还是rMC-1细胞系中,谷氨酸的细胞毒性作用均呈剂量依赖趋势。原代Muller细胞中,12 mmol/L谷氨酸使细胞活力降低15.5%,10 mmol/L谷氨酸使rMC-1细胞活力降低15.6%;此时细胞凋亡明显增加,GLAST表达降低。0.2 U/ml和0.5 U/ml EPO分别对原代Muller细胞和rMC-1细胞的保护作用最佳,细胞凋亡数量显著减少,并防止了GLAST蛋白水平的降低。结论体外高浓度谷氨酸可损伤Muller细胞,EPO可以通过维持谷氨酸转运体GLAST水平等机制维持Muller对谷氨酸的正常摄取、抑制凋亡发生。Objective To determine the mechanism of the protective effect of erythropoietin （EPO） on Miiller cells, with special focus on its effects on expression of glutamate-aspartate transporter （GLAST） under glutamate stress. Methods Rat retinal Mtiller cell line （rMC-1） and primary Miiller cells were employed in this study. MTT assay was used to exam the effects of different dosage of glutamate on viability of rMC-1 and primary MUller cells, and to evaluate EPO＇s protection under glutamate stress. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling （TUNEL） method was used to detect cell apoptosis. GLAST level was studied with Western blotting method. Results Glutamate toxicity presented a dose-dependent manner on both primary Mtiller cells and rMC- 1 cells; 12 mmol/L glutamate caused 15.5% of the primary Mfiller cells to die and 10 mmol/L glutamate caused similar death rate in rMC-1 cells. TUNEL staining examination confirmed such cell death by demonstrating the increased apoptosis in both cell types and Western blotting showed a decreased GLAST expression in glutamate-treated cells. EPO （0. 2 U/ml ） exerted its optimal protection on primary MUller cell against apoptosis induced by 12 mmol/L glutamate and also maintained GLAST at normal level. Similarly, rMC-1 cells were protected by EPO but had a better response to 0.5 U/ml EPO. Conclusion The present study demonstrates that EPO might protect Mtiller cells by maintaining their normal GLAST expression so that the glutamate was uptake into the MUller cells.

关 键 词：促红细胞生成素 谷氨酸 M(u|¨)ller细胞 细胞凋亡 谷氨酸转运体 

分 类 号：R774.1[医药卫生—眼科]