山东大学耳鼻喉眼学报 ›› 2015, Vol. 29 ›› Issue (3): 65-71.doi: 10.6040/j.issn.1673-3770.0.2014.293

• 论著 • 上一篇    下一篇

促红细胞生成素抑制氧化损伤诱导的人眼Müller细胞凋亡

陈春丽1, 宋宗明2, 贾新国3, 周仲楼2, 汪朝阳3   

  1. 1. 胜利油田中心医院眼科, 山东 东营 257034;
    2. 温州医科大学附属眼视光医院, 浙江 温州 325027;
    3. 上海交通大学医学院附属新华医院, 上海 200092
  • 收稿日期:2014-09-01 修回日期:2015-04-04 发布日期:2015-06-16
  • 通讯作者: 汪朝阳。E-mail:zhaokekewzy@hotmail.com E-mail:zhaokekewzy@hotmail.com
  • 作者简介:陈春丽。E-mail:chenchunli125@163.com
  • 基金资助:
    国家自然科学基金(81100677);国家自然科学基金(面上项目)(81371040);上海市青年科技启明星计划(12QA1402200)

Erythropoietin inhibits Müller cell apoptosis induced by oxidative damage

CHEN Chunli1, SONG Zongming2, JIA Xinguo3, ZHOU Zhonglou2, WANG Zhaoyang3   

  1. 1. Shengli Oilfield Central Hospital, Dongying 257034, Shandong, China;
    2. The Eye Hospital of WMU, Wenzhou 325027, Zhejiang, China; 3. Xinhua College of Clinical Medicine, Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China
  • Received:2014-09-01 Revised:2015-04-04 Published:2015-06-16

摘要: 目的 探讨促红细胞生成素(EPO)抑制氧化损伤导致的Müller凋亡的可能性及其分子机制。方法 对培养的人眼Müller细胞系MIO-M1细胞采用BrdU标记法和MTT比色法观察正常及过氧化氢(H2O2)或葡萄糖氧化酶(GO)氧化损伤条件下0、0.01、0.1、1、10、30、100 U/mL EPO作用24、48、72 h后对Müller细胞增殖、迁移的影响;采用MTT比色法观察加PI3K/PDK1/PKB(Akt)信号传导通路阻断剂LY294002后Müller细胞增殖的变化;采用ELISA实验观察Müller细胞对EPO的表达和分泌;通过Western-blotting技术检测体外不同条件培养下EPO对ERK1/2及Akt信号传导通路的作用。结果 正常培养条件下, EPO有轻度促进Müller细胞增殖迁移的作用, 但差异无统计学意义;正常培养条件下, Müller细胞自身不分泌EPO, 0.4 mmol/L H2O2致Müller细胞损伤80%时, 其培养液内EPO的表达量为正常培养液下的1.42倍;氧化损伤状态下, 0.08 mmol/L H2O2或8 U/L GO作用Müller细胞24 h后导致其半数死亡且Akt信号通路激活;提前2 h加入外源性EPO后, 发现30 U/mL EPO对抗氧化所致Müller细胞的损伤作用最明显;同时提前2 h加入Akt信号通路阻断剂LY294002后, Akt信号传导通路的保护作用被阻断减弱。结论 EPO对体外正常培养的Müller细胞不具有促进增殖迁移作用;正常培养条件下Müller细胞自身不表达并且不分泌EPO, 氧化损伤条件下Müller细胞自身可低分泌EPO;加入外源性EPO后, EPO可能通过Akt信号传导通路对H2O2损伤的Müller细胞发挥保护作用。

关键词: ller细胞, Mü, 增殖, 年龄相关性黄斑变性, 促红细胞生成素

Abstract: Objective To demonstrate whether EPO can inhibit Müller cell apoptosis induced by oxidative damage and to investigate the molecular mechanism. Methods The proliferation effect of EPO on cultured human Müller cell-MIO-M1 was determined by BrdU marking and MTT assay both under normal condition and exposed to H2O2 or GO with different concentration (0 U/mL, 0.01 U/mL, 0.1 U/mL, 1 U/mL, 10 U/mL, 30 U/mL and 100 U/mL) at 24 h, 48 h and 72 h after exposure. The Müller cell proliferation changes after the use of PI3K/PDK1/PKB(Akt) signal pathway blocker-LY294002 was surveyed by MTT assay. The Müller cell function on EPO expression and secretion was observed by ELISA. The EPO effect on ERK1/2 and Akt signal pathway was detected under different cultured condition in vitro by Western-blotting. Results EPO had no effect on Müller cell proliferation under normal culture condition. The 80% damaged Müller cell caused by 0.4 mmol/L H2O2 can secrete 1.42 times EPO as that under normal culture condition. Under oxidative damage condition, 0.08 mmol/L H2O2 or 8 U/L GO could cause half of Müller cells to be dead and activate Akt signal pathway. While adding EPO 2 h earlier, 30 U/mL of EPO can reduce the oxidative damage to the minimum. The protection effect of Akt signal pathway could be reduced by its blocker LY294002 spontaneously. Conclusion EPO has no proliferation and migration effects on Müller cells cultured in vitro. Under normal culture condition, Müller cells did not express and secrete EPO. While under oxidative damage, Müller cells can secret EPO which can protect Müller cells by Akt signal pathway.

Key words: ller cells, Age-related macular degeneration, Mü, Erythropoietin, Proliferation

中图分类号: 

  • R774.1
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