【摘要】 目的:研究烟焦油对体外培养的人视网膜色素上皮细胞(human retinal pigment epithelium，HRPE)超微形态结构的影响，以及趋化因子MCP1的细胞外表达水平。方法:将体外培养的HRPE分为空白对照组、二甲亚砜（dimethyl sulfoxide, DMSO）对照组和焦油实验组,其中焦油实验组中焦油浓度分别为1，10，50mg/L，经48h培养后，与空白对照组及DMSO对照组进行比较，使用倒置显微镜及透射型电子显微镜观察HRPE的形态以及超微结构变化，并用ELISA法选择检测各组培养上清中细胞因子MCP1的表达量。结果:倒置显微镜显示焦油实验组随着焦油浓度的增大，发生变性坏死的HRPE细胞数量逐渐增多，并且细胞的形态发生明显改变；电镜显示焦油实验组随着焦油浓度的增大，RPE细胞形态扁平，微绒毛减少，胞质稀疏、减少，细胞核细长、核染色质稀疏，染色质浓缩，可见空泡，细胞呈萎缩状态；ELISA结果显示随着焦油浓度的增大，RPE分泌至胞外的MCP1逐渐减少，差异有统计学意义（P＜0.05）。结论:烟焦油可改变视网膜色素上皮细胞正常的形态和生理功能，造成色素上皮细胞的严重损害。
【关键词】 吸烟；烟焦油；年龄相关性黄斑变性；视网膜色素上皮细胞；MCP1
Damage of cigarette tar on human retinal pigment epithelium in vitro
YuanYuan Hu, LingLing Yang, HuaQing Gong
Foundation item:Shandong Provincial Medical Scientific Academy Science and Technology Rroject（No.200637）
State Key Lab Cultivation Base, Shandong Provincial Key Lab of Ophthalmology, Shandong Eye Institute, Qingdao 266071, Shandong Province,China
Abstract
AIM: To investigate the effect of cigarette tar on the ultrastructure and the expression of monocyte chemotactic protein1 (MCP1) in human retinal pigment epithelium(HRPE) in vitro.
METHODS: The cultured HRPE were divided into blank group, DMSO group and cigarette tar experimental group. The concentrations of tar in cigarette tar experimental group were 1，10，50mg/L. After HRPE were treated with cigarette tar for 48h, the cellular morphology of HRPE was observed by inverted microscope and their ultrastructure was analyzed by transmission electron microscope, while ELISA kit is used to determine the levels of MCP1 in the supernatants in the three groups.
RESULTS: The quantity of HRPE, which degenerated or necrosed in cigarette tar experimental group, increased with the rising of tars concentration in inverted microscope. The transmission electron microscope showed that HRPE became flat in morphous with reduced microvilli, their cytoplasm were rarities and diminished while their cellular nucleus became slender, with rarities and psychotics chromatin. And the expression of MCP1 in the supernatants was reduced significantly with the rising of tar’s concentration(P<0.05).
CONCLUSION: Cigarette tar can change HRPE’s normal morphous and physiological function，and it can cause HRPE serious damage. Smoking may damage retinal by affecting retinal pigment epithelium.
KEYWORDS: smoking; cigarette tar; agerelated macular degeneration; retinal pigment epithelium; monocyte chemoattractant protein1
0引言
年龄相关性黄斑变性（agerelated macular degeneration，AMD）是目前西方发达国家中老年人群首要的致盲性眼病,而且随着人口老龄化的加剧,其发病率亦随之逐年上升[1,2]。视网膜色素上皮(retinal pigment epithelium,RPE)对维持视网膜的正常功能具有重要意义,其与许多视网膜病理改变相关,研究表明其在AMD的发病机制中具有重要的作用[3,4]。AMD早期临床表现包括RPE细胞的色素异常，RPE细胞基底部有玻璃膜疣（Drusen）形成，导致其上和邻近RPE细胞结构和功能失常，或引起脉络膜血管增生，最后引起相应部位的视细胞功能丧失。当病变发生在黄斑中心部时，患者中心视力严重受损，甚至失明[5]。流行病学研究表明，AMD是多基因和环境危险因素共同作用而导致的疾病[6,7]。吸烟可增加AMD的患病率,其危险度在1.20～3.29之间[8]。吸烟在AMD中发挥作用的阶段并不局限于加重脉络膜血管新生，而可能是在更早阶段，比如通过影响RPE细胞的功能发挥作用[9]。由于引起AMD的致病因子和具体的发病机制尚不明了，吸烟在AMD发生发展中的作用机制也不得而知。我们探讨烟焦油对体外培养的人视网膜色素上皮细胞（human retinal pigment epithelium，HRPE）的结构和功能的影响，揭示吸烟对视网膜的损害，为阐明吸烟对AMD的发生、发展提供理论依据。
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