胰岛素样生长因子Ⅰ对豚鼠巩膜成纤维细胞增殖的影响  

Effects of insulin-like growth factor-Ⅰ on the proliferation of scleral fibroblasts in guinea pigs

作　　者：朱子诚[1] 张金嵩[1] 李秀娟[1] 籍雪颖[1] 王艳婷[1] 李元元[1] 

机构地区：[1]郑州大学第一附属医院眼科,河南省郑州市450052

出　　处：《中国组织工程研究与临床康复》2008年第11期2167-2169,共3页Journal of Clinical Rehabilitative Tissue Engineering Research

摘　　要：目的：研究表明，体内一定浓度的生长因子是保证眼球正常发育的必要条件，过少或过量都会引起巩膜的异常改变。实验拟进一步验证胰岛素样生长因子Ⅰ对体外培养的豚鼠巩膜成纤维细胞生长的影响。方法：实验于2006-09／2007—03在郑州大学基础医学院省肿瘤病理学重点实验室完成。①实验材料：出生3d内3色豚鼠后部巩膜组织由郑州大学基础医学院动物实验中心提供；实验用胰岛素样生长因子Ⅰ为Sigma公司产品。②实验过程及评估：采用器官组织块培养法获取原代巩膜成纤维细胞，选取传3，4代的豚鼠巩膜成纤维细胞用于实验；采用倒置相差显微镜下观察细胞形态并采用免疫化学染色法进行细胞鉴定；应用四甲基偶氮唑盐比色法和流式细胞仪分析技术，观察不同质量浓度胰岛素样生长因子Ⅰ（0．1，0．5，5，10，50μg／L）对体外培养的豚鼠巩膜成纤维细胞增殖和细胞生长周期的影响。结果：①巩膜成纤维细胞鉴定：倒置相差显微镜下观察，细胞透亮呈梭形，在细胞密度较低时细胞排列成网状，密度较高时呈束状；波形蛋白染色阳性。胞浆内可见棕黄色阳性反应产物，证实所培养的细胞为巩膜成纤维细胞。②体外培养的豚鼠巩膜成纤维细胞增殖和细胞生长周期：0．5，5，10μg／L的胰岛素样生长因子Ⅰ能促进豚鼠巩膜成纤维细胞增殖，呈剂量依赖性（F=84．510，P=0．000〈0．05），最佳质量浓度为10μg／L。流式细胞仪结果显示，胰岛素样生长因子Ⅰ处理48h后，与对照组比较，巩膜成纤维细胞GO—G1期百分比降低（78．2％下到64．5％，P〈0．05），而S期百分比显著升高（10．4％上升到16．4％，P〈0．05），增殖指数百分比增高（27.8％增高到56．5％，P〈0．05）。结论：在一定浓度范围内外源性胰岛素样生长因子Ⅰ可以促进体外培养的豚鼠巩�AIM： Growth factors at certain concentration in body are essential for the normal development of eyeball; either less or more can result in scleral abnormity. This study investigated the effects of insulin-like growth factor-Ⅰ（IGF-Ⅰ） on the proliferation of scleral fibroblast of guinea pigs. METHODS： The experiment was performed at tumor pathologic laboratory, the Basic Medicine School of Zhengzhou University from September 2006 to March 2007. ①Primary culture and subculture of scleral fibroblast （three-color guinea pigs, animal experimental center of Zhengzhou University） were established in vitro firstly. The third and fourth generation scleral fibroblast were collected. Cell morphous was observed using inverted phase contrast microscope and identified with immunochemistry. The effects of IGF-Ⅰ （Sigma） by various concentrations of 0.1, 0.5, 5, 10, 50 μg/L （final concentration） on cells proliferation and cell cycle were determined by methylthiazolyl tetrazolium （MTT） assay and flow cytometry. RESULTS： ①The cells showed fusiform shape, reticulate at low density and fasciculate at high density under inverted phase contrast microscope. Mouse anti-Vimentin was positive. Buffy positive reaction product was observed in cytoplasm, which confirmed the cultured cells were scleral fibroblast cells. ②IGF- Ⅰ （0.5, 5, and 10 μg/L） could significantly promote the proliferation of scleral fibroblast in a dose-dependent manner （F =84.510, P =4）.000 〈 0.05）, especially at the concentration of 10 μg/L, The results of flow cytometry analysis showed that compared with control group, the percentage of scleral fibroblast in S phase （from 10.4% to 16.4%, P 〈 0.05） and proliferative index （from 27.8% to 56.5 %, P 〈 0.05） was significantly increased and that in G0-G1 was decreased from 78.2% to 64.5% （P 〈 0.05） when the cells were stimulated with IGF-Ⅰ for 48 hours. CONCLUSION： IGF-Ⅰ within certain concentration can stimulate the proliferation of sclera

关 键 词：胰岛素样生长因子Ⅰ 巩膜 成纤维细胞 细胞增殖 细胞周期 

分 类 号：R392.114[医药卫生—免疫学]