反义寡核苷酸对转化生长因子β2刺激的人眼球筋膜囊成纤维细胞结缔组织生长因子表达的影响  

Influence of ASODN to the human Tenon＇s fibroblasts in expressing CTGF induced by transforming growth factor β2

作　　者：胡义珍[1] 王育红[2] 曹阳[1] 张明昌[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院眼科,武汉430022 [2]武汉爱尔眼科医院

出　　处：《中华眼科杂志》2008年第2期157-162,共6页Chinese Journal of Ophthalmology

摘　　要：目的研究结缔组织生长因子（CTGF）在滤过泡瘢痕化形成中的作用。方法对照实验研究。应用二甲基四氮唑盐（MTT）法检测脂质体对人眼球筋膜囊成纤维细胞（HTF）活性的影响；用脂质体包裹CTGF的反义寡核苷酸（ASODN）处理经转化生长因子β2（TGF-β2）刺激的HTF，然后用逆转录聚合酶链反应（RT—PCR）技术和免疫细胞化学法检测纤维连接蛋白（Fn）的合成，用RT-PCR和免疫印迹法检测CTGF的表达。结果3个不同浓度组的脂质体作用HTF后，其吸光度（A）值分别为0．178±0．069、0．196±0．071、0．141±0．036，与正常对照组（0．202±0．073）比较，差异无统计学意义（F=1．65，P〉0．05）；单纯的TGF-β2刺激组（T组）与加入了脂质体的TGF-β2刺激组（D组）分别从基因和蛋白水平比较CTGF和Fn的表达，差异无统计学意义（t=0．90，2．32,0．75，2．11；P〉0．05）；转染12h后CTGF与Fn的mRNA水平比较，脂质体介导的CTGF反义寡核苷酸治疗组（A组）的相对灰度RI值明显高于脂质体介导的CTGF正义寡核苷酸对照组（S组）和D组（F=15．25,19．73；P〈0．05）；CTGF与FN的蛋白表达水平与mRNA表达水平相一致。结论CTGF的ASODN能够抑制TGF-β2对HTF表达CTGF和Fn的刺激作用。Objective To investigate the effect of connective tissue growth factor＇s antisense oligonucleotides （ASODN） on the growth of human tenon＇ s capsule fibroblasts （HTF） induced by transforming growth factor β2 （TGF-β2） in vitro. Methods It was a experimental study. HTF was collected from glaucoma patients and cultured. The 5-6 passage was used for experiments. The HTF induced by TGF- [32 was divided into the following groups： N group： normal HTF; T group： HTF induced by TGF-β2 ; A group ： CTGF ASODN antisense ：5＇-TACTGGCGGCGGTCAT-3＇ encapsulated with liposome ; S group ： sense 5＇-ATGACCGCCGCCAGTA-3＇ encapsulated with liposome; D group： HTF encapsulated with liposome only. The activity of HTF treated by different concentrations of liposome was detected using methylthianolyldiphenyl tetrazolum bromide （MTT） colorimetry. The expression of CTGF was detected by reverse transcription- polymerase chain reaction（RT-PCR） and immunocytochemistry assays. The expression of fibronectin （Fn） was examined by Western blot and immunocytochemistry assays. Results Liposome-ASODN （A group） significantly （F = 15.25, 204. 88, 19.73, 90. 00 ; P 〈 0.05 ） inhibit the expression of CTGF and Fn in HTF induced by TGF-β2 compared with S and D group. However, Liposome alone （ T group） has no significant impact in HTF growth compared with T group （ t = 0.90, 2. 32, 0.75, 2.11 ; P 〉 0.05 ）. Conclusion CTGF-ASODN inhibits the CTGF and Fn expression of HTF induced by TGF-β2, which may delay the formation of scar in glaucoma filtering surgery.

关 键 词：即早蛋白质类 胞间信号肽类和蛋白质类 寡核苷酸类 反义 转化生长因子β 纤连蛋白类 成纤维细胞 

分 类 号：R686[医药卫生—骨科学]