肺成纤维细胞三维立体培养条件对肺纤维化组织重塑的影响(英文)  

Three-dimensional culture of human lung fibroblasts inpulmonary fibrosis remodeling process

作　　者：朱运奎[1] 肖永久[1] 薛庆亮[1] 王晓芹[1] 李继东[1] 汪莉[1] 刘卫[1] 牟玉兰[1] 

机构地区：[1]解放军兰州军区兰州总医院呼吸内科,甘肃省兰州市730050

出　　处：《中国组织工程研究与临床康复》2007年第23期4650-4653,共4页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：吴阶平医学基金会临床科研资助基金(2003-79-B)~~

摘　　要：背景:组织细胞三维立体培养是肺纤维化破坏与组织收缩研究的理想模型,特别是在纤维化的进展过程中,几种细胞因子如肿瘤坏死因子、白细胞介素、前列腺素、胰岛素以及渗出的血浆成分可能在纤维化的损伤修复和组织重塑中起重要的作用。目的:观察三维立体培养条件下肺成纤维细胞在血清和血清蛋白、胰岛素和细胞因子前列腺素E2、转移生长因子β、肿瘤坏死因子α的细胞外胶原基质收缩和细胞凋亡,探讨肺纤维化过程中细胞因子、胰岛素、血清及血清蛋白对肺组织重塑和纤维化形成的影响。设计:随机对照实验。单位:解放军兰州军区兰州总医院呼吸内科。材料:实验于2005-08/2006-01在解放军兰州军区兰州总医院呼吸内科实验室完成。人胚肺成纤维细胞(AmericanTypeCultureCollection),DMEM培养液和胎牛血清(GIBCO),胰岛素、转移生长因子(R&D),前列腺素E2(Sigma),Ⅰ型胶原提取自大鼠尾部肌腱。方法:为观察初始浓度对成纤维细胞收缩力的影响及细胞数量对胶原收缩的影响,提取大鼠尾部的胶原,与双蒸水、4×DMEM及成纤维细胞混合成胶原含量为0.75￣1.5g/L的立体组织胶原,细胞浓度为0.2×107￣4×107L-1,置入含体积分数0.05的CO2培养箱37℃培养。每天测定胶原的面积,最后计算终面积与初始面积的比值。将0.01%￣0.5%血清、0.1%的血清白蛋白和0.1%球蛋白分别加入到培养液中,观察胶原的收缩。加入10mg/L转移生长因子、10mg/L白细胞介素1、1mmol/L胰岛素和0.1μmol/L前列腺素E2,观察细胞因子对成纤维细胞介导胶原收缩的影响,对胶原中DNA含量进行测定及细胞存活率检测。主要观察指标:不同细胞因子或血清成分对肺成纤维细胞介导胶原收缩的影响,胶原内成纤维细胞数量对胶原收缩的影响,以及不同浓度胶原对胶原收缩和对细胞增殖和凋亡的影响。结果:①胶原的收缩有细胞依赖性BACKGROUND：Contraction of three-dimensional collagen gels has been used as a model for the contraction which characterizes both normal wound healing and the development of fibrosis in the tissue. Several factors and cytokines,such as tumor necrosis factor alpha （TNF-α）, interleukin-1, prostaglandin E2 and insulin have been proved to play important roles in collagen remodeling in vitro as well as serum extravasation during the fibrotic progress.OBJECTIVE： To observe extracellular collagen matrix contraction and apoptosis of fetal lung fibroblasts in TNF-α,interleukin-1, insulin, prostaglandin E2, albumin and globum under three-dimensional culture, and investigate the effects of cytokines, insuin, serum and serum protein on the remodeling and fibrotic formation of lung tissue.DESIGN： A randomized controlled experiment.SETTING： Department of Respiratory Medicine, Lanzhou General Hospital of Lanzhou Military Area Command of Chinese PLA.MATERIALS： The experiment was carried out The experiments were carried out in the respiratory laboratory of Lanzhou General Hospital of Lanzhou Military Area Command of Chinese PLA from August 2005 to January 2006. Human fetal lung fibroblasts （American Type Culture Collection）, Dulbecco＇s modified Eagle＇s medium （DMEM） and fetal bovine serum, insulin, transforming growth factor （TGF） （R＆D）, prostaglandin E2, type Ⅰ collagen was extracted from rat-tail tendons.METHODS： In order to investigate the effect of initial collagen concentration in the gels on the contractility of fibroblasts,the appropriate amount of collagen was mixed with distilled water, four fold-concentrated DMEM, and cells were suspended so that the mass concentration was 0.75-2.0 g/L, with a physiological ionic strength and the desired cell concentration. In order to investigate the effect of cell number in the gels on the contraction, the cellular concentration fibroblasts in the gels were prepared to 0.2×10^7-4×10^7 L^-1. The areas of floating gels were measured daily and th

关 键 词：肺成纤维细胞 三维立体 肺纤维化 

分 类 号：R329.4[医药卫生—人体解剖和组织胚胎学]