IL-13和乙醇促进人肺成纤维细胞胶原表达

发布时间：2018-06-21 18:58

本文选题：IL-13 + IL-13Rα1　；参考：《南昌大学》2011年硕士论文

【摘要】：目的：白细胞介素-13(Interleukine-13, IL-13)是调控肺纤维化的主要TH2细胞因子,IL-13与IL-13受体al(IL-13Rα1)、IL-4受体α(IL-4Rα)相结合产生促纤维化作用,IL-13受体α2(IL-13Rα2)以高亲和力结合IL-13并抑制IL-13的促纤维化作用。本实验检测乙醇对人肺成纤维细胞(HFL-1)增殖以及HFL-1细胞IL-13受体基因表达的影响,研究乙醇和1L-13对HFL-1细胞Ⅰ、Ⅲ胶原基因(COL1A1、COL3A1 mRNA)的表达以及胶原蛋白合成的影响,从分子水平和蛋白水平探讨乙醇对IL-13介导的肺纤维化的作用机制。方法： 1.台盼蓝染色检测HFL-1细胞活性。2.HE染色观察HFL-1细胞形态。3.细胞代谢测定(MTT比色法)检测乙醇和/或IL-13刺激HFL-1细胞后的增殖：细胞分组：①空白组②IL-13组(10ng/ml、20ng/ml和50 ng/m1)③乙醇组(25 mM、50 mM、100 mM、200 mM、400 mM、800 mM、1600 mM)④乙醇(25 mM、50 mM、100 mM、200 mM)和IL-13 (10ng/ml、20 ng/ml和50 ng/m1)混合组4.采用实时荧光定量RT-PCR (Real time RT-PCR)分析IL-13 (10 ng/ml、20 ng/ml和50 ng/m1)和/或乙醇(25 mM.50 mM、100 mM、200 mM)对HFL-1细胞刺激前后各组IL-4Ra mRNA, IL-13Rα1 mRNA和IL-13Ra2 mRNA以及COL1A1、COL3A1 mRNA表达的影响。5.ELISA方法检测IL-13(10 ng/ml、20 ng/ml和50 ng/m1)和/或乙醇(25 mM、50 mM、100 mM、200 mM)对HFL-1细胞刺激前后各组细胞培养液中Ⅰ型胶原蛋白的表达的影响。结果： 1.HFL-1细胞活性良好,低浓度乙醇(200 mM)对HFL-1细胞无毒性作用。2.HE染色观察HFL-1细胞为长梭形,胞质被染成粉红色,胞核为椭圆形被染成蓝色。3.低浓度乙醇(200 mM)不会影响HFL-1细胞的增殖,但与IL-13(10 ng/ml、20 ng/ml和50 ng/m1)共同刺激下对HFL-1细胞的增殖有显著的促进作用,IL-13(10 ng/ml、20 ng/m1和50 ng/m1)对HFL-1细胞有促增殖作用,且有浓度依赖性。4. Real time RT-PCR检测结果显示,低浓度乙醇(50 mM、100 mM、200 mM)刺激HFL-1细胞后IL-13Ral mRNA, IL-4Ra mRNA表达都显著增强,同时IL-13Rαt2 mRNA表达显著减弱。低浓度乙醇(25 mM、50 mM、100 mM、200 mM)刺激HFL-1细胞前后COL1A1 mRNA、COL3A1 mRNA并无表达差异,IL-13(10ng/ml、20 ng/ml、50 ng/ml)刺激HFL-1细胞COL1A1 mRNA、COL3A1 mRNA表达显著增强,IL-13(10 ng/ml、20 ng/ml、50 ng/ml)和200 mM乙醇共同刺激HFL-1细胞后比单独相应浓度IL-13刺激后COL1A1 mRNA、COL3A1 mRNA表达显著增强。5.ELISA检测结果显示：低浓度乙醇(25 mM、50 mM、100 mM和200 mM)刺激HFL-1细胞前后Ⅰ型胶原蛋白并无表达差异,IL-13(10 ng/m1、20 ng/m1、50ng/ml)和200 mM乙醇共同刺激HFL-1细胞后比单独相应浓度的IL-13刺激后Ⅰ型胶原蛋白表达显著增强。结论： 1.低浓度乙醇不影响HFL-1细胞的增殖,但与IL-13共同刺激下将对HFL-1细胞的增殖有显著的促进作用。2.低浓度乙醇刺激HFL-1细胞对IL-13Rαl mRNA、IL-4RαmRNA的表达起着上调作用,下调IL-13Rα2 mRNA的表达。3.低浓度乙醇刺激HFL-1细胞后COL1A1 mRNA、COL3A1 mRNA的表达无影响。4.IL-13能促进HFL-1细胞的COL1A1 mRNA、COL3A1 mRNA的表达且有一定的浓度依赖性,IL-13和乙醇共同刺激HFL-1细胞可以显著上调COL1A1 mRNA、COL3A1mRNA和Ⅰ型胶原蛋白的表达。
[Abstract]:Purpose :

The effects of ethanol and 1L - 13 on the proliferation of human lung fibroblasts ( HFL - 1 ) and the expression of IL - 13 receptor genes in HFL - 1 cells were investigated . The effects of ethanol and 1L - 13 on the expression of IL - 13 receptor genes in human lung fibroblasts ( HFL - 1 ) and the effects of ethanol and 1L - 13 on the expression of IL - 13 receptor genes in human lung fibroblasts ( HFL - 1 ) and the effects of ethanol on the expression of collagen gene ( COL1A1 , COL3A1 mRNA ) and collagen synthesis were investigated .

Method :

1 . The expression of IL - 13 ( 10 ng / ml , 20 ng / ml and 50 ng / ml ) and / or ethanol ( 25 mM , 50 mM , 100 mM , 200 mM , 400 mM , 800 mM , 100 mM , 200 mM ) and IL - 13 ( 10 ng / ml , 20 ng / ml , and 50 ng / ml ) were detected by trypan blue staining . The effects of IL - 13 ( 10 ng / ml , 20 ng / ml and 50 ng / ml ) and / or ethanol ( 25 mM . 50 mM , 100 mM , 200 mM ) on the expression of IL - 4Ra mRNA , IL - 13伪 1 mRNA and IL - 13Ra2 mRNA and COL1A1 , COL3A1 mRNA were analyzed by real time RT - PCR . Effects of 20 ng / ml and 50 ng / ml ) and / or ethanol ( 25 mM , 50 mM , 100 mM , 200 mM ) on the expression of type I collagen in each group of cells before and after stimulation of HFL - 1 cells .

Results :

1 . HFL - 1 cells were well - activated and low - concentration ethanol ( 200 mM ) had no toxic effect on HFL - 1 cells . IL - 13 ( 10 ng / ml , 20 ng / ml , 50 ng / ml ) stimulated the expression of COL1A1 mRNA and COL3A1 mRNA in HFL - 1 cells . The levels of IL - 13 ( 10 ng / ml , 20 ng / ml , 50 ng / ml ) and 200 mM ethanol stimulated HFL - 1 cells .

Conclusion :

2 . The expression of COL1A1 mRNA and COL3A1 mRNA in HFL - 1 cells stimulated by low - concentration ethanol could significantly increase the expression of COL1A1 mRNA , COL3A1 mRNA and the expression of COL1A1 mRNA , COL3A1 mRNA and type I collagen in HFL - 1 cells .
【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R363

【共引文献】