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转化生长因子β1诱导皮肤成纤维细胞向肌成纤维细胞表型转分化的机制研究
熊杰,夏照帆*,吕开阳,王钰,韩姝
0
(第二军医大学长海医院烧伤科,全军烧伤研究所,上海 200433)
摘要:
目的:探讨TGFβ1诱导皮肤成纤维细胞(FB)向肌成纤维细胞转分化的可能途径和调控机制。方法:将野生型和Smad3基因敲除(KO)型小鼠皮肤FB分为9组:野生型FB组、野生型FB+TGFβ1组、野生型FB+SB431542组、野生型FB+SB431542+TGFβ1组、Smad3 KO FB组、Smad3 KO FB+TGFβ1组、野生型FB+SB203580+TGFβ1组、野生型FB+PD98059+TGFβ1组和野生型FB+SP600125+TGFβ1组。各组细胞经同步化处理后,直接以TGFβ1刺激或经上述各激酶抑制剂预处理后再以TGFβ1刺激。收集细胞,一部分以单细胞RTPCR检测αSMA阳性表达百分比,另一部分细胞抽提总RNA后采用实时荧光定量RTPCR检测αSMA的表达水平变化。结果:Smad3 KO组与SB431542组的αSMA表达水平和阳性百分比显著升高(Smad3 KO FB组vs野生型FB组;野生型FB+SB431542+TGFβ1组vs 野生型FB+SB431542组;Smad3 KO FB+TGFβ1组vs Smad3 KO FB组,P<0.01),而SB203580组和SP600125组中αSMA表达水平和阳性百分比升高的作用则被显著抑制(野生型FB+SB203580+TGFβ1组、野生型FB+SP600125+TGFβ1组vs 野生型FB+TGFβ1组,P<0.05)。结论:在TGFβ1诱导成纤维细胞向肌成纤维细胞的转分化过程中,Smad3途径介导抑制作用,而p38/MAPK、JNK/MAPK途径则介导正向调节作用。
关键词:  转化生长因子β  Smad3  成纤维细胞  肌成纤维细胞  转分化  小鼠,基因敲除
DOI:10.3724/SP.J.1008.2007.01175
投稿时间:2007-06-18修订日期:2007-09-29
基金项目:科技部“973”项目子课题(2005CB522603).
TGF-β1-induced transdifferentiation of derma fibroblasts into myofibroblasts:a study of mechanism
XIONG Jie,XIA Zhao-fan*,,LvKai-yang,WANG Yu,HAN Shu
(Department of Burns,Burn Institute of PLA,Changhai Hospital,Second Military Medical University,Shanghai 200433,China)
Abstract:
Objective:To explore the possible pathways and regulatory mechanism of TGFβ1induced transdifferentiation of derma fibroblasts(FB) into myofibroblasts. Methods: Mice Wildtype and Smad3 knockout(Smad3 KO) derma FB were divided into 9 groups,namely,A:Wildtype FB; B:Wildtype FB+TGFβ1; C:Wildtype FB+SB431542; D:Wildtype FB+SB431542+TGFβ1; E:Smad3 KO FB; F:Smad3 KO FB+TGFβ1; G:Wildtype FB+SB203580+TGFβ1; H:Wildtype FB+PD98059+TGFβ1; and I:Wildtype FB+SP600125+TGFβ1. After synchronization treatment,the cells were treated with TGFβ1 with or without pretreatment with above mentioned kinases inhibitors. Then the cells were collected for RNA extraction and the expression of αSMA was detected by real time quantitative RTPCR; some cells were analyzed by single cell RTPCR to test the positive expression rate of αSMA.Results: The expression and positive rate of αSMA in SB431542 group and Smad3 knockout group were significantly increased(group E vs. group A;group D vs. group C;group F vs. group E,P<0.01) and those in SP600125 group and SB203580 group were significantly inhibited(group G and I vs. group B,P<0.05). Conclusion: In TGFβ1induced derma fibroblasts transdifferentiation into myofibroblasts,Smad3 pathway plays a negative regulatory role and p38/MAPK and JNK/MAPK pathway play a positive regulatory role.
Key words:  transforming growth factor beta  Smad3  fibroblast  myofibroblast  transdifferentiation  mice,knockout