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血管紧张素II诱导心肌成纤维细胞表达上调基因的分离和鉴定

王新凤, 高广道, 杨予白, 周娟, 王亚文, 苏兴利, 王琰, 韩锋产, 白玉杰

西安交通大学医学院病理生理教研室.陕西,西安 710061；陕西医学高等专科学校病理生理教研室.陕西,西安 710068；第四军医大学基因诊断研究所.陕西,西安 710032

摘要

为了对成年大鼠心肌成纤维细胞（cardiac fibroblasts, CF）受血管紧张素Ⅱ（AngiotensinⅡ, AngⅡ）刺激后上调基因表达谱进行筛选及分析，以受AngⅡ刺激CF为实验方，未刺激CF为驱动方，进行抑制消减杂交（suppression subtractive hybridization, SSH）建立消减cDNA文库。经斑点杂交筛选文库后将表达变化显著的部分阳性克隆测序及同源性分析，共获得19条上调表达的基因，分别与细胞外基质、细胞周期、胞内信号转导、细胞骨架及细胞代谢相关，并克隆到7条新的基因表达序列标签（expression sequence tags, EST）。作者的数据证实了SSH可以有效地克隆成年大鼠CF受AngⅡ刺激后上调表达基因，对这些基因的研究将有助于阐明心肌重塑的分子机制。

关键词： 重塑; 血管紧张素Ⅱ; 成纤维细胞; 抑制消减杂交

Identification of up--regulated genes induced by AngiotensinⅡin cardiac fibroblasts

Wang Xinfeng, Gao Guangdao, Yang Yubai, Zhou Juan, Wang Yawen, Su Xingli, Wang Yan, Han Fengchan, Bai Yujie

Department of Pathophysiology, Medical college of Xi'an Jiaotong University.Xi'an 710061,Shaanxi；China；Institute of Genetic Diagnosis, Forth Military Medical University.Xi'an 710032,Shaanxi

Abstract

To identify up-regulated genes in adult rat cardiac fibroblasts (CF) induced by AngiotensinⅡ (AngⅡ). Suppression subtractive hybridization (SSH) was performed between the CF stimulated by AngⅡ (tester) and unstimulated CF (driver) to generate subtractive cDNA library. The library was screened with dot blots hybridization to further verify the differentially expressed cDNA clones. Partial positive clones (19 up-regulated genes) were sequenced and BLAST analyzed. Twelve up-regulated genes related to extracellular matrix , cell cycle, intracellular signal transduction, cell cytoskeleton, cell metabolism and 7 new expression sequence tags (EST) were acquired （Genebank accession number：CN382808, CN382809, CN382810, CN382811, CN382812, CN382813, CN382814）. Our data reveal that SSH is a powerful technique of high sensitivity for the detection and clone of up-regulated gene expressed in CF induced by AngⅡ, which may be helpful to clarify the mechanism of cardiac remodeling.

Key words: remodeling;angiotensinⅡ;Fibroblasts;suppression subtractive hybridization

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引用本文：

王新凤, 高广道, 杨予白, 周娟, 王亚文, 苏兴利, 王琰, 韩锋产, 白玉杰. 血管紧张素II诱导心肌成纤维细胞表达上调基因的分离和鉴定[J]. 生理学报 2005; 57 (5): .

Wang Xinfeng, Gao Guangdao, Yang Yubai, Zhou Juan, Wang Yawen, Su Xingli, Wang Yan, Han Fengchan, Bai Yujie. Identification of up--regulated genes induced by AngiotensinⅡin cardiac fibroblasts . Acta Physiol Sin 2005; 57 (5): (in Chinese with English abstract).