大鼠胸主动脉球囊损伤对细胞凋亡及凋亡相关基因Fas、Bcl-2表达的影响

Effect of balloon withdrawal injury on aortic cell apoptosis and Fas/Bcl-2 gene expression in rats

目的探讨大鼠胸主动脉球囊损伤后细胞凋亡和凋亡相关基因表达的变化规律。方法将30只400～500 g的雄性SD大鼠随机分为2组,手术组（n=24）行球囊扩张损伤大鼠胸主动脉术;对照组（n=6）不行球囊损伤,作为正常对照。分别于术后2、7、14、28 d取胸主动脉应用HE染色、TUNEL法、免疫组化和计算机图像分析仪进行形态学、细胞凋亡、增殖细胞核抗原（PCNA）、凋亡基因Fas;抗凋亡基因Bcl-2表达水平检测。结果对照组管壁处于非增殖状态;手术组球囊损伤后7 d形成新生内膜,血管平滑肌细胞（VSMC）增殖活跃;14 d内膜明显增厚,但VSMC增殖已减弱;28 d内膜继续缓慢增厚,管腔明显狭窄。动脉损伤后Fas表达和TUNEL法测定的凋亡规律一致,两周内凋亡较明显,但细胞凋亡高峰时间（中膜7 d、内膜14 d）迟于增殖高峰（中膜2 d、内膜7 d）,两周后凋亡与增殖均明显下降。动脉损伤后抗凋亡基因Bcl-2表达下调,在中膜和内膜分别在7 d1、4 d达最低水平,后回升,与凋亡基因Fas表达呈明显负相关（r=-0.878,P〈0.001）。结论动脉球囊损伤后,平滑肌细胞的凋亡呈现规律性变化,可能在管腔狭窄的病理过程中具有重要作用。

Objective To investigate the effect of balloon withdrawal injury on aortic cell apoptosis and Fas/Bcl-2 gene expression in rats.Methods Thirty SD male rats were randomly divided into 2 groups： Group 1（n=24） received balloon induced endothelium denudation in the thoracic aorta and were respectively sacrificed at day 2,7,14,28 after artery injury;Group 2（control group,n=6） received no injury procedure and were sacrificed 28 days after procedure.After the thoracic aorta were harvested,immunohistochemistry staining was performed to measure the protein expression of proliferation cell nuclear antigen（PCNA） and Fas/Bcl-2 gene.Vessel morphological measurements were also taken.Results In normal rat arteries,PCNA was not detected.After balloon injury,the proliferation of vascular smooth muscle cells（VSMCs） in media reached its peak at day 2 and then decreased quickly,while in neointima,VSMCs proliferated extremely at day 7 and then decreased.At day 28 after balloon injury,remarkable neointima and luminal narrowing were found.After balloon injury,the expression of Fas gene was accorded with the apoptosis rule measured by TUNEL method.The peak time of apoptosis（7d in media,14 d in neointima） was posterior to the peak time of proliferation（2 d in media,7 d in neointima）.After 14 days the apoptosis and proliferation decreased quickly.After balloon injury,Bcl-2 expression in media reached the lowest level at day 7 and then up-regulated;while Bcl-2 expression in neointima reached the lowest level at day 14.The expressions of Fas and Bcl-2 were in a remarkable negative correlation（r=-0.878,P〈0.001）.Conclusions After artery balloon withdrawal injury,there was regular change of apoptosis,which maybe play an important role in the process of luminal narrowing.