摘要
目的 :研究重组腺病毒介导的人内皮型一氧化氮合酶基因 (eNOS)表达生成的一氧化氮合酶 (NOS)对球囊损伤后大鼠颈总动脉新生内膜的抑制作用。方法 :在 2 93细胞内扩增、纯化Ad LacZ和Ad eNOS ,鉴定其是否携带有LacZ和eNOS基因。建立大鼠颈总动脉球囊损伤模型后 ,将磷酸缓冲液 (PBS)、Ad LacZ和Ad eNOS在体内分别转染到损伤血管段 ,以X gal染色、苏木精 伊红染色 ,免疫组化及计算机图像分析处理等方法观察转染动脉节段外源性eNOS蛋白表达及其对新生内膜的影响。结果 :重组腺病毒携带有eNOS基因 ,并且在损伤血管段得到有效表达。转染后PBS组、Ad LacZ组和Ad eNOS组的新生内膜面积分别为 (0 .187± 0 .0 18)、(0 .134± 0 .0 6 1)和 (0 .0 6 3± 0 .0 2 6 )mm 2 ,新生内膜与中膜面积比值 (I/M)分别为 1.5 76± 0 .2 73、1.342± 0 .35 7和 0 .5 6 0± 0 .16 1。与PBS组、Ad LacZ组相比Ad eNOS组无论新内膜面积 ,还是管腔狭窄程度都明显减小。结论 :腺病毒介导的eNOS基因转染能有效抑制球囊损伤后血管内膜的增生 。
Objective:To study the effects of adenovirus mediated human endothelial nitric oxide synthase gene(eNOS) transfer on neointimal proliferation of rat carotid artery after balloon injury.Methods:The recombinant viruses were amplified on confluent 293 cells and purified by discountinous CsCl gradient centrifugation. The morphology of 293 cells, X gal staining on 293 cells and PCR of eNOS gene cDNA segment were employed. After balloon injury, rats' carotid arteries were immediately transduced with phosphate buffered saline(PBS) 0.2 ml, 1.02 ×10 12 pfu/ml Ad LacZ 0.2 ml and 1.24 ×10 12 pfu/ml Ad eNOS 0.2 ml in vivo respectively. X gal staining, HE staining, immunochemistry and computer image analgsis system were used to analyze the expression of exogenous eNOS protein and its effects on neointimal proliferation.Results:The transfected 293 cells were stained blue with X gal. The eNOS cDNA was confirmed in the Ad eNOS genome by PCR. After 2 weeks, eNOS protein were confirmed by immunochemistry in Ad eNOS infected segments. In PBS?Ad LacZ infected and eNOS infected segments, the neointimal area was ( 0.187 ± 0.018 ) mm 2, ( 0.134 ± 0.061 ) mm 2 and( 0.063 ± 0.026 ) mm 2 respectively, the ratio of neointima to media was 1.576 ± 0.273 , 1.342 ± 0.357 and 0.560 ± 0.161 respectively. The neointimal area and the I/M ratio of Ad eNOS infected were significantly reduced.Conclusion:Adenovirus mediated NOS gene transfer might reduce arterial restenosis after balloon angioplasty.
出处
《临床心血管病杂志》
CAS
CSCD
北大核心
2003年第7期406-409,共4页
Journal of Clinical Cardiology
基金
湖北省科技厅重点支持项目 (NO .993 D0 9)