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目的探讨联合转染血管内皮生长因子(VEGF)基因和增殖细胞核抗原反义寡核苷酸(PCNA-ASODN)对血管成形术后再狭窄的影响。方法 32只新西兰纯种大白兔随机平均分为对照组、转染VEGF组、转染PCNA-ASODN组、联合转染VEGF和PCNA-ASODN组(联合转染组)。构建兔的股髂动脉球囊损伤术后再狭窄模型,以医用生物蛋白胶作为缓释载体,将300μg PCNA-ASODN或VEGF均匀喷布于处理血管段外膜。取损伤处理血管段标本,光学显微镜观察损伤血管内膜、中层结构和局部血栓形成情况;电子显微镜观察新生内膜内皮细胞及内膜、中膜,计算内膜/中膜(I/M)面积比与厚度比;采用RT-PCR法检测VEGF与PCNA基因的表达,采用免疫组化从蛋白质水平检测VEGF与PCNA的表达。结果联合转染组与其他组相比,其血管内皮较完整、光滑,内膜轻度增生,平滑肌细胞排列规则,外膜无明显变化;I/M面积比与厚度比明显较低(P<0.05);RT-PCR检测VEGF表达水平明显升高(P<0.05);免疫组化检测血管组织中VEGF蛋白表达阳性率较高(P<0.05)。结论联合转染VEGF和PCNA-ASODN能更有效地抑制球囊损伤术后的血管内膜增生和再狭窄的发生。
Objective To investigate the effect of combined transfection of vascular endothelial growth factor (VEGF) gene and proliferating cell nuclear antigen antisense oligonucleotide (PCNA-ASODN) on restenosis after angioplasty. Methods Thirty-two New Zealand purebred rabbits were randomly divided into control group, VEGF group, PCNA-ASODN group and VEGF-PCNA-ASODN group. To construct rabbit model of restenosis after balloon injury of iliac arteries, 300μg of PCNA-ASODN or VEGF was uniformly sprayed on the outer membrane of vascular segment with medical bioprotein gel as a sustained-release carrier. The injured vascular segments were examined by light microscope, the intima-media, middle-layer structure and local thrombus formation were observed by optical microscope. The neointimal endothelial cells and intima-media were observed under electron microscope. The ratio of VEGF to PCNA was detected by RT-PCR. The expression of VEGF and PCNA was detected by immunohistochemistry at the protein level. Results Compared with other groups, the co-transfection group showed a more complete and smooth vascular endothelium, a mild hyperplasia of the endometrium and a smooth arrangement of smooth muscle cells with no significant changes in the outer membrane. The ratio of I / M area to thickness was significantly lower (P < 0.05). The VEGF expression level was significantly increased by RT-PCR (P <0.05). The positive rate of VEGF protein expression by immunohistochemistry in vascular tissue was higher (P <0.05). Conclusion Combined transfection of VEGF and PCNA-ASODN can effectively inhibit the intimal hyperplasia and restenosis after balloon injury.