C-myc siRNA抑制大鼠自体移植静脉再狭窄的研究

Small interfering RNA targeting c-myc inhibits vein graft restenosis in rats

目的静脉移植术后c-myc基因持续高表达是导致血管内膜增生进而再狭窄的原因之一。文中探讨通过局部给药方法给予针对c-myc基因的小分子干扰RNA(siRNA)对自体移植静脉内膜增生的影响。方法建立SD大鼠自体颈外静脉-颈总动脉移植模型。32只大鼠按移植静脉外膜局部给药不同处理方式方法分为未处理组(No treatment组)、凝胶组(Gel组)、无义序列组(scramble sequence group,SCR组)、c-myc RNA干扰组(C-myc siRNA组),于术后3周处死大鼠取移植静脉,光学显微镜下测量内膜厚度,免疫组化染色检测移植静脉增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)、c-myc蛋白的表达,Q-RT-PCR检测移植静脉c-myc mRNA的含量。结果 C-myc siRNA组内膜厚度[(15.0±2.2)μm]显著低于No treatment组[(58.3±8.3)μm]、SCR组[(60.5±6.1)μm]和Gel组[(63.5±5.4)μm]。C-myc siRNA组静脉内膜区和中膜区PCNA阳性细胞率显著低于SCR组[分别为(8.4±2.2)%vs(68.5±5.0)%,P<0.05和(15.0±3.1)%vs(55.2±7.5)%,P<0.05],c-myc蛋白阳性细胞率C-myc siRNA组显著低于SCR组[(30.6±2.6)%vs(48.5±3.5)%,P<0.05]。C-myc siRNA组移植静脉中c-myc mRNA的表达相对量显著低于SCR组(0.48±0.05 vs 1.00±0.12,P<0.05)。结论C-myc siRNA能抑制移植静脉中平滑肌细胞的增殖,抑制内膜的增生,减轻移植静脉再狭窄程度。

Objective Persistent high expression of the c-myc gene after vein transplantation may lead to intimal hyperplasia and restenosis of the grafted veins.The present study aims to evaluate the effect of locally administrated small interfering RNA（siRNA） targeting the c-myc gene on intimal hyperplasia after vein autotransplantation.Methods We performed autologous jugular vein to carotid artery reverse interposition on 32 male SD rats,which were equally divided into an untreated group（vein grafts untreated）,a gel group（vein grafts treated with gel only）,an SCR group（vein grafts treated with gel containing scrambled siRNA） and a c-myc siRNA group（vein grafts treated with gel containing c-myc siRNA）.The rats were sacrificed 3 weeks after surgery,and the grafted veins harvested for measurement of the intimal thickness under the microscope,determination of the expressions of the proliferating cell nuclear antigen（PCNA） and c-myc protein by immunohistochemistry,and detection of the content of c-myc mRNA by Q-RT-PCR.ResultsThe intimal thickness was（15.0±2.2） μm in the c-myc siRNA group,significantly less than in the untreated（[58.3±8.3]μm）,SCR（[63.5±5.4]μm） and gel group（[60.5±6.1]μm）.The percentages of PCNA positive cells in the intima and media were significantly lower in the c-myc siRNA group（[8.4±2.2]% and [15.0±3.1]%） than in the SCR group（[68.5±5.0]% and [55.2±7.5]%）（P〈0.05）,and so were the percentage of c-myc positive cells and the relative expression of c-myc mRNA in the c-myc siRNA than in the SCR group（[30.6±2.6]% vs [48.5±3.5]% and 0.48±0.05 vs 1.00±0.12,P〈0.05）.Conclusion Local administration of siRNA targeting c-myc can effectively inhibit the proliferation of vascular smooth muscle cells,suppress intimal hyperplasia,and thus ameliorate restenosis of the vein graft.