核因子κB的小干扰RNA防治自体移植静脉再狭窄的实验研究

目的   探讨核因子κB（NF-κB）的小干扰RNA（small interfering RNA，siRNA)对大鼠自体静脉移植术后血管内膜增殖及相应炎性细胞因子表达的影响。方法    雄性Wistar大鼠80只，随机分为空白组（A组，n=10）；自体静脉移植组（B组，n=18）；空载体组（阴性质粒脂质体医用蛋白胶复合物转染移植静脉，C组，n=26）和基因转染组（NF-κB siRNA阳离子脂质体医用蛋白胶复合物转染自体移植静脉，D组，n=26）。B、C及D组需制作大鼠自体颈外静脉——腹主动脉移植模型，每组4个时点( 3,7,14,21d)，相应时间点取移植静脉观察病理形态学变化，免疫组化检测增殖细胞核抗原（PCNA），PCR测定单核细胞趋化因子（MCP-1）和肿瘤坏死因子-α（TNF α）的mRNA含量，Western blot测定NF-κB p65蛋白的表达。结果    自体静脉移植术后，B、C和D组移植静脉血管桥均出现内膜增生变厚，新生内膜有大量PCNA阳性细胞，中膜平滑肌细胞增生活跃。术后3d，B组和C组 MCP-1 mRNA及TNF-αmRNA表达水平明显高于A组（P＜0.05），但D组水平明显低于C组（P＜0.05）。术后7d，B、C组NF-κB p65蛋白表达水平明显高于A组（P＜0.05），与C组相比，D组NF κB p65蛋白水平明显降低（P＜0.05）。结论     NF-κB的基因表达产物和MCP-1mRNA、TNF-αmRNA的表达有一定相关性，自体静脉移植术后新生内膜增生及炎性细胞因子表达在不同时相点呈动态变化。转染NF-κB siRNA阳离子脂质体复合物可抑制NF-κB的表达，减少MCP-1及TNF-αmRNA的表达，从而抑制移植静脉内膜增生和VSMC增殖，减轻再狭窄。

Research of nuclear factor-κB siRNA to prevent proliferation and restenosis of vein grafts

Objective    To investigate the effects of nuclear factor κB small interfering RNA (NF-κB siRNA) on intimal proliferation and expressions of inflammatory factors in autologous vein grafts. Methods    A total of 80 male Wistar rats(weight, 300g to 350g)were randomly divided into four groups: 1) Group A( n=10), normal group without transplantation; 2)Group B(n=18), vein transplantation only; 3) Group C (n=26),  vein transplantation but the graft was processed by blank plasmid liposome complexes; 4) Group D(n=26), vein transplantation but the graft was transfected by NF-κB siRNA liposome complexes. The transplantation was to replace an  abdominal aortic vein with the autologous right external jugular vein.  At four time points (3rd ,7th,14th,21st day),  autografts were processed to determine intima hyperplasia, PCNA, MCP-1 mRNA, TNF-α mRNA and NF-κB p65 protein. Results    Intimal hyperplasia and VSMC obviously proliferated in groups B,C and D. At the 3rd day, expressions of MCP-1mRNA and TNF-αmRNA in group D was significantly lower than group C (P＜0.05). At the 7th day, the levels of NF-κB p65 protein in group B and C were significantly higher than in group A (P＜0.05). Compared with group C, the levels of NF κB p65 protein in group D were lower(P＜0.05). Conclusion     Expressions of MCP-1 mRNA and TNF-α mRNA have some relationships with NF-κB p65 protein. The proliferation of vascular endothelial cells and expression of inflammatory cytokines are changing and correlative in the vein graft. Transfection of NF-ΚB siRNA liposome complexes on the autologous vein graft can inhibit its proliferation and reduce expressions of NF-κB, MCP-1 and TNF-α.