雌激素-胶原蛋白海绵对移植静脉再狭窄的影响及机制研究

Effect and Influence of Estrogen-collagen Sponge on Restenosis in Autologous Vein Graft Protein Expression

背景自体静脉行冠状动脉旁路移植术后,桥静脉再狭窄发生率高,影响远期疗效。目的观察雌激素-胶原蛋白海绵在预防移植静脉再狭窄中的作用并探讨其可能机制。方法 2016年1—5月,选取雄性新西兰家兔80只,采用随机数字表法将家兔分为A^D共4组,各20只。建立颈外静脉-颈总动脉旁路移植动物模型,游离并取颈外静脉1.5 cm,将其两端倒置,分别与颈总动脉行端侧吻合。A组不进行特殊处理;B组采用胶原蛋白海绵敷于吻合口周围;C组以17β-雌二醇均匀涂抹于吻合口周围;D组将胶原蛋白海绵浸泡于17β-雌二醇溶液,然后敷于吻合口周围。分别于术后6 h及7、14、28 d各取5只家兔,完整取下桥静脉。每个标本选3个不同切面,计算机图像处理系统测量内膜、中膜厚度,采用Western blotting法检测增殖细胞核抗原(PCNA)、细胞外调节蛋白激酶1/2(ERK1/2)、磷酸化细胞外调节蛋白激酶1/2(p ERK1/2)表达水平。结果 C组、D组术后14、28 d桥静脉内膜厚度小于A组、B组,且D组术后14、28 d桥静脉内膜厚度小于C组(P<0.05)。C组、D组术后14、28 d桥静脉中膜厚度小于A组,C组术后28 d及D组术后14、28 d桥静脉中膜厚度小于B组,D组术后28 d桥静脉中膜厚度小于C组(P<0.05)。C组和D组术后6 h桥静脉PCNA表达水平低于A组、B组,C组术后7 d桥静脉PCNA表达水平低于A组、B组,D组术后7、14、28 d桥静脉PCNA表达水平低于A组、B组和C组(P<0.05)。C组、D组术后6 h桥静脉p ERK1/2与ERK1/2比值低于A组、B组,C组术后7 d桥静脉p ERK1/2与ERK1/2比值低于A组、B组,D组术后7、14、28 d桥静脉p ERK1/2与ERK1/2比值均低于A组、B组和C组(P<0.05)。结论雌激素用于自体移植静脉外膜可降低PCNA的表达水平和ERK1/2的活化,抑制细胞增殖和移植静脉内膜、中膜增生;胶原蛋白海绵具有缓释作用,提高了雌激素在局部的药物浓度,延长了药物的作用时间。

Background After coronary artery bypass grafting in autologous veins,the high incidence rate of restenosis in bridging veins will influence forward curative effects.Objective To observe the effect of estrogen-collagen sponge on preventing vein from restenosis in graft and explore its possible underlying mechanism.Methods From January to May in2016,eighty male New Zealand rabbits were selected and randomly divided into A,B,C and D group by random number table method,20 rabbits each group.The external jugular vein-common carotid artery bypass transplantable animal model was established,freed and the 1.5 cm of the external jugular vein was taken,and then ends were inverted and was connected to common carotid artery by end-to-side anastomosis respectively.No special processing was made to A group; collagen sponge was applied to anastomotic stoma peripheries in B group,17β-estradiol was evenly applied to anastomotic stoma peripheries in C group; and in D group,collagen sponge was steeped in 17β-estradiol solution and applied it to anastomotic stoma peripheries.Five rabbits were got at time points of 6 hours,7 days,14 days and 28 days after operation respectively,and the bridging veins were completely removed.Three different sections were selected from each specimen sample.Computer image processing system was used to measure intima and media thickness,and Western blotting was adopted to detect the expression level of proliferating cell nuclear antigen（ PCNA）,ERK1 /2 and p ERK1 /2.Results Fourteen days and twenty-eight days after operation,the intimal thickness of bridging veins in group C and D were significantly less than that in group A and B,and the intimal thickness of bridging veins in group D was less than that in group C（ P〈0.05）.Fourteen days and twenty-eight days after operation,the medial thickness of bridging veins in group C and D were significantly less than that in group A,the medial thickness of bridging veins in group C 28 days after operation and group D 14 days and 28 days after operation were significantly less than that in group B,and the medial thickness of bridging veins in group D 28 days after operation were significantly less than that in group C（ P〈0.05）.The PCNA expression level of bridging veins in group C and D six hours after operation was significantly lower than that in group A and B,the PCNA expression level of bridging veins in group C seven days after operation was significantly lower than that in group A and B,the PCNA expression level of bridging veins in group D seven days,14 days and 28 days after operation was significantly lower than that in group A,B and C（ P〈0.05）.The ratio of p ERK1 /2 to ERK1 /2 of bridging veins in group C and D six hours after operation was lower than that in group A and B,the ratio of p ERK1 /2 to ERK1 /2 of bridging veins in group C seven days after operation was lower than that in group A and B,and the ratio of p ERK1 /2 to ERK1 /2 of bridging veins in group D seven days,14 days and 28 days after operation was lower than that in group A,B and C.Conclusion The estrogen in vein membrane of autologous graft can decrease the expression level of PCNA and the activation of ERK1 /2,and suppress the proliferation of cells and thickness of intima and media of vein grafts.The collagen sponge with sustained release effect increases the local drug concentration of estrogens and prolongs the acting time of drugs.