外膜涂抹益气活血方防治静脉移植物再狭窄的实验研究

Study on the prevention of autologous vein graft restenosis by smearing Yiqihuoxue Prescription on the testing vessel adventitia

目的探讨术中局部应用益气活血方对自体移植静脉再狭窄的影响及其作用机制。方法建立家兔颈总动脉自体颈外静脉移植模型,随机分为:(1)A组,手术组;(2)B组,手术+抹胶(pluronic-F-127 gel)组;(3)C组,手术+抹益气活血方剂(由黄芪、川穹、三七组成)组,益气活血方由pluronic-F-127gel携带,涂抹于静脉移植物外膜。分别于术后7、14、28 d用免疫组织化学方法检测移植静脉增殖细胞核抗原(PCNA)的表达情况;HE、Masson染色后应用计算机图像分析系统检测术后14 d移植静脉内膜、中膜增生情况及内膜厚度/中膜厚度(I/M)值;应用流式细胞仪检测移植静脉DNA含量,观察血管平滑肌细胞(VSMC)的细胞周期,确定各期细胞所占的百分比。结果术后14 d,C组移植静脉内膜厚度、中膜厚度、I/M值明显低于A组和B组,差异有统计学意义(P<0.05);术后7、14、28 d,C组的VSMC增殖率低于A组和B组,差异有统计学意义(P<0.05);流式细胞仪分析,C组G0/G1、S、G2/M期细胞所占比例逐渐减少,A组和B组G0/G1、S、G2/M期细胞所占比例逐渐增多,与C组比较差异有统计学意义(P<0.05)。结论用pluronic-F-127 gel携带益气活血方涂抹自体移植静脉外膜可使移植静脉内膜、中膜增生减轻,对防治静脉移植物术后早期再狭窄有良好作用,其机制可能在于益气活血方能显著抑制VSMC的过度迁移增殖,减少细胞外基质的大量合成,从而减轻移植静脉内膜、中膜增生,对抗血管重构。

Objective To investigate the effect of smearing Yiqihuoxue Prescription locally in the management of vein graft failure and its mechanism. Methods A rabbit model of jugular veins grafted into the common carotid arteries was built. Animals were randomly divided into operation group （ group A）, operation ＋ smearing pluronic-F-127 gel group（ group B） and operation ＋ Yiqihuoxue Prescription group（ group C）. Yiqihuoxue Prescription mixed with pluronic-F-127 gel was smeared on the adventitia of autologous vein grafts in group C. Proliferating cell nuclear antigen（PCNA） were localized in 7,14,28 days by immunohistochemical procedure. Fourteen days after the operation, the thickness of neointima and media and neointima/media （I/M） were calculated from the sections stained with HE and Masson by computer image analysis. Flow cytometry was used to detect the DNA content and the ratio of cell cycles. Results The thickness of neointima, media and I/M in group C were significantly less than those in group A and B fourteen days after the operation （ P 〈 0.05 ）. The proliferating rate of vascular smooth muscle cell in group C decreased more significantly than those in group A and B in 7,14,28 days after operation. Analysis of flow cytometry showed that the ratio of cells in G0/G1, S, G2/M phase decreased gradually in group C and increased gradually in A and B group（P 〈 0.05）. Conehtsion Smearing Yiqihuoxue Prescription mixed with the pluronic-F-127 gel on the autologous vein grafts can inhibit the proliferation of the vessel proliferating cells, at least partly by inhibiting the adventitia activation, alleviating the hyperplasia of both neointima and media. It is an operative therapeutic method to prevent the autologous vein grafts restenosis by smearing Yiqihuoxue Prescription with pluronic-F-127 gel on the adventitia.