组织型纤溶酶原激活物基因疗法防治血管吻合口血栓栓塞

t-PA gene prevents thrombo embolism at vascular anastomotic sites

目的 探讨防止血管吻合口血栓栓塞的新方法 ,了解组织型纤溶酶原激活物基因疗法防止吻合口血栓栓塞的效果 .方法 1将含有 t- PA基因的重组质粒 p Ad CMVt- PA与缺陷型腺病毒大框架 p JM1 7共转染病毒包装细胞 2 93细胞 ,获得有感染能力但复制缺陷的重组腺病毒 Ad CMVt- PA.2实验动物随机分为对照组和治疗组 ,均用 11- 0尼龙缝线行大鼠颈总动脉原位端端吻合 .治疗组吻合口注射 Ad CMVt- PA溶液 ,对照组注射 Ad CMV(不含 t- PA的空载体 ) .提取吻合口组织的总 RNA行 RT- PCR,应用发色底物法检测吻合口组织的纤溶酶活力 .结果 治疗组 RT- PCR在 2 .0 kb前面有一条带 ,对照组则没有 .治疗组术后第 2 ,3,5 ,7和 14日均可检测到纤溶酶活力 ,但第 3日活力最高 ,对照组未检测到纤溶酶活力 .结论 克隆入腺病毒载体的 t- PA基因可在吻合口组织中表达出有生物学活性的 t- PA,可能有效地抑制吻合口血栓形成 。

AIM To develop a new way to prevent vascular anastomotic sites from thrombo embolism and to study the effect of gene therapy with tissue type plasminogen activator (t PA ) gene on the formation of thrombo embolism of vascular anastomotic sites. METHODS 1: Recombinant plasmid pAdCMVt PA was cotransfected into 293 cells with pJM 17 , and infectious but replication deficient AdCMVt PA was generated. 2:Rats were randomly divided into control and treatment groups. 11 0 nylone medical suture was applied to perform rat carotid artery end to end anastomoses. In the treatment group, AdCMVt PA solution was injected into vascular anastomotic site while AdCMV (no containing t PA DNA) solution was injected into the control group. RESULTS When the isolated RNA was performed with RT PCR, 1.69 kb band appeared in the treatment group, but band could not be found in the control group. The t PA activity could be detected postoperation on the 2nd, 3rd, 5th, 7th and 14th day in the treatment, but could not be detected in the control group. CONCLUSION t PA gene can produce t PA having biological activity at anastomotic sites, possibly preventing the formation of thrombo embolism effectively and developing the anastomotic patency.