导入外源hTERT基因对大鼠供肝冷灌注损伤防护作用的研究

Alleviation of ischemia-reperfusion injury in rat liver donors by induction of exogenous hTERT gene

目的探讨外源hTERT基因转染对大鼠供肝冷缺血再灌注损伤的防护作用。方法将表达hTERT蛋白的重组腺病毒rAdhTERT转染试验组大鼠，同时设空载体对照组及空白对照组。组1（n=25）大鼠供体取肝前48h前阴茎背静脉目的基因干预实验组；组2（n=25）空载体病毒干预实验组；组3（n=25）注射等体积生理盐水；供肝切取置于uw液中修整保存6h后移植给同种大鼠。移植后3，6，12，24，48h分别取材进行相关检测，检测各组血清内ALT水平，端粒酶活性的测定，普通光镜和电镜下观察组织学变化，TUNEL法检测细胞凋亡。结果hTERT预处理组术后3，6，12，24，48h血清ALT较生理盐水组明显降低（P〈0．05），而空载体组和生理盐水组之间无差异；hTERT预处理可以显著提高供肝的端粒酶的活性，24h达到空载体对照组的（177．03±7．24）％（P〈0.05）；和hTERT组相比较，其他两组移植肝均表现出严重的小梁结构的破坏，门静脉周围的水肿以及空泡样变性。电镜检测也发现生理盐水组、空载体组的肝细胞内有大量的空泡，线粒体严重肿胀的现象；和生理盐水组及空载体组相比较，hTERT组的TUNEL阳性细胞明显减少，其凋亡指数和其他两组相比亦明显减少（P〈0.05）。结论腺病毒载体可成功介导hTERT基因对大鼠移植肝的转染；hTERT基因转染预处理能够减轻冷缺血再灌注损伤，hTERT的作用主要是通过上调端粒酶活性而抑制细胞凋亡来实现的。

Objective To determine if the ischemia reperfusion injury of liver can be alleviated by extrogen hTERT gene. Methods SD rats （16 months） were randomized into 3 groups： the donors in group A were pretreated with extrogen hTERT gene, those in group 13 were pretreated with none aim gene vector and the control ones in group C received physiological saline 48hr before the harvest. Liver were harvested, stored at University of Wisconsin solution （4℃） for 6h and then transplanted to syngeneic rats. Results The serum levels of AI.T and AST were significantly reduced in group A （P（0.05）. In 24 h after reperfusion, the telomerase activity was markedly higher in group A than in group B and C （P〈0.05） . The frequency of （TUNEL＋） liver cells was remarkably lower in group A and B than in group C. Conclusion Extrogen hTERT gene overexpression provides potent protection against cold ischemia-reperfusion injury. This effect depends on extrogen hTERT gene-mediated inhi bition of antiapoptotic mechanism.