摘要
目的：观察缺血后处理(ischemic postconditioning，Post-con)对大鼠移植肝细胞凋亡及 Caspase-3蛋白表 达的影响，探讨在体内条件下，缺血后处理对大鼠移植肝脏凋亡的保护机制。方法：采用 SD 大鼠原位肝移植动物模 型，供肝冷保存时间100min，无肝期控制于21min 内。48只雄性健康 SD 大鼠随机分为2组。对照组受体大鼠供肝 切除前及移植后无特殊处理；后处理组供肝植入后完全再灌注前，给予多次短暂复灌、复停作为缺血后处理。各组受 体一半(n=6) 于再灌注后2h 留取血液，检测血清肝功能指标和 TNF-α水平，另一半(n=6) 于再灌注后6h 取肝脏， 采用电镜、缺口末端标记法检测肝脏凋亡细胞，利用光学显微镜进行细胞计数；免疫组织化学检测 Caspase-3基因蛋 白表达，利用图像分析系统测量平均灰度值进行定量分析。结果：后处理组血清 AST、ALT、LDH 和 TNF-α含量均明 显低于对照组(P＜0． 05) ；组织的病理改变也明显轻于对照组；对照组凋亡细胞数为(112． 25±11． 73) 个/视野，后处理组 为(70． 36±18． 52) 个/视野，两组之间有显著差异(P＜0． 01) ；后处理组 Caspase-3蛋白表达明显下降(P＜0． 01) 。结论：缺 血后处理能够减轻再灌注损伤对移植肝的损害，明显抑制移植肝脏的细胞凋亡。这一作用可能是通过降低炎性细胞 因子水平，从而减少 Caspase-3蛋白表达而实现。
Objective To observe the effect of ischemic postconditioning on apoptosis of hepatocytes and expression of caspase-3 gene protein of rat liver graft, and to elucidate the possible mechanisms. Methods Model of orthotopic liver transplantation was adopted by using male Sprague-Dawley rats. The time of cold preservation and anhepatic phrase were 100 min and within 21 min respectively. Forty-eight rats were randomly divided into two groups： ① control group： there was no additive intervention on the donor liver either before being harvested or after implantation;②ischemic postconditioning group： immediately before the onset of reperfusion after donor liver was transplanted, several brief reperfusion-ischemia were given repeatedly. Half of the recipients of each group were used to take blood samples after 2 hours of reperfusion, and rest of the recipients were used to take liver tissue samples after 6 hours of reperfusion. The apoptotic hepatocytes were confirmed transmission electron microscope and counted with in situ nick labeling （TUNEL） method and light microscopy. The expression of caspase-3 gene protein was demonstrated by immunohistochemical staining and quantitatively examined by image analysis system. Results Compared with the control group, the contents of ALT, AST, LDH and TNF-α in serum of ischemic postconditioning group were significantly decreased（P〈0.05, respectively）; histological findings showed less severe injury in the ischemic postconditioning group than that in the control group. The number of apoptotic cells were （112.25±11.73） field in the control group, and （70.36±18.52） field in the ischemic postconditioning group; the difference was significant （P〈0.01）. The of expression caspase-3 gene was decreased significantly in the ischemic postconditioning group as compared with the control group （P〈0.01）. Conclusions Ischemic postconditioning could relieve to a certain degree the ischemiceperfusion injury in hepatocytes of the implanted liver and significautly inhibit the degree of apoptosis of the transplanted hepatocytes. Ischemic postconditioning can decrease the contents of cytokine and inhibit the expression of caspase-3, and thus producing its protective effect.
出处
《外科理论与实践》
2005年第6期535-538,共4页
Journal of Surgery Concepts & Practice
关键词
缺血后处理
大鼠
肝移植
再灌注损伤
细胞凋亡
CASPASE-3
Ischemi postconditioning
Rat
Orthotopic liver transplantation
Ischemic reperfusion injury
Apoptosis
Caspase-3
