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目的探讨自制肝保护液对梗阻性黄疸大鼠门静脉阻断所致肝细胞损伤的作用及对肝细胞MAPKs信号途径的影响。方法 72只Wistar系大鼠完全随机分为4组:A组:胆道再通、门静脉周围游离、切除肝尾状叶,90 min后关腹腔;B组:胆道再通、肝尾状叶分流门静脉、余肝门静脉血流阻断90 min、恢复正常门静脉血流、切除肝尾状叶,关腹;C组:B组+门静脉肝侧灌注4℃乳酸林格氏液;D组:B组+门静脉肝侧灌注4℃自制肝保护液(home-made liver solution,HMLS)。术后0、6、24 h获取各组大鼠组织标本。采用RT-PCR和Western blot分别测定肝组织A20 mRNA和蛋白表达,测定肝组织中p-JNK、p-ERK、p-p38、Bcl-2、Bax和Caspase-3的蛋白表达情况,TUNEL法检测肝细胞凋亡指数。结果 D组大鼠A20 mRNA和蛋白表达均明显增加(P<0.05),p-JNK、Bax、Caspase-3蛋白表达明显下降(P<0.05),而p-ERK、Bcl-2蛋白表达明显增加(P<0.05)。C、D组大鼠与B组相比肝细胞凋亡率明显下降(P<0.05),其中D组下降更明显。结论自制肝保护液经门静脉在体持续低温灌注肝脏,可以促进A20 mRNA及其蛋白的高表达,由此激活ERK信号转导通路,抑制JNK信号转导通路,反向调节线粒体途径Bcl-2/Bax蛋白平衡的逆转,降低了其下游的凋亡执行蛋白Caspase-3,从而起到抗凋亡、保护肝细胞的作用。
Objective To investigate the effect of self-made hepatoprotective fluid on hepatocyte injury induced by portal vein occlusion in obstructive jaundice rats and its effect on the signal transduction pathway of hepatocellular MAPKs. Methods Totally 72 Wistar rats were randomly divided into 4 groups: Group A: biliary recanalization, free portal vein around the tail, removal of the caudate lobe and closure of the abdominal cavity after 90 min; Group B: recanalization of the biliary tract, , The remaining hepatic portal vein blood flow was blocked for 90 min, the normal portal vein was recovered, the caudate lobe of the liver was resected, and the abdomen was closed. Group C: Group B + portal vein perfusion of lactated Ringer’s solution at 4 ℃; Group D: Group B + The portal vein was infused with home-made liver solution (HMLS) at 4 ° C. At 0, 6 and 24 h after operation, the tissue samples of each group were obtained. The expression of A20 mRNA and protein in liver tissue were detected by RT-PCR and Western blot respectively. The protein expressions of p-JNK, p-ERK, p-p38, Bcl-2, Bax and Caspase-3 in liver tissue were detected by TUNEL assay Hepatocyte apoptosis index. Results The mRNA and protein expressions of A20 in group D were significantly increased (P <0.05), the expressions of p-JNK, Bax and Caspase-3 were significantly decreased, while the expressions of p-ERK and Bcl- (P <0.05). Compared with group B, the apoptosis rate of hepatocytes in group C and group D was significantly decreased (P <0.05), and the decrease in group D was more obvious. Conclusions Self-made hepatoprotective fluid is able to promote the expression of A20 mRNA and its protein in the liver via continuous portal vein hypothermia, which can activate ERK signal transduction pathway, inhibit JNK signal transduction pathway and reversely regulate mitochondrial pathway Bcl-2 / The reversal of Bax protein balance reduces the downstream apoptotic executive protein Caspase-3, thereby preventing apoptosis and protecting hepatocytes.