全胃肠外营养对乳猪肝脏细胞死亡受体活化信号传导通路的影响

The Investigation of Total Parenteral Nutrition Induced Liver Apoptosis Mediated by Fas Pathway

目的了解全胃肠外营养对新生乳猪肝脏细胞死亡受体活化途径的影响，探讨其引起肝脏损伤的作用机制。方法以胃肠内营养乳猪作为对照，建立全胃肠外营养新生乳猪实验动物模型，喂养7d后获取肝脏组织，并采用胶原酶灌流法分离单个肝细胞，应用琼脂糖凝胶电泳检测肝细胞DNA凋亡条带，凋亡蛋白活性测定、Western blot印记法等检测全肠外营养诱导肝脏损伤，细胞死亡受体凋亡途径的蛋白表达。结果全肠外营养乳猪肝细胞活率为（49±21）％，明显低于肠内营养（s8±14）％（P〈0．05）。全胃肠外营养乳猪肝组织DNA电泳呈现典型梯形凋亡条带，细胞凋亡ELISA检测发现细胞凋亡定量高达对照组的2.6倍，Western blot分析提示全胃肠外营养组肝组织中pro caspase-8、-7和PARP凋亡蛋白酶原被活化；Fas蛋白量表达上调。凋亡蛋白酶caspase-3活性也明显升高，与对照组相比，增高了9．9倍（P〈0．01）。结论全胃肠外营养损伤乳猪肝脏组织，并可通过死亡受体信号传导通路诱导肝细胞凋亡。

Objective Using neonatal piglets as an animal model to investigate total parenteral nutrition（TPN） induced apoptosis mediated by fas pathway. Methods Neonatal piglets were sacrificed following 7 days on TPN. Control animals received enteral nutrition （EN）. Following removal of small liver segments for molecular analysis, Piglet hepatocytes （PH） were isolated by perfusion with collagenase and subjected to functional and metabolic studies. We used DNA fragmentation,cell death detection and western blot tO assess molecular events in the development of TPN-mediated liver injury. Results Viability of PH following cell isolation was dramatically decreased： between （49±21）% in TPN-fed piglets as compared to （88±14）% in EN-fed piglets（P〈0.05）. DNA fragmention in TPN-fed piglet cells increased 2.6-fold and caspase-3 activation was 9.9-fold higher （P〈0.01） than EN piglets. By western blot, and cleavage of PARE caspase-8 and -7 was higher in TPN liver tissues than in EN. Fas was upregulated in TPN-fed livers. Conclusion TPN induces significant oxidative stress, which results in apoptosis mediated by Fas pathway.