PI3K-p85与Caspase-3在脂多糖/氨基半乳糖诱导小鼠急性肝损伤的实验研究

Experimental study of PI3K-p85 and Caspase-3 on LPS/GalN-induced acute liver injury in mice

①目的观察凋亡标志蛋白在脂多糖/氨基半乳糖(LPS/GalN)诱导的小鼠急性肝损伤模型肝脏中表达、意义,探讨可能的信号转导通路。②方法小鼠急性肝损伤模型通过脂多糖(LPS)联合氨基半乳糖(GalN)腹腔注射诱导复制;实验分为损伤1、3、6小时组(腹腔联合注射LPS和GalN 0.2mL/只),对照组(只腹腔注射同等量的生理盐水);采用谷丙转氨酶/丙氨酸氨基转移酶(ALT/GPT)试剂盒测血清ALT含量,苏木精-伊红(HE)染色观察肝脏组织形态学变化,Western blot法检测肝脏组织PI3K-p85及Caspase-3蛋白表达水平。③结果与对照组比较,损伤1小时组ALT活性差异无统计学意义(P>0.05),3小时组ALT活性显著升高(P<0.05);损伤6小时组较损伤1小时组、损伤3小时组ALT活性均显著升高(P<0.05)。HE染色显示,损伤6小时组的肝脏结构紊乱,细胞核浓缩、深染明显增加。与损伤1、3小时组比较,损伤6小时组Caspase-3表达显著升高(P<0.05)而PI3K-p85表达明显降低(P<0.05)。与对照组比较,损伤1小时组PI3K-p85表达显著升高(P<0.05),而Caspase-3表达未明显升高(P>0.05)。④结论在肝损伤早期PI3K-p85表达增强而在肝损伤晚期PI3K-p85表达明显减弱,在肝损伤早期Caspase-3表达开始增强在肝损伤晚期Caspase-3表达最强,推测PI3K通路抑制、凋亡增强参与了晚期肝损伤过程。

Objective To observe the expression and significance of apoptotic marker protein in the liver of LPS/GalN-induced acute liver injury model in mice,and to explore possible signal transduction pathways.Methods Induced replication of acute liver injury model in mice by intraperitoneal injection of LPS and GalN.The mice were divided into groups of 1 h,3 h and 6 h(LPS combined with GalN 0.2mL/mice)and control group(only intraperitoneal injection of the same amount of saline).ALT/GPT kit was used to measure the content of serum ALT,HE staining was used to observe the morphological changes of liver tissue,and Western blot was used to detect the expression of PI3K-p85 and Caspase-3 protein in liver tissue.Results Compared with the control group,there was no significant difference in ALT activity in the 1H injury group(P>0.05),but significant increase in ALT activity in the 3H injury group(P<0.05),and significant increase in ALT activity in the 6h injury group compared with the control group and the 3H injury group(P<0.05).HE staining showed that the structure of liver was disordered and the concentration and hyperchromatism of nucleus increased significantly in 6h group.Compared with the 1H and 3H groups,the 6h group significantly increased Caspase-3 expression(P<0.05),while PI3K-p85 expression was significantly decreased(P<0.05).Compared with the control group,the expression of PI3K-p85 in the 1h groups increased significantly(P<0.05),while Caspase-3 expression was not significantly increased(P>0.05).Conclusion In the early stage of liver injury,the expression of PI3K-p85 increased,but in the late stage of liver injury,the expression of PI3K-p85 decreased;Caspase-3 expression began to increase in the early stage of liver injury,and it was the strongest in the late stage of liver injury.It is suggested that the inhibition of PI3K pathway and the enhancement of apoptosis are involved in the late stage of liver injury.