对乙酰氨基酚活化巨噬细胞HO-1/ROS途径诱导急性肝损伤的实验研究

Acetaminophen-induced acute liver injury through activation of HO-1/ROS pathway in macrophages

背景与目的:对乙酰氨基酚(APAP)是一种安全有效的退热镇痛抗炎药,但过量使用会迅速引发急性肝损伤和肝功能衰竭,从而导致患者死亡或需要进行肝移植。研究表明,巨噬细胞在维持肝脏稳态以及调控急慢性肝损伤的进展中发挥着重要作用,因此,本研究探讨巨噬细胞功能变化在APAP所致急性肝损伤中的作用与机制。方法:将BALB/c雄性小鼠随机分为对照组(生理盐水灌胃)、药物性肝损伤模型组(APAP组,600 mg/kg APAP灌胃)、药物性肝损伤模型+巨噬细胞清除剂氯膦酸盐脂质体(CL)组(APAP+CL组,APAP灌胃前12 h尾静脉注射CL)。APAP灌胃3 h后收集各组小鼠血清和肝脏组织标本。检测血清中碱性磷酸酶(ALP)、丙氨酸氨基转移酶(ALT)和天门冬氨酸氨基转移酶(AST)水平。观察肝脏组织病理改变,检测肝组织中活性氧(ROS)水平与血红素氧化酶1(HO-1)表达。将RAW246.7细胞分别用APAP和APAP+HO-1抑制剂锌原卟啉Ⅸ(ZnPPⅨ)孵育后,以无处理的RAW246.7细胞为对照,观察细胞ROS水平与HO-1表达的变化。结果:动物实验结果显示,与对照组比较,APAP组肝脏出现明显肝损伤病理改变,血清ALP、ALT和AST明显升高,肝组织ROS水平与HO-1表达明显升高;APAP+CL组以上指标的变化均被明显抑制,所有定量指标的差异均有统计学意义(均P<0.05)。细胞实验结果显示,APAP孵育后,RAW246.7细胞的ROS水平与HO-1表达明显升高,而同时用ZnPPⅨ孵育,APAP诱导的ROS水平与HO-1表达升高被明显抑制(均P<0.05)。结论:APAP可能通过诱导HO-1表达促进巨噬细胞ROS生成,从而导致急性肝损伤,对该途径的干预是临床急性肝损伤防治的新思路。

Background and Aims:Acetaminophen(APAP)is a safe and effective antipyretic,analgesic,and antiinflammatory drug.However,excessive use of APAP can rapidly induce acute liver injury and liver failure,leading to patient death or the need for liver transplantation.Studies have shown that macrophages are essential in maintaining liver homeostasis and regulating the progression of acute and chronic liver injuries.Therefore,this study investigated the role and mechanisms of macrophage functional changes in APAP-induced acute liver injury.Methods:Male BALB/c mice were randomly divided into the control group(gavage with normal saline),drug-induced liver injury model group(APAP group,600 mg/kg APAP gavage),and druginduced liver injury model plus macrophage depletion agent clodronate liposomes(CL)group(APAP+CL group,intravenous injection of CL 12 h before APAP gavage).After 3 h of APAP gavage,serum and liver tissue samples were collected from each group of mice.The serum levels of alkaline phosphatase(ALP),alanine aminotransferase(ALT),and aspartate aminotransferase(AST)were measured.Liver tissue pathological changes were observed,and the levels of reactive oxygen species(ROS)and heme oxygenase-1(HO-1)expression in the liver tissue were detected.RAW246.7 cells were cultured with APAP or APAP plus HO-1 inhibitor zinc protoporphyrinⅨ(ZnPPⅨ)using untreated RAW246.7 cells as a control.Then,the changes in cellular ROS levels and HO-1 expression were observed.Results:The results of the animal experiment showed that compared to the control group,the APAP group exhibited significant pathological changes in the liver,with significant increases in serum levels of ALP,ALT,and AST,as well as elevated levels of ROS and HO-1 expression in liver tissue.However,in the APAP+CL group,these changes were significantly suppressed,and the differences in all quantitative indicators were statistically significant(all P<0.05).The cell experiment results showed that ROS levels and HO-1 expression in RAW246.7 cells were significantly increased after incubation with APAP.However,when co-incubated with ZnPPⅨ,the APAP-induced ROS levels and HO-1 expression elevation were significantly inhibited(all P<0.05).Conclusion:APAP may promote macrophage ROS generation by inducing HO-1 expression,causing acute liver injury.Intervention targeting this pathway may provide a new approach to preventing and treating clinical acute liver injury.