清肝活血方调控Caspase-4/Caspase-3/GSDME介导的细胞焦亡改善酒精性肝损伤

Qinggan Huoxue Recipe regulates the cell pyroptosis mediated by Caspase-4/Caspase-3/GSDME to improve alcoholic liver injury

[目的]研究清肝活血方(QGHXR)通过调控半胱天冬酶(Caspase)-4/Caspase-3/GSDME细胞焦亡通路防治酒精性肝病(ALD)的作用机制。[方法]将18只C57BL/6小鼠随机分为对照组、模型组和QGHXR组复制ALD小鼠模型。检测血清肝功能、血脂水平;苏木精-伊红(HE)染色及油红O染色观察肝脏病理改变及肝脏脂质沉积状况;实时荧光定量逆转录聚合酶链反应(qRT-PCR)、蛋白免疫印迹(Western Blot)法及免疫荧光染色检测Caspase-4/Caspase-3/GSDME通路相关基因及蛋白表达情况。[结果]与空白对照组比较,模型组小鼠肝组织三酰甘油(TG)水平明显升高(P<0.01),血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)水平升高(P<0.05),血清白介素-1β(IL-1β)、白介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)水平升高(P<0.05);肝脏组织出现病理改变及脂质沉积现象;肝组织IL-1B、IL-6、TNF-α、NLRP3 mRNA表达水平升高。与模型组比较,QGHXR组小鼠肝组织TG水平明显降低(P<0.01),血清ALT、AST、总胆红素(TBIL)、总胆汁酸(TBA)水平降低(P<0.05),血清TG、IL-6、TNF-α水平降低(P<0.05)以及血清高密度脂蛋白(HDL)水平升高(P<0.01);血清中血脂、肝脏病理改变及脂质沉积现象缓解;GsdmeN、Caspase-4、Caspase-3和C-Caspase-3基因及蛋白水平降低(P<0.05)。[结论]QGHXR可通过调节Caspase-4/Caspase-3/GSDME介导的细胞焦亡通路改善ALD小鼠肝损伤和脂质沉积。

[Objective]To investigate the mechanism of Qinggan Huoxue Recipe(Qinggan Huoxue Recipe,QGHXR)in the prevention and treatment of alcoholic liver disease(ALD)by regulating Caspase-4/Caspase-3/GSDME pyroptosis pathway.[Methods]Eighteen C57BL/6 mice were randomly divided into control group,model group and QGHXR group to replicate the alcoholic liver disease model.Serum liver function and lipid levels were detected.Hematoxylin-eosin(HE)staining and oil red O staining were used to observe the pathological changes and lipid deposition in the liver.The expression of Caspase-4/Caspase-3/GSDME pathway related genes and proteins were detected by q-RT PCR,Western Blot and immunofluorescence staining.[Results]Compared with the control group,the liver TG level in model group was significantly increased(P<0.01),the serum ALT and AST levels were increased(P<0.05),and the serum IL-1β,IL-6 and TNF-αlevels were increased(P<0.05).Liver tissue showed pathological changes and lipid deposition.The mRNA expression levels of IL-1β,IL-6,TNF-αand NLRP3 in liver tissue were increased.Compared with model group,the liver TG level in QGHXR group was significantly decreased(P<0.01),and the serum ALT,AST,TBIL and TBA levels were decreased(P<0.05).Serum TG,IL-6 and TNF-αlevels were decreased(P<0.05)and serum HDL levels were increased(P<0.01).Serum lipid,liver pathological changes and lipid deposition were relieved.The levels of GsdmeN,Caspase-4,Caspase-3 and C-Caspase-3 genes and proteins were decreased(P<0.05).[Conclusion]QGHXR alleviates liver damage and lipid deposition in ALD mice by modulating the Caspase-4/Caspase-3/GSDME-mediated pyroptosis pathway.