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

关键词:  清肝活血方  酒精性肝病  焦亡  Caspase-4/Caspase-3/GSDME通路  中药复方

Key words:  Qinggan Huoxue Recipe  alcoholic liver disease  pyroptosis  Caspase-4/Caspase-3/GSDME pathway  traditional Chinese medicine compound

摘要:

[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。

Abstract:

[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-α levels were increased（P<0.05）. Liver tissue showed pathological changes and lipid deposition. The mRNA expression levels of IL-1β，IL-6，TNF-α and 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-α levels 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.