祛湿化瘀方对非酒精性脂肪性肝病小鼠肝组织谷胱甘肽合成的影响

Effects of Qushi Huayu Formula on glutathione synthesis in non-alcoholic fatty liver disease mice

目的:观察祛湿化瘀方对非酒精性脂肪性肝病(NAFLD)小鼠肝脏谷胱甘肽(GSH)合成的影响。方法:C57BL/6J小鼠随机分为正常组、模型组、祛湿化瘀方组和奥贝胆酸组,除正常组外,均予高脂饮食。12周末,各药物组分别灌胃祛湿化瘀方(2.14 g·kg^(-1)·d^(-1))和奥贝胆酸(10 mg·kg^(-1)·d^(-1))4周,其余组予等量饮用水。观察血清丙氨酸转氨酶(ALT)及肝脏病理、甘油三酯(TG)、过氧化氢(H_(2)O_(2))、GSH,RT-PCR和Western Blot法分别检测肝脏超氧化物歧化酶1(SOD1)、谷胱甘肽过氧化物酶4(GPX4)、酰基-CoA合成酶长链家族成员4(ACSL4)、谷氨酸半胱氨酸连接酶催化亚基(GCLC)与调节亚基(GCLM)、谷胱甘肽合成酶(GSS)mRNA及GCLC蛋白表达。结果:与正常组比较,模型组小鼠肝组织可见脂肪沉积、炎症反应、气球样变;血清ALT、血清胰岛素及肝组织TG及H_(2)O_(2)含量显著升高(P<0.01);SOD1、GPX4、GCLC mRNA表达显著降低(P<0.05,P<0.01)。与模型组比较,祛湿化瘀方组小鼠肝组织脂肪变性、炎症细胞浸润及肝细胞气球样变均减轻;血清ALT、血清胰岛素及肝组织TG含量和H_(2)O_(2)含量显著降低(P<0.01);肝组织中GSH含量显著增加(P<0.01);SOD1、GPX4、ACSL4、GCLC、GCLM、GSS mRNA表达显著升高(P<0.01,P<0.05);GCLC蛋白表达显著升高(P<0.05)。结论:祛湿化瘀方治疗NAFLD与其促进GSH合成、改善氧化损伤密切相关。

Objective:To investigate the effects of Qushi Huayu Formula(QHF)on the synthesis of glutathione(GSH)in non-alcoholic fatty liver disease(NAFLD)in mice.Methods:C57BL/6J mice were randomly divided into the control group,model group,QHF group and Obeticholic Acid group.Mice in the control group were fed with control diet and the others were fed with high-fat diet.At the end of 12th week,mice in the QHF or Obeticholic Acid group were administrated with QHF(2.14 g·kg^(-1)·d^(-1))or Obeticholic Acid(10 mg·kg^(-1)·d^(-1)),respectively till to the end of 16th week,and the others were administrated with distilled water.Liver histopathology(hematoxylin-eosin staining and oil red staining),serum alanine aminotransferase(ALT),hepatic triglyceride(TG),GSH and hydrogen peroxide(H_(2)O_(2))were measured.The mRNA expression of superoxide dismutase1(SOD1),glutathione peroxidase 4(GPX4),acyl-CoA synthetase long chain family member 4(ACSL4),glutamate-cysteine ligase catalytic subunit(GCLC),glutamate-cysteine ligase modifier subunit(GCLM)and glutathione synthetase(GSS)in liver tissue were detected by real-time polymerase chain reaction(RT-PCR).The protein expression of GCLC was detected by Western Blot.Results:Compared with the control group,lipid deposition,ballooning degradation and the lobular inflammation were observed in the live tissue in the model group;serum ALT,serum insulin,hepatic TG and H_(2)O_(2)contents in model group mice were significantly increased(P<0.01);the mRNA expression of SOD1,GPX4,GCLC in the model group were significantly decreased(P<0.05,P<0.01).Compared with the model group,lipid deposition,ballooning degradation and the lobular inflammation were ameliorated in the QHF group;serum ALT,serum insulin,hepatic TG and H_(2)O_(2)contents in the QHF group mice were significantly decreased(P<0.01);hepatic GSH content was significantly increased(P<0.01);the mRNA expression of SOD1,GPX4,ACSL4,GCLC,GCLM,GSS were significantly increased(P<0.01,P<0.05);the protein expression of GCLC was significantly increased(P<0.05).Conclusion:QHF ameliorates NAFLD,which is probably associated to increasing hepatic GSH synthesis and improve oxidative stress.