Molecular Mechanism Of Lingguizhugan Decoction In The Treatment Of Non-alcoholic Fatty Liver Disease Through Inhibition Of STING-mediated Pathway

Background:The prevelance of non-alcoholic fatty liver disease(NAFLD)has been increasing year by year and has become a serious social burden worldwide.Due to the complex pathogenesis,there is a lack of recognized and effective treatment measures currently.During the recent years,the role of Traditional Chinese medicine(TCM)in the prevention and treatment of NAFLD has been recognized gradually.For example,the classic prescription Linggui Zhugan Decoction(LGZG)has a significant effect on NAFLD,though the mechanism of action is not yet clearObjectives:NAFLD model induced by leptin knockout ob/ob mice and high-fat-feeding mice were established to further explore the mechanism of LGZG through HNF1b/DPP4/NOX1 signaling pathway to alleviate insulin resistance as well as STING-TBK1-NF-κB pathway in liver macrophages to reduce liver inflammation.Methods:In animal study,leptin knockout ob/ob mice and high-fat feeding mice were used to establish NAFLD model.After the model was established,treatment with LGZG and positive drugs started.Body weight and blood glucose indexes were monitored every other week.Besides that,intraperitoneal glucose tolerance(IPGTT)and intraperitoneal insulin tolerance(IPITT)were detected every four weeks to observe changes in glucose tolerance and insulin resistance in mice before and after intervention.The content of serum total cholesterol(TC)and triglyceride(TG)in mice were detected to observe the effects of LGZG on lipid metabolism in mice.Whether LGZG could improve liver function caused by NAFLD in mice were observed by measuring the index of ALT,AST and γ GT.Liver Red O and HE staining were used to assess the severity of NAFLD in mice.Western blot was adopted to detect the macrophage infiltration in Kupper cells as well as the protein expression of STING-mediated pathway.Results:The mice weight in the model group significantly increased compared with the blank control group.However,compared with the model group,LGZG had no significant effect on the weight of mice(P>0.05).Compared with the blank control group,the fasting blood glucose of the model group was significantly increased(P<0.05).LGZG could reduce the hyperglycemia induced by high-fat diet but without significant statistical dereference compared with the model group(P>0.05).Compared with the blank control group,the insulin tolerance of the model group was significantly decreased with the serum ALT and AST increased dramatically(P<0.05).Compared with the model group,LGZG could reduce the y GT level with significant difference,but the there was no significant difference in serum ALT and AST.The results of Oil Red and HE showed that the liver lipid accumulation in the model group was obvious compared with the control group.The liver exhibited significant pathological changes of non-alcoholic steatohepatitis,with swollen hepatocytes as well as single or scattered cells that were swollen and necrotic.In addition,the Mallory Denk body appeared.After LGZG intervention,the accumulation of liver lipid droplets was reduced significantly and only small amounts of liver cells were swollen and necrotic.WB results showed that the expression of STING,TBK1,TNF-,IL-β in the liver of mice decreased significantly after administration(P<0.05).Conclusions:LGZG could significantly improve glucose and insulin tolerance in HF-C57 mice.In addition,LGZG can reduce fasting blood glucose,improve liver fat deposition as well as alleviate NAFLD symptoms in HF-C57 mice.This may be due to the mechanism of inhibiting the STING-TBK1-NF-κB pathway in Kupffer cells,reducing liver inflammation,thereby slowing down the deterioration process of NAFLD.