Clinical Observation And Mechanism Study On The Treatment Of Metabolic Fatty Liver Disease (Liver Depression And Spleen Deficiency) With Sini Xiexin Formula

BackgroundMetabolic associated fatty liver disease(MAFLD)is a liver disease caused by factors such as obesity,insulin resistance,and high fat,excluding excessive alcohol consumption and other stimulating factors.At first,the disease can only be manifested as steatosis in the liver,and gradually it can appear inflammatory reaction,and later it can evolve into liver fibrosis,mainly including lipid accumulation in liver cells and diffuse alveolar steatosis of liver cells.MAFLD does great harm.With urbanization and changes in diet structure,MAFLD has become the most common cause of liver fibrosis,cirrhosis,liver cancer,type 2 diabetes,etc.in developed countries.With the development of the country,great changes have taken place in citizens’ eating habits and ways of work and rest.MAFLD is climbing,which has become the second largest cause of liver disease in China except chronic hepatitis B.MAFLD has seriously endangered human health,and the exploration of its pathogenesis and the development of diagnostic and therapeutic drugs are currently urgent issues to be addressed.The regulation of autophagy by endoplasmic reticulum Stress(ERS)is closely related to the pathogenesis and progress of MAFLD.The treatment of MAFLD by regulating autophagy through intervention of ERS by traditional Chinese medicine has theoretical support.The Sini Xie Xin Formula is an empirical formula for treating MAFLD(Liver Depression and Spleen Deficiency Type)by Professor Guo Peng,a mentor.It is a combination of Sini San and Banxia Xie Xin Tang,which has good clinical efficacy,especially in improving discomfort symptoms such as flank pain and abdominal distension.Modern pharmacological research has found that the drugs in the Sini Xiexin Formula have multiple therapeutic effects such as anti-inflammatory reactions,regulating lipid metabolism,alleviating liver fibrosis progression,and also have the effects of regulating ERS and autophagy.Therefore,this study proposes a hypothesis that the mechanism of treating MAFLD with Sini Xie Xin Formula is to regulate autophagy by intervening in ERS.This study designed two protocols simultaneously,clinical randomized controlled trials and animal experiments,which combined the two.On the one hand,to evaluate the effectiveness and safety of Sini Xie Xin Formula in the treatment of MAFLD(Liver Depression and Spleen Deficiency Type),and provide preliminary experimental evidence for its application.On the other hand,by analyzing the effects of Sini Xie Xin Formula on liver tissue ERS,autophagy,and mediator signaling pathway indicators in high-fat feed fed MAFLD model rats,to explore the mechanism of action of Sini Xie Xin Formula in the treatment of MAFLD,and to provide new experimental basis for drug research and clinical application.Purpose1.Evaluate the clinical efficacy and safety of Sini Xiexin Formula in the treatment of MAFLD(liver depression and spleen deficiency type).2.To evaluate the protective effect of Sini Xiexin Formula on MAFLD model rats fed with high-fat diet,and explore the regulatory mechanism of Sini Xiexin Formula based on AMPK signal pathway mediated endoplasmic reticulum stress regulation autophagy.MethodIn the clinical trial,a single center,exploratory,randomized,and controlled design was used to include 136 patients with MAFLD(liver depression and spleen deficiency type).They were randomly divided into the Sini Xie Xin Formula group and the Silybin group 1:1.By comparing the efficacy indicators such as TCM symptom quantification score,serum liver enzymology,blood bilirubin,and blood lipids,the baseline and horizontal differences between the two groups before and after treatment were analyzed,Evaluate the effectiveness and safety of Sini Xie Xin Formula in treating MAFLD(Liver Depression and Spleen Deficiency Type).In the animal experiment section,a MAFLD rat model was constructed by feeding it with high-fat feed,and intervention was given by gavage of high and low concentrations of Sini Xie Xin Formula Formula.To evaluate the protective effect of Sini Xiexin Formula on MAFLD rats by comparing the weight changes,liver tissue staining pathology,blood liver function,blood bilirubin,blood lipid indicators,and inflammatory factors of normal group,model group,high-dose Sini Xiexin Formula group,and control drug group rats.In addition,q-PCR and West bolt techniques were used to explore the expression of various indicators of endoplasmic reticulum stress,autophagy and mediated AMPK signaling pathway in rats of normal group,model group,Chinese medicine intervention group and control group before and after highfat feed induction modeling and before and after the intervention of Sini Xiehe Formula.Result1.In terms of improving traditional Chinese medicine symptoms,the total effective rate of the Sini Xie Xin Formula in treating MAFLD(liver depression and spleen deficiency type)is 96.9%,which is better than silybin(P<0.0001).The Sini Xiexin Formula formula can significantly improve various symptoms of MAFLD(liver depression and spleen deficiency type)patients,and is superior to silybin(P<0.05),especially in the main symptoms of rib distension and pain,loose stools and diarrhea,and chest and abdominal distension(P<0.001).Both Sini Xie Xin Formula and Silybin can improve the total score of TCM symptom quantification,and Sini Xie Xin Formula is significantly better than Silybin(P<0.001).2.Sini Xie Xin Formula can improve serum ALT(P=0.009),AST(P=0.039),GGT(P=0.010),TBIL(P=0.001),DBIL(P=0.002),IBIL(P=0.003),TBA(P=0.001),CHOL(P<0.001),and TG(P<0.001)in patients with MAFLD(liver depression and spleen deficiency type).This study did not observe the regulatory effect of Sini Xie Xin Formula on HDL-C,LDL-C,and VLDL.3.Serum ALT(P<0.05),AST(P<0.05),GGT(P<0.0001),ALP(P<0.0001),TBA(P<0.01),TC(P<0.01),LDL(P<0.001),TNF of MAFLD rats fed with high-fat feedα(P<0.001)、IL-1β The(P<0.01)level was significantly increased compared to the normal group,while the serum HDL(P<0.05)level was significantly reduced compared to the normal group.However,there were no significant changes in serum TBIL(P>0.05),DBIL(P>0.05),IBIL(P>0.05),TG(P>0.05),and IL-6(P>0.05).4.High dose Sini Xie Xin Formula can reduce serum ALT(P<0.05),AST(P<0.05),LDL(P<0.05),and TNF in MAFLD rats-α(P<0.0001)、IL-1β(P<0.05)level,but there was no significant effect on GGT,TBIL,DBIL,IBIL,ALP,TBA,TC,HDL(P>0.05)levels.5.Low dose Sini Xie Xin Formula can reduce serum LDL(P<0.05)and TNF in MAFLD rats-α(P<0.001)level,but there was no significant effect on the levels of ALT,AST,GGT,TBIL,DBIL,IBIL,ALP,TBA,TC,HDL(P>0.05).6.PPARG mRNA(P<0.0001)and protein expression level(P<0.0001),SREBP1 mRNA(P<0.0001)and protein expression level(P<0.0001),IRE1 in liver tissue of MAFLD rats fed with high-fat feed a MRNA(P<0.0001)and protein expression levels(P<0.0001),XBP1 mRNA(P<0.0001)and protein expression levels(P<0.0001),ATF6 mRNA(P<0.0001)and protein expression levels(P<0.0001),GRP78 mRNA(P<0.0001)and protein expression levels(P<0.0001),PERK mRNA(P<0.0001)and protein expression levels(P<0.0001),LC3 mRNA(P<0.0001)and protein expression levels(P<0.0001)Beclin-1 mRNA(P<0.0001)and protein expression level(P<0.0001),AMPK a The levels of mRNA(P<0.01)and protein expression(P<0.0001)were significantly increased compared to the normal group,while the levels of p62 mRNA(P<0.0001)and protein expression(P<0.0001),mTOR mRNA(P<0.0001),and protein expression(P<0.0001)were significantly reduced compared to the normal group.7.High dose Sini Xie Xin Formula can reduce PPARG mRNA(P<0.0001)and protein expression levels(P<0.0001),SREBP1 mRNA(P<0.0001)and protein expression levels(P<0.0001),and IRE1 in liver tissue of MAFLD rats a MRNA(P<0.0001)and protein expression levels(P<0.0001),XBP1 mRNA(P<0.0001)and protein expression levels(P<0.0001),ATF6 mRNA(P<0.0001)and protein expression levels(P<0.0001),GRP78 mRNA(P<0.0001)and protein expression levels(P<0.0001),PERK mRNA(P<0.0001)and protein expression levels(P<0.0001),LC3 mRNA(P<0.0001)and protein expression levels(P<0.0001)Beclin-1 mRNA(P<0.001)and protein expression level(P<0.001),AMPK α 1 mRNA(P<0.01)and protein expression levels(P<0.0001)can increase p62 mRNA(P<0.01)and protein expression levels(P<0.0001),mTOR mRNA(P<0.05)and protein expression levels(P<0.0001).8.Low dose Sini Xie Xin Formula can reduce PPARG mRNA(P<0.0001)and protein expression levels(P<0.0001),SREBP1 mRNA(P<0.0001)and protein expression levels(P<0.0001),and IRE1 in liver tissue of MAFLD rats α Protein expression level(P<0.0001),XBP1 mRNA(P<0.0001)and protein expression level(P<0.0001),ATF6 protein expression level(P<0.0001),GRP78 mRNA(P<0.0001)and protein expression level(P<0.0001),LC3 mRNA(P<0.05)and protein expression level(P<0.0001),Beclin-1 mRNA(P<0.0001)and protein expression level(P<0.0001),AMPK α 1.Protein expression level(P<0.0001)can increase p62 mRNA(P<0.01),protein expression level(P<0.0001),and mTOR protein expression level(P<0.0001),but it can affect IRE 1 α mRNA、ATF6 mRNA、PERK mRNA、AMPK a The levels of mRNA and mTOR mRNA(P>0.05)showed no significant effect.Conclusion1.Si Ni Xie Xin Formula can improve the discomfort symptoms of rib pain,fatigue,abdominal distension and pain,loose stools and diarrhea,chest and abdominal distension,and Xi Shan Tai Xi in patients with MAFLD(liver stagnation and spleen deficiency type),reduce the total score of traditional Chinese medicine symptoms,and lower serum levels of ALT,AST,GGT,TBIL,DBIL,IBIL,TBA,CHOL,and TG,with good safety.2.Sini Xiexin Formula can provide positive protection to MAFLD rats fed with highfat diet,improve liver steatosis,inflammation and fibrosis,and reduce serum ALT,AST,LDL and TNF-α、IL-1β,The effectiveness and safety of the Sini Xie Xin Formula in treating MAFLD were demonstrated from an experimental level.3.Sini Xiexin Formula can down regulate the stress index IRE1 of endoplasmic reticulum in MAFLD rats α、XBP1,ATF6,GRP78,PERK,inhibit endoplasmic reticulum stress,down regulate the intermediary pathway indicator AMPK,up regulate the intermediary pathway indicator mTOR,up regulate autophagy indicator p62,down regulate autophagy indicators LC3,Beclin-1,restore autophagy balance,suggesting that Sinixiexin Formula may regulate autophagy through AMPK signal pathway mediated endoplasmic reticulum stress to treat MAFLD.