| The gut microbiota is a large and complex micro-ecosystem,which plays a very important role in the health of the host.In recent years,many studies have shown that the gut microbiota has a significant effect in the occurrence and development of various diseases.And there are growing evidences that the gut microbiota has a crucial action in the treatment of various diseases with traditional Chinese medicine(TCM),known as 'the key to open the treasure house of TCMs'.TCMs are rich in chemical components,which inevitably interact with gut microbiota in the process of absorption,distribution,metabolism and excretion in vivo.Gut microbiota can affect the metabolism and absorption of TCMs ingredients,while TCMs ingredients can regulate the composition,structure and metabolism of gut microbiota,thus play a variety of roles.The occurrence and development of different diseases are often accompanied by disorders of gut microbiota,or metabolic abnormalities of gut microbiota.Therefore,the interaction between TCMs and gut microbiota may be one of the relevant mechanisms of TCMs for treating diseases.Cassiae Semen,the seeds of Cassia obtusifolia L.and Cassia tora L,named 'Jue Ming Zi' in China,originally published in 'Sheng Nong Ben Cao Jing',it has the traditional effect of clearing heat and improving vision,moistening the intestines to relieving constipation.As a medicinal and edible plant,Cassiae Semen has a medicinal history of more than 2000 years in China.It is mainly used to protect liver,relieve constipation,reduce blood fat,etc.At present,there are many studies on the traditional effect of Cassiae Semen to moisten intestines and relieve constipation,its mechanism is relatively clear;and many pharmacological studies have proved that Cassiae Semen has a liver-protective effect,but the research on its mechanism is still insufficient.Based on the research on the efficacy of Cassiae Semen in treating NAFLD,this article studied the mechanism of 'protective liver' of Cassiae Semen mediated by gut microbiota,for a better understand the mechanism of Cassiae Semen's traditional effect of 'clearing liver' and protecting liver,and provided a reference for the mechanism of Cassiae Semen treatment on liver disease.This thesis mainly included the following sections:1.Literature review.By searching CNKI,Wanfang Med Online,Cqvip,Pubmed,Web of Sciense and other commonly used literature databases,and consulting related books,to reviewed and summarized the research status of the hepatoprotective effect of Cassiae Semen and the application of gut microbiota in the studies of TCMs.The mechanism of the extract of Cassiae Semen or active ingredients in hepatoprotective effect mainly includes:i.reducing blood lipid and regulating lipid metabolism;ii.improving lipid peroxidation;iii.alleviating inflammatory reaction;iv.regulating intestinal flora and improving intestinal wall barrier,playing a role based on intestinal liver axis;v.immune regulation;vi.possibly related to defecation.The studies on mechanisms of lipid lowering and antioxidation of Cassiae Semen were relatively clear,but the other four possible mechanisms of the hepatoprotective effect of Cassiae Semen were not enough,and more experiments are needed to supplement.As for the application of gut microbiota in the studies of TCMs,this paper summarized the basic situation of gut microbiota(including classification and physiological functions,common metabolites),the effects of gut microbiota on the metabolism of TCMs components,the regulation of TCMs on gut microbiota,and the related mechanism of TCMs in the treatment of diseases based on gut microbiota.At present,the identified human intestinal flora is mainly composed of 9 bacteriums,which are dominated by two dominant bacteriums,Firmicutes and Bacteroides.According to their influence on host health,it can be divided into physiological bacteria,conditional pathogens and passing bacteria(mostly pathogenic bacteria).The main metabolites of gut microbiota include bile acids,short-chain fatty acids,trimethylamine and its oxides,and lipopolysaccharide,which have different function in maintaining host health in the intestinal tract.In addition,the gut microbiota can secrete many kinds of enzymes,which can carry out the disaccharide metabolism of the components contained in TCMs,and metabolize the components of TCMs by the way of 'hydrolysis mainly,oxidation-reduction as auxiliary'.Various components contained in TCMs can also regulate the composition of intestinal flora by changing the growth conditions or nutritional conditions of intestinal flora,and affect the metabolism of intestinal flora.These interactions between TCM and intestinal flora can partly explain the mechanism of TCMs in the treatment of many diseases.Many diseases are accompanied by gut microbiota disorders or changes in metabolites,such as gastrointestinal diseases(ulcerative colitis,gastrointestinal tumors,bowel stress syndrome,etc.),liver diseases(nonalcoholic fatty liver disease,cirrhosis,Liver cancer,etc.),metabolic diseases(diabetes,obesity,etc.),neurological diseases(depression,Parkinson's disease,Alzheimer's disease,etc.),cardiovascular diseases(coronary heart disease,etc.).In the treatment of these diseases,TCMs can often alleviate the symptoms of diseases by regulating the composition of gut microbiota and its metabolism.However,most of the current studies focused on the regulation of disease-related gut microbiota by TCMs,and there were few studies on the regulation mechanism of the host,the relevant target pathways involved in the regulation and the specific role of gut microbiota.2.Study on hepatoprotective effect of Cassiae Semen.Mice models of nonalcoholic fatty liver disease(NAFLD)were established by feeding with High Fat Diet(HFD-fed).and the extract of Cassiae Semen(JMZ),the extract of total aglycone of Cassiae Semen(TA),two main active monomers in Cassiae Semen:rubrofusarin-6-?-gentiobioside(RG)and aurantio-obtusin(AO)were given,respectively.To observe the effects of them on lipid accumulation,liver damage and liver inflammation.intestinal flora disorders,and intestinal wall barrier damage in NAFLD mice.The results showed that:i.Both JMZ and TA could reduce the liver weight index,total cholesterol(TC),triglycerides(TG),free fatty acids(FFA)and low-density lipoprotein cholesterol(LDL-c)of the model mice.RG and AO can reduce TC and LDL-c,but without significant change of TG and FFA.ii.Both JMZ and TA can reduce the serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)in NAFLD mice,but the two monomer components can only reduce the serum ALT and have no obvious effect on AST.iii.The four drugs can significantly reduce tumor necrosis factor-alpha(TNF-?),interleukin-6(IL-6)and increase anti-inflammatory factor IL-10 in model mice liver homogenate.iv.Compared with the control group,the gut microbiota diversity of the model group decreased,after administration,the gut microbiota diversity of mice in the model group increased,the relative abundance of Firmicutes and Bacteroidetes increased,and the relative abundance of Proteobacteria decreased.v.The relative expression levels of Occludin and ZO-1 m RNA in ileum of model mice significantly increased after JMZ,TA,RG and AO were given respectively,and the expression levels of Occludin and zo-1 proteins slightly increased after administration,but not significantly.The above results showed that these extract or monomer of Cassiae Semen can improve the lipid metabolism disorder,liver injury,liver inflammation,intestinal flora imbalance,and intestinal wall barrier damage in NAFLD mice to varying degrees.3.Transfered effect of fecal microorganism transplantation on the hepatoprotective effect of Cassiae Semen.In order to verify that the hepatoprotective effect of Cassiae Semen is exerted through the gut microbiota,we designed a fecal microbial transplantation experiment.Specifically,the fecal microorganisms of control group,model group,JMZ group and RG group were extracted and transplanted into HFD-fed mice in time to observe the changes of various detection indexes of each recipient group mice.And then found that the lipid accumulation,liver injury,liver inflammation,flora disorder and intestinal barrier damage of HFD-fed mice which accepted the fecal microbial transplantation from control group,JMZ group,RG group mice has certain improvement.It may be that the normal gut microbiota structure of control group and the improved gut microbiota structure of mice after administration affected and changed the gut microbiota structure of HFD-fed mice through fecal microbial transplantation,and thus played a role in protecting the live.However,it was impossible to exclude whether the drug prototype components or(and)its metabolites in the feces play a role.In conclusion,it can be preliminarily proved that the therapeutic effect of JMZ and RG on NAFLD mice will be transferred with fecal bacteria transplantation,and its liver protection depends partly on intestinal flora.4.Effects of antibiotic-induced gut microbiota disorder on the pharmacodynamics of Cassiae Semen.In order to further verify that the pharmacodynamic effects of Cassiae Semen depend on the intestinal flora,we designed an experiment in which antibiotics were given to induce the bacterial flora disorder of HFD-fed mice before administration,and observed the effects of antibiotic-induced bacterial flora disorder on the pharmacodynamics of JMZ.First of all,HFD-fed mice were given different antibiotics orally to cause gut microbiota disorder before administration,and freely drank sterilized drinking water containing antibiotics to maintain bacterial flora disorder.The results showed that the lipid accumulation,liver injury,liver inflammation,flora disturbance and intestinal barrier damage of HFD-fed mice were not improved after antibiotic treatment,and some indexes were even aggravated.These results proved that antibiotic induced gut microbiota disorder can weaken or even eliminate the efficacy of Cassiae Semen,and further proved our hypothesis that the efficacy of Cassiae Semen depends on gut microbiota.5.The key gut microbiota of Cassiae Semen change and its potential function prediction.In order to identify the key gut microbiota that played a role in JMZ's treatment of NAFLD and its functions,we used Metastats analysis to determine the type of flora system that changed significantly after HFD-fed and administering with JMZ,and explored the potential function of the microbiome with PICRUSt algorithm.The results showed that the gut microbiota enriched in the model group is mainly 10 genera under Proteobacteria,which were Sutterella of the Alcaligenaceae,Janthinobacterum of the Oxalobacteraceae,and 8 genera of Enterobacteriaceae,which were Enterobacter,Erwinia,Escherichia.Klebsiella,Morganella,Salmonella,Serratia and Trabulsiella;The abundance of these genera was reversed after administration,and JMZ significantly reduced the abundance of Erwinia,Klebsiella,Morganella,Trabulsiella.Dehalobacterium,Adlercreutzia,Odoribacter,Rikenella,Mucispirillum,Clostridium,Dorea,Lachnobacterium,Oscillospira and Ruminococcus,whose abundance decreased in the model group,increased after administration,especially Dehalobacterium,Oscilillospira,and Ruminococcus after JMZ treatment.The enriched flora in the model group was positively correlated with most disease-related indicators(except Occludin,ZO-1),while the enriched flora after administration was just the opposite.The bacteria genera with increased abundance in the model mice had the potential function of increasing metabolites related to endotoxemia,inflammatory response,obesity and insulin resistance,such as LPS,branched amino acids and aromatic amino acids,etc.On the other hand,the bacteria genera with increased abundance in the drug administration group had positive effects on lipid metabolism,bile acid production,etc.,while the bacteria with increased abundance in the model group had opposite effects,and inhibited the effects of these bacteria after drug administration.In concusion,these results indicated that Cassiae Semen and its components may regulate the abundance and composition of gut microbiota with specific metabolic functions,induce the production of metabolites beneficial to NAFLD,reduce harmful metabolites,improve endotoxemia and lipid accumulation in NAFLD mice,and reduce the intestinal barrier damage,liver damage and inflammatory response caused by lipid toxicity,bacterial translocation and other factors. |
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