The Correlation and Mechanism between Improved Endotoxemia and Gut Microbiota by Bitter Melon Intervention in Obese Rats

Author:Bai Juan

Supervisor:dong ying


Degree Year:2019





With the rapid development of economy and the changes of human lifestyle,obesity has become one of the most important non-communicable chronic diseases which affect human health.Obesity can increase the risk of type 2 diabetes(T2DM),cardiovascular disease,nonalcoholic fatty liver disease and so on.Obesity is a chronic,low-grade inflammatory state.Studies have confirmed that the endotoxin from intestinal bacterial is one of the important factors triggering obesity-related low-grade inflammation,namely"metabolic endotoxemia".Recently,intervention in the obesity-related inflammatory process from the perspective of gut microbiota has been regarded as a new strategy to prevent and improve obesity and related metabolic diseases.Momordica charantia L.,a medicinal and edible plant,which is rich in a variety of bioactive components,possesses the functions of regulating glucose and lipid metabolism and improving insulin resistance.However,little was known about bitter melon alleviating the metabolic endotoxemia in obesity by regulating gut microbiota.In this present paper,bitter melon was selected as the raw material,and bitter melon powder(BMP)was prepared by freezing-drying and pulverization technology.It was aimed to study the effect of BMP on metabolic endotoxemia induced by high fat diet and its possible mechanism by establishing the obese rat model.The main contents and conclusions were presented as the following:1.The effects of BMP on insulin resistence and metabolic endotoxemia in obese rats were analyzed.Results were as below:(1)BMP could significantly reduce body weight and Lee’s index of obese rats,improve glucose tolerance,decrease the levels of serum fasting glucose,fasting insulin and insulin resistance index(HOMA-IR).Additionally,BMP could obviously reduce the level of total triglycerides(TG)and total cholesterol(TC)in serum,increase the level of high-density lipoprotein(HDL),as well as decrease the size of epididymal adipose cells and slow down the follicular lipid degeneration of liver cells.Therefore,BMP could slow down the weight gains,improve the impaired metabolism of glucose and lipid,thereby alleviating insulin resistance in obese rats.(2)BMP could remarkably lower the cytotoxicity of colonic contents towards HT-29 cell lines,increase the depth of the crypt and regularity of colonic villi,reduce the intestinal permeability by up-regulating the mRNA expression of colonic tight binding protein ZO-1 and Occludin,thereby lessening the enterogenous toxic substances entering the circulation system.This suggested that BMP may ameliorate metabolic endotoxemia by improving the disorder of colonic environment and repairing the intestinal barrier in obese rats.(3)BMP could significantly reduce the level of plasma endotoxin lipopolysaccharide(LPS),serum endotoxin binding protein(LBP)and pro-inflammatory factors(TNF-α,IL-6 and MCP-1),and blunt the inflammatory state in adipose,liver and muscle tissues.In addition,BMP could alleviate adipose inflammation by inhibiting the nuclear transcription factor kappa-B complex IκBα/NF-κB)and the mitogen-activated protein kinase(MAPKs)signaling pathway mediated by and c-jun amino-terminal kinase(JNK)/p38.Results indicated that BMP could effectively mitigate metabolic endotoxemia,thereby improving insulin resistance in obese rats.2.The effects of BMP on the the composition and function of gut microbiota in obese rats were analyzed by using Illumina Miseq high-throughput sequencing technology.Results were as below:(1)BMP could regulate the composition of gut microbiota in obese rats by increasing the relative abundance of Verrucomicrobia in the phylum level,reducing the abundance of Enterobacteriaceae,and selectively increasing the abundance of SCFAs-producers(Blautia,Allobaculum,Butyricicoccus,etc.)and Akkermansia which can protect the intestinal barrier.Resuls indicated that BMP could make the gut much healthier by changing the composition of gut microbiota in obese rats.(2)Based on the composition of gut microbiota,the KEGG function prediction showed that BMP could regulate the biosynthesis metabolism of carbohydrate,lipid and polysaccharide,such as tricarboxylic acid cycle,LPS synthetic protein and propionic acid metabolism,thereby improving the metabolic endotoxemia.3.The effects of BMP on the gut-generated metabolite profile in obese rats and its correlation with gut microbiota were studied by ~1H NMR technique.Results were as below:(1)BMP could change the profile of gut-generated metabolites in obese rats,which was significantly different from that of the obese control group.Specifically,BMP could notably increase the level of SCFAs(namely acetate,propionate and butyrate),glucose and ribose in sugars,taurine in amino acids and reduce the level of lactate and valerate.Pathway prediction showed that BMP made a significant effect on the metabolic pathways of butanoate,taurine and hypotaurine,and propanoate,which indicated that BMP could improve metabolic endotoxemia by regulating relevant metabolic pathways via altering the composition of gut-generated metabolites.(2)There were some correlations between the gut-generated metabolites and gut microbiota changed by BMP:butyric acid and propionic acid were positively correlated with SCFAs-producers Blautia and Coprococcus,respectively;valerate was negatively correlated with Blautia;both of glucose and ribose were positively correlated with Coprococcus;proline was positively correlated with Allobaculum.Results indicated that BMP could intervene metabolic endotoxemia by changing the gut metabolic profile in obese rats,which was correlated with the variation of gut microbiota.4.The effects of BMP on the gene expression profile of colonic mucosa in obese rats and its correlation with gut microbiota were analyzed by RNA-Seq technology based on Illumina HiSeq platform.Results were as below:(1)RNA-Seq screening revealed that there were 46 differentially expressed genes in the colonic mucosa between the normal and the obese model groups,while87 differentially expressed genes between the obese model and the BMP groups.Results indicated that BMP could significantly change the gene expression profile of colonic mucosa in high fat diet induced obese rats.(2)BMP could affect some pathways related with lipid metabolism,inflammatory response and glucose metabolism,such as arachidonic acid metabolismpathway,MAPK signaling pathway,phosphatidylinositol 3-kinase/protein kinase B(PI3K-Akt)pathway and so on.In addition,BMP could also regulate GO functions,such as remodeling of VLDL,negative regulation of sugar transport,lipoprotein metabolism and so on.Analysis of the interaction between genes and pathways showed that Prkcβand Pla2g2a were the core genes in the network,which were closely related with inflammatory response and lipid metabolism pathways.(3)There were some correlations between the differentially expressed genes related inflammatory response in colonic mucosa and gut microbiota.For example,Prkcβ,Nr4a1 and Mcpt1l3 were negatively correlated with Paraprevotella,Blautia and Allobaculum,respectively.Results indicated that BMP could ameliorate metabolic endotoxemia by changing the gene expression profile of colonic mucosa in obese rats,which was correlated with the variation of gut microbiota.