The Effects And Mechanism Of Capsaicin On Hypolipidemic By Affecting Gut Microbiota In Obese Mice

With the improvement of people's living standard,the incidence of obesity has been increasing for many years,as a result living standards improvements,lifestyle changes,excessive calorie intake,and physical inactivity.The world federation estimated that roughly 60 percent of the world's population will be overweight or obese by 2030,with at least 41 million children under the age of five.Obesity and its complications not only seriously endanger people's health and threaten people's lives,but also impose a heavy medical investment and economic burden on society,families and individuals.Therefore,the prevention and treatment of obesity has become a major public health challenge facing all countries in the world.Capsaicin,as an active ingredient extracted from natural food,has been found to have good anti-obesity effect and fewer side effects,and has been widely concerned by more and more researchers.In the research on the lipid-lowering mechanism of capsaicin,the activation of the instantaneous receptor potential receptor cation channel(TRPV1)is considered to be the key to its lipid-lowering effect,and TRPV1 is involved in many of its mechanisms.At the same time,more and more studies have shown that gut microbiota is closely related to the occurrence and development of obesity,and capsaicin has been reported to affect the composition of gut microbiota,which is more conducive to lipid-lowering.So,the regulation of gut microbiota has become one of capsaicin's lipid-lowering mechanisms.However,there is a lack of systematic and in-depth research on this mechanism.Current studies only focus on the effect of capsaicin on the species and quantity of gut microbiota,while there are few in-depth research on how capsaicin causes changes in gut microbiota to regulate the absorption and metabolism of lipids in the body,which mechanism is not very clear requiring further exploration.And none of these studies ruled out a role for TRPV1 in lipid-lowering.Thus,the aim of this study was elucidate hypolipidemic mechanism of capsaicin acting on the gut and gut microbiota,which further enrich lipid-lowering mechanism of capsaicin and provide a scientific basis for the application of capsaicin as lipid-lowering drugs or health care products.In this study,TRPV1 knockout mice and caco-2 intestinal epithelial cells were used as animal models to exclude the interference of TRPV1 cation channel pathway.The main conclusions of the research are as follows:(1)The research has been conducted on capsaicin played a lipid-lowering role by influencing the composition and fermentation products of gut microbiota in obese mice.Eight-week-old female TRPV1 knockout mice(KO)and wild type mice(WT)were each categorized into three groups(8 mice/group,a total of 6 groups): control group,model group and dose group.Control group was fed a standard fat diet,model group was fed a high fat diet and dose group was fed a high fat diet and orally treated with 2mg/kg·bw of capsaicin on alternate days for 12 weeks.The gut microbiota composition and intestinal short chain fatty acids(SCFAs)composition were measured respectively by high-throughput sequencing and gas chromatography-mass spectrometry(GC-MS).At the same time,the weight,feed intake,blood lipid and blood sugar were also measured to explore the effects of capsaicin on the gut microbiota,SCFA production,and host energy metabolism with and without TRPV1 cation channel activation.We observed significantly lower gains in body mass,TGs,cholesterol,and insulin,and a smaller glucose AUC in WT and KO mice of dose group compared with WT and KO mice of model group,which suggests that intragastrical administration of capsaicin significantly inhibits HFD-induced body mass gain,but this effect is smaller when TRPV1 channels are not activated.The effect of CAP on weight gain and food intake may be mediated by an increase in the relative abundances of Bacteroides,Coprococcus,Prevotella,Akkermansia,Odoribacter,Allobaculum and S24-7,which are beneficial for weight loss in obese mice.This change in the relative abundances of those SCFA-producing bacterial species results in enhanced acetate and propionate production in the gut,which also are beneficial for energy balance in obese mice.The effect of CAP may be mediated by reducing the relative abundances of Proteobacteria spp.,such as Desulfovibrio,Escherichia,Sutterella and Helicobacter,which can generate a chronic,low-grade inflammatory response associated with higher risks of obesity.(2)The research has been conducted on capsaicin played a lipid-lowering role by inhibiting the appetite of obese mice through the Microbiome-Gut-Brain Axis(MGBA)pathway.Eight-week-old female KO mice were categorized into three groups(10 mice/group): control group,model group and dose group.Control group was fed a standard fat diet,model group was fed a high fat diet and dose group was fed a high fat diet and orally treated with 2mg/kg·bw of capsaicin on alternate days for 12 weeks.The weight,feed intake,blood lipid,blood sugar,intestinal hormones and the related indicators of the MGBA pathway were measured.The results showed that capsaicin affected the concentration of acetic acid and propionic acid in intestinal tract by increasing the relative abundances of Bacteroides,Coprococcus,Prevotella,Akkermansia,Odoribacter,Allobaculum and S24-7,which produces acetic acid and propionic acid.Increased concentrations of acetic acid and propionic acid in the intestine upregulated the expression of SCFAs receptor(G protein-coupled receptor 43,FFAR2)in intestinal epithelial cells.Up-regulated FFAR2 stimulated the secretion of glucagon-like peptid-1(GLP-1)and peptide YY(PYY)in L cells of the colon.GLP-1 mainly stimulated the vagal afferent neurons through the paracrine system,and transmitted signals to the hypothalamus through the vagus nerve,and upregulated the expression of GLP-1 receptor(GLP-1R)in the hypothalamus.PYY circulated through the blood to the hypothalamus,and upregulated the expression of PYY receptor(NPY2R)in the hypothalamus.The up-regulated GLP-1R and NPY2 R inhibited the NPY and AgRP neurons promoting foraging in the ARC of hypothalamus,and activated the POMC and CART neurons inhibiting foraging,thus increasing the sense of satiety,reducing appetite,and resulting in lower food intake in obese mice.Finally,capsaicin can reduce fat and reduce weight through this MGBA pathway.(3)The research has been conducted on capsaicin played a lipid-lowering role by reducing chronic,low-degree inflammatory response(CLGI).Eight-week-old female KO mice were categorized into three groups(10 mice/group): control group,model group and dose group.Control group was fed a standard fat diet,model group was fed a high fat diet and dose group was fed a high fat diet and orally treated with 2mg/kg·bw of capsaicin on alternate days for 12 weeks.The express ion of Toll-likereceptor4(TLR4),tumor necrosis factor alpha(TNF-?),interleukin 6(IL-6),cytoplasmic protein ZO-1 and transmembrane protein Occludin were measured.The concentration of lipopolysaccharide(LPS),IL-6,TNF-? and the related indicators of CLGI in the blood were also measured.The results showed that capsaicin reduced the concentration of LPS in the intestinal tract by reducing the relative abundances of Desulfovibrio,Escherichia,Sutterella and Helicobacter,which belonged to Proteobacteria.Increased concentrations of LPS downregulated the expression of IL-6 and TNF-?mediated by TLR4,which weakened the local inflammation in the intestinal tract.Weakened local inflammation leaded to increased expression of tight junction proteins ZO-1 and Occludin,which restored intestinal barrier function,reduced intestinal permeability and reduced LPS concentration in blood circulation.Weakened endotoxemia leaded to reduced concentrations of IL-6 and TNF-?in the blood,weakening CLGI and ultimately achieving the lipid-lowering and weight loss effect of capsaicin.(4)The research has been conducted on the effect of capsaicin on inflammatory response and barrier function of intestinal epithelial cells in vitro experiments,which were used to verify the lipid-lowering effect of capsaicin by weakening CLGI.TRPV1 knockout intestinal epithelial cell line(Caco-2,KO)was successfully build using CRISPR/Cas9 technology,and were categorized into three groups : control group,model group and dose group.Model group was treated with 1?g/mL of LPS and dose group was treated with 1?g/mL of LPS and 75?M of capsaicin.The expression of TLR4,Occludin and ZO-1 in cell,the concentration of IL-6 and TNF-?in the cell supernatant and transepithelial electrical resistance were measured.It was found that capsaicin did not inhibit the expression of TLR4 induced by LPS and reduce the concentration of IL-6 and TNF-?in the cell supernatant due to deletion of TRPV1 gene in cell,which indicating that capsaicin weakened local inflammation by reducing the concentration of LPS in vivo mice test.Capsaicin can stimulate the expression of zo-1 and Occludin,improve the transepithelial electrical resistance value of cells,indicating that capsaicin can repair the intestinal barrier function.Capsaicin reduced intestinal permeability and the concentration of LPS entering the bloodstream by repairing the intestinal barrier function,thereby weakening endotoxemia and CLGI.It was further confirmed the lipid-lowering effect of capsaicin by weakening CLGI in vitro experiments.In summary,capsaicin plays a lipid-lowering role by influencing the diversity of gut microbiota,increasing the number of lipid-lowering bacteria and the relative abundance of bacteria producing acetic acid and propionic acid through fermentation,and reducing the number of harmful bacteria belonging to Proteobacteria in the intestine.Capsaicin plays a lipid-lowering role by reducing appetite and feeding through the MGBA pathway which transmitted the signals of the gut sensory nerve to the neurons controlling appetite in the ARC of hypothalamus.Capsaicin plays a lipid-lowering role by weakening CLGI caused by LPS in the intestinal tract,which produced by harmful bacteria belonging to Proteobacteria.Capsaicin plays a lipid-lowering role by repairing the intestinal barrier function,which weakened CLGI.In this study,the mechanisms of lipid-lowering effect through influencing gut microbiota were explored,which are complementary to the lipid-lowering mechanism of capsaicin in addition to the TRPV1 cation channel.