Study On The Anti-atherosclerotic Mechanism Of Polydatin Based On Intestinal Flora And Lipid Metabolis

Polygonum cuspidatum is a traditional Chinese medicine with detoxification and blood activating effects selected for the treatment of atherosclerosis based on the theory of "blood stasis and toxin" of Academician Chen Keji,a master of Chinese medicine.The research of its main effective component,polydatin in anti atherosclerosis has gradually increased,mainly focusing on regulating lipid metabolism,anti-inflammatory and antioxidant stress.The changes in gut microbiota and lipid metabolism are closely related to various diseases such as AS,and have also become an important content of modern research on traditional Chinese medicine.There is a good agreement between the role of gut microbiota and its lipid metabolism in the occurrence and development of AS and the traditional Chinese medicine theory of "stasis and toxin".On the one hand,the imbalance of gut microbiota leads to pathogenic microorganisms and other harmful components acting as antigens directly stimulating the body to induce inflammatory reactions,exacerbating the development of AS,which is similar to the pathogenic characteristics of "external toxin";on the other hand,the accumulation of lipid abnormal metabolites and metabolic disorder caused by intestinal microbiota imbalance indirectly affect the body’s inflammatory response and promote the progression of AS,which coincides with the pathogenic characteristics of traditional Chinese medicine’s "internal toxicity" and "blood stasis".As the main effective component of traditional Chinese medicine with detoxification and blood circulation promoting effects,studying its regulatory effects on lipid metabolism and intestinal microbiota is of great significance for further revealing the connotation and mechanism of polydatin anti AS effect.In order to further explore the anti atherosclerosis effect and mechanism of polydatin,this study conducted relevant experimental studies.Objective:To observe the effect of polydatin,an effective component of traditional Chinese medicine with detoxification and blood activating effect,on the pathomorphology of atherosclerosis,serum lipid group and gut microbiota in ApoE-/-model mice,and to explore the mechanism of polydatin against arteriosclerosis based on gut microbiota,lipid metabolism and their interaction.Methods:1.General research methods based on network pharmacology,to screen Polygonum cuspidatum and related targets of AS,construct Polygonum cuspidatum component as target protein interaction network,perform enrichment analysis of Polygonum cuspidatum component as target function and pathway,construct Polygonum cuspidatum component as target pathway network diagram,and analyze the core targets and the main active ingredients that exert drug efficacy;2.ApoE-/-male mice maintained on a high-fat diet were used as animal models of AS,while congenic C57BL/6J male mice were treated with low,medium,and high-dose polydatin intervention and simvastatin as positive control by gross oil red staining for HE,Oil Red O,Movat,CD68 and α-SMA immunohistochemical staining the experimental mice were pathologically stained to observe the effect of polydatin on atherosclerosis;3.To observe the pathological staining of the liver in ApoE-/-mice using HE staining combined with Oil Red O staining,to detect the serum lipid metabolism level using a biochemical instrument,and to investigate the effect of polydatin on the serum lipidome using a combination of untargeted lipidomics and targeted lipidomics;4.Using 16S rRNA with metagenomic sequencing,sequencing analysis of the gut microbiota of experimental mice;5.Statistical methods were used to analyze the correlation between serum lipids and gut microbiota and to explore the mechanism of anti atherosclerotic effect of polydatin.Results:1.Resveratrol,catechins,quercetin,and polydayin are the main effective components of Polygonum cuspidatum that exert anti AS effects,and their mechanism of action may involve targets such as AKT1,MAPK family,TNF,and IL6;The molecular docking results showed that polydatin and emodin were the most effective active ingredients and targets for docking;According to the literature and previous research results,polydatin is selected as the research object to carry out experimental research on anti atherosclerosis;2.Polydatin can dose-dependently reduce the area of AS plaques in the general aorta and aortic root of ApoE-/-model mice,reduce lipid deposition and macrophage content in the aortic root valve,increase collagen fiber and smooth muscle cell content,reduce plaque vulnerability index,and stabilize plaques.Among them,high-dose polydatin intervention has the best effect;3.Polydatin can dose-dependently reduce liver fat deposition and degeneration in ApoE-/-model mice,reduce the concentration of TC,LDL-C,TG in the main serum of model mice,and increase the concentration of HDL-C.The high-dose polydatin intervention group has the best effect,while polydatin significantly changes the serum lipid group of mice;Further differential analysis showed that polydatin could regulate the three signal pathways of glycerol phospholipid metabolism,sphingolipids metabolism and choline metabolism,and lactoceramide,ASGP,phosphatidylcholine,N-acylsphingosine and sphingomyelin might be potential biomarkers;The content of ASGP,N-acylsphingosine,galactose ceramide and lactoceramide in ApoE-/-model mice fed with high-fat diet significantly increased,and the content of phosphatidylcholine significantly decreased.The intervention of polydatin can reduce the content of galactose ceramide,ASGP,N-acylsphingosine and lactoceramide,and increase the content of phosphatidylcholine.The intervention effect of polydatin is dose dependent;4.There was no significant change in the alpha diversity of the gut microbiota in the six groups of mice,indicating that high-fat feeding and drug intervention did not significantly disrupt the richness,uniformity,and diversity of the gut microbiota in the experimental mice;However,the OTU analysis results showed that the OTU levels in the intestinal microbiota of AS model mice changed compared to the CN group,and the intervention of polydatin also changed the OTU levels in the intestinal microbiota of AS model mice.The comprehensive principal coordinate analysis results showed that high-fat feeding changed the intestinal microbiota composition of ApoE-/-model mice,and the intervention of polydatin also changed the intestinal microbiota composition of ApoE-/-model mice.Multiple group differences analysis showed that in the thick walled bacteria At the level of seven phyla including Bacteroides,there were significant differences in gut microbiota.The intervention of polydatin increased the abundance of Firmicutes in the mouse gut microbiota,while reducing the abundance of Bacteroides;In terms of species and function identification and difference analysis of intestinal flora,this study screened Lactobacillus as beneficial bacteria,and Muribaculum and Saccharibacteria as harmful bacteria.The gene functions involved include macB gene,tetA gene,and evgS gene.The related metabolic pathways are mainly lipid metabolism,starch and sucrose metabolism.At the same time,polydatin can significantly inhibit the effects of Capsule virulence factor and PhoP/R virulence factor,Its ability as a potential antibiotic has also been demonstrated;5.The study on the correlation between intestinal flora and lipid metabolism showed that the ordinal regression analysis showed that sphingomyelin,glucosinolate,galactose ceramide,N-acylsphingosine,and LDL-C were not related to the abundance of intestinal flora PCA1,while lactoceramide,phosphatidylcholine,ASGP,TG,HDL-C,and TC were significantly related to the abundance of intestinal flora PCA1,in which HDL-C and phosphatidylcholine were significantly positively related,and TC,TG,ASGP,and lactoceramide were significantly negatively related;Saccharibacteria was positively correlated with TC,LDL-C,galactose ceramide,ASGP,N-acylsphingosine and lactose ceramide,and negatively correlated with HDL-C;Muribaculum was positively correlated with TG,LDL-C,galactose ceramide,ASGP,N-acylsphingosine,and lactose ceramide,and negatively correlated with HDL-C;Lactobacillus was negatively correlated with TC,LDL-C,galactose ceramide,ASGP,N-acylsphingosine and lactoceramide.Conclusion:Polygonum cuspidatum is a traditional Chinese medicine with anti atherosclerotic effect under the guidance of the "blood stasis and toxin" theory.Its effective component,polydatin,can reduce the area of atherosclerotic plaque in ApoE-/-model mice and play a role in stabilizing plaque,and change the serum lipid group of ApoE-/-model mice and intestinal flora,and the interaction between intestinal flora and lipid metabolism plays a key role in the anti atherosclerotic process of polydatin.