Effects Of Long-term Exposure To DEHP On Cholesterol Metabolism In Rats Based On Intestinal Metabolomics

Di(2-ethylhexyl)phthalate(DEHP)is a widely used synthetic plasticizer.It is produced in large quantities and used in a variety of plastic products,and it is easy to seep out from plastic products.DEHP is a recognized environmental endocrine disruptor,and this study based on ultra-high performance liquid chromatography(HPLC)-quadrupole time-of-flight mass spectrometry technology of targeted metabonomics technology,to explore DEHP on cholesterol metabolism from the perspective of the intestinal metabolism,and the effects of host for protection and metabolic disease diagnosis and treatment of environmental endocrine disruptors.Methods:SD rats were gavaged with 0.5 mg/(kg·d)DEHP for 23 weeks.The effects of long-term exposure to DEHP on body weight and liver weight of rats were determined,and lipid-related indicators,including high density and low density lipoprotein,total triglycerides and total cholesterol,were measured from serum samples using the appropriate kit.Subsequently,the cecal tissues of the experimental group(D0.5)and the control group(D0)were analyzed by LC-MS non-target metabolomics.After the completion of the experiment,bioinformatics analysis was performed on the metabolomics results to identify the metabolites related to cholesterol metabolism in the intestines of rats in different treatment groups after long-term exposure to DEHP.Finally,fluorescent quantitative PCR and western blot were used to verify the genes and proteins involved in regulating differential metabolites.Results:1.The average body weight of the infected group was not significantly different from that of the control group,but the liver weight was significantly increased(P < 0.05).Hematoxylin-eosin staining(H&E)of liver slices showed an increased accumulation of lipids in the livers of DEHP-infected rats compared with the control group.2.Compared with the control group,the serum total cholesterol and low density lipoprotein in exposure group were significantly increased(P < 0.05).3.Appendix organization of non-target metabolomics results were identified five kinds of metabolites.PCA analysis shows that levels of metabolites significant difference between two groups of samples,which,compared with the control group,the infected group have three metabolites(cholic acid and deoxycholic acid and lithocholic acid)content increased,and the two kinds of metabolites(bovine sulfonated goose deoxycholic acid,taurocholic acid)content is lower.Bile acids composition were significantly different between the infected group and the control group,among which "primary bile acid biosynthesis" related to cholesterol metabolism was one of the most important way to enrich the differentially regulated metabolites.4.The validation results for the expression levels of genes related to cholesterol metabolism showed that the m RNA and protein expression levels of the rate-limiting enzyme CYP7A1 involved in bile acid synthesis in the liver of rats were significantly decreased after long-term exposure to DEHP(P < 0.05).MRNA and protein expression levels of FGF15 and FXR in small intestine were significantly increased(P < 0.05).Conclusions:Long-term exposure to DEHP can affect cholesterol metabolism in the intestinal tract of rats,activate FXR in the intestinal tract,induce the expression of FGF15,thereby inhibiting CYP7A1 in the liver and inhibiting bile acid synthesis,resulting in the imbalance of cholesterol metabolism in rats.