Explore The Protective Mechanism Of Metformin On Bisphenol A-induced Liver Injury In Rats Based On Metabolomics

Background:Food health has gradually been attracted person's attention.Bisphenol A(BPA),as an organic material,has been extensively used in a variety of products,including baby bottles and food storage containers,etc.Moreover,a plethora of evidence has shown that long-term extensive exposure to BPA could cause reproductive toxicity,immune damage,and neurotoxicity,etc,The accumulation of BPA could easily cause oxidative stress and cause liver damage in vivo.Recently,Metformin hydrochloride,the first-line therapy for oral hypoglycemic agent,has undergone in-depth research in anti-inflammatory,anti-oxidant,anti-tumor and etc,.Excitingly,accumulated evidence has demonstrated that Metformin hydrochloride could can exert its pharmacological effects by enhancing the body's antioxidant capacity.Given the previous evidence,it may be of high interest to investigate the potential protective effect of Metformin hydrochloride on BPA-induced chronic liver injury.Purposes:We attempt to investigate the protective effect of Metformin hydrochloride on BPA-induced liver injury and analyze the molecular mechanism of its protective effect in rats.Method:1.Model:The rats were randomly divided into the following groups(n=7),(1)saline group(control),(2)corn oil group(vehicle),(3)Metformin hydrochloride group(Met,220 mg/kg),(4)bisphenol A group(BPA,500 mg/kg),(5)bisphenol A+Metformin hydrochloride group(BPA+Met,500+220 mg/kg)and(6)bisphenol A+diammonium glycyrrhizinate(positive control)group(BPA+DG,500+40 mg/kg),respectively.All rats were administered once a day for 14 days.2.Biochemical analysis and histopathology examination:The enzyme levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),alkaline phosphatase(ALP)and y glutamyl transpeptidase(GGT)and the histopathological examination of liver tissue were used to determine liver damage.Furthermore,the content of malondialdehyde(MDA),the activity of glutathione peroxides(GSH-Px)and the capacity of total antioxidant(T-AOC)in liver tissue was determined by using MDA,T-AOC and GSH-Px enzyme activity assay kits.3.Mechanism study:UPLC-MS/MS metabolomics was used to determine the changes of endogenous small molecule compounds of serum and liver samples in rats,UPLC-MS/MS was used to determine the change of homocysteine(Hcy),cysteine(Cys)and glutathione(GSH)in rats serum and liver samples;the expression of cystathionine ? Synthetic enzyme(CBS)and cystathionine y lyase(CSE)were quantitative determined by enzyme-linked immunosorbent assay(ELISA)in rat liver tissue.Results:1.From the results of biochemical analysis,we could find that the level of ALT increased from 28.93±3.56 U/L to 52.71±13.14 U/L,and the level of AST increased from 106.11±23.68 U/L to 147.71±32.12 U/L,the level of ALP increased from 248.72±60.10 U/L to 321.74±47.76 U/L,and the level of GGT increased from 0.38±0.24 U/L to 0.80±0.26 U/L after BPA-treated rats,compared with vehicle group rats.Compared with the BPA group rats,the level of ALT,AST and GGT decreased by 1.40,1.30 and 2 fold after Metformin hydrochloride administration,respectively.But the level of ALP did not decrease significantly.And the results indicated that hepatocyte protection effect of metformin on BPA-exposed rats was more significance than that of diammonium glycyrrhizinate.Histopathological results showed that metformin could ameliorate the irregular arrangement and inflammatory cell infiltration of rat liver cells caused by BPA.In addition,administration of metformin significantly ameliorated the endogenous anti-oxidant system as demonstrated by increased abilities of GSH-Px and total antioxidant capacity,and declined level of MDA in liver tissue in BPA rats.2.Metabolomics results showed that 21 differential compounds in serum and 26 differential compounds in liver were screened out after metformin treatment,respectively.Metabolomics data analysis of endogenous small molecule metabolites both in serum and liver tissue demonstrated that such beneficial action of metformin was mediated by modulating the dysregulated metabolism pathways,including the metabolism of cysteine and methionine metabolism,purine metabolism,arginine biosynthesis,and glutathione metabolism.3.The content measurement result showed that BPA-treated could increase the content of Hcy in the serum samples,and reduce the content of Cys and GSH in serum and liver samples.While after metformin administration,the content of Hcy was significantly reduced,and at the same time,the content of Cys and GSH were also increased.4.Enzyme-linked immunosorbent assay(ELISA)result showed that metformin could enhance the expression of CBS and CSE,and activate the homocysteine transsulfide pathway in the cysteine and methionine pathways.Conclusion:1.Metformin could exert the liver protective via reduce the levels of AST,ALT and GGT,and ameliorate the disorder of liver cell arrangement and inflammatory cells caused by BPA,as well as enhance the activity of antioxidant enzymes in BPA rats.2.Metformin up-regulated the endogenous anti-oxidant system mainly by enhancing the metabolism of cysteine and methionine as demonstrated by reducing the content of Hcy and increase the content of Cys and GSH,as well as the improved activities of CBS and CSE in BPA rats.As this,metformin could exert liver protection effect.