Protective Effect And Mechanism Of Sweet Potato Polysaccharide On Liver Injury Induced By Tris-(2,3-dibromopropyl) Isocyanurate

Tris-(2,3-dibromopropyl)isocyanurate(TBC)is a bromine compound containing a triazine ring structure and is one of the perfluorooctanoic acid substitute products.According to the requirements of the Stockholm Convention on Persistent Organic Pollutants,except for special purposes,China has completely banned PFOS.At present,TBC is widely used in the synthesis of flame retardants,plastics such as polystyrene,polyolefin,unsaturated polyester,polyurethane,textiles and rubber.With the increasing use of TBC,people inevitably come into contact with them through direct or indirect pathways.Therefore,it is of great strategic significance and research value to elucidate the hepatotoxicity of TBC and find effective preventive measures.In the previous study,our research team found that TBC has different degrees of damage to the liver,lung and kidney of rats,among which liver damage is the most serious.Purple sweet potato polysaccharide(PSPP)is a plant polysaccharide isolated and purified from the Sweet Potato Research Institute of the Chinese Academy of Agricultural Sciences "Sushu No.8" by the "Natural Anti-Tumor Research Center of the Ministry of Education".It has been reported in the literature that PSPP has a hepatoprotective effect.In the treatment of damaged liver with PSPP,serum liver function indicators such as alanine aminotransferase(GPT),aspartate aminotransferase(GOT)and lactate dehydrogenase(LDH)have a certain degree.The recovery,serum and liver antioxidant enzymes(SOD,GSH-Px and CAT)activity and total antioxidant capacity(T-AOC)increased,while the peroxidation product malondialdehyde(MDA)content decreased.Wu et al.reported that PSPP promotes energy metabolism in rats and increases the synthesis of amino acids and glutathione in rats.Therefore,combined with the biochemical mechanism of hepatic oxidative damage and the pharmacological mechanism of PSPP,we speculate that PSPP may also have a protective effect on oxidative liver injury induced by TBC.METHODS:(1)MTT assay was used to detect the effect of PSPP on the apoptosis of Hep G2 cells by TBC.(2)Morphological observation was used to observe the morphological changes of Hep G2 cells induced by PSPP.(3)Flow cytometry was used to determine the apoptosis rate of Hep G2 cells induced by PSPP.(4)Study on the mechanism of PSPP intervention in TBC-induced apoptosis of Hep G2 cells using PCR technology.(5)Western blot was used to detect the expression of related proteins induced by apoptosis.The mechanism of PSPP intervention on THP-induced apoptosis of Hep G2 cells was investigated from mitochondrial pathway and death receptor pathway,and the endoplasmic reticulum stress response was tentatively explored.(6)Pearson correlation,cluster analysis,and network analysis were used to explore the correlation of proteins and genes in the apoptosis pathway of Hep G2 cells induced by PSPP in TBC.RESULTS:(1)After different concentrations of PSPP,TBC had different degrees of attenuation on Hep G2 cell apoptosis.When the concentration of PSPP was 160?g/m L,the inhibition rate was only 6.38%.(2)Observe the state of Hep G2 cells administered for 72 h under an inverted microscope,As the dose of PSPP was increased,the adherent cells gradually returned to normal morphology.Observe the state of Hep G2 cells after 72 hours of administration under a fluorescent microscope,As the dose of PSPP is increased,the cells become larger and the rupture of the cell membrane weakens.The clearer the intercellular contours,the dead bodies around the cells gradually disappear.(3)The results of flow cytometry PI single staining showed that after PSPP intervention,TBC caused a decrease in apoptosis of Hep G2 cells.(4)PCR technology was used to detect the genes involved in the apoptosis of Hep G2 cells induced by PSPP in TBC.It was found that the expression of BCL2,BCL2A1,BCL2L1,BCL2L10,BCL2L11,and BCL2L2 genes in the mitochondrial apoptotic pathway increased with the increase of PSPP concentration.The changes of BAX CASP3,CASP9,APAF1,and CYCS gene expression decreased with the increase of PSPP concentration.In the cell death receptor apoptosis pathway,FADD,FAS,FASLG,CASP8,CD27,CD70,TNFRSF10 A,and TNFRSF10 B genes all decreased with the increase of PSPP concentration.(5)Western blotting was used to detect changes in the expression of apoptosis-related proteins in Hep G2 cells induced by PSPP in TBC by Western Blot in the mitochondrial pathway.After PSPP,the expression levels of Bcl-2 and Bcl-x L proteins increased with the increase of drug concentration.Increased;Bax,Caspase-3,Caspase-9,and Cytochrome C protein expression levels decreased with increasing drug concentration,and the Bcl / Bax protein expression ratio increased;in the death receptor pathway,after the action of PSPP The expression level of PARP protein increased with the increase of the concentration;CD95 / FAS,DR4,DR5,TRAIL,FADD,Caspase-8 protein expression level decreased with the increase of the concentration;endoplasmic reticulum In the stress response pathway,after the action of PSPP,the expression level of Caspase-12 protein increased with the increase of drug concentration.Conclusion: PSPP can inhibit the apoptosis of Hep G2 cells induced by TBC.In the mitochondrial pathway,PSPP attenuates proteolysis of members of the Caspases family by increasing the expression of Bcl/Bax protein,thereby preventing apoptosis.In the death receptor pathway,PSPP prevents apoptosis by reducing receptor proteins.In the endoplasmic reticulum stress response,the promoter Caspase-12 is prevented from being activated and inhibits apoptosis.PSPP can inhibit the apoptosis of Hep G2 cells induced by TBC through mitochondrial pathway,death receptor pathway and endoplasmic reticulum stress response.