The Study On The Role And Mechanism Of Active Peptides In Protecting The Liver

Objective: Metabolic associated fatty liver disease(MAFLD)is a condition that causes lipid degeneration due to excessive accumulation of liver fat in the body,and is accompanied by liver damage.Clinically,the symptoms and pathological changes of MAFLD include simple fatty liver(NAFL),non-alcoholic steatohepatitis(NASH),cirrhosis and even liver cancer.With the continuous improvement of people’s material quality of life,people’s dietary habits and life structure have changed,the number of obese people has increased,resulting in a rising incidence of MAFLD.However,there are no treatment has been approved for MAFLD.At the same time,since oxidative stress is a key factor in the pathogenesis of MAFLD,finding antioxidants that have protective effects on the liver is expected to become one of the new treatment strategies for MAFLD.Peptides are ubiquitous in various organisms and have many biological activities,including antioxidant activity.Active peptides have attracted the attention of many drug developers due to their low toxicity and easy absorption.So,we screened peptides with anti-oxidant activity in the early stage to explore the protective effect of active peptides on oxidatively damaged liver and related mechanisms.Method: In this study,AAPH-induced oxidative damage L02 cell model and CCL4-induced acute liver injury model in mice were used as research objects.The effects and mechanisms of active peptides were evaluated in vitro and in vivo respectively.In vitro experiments,we screened peptides with antioxidant activity by MTT assay,and then tested the effects of peptides on ROS level,mitochondrial membrane potential,apoptosis and cell cycle block of oxidatively damaged L02 cells by flow cytometry.And the related oxidation indicators(SOD,MDA)and ATP levels were detected.In vivo experiments,we divided mice into a normal control group,a CCl4 model,and a peptide administration group.The peptide a at a dose of 50 mg / kg was administered by intraperitoneal injection once a day for a week.One hour after the last administration,0.2% CCl4 was used for modeling.Subsequently,we tested the serum levels of ALT,AST,ALB,ALP,TC and TG,as well as the levels of inflammatory factors IL-1β and IL-6.Next,we went to the right lobules of each group of mice for paraffin embedding,and observed the pathological changes of the liver by HE staining.Finally,Western Blotting was used to detect the expression of oxidative stress,mitochondrial apoptotic pathway,and mitochondrial autophagy-related proteins in mouse liver tissues.Results: MTT results showed that peptide a can increase the survival rate of AAPH-induced oxidatively damaged LO2 cells.Compared with the AAPH model,peptide a can reduce intracellular MDA levels and increase SOD activity and ATP levels.The results of flow cytometry showed that compared with the AAPH model,peptide a can reduce the level of intracellular ROS,which increases the cell mitochondrial membrane potential,thereby reducing apoptosis,and can also reduce the S-phase cycle block proportion.The results of animal experiments show that compared with the CCL4 model,peptide a can significantly reduce the levels of ALT and AST in serum and significantly improve liver pathological changes,and can reduce the levels of IL-β1 and IL-6 in serum.Western Blotting results showed that compared with the CCL4 model,the expression of Nrf2 pathway-related proteins Nrf2 and NQO1 was increased in the liver tissue of mice given the peptide a preventively.The expressions of mitochondrial apoptotic pathway-related proteins p53,Bax,cytochrome c(cyto-c),Caspase3 and Cleaved PARP were reduced,and Bcl-2 expression is increased.The expression of mitochondrial autophagy related proteins AMPKα,PINK1,Parkin,HSP60 and COX IV were increased.At the same time,the expression of inflammation-related factors NF-κB increased.Conclusions: In general,peptide a has anti-oxidative activity in oxidatively damaged liver cells and liver tissues.Peptide a regulates the activity of antioxidant enzymes through the Nrf2 pathway,reduces the level of ROS in cells,reduces the p53-mediated mitochondrial-dependent apoptosis induced by oxidative damage,and induced mitochondria in liver cells mediate AMPKα-PINK1 / Parkin pathway for autophagy and reduce inflammation,thereby protecting the liver from oxidative damage.These results provide more experimental evidence and possibilities for antioxidant peptides as candidates for MAFLD.