Study On The Therapeutic Effect Of Chicken Biliary Exosomes On Hepatic Steatosis

Exosomes are teacup shaped extracellular vesicles of 50-100 nm secreted by a variety of cells.They play an important role in various physiological and pathological processes such as lipid generation and metabolism,inflammation,apoptosis,and information transmission,and can be found in a variety of body fluids(bile,milk,blood,urine,etc.).Bile exosomes are mainly secreted by hepatocytes and bile duct epithelial cells.Our laboratory has previously found that chicken bile exosomes have certain targeting to liver injury,which can enhance liver immune function and inhibit liver injury.Hepatocellular steatosis mainly occurs during the peak period of laying hens in the poultry industry,which is manifested as excessive obesity,liver fat accumulation and decreased laying rate,which seriously affects the development of laying hens.Some studies have found that exosomes are closely related to lipid production and metabolism.In order to explore the therapeutic effect of chicken bile exosomes on hepatic steatosis and its mechanism,this study administered exosomes to evaluate the therapeutic effect by artificially inducing steatosis of chicken hepatocytes in vivo and in vitro.First,480 m L bile was collected from the farm,and exosomes were extracted by differential centrifugation.Then,specific proteins were detected by transmission electron microscopy and Western Blot.To establish in vitro and in vivo hepatocyte steatosis models.In vitro: Different concentrations of Oleic acid(OA)were applied to liver cells.CCK-8 method,alanine aminotransferase(ALT),aspartate aminotransferase(A ST)and Triglyceride(TG)assay boxes,q RT-PCR and red oil O staining were used to test the activity of liver cells.ALT and AST enzyme activities,TG content,relative transcription levels of endoplasmic reticulum stress and lipid transport-related factors.After staining hepatocytes with red oil O,the general content of lipid droplets in hepatocytes was observed under an inverted microscope.In vivo: Diets with different doses of soybean oil were fed to build models.Serum samples were collected at different time points and assay kit for ALT,AST and Triglyceride(TG)was used to detect the activity of ALT and AST enzymes and TG content in serum.After the in vivo and in vitro hepatic steatosis model was established,chicken bile exosomes were given,and red oil O staining,alanine aminotransferase(ALT),aspartate aminotrans ferase(AST)and TG detection kit,q RT-PCR,gross anatomy and histopathology were used.ALT and AST enzyme activities and TG contents in cell supernatant and serum,morphology of liver cells and histopathological observation of liver tissue,as well as endoplasmic reticulum stress and relative m RNA expression levels of lipid trans port and lipid generation related factors in liver cells and liver tissue were detected.The results showed that the chicken bile exosomes extracted by transmission electron microscopy showed typical teacup vesicles,and the carrier exosome specific protein Tsg101 was successfully extracted from chicken bile exosomes.In the in vivo and in vitro models of hepatic steatosis,TG content,ALT and AST activities were significantly increased compared with the normal group,and the relative expression level of endoplasmic reticulum stress factor(IRE1)was significantly decreased in the in vitro model.Therefore,hepatocyte steatosis models were successfully constructed in vitro(OA was 0.50 mmol/m L)and in vitro(soybean oil was 20% of the diet).Hepatocyte steatosis was induced.In the case of hepatic steatosis,the activities of ALT and AST in the supernatant and serum of chicken bile exosomes were significantly decreased after liver cells ingestion(P<0.001),TG content decreased(<0.001),by promoting the expression of IRE1,the expressions of lipid transport proteins(PDI,MTP,VLDL)were significantly increased,while the expressions of lipid generating factors(XBP1,ACC2,SCD1,Dgat2)were decreased.In conclusion,this study successfully extracted chicken bile exosomes and constructed the steatosis model of chicken hepatocytes in vitro and in vivo,confirming that chicken bile exosomes can promote lipid metabolism,repair abnormal lipid metabolism of liver cells,and improve lipid metabolism of liver tissue through endoplasmic r eticulum stress.Therefore,bile exosomes have a significant therapeutic effect on hepatocyte steatosis.