Blueberry-derived Exosomes-like Nanoparticles Antagonize Oxidative Stress And Its Associated Molecular Mechanisms In The Hepatocytes Of HFD-fed Mice

Nonalcoholic fatty liver disease（NAFLD）is a common chronic liver disease in worldwide,which is characterized by the accumulation fat of liver and is associated with obesity,hyperlipidemia and insulin resistance.Accumulating evidence indicates that mitochondrial dysfunction and oxidative stress play a pivotal role in the initiation and progression of NAFLD.Antioxidant supplementation may helpful to improve the pathological symptoms of NAFLD.In this study,the antioxidant activity of exosomes-like nanoparticles（ELNs）from14 different plants was evaluated.The results indicated that,blueberry-derived exosomes-like nanoparticles（BELNs）significantly increased the cell viability in H₂O₂-treated human Hep G2 hepatocytes.Based on the analysis of morphology,particle size,Zata potential,RNA,lipid,protein and the uptake of Hep G2 cells,BELNs displayed exosomes-like characteristics.In addition,pre-incubation with BELNs could alleviate the cytotoxicity of rotenone（an inhibitor of mitochondrial complexⅠ）by increasing the cell viability in Hep G2 cells in a dose-dependent manner.We also observed that BELNs significantly increased the content of mitochondria,inhibited the production of ROS and significant reduction of mitochondrial membrane potential in a dose-dependent manner in rotenone-treated Hep G2 cells.Furthermore,BELNs attenuated the cell apoptosis via affecting the distribution of nuclear factor E2-related factor 2（Nrf2）in the cytoplasm and cell membrane to increase the expression of anti-apoptotic proteins such as Bcl-2 and HO-1,and reducing the content of Bax protein.After intragastric administration of BELNs for continued 4 weeks,BELNs improved the insulin resistance,reduced fasting blood glucose and insulin content in a high-fat diet mouse model of NAFLD.BELNs also improved the liver dysfunction by reducing the expression of fatty acid synthesis-related genes including acyl-Co A carboxylase（ACC）,fatty acid synthase（FAS）and the contents of aspartate aminotransferase（AST）and alanine aminotransferase（ALT）,prevent the formation of vacuoles in liver tissue and reduce the accumulation lipid droplets of liver.Further studies found that BELNs can mediate the activation of Nrf2 in the nucleus to regulate the expression of antioxidant genes,regulate the activities of superoxide dismutase（SOD）and glutathione（GSH）in the liver tissue of obese mice,and reduce the content of malondialdehyde（MDA）to inhibit hepatocytes apoptosis.The present data revealed BELNs regulated the distribution of Nrf2 in the cytoplasm and cell nucleus to influence the expression of anti-apoptotic proteins to antagonism the injury and apoptosis in rotenone-induced Hep G2.Furthermore,BELNs also improved insulin resistance in high-fat diet induced mouse model of NAFLD.BELNs regulated the expression of fatty acid synthesis-related genes and the distribution of Nrf2 in liver to attenuate the hepatic oxidative stress.All the findings of this study indicate that BELNs can be used as a novel antioxidant and has a wide perspective application in the prevention and treatment of NAFLD.