Effect Of Sodium Butyrate On Redox State And Mitochondrial Energy Metabolism In HepG2 Cells Under H₂O₂-induced Oxidative Stress

Objectives: Oxidative stress leads to mitochondrial dysfunction and plays an important role in the occurrence and development of the metabolic syndrome.Butyrate,a volatile short chain fatty acids,is the byproduct of microbial fermentation of dietary fiber in the gastrointestinal tract and easily combines with sodium to form sodium butyrate?NaBu?.Previous studies show that NaBu modulates several antioxidant enzymes and glucose and lipid metabolism,which suggest that NaBu regulates the cellular antioxidant system and mitochondrial metabolism.However,the specific mechanism is still undefined.In the present study,we established a model of oxidative stress in HepG2 cells and investigated the effect of NaBu on redox staus and mitochondrial energy metabolism,and the related mechanism.Methods: HepG2 cells were exposed to 400 ?M,600 ?M,800 ?M H₂O₂ for 4 h to establish oxidative injury model.The experiment was divided into four groups: control,H₂O₂ group,NaBu+H₂O₂ group and LA+H₂O₂ group.After treatment,we detected ROS level,survival rate and cellular redox index by DCFH-DA assay,MTT assay and appropriate kits,respectively.Lactate level,NADH/NAD+ ratio,acetyl coenzyme A and ATP content were detected.The mitochondrial membrane potential was tested by flow cytometric.The mRNA expression of Nrf2 pathway,glycolysis,?-oxidation and TCA cycle,and mitochondrial DNA copy number were analyzed by Real-time PCR assay.TFAM and PGC-1? protein level,GSK-3? ser9 phosphorylation level and Nrf2 nuclear-cytoplasmic ratio were measured by western blot assay.Results:?1?NaBu of 0.3 mM treated HepG2 cells for 96 h significantly reduced the oxidative stress,and induced an increase of T-AOC,the activity of SOD,CAT and GPX,GSH/GSSG ratio and survival rate and a decrease of ROS level and MDA content in oxidative injured HepG2 cells.?2?NaBu of 0.3 mM treatment upregulated the mRNA expression of PI3 K,Nrf2,HO-1 and NQO1,and increased the phosphorylation of GSK-3? ser9 and the nucleus to cytoplasm ratio of Nrf2 in oxidative injured HepG2 cells.?3?NaBu treatment inhibited the mRNA expression of GLUT4,HK I,HK II and PYK,and upregulated the mRNA expression of CPT1 A,HADH and ACADS,which resulted a decrease of lactate content and the increase of acetyl coenzyme A content in oxidative injured HepG2 cells.?4?NaBu protected oxidative injured mitochondrial function,which consist of a increase of the activity of mitochondrial MnSOD and GPX,mitochondrial membrane potential,the mRNA experssion of 2-OGDH and MDH2,mitochondrial DNA copy number,NADH/NAD+ ratio and ATP content in oxidative injured HepG2 cells.The mechanism may be related to the improvement of PGC-1? and TFAM.Conclusions: NaBu protectes cells from oxidative stress and maintains cellular redox homeostasis and mitochondrial energy metabolism in H₂O₂-induced oxidative injured HepG2 cells,which may be partly through a regulation of GSK-3?/Nrf2 pathway and an increase of nuclear translocation of Nrf2 and the expression of PGC-1? and TFAM.