Effect Of Antiproliferative Protein Prohibitin1 On Mitochondrial Function And F0F1-ATP Synthase In C2C12 Mouse Myoblasts

Objective:To regulation the effect of PHB1 on the content of F0F1-ATPase and its synthesis activity in C2C12 cells.The effects of PHB1 on energy metabolism,oxidative stress(ROS)and ATP content were observed.To investigate the mechanism of PHB1 regulation of intracellular F0F1-ATPase and its effect on exercise ability,to reveal the molecular mechanism of mitochondrial self-regulation during exercise and whether PHB1 can be used as a scientific basis for regulating energy metabolism.Methods: The culture system of PHC1 overexpression and RNA interference was constructed,and a good C2C12 cell culture system was established.The transfection efficiency was determined by flow cytometry.Flow cytometry was used to determine the transfection efficiency.The expression of PHB1 in mouse skeletal muscle cell line C2C12 was detected by Western blot.It was confirmed that the two vectors increased and inhibited intracellular PHB1 The effect of expression.The expression of PHB1 in the skeletal muscle cell line C2C12 was detected by real-time fluorescence quantitative PCR.The expression of F0F1-ATPase in the cells was increased and the expression of PHB1 in C2C12 cells was decreased by Western blot.F0F1-ATPase protein expression levels.The changes of F0F1-ATPase activity in the cells were detected by mitochondrial respiratory chain complex V activity kit.The changes of ROS in the cells were detected by reactive oxygen species(ROS)(OCR)in C2C12 cells was detected by XF cell mitochondrial stress test kit.The changes of ATP content in C2C12 cells were detected by XF cell mitochondrial stress test kit.Results:1.Adenovirus vector(Ad-GFP)was transfected into C2C12 cells.The optimal transfection conditions were MOI = 150 and 48 h.2.The expression level of PHB1 was significantly increased in adenovirus high expression vector Ad-phb1 transfected with C2C12 cells,and the expression level of PHB1 was significantly decreased by adenovirus RNA interference vector Ad-phb1 shRNA transfected with C2C12 cells.3.Compared with the control group,the expression of PHB1,the level of FOF1-ATPase and the expression of F0F1-ATPase protein in C2C12 cells were significantly higher than those in the control group.Compared with the control group,the expression of PHB1 The expression of FOF1-ATPase mRNA and F0F1-ATPase protein in C2C12 cells were significantly decreased.4.Compared with the control group,the expression of PHB1 and the expression of F0F1-ATP synthase in C2C12 cells were significantly higher than those in the control group.Compared with the control group,the expression of PHB1 was decreased,and the expression of F0F1-ATP in C2C12 cells Synthetic enzymes were significantly reduced.5.Compared with the control group,the expression of PHB1,the content of ATP and the oxygen consumption rate(OCR)in C2C12 cells were significantly higher than those in the control group.The cell respiration was enhanced and the energy metabolism of the cells was increased.The expression of PHB1,the content of ATP and the oxygen consumption rate(OCR)in C2C12 cells were significantly decreased,the cell respiration was decreased and the energy metabolism of the cells was decreased.6.Compared with the control group,the expression of PHB1 in the C2C12 cells was significantly lower than that in the control group.Compared with the control group,the expression of PHB1 was decreased and the ROS production in C2C12 cells was significantly increased.Conclusion:1.By regulating the expression of PHB1,PHB1 can be found to increase the expression and activity of F0F1-ATP synthase,thereby enhancing the energy metabolism of cells.2.By regulating the expression of PHB1,the production of ROS in the cells decreased,indicating that PHB1 may be related to the stable mitochondrial structure.