Effect Of Electrical Stimulation On Energy Metabolism Of Aging Myotube Cells Induced By H₂O₂

OBJECTIVE:To treat skeletal muscle myotube cells with H₂O₂,and obtain skeletal muscle myotube cell senescence model,and then use electrical stimulation to simulate exercise to interfere with aging myotube cells.Experimental observation of changes in the differentiation of skeletal muscle C₂C₁₂ cells into myotubes;H₂O₂ treatment of senescence of skeletal muscle myotubes;re-intervention of H₂O₂-treated skeletal muscle myotubes by electrical stimulation of cell movement to reveal movement for aging bones The influence of the expression of energy metabolism related indicators of muscle myotubes,and then supplement the relevant research results on aging skeletal muscle myotubes.METHODS:In experiment 1,skeletal muscle C₂C₁₂ cells were resuscitated and placed in DMEM medium（10%FBS）for culture.When the confluency was 70%-80%,the cells were digested with 0.25%trypsin,and then the cells were transferred to In the differentiation medium,the myotube cells after differentiation for 4 days were used for subsequent experiments;the H₂O₂ was diluted to a different concentration gradient in DMEM complete medium in 96-well plates,and the myotube cells were treated for 2 h and then cultured for 24h,usingβ-The galactosidase staining kit was used to detect and observe H₂O₂-treated myotube cells.In the second experiment,the experimental cells were divided into the control group（C group）,the control electrical stimulation group（CE group）,the H₂O₂ treatment group（H group）,and the H₂O₂ treatment electrical stimulation group（HE group）,and the RM6240 type electrical stimulator was used to in vitro.The cultured skeletal muscle myotube cells were subjected to electrical stimulation.The ATP level test kit was used to detect the ATP level of each group,and the PGC-1αprotein of skeletal muscle myotube cells was detected by Western blot.The experimental experiment of functional electrical stimulation was set to a current of 100μA,a frequency of 2.5 Hz,a pulse width of 20 ms,and a stimulation duration of 20 min.RESULTS:Results 1.There was no difference in myotube cell activity between the skeletal muscle myotubes and the C group after treatment with 100μmol/L H₂O₂.Aging model was used to treat skeletal muscle myotubes with 100μmol/L concentration of H₂O₂ for senescence,and a senescent cell model was obtained after 24 hours of culture.Results 2.In this experiment,the ATP level of group H was significantly lower than that of untreated group C（P<0.01）.The ATP level of HE group treated with H₂O₂ and then treated with electrical stimulation was significantly higher than that of H treated with H₂O₂ only.The expression of ATP in CE group was significantly higher than that in group C（P<0.01）.The expression of PGC-1αprotein in group H and group C was extremely significant（P<0.01）;There was a significant difference in the expression of PGC-1αprotein between H group and HE group（P<0.01）.There was no significant difference in PGC-1αprotein expression between C group and CE group（P>0.05）.Conclusion:H₂O₂ intervention is an effective way to promote aging of skeletal muscle myocytes.The ATP level expression of the aging model was significantly lower than that of the control model,and the electrical stimulation simulated exercise could increase the ATP level expression of skeletal muscle myotube cells.The expression of PGC-1αprotein in aging myotubes was significantly lower than that in the control model.After electrical stimulation simulated exercise intervention,there was a significant difference in the expression of PGC-1αprotein between aging electrical stimulation model and aging model.