Mechanisms Of Skeletal Muscle Mitochondrial Fission Induced By A Heavy Load Eccentric Exercise And Acupuncture Intervention

Objective: To observe the changes of mitochondrial fission indicators in skeletal muscle after a heavy-load exercise in rats,and clarify the phenomenon of mitochondrial fission at different phase after exercise and its molecular mechanism.Exploring the effects of acupuncture intervention on mitochondrial fission in skeletal muscle after a heavy-load exercise.Method: The male SD rats were randomly grouped,experiment consisted of control group(C,n=8),acupuncture group(A,n=56),exercise group(E,n=56),exercise and acupuncture group(EA,n=56).E group and EA group were carried out a downhill eccentric exercise,and the others in A group and EA group were conducted acupuncture intervention.Except for C group,according to the phase points,each group was divided into 0h,12 h,24h,48 h and 72 h groups at the time of the intervention.The activity of mitochondrial function indicator of SQR and Cyt-c content of cytoplasm were determined by ELISA;fluorescein labeling technology was used to detecte the fluorescene ratio of JC-1;mitochondrial fragmentation in skeletal muscle was observed by transmission electron microscopy;the co-localization of TOMM20/DRP1 was detected by immunofluorescence;western blot was used to detecte the expression of FUNDC1 and DRP1,markers of fission in global cell,cytoplasm and mitochondria;the interaction between FUNDC1 and OPA1,CNX and DRP1 protein were detected by co-immunoprecipitation.The experimental data were anaylyzed by two-way ANOVA and one-way ANOVA.Results:(1)The mitochondrial SQR activity of skeletal muscle was decreased at various phase after a heavy-load exercise,mitochondrial fragment was observed under transmission electron microscope.The co-localization of TOMM20/DRP1 and the protein level of DRP1 in mitochondria increased significantly(P< 0.05),which reached the peak value at 12 h were significantly higher than C group(P<0.01).(2)There was no significant change in the expression of DRP1 in global cell after a heavy-load exercise(P>0.05);the change of DRP1 in cytoplasm was opposite to that in mitochondria,and there was no significant change of FUNDC1 in global cell observed(P> 0.05);the binding effect of FUNDC1 and OPA1,CNX,DRP1 appeared,and reached the peak at quiet state,2h and 12 h after exercise,respectively,and significantly higher than other phase(P< 0.05).(3)Acupuncture intervention was used after exercise,the content of Cyt-c in cytoplasm decreased significantly(P< 0.01);the fluorescence intensity of JC-1 increased significantly(P< 0.01);the fragment of mitochondria decreased and the co-localization of TOMM20/DRP1,the percentage of mitochondria DRP1 protein,the binding effect of FUNDC1/DRP1 was significantly decreased(P<0.05),and the mitochondrial SQR activity increased after exercise.Conclusion:(1)A heavy-load exercise could lead to abnormal mitochondrial distribution and structure in skeletal muscle,and mitochondrial function decreased,the phenomenon of mitochondria fission appeared,which reached the peak at 12 h after exercise and continued unitil 24 h,then gradually recovered.(2)The mechanism of mitochondrial fission in skeletal muscle induced by a heavy-load exercise in rats is that,FUNDC1/OPA1 binds to each other under normal conditions.After a heavy-load exercise,FUNDC1 and OPA1 began to separate,and then bound to CNX,and then the binding capacity decreased.FUNDC1 gradually bound to DRP1 and aggregated DRP1 to mitochondria,leading to fission.(3)Acupuncture after exercise significantly reduced the damage of mitochondria,mitigated the damage of mitochondria caused by a heavy-load exercise,and inhibited the mitochondrial fission through FUNDC1,which may contribute to the recovery of mitochondrial function to some extent.