Effects Of One-time High-load Eccentric Exercise On Rats’ Skeletal Muscle Mitochondria-associated Endoplasmic Reticulum Membrane(MAM)

Objective:Explore the relationship between mitochondrial-endoplasmic reticulum coupling changes and skeletal muscle ultrastructure injury caused by a heavy-load eccentric exercise by detecting the changes of mitochondrial-endoplasmic reticulum coupling degree,mitochondrial calcium concentration and MAM tether protein Mfn2,VAPB and PTPIP51 protein expression.Methods:48 healthy male SD rats aged 8 weeks were randomly divided into a control group（C）and an exercise group（E）.According to the different phases after exercise,they are divided into the immediate group after exercise（E0）,the 12h group after exercise（E12）,the 24h group（E24）,the 48h group（E48）,and the 72h group（E72）.8animals in each group.The Armstrong centrifugal exercise model was selected for one-time exercise intervention.The slope of the treadmill was set to-16°and the training time was 90 minutes.After that,we used transmission electron microscopes to observe the change of MAM quantity per unit area,used dual immunofluorescence labeling method to detect the Co-localization of TOMM20 and RyR1.Besides,Fluorescence Probe Method was used to detect the change of[Ca²⁺]_mt.The expression of MAM tether protein Mfn2、VAPB and PTPIP51was detected by Western Blot method.One-way analysis of variance was performed on the data results using SPSS20 software.When the variance is uniform,the LSD method is used for multiple comparisons between groups,and when the variance is uneven,the Tamhanes method is used.Results:（1）After one-time high-load centrifugal exercise,the number of MAM in the skeletal muscle unit area first increased and then decreased,and the maximum value appeared at 12 hours after exercise（p<0.05）.The co-localization coefficient of TOMM20 and RyR1 increased and then decreased after a one-time centrifugal exercise,and the maximum value appeared at 24 hours after exercise（p<0.01）;（2）After one-time high-load centrifugal exercise,[Ca²⁺]_mtincreased at first and then gradually decreased,and the maximum value appeared at 12 hours after exercise（p<0.01）;（3）The expression of MAM tether protein Mfn2 increased immediately after one-time centrifugal exercise and then decreased,and then increased again at 72hours,There was no significant difference between the groups（p>0.05）;The expressions of VAPB and PTPIP51 protein increased significantly after exercise,generally speaking,they increased at first and then decreased,and the maximum value appeared at 24 hours after exercise（p<0.05）.Conclusions:（1）The increased degree of mitochondrial endoplasmic reticulum coupling after one-time high-load centrifugal exercise leads to increased calcium transport from the endoplasmic reticulum to the mitochondria;（2）The expression of MAM tether protein VAPB and PTPIP51 in skeletal muscle increased after one-time high-load centrifugal exercise.VAPB and PTPIP51 can promote mitochondrial endoplasmic reticulum coupling.The effect of Mfn2 on mitochondrial endoplasmic reticulum coupling is still unclear.