The Effects Of Exercise Pre-Conditioning On Muscle Atrophy And Oxidative Stress Induced By DEX Injection In Mice

Background: Glucocorticoids are widely used clinically because of their antiinflammatory and immunosuppressive properties.However,it has strong side effects that cause significant physical and life impairments for patients,such as osteoporosis,hyperglycemia,insulin resistance,fat deposition disorders,high blood pressure and skeletal muscle atrophy.The mechanism of glucocorticoid-induced skeletal muscle atrophy is still under study.Exercise has the effect of improving skeletal muscle atrophy,but the effect of exercise preconditioning on dexamethasone-induced muscle atrophy is not yet fully understood.Recent studies have found that high-intensity interval exercise(HIIT)might exert a better metabolic effect than traditional aerobic exercise,but it is not clear whether HIIT has a better preventive effect on muscle atrophy caused by DEX.Swimming is a relatively mild and practical type of exercise intervention,especially for patients with muscle atrophy.Therefore,swimming pre-conditioning intervention methods were used in the present study to explore the effects of moderateintensity swimming and high-intensity intermittent swimming on DEX-induced skeletal muscle atrophy in mice,as well as its molecular mechanism.Objective: This study aims to explore the effects of exercise pre-conditioning on DEX-induced skeletal muscle atrophy in mice,and reveal the mechanism of exercise prevention of skeletal muscle atrophy from the perspectives of protein synthesis signaling pathways and protein breakdown signaling pathways,as well as the role of oxidative stress in it.It also compares the prevention effects of moderate-intensity swimming and high-intensity intermittent swimming on DEX-induced skeletal muscle atrophy,and provides a new idea for exercise pre-conditioning to prevent skeletal muscle atrophy.Methods: Forty 6-week-old C57BL/6 male mice were purchased from Animal Research Center of East China Normal University.All mice were kept in an SPF-grade animal breeding room with an indoor temperature of 22 ± 2 ℃,an air humidity of 50 ±10%,and a 12-hour day-night cycle.The mice were fed standard feed and freely ingested food and water.All mice were randomly divided into 4 groups after 1 week of adaptive feeding.The groups are as follows:(1)Group C: SED+CON(n=10);(2)Group D: SED+DEX(n=10);(3)Group M: MICT+DEX(n=10);(4)Group H:HIIT+DEX(n=10).Among them,the mice in group C and D maintained a quiet lifestyle,group M performed moderate-intensity swimming for 8 weeks,and group H performed high-intensity intermittent swimming for 8 weeks,training 5 days a week.After 8 weeks of exercise pre-conditioning,group C received intraperitoneal injection of normal saline(0.5 mg/kg BW)for 10 consecutive days;group D,M,and H received intraperitoneal injection of DEX(0.5 mg/kg BW)daily for 10 consecutive days.During this period,group M and group H continued the original exercise intervention.During the whole exercise intervention and DEX injection,the mice’s body weight and fasting blood glucose were measured regularly every week.The mice in each group were tested for insulin tolerance the day before being sacrificed.In terms of morphology,hematoxylin-eosin experiment technique was used to stain mouse gastrocnemius muscle,and quantitatively analyze the cross-sectional area of gastrocnemius muscle.In terms of serum indicators,total cholesterol,triglycerides,glucose,and insulin in the serum were measured.Use the kit to detect oxidative stress indicators of skeletal muscle,such as SOD,MDA,CAT,GSH-PX,T-AOC.Real-time PCR was used to detect the mRNA level of FOXO3 a,AKT,mTOR,p70S6 k,S6,Atrogin-1,MURF1,MYOD,MYOG,MSTN in skeletal muscle.Western Blotting was used to detect the protein level of FOXO3 a,p-FOXO3 a,AKT,p-AKT,mTOR,p-mTOR,p70S6 k,p-p70S6 k,S6,p-S6,Atrogin-1,MURF1 in skeletal muscle.Results:(1)After 8 weeks of exercise preconditioning,the body weight of mice in group C,group D,group M,and group H increased steadily over time.Compared with group C,the body weight of mice in group D decreased significantly(p<0.05)at the 9th and 10 th week,while the body weight of mice in group M and group H began to decrease.(2)Compared with group C,the fasting blood glucose level of group D at the 9th and10 th week were significantly higher(p<0.05).The fasting blood glucose level of mice in group M and H were basically stable at 6-8mmol/L within 10 weeks.(3)Compared with group C,the area under the curve of IPITT in group D and group H were significantly increased(p<0.01);compared with group D,the area under the curve of IPITT in group M(p<0.01)and group H(p<0.05)were significantly decreased.Compared with group C,the levels of mice glucose in group D(p<0.01),glucose in group M(p<0.01),insulin in group D(p<0.01),and insulin in group M(p<0.05)were significantly increased;compared with group D,the levels of mice glucose in group M(p<0.01),glucose in group H(p<0.01),insulin in group M(p<0.01),insulin in group H(p<0.01)were significantly decreased.(4)Compared with group C,the levels of triglyceride and total cholesterol in group D were significantly increased(p<0.01);compared with group D,the levels of triglyceride and total cholesterol in group M and H were significantly decreased(p<0.01).The iwat mass of mice in group D and M were significantly higher than that in group C(p<0.05);the i-wat index of mice in group D,M,and H were significantly higher than that in group C(p<0.05).The e-wat mass of mice in group D was significantly higher than that of group C(p<0.01);the e-wat mass of mice in group M and H were significantly lower than that in group D(p<0.01).The e-wat index of mice in group D were significantly higher than that of group C(p<0.05);the e-wat index of mice in group H was significantly lower than that of group D(p<0.01).(5)Compared with group C,MDA of gastrocnemius in group D was significantly increased(p<0.05);compared with group D,MDA of gastrocnemius of mice in group M and H were significantly decreased(p<0.05).Compared with group C,SOD(p<0.05)and GSH-PX(p<0.05)of gastrocnemius in group D were decreased significantly;compared with group D,SOD(p<0.01),GSH-PX(p<0.05)and T-AOC(p<0.05)of gastrocnemius in group M were increased significantly,and SOD of gastrocnemius muscle in group H was increased significantly(p<0.05).(6)Compared with group C,the mRNA level of AKT in gastrocnemius of mice in group D,group M and group H were increased significantly(p<0.01).Compared with group C,the mRNA level of mTOR in gastrocnemius of mice in group D was significantly decreased(p<0.05).Compared with group D,the mRNA level of mTOR in gastrocnemius of mice in group M was significantly increased(p<0.05).Compared with group C,the mRNA level of p70S6 k in gastrocnemius of mice in group D and group H were significantly decreased(p<0.05).Compared with group D,the mRNA level of S6 in gastrocnemius of mice in group M was significantly increased(p<0.05).Compared with group C,the mRNA level of FOXO3a(p<0.05),Atrogin-1(p<0.05),and MURF1(p<0.01)in gastrocnemius of mice in group D was significantly increased;compared with group D,the mRNA level of FOXO3a(p<0.05),Atrogin-1(p<0.05),and MURF1(p<0.05)in gastrocnemius of mice in group M and group H were significantly decreased.Compared with group C,the mRNA level of MSTN in gastrocnemius of mice in group D was significantly increased(p<0.05).Compared with group D,the mRNA level of MSTN in gastrocnemius of mice in group H and the mRNA level of MYOD in gastrocnemius of mice in group H were significantly decreased(p<0.05).(7)Compared with group C,the wet weight and index of gastrocnemius in group D(p<0.01)and the wet weight and index of tibial anterior in group D(p<0.05)were significantly decreased;compared with group D,the wet weight and index of gastrocnemius in group M(p<0.01),the wet weight and index of gastrocnemius in group H(p<0.01),the wet weight and index of tibialis anterior in group M(p<0.01),the wet weight of tibialis anterior in group H(p<0.01)and the index of the tibialis anterior in group H(p<0.05)were increased significantly.Compared with group C,the cross-sectional area of gastrocnemius muscle fibers in group D was significantly decreased(p<0.05);compared with group D,the cross-sectional area of gastrocnemius muscle fibers in group M and group H were significantly increased(p<0.01).Compared with group C,the phosphorylated protein level of FOXO3 a in group D was increased(p<0.05);compared with group D,the phosphorylated protein level of FOXO3 a in group H was decreased significantly(p<0.05).Compared with group C,the protein level of Atrogin-1 in group D was increased(p<0.05);compared with group D,the protein level of Atrogin-1 in group M and group H were significantly decreased(p<0.05).Conclusion:(1)The 8-week medium-intensity swimming pre-conditioning and the 8-week highintensity intermittent swimming pre-conditioning can effectively prevent DEX-induced oxidative stress in skeletal muscle of mice,and improve the symptoms of skeletal muscle atrophy.The 8-week moderate-intensity swimming pre-conditioning has a better preventive effect on skeletal muscle oxidative stress.(2)The 8-week medium-intensity swimming pre-conditioning and the 8-week highintensity intermittent swimming pre-conditioning may prevent DEX-induced skeletal muscle atrophy in mice through the FOXO3a/Atrogin-1/MURF1 signaling pathway.There was no significant difference in the prevention of skeletal muscle atrophy between 8-week medium-intensity swimming pre-conditioning and 8-week highintensity intermittent swimming pre-conditioning.