The Effect Of Weight-bearing Treadmill Exercise And Low-intensity Pulsed Ultrasound On The Skeletal Muscle Of Ovariectomized Rats And Its Molecular Mechanism

Objective:For postmenopausal women,with the increase of their ages,the function of their ovaries degenerates and the secretion level of their endogenous estrogen decreases,which can lead to a series of metabolic diseases.Being an important component of the motor system,skeletal muscle cooperates closely with the nervous system and draws the attached bone during the contraction of the skeletal muscle,which is to produce movements and complete the daily actions of human’s body.The clinical diagnoses for postmenopausal women are accompanied with low quality and/or function of skeletal muscle.And it also manifests as a decrease of the number,function and muscle strength(especially explosive force)of skeletal muscles,which seriously affect daily life and activities.In addition,exercise has a good effect on improving muscle strength,so the influence of different exercise modes on muscle and its mechanism are still a hotspot in this research field.As one treatment,low-intensity pulsed ultrasound(LIPUS)is very popular due to its non-invasion,safety and convenience.Some studies have shown that LIPUS has a good effect on the treatment of muscle,tendon and soft tissue injury because it can promote the proliferation of muscle cells and protein synthesis.Exercise can inhibit the expression of negative muscle regulatory factor Myostatin(MSTN),and then promote skeletal muscle to be hypertrophic.It has been proved that MSTN is able to activate the key molecule Akt,thereby to activate the downstream molecule mTOR,to inhibit the molecules FoxO1 and finally to promote the protein synthesis.However,rare researches focus on the effects of the two different methods,i.e.,exercise and LIPUS on postmenopausal skeletal muscle,and the combined intervention of these two methods even less.Therefore,by means of the combined intervention of the weight-bearing treadmill exercise and LIPUS,this experiment makes a research on the effects of different intervention methods on gastrocnemius muscle in ovariectomized rats and its molecular mechanism from different research aspects including physiology,biochemistry,protein and gene.By doing so,the author attempts to find an effective adjuvant therapy for postmenopausal muscular atrophy.Methods:This experiment adopts the Sprague-Dawley(SD)74 female rats,3 month old,weight 220-240 g.fed 1 W after bilateral oophorectomy(OVX)to establish postmenopausal model.Wound healing for 2 W,were randomly divided into normal control group(group NC,18 rats),model group(OVX group,28)and exercise group(group EX,28 rats).The normal control group and model group fed normal weight-bearing exercise group treadmill exercise,termination of exercise intervention for 10 W.The model group(OVX group)and exercise group(EX group)were randomly selected each half were of LIPUS and weight treadmill exercise and LIPUS combined with intervention group(EX+LIPUS).The rats in the normal group and model group rats without any intervention,the intervention group of LIPUS and weight treadmill with low in LIPUS combined with intervention group(EX LIPUS)rats intervention the use of ultrasound in the treatment of treatment.For every day for 20 min weekly for 6 d for 3 W.In this experiment sonostat strength is 30 mW/cm2 and the duty ratio is 20%,the ultrasonic frequency is 1 MHz,pulse frequency of 100 Hz.morphology was observed in rats,and living habits of the rats weight and grip strength index change.By conventional biochemical reagent kit for each rat serum and muscle lactate dehydrogenase(LDH),muscle glycogen,serum inorganic phosphorus(P),creatine kinase(CK),serum calcium(Ca)detection and other biochemical indexes;by Western Blot and real-time fluorescence quantitative PCR(Quantitative Real-time PCR)from the protein level and gene level change detection of MSTN/Akt signal pathway of different signal molecules,observe the effect to improve to the gastrocnemius muscle of the rat ovary and its molecular mechanism.Results:1.the weight of ovariectomized rats was significantly higher than that of group NC(P<0.01),and the body weight of group EX was significantly lower than that of group OVX(P<0.01),and the weight of EX LIPUS group was decreased and the effect of improvement was the best(P<0.01).2.Ovariectomized rats grip and gastrocnemius muscle weight was significantly lower than that of group NC(P<0.01,P<0.01),intermittent weight training and LIPUS intervention can effectively increase the holding power of rats(P<0.01)and gastrocnemius muscle weight(P<0.01),the above results suggest that intermittent weight training and LIPUS intervention can really improve the rat skeletal muscle weight and increase muscle strength,and the two kinds of interventions combined than single intervention effect is obvious.3.Weight training and running LIPUS intervention,and NC group in comparison with the concentration of calcium in serum of rats,the OVX group,the EX and LIPUS group rats serum calcium concentration increases coexist with significant sex differences(P<0.01,P<0.05,P<0.05),EX + LIPUS group rats serum calcium levels but there is no significant difference,compared with OVX group of rats,the EX +LIPUS group rats serum calcium levels and there are significant differences(P<0.05);Compared with NC group of rats,the OVX group,the EX,LIPUS group and the EX +LIPUS group rats serum phosphorus levels,the OVX group and LIPUS group there are significant sex differences(P<0.01,P<0.05),compared with OVX group of rats,the EX and EX + LIPUS group rats serum phosphorus levels has extremely significant sex differences(P<0.01,P<0.01);And NC group rats creatine kinase(CK activity compared with OVX group,the EX,LIPUS group and the EX + LIPUS group rats CK elevated vitality has significant sex differences(P<0.01,P<0.01,P<0.01,P<0.05),compared with OVX group of rats,the EX and EX + LIPUS group CK activity in rats has significant correlation(P<0.05,P<0.01);Lactate dehydrogenase activity of LDH and NC group rats compared with OVX group,the EX,LIPUS group and the EX +LIPUS group activities of LDH coexist in rats have significant sex differences(P<0.01,P<0.01,P<0.01,P<0.01),compared with OVX group,the EX,LIPUS group and the EX + LIPUS group rats LDH activity has significant sex differences(P<0.01,P<0.05,P<0.05);Compared with NC group of muscle glycogen content in rats,the OVX group,the EX,LIPUS group and the EX + LIPUS group rats decreased muscle glycogen content has significant sex differences(P<0.01,P<0.01,P<0.01,P<0.05),compared with OVX group of rats,the EX and EX + LIPUS group rat muscle glycogen content increased coexist with significant sex differences(P<0.05,P<0.05),in which the EX + LIPUS group best increase in muscle glycogen content in rats.Different interventions on the concentration of serum calcium,phosphorus concentration,CK,LDH activity of muscle glycogen and muscle cell have certain improvement effect,improve the energy metabolism of muscle cells,and thus improve the skeletal muscle tissue.4.The expression of gastrocnemius-associated proteins and genes was detected by Western blot and RT-qPCR.The level of mRNA and protein expression of MSTN were significantly decreased by intermittent weight-bearing training,LIPUS intervention and combined intervention of both methods(P<0.01),and the expression of MSTN receptor ActR II B protein was significantly decreased(P<0.01),while intermittent weight-bearing training and low-intensity pulse were significantly decreased(P<0.01).The levels of mRNA and protein expression of Akt and its phosphorylated molecule P-Akt were significantly increased by impulse ultrasound intervention.The protein expression of FoxO1 and GSK-3 β in the ex LIPUS group was significantly lower than that in the OVX group,and the expression of mRNA and protein in the OVX group was significantly higher than that in the OVX group,and the expression of FoxO1 and GSK-3 β in the ex LIPUS group was significantly higher than that in the OVX group.Compared with exercise intervention and LIPUS therapy,The expression of related proteins was better than that of single ultrasound and exercise intervention in ovariectomized rats.Conclusion:1.Intermittent weight-bearing treadmill training,low-intensity pulsed ultrasound and two methods of combined intervention can effectively slow down the weight gain and improve the quality of skeletal muscle caused by ovariectomized estrogen secretion in rats.Promote the proliferation and differentiation of skeletal muscle cells,thereby increasing skeletal muscle strength and improving muscle atrophy symptoms.2.Intermittent weight-bearing treadmill training,low-intensity pulsed ultrasound intervention and combination of the two methods all improved the symptoms of muscular atrophy in ovariectomized rats,and promoted protein synthesis by enhancing the MSTN/Akt signaling pathway mediated and activated by Akt/FoxO1 pathway downstream of Akt.Improve muscle atrophy in ovariectomized rats.3.On the whole,the effect of intermittent weight-bearing training and combined intervention on improving muscle atrophy of ovariectomized rats was better than that of single low-intensity pulsed ultrasound intervention,and the combined intervention effect was better than that of single intervention.4.Exercise has a good auxiliary effect on improving muscle atrophy in postmenopausal female group,while LIPUS is an economical,safe,non-invasive and effective treatment for the incapacitated population.As an innovative model,intermittent weight-bearing training and low-intensity pulsed ultrasound intervention can also provide a new regulatory approach for clinical treatment of postmenopausal muscular atrophy.