| Objective:Massage therapy has effects of improving metabolism and reducing turbidity and generating muscle.The T2DM rat model and T2DM patients were treated with massage intervention.And from the massage therapy to improve skeletal muscle glucose metabolism,regulate skeletal muscle autophagy,promote myoblastic differentiation of muscle satellite cells and other aspects to explore the effect of massage to reduce turbidities on the improvement of limb muscle in type 2 diabetes patients with sarcopenia and its possible biological mechanism.The T2DM rat model and T2DM patients were treated with massage intervention.And from the massage therapy to improve skeletal muscle glucose metabolism,regulate skeletal muscle autophagy,promote myoblastic differentiation of muscle satellite cells and other aspects to explore the effect of massage to reduce turbidities on the improvement of limb muscle in obese type 2 diabetes patients with sarcopenia and its possible biological mechanism.It also provides high-quality experimental evidence for the clinical application of massage therapy in T2DM sarcopenia and other related skeletal muscle mass loss and functional decline diseases.Methods:（1）Effect of massage on skeletal muscle mass and function in patients with T2DM.72T2DM patients were randomly divided into treatment group and control group.The control group received daily blood glucose monitoring and oral metformin hydrochloride tablets to control blood glucosethe massage group received blood glucose monitoring and oral metformin hydrochloride tablets combined with Sheng Ji Jiang Zhuo massage therapy,0min/time,6 times/week,a total of 8 weeks of intervention.The control group received daily blood glucose monitoring and oral metformin hydrochloride tablets to control blood glucose;the massage group received blood glucose monitoring and oral metformin hydrochloride tablets combined with Shengji turbidity reducing massage therapy,40min/time,6 times/week,a total of 8 weeks of intervention.（2）The effect of massage on muscle strength and motor function of skeletal muscle in T2DM rats.40 Wistar male rats were randomly divided into blank group,model group,metformin group and massage group,with 10 rats in each group.Except blank group,other groups were fed with high-fat diet plus STZ injection to establish T2DM animal model.The massage group began to use the experimental animal massage instrument to massage the limbs of rats on the 1stday after modeling,once a day,6 times a course,with a rest of 1 day between courses,a total of 8 courses of treatment.Rats in metformin group were administrated with metformin aqueous solution by gavage(250mg·kg-1·d-1)on the 1stafter modeling.Blank group and model group were observed without any intervention.And conducted the following studies.The rats in each group were given the running exhaustion test,the test of rat grip strength and the general observation before and after treatment.After treatment,the lower limb gastrocnemius muscles of rats were taken for wet weight weighing.The improvement of skeletal muscle function in different groups of rats at different time points and the difference of skeletal muscle mass after intervention were observed,and the effects of massage on skeletal muscle function and treatment duration in obese T2DM rats were explored.（3）Effects of massage on skeletal muscle ultrastructure in T2DM rats.After the intervention,HE staining,ATP-ase staining and transmission electron microscopy were used to observe the changes of muscle morphology,muscle fiber type proportion and ultrastructure of the gastrocnemius of each group.（4）Effects of massage on blood glucose and myosatellite cell myogenic differentiation in T2DM rats.After the intervention,insulin sensitivity index,oral glucose tolerance,changes of muscle and liver glycogen,protein and m RNA expression levels of myogenic adipogenic differentiation factors Pax7,Myo D,Myo G,Myf5 and Myostatin were detected by rat glucose metabolism detection,Western Blot and RT-PCR.To investigate the effect of massage on blood glucose and myosatellite cell myogenesis differentiation in T2DM rats.（5）Effects of massage on skeletal muscle mitochondria autophagy in T2DM rats.After the intervention,the protein expression m RNA expression levels of LC3,P62,PINK1,PRKN and PGC-1αwere detected by Western blot and RT-PCR,so as to explore how massage can promote the recovery of skeletal muscle function by improving skeletal muscle autophagy.Results:（1）There was no significant difference between the control group and the treatment group in gender,age,course of disease and other baseline data.Comparison between the two groups showed no significant difference in FBG,FINS,Hb A1c,HOMA-IR,SMI grip strength（IPAQ score and daily walking speed）before treatment,FBG and Hb A1c in the treatment group were significantly lower than those in the control group after treatment（P<0.05）,FINS and HOMA-IR were significantly lower than those in control group（P<0.01）,the SMI grip strength IPAQ and daily walking speed in the treatment group were significantly higher than those in the control group（P<0.01）;Intragroup comparison showed that there were significant differences in FBG,FINS,Hb A1c,HOMA-IR,SMI grip strength,IPAQ score and daily walking speed between the two groups before and after treatment（P<0.05）.（2）The running exhaustion time,grasping power and wet weight of gastrocnemius in model group,metformin group and massage group were significantly lower than those in control group on day 0（P<0.01）,indicating that the model of sarcopenia in T2DM rats was successful.Compared with the model group,the exhaustion time of running and the grasping power of rats began to increase significantly in the massage group on the 28th day（P<0.01）,and the wet weight of gastrocnemius muscle of rats was significantly increased after treatment（P<0.01）,while the exhaustion time of running,the grasping power of rats and the wet weight of gastrocnemius muscle after treatment did not increase significantly in the metformin group（P>0.05）.（3）After 56d of treatment,HE staining results of the gastrocnemius muscle in each group showed that,compared with the blank group,the model group,metformin group and massage group had disordered skeletal muscle tissue arrangement,muscle cell atrophy and edema,inflammatory cell infiltration was more obvious,muscle fiber autolysis was a small amount,fiber bundle membrane was slightly broken,and a small number of lymphocytes were infiltrated,indicating more obvious muscle tissue injury.Compared with the model group,the massage group skeletal muscle cells arranged neatly,visible local focal myocyte atrophy,visible myocyte edema,but the degree is light,small area,accompanied by a small number of inflammatory cell infiltration,occasionally seen myocyte fracture,necrosis.The results of ATP-ase staining showed that compared with the blank group,the two types of muscle fibers in the model group and the metformin group were mainly light-colored fast-twitch type II fibers,and the proportion of type II muscle fibers was significantly increased.Compared with the model group,the proportion of slow-twitch type I fibers in the massage group was significantly increased.The results of transmission electron microscopy showed that compared with the blank group,the muscle fibers in the model group were seriously damaged,and the mitochondria and sarcoplasmic reticulum were swollen and dissolved.Compared with the model group,the ultrastructure of skeletal muscle in the metformin group was not significantly improved,but that in the massage group was superior to the model group.（4）The results of glucose and lipid metabolism of rats showed that from 14 weeks after intervention to the end of intervention,fasting blood glucose of rats in metformin group and massage group was significantly decreased compared with model group（P<0.01）.From 14 days after intervention to the end of intervention,the oral insulin level of rats in metformin group and massage group was significantly decreased compared with model group（P<0.01）.After intervention,the OGTT level of rats in model group,metformin group and massage group was significantly increased compared with blank group（P<0.01）,while the OGTT level of rats in metformin group and massage group was significantly decreased compared with model group（P<0.01）.After intervention,the insulin sensitivity level of rats in model group and massage group was significantly increased compared with blank group（P<0.01）,and the insulin sensitivity level of rats in metformin group and massage group was significantly decreased compared with model group（P<0.01）.Western Blot and RT-PCR assay results showed that compared with blank group,protein and m RNA expressions of Pax7,Myo D,Myo G,Myf5 in model group were significantly decreased（P<0.01）,the expression of MSTN was significantly increased（P<0.05）Compared with model group,the protein and m RNA expressions of Pax7,Myo D,Myo G,Myf5 in massage group were significantly increased（P<0.05）,there was no significant difference between metformin groups.（5）The results of Western Blot and RT-PCR detection of autophagy related factors in gastrocnemius of rats showed that the protein and m RNA expressions of LC3,PRKN and PGC-1αwere significantly decreased in model group after intervention compared with blank group（P<0.01）,while the expressions of P62 and PINK1 were significantly increased in model group（P<0.05）.Compared with model group,the protein and m RNA expressions of LC3,PRKN and PGC-1αin massage group were significantly increased（P<0.05）,while the expressions of P62 and PINK1 were significantly decreased（P<0.05）,and there were no significant changes in drug group.Conclusion:1.Massage combined with oral metformin can more effectively maintain the stability of blood glucose in patients with T2DM,and significantly improve limb movement ability,skeletal muscle strength,skeletal muscle content and patients’daily living ability.2.Massage can improve a series of behavioral changes of limb muscle function in T2DM rats,and can significantly improve limb skeletal muscle mass,with obvious musculogenesis effect.3.Massage can improve the type and distribution of muscle fibers in skeletal muscle of T2DM rats,and reduce the lipid differentiation of skeletal muscle.4.Massage can promote myogenic differentiation,increase the content of skeletal muscle,regulate the decomposition and conversion of blood glucose by skeletal muscle,and improve the function of skeletal muscle by regulating the expression of muscle myogenic factors in the limbs of T2DM rats.5.Massage can promote autophagy of skeletal muscle in T2DM rats,thereby improving energy metabolism of skeletal muscle and improving skeletal muscle function. |
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