| Objective:The rabbit model of contusion was interfered by massage at different time points,and the corresponding tissues were analyzed by light lens,swelling measurement,ELISA,WB and other detection methods.To observe the mechanism of massage in different time after skeletal muscle contusion,in order to explore the best intervention time of massage treatment skeletal muscle contusion.Methods:56 New Zealand male rabbits were purchased,and 40 rabbits were divided into a blank group(8 rabbits)and a modeling group(48 rabbits).After successful modeling,rabbits were random Ly divided into 6 groups according to the random number table method,with 8 rabbits in each group: massage group 30 min after injury,massage group 2hours after injury,massage group 24 hours after injury,massage group 48 hours after injury,ice control group and model group.A total of 7 groups were sampled on the 7th and 14 th day after injury.The rabbits in the modeling group used a hammer to align the mark with the free fall along the air pipe causing blow injuries.Hammer 500 g,height 50 cm,repeat 3times.20 minutes after the blow,the right lower limb swelling and no fracture were considered as successful modeling.Seven groups of rabbits were raised in a standard animal house environment.Massage intervention: the rabbit is fixed in the rabbit box,72 hours with kneading method according to kneading rabbit Weizhong point 1min,then use the push wipe method(to avoid the damage site,push to wipe),both operation pressure to slightly drive the skin is appropriate,frequency 40 times/ min,10 min /time,1 time / d;After 72 h,the rabbit Weizhong point was kneaded by kneading method for 1min,and then kneaded the injured site,10 min/ time,once /d;The intervention lasted for 7 days and 14 days.Ice treatment: Ice treatment was performed immediately after injury.Crushed ice was wrapped in a plastic bag and placed on the injured soft tissue for 5 min.Model group: No treatment was given after modeling.After 7 and 14 days of intervention,all the rabbits were killed by air embolization of the auricular vein,and bilateral gastronemius were stripped.The protein expression levels of 5-HT,IL-6,PEG2,TNF-α,Myo D and PAX 7 in rabbit skeletal muscle tissue were detected by ELISA,HE staining and Western Blot.The morphological changes of muscle fibers in rabbit skeletal muscle were observed by HE staining.Results:1.ELISA was used to detect the serum CK and PGE2 levels at 20 min after gastrocnemius injury,and the serum CK level at 20 min after injury was higher than that before injury(P < 0.01);The level of serum PGE2 in 20 min after injury was significantly higher than that before injury(P < 0.05).The ratio of muscle circumference of lower extremity before and after injury was measured.The right lower extremity of rabbits was significantly swollen 20 min after modeling,and the ratio was significantly higher than that before modeling(P < 0.0001).The results showed that the rabbit skeletal muscle blunt contusion model was successfully prepared.2.Changes in body weight: 7 days after injury,the body weight of rabbits in the model group decreased after modeling(P > 0.05).Other intervention groups did not show significant body mass loss after injury.The body weight of rabbits in each group increased steadily without significant difference.3.Change of muscle circumference: muscle circumference increased rapidly 20 min after injury,and all injury groups reached the peak muscle circumference 1 day after injury.The swelling degree in the 30 min group was lower than that in the model group 7 days after modeling(P < 0.05).The 48 hours group showed a slight increase in swelling on the second day after the first intervention.Massage intervention 30 min after injury can reduce the swelling of the injured site,and massage intervention 48 hours after injury for the first time will cause swelling aggravation the next day.4.Wet weight of gastrocnemius muscle: 7 days after injury,relative wet weight of gastrocnemius muscle in all groups was lower than that in model group(P < 0.05).On day14 after injury,the relative wet weight of all groups decreased compared with day 7.The wet weight of gastrocnemius muscle in 24 hours group,48 hours group and model group was lower than that in blank group(P < 0.05).This indicates that in the absence of intervention,natural recovery will produce certain muscular atrophy,and ice intervention will lead to muscle atrophy.5.ELISA results showed that the 5-HT concentration in the 30 min group was lower than that in the model group at 7 days after injury(P < 0.05).The level of PGE2 in model group was higher than that in other intervention groups(P < 0.05).The IL-6 level in the model group was higher than that in other intervention groups except 2 hours group(P <0.05),and the IL-6 level in the 24 hours group was lower than that in the ice group,indicating that massage and ice were beneficial to control the level of inflammation,and 30 min intervention significantly reduced the content of inflammatory factors,but 2 hours intervention was not effective in controlling the level of inflammation.The 5-HT concentration in 14 d,30 min and 24 hours after injury group was lower than that in model group(P < 0.05).Except for blank group,PGE2 level increased compared with that on day7.The level of IL-6 in model group was hoursigher than that in all intervention groups(P <0.05).This indicated that the pain caused by contusion was still obvious on day 14,and the concentration of inflammatory factors was still maintained at a relatively high level.6.HE staining results showed that different degrees of muscle fiber necrosis and inflammatory cell infiltration occurred in the injured group at 7 days,and the necrotic muscle fiber nucleolysis disappeared,among which,a small amount of blue and purple calcification masses were observed in the 30 min group,the 48 hours group and the ice group,while large areas of calcification masses were observed in the 2 hours group and the24 hours group.On day 14,the damage in the model group was aggravated with the increase of time.Most of the necrotic areas of muscle fibers in each group had been phagocytic and cleared by inflammatory cells,and some necrotic muscle fibers that could not be cleared by phagocytic had calcified.The 30 min manipulation group had the best recovery,with no calcification foci observed,while the 2 hours manipulation group and the ice group still had different degrees of calcification,and the recovery was not ideal.7.Western blot test results showed that the expression of Myo D protein in the 30 min,48 hours and ice groups was lower than that in the model group,with no statistical significance(P > 0.05).PAX 7 protein expression in ice group was significantly lower than that in other groups(P < 0.05).24 hours group was higher than model group,(P < 0.05).Myo D protein expression at 14 days tended to the level of blank group except for 24 hours group,and was generally lower than that at 7 days.Myo D protein expression in 24 hours group was higher than that in ice group and model group(P < 0.05).This indicates that skeletal muscle regeneration has been basically completed at 14 d,which may be due to inflammatory stimulation that leads to the massage intervention group except the 30 min group,especially the satellite cells in the 24 hours group are still proliferating,and the proliferating muscle satellite cells are not transformed into myoblasts in time to participate in the repair and regeneration of skeletal muscle.Conclusion:1.Massage is beneficial to reduce swelling,reduce atrophy,and reduce inflammatory response.2.Massage intervention 30 min after injury is beneficial to reduce the swelling of the injured site,effectively reduce inflammatory response,shorten the skeletal muscle repair and regeneration cycle,and promote skeletal muscle repair.3.After injury2 hours,48 hours of massage intervention,will delay the skeletal muscle repair time.4.Massage intervention 24 hours after injury will prolong the cycle of muscle satellite cell regeneration and delay differentiation,which is not conducive to skeletal muscle repair.5.Ice immediately after injury can reduce swelling at the injured site and reduce inflammation,but can lead to muscle atrophy. |
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