The Effect And Mechanism Of Exercise And Etanercept On Moderate Brain Injury In Mice

Traumatic brain injury(TBI)is the main cause of death and disability in children and adolescents.Despite preventive measures,the incidence of TBI associated with motor vehicle accidents,falls,assaults,and various intense sports is still high,and it poses a serious threat to public health.Children,adolescents and TBI patients are often accompanied by emotional,behavioral,adaptive and cognitive dysfunction during their growth and adulthood.At present,the central mechanism for emotional and behavioral disorders is still not fully understood.In this study,randomized exercise and intraperitoneal injection of TNF-a antagonist Etanercept intervention before and after trauma were used to explore the effects of these two interventions on peripheral inflammatory cytokines and damage of central cortex and hippocampal neurons after moderate TBI in mice.At the same time,it examines its effect on behavior and emotions after TBI.The aim is to provide a new idea for the prevention and treatment of TBI sequelae,so as to provide a theoretical basis for the prevention and treatment of brain injury.Research methods:1.Wild-type male C57BL/6 mice(30-week-old,30 animals,body weight 9-11g)were randomly divided into Sham group(Sham,n=6),brain trauma injury group(TBI,n=8),and brain trauma injury + etanercept group(TBI + ETA,n = 8),rolling cage exercise + brain trauma injury group(RCE + TBI,n = 8).The mice in the RCE + TBI group were preliminarily subjected to cage training for 4 weeks before brain injury.C57 mice in the 4 weeks of free cage movement,their active time is 7pm-12pm daily,the running distance is mostly completed during this period.The peak activity period is between 7pm-9pm and the average running speed is about 12m/min[1,2].Therefore,we opted for a period of 4 weeks,at 7pm-9pm each day,to allow the exercise group to undergo a 40-minute cage training.This amount of exercise is equivalent to human moderate-intensity aerobic exercise.A hydraulic strike device was used to hit the right motor cortex,resulting in an animal model of moderate brain injury(strike strength of 2.4-2.8 atm,strike angle of 17 degrees).In the TBI + ETA group,the TNF-? blocker ischemic(20 mg/kg body weight)was injected intraperitoneally immediately after TBI,and was injected once more on the first day after the wound(ie,24 hours after the first injection of esablex).After 48 hours of TBI,serum inflammatory cytokines were detected by ELISA kit,and the expression of brain-related proteins was detected by Western Blot,2.Wild-type male C57BL/6 mice(30-week-old,30,body weight 9-11g)were randomly divided into Sham group(Sham,n=6),brain trauma injury group(TBI,n=8),and brain trauma injury + etanercept group(TBI + ETA,n = 8),rolling cage exercise + brain trauma injury group+ motor skills training(RCE + TBI + MST,n =8).The mice in the RCE + TBI + MST group were preliminarily subjected to four-week cage training(40 min/day)prior to brain injury.A hydraulic strike device was used to hit the right motor cortex,resulting in an animal brain injury model(strike strength of 2.4-2.8 atm,attack angle of 17 degrees).The trauma-administered group and the sports-trauma-administered group were intraperitoneally injected with the TNF-? blocker isepram(20 mg/kg body weight)immediately after TBI,and each was injected on the first to sixth days after the wound.After TBI,the first ethological test on days 1-6.After 7 to 20 days after TBI,the RCE + TBI + MST group had skill training.The second behavioral test and histological examination at days 21-26 after TBI.Result:1.Weight testFrom the 2nd week of exercise training,the weight of mice in the RCE+TBI group was smaller than that of the control group(week 2,p?0.05;week 3,p?0.01;week 4,p<0.05).2.ELISA detection of serum inflammatory factorsAfter brain injury,the TNF-a(p?0.01),IL-1?(p?0.05),IL-6(p?0.01),and IL-4(p?0.01)concentrations of the inflammatory cytokines in the TBI group correlated with the Sham group.The ratio is significantly higher.Exercise(elevated levels of anti-inflammatory cytokines IL-4 and IL-6,p?0.01)and elisa(decreased pro-inflammatory cytokines TNF-a,p<0.01;anti-inflammatory cytokines IL-4 and IL-6 concentrations Elevated(p?0.01)improved the expression of inflammatory factors after TBI.3.Western Blot detection of brain tissue protein expressionAfter brain injury,hippocampal GFAP expression was up-regulated in the TBI group(p?0.01),and GFAP and LC3-? expression in the cerebral cortex and hippocampus was also upregulated(p?0.01).Yisaipu up-regulated the expression of GFAP in cerebral cortex(p?0.05)and hippocampus(p?0.01)after TBI,and down-regulated the expression of LC3-? in cerebral cortex and hippocampus after TBI(p?0.01).Exercise increased the expression of GFAP in the hippocampus after TBI(p?0.01)and down-regulated the expression of LC3-? in the cerebral cortex and.hippocampus after TBI(p?0.01).4.Behavior testing(1)Neurological score tests found that after trauma,the neurological scores of the TBI group increased(DPI1,DPI1,p?0.05;DPI14,DPI21,p?0.01).In the DPI21 test,exercise(p?0.05)and epleprotype(p?0.01)both reversed the increase in neurological score after TBI.(2)In the crossbar test,after the two tests of trauma,the crossbar time in the TBI group increased(p?0.01).In the first test,exercise suppressed the increase in crossbar time after TBI(p?0.05).In the second test,both exercise and Yisaipu were able to reverse the increase in crossbar time after TBI(p?0.05).(3)In the rotarod test,the test results of the two speeds are basically the same.TBI shortens the mouse's stay on the rotarod.The first test results showed that exercise can increase the residence time of mice after TBI(20 rpm/60 seconds,p?0.05).The second test showed that exercise(20 rev/60 sec,p?0.01;40 rev/60 s,p?0.05)and escites(p?0.05)both increased the mice's residence time.(4)In the open field test,the percentage of total distance in the middle quarter of the open field after TBI was significantly decreased in the TBI group(DPI3,DPI23,p?0.01).Both exercise and etanercept were able to increase the percentage of post-traumatic mouse activity in the middle quarter of the open field(DPI3,DPI23,p?0.01).(5)In the tail-tail test,the time of immobility of the mice in the TBI group was prolonged(DPI4,DPI24,p?0.05).In the first test,the effects of exercise and etanercept were not significant(p?0.05).In the second test,exercise training reduced the immobility time of post-traumatic mice.(6)Y-maze test is divided into two experiments,TBI will cause the mouse's automatic rotation percentage and resolution coefficient decreased(p?0.01).In the auto-rotation percentage test,etanercept and exercise training had significant effects at DPI5 and DPI25,respectively(p?0.05).In the resolution coefficient test,etanercept(p?0.05)and exercise training(p<0.01)could increase the resolution of the mice after DPI26,but the effect of DPI6 was not significant(p?0.05).5.Histological detection(1)Injury measured the number of neurons in cerebral cortex TBI group?TBI+ETA group?RCE+TBI+MST group?Sham group,and the arrangement of cortical neurons in TBI group was disordered,vacuoles were severely vacuolated,and cortical neurons in Sham group.The order of the neurons was clear and the nucleolus were clear.The neuronal forms of the two groups were between the two.(2)Observing the morphology of hippocampal cells under 200 magnifications,we found that the TBI group's lesions measured the glycogen in the DG region of the hippocampus as cell body pyknosis,and the neurons became irregular lumps and could not distinguish the nucleoli.There were fewer pyknosis neurons in Sham group,TBI+ETA group and RCE+TBI+MST group.Conclusion:The improvement of moderate brain injury in mice by exercise training and etanercept is to improve the immune response and autophagic processes in the brain by reducing the level of inflammatory cytokines in the peripheral blood circulation,thereby altering the traumatic cerebral cortex and hippocampal dentition.The morphology and number of back neurons ultimately improve the behavior and mood of mice.However,the rehabilitation effects of exercise and escite on brain injury are related to the training time and the total dose of the injection.Therefore,further studies are needed to determine the optimal time and dose.