| Objective:Post-traumatic arthritis(PTOA)is a chronic degenerative joint disease involving articular cartilage and subchondral bone.Abnormal mechanical stress is often believed to be one of the main reason for PTOA.Abnormal mechanical stress may be the common inducement.Abnormal intra-articular mechanical stress can be caused by excessive weight gain,excessive exercise,and excessive external load.However,the specific pathogenesis of PTOA induced by abnormal mechanical stress is still unclear.Studies have found that the abnormal expression of miR-214 can cause abnormal growth regulation of bone and cartilage,and then lead to the occurrence of OA.It is speculated that miR-214 may mediate the process of cartilage degeneration in OA.Therefore,this study attempts to establish mice PTOA model through high-intensity treadmill exercise,and then carry out moderate-intensity treadmill exercise on PTOA mice for rehabilitation,to explore the relationship between miR-214 expression and PTOA cartilage damage and degeneration,to provide experimental basis for further exploration of exercise intervention in the prevention and treatment of PTOA and miR-214 related PTOA therapeutic targets.Methods:Forty male C57BL/6 mice aged 20 weeks were randomly divided into model control group(CM group,n=8),model group(M group,n=8),control group(CK group,n=8),model rehabilitation group(MK group,n=8)and model resting group(MJ group,n＝8).The CM group and CK group were fed routinely without any exercise intervention.In group M,MK,and MJ,a week of pre-adapted treadmill exercise with a speed of 10m/min-18m/min was performed,and then a week of high-intensity treadmill exercise with a speed of 20m/min was performed.At the age of 25 weeks,the mice in CM group and M group were sacrificed and sampling.After 1 week of static rehabilitation,MK group and MJ Group were given moderate-intensity treadmill exercise at a speed of 8 m/min for 4 weeks and static feeding for 4 weeks respectively.At the age of 30 weeks,the mice in CK group,MK group,and MJ Group were sacrificed and sampling.HE staining and safranin O staining were used to observe the pathological changes of knee cartilage;Micro-CT was used to detect the changes of subchondral bone;RT-qPCR was used to detect the mRNA levels of inflammatory factors,anabolic and catabolic factors,and miR-214 related target genes.All the experimental results were expressed as mean±standard deviation(X±SD).SPSS 20.0 software was used for data analysis and processing.Independent sample t-test and one-way ANOVA were used for pairwise comparison,and LSD method was used for post test.P<0.05 indicated that the results were statistically different.Results:(1)Effect of exercise on the weight of mice:During the conventional feeding period,the weight of mice in CM group and CK group showed an overall growth trend,but there was no significant difference in the weight change between weeks.The weight of mice in M group decreased during the period of high-intensity treadmill modeling,which was significantly lower than that of mice in CM group at 24 weeks of age(P<0.05).The weight of mice in MK and MJ groups decreased during high-intensity treadmill exercise(20-24w).After the end of high-intensity treadmill intervention,the weight of mice in MK group continued to decrease during 26-29 weeks of moderate-intensity treadmill exercise,but the weight of mice in MJ group increased after 26-29 weeks of rest.There was a significant difference between the two groups at 29 weeks(P<0.05).(2)Effect of exercise on histomorphology of mice1)The effect of exercise on the histomorphology of articular cartilage:HE staining showed that there were micro cracks on the surface of knee cartilage in M group,which was more disordered than that in CM group.The results of safranin o-green staining showed that the cartilage loss and ossification of the knee joint in M group were more obvious than those in CM group.There were significant differences in Mankin’s score and OARSI score between the two groups.The score of M group was significantly higher than that of CM group(P<0.05).After four weeks of moderate-intensity treadmill exercise,HE staining and safranin o-green staining of knee cartilage in MK group showed that cartilage loss decreased and cells gradually returned to normal arrangement,which partially reversed the pathological changes of knee joint OA in M group.The staining of MJ Group was similar to that of MK group,but the Mankin’s and OARSI scores of MK group were significantly lower than those of MJ Group(P<0.05).2)The effect of exercise on the morphological changes of subchondral bone:Micro-CT results of mice knee joint showed that BV/TV of subchondral bone of femoral condyle in M group was significantly higher than that in CM group after high-intensity treadmill exercise(P<0.05).After 4 weeks of moderate-intensity treadmill rehabilitation training,BMD of subchondral bone of femoral condyle in MK group was significantly higher than that in CK group(P<0.01).The results showed that the BMD,TB.N,BV/TV of MK group were significantly higher than those of MJ group,while the BMD,TB.N,BV/TV of MK group were significantly higher than those of MJ Group.Moreover,Tb.Sp of MK group was significantly less than MJ group(P<0.05).(3)Effects of exercise on gene expression of knee cartilage in mice1)Effects of exercise on cartilage metabolism and inflammatory expression of knee joint in miceAfter high-intensity treadmill exercise,compared with CM group,the expressions of MMP-13 and TNF-α were significantly increased in M group,while the expressions of COL2A and ACAN were significantly decreased.After 4 weeks of moderate intensity treadmill training,the expression of TNF-α and MMP-13 in articular cartilage of MK group was significantly lower than that of and MJ Group,and the expression of ACAN and COL2A was significantly increased(P<0.05).2)Effects of exercise on expression of miR-214 and its related target genes in mouse knee cartilageAfter high-intensity treadmill exercise,the expression of miR-214 in M group was significantly lower than that in CM group,while the expression of Wnt and β-catenin was significantly increased(P<0.05).After 4 weeks of moderate intensity treadmill training,the expression of miR-214 increased in MK group,which was significantly higher than that in MJ Group,while the expression of β-catenin decreased,which was significantly lower than that in MJ group.increased(P<0.05).At the same time,the expression of miR-214 in MK group was also higher than that in MJ Group,while the expression of Wnt and β-catenin were decreased and lower than that in MJ Group.Conclusion:Four weeks of high-intensity treadmill exercise can induce PTOA in C57BL/6 mice,and reduce the expression of miR-214 meanwhile increase the expression of Wnt andβ-Catenin in articular cartilage;moderate-intensity treadmill rehabilitation training can alleviate the exercise-induced OA cartilage injury,increase the expression of miR-214 and decrease the expression of Wnt and β-catenin in articular cartilage.MiR-214 may play a role in cartilage protection by inhibiting Wnt/β-catenin signaling pathway and promote the repair of articular cartilage injury. |
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