基于胰岛素样生长因子Ⅰ-磷脂酰肌醇3-激酶-蛋白激酶B信号通路探讨推拿疗法对运动性骨骼肌损伤大鼠运动功能的影响及作用机制OA

Objective:To investigate the effects of Tui Na therapy on motor function in rats with exercise-induced muscle damage(EIMD),and to explore its underlying mechanisms based on the insulin-like growth factor Ⅰ(IGF Ⅰ)-phosphatidylinositol 3-kinase(PI3K)-protein kinase B(PKB)signaling pathway.Methods:Thirty 8-week-old male Sprague-Dawley(SD)rats were selected and randomized into blank group,model group,Tui Na group,NVP-AEW541 group,and Tui Na combined with NVP-AEW541 group,with 6 ones in each group.All rats but the ones in the blank group underwent exhaustive treadmill training to induce EIMD.Following the end of modeling,the rats in the Tui Na group underwent Tui Na therapy on the right hindlimb;those in the NVP-AEW541 group received intragastric administration of IGF-1 re-ceptor inhibitor NVP-AEW541;those in the Tui Na combined with NVP-AEW541 group received both Tui Na therapy on the right hindlimbs and intragastric administration of NVP-AEW541;whereas those in the blank group and model group received only intragastric administration of an equal volume of normal saline.All rats in the 5 groups were intervened for consecutive 14 days.After the end of the intervention,the motor coordination and balance ability were assessed using the beam walking test.After that,the gastrocnemius tissues were harvested for observing the histopathological changes via HE staining and detecting the apoptosis rate via the TUNEL assay.Meanwhile,the serum sam-ples were collected,and the levels of inflammation-and oxidative stress-related markers in the serum samples were detected by ELISA.Fur-thermore,another gastrocnemius tissues were obtained,and the expression levels of IGF Ⅰ-PI3K-PKB signaling pathway-related proteins in the gastrocnemius tissues were detected by Western Blot.Results:①The motor coordination and balance ability.In the beam walking test,the model group showed significantly longer crossing time and more paw slips than the blank group(P=0.000,P=0.000);the Tui Na group exhibited significantly shorter crossing time and fewer paw slips than the model group(P=0.000,P=0.000);while the NVP-AEW541 group showed longer crossing time and more paw slips than the model group(P=0.032,P=0.002);furthermore,the Tui Na combined with NVP-AEW541 group demonstrated significantly longer crossing time and more paw slips than the Tui Na group(P=0.000,P=0.000).②Histopathological changes in gastrocnemius tissues.In the blank group,the gastrocnemius muscle fibers were arranged neat-ly,with clear and intact cross-striations and preserved muscle bundle structures;no inflammatory cell infiltration was observed.In the model group,the muscle fibers were disordered,with extensive inflammatory cell infiltration observed within the tissues.Compared with the model group,these abnormalities were markedly alleviated in the Tui Na group but aggravated in the NVP-AEW541 group.Compared with the Tui Na group,the Tui Na combined with NVP-AEW541 group also demonstrated aggravation.③The apoptosis rate in the gastrocnemius tissues.The apoptosis rate in the gastrocnemius tissues was significantly higher in the model group than that in either the blank group or the Tui Na group(P=0.000,P=0.000),but was lower than that in the NVP-AEW541 group(P=0.029).Furthermore,the Tui Na combined with NVP-AEW541 group showed a higher apoptosis rate compared with the Tui Na group(P=0.000).④The serum levels of inflammation-and oxidative stress-related markers.The serum interleukin(IL)-1β,IL-6,and malondialdehyde(MDA)levels were higher,but the superoxidase dismutase(SOD)level was lower in the model group compared to the blank group(P=0.000,P=0.000,P=0.000,P=0.000).The ser-um IL-1 β,IL-6,and MDA levels were lower,but the SOD level was higher in the Tui Na group compared to the model group(P=0.000,P=0.000,P=0.000,P=0.000).Conversely,the serum IL-1 β,IL-6,and MDA levels were higher,but the SOD level was lower in the NVP-AEW541 group compared to the model group(P=0.047,P=0.032,P=0.041,P=0.046).Furthermore,the Tui Na combined with NVP-AEW541 group exhibited significantly higher serum IL-1 β,IL-6,and MDA levels,but lower SOD level compared to the Tui Na group(P=0.000,P=0.000,P=0.000,P=0.000).⑤The expression levels of IGF Ⅰ-PI3K-PKB signaling pathway-related proteins in the gastrocnemius tissues.In the gastrocnemius tissues,the protein expression level of IGF-1,as well as the ratios of phosphorylated phosphati-dylinositol 3-kinase(p-PI3K)to PI3K and phosphorylated protein kinase B(p-PKB)to PKB,was lower in the model group compared to the blank group(P=0.000,P=0.000,P=0.000),notably,compared with the model group,these parameters were increased in the Tui Na group(P=0.000,P=0.000,P=0.000),but decreased in the NVP-AEW541 group(P=0.045,P=0.037,P=0.033),furthermore,the Tui Na combined with NVP-AEW541 group showed significantly lower values than the Tui Na group(P=0.000,P=0.000,P=0.000).Conclusion:Tui Na therapy can inhibit skeletal muscle cell apoptosis,and reduce serum inflammation and oxidative stress levels via acti-vating the IGF Ⅰ-PI3K-PKB signaling pathway,thereby promoting the skeletal muscle repair and improving motor function in EIMD rats.