伤科灵喷雾剂治疗急性软组织损伤的作用机制研究OA

Objective To observe the therapeutic efficacy of Shangkeling spray in treating acute soft tissue injury(ASTI)in rats and delve into its possible mechanism by which it can intervene in ASTI.Methods In this study,a reproducible closed soft tissue injury model was established in the right hind limb of male SD rats using the free-fall method.A total of 45 SD rats were randomly allocated into 5 groups:The normal group,the model group,the Yunnan Baiyao group,the Futalin group,and the Shangkeling group,with 9 rats in each group.The rats received daily administration of the respective treatments.The degree of limb swelling in rats was assessed by measuring the circumference of the right hindlimb gastrocnemius muscle on a daily basis.The success of the modeling process was evaluated using the difference in weight-bearing capacity between the two feet of rats as an indicator of pain.On the first,third,and eighth days post-treatment,hematoxylin-eosin(H&E)staining was employed to evaluate the infiltration of inflammatory cells,necrosis and regeneration of muscle fibers at the injury site.On the third day after the start of the treatment,enzyme-linked immunosorbent assay(ELISA)was utilized to measure the levels of pro-inflammatory factors,namely tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β),as well as anti-inflammatory factors,including interleukin-10(IL-10)and transforming growth factor-β1(TGF-β1)in the serum.The potential molecular mechanism of Shangkeling spray in treating soft tissue injury was analyzed based on transcriptome data.The phenotypic polarization of macrophages was analyzed through immunofluorescence staining of the damaged muscle tissue with CD86(a marker for M1 macrophages)and CD206(a marker for M2 macrophages).On the eighth day post-treatment,immunohistochemistry was used to detect the expression of vascular endothelial growth factor(VEGF)and Platelet endothelial cell adhesion molecule-1(CD31)to assess angiogenesis.Results On the eighth day post-treatment,the model group still exhibited severe tissue edema,whereas the swelling rates in each treatment group were significantly reduced,showing no significant difference compared to the normal group.Compared with the normal group at the same time point,the difference in bipedal weight-bearing in rats across all groups significantly increased after modeling(P<0.0001),indicating successful modeling.Following treatment,the pain in each treatment group gradually diminished and eventually recovered.Compared with the model group,the H&E staining results of the treatment groups revealed a marked reduction in the inflammatory response after injury,a decrease in fibrosis levels,an increase in the number of newly formed muscle fibers,and enhanced muscle regeneration.The ELISA results suggested that Shangkeling spray could alleviate the inflammatory response by decreasing the levels of serum pro-inflammatory factors TNF-α and IL-1β,while increasing the levels of anti-inflammatory factors TGF-β1 and IL-10.The transcriptome results indicated that Shangkeling spray might exert its therapeutic effects by regulating key factors involved in macrophage polarization.Immunofluorescence results demonstrated a significant inhibition of CD86 expression and an elevation in CD206 expression levels,suggesting a polarization in macrophage from the M1 to the M2 type.Concurrently,the positive staining signal for VEGF in the Shangkeling group was higher than that in the model group,promoting vascular repair in the injured tissues.Conclusion Shangkeling spray can effectively suppress excessive inflammatory reactions,enhance VEGF-mediated angiogenesis,and ultimately accelerate the repair of soft tissue injuries by promoting the transition of macrophages from the pro-inflammatory M1 phenotype to the anti-inflammatory M2 phenotype.