Protective effects of Wuweixiaoduyin against chronic osteomyelitis through Foxp3+CD25+CD4+Treg cells via the IL-2/STAT5 signaling pathway

To explore the efficacy and safety of the Chinese medicine prescription Wuweixiaoduyin in inhibition of chronic osteomyelitis via regulatory T cells signaling. The effective components of Wuweixiaoduyin and osteomyelitis related genes were screened. Target proteins were cross-validated using the Venny database. GO and KEGG pathway analyses were performed for target proteins as well as pharmacological network was constructed. The bone properties were analyzed by HE staining and the concentration of immune factors were measured by ELISA. Real-time PCR and western blot were used to analyze mRNA and protein expression level. FACS assay was used to analyze the rate of cells. 117 genes overlapped between 785 target genes of active compounds of WWXDY and 912 Osteomyelitis related genes. Inflammation-related genes, including IL-6, TNFa, IL-1b and IL-2 showed high connection degree in drug-compounds-disease-targets network. GO function and KEGG pathway analyses revealed that 117 intersection genes mainly enriched in virus infection related pathways, immune related pathways and Chemokine signaling pathway. Besides, the development of chronic osteomyelitis was suppressed in rat models after treatment of Wuweixiaoduyin granule and meanwhile the concentration of IL-2 and CD4+CD25+Foxp3 Treg rates together with the levels of p-STAT5, CTLA-4 and Foxp3 were also downregulated. Further studies also showed that IL-2 and Wuweixiaoduyin drug-containing serum showed an opposite effect on regulating IL-2, IL-10, TGF-β1 Foxp3, CTLA4 and STAT5. In addition, STAT5 phosphorylation inhibitor could also suppress the expression of Foxp3 and CTLA-4. Wuweixiaoduyin could treat chronic osteomyelitis via suppressing the main regulating factors of Tregs and further interfere its immunodepression. Our results bring new solution for chronic osteomyelitis.