Abstract:

AIM: To study the therapeutic effect of Yiqi Huoluo formula on collagen-induced arthritis (CIA) rats and its effect on PI3K/Akt/mTOR signaling pathway. METHODS: A total of 60 SD rats were randomly divided into blank group, model group, methotrexate group (1 mg/mL), and Yiqi Huoluo Fang low, medium and high dose groups (9.45 g/kg, 18.9 g/kg, 37.8 g/kg), with 10 rats in each group. Except for the blank group, CIA rat models were established for the rest of the rats. After 28 days of continuous intervention after booster immunization, the samples were collected. During the period, the general condition, ankle swelling and arthritis index score of the rats were observed. Enzyme linked immunosorbent assay (ELISA) was used to detect the levels of TNF-α, IL-1β, IL-6 and IL-10 in the serum of rats. Hematoxylin-eosin staining (HE) was used to observe the histopathological changes of the synovial synovium of the ankle joint in rats. Quantitative Real-time polymerase chain reaction (RT-qPCR) was used to detect the mRNA expressions of PI3K, Akt, mTOR, CD86 and CD206 in the synovial tissue of rat knee. Western blot was used to detect the expression levels of p-PI3K, p-Akt and p-mTOR proteins in the synovial tissue of rat knee. Immunohistochemistry was used to determine the positive expression of CD86 and CD206 in rat ankle joints. RESULTS: Compared with the blank group, the joint swelling and AI index of the model group increased (P<0.01), a large number of inflammatory cell infiltration, fibrous tissue and synovial cell proliferation and disordered arrangement were seen in the pathological sections of the ankle joint, a small number of capillaries were found around the synovial tissue, the serum levels of TNF-α, IL-1β and IL-6 were increased, and the levels of IL-10 were decreased (P<0.05, P<0.01), the expression levels of PI3K, Akt, mTOR, CD86 and mRNA in the synovial tissue of knee joint were increased, while the expression of CD206 mRNA was decreased (P<0.05, P<0.01), the expression levels of p-PI3K, p-Akt and p-mTOR proteins in synovial tissue increased (P<0.05, P<0.01), and the expression of CD86 protein and CD206 protein decreased in synovial membrane of ankle joint (P<0.05, P<0.01). Compared with the model group, the degree of ankle swelling in the methotrexate group, the Yiqi Huoluo formula and the high-dose group decreased (P<0.05, P<0.01), the AI index decreased (P<0.05, P<0.01), the serum levels of TNF-α, IL-1β and IL-6, the levels of IL-10 increased (P<0.05, P<0.01), the expression levels of PI3K, Akt, mTOR, CD86 and mRNA in the synovial tissue of the knee joint decreased, and the expression of CD206 mRNA increased (P<0.05, P<0.01), the expression levels of p-PI3K, p-Akt and p-mTOR proteins in synovial tissue decreased (P<0.05), the expression of CD86 protein decreased in synovial membrane of ankle joint, and the expression of CD206 protein increased (P<0.05) and can improve synovial inflammation. CONCLUSION: The Yiqi Huoluo formula alleviates synovial inflammation in rheumatoid arthritis (RA) and reduces joint pathological damage by down-regulating the PI3K/Akt/mTOR pathway, modulating macrophage polarization, decreasing pro-inflammatory factors, and promoting the expression of anti-inflammatory factors.

Key words: phosphatidylinositol 3-kinase/protein kinase B/mammalian rapamycin target pathway, rheumatoid arthritis, synovitis, Yiqi Huoluo formula