Objective To investigate the potential mechanism of the ethanol extract of Juanbi Granules (JBKL) on T cell activation via the mitogen-activated protein kinase (MAPK) signaling transduction pathway, and to reveal the mechanism of immunosuppression on the application of Yang-aided theory in rheumatoid arthritis(RA) treatment. Methods T cells and B cells proliferation assay were induced by concanavalin A(Con A) and lipopolysaccharide(LPS), respectively. Splenocyte proliferation and cytotoxicity were detected by MTT assay. The T cells were further stimulated in vitro with Anti-CD3 mAb to establish Anti-CD3 mAb-induced T cells activation; ELISA assay was used to measure the supernatant concentrations of interferon-γ (IFN-γ), interleukin (IL)-17A, IL-10 and IL-6. The expression of MAPK signaling pathway related proteins (p-ERK and p-p38) in purified CD4+ T cells were detected by Western blot assay. Results Compared with the vehicle control group, JBKL (30 μg/mL) significantly inhibited the proliferation of T cells induced by Con A and Anti-CD3 mAb, and the proliferation of B cells responded to LPS. Further, JBKL groups had no remarkable cytotoxicity in normal mouse spleen lymphocytes. The levels of IFN-γ L-17A, IL-6 content were obviously decreased in the JBKL(3~30 μg/mL) group and the IL-10 content was up-regulated. The phosphorylation levels of ERK and p38 proteins were lower in the JBKL(100 μg/mL) group than those in the vehicle group. Conclusion JBKL can significantly restrain the activation and proliferation of CD4+ T cells, which may be related to the inhibition of p-ERK and p-p38 MAPK signaling pathways. This may be an important molecular mechanism contributes to the effect of JBKL in rheumatoid arthritis treatment.