Abstract: OBJECTIVE To explore the inhibitory effects of pseudolycorine on activated T cell proliferation and its mechanism. METHODS Human T cells were isolated and purified by density gradient centrifugation and the immunomagnetic microbeads, and activated by anti-CD3/CD28 mAbs or phytohemagglutinin(PHA). Cell proliferation, apoptosis, CD25 expression and cell cycle were detected by flow cytometry. Level of cytokines IL-2, IL-6, IL-17A and IFN-γ were measured by ELISA. RESULTS Pseudolycorine inhibited human T cell proliferation with anti-CD3/CD28 mAbs stimulation with an IC₅₀ value of (0.97±0.22) μmol·L^-1 and PHA stimulation with an IC₅₀ value of (0.82±0.07) μmol·L^-1. Pseudolycorine did not induce apoptosis of activated T cells and had no significant effect on the cell viability of resting T cells with the same concentration of inhibiting activated T cell proliferation. Pseudolycorine did not affect CD25 and IL-2 expression, but induced cell cycle arrest in G0/G1 phase. Pseudolycorine significantly inhibited IL-6, IL-17 and IFN-γ expression. CONCLUSION Pseudolycorine does not affect the activation of T cells, but inhibits the proliferation of T cells by blocking the cell cycle in G0/G1 phase. Pseudolycorine represents a potential lead compound for the design and development of new immunosuppressive drugs.