Curcumin inhibits mitogen stimulated lymphocyte proliferation, NFκB activation, and IL-2 signaling

Abstract

T cell mediated acute rejection of transplanted organ continues to be a noticeable problem in solid organ transplantation. We showed that Curcumin is a potent inhibitor of Cyclosporin A resistant T cell CD28 co-stimulation pathway. Here we report the inhibitory effects of Curcumin on mitogen-stimulated lymphocyte proliferation, IL-2 synthesis/signaling, and NFkappaB (transcription factor of IL-2 promoter) activation. Human lymphocytes were isolated from fresh human spleen (SP-L). Mitogens [final concentrations of 2 microg/ml concanavalin A (Con A), 5 microg/ml phytohemagglutinin (PHA), and 20 ng/ml of phorbol-12-myristate-13-acetate (PMA)] were added to the designated wells in a 96-well plate with 0.2 million SP-L and cultured for 48 h and then assayed for IL-2 synthesis by ELISA and 3H-thymidine uptake. In another parallel experiment we added IL-2 (0.5 nM) to stimulate the cells to check if Curcumin's inhibition of IL-2 synthesis is the sole reason for inhibition of cell proliferation. Electrophoretic mobility shift assay (EMSA) was performed in PMA (20 ng/ml, 1 h) stimulated cells with or without Curcumin to assay NFkappaB activation. Curcumin at 2.5 microg/ml inhibited Con A, PHA, and PMA stimulated SP-L proliferation at 77, 23, and 48%, respectively, over controls and Curcumin at 5 microg/ml completely (nearly 100%) inhibited the mitogen stimulated proliferation. Curcumin inhibited IL-2 synthesis in Con A, PHA, and PMA stimulated SP-L in a concentration-dependent manner with an ED50 (concentration required for 50% inhibition) measured at 3.5 microg/ml. Exogenous IL-2 stimulated SP-L proliferation was also inhibited by Curcumin in a concentration-dependent manner with an ED50 of 2 microg/ml. EMSA assay indicated that PMA at 20 ng/ml stimulated NFkappaB activation 253% over control, which was inhibited by 24, 38, and 73%, respectively, with Curcumin at final concentrations of 2.5, 5, and 10 microg/ml, respectively. Curcumin has profound immunosuppressive effects mediated via inhibition of IL-2 synthesis, mitogen, and IL-2 induced activation of human lymphocytes. This effect may be mediated via NFkappaB inhibition.