Figure 1

IL-17 enhances the immunosuppressive effect of MSCs. (a) Cloned MSCs were first treated with the indicated combinations of recombinant cytokines IFNγ, TNFα, and IL-17 (2 ng/ml each) for 12 h, then co-cultured with T-cell blasts at a 1 : 20 ratio (MSC: T cells), and proliferation was assessed by ³H-thymidine incorporation after an additional 12 h. (b) The mRNA expression of IL-17 receptor family members in MSCs or cell line Raw 264.7 were examined by quantitative RT-PCR. (c) Surface expression of IL-17RA was detected by flow cytometry with cloned MSCs. (d and e) MSCs were first treated with IFNγ and TNFα, with or without IL-17 (10 ng/ml). IFNγ and TNFα were supplemented at different cytokine concentrations, for 12 h, and then co-cultured with T-cell blasts (d) or T-cell hybridoma A1.1 cells (e) at a ratio of 1 : 20 for 12 h. T-cell proliferation was measured by ³H-thymidine incorporation. (f) MSCs were first treated with IFNγ and TNFα (2 ng/ml) with graded concentrations of IL-17 for 12 h, and then co-cultured with T-cell hybridoma A1.1 cells at a ratio of 1 : 10 for 12 h. T-cell proliferation was measured by ³H-thymidine incorporation. (g) MSCs were co-cultured with fresh C57BL/6 splenocytes activated with anti-CD3 and anti-CD28, or in the presence of antibodies against IL-17A, at a 1 : 20 or 1 : 40 ratio (MSCs: splenocytes), for 48 h, and then cell proliferation was assessed by ³H-thymidine incorporation. (h) MSCs were first treated with IFNγ and TNFα (2 ng/ml), or together with IL-17 for 12 h, and then co-cultured with T-cell hybridoma A1.1 cells at a ratio of 1 : 10, in the presence or absence of L-NMMA, for 12 h. T-cell proliferation was measured by ³H-thymidine incorporation. Proliferation values represent means±S.E.M. of three wells from a representative of three independent experiments