Fig. 3

Expression of HLA-DR and costimulatory molecules as well as stimulation and proliferation index. A MSCs from P1 of 3 donors were expanded in 10% PL-240 for 7 days. The percentages of HLA-DR, CD80 and CD86 are presented as the mean values of 3 experiments. B Immunogenicity of MSCs. MSCs from 3 donors (MSC-D1, MSC-D2 and MSC-D3) were cocultured at a ratio of 1:1 with PB-MNCs isolated from buffy coats. In the other wells, MSCs were cocultured at a ratio of 1:1 with PB-MNCs isolated from buffy coats that were stimulated with anti-CD28 antibodies. PB-MNCs stimulated with anti-CD3 and anti-CD28 antibodies were used as a positive control. On day 5, the proliferation of PB-MNCs was assessed by flow cytometry, and the results are presented as the stimulation index (SI), which is expressed as the number of plated PB-MNCs (± anti-CD28) + MSCs on day 5/number of PB-MNCs (± CD28) on day 5. The results are presented as the mean ± SD of 3 experiments. There was no proliferation of either stimulated or not-stimulated MNCs with anti-CD28 antibody in the presence of MSCs. C Immunosuppressive effect of MSCs from 3 donors (MSC-1, MSCs-2 and MSC-3) cocultured with anti-CD3- and anti-CD28-stimulated PB-MNCs at a 1:1 ratio for 5 days. This effect is presented as the proliferation index (PI). The results were tested with a t test under the assumption of a normal distribution (N = 3). SI Index = MNCs ± anti-CD28 + MSCs/MNCs ± anti-CD28 presented in fold of proliferation. PI index = MNCs stimulated with anti-CD3/anti-CD28 (S) + MSCs/MNCs stimulated with anti-CD3/anti-CD28 (S) presented as fold changes in proliferation. MNC-S = stimulated with anti-CD3/anti-CD28; MNC-NS = not stimulated