Experiments of MSC induced immunosuppressive activity. CSFE dilution profile of stained responder (R) cells in MLC cultured without MSC (a) and with MSC, with R cells to MSC ratio of 20/1 (R/MSC 20/1) (b). M1 is the peak of cells not proliferating, M2 is the peak of 1st daughter proliferating cells and M3 is the peak of 2nd daughter proliferating cells. c) Percentage of R proliferating cells in MLC moving respectively to 1st and 2nd daughter generations either in absence or in presence of different MSC concentrations, in a representative experiment. d) Inhibition of cell proliferation in MLC expressed as percentage of study R cells (co-cultured with MSC) not moving from 1st daughter to 2nd daughter generation, compared to control R cells (cultured without MSC; see also methods). The experiments were performed using PL1- and PI-PL1derived MSC3, PL2- and PI-PL2-derived MSC8, PL3- and PI-PL3-derived MSC8. PBMC used as R cells in MLC were also co-cultured at R/MSC 20/1 without stimulator cells with PHA at a concentration of 50 μg/ml. Data show that with a ratio of R cells to MSC of 20/1 (R/MSC 20/1) the highest cell proliferation inhibition was observed, without any significant difference among MSC populations (as judged by two-tailed paired Student’s T test, specifically applied to assess differences between PL- and PI-PL-derived identical MSC populations). Cultures carried out in the presence of PHA produced less inhibition for any MSC population than the corresponding MLC at R/MSC 20/1. MSC, mesenchymal stem cells; R, responder cells; S, stimulator cells; MSC 50000 corresponds to R/MSC 20/1, MSC 100000 to R/MSC 10/1, MSC 200000 to R/MSC 5/1.