MSC^wt and MSC^IFNγR1- inhibit proliferation of PBMCs in the absence of IDO—contribution of IGFBPs. Proliferation of HLA-mismatched PBMCs in the absence (■) or presence (▩) of MSC^wt compared with PBMCs in the presence of 1 mM of the IDO inhibitor 1-methyltryptophan (□). (A) 1-methyltryptophan partially restored the proliferation of PBMCs cocultured with MSC^wt (representative of n = 5). (B) By contrast, 1-methyltryptophan had no effect on cocultures of MSC^IFNγR1− and PBMCs (representative of n = 3). (C) RT-PCR of RNA isolated from MSCs. IDO was not constitutively expressed (lane a) in either MSC^wt or MSC^IFNγR1−. IDO was induced in MSC^wt when they were cocultured with PBMCs in the presence of IL-2 and OKT3 (lane b, separated by a semipermeable membrane; lane c, in direct cell-cell contact) or when IFNγ was added (lane d) (representative of n = 3). (D) IL-2/OKT3-induced proliferation of PBMCs in media conditioned by PBMCs alone, by cocultures of MSC^wt/PBMCs or by MSC^IFNγR1−/PBMCs, respectively, for 3 days. Media conditioned by MSC^wt/PBMCs and by MSC^IFNγR1−/PBMCs inhibited proliferation of PBMCs (▩). Inhibition was partially reverted by depletion of free IGFBP from the media of both cocultures, MSC^wt/PBMCs and MSC^IFNγR1−/PBMCs (□) (representative of n = 3). Data are shown as means of triplicates (±SD) of one representative experiment.