Replicative senescence associated impairment of immunosuppressive properties of Mesenchymal stromal cells

Industry sponsored clinical trials differ from academic clinical trials by using Mesenchymal stromal cells (MSCs) expanded heavily in vitro. Prolonged expansion leads to replicative senescence of MSCs and the effect of this process on the immunobiology of MSCs is unknown. We passaged MSCs from the bone marrow of two independent donors for a period of 60 days. Real time PCR analysis demonstrated that shortening of telomere length in MSCs is directly associated with prolonged culture expansion mediated replication defect. Further phenotypical analysis demonstrated that replication defective MSCs not only exhibit cellular hypertrophy also express senescent marker, βgalactosidase. In vitro T cell suppressive assays demonstrate that senescent MSCs display defect in inhibiting T cell proliferation and cytokine secretion. In addition, replication competent MSCs maintain their inhibitory potential upon coculture with senescent MSC. This suggests senescent MSCs lack bystander proinflammatory role to other fit cells in a heterologous cell preparation where senescent and non-senescent MSC are mixed. Interestingly, senescent MSCs upregulate IDO upon stimulation with IFNγ, which suggests that IDO response is intact in these cells despite defective immunesuppression. Further analysis demonstrates that IFNγ upregulates MHC-I, B7-H1 and B7DC but not the costimulatory molecules B7-1 and B7-2 on both MSC populations. However, unlike fit MSCs IFNγ does not upregulate HLADR on senescent counter parts. In addition senescent MSCs display defect (2log) in upregulating the HLA class II transactivator (CIITA) upon stimulation with IFNγ. In summary, senescent MSCs display defective immunosuppressive properties, lacking HLADR upregulation despite intact IDO response, suggesting the tolerizing role of HLADR in MSC’s immunobiology. Our results suggest the advantage of using non-senescent MSCs in clinical trials.