Autologous bone marrow-derived mesenchymal stromal cells (MSCs) for adoptive cell therapy of luminal Crohn's disease (CD) are being tested in clinical trials. However, CD is associated with dysregulation of autophagy and its effect on MSC's immunobiology is unknown. Here, we demonstrate no quantitative difference in phenotype, in vitro growth kinetics and molecular signatures to IFNγ between MSCs derived from CD and healthy individuals. CD MSCs were indistinguishable from those derived from healthy controls at inhibiting T-cell proliferation through an indoleamine 2,3-dioxygenase (IDO)-dependent mechanism. Upon IFNγ prelicensing, both MSC populations inhibit T-cell effector functions. Neither a single-nucleotide polymorphism (SNP) rs7820268 in the IDO gene, nor a widely reported CD predisposing SNP ATG16L1rs2241880 modulated the suppressive function of MSCs carrying these haplotypes. IFNγ stimulation or coculture with activated T cells upregulated the expression of autophagy genes and/or vacuoles on MSCs. Pharmacological blockade of autophagy pathway did not reverse the immunosuppressive properties and IFNγ responsiveness of MSCs confirming the absence of a functional link between these two cell biochemical properties. We conclude that autophagy, but not IDO and IFNγ responsiveness, is dispensable for MSC's immunosuppressive properties. MSCs from CD subjects are functionally analogous to those of healthy individuals.
Industrial-scale expansion of mesenchymal stromal cells (MSCs) is often used in clinical trials, and the effect of replicative senescence on MSC functionality is of mechanistic interest. Senescent MSCs exhibit cell-cycle arrest, cellular hypertrophy, and express the senescent marker β-galactosidase. Although both fit and senescent MSCs display intact lung-homing properties in vivo, senescent MSCs acquire a significant defect in inhibiting T-cell proliferation and cytokine secretion in vitro. IFNγ does not upregulate HLA-DR on senescent MSCs, whereas its silencing did not reverse fit MSCs’ immunosuppressive properties. Secretome analysis of MSC and activated peripheral blood mononuclear cell coculture demonstrate that senescent MSCs are significantly defective in up (vascular endothelial growth factor [VEGF], granulocyte colony-stimulating factor [GCSF], CXCL10, CCL2) or down (IL-1ra, IFNγ, IL-2r, CCL4, tumor necrosis factor-α, IL-5) regulating cytokines/chemokines. Unlike indoleamine 2,3 dioxygenase (IDO), silencing of CXCL9, CXCL10, CXCL11, GCSF, CCL2, and exogenous addition of VEGF, fibroblast growth factor-basic do not modulate MSCs’ immunosuppressive properties. Kynurenine levels were downregulated in senescent MSC cocultures compared with fit MSC counterparts, and exogenous addition of kynurenine inhibits T-cell proliferation in the presence of senescent MSCs. IFNγ prelicensing activated several immunomodulatory genes including IDO in fit and senescent MSCs at comparable levels and significantly enhanced senescent MSCs’ immunosuppressive effect on T-cell proliferation. Our results define immune functional defects acquired by senescent MSCs, which are reversible by IFNγ prelicensing.
Utilization of mesenchymal stromal cells (MSCs) for the treatment of Crohn's disease and ulcerative colitis is of translational interest. Safety of MSC therapy has been well demonstrated in early phase clinical trials but efficacy in randomized clinical trials needs to be demonstrated. Understanding MSC mechanisms of action to reduce gut injury and inflammation is necessary to improve current ongoing and future clinical trials. However, two major hurdles impede the direct translation of data derived from animal experiments to the clinical situation: (1) limitations of the currently available animal models of colitis that reflect human inflammatory bowel diseases (IBD). The etiology and progression of human IBD are multifactorial and hence a challenge to mimic in animal models; and (2) Species specific differences in the functionality of MSCs derived from mice versus humans. MSCs derived from mice and humans are not identical in their mechanisms of action in suppressing inflammation. Thus, preclinical animal studies with murine derived MSCs cannot be considered as an exact replica of human MSC based clinical trials. In the present review, we discuss the therapeutic properties of MSCs in preclinical and clinical studies of IBD. We also discuss the challenges and approaches of using appropriate animal models of colitis, not only to study putative MSC therapeutic efficacy and their mechanisms of action, but also the suitability of translating findings derived from such studies to the clinic.
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