Induced CD4⁺CD25⁺ FOXP-3⁺ and CD8+CD25⁺ FOXP-3⁺t Cells Exhibit a Concurrent Expression of Effector and Selective Suppressive Capacities

Naturally occurring regulatory T cells (Tregs), identified as CD4⁺CD25⁺FoxP-3⁺ cells, are known to modulate immunologic tolerance and inhibit primary T cell activation, and therefore, may protect against graft-versus-host disease (GvHD) following allograft. However, having a non-selective suppressive effect, Tregs administration following transplantation may also result in delayed immunological reconstitution and diminished graft-versus-tumor effect, leading to an increased risk of infections and disease relapse. The generation of suppressive T cells that could selectively inhibit GvHD whilst preserving immunity against malignant cells and infection would promote significant contribution to GvHD prevention and therapy. The aim of this study was to investigate the characteristics of T cells undergoing allogeneic stimulation, looking for a subpopulation with a more specific inhibition of host immunity. A negative selection of PBMCs with anti-CD25 magnetic beads resulted in the production of CD25⁺ cells (-Naturally occurring) and CD25⁻PBMCs. CD25⁻ PBMCs were than stimulated with allogeneic DC for 5–6. The resultant CD4⁺CD25⁺ or CD8⁺CD25⁺ populations were isolated with magnetic beads and investigated for immunophenotypical markers, cytokine expression profile (FACS), cell proliferation, inhibitory capacity and anti-viral response (INF-gamma). The current study has shown that stimulation of CD25⁻ T cells with allogeneic peripheral blood mononuclear or dendritic cells (DCs) induces generation of a large fraction of CD4⁺CD25⁺ and CD8⁺CD25⁺ T-cells. This process was found to be more rapid and prominent following DCs stimulation. The resultant cells exhibit a mixed phenotype, demonstrating expression of the regulatory marker, FoxP3, as well as the inflammatory cytokines, IL-2 and IFNγ. Similar to naturally occurring Tregs, the induced CD4⁺CD25⁺ and CD8⁺CD25⁺ have markedly inhibited alloreactive T cell expansion. However, in contrast to naturally occurring Tregs, the induced populations, did not suppress proliferation against the cytomegalovirus (CMV) recombinant protein, suggesting their potential for more selective inhibition of host immunity.

According to these data, CD25⁻ derived CD4⁺CD25⁺FOXP-3⁺ and CD8⁺CD25⁺ FOXP-3, appear to have a dual expression of both effector and selective suppressive capacities, and therefore, might be potentially ideal for the use as post-transplant GvHD prophylaxis.