Immature DC isolated after co-culture with PUVA-treated peripheral blood mononuclear cells downregulate graft-versus-host reactions in the human skin explant model.

Graft-versus-host disease (GvHD) remains the major barrier to successful allogeneic hematopoietic stem cell transplantation (HSCT). Extracorporeal photopheresis (ECP) is a potent immunomodulatory treatment option for GvHD. In contrast to conventional immunosuppressants, ECP is considered not to increase relapse and infection rates resulting from generalised immunosuppression. ECP involves the mechanical separation of 5-10% of patient peripheral blood mononuclear cells, which are then exposed to psoralen and UVA light (PUVA) before they are returned to the patient. ECP has been shown to induce apoptosis in various cell types, in particular lymphocytes. Several studies describe downregulation of pro-inflammatory cytokines as well as promotion of peripheral tolerance through enhanced production of T regulatory cells in the course of ECP-treatment. Modulation of antigen-presenting cells such as dendritic cells (DC) by PUVA-treated lymphocytes might be implicated in these regulatory processes. We evaluated the impact of PUVA-treated lymphocytes on immature DC and further demonstrated the functional capacity of such modified DC to modulate GVH reactions using a well-established human skin-explant model of GvHD. Addition of immature DC isolated after co-culture with PUVA-treated but not untreated MLR cells significantly downregulated skin-GvH reactions (p=0.023, Mann-Whitney-Test). IFN-gamma levels were non-significantly decreased in MLR and skin supernatants. We observed a non-significant increase in PD-L1 expression in iDC after co-culture with PUVA-treated MLR cells whereas expression levels of IDO and ILT-3 were not affected. We conclude that iDC modulated by PUVA-induced apoptotic cells potently downregulate allogeneic immune responses possibly through PD-L1- dependent signaling.

Impact of psoralen/UVA-treatment on survival, activation, and immunostimulatory capacity of monocyte-derived dendritic cells.

BACKGROUND: Extracorporeal Photopheresis (ECP) has been shown to be an effective treatment of graft-versus-host disease, solid organ graft rejection, and other T-cell-mediated diseases. The mechanisms of action of ECP include lymphocyte apoptosis, cytokine modulation, and the induction of regulatory T cells. It has been suggested that dendritic cells (DCs) are more resistant to ECP-induced apoptosis and might be directly modulated by ECP. We tested this hypothesis using in vitro Psoralen/UVA (PUVA) treatment as an in vitro model of ECP. METHODS: Monocyte-derived DCs (mo-DCs) were treated with 8-methoxypsoralen /UVA and analyzed for surface molecule expression, apoptosis markers, endocytosis, and migratory and immunostimulatory capacity. Mo-DC phenotype and cytokine secretion was tested after CD40L stimulation. Naive T cells stimulated with PUVA-treated mo-DCs were tested for Th1/Th2 cytokine secretion and associated chemokine receptor patterns. RESULTS: DCs underwent apoptosis after in vitro PUVA and in vivo ECP. In vitro, the induction of apoptosis was preceded by partial maturation of immature mo-DCs. PUVA-treated immature mo-DCs also exhibited enhanced migratory and immunostimulatory capacity. However, mo-DCs stimulation through CD40 ligation was abrogated and interleukin (IL)-12 secretion was abolished 24 hr after PUVA treatment. PUVA-treated mo-DCs skewed naive T cells toward a Th2 response as defined by increased IL-4, IL-10, and IL-13 and decreased interferon-gamma levels, and the expression of the Th2-associated chemokine receptors CCR4 and CCR10. The observed Th2 shift was partially reversed by exogenous IL-12. CONCLUSION: These data suggest that direct modulation of DC function as well as apoptosis contribute to the immunoregulatory effects of ECP.

Technology insight: ECP for the treatment of GvHD--can we offer selective immune control without generalized immunosuppression?

Hematopoietic stem-cell transplantation remains an important curative therapy for many conditions and its use is increasing annually. Graft-versus-host disease (GvHD) is the major cause of mortality and suffering following allogeneic hematopoietic stem-cell transplantation. Conventional treatments are associated with multiple side effects and are often ineffective. New therapeutic approaches for the control of GvHD are desperately required. Extracorporeal photochemotherapy (ECP) was developed in the 1970s for the treatment of cutaneous T-cell lymphoma and was approved by the FDA as the first selective immunotherapy for a cancer. ECP has also proved an effective therapy for immune-related conditions, particularly GvHD, even in patients refractory to conventional therapies. The treatment involves the mechanical separation of circulating white cells, which are exposed to psoralen and UVA light and then returned to the patient. ECP is extremely well tolerated with minimal side effects and is not associated with the increased rates of infection or relapse of malignant disease typical of conventional immunosuppressive agents. Thus, ECP appears to offer selective immune modulation without generalized immunosuppression, but its mechanism of action remains poorly understood. This review discusses the development of ECP, its use in the treatment of GvHD, as well as current theories of its mechanism of action.