Early reduction in number of T cells producing proinflammatory cytokines,observed after extracorporeal photopheresis,is not linked to apoptosis induction

Immediately following ECP, a significant number of lymphocytes become apoptotic and the number of T cells producing TNFalpha and IFNgamma is reduced. This study sought to determine if the cytokine down-regulation was a direct consequence of apoptosis induction. METHODS: Samples were obtained from 6 graft versus host disease (GvHD) and 5 cutaneous T cell lymphoma (CTCL) patients immediately pre-ECP and from the leucocyte collection bag following 8-MOP/UVA exposure, but prior to re-infusion. Separated peripheral blood mononuclear cells (PBMC) were placed in cell culture and stimulated for 6 hours with phorbol myristate acetate (PMA), Ionomycin and Brefeldin A. Using flow cytometry, T cells were identified by CD3 expression and apoptotic T cells sub-selected by Annexin V staining. Both apoptotic and non-apoptotic T cells were evaluated for their intracellular expression of IL2, IL4, IL10, IFNgamma and TNFalpha. RESULTS: Neither patient group demonstrated a significant change in IL4 or IL10 expression post ECP. However the number of T cells expressing IL2, IFNgamma and TNFalpha was reduced in both the Annexin V-positive and -negative T cell populations (P <.05). The nonapoptotic T cells from GvHD patients demonstrated the greatest reduction in cytokine expression. CONCLUSIONS: Since proinflammatory cytokines play a major role in the pathology of GvHD, their down-regulation post-ECP may produce a direct clinical benefit. The lowest number of IL2-, IFNgamma- and TNFalpha-expressing T cells occurred within the apoptotic population; however, Annexin V-negative T cells also demonstrated a marked reduction post-ECP. However, the lack of an increase in IL4 and IL10 expression indicates that this process was not a consequence of skewing toward a Th2 cytokine profile.