Persistent Recipient Antigen-Presenting Cells in Human Hematopoietic Stem Cell Transplantation: Identification of a New Dermal Subset That Outlives Epidermal Langerhans Cells.

Graft versus host disease (GVHD), a potentially lethal complication of hematopoietic stem cell transplantation (HSCT), is initiated by recipient antigen presenting cells (APC) priming donor T cells. The skin, a commonly affected organ, contains diverse APC including epidermal Langerhans cells (LC) and heterogeneous dermal dendritic cells (DC) whose phenotype, turnover and contribution to GVHD is not well described. The persistence of LC in mice and to some extent humans, is well known, yet the importance of LC relative to dermal DC in the lichenoid dermal infiltration of GVHD remains unproven. In this study we have:

1. analyzed the depletion of dermal APC by conditioning, demonstrating a resistant DR+ CD14+ subset;

2. identified a DR+ CD14+ fXIIIa+ dermal cell that remains recipient in origin for up to 1 year post HSCT and may therefore contribute to the sensitization of donor T cells and promotion of GVHD.

From 2003 to 2007 we obtained 22 pairs of 2mm skin biopsies pre and post conditioning (high dose BuCy or CyTBI and reduced intensity Flu/Mel) and 85 biopsies at day 40, 100 and 365 post HSCT from patients with sex-mismatched donors. Pre and post conditioning samples were digested with dispase and collagenase followed by single step B–D Trucount analysis with CD45, HLA-DR, CD14 and CD1a antibodies. Chimerism was determined on cytospins of spontaneously migrated dermal APC and collagenase digested dermal cells using sequential four-colour confocal microscopy and X/Y fluorescence in situ hybridization. We find 3 populations of CD45+ DR+ dermal cells: CD14+ CD1a- fXIIIa-; CD14+ CD1a- fXIIIa+; CD1a+ CD14- fXIIIa-. Characterization of these cells in vitro shows that CD14+ cells are more adherent and phagocytic than CD1a+ cells and that the fXIIIa+ component is in addition, heavily granulated with ingested melanin. CD1a+ APC are more sensitive to depletion by conditioning (201 pre /97 post; mean cells per mm²; P 0.03) compared with CD14+ cells (210 pre /213 post; cells per mm²; P 0.86). Both are reduced in HSCT patients compared with controls (706 CD14+ and 468 CD1a+ cells per mm²; both P 0.01), in contrast to LC that are relatively well preserved in HSCT patients. CD1a+ cells are equally sensitive to depletion by both high dose and reduced intensity HSCT. Chimerism analysis at 40, 100 and 365 days post HSCT has shown regimen-dependent but nearly complete donor LC engraftment by 100 days. In dermal cells, two strikingly different patterns of engraftment are seen. CD1a+ and CD14+ fXIIIa- cells, obtained either by migration or digestion, engraft rapidly in all patients, ahead of LC, reaching medians of 99% and 100% donor, respectively, at day 40. In contrast, CD14+ fXIIIa+ cells, which are obtained only from digested dermis, are very slow to engraft in the absence of GHVD, reaching only 63% median donor after high dose and 10% median donor chimerism after reduced regimens, at 1 year. Acute GVHD promotes engraftment with 100% donor chimerism seen in nearly all patients at day 100. Preliminary data indicate that these cells are not actively synthesizing DNA, suggesting a different means of survival compared with persistent recipient LC. In conclusion, we have identified a new subset of human dermal APC with protracted survival after high dose and reduced intensity HSCT, sensitivity to GVHD and potential importance in promoting donor T cell reactivity to host antigens.