INTRAVENOUS INFUSION OF APOPTOTIC CELLS SIMULTANEOUSLY WITH ALLOGENEIC HEMATOPOIETIC GRAFT ALTERS ANTI-DONOR HUMORAL IMMUNE RESPONSES

Abstract

O106* Aims: Use of reduced intensity conditioning (RIC) regimens before allogeneic hematopoietic cell (HC) transplantation leads to limited toxicity allowing the extension of HC transplantation to organ transplantation. In contrast to myeloablative allografts, RIC regimens were associated with longer host immune cell persistence leading to increased HC graft rejection. We have previously described that intravenous infusion of apoptotic leukocytes with an allogeneic bone marrow (BM) graft favors engraftment across major histocompatibility barriers without inducing auto-immunity. This graft-facilitating effect observed is independent of the apoptotic cell origin (donor, recipient or third party). Apoptotic cells may induce allo-immune responses against themselves, but also against co-infused viable cells (e.g. BM cells) due to their adjuvant properties. Here we evaluated the risk of anti-donor immunization. Methods: Mice receiving allogeneic BM cells with or without apoptotic cells after a RIC regimen were analyzed 6-9 weeks after BM transplantation (BMT) for engraftment. Sera were collected for the detection of allo-Ab directed against BM and/or apoptotic cell determinants, using complement dependent cytotoxicity assay as well as flow cytometry. Skin graft was also performed. Results: Six to 9 weeks post-BMT, only 2% of recipient mice receiving donor or third party apoptotic cells with their BM graft presented cytotoxic Abs directed against apoptotic cell determinants. This absence of immune responses was confirmed using more sensitive methods: flow cytometry and skin graft. Tolerance (as attested by skin graft survival) was restricted to BM donor Allo-Ags. When now testing the allo-immunization against the BM donor, apoptotic cell infusion did not favor such immunization. As expected in view of the engraftment effect, apoptotic cell infusion considerably reduced the frequency of mice immunized against the BM donor (45% of allo-immunized recipients receiving only BM versus 5% of allo-immunized recipients in presence of apoptotic cells). Surprisingly and most importantly, we found that apoptotic cell infusion at the time of BMT could successfully prevent humoral allo-immunization against the donor BM cells in recipients having rejected their marrow. Among these recipients, prior apoptotic cell infusion was associated with a near absence of cytotoxic Ab-mediated allo-responses (2/50, 4%), while such immunization was frequently observed in non-engrafted recipients that had not received apoptotic cells (29/62, 47%, P<.05). Such cytotoxic Abs were identified as Th1-associated IgG2a isotype. In vivo anti-TGF-beta Ab treatment resulted in the loss of this apoptotic cell infusion-associated protective effect on humoral allo-responses. Infusion of neutralizing anti-TGF-beta simultaneously to apoptotic cells was indeed associated with humoral anti-BM donor-immunization among most of the non-engrafted mice (80%, 4/5, P<.05). Conclusions: Overall, we demonstrate that apoptotic cell infusion with an allogeneic graft can, in addition to enhancing engraftment, also prevent the Ab-mediated allo-immunization otherwise frequently observed in recipients rejecting their graft. Because graft rejection is mainly T- and/or NK-cell-mediated, our results suggest that apoptotic cell infusion-induced immunomodulation can differentially affect cellular and humoral allo-immune responses. This effect of apoptotic cells may be of interest to prevent deleterious Ab-mediated allo-responses in various transplantation settings.