Mechanisms of early peripheral CD4 T-cell tolerance induction by anti-CD154 monoclonal antibody and allogeneic bone marrow transplantation: evidence for anergy and deletion but not regulatory cells

Abstract

Anti-CD154 (CD40L) monoclonal antibody (mAb) plus bone marrow transplantation (BMT) in mice receiving CD8 celldepleting mAb leads to long-term mixed hematopoietic chimerism and systemic donor-specific tolerance through peripheral and central deletional mechanisms. However, CD4 ؉ T-cell tolerance is demonstrable in vitro and in vivo rapidly following BMT, before deletion of donorreactive CD4 cells is complete, suggesting the involvement of other mechanisms. We examined these mechanisms in more detail. Spot enzyme-linked immunosorbent (ELISPOT) analysis revealed specific tolerization (within 4 to 15 days) of both T helper 1 (Th1) and Th2 cytokine responses to the donor, with no evidence for cytokine deviation. Tolerant lymphocytes did not significantly down-regulate rejection by naive donor-reactive T cells in adoptive transfer experiments. No evidence for linked suppression was obtained when skin expressing donor alloantigens in association with third-party alloantigens was grafted. T-cell receptor (TCR) transgenic mixing studies revealed that specific peripheral deletion of alloreactive CD4 T cells occurs over the first 4 weeks following BMT with anti-CD154. In contrast to models involving anti-CD154 without BMT, BMT with anti-CD154 leads to the rapid induction of anergy, followed by deletion of pre-existing donorreactive peripheral CD4 ؉ T cells; the rapid deletion of these cells obviates the need for a regulatory cell population to suppress CD4 cell-mediated alloreactivity.

Disruption of CD40/CD40-Ligand Interactions in a Retinal Autoimmunity Model Results in Protection without Tolerance

“…Interventions interfering with the CD40/CD40L interaction have been examined in a number of models, from transplantation, through autoimmune diabetes (spontaneous and lymphocytic CMV-induced), autoimmune glomerulonephritis, experimental lupus erythematosus, through EAE, an induced disease model that shares similar mechanisms with EAU. Diverse mechanisms have been reported, even in the same model, including inhibition of autoreactive T effector priming via T cell-APC interference (42,43), inhibition of Th1 effector function/expansion/migration (21,44), immune deviation/cytokine blockade (45,46), induction of anergy and/or deletion (47,48), and elicitation of regulatory cells (35,49).…”

Roles of Deletion and Regulation in Creating Mixed Chimerism and Allograft Tolerance Using a Nonlymphoablative Irradiation-Free Protocol

“…To deplete CD25 ϩ T cells, recipients were treated with 3 doses of anti-CD25 mAb (PC 61) at days Ϫ7, Ϫ5, and Ϫ3 before BMT and skin transplantation, and the depletion determined by FACS analysis on the day of BMT was Ͼ99% (21). For comparison purposes, in some mice MC was induced by administering 1.5-2.5 ϫ 10 8 BMC, anti-CD154 (0.5 mg on day 0), depleting anti-CD8 mAb 2.43 (1.44 mg on day Ϫ1), and nonmyeloablative (3 Gy) TBI as described previously (22).…”

Deletional and regulatory mechanisms coalesce to drive transplantation tolerance through mixed chimerism

“…While some form of (local) irradiation was universally required even when very profound doses of T-cell depleting antibodies (anti-CD4 and anti-CD8 mAb) were given as part of the conditioning [9,10], costimulation blockade allowed the elimination of any irradiation from recipient conditioning if very high doses (mega doses) of allogeneic BM were transplanted [7,11]. Anti-CD40L is sufficiently effective alone (without CTLA4Ig) in strain combinations without minor antigen barriers (e.g., donor B10.A → recipient C57BL/10) [12,13] [8,14,18,19]. As long as no effective anti-CD40L mAb is available for clinical use, its elimination from recipient conditioning would be desirable.…”

“…While some form of (local) irradiation was universally required even when very profound doses of T-cell depleting antibodies (anti-CD4 and anti-CD8 mAb) were given as part of the conditioning [9,10], costimulation blockade allowed the elimination of any irradiation from recipient conditioning if very high doses (mega doses) of allogeneic BM were transplanted [7,11]. Anti-CD40L is sufficiently effective alone (without CTLA4Ig) in strain combinations without minor antigen barriers (e.g., donor B10.A → recipient C57BL/10) [12,13], in particular if CD8 + T cells are depleted [14], but CTLA4Ig is required for an optimal outcome in fully mismatched combinations (MHC plus minor histocompatibility antigen disparities) without T-cell depletion [15]. A second generation CTLA4Ig (belatacept) has since been approved for immunosuppression in renal transplant recipients [16].…”

“…Clonal deletion has mostly been assessed by following the frequency of T cells/thymocytes expressing certain Vβ subunits on their TCR [8,59]; this is done in donor-recipient strain combinations in which the presentation of endogenous retroviral superantigens requires donor MHC class II (e.g., Vβ5-and Vβ11-positive T cells recognize superantigen presented by donor I-E in the Balb/c to B6 combination). In selected murine studies, peripheral deletion of truly alloreactive CD4 + and CD8 + T cells was directly measured by following syngeneic T cells expressing a transgenic TCR for donor MHC class II or class I, respectively, which were adoptively transferred into the recipient before BMT with costimulation blockade [12,60]. Very recently, clonal deletion was reported to also occur in tolerant patients in a pilot study of combined kidney and HSC transplantation [61].…”