We identified TIM-1 as an inclusive marker for IL-10+ regulatory B cells (TIM-1+ Bregs), which prolong islet allograft survival (GS). We recently found that TIM-4, a ligand for TIM-1, is expressed on a subset of B cells enriched for IFN-γ+ B cells (B effector 1; Be1). Adoptive transfer of WT TIM-4+ B cells from alloimmunized mice into B cell-deficient μMT (B6) recipients accelerates rejection of BALB/c islets compared to control recipients with no B cell transfer (MST 11d vs. 15 d; p<0.05), augments Th1 and Th17 responses. In contrast, transfer of alloimmunized IFN-γ-/- TIM-4+ B cells into μMT recipients prevents accelerated rejection and GS is prolonged (MST 33d vs. control 15d; p<0.05). Thus, IFN-γ plays a role in Be1 cell function.

Now we show that TIM-4+B cells also express IL-17. In alloimmunized mice (d7), ∼15% of TIM-4+ B cells expressed IL-17 (7-fold enriched compared to TIM-4– B cells, and 4-fold enriched compared to CD4 cells). Transfer of IL-17-/- TIM-4+ B cells from alloimmunized B6 mice into μMT recipients of BALB/c islets induced long-term GS in >85% of recipients (MST >100d; p<0.01). indicating that TIM-4+B cells exhibit potent Breg activity in the absence of IL-17. Moreover, mixed bone marrow chimeric “B-IL-17−/−” mice specifically lacking IL-17 in B cells (20% IL17-/- BM:80% μMT BM in irradiated μMT mice) exhibit markedly prolonged GS compared to control “B-WT” chimeras (MST; B-IL-17−/− 45d with 33% long-term GS vs. B-WT MST 20d; p<0.05). This implies that IL-17 production by B cells is a major contributor to transplant rejection.

To study the role of B cell IL-17 in T cell responses in vivo, CFSE labeled OTII CD4+ T cells were adoptively transferred into B-IL-17−/− vs. B-WT chimeric mice and their response to immunization with CB6F1 allogeneic cells expressing act-mOVA was examined. Compared to their responses in B-WT mice, OTII cells in B-IL-17−/− chimeric hosts exhibit impaired proliferation (1.5×⇓, p<0.01), reduced Th1 and Th17 differentiation (IFNγ and IL-17 both ⇓2.5X, p<0.05), and increased Th2 and IL-10 differentiation (IL-4 ↑2X and IL-10, ↑1.8X; p<0.05). Similar findings were observed with transferred OTI cells (30%⇓ proliferation, 60% ⇓IFNγ, 60%⇓IL-17 and 25%↑IL-10 and 50%↑IL-4; p<0.05)

Collectively, our results indicate that B cell IL-17 plays a critical role promoting proinflammatory Teff responses that lead to rejection. Targeting TIM-4+ B cells and/or B cell IL-17 may reduce rejection and unleash potent Breg activity.

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