Induction of active tolerance after islet transplantation in longstanding diabetic NOD mice by L. lactis-based vaccine combined with low-dose anti-CD3

The long-term success of islet transplantation as a diabetes treatment depends on halting underlying recurrence of autoimmunity driven by pre-primed autoreactive effector T cells. Low-dose anti-CD3 monoclonal antibodies combined with live Lactococcus lactis secreting proinsulin and IL-10 (LL-PINS+IL-10) restore peripheral tolerance in new-onset diabetic non-obese diabetic (NOD) mice. Therapeutic success depends on residual ?-cell mass at time of intervention, but ?-cell mass rapidly declines at disease onset.  Thus, islet implantation may be used to both enlarge ?-cell mass during treatment, improving likelihood for success in early stage disease, and completely replace ?-cells in more advance disease. In NOD mice with existing autoimmunity and hyperglycemia, kidney capsular islet implantation was performed simultaneously with anti-CD3 treatment alone (monotherapy) or combined with LL-PINS+IL-10 (combination).  Islet-transplantation successfully maintained normoglycemia in 45% of combination treated mice, 28% in those under monotherapy (**p<0,01), and 5% in untreated controls (****p<0,0001 for both therapies). Disease duration prior to intervention in islet recipient mice also affected therapeutic success. Successful mono- and combination therapy protected ?-cell function of islet grafts and preserved remaining endogenous islets. Islet grafts from untreated mice and those where therapy failed were almost completely destroyed and showed striking lymphocytic infiltration. Remarkably, mononuclear infiltration of CD4+ and CD8+ T cells was also present in grafts from cured mice and to the same extent in both treatment groups. CD4+Foxp3+ regulatory T cells (Tregs) were increased in grafts from mice treated with either therapy compared to untreated controls and higher in combination cured vs monotherapy cured mice (**p<0,01).  Pathogenic CD8+IGRP+ effector T cells were blocked from early entry into the islet graft, a phenomenon persisting at least until 6 weeks post-transplantation in therapy-responsive mice. LL-based combination therapy induces peripheral tolerance in longstanding diabetic mice by inducing of Tregs and prevents graft infiltration by autoreactive CD8+ T cells.