Immune modulators can arrest loss of insulin secretion in type 1 diabetes mellitus (T1DM), but they have not caused permanent disease remission or restored normal insulin secretion. We tested whether exendin-4, a glucagon-like peptide-1 receptor agonist, would enhance remission of T1DM in NOD mice treated with anti-CD3 monoclonal antibody (mAb) and studied the effects of exendin-4 treatment on cellular and metabolic responses of beta-cells. Diabetic NOD mice treated with anti-CD3 mAb and exendin-4 had a higher rate of remission (44%) than mice treated with anti-CD3 mAb alone (37%) or exendin-4 (0%) or insulin or IgG alone (0%) (P < 0.01). The effect of exendin-4 on reversal of diabetes after anti-CD3 mAb was greatest in mice with a glucose level of less than 350 mg/dl at diagnosis (63 vs. 39%, P < 0.05). Exendin-4 did not affect beta-cell area, replication, or apoptosis or reduce the frequency of diabetogenic or regulatory T cells or modulate the antigenicity of islet cells. Reversal of T1DM with anti-CD3 mAb was associated with recovery of insulin in glucose transporter-2(+)/insulin(-) islet cells that were identified at diagnosis. Glucose tolerance and insulin responses improved in mice treated with combination therapy, and exendin-4 increased insulin content and insulin release from beta-cells. We conclude that treatment with glucagon-like peptide-1 receptor agonist enhances remission of T1DM in NOD mice treated with anti-CD3 mAb by enhancing the recovery of the residual islets. This combinatorial approach may be useful in treatment of patients with new-onset T1DM.