Type I diabetes is an autoimmune T-cell-mediated disease associated with overexpression of inflammatory mediators and the disturbance of different T-cell subsets. Vasoactive intestinal peptide (VIP) is a potent anti-inflammatory agent with regulatory effects on activated T cells. As the equilibrium between different T-cell subsets is involved in the final outcome, leading to tolerance or autoimmunity, we studied the evolution of markers for T cells in nonobese diabetic (NOD) mice. The study of different transcription factors, cytokines or cytokine receptors, shows that VIP interferes with functional phase of T helper 17 (Th17) cells and prevents the increase in the proportion of Th1 to Th17 cells. On the other hand, VIP-treated NOD mice show an increase in the proportion of CD4(+)CD25(+) cells in the spleen. Thus, VIP switches the Tregs/Th17 ratio leading to tolerance in NOD mice. Similarly, VIP reverses the ratio of Th1-/Th2-cell subsets associated with autoimmune pathology. All these effects on the ratio of T-cell subsets and the anti-inflammatory effect of VIP in decreasing proinflammatory mediators result in a reduction of β-cell destruction in pancreas. Taken together, these results show that VIP provides significant protection against spontaneous diabetes by modulating T-cell subsets and counterbalancing tolerance and immunity.