AIMS/HYPOTHESIS: 1,25-dihydroxyvitamin D(3), the active form of vitamin D, prevents Type 1 diabetes in non-obese diabetic (NOD) mice. Epidemiological data show a threefold increase in human Type 1 diabetes when vitamin D deficiency was present in the first months of life. To evaluate whether a similar dietary deficiency affects diabetes incidence in NOD mice, we generated NOD mice with vitamin D deficiency in early life. METHODS: Breeding pairs of NOD mice, as well as their offspring (test mice), were kept in surroundings devoid of ultraviolet light and were fed a vitamin D-depleted diet for 100 days. Mice were followed for 250 days. RESULTS: At 250 days, 35% (12/35) male and 66% (22/33) female vitamin D-deficient mice were diabetic compared to 15% (6/40, p=0.05) and 45% (13/29, p<0.01) of the control mice. At 100 days no difference in insulitis was seen, but more vitamin D-deficient mice were glucose intolerant. Higher IL1 expression was detected in islets of vitamin D-deficient mice and their peritoneal macrophages had an aberrant cytokine profile (low IL1 and IL6, high IL15). Thymus and lymph nodes of vitamin D-deficient mice contained less CD4(+)CD62L(+) cells. CONCLUSION/INTERPRETATION: Vitamin D status increases the expression of Type 1 diabetes in NOD mice. Our data in NOD mice, as well as human epidemiological data, point to the importance of preventing vitamin D deficiency in early childhood. Controlling this dietary factor could be an easy and safe way to reduce the incidence of Type 1 diabetes in subjects who are genetically at risk.