Abstract This study was designed to evaluate the potential therapeutic efficacy of vitamin D (Vit D) in averting the harmful effects of type 2 diabetes mellitus (T2D). Forty male Wistar rats were allotted into four groups: (1) the control, (2) Vit D, (3) streptozotocin (STZ), and (4) STZ + Vit D groups. Rats co-treated with Vit D had significantly (p < 0.05) decreased levels of cortisol; proinflammatory cytokines, including interleukin-6 (IL-6); and malondialdehyde (MDA). Meanwhile, the levels of insulin significantly (p < 0.05) increased, whereas the activity of the antioxidant system, including glutathione (GSH), superoxide dismutase (SOD), catalase, and total antioxidant capacity (TAC), significantly (p < 0.05) decreased. Histopathological examination revealed the destruction of beta cells in the islets of Langerhans in rats with diabetes. Meanwhile, immunoexpression revealed an increase in the immunoreactivity of caspase-3 and endothelial nitric oxide synthase and a reduction in the immunoreactivity of insulin in rats with diabetes. In conclusion, Vit D ameliorated the harmful biochemical impact of diabetes mellitus, probably by increasing insulin secretion and sensitivity, ameliorating β-cell function, and decreasing cortisol levels; also, the anti-inflammatory effect of Vit D reduces the number of proinflammatory cytokines (e.g., IL-6) and increases the activity of the antioxidant system, such as GSH, SOD, TAC, and catalase while reducing lipid peroxidation enzymes (e.g., MDA).