Background: The aim of the current trial was to investigate the effect of Vitamin D treatment on metabolic markers in people with Vitamin D deficiency and thyroid autoimmunity. Materials and Methods: In this double-blind, randomized, placebo-controlled clinical trial, 65 Vitamin D deficient euthyroid or hypothyroid patients with positive TPO-Ab were enrolled. They randomly allocated into two groups to receive oral Vitamin D3 (50000 IU weekly) and placebo for 12 weeks. Serum concentration of calcium, phosphorus, albumin, C-reactive protein, blood urea nitrogen, creatinine, glycated hemoglobin (HbA1c), insulin, fasting plasma glucose (FPG), triglyceride (TG), total cholesterol, and high-density lipoprotein were measured in both groups before and after the trial. Homeostasis model assessment estimates of beta cell function (HOMA-B) and HOMA-insulin resistance (HOMA-IR) were calculated before and after trial in both groups. Results: Thirty-three and thirty-two participants were allocated to Vitamin D-treated and placebo-treated groups, respectively. Mean(standard error) level of Vitamin D increased significantly in Vitamin D-treated group (45.53[1.84] ng/mL vs. 12.76[0.74] ng/mL, P= 0.001). The mean of HbA1c and insulin was increased significantly both in Vitamin D-treated and placebo-treated groups (P< 0.05). Other variables did not meet a significant change after trial (P= NS). In between-group comparison, there was not any significant difference between Vitamin D-treated and placebo-treated groups regarding measures of HOMA-B, HOMA-IR, FPG, HbA1c, and TG(P= NS). Conclusion: Our findings showed that weekly 50000 IU oral Vitamin D3 for 12 weeks did not improve metabolic markers, IR, or insulin secretion in Vitamin D deficient patients with Hashimoto thyroiditis.