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Vitamin D is considered to modulate T-cell function, which has been implicated in the treatment of inflammatory conditions. However, there is limited knowledge on the effects of vitamin D and its influences on circulating T-cell profiles in humans, particularly in overweight Black individuals who are more likely to be vitamin D insufficient (serum 25(OH)D concentrations of ≤20 ng/mL). Thus, this study tested the hypothesis that vitamin D supplementation modulates T-cell composition, which is in a dose-dependent manner. A 16-week randomized, double-blinded, placebo-controlled trial of vitamin D₃ supplementation was undertaken in 70 overweight/obese Black people (mean age = 26 years, 82% female) with 25 hydroxyvitamin D ≤ 20 ng/mL at baseline. Subjects were randomly assigned a supervised monthly oral vitamin D₃ equivalent to approximately 600 IU/day (n = 17), 2000 IU/day (n = 18), 4000 IU/day (n = 18), or a placebo (n = 17). Fresh peripheral whole blood was collected and CD3⁺, CD4⁺ and CD8⁺ cell counts and percentages were determined by flow cytometry at baseline and at 16 weeks, among 56 subjects who were included in the analyses. A statistically significant increase in CD3⁺% in the 2000 IU/day vitamin D₃ supplementation group, and increases in CD4⁺% in the 2000 IU/day and 4000 IU/day vitamin D₃ supplementation groups were observed (p-values < 0.05) from the changes in baseline to 16 weeks. Further adjustments for age, sex and BMI showed that 2000 IU/day vitamin D₃ supplementation increased in CD3⁺ count, CD3%, CD4 count, and CD4%, as compared to the placebo group (p …