The present study investigated the effects of vitamin D deficiency on T cell subsets in patients with spinal tuberculosis. In addition, the influence of vitamin D deficiency was investigated on the expression of cytokines IL-1β, IL-6 and TNF-α in intervertebral disc lesions of patients. One hundred and seventeen patients with spinal tuberculosis who received operative treatment in the Department of Orthopedics in Wuhan City Third Hospital from March 2012 to March 2015 were collected. The patients were divided depending upon vitamin D content into the control group (64 cases, vitamin D content <25 nmol/l) and experimental group (53 cases, vitamin D content >50 nmol/l). Immunofluorescence method was applied to determine the content of T cell subsets in both groups of patients. Intervertebral disc lesion tissues of two groups of patients were obtained during surgery then treated with HE staining and immunohistochemical staining. The values of average optical density obtained under light microscope were observed as the expression quantities of IL-1β, IL-6 and TNF-α, to explore the relationship between vitamin D and the expression of cytokines. When vitamin D is lacking, the expression of T lymphocyte subsets in patients with spinal tuberculosis significantly decreased. Compared with experimental group, the difference was statistically significant (P<0.05). Further, the expression of cytokines IL-1β, IL-6 and TNF-α in intervertebral disc lesion tissues of patients with spinal tuberculosis were significantly higher than those of patients with spinal tuberculosis whose vitamin D content was normal (P<0.05). In the control group, vitamin D content was negatively correlated with the expression of IL-1β, IL-6 and TNF-α. The expression of T lymphocyte subsets in patients with vitamin D deficiency was significantly reduced, and the immune function decreased. The expression of IL-1β, IL-6 and TNF-α in lesions were significantly higher than those of patients with normal vitamin D content. In addition, the lower the content of vitamin D was, the more active the expression of inflammatory factors were, which was not conducive to the recovery of tuberculosis lesions.