In older persons, vitamin D insufficiency and a subclinical chronic inflammatory status frequently coexist. Vitamin D has immune-modulatory and in vitro anti-inflammatory properties. However, there is inconclusive evidence about the anti-inflammatory role of vitamin D in older subjects. Thus, we investigated the hypothesis of an inverse relationship between 25-hydroxyvitamin D (25(OH)D) and inflammatory markers in a population-based study of older individuals. After excluding participants with high-sensitivity C-reactive protein (hsCRP) ≥ 10 mg/dl and those who were on chronic anti-inflammatory treatment, we evaluated 867 older adults ≥65 years from the InCHIANTI Study. Participants had complete data on serum concentrations of 25(OH)D, hsCRP, tumor necrosis factor (TNF)-α, soluble TNF-α receptors 1 and 2, interleukin (IL)-1β, IL-1 receptor antagonist, IL-10, IL-18, IL-6, and soluble IL-6 receptors (sIL6r and sgp130). Two general linear models were fit (model 1-adjusted for age, sex, and parathyroid hormone (PTH); model 2-including covariates of model 1 plus dietary and smoking habits, physical activity, ADL disability, season, osteoporosis, depressive status, and comorbidities). The mean age was 75.1 ± 17.1 years ± SD. In model 1, log(25OH-D) was significantly and inversely associated with log(IL-6) (β ± SE = -0.11 ± 0.03, p = <0.0001) and log (hsCRP) (β ± SE = -0.04 ± 0.02, p = 0.04) and positively associated with log(sIL6r) (β ± SE = 0.11 ± 0.04, p = 0.003) but not with other inflammatory markers. In model 2, log (25OH-D) remained negatively associated with log (IL-6) (β ± SE = -0.10 ± 0.03, p = 0.0001) and positively associated with log(sIL6r) (β ± SE = 0.11 ± 0.03, p = 0.004) but not with log(hsCRP) (β ± SE = -0.01 ± 0.03, p = 0.07). 25(OH)D is independently and inversely associated with IL-6 and positively with sIL6r, suggesting a potential anti-inflammatory role for vitamin D in older individuals.