Abstract Background Glucocorticoids are widely used in treating inflammatory diseases. The contribution of adrenal glucocorticoids to inflammatory regulation is unknown. Endogenous glucocorticoids, as distinct from synthetic analogues, not only suppress but also enhance immune functions. Elevated circulating cortisol levels are characteristic of injured patients. In a model of trauma, an early glucocorticoid surge occurs concomitantly with decreased cellular cytokine responses. Cytokine production elevated late after injury is associated with increased mortality. We hypothesized that this glucocorticoid surge mediates the later heightened macrophage responses. Materials and methods The murine macrophage like cells RAW 264.7 were incubated with corticosterone (35 ng/mL), or vehicle control, for 1 h, after which the cells were washed and corticosterone-free medium added. At 0, 3, 6, 12, and 24 h after removal of the corticosterone, the cells were stimulated with lipopolysaccharide (LPS) and interferon-γ. Supernatant tumor necrosis factor (TNF)-α, interleukin (IL)-6, and nitrite levels were measured. In separate experiments the effect of pretreatment with corticosterone on TNF-α, IL-6, and nitrite mRNA expression as well as nuclear factor-κB and glucocorticoid receptor activity was determined. CD14 receptor expression was determined by flow cytometry. Results Glucocorticoid pretreatment caused significantly increased RAW 264.7 cell production of nitrite, IL-6 and TNF-α. mRNA for these inflammatory mediators was induced 6 h after the corticosterone pretreatment, and was associated with activation of nuclear factor-κB in the presence of activated glucocorticoid receptor. Cell surface-expression of CD14 was likewise increased. Conclusions The results of this study demonstrate a novel role for glucocorticoids and provide a mechanism for the late upregulation in macrophage function after injury.