Tumor necrosis factor alpha (TNF-alpha) is a proinflammatory cytokine, which participates in a wide range of immunoregulatory activities. It is generally produced at highest levels by cells of the myeloid lineage in response to activation of pathogen recognition receptors such as Toll-like receptors. Impaired production predisposes to infection with intracellular organisms, and overproduction results in systemic or organ-specific inflammation. Control of expression is essential to maintain homeostasis, and this control is mediated via multiple strategies. We examined two separate aspects of chromatin accessibility in this study of the human TNF-alpha promoter. We examined the role of histone acetylation and chromatin remodeling in cell lines and primary cells and identified two individual steps associated with activation of TNF-alpha production. Histone H3 and H4 acetylation was found to be strongly dependent on the developmental stage of human monocytes. It did not appear to be regulated by acute stimuli, and instead, chromatin remodeling was found to occur after acute stimuli in a cell line competent to produce TNF-alpha. These data suggest that there is a hierarchy of controls regulating expression of TNF-alpha. Acetylation of histones is a prerequisite but is insufficient on its own for TNF-alpha production.