The success of anti-tumor necrosis factor alpha (TNFα) therapies has led to increased interest as to the mechanisms and consequences of TNFα blockade. The aim of the study was to examine the effects of TNFα blockade by etanercept on lipopolysaccharide (LPS) or peptidoglycan (PG)-induced maturation of human monocyte-derived dendritic cells (MDDCs). MDDCs grown from peripheral blood of healthy donors were stimulated by LPS or PG with/without the presence of etanercept. Concentrations of TNFα in cell supernatants were assessed by ELISA, while the cells were stained with monoclonal antibodies to CD83, CD80, CD86, CD11c, CD40, HLA-DR, and annexin-V and acquired using a flow cytometer. Etanercept significantly decreased the stimulated cell surface expression of HLA-DR, CD80, CD86, CD40 and CD83 on MDDCs in all examined samples. Etanercept in the same dose, but denatured to loss of specificity for TNFα, failed to change any of the aforementioned markers. In the presence of etanercept, concentrations of TNFα in cell supernatants were decreased by 53% on average, with a range of 25%–87%. Etanercept impaired the stimulated maturation of MDDCs by neutralizing the induced TNFα, produced by the same MDDCs after antigenic stimulation. The reported data confirms that TNFα blockade may have a direct effect on DCs, with a wide spectrum of potential secondary effects downstream. The data also suggests the presence of TNFα-mediated autocrine signaling, serving to accelerate or catalyze the maturation process of MDDCs.