Abstract Mycophenolic acid (MPA) is an immunosuppressive drug which induces resistance to several maturation signals in human dendritic cells (DC) by unknown mechanisms. As mitogen-activated protein kinases (MAPK) are involved in the maturation process, we studied whether MPA affected p38MAPK and extracellular signal-regulated kinase (ERK1/2) in human DC. We first showed that MPA reduced TNFα-induced phenotype maturation, whereas it had no effect after LPS activation, suggesting that MPA preferentially affects the signaling pathway used by TNFα. We found that TNFα preferentially used p38MAPK to induce phenotype maturation in DC, whereas LPS preferentially activated NF-κB. Importantly, we showed that MPA more strongly inhibited p38MAPK phosphorylation induced by TNFα than by LPS. This difference in inhibition may therefore explain its different effect on DC phenotype. Interestingly, MPA inhibited the inflammatory cytokine synthesis and allostimulatory capacity induced by both stimuli. Exogenous guanosine antagonized the effect of MPA on the phenotype of TNFα-matured-DC as well as the IL-12p70 and IFNγ secretion induced by both stimuli, without affecting p38MAPK phosphorylation. The action of MPA on human DC phenotype maturation appears mainly to be due to its ability to inhibit p38MAPK. Furthermore, the difference between LPS and TNFα emphasizes that the DC microenvironment strongly influences DC sensitivity to MPA.