The consumption of caffeine from some common beverages has been associated with low bone mass by inducing urinary calcium loss and deceasing bone mineral density. However, the effect of caffeine on osteoclast differentiation is still unclear. Here, we demonstrate that caffeine directly enhances osteoclast differentiation and maturation. TRAP staining showed that the number of larger (>. 100. μm) osteoclastic cells as well as of TRAP-positive multinucleated cells was increased by caffeine treatment. Among the MAP kinases, caffeine specifically activated p38 MAP kinase, which in turn, controlled osteoclast differentiation and maturation. This is evidenced by the abolishment of activated p38 MAP kinase by pretreatment with SB203580, a p38-specific inhibitor, resulting in suppressed osteoclast differentiation and maturation that should be increased by caffeine. Caffeine significantly induced the expression of Mitf and pretreatment with SB203580 markedly suppressed the expression of Mitf induced by caffeine. Whereas it failed to regulate the expression of NFATc1 and Oscar, the expressions of Cathepsin K and TRAP were induced by caffeine treatment in primary preosteoclasts. Real-time PCR and luciferase assays showed that the increase of osteoclastic cell-cell fusion by caffeine was through the transcriptional up-regulation of DC-STAMP expression but not of Atp6v0d2. These results strongly suggest that caffeine directly enhances osteoclast differentiation and maturation through p38 MAP kinase activation, thus inducing Mitf expression and transcriptional activation of DC-STAMP, and finally CtsK and TRAP.