Propionibacterium acnes is a major etiological factor of acne, triggering an inflammatory response in part through the activation of TLR2. In this study, we demonstrate that activation of peripheral blood monocytes with P. acnes in vitro induced their differentiation into two distinct innate immune cell subsets, CD209(+) macrophages and CD1b(+) dendritic cells. Furthermore, P. acnes induced expression of mRNA for the cytokines IL-15 and GM-CSF, which differentiate CD209(+) and CD1b(+) cells, respectively. The CD209(+) cells were more effective in uptake of P. acnes, compared with the CD1b(+) cells, and demonstrated a 2-fold greater antimicrobial activity against the phagocytosed bacteria. Although CD1b(+) cells secreted inflammatory cytokines in response to both P. acnes and a TLR2 ligand control, the CD209(+) cells responded only to P. acnes. The addition of all-trans retinoic acid, a commonly used agent for the treatment of acne, directly induced differentiation of monocytes into CD209(+) macrophages and enhanced the P. acnes-mediated differentiation of the CD209(+) subset. Therefore, the differentiation of monocytes into CD209(+) macrophages and CD1b(+) dendritic cells distinctly mediate the innate immune response to P. acnes.