Murine polymicrobial sepsis is associated with a sustained reduction of dendritic cell (DC) numbers in lymphoid organs and with a dysfunction of DC that is considered to mediate the chronic susceptibility of post-septic mice to secondary infections. We investigated whether polymicrobial sepsis triggered an altered de novo formation and/or differentiation of DC in the bone marrow. BrdU labeling experiments indicated that polymicrobial sepsis did not affect the formation of splenic DC. DC that differentiated from bone marrow (bone marrow-derived DC [BMDC]) of post-septic mice released enhanced levels of IL-10 but did not show an altered phenotype in comparison with BMDC from sham mice. Adoptive transfer experiments of BMDC into naive mice revealed that BMDC from post-septic mice impaired Th1 priming but not Th cell expansion and suppressed the innate immune defense mechanisms against Pseudomonas bacteria in the lung. Accordingly, BMDC from post-septic mice inhibited the release of IFN-γ from NK cells that are critical for the protection against Pseudomonas. Additionally, sepsis was associated with a loss of resident DC in the bone marrow. Depletion of resident DC from bone marrow of sham mice led to the differentiation of BMDC that were impaired in Th1 priming similar to BMDC from post-septic mice. Thus, in response to polymicrobial sepsis, DC precursor cells in the bone marrow developed into regulatory DC that impaired Th1 priming and NK cell activity and mediated immunosuppression. The absence of resident DC in the bone marrow after sepsis might have contributed to the modulation of DC differentiation.