CD103 or CX 3 CR1 surface expression defines distinct dendritic cells (DCs) and macrophages in the murine lamina propria of the colon (cLP). We investigated the surface marker and functional phenotype of CD103+ and CX 3 CR1+ cLP DCs and their role in transfer colitis. cLP CD11c+ cells were isolated from specific pathogen-free or germ-free mice to elucidate the role of the commensal flora in their development. The cLP CD11c+ cells are a heterogeneous cell population that includes 16% CX 3 CR1+, 34% CD103+, 30% CD103− CX 3 CR1− DCs, and 17% CD68+/F4/80+ CX 3 CR1+ CD11c+ macrophages. All DCs expressed high levels of MHC II but low levels of costimulatory (CD40, CD86, and CD80) and coinhibitory (programmed death ligand-1) molecules. Ex vivo confocal microscopy demonstrated that CX 3 CR1+ CD11c+ cells, but not CD103+ DCs, were reduced in the cLP of germ-free (CX 3 CR1-GFP) mice. The absence of the enteric flora prevents the formation of transepithelial processes by the CX 3 CR1+ DCs. CX 3 CR1+ DCs preferentially supported Th1/Th17 CD4 T cell differentiation. CD103+ DCs preferentially induced the differentiation of Foxp3-expressing regulatory T cells. The stimulation of cLP DCs with fractalkine/CX 3 CL1 increased the release of IL-6 and TNF-α. In the absence of CX 3 CR1, the CD45RB high CD4 transfer colitis was suppressed and associated with reduced numbers of DCs in the mesenteric lymph nodes and a reduction in serum IFN-γ and IL-17. The local bacteria-driven accumulation of CX 3 CR1+ DCs seems to support inflammatory immune responses.