CCR2⁺CD103⁻ intestinal dendritic cells develop from DC-committed precursors and induce interleukin-17 production by T cells

The identification of intestinal macrophages (m phi s) and dendritic cells (DCs) is a matter of intense debate. Although CD103(+) mononuclear phagocytes (MPs) appear to be genuine DCs, the nature and origins of CD103(-) MPs remain controversial. We show here that intestinal CD103(-)CD11b(+) MPs can be separated clearly into DCs and m phi s based on phenotype, gene profile, and kinetics. CD64(-)CD103(-)CD11b(+) MPs are classical DCs, being derived from Flt3 ligand-dependent, DC-committed precursors, not Ly6C hi monocytes. Surprisingly, a significant proportion of these CD103(-)CD11b(+) DCs express CCR2 and there is a selective decrease in CD103(-)CD11b(+) DCs in mice lacking this chemokine receptor. CCR2(+)CD103(-) DCs are present in both the murine and human intestine, drive interleukin (IL)-17a production by Tcells in vitro, and show constitutive expression of IL-12/IL-23p40. These data highlight the heterogeneity of intestinal DCs and reveal a bona fide population of CCR2(+) DCs that is involved in priming mucosal T helper type 17 (Th17) responses

Isolation of Rat Intestinal Lymph DC

Dendritic cells (DCs) migrate constitutively from the intestine via the lymph to the mesenteric lymph nodes. These migrating intestinal lymph DCs (ilDCs) carry antigens acquired in the intestine and play important roles in both the initiation of immune responses and the maintenance of oral tolerance. The ilDC population is made up of at least three functionally different DC subsets. Like many DC populations, ilDCs are exquisitely sensitive to their environment, changing their phenotype and maturing in response to the procedures associated with their extraction from solid tissues. We have developed and refined a method for collecting and purifying these DC subsets from rats, without inducing them to mature. This method involves two separate surgical procedures, separated by at least 6 weeks. Initially, mesenteric lymph nodes are removed. After the animals have fully recovered we cannulate the thoracic duct and collect the ilDCs on ice, minutes after they have left the lymph vessel. The DCs are then enriched using magnetic beads and purified by flow cytometric sorting. We describe this method here, including our recent refinements to limit the use of the restraining "Bollman" cage