Figure 3. Intestinal dendritic cells mediate cross talk between commensal bacteria and IL22⁺ILC3 innate lymphoid cells and induce ILC3 to traffic to the murine newborn lung.

(A) Representative flow cytometry histograms showing expression of C-C chemokine receptor (CCR) 4, 6, 7 and 9, C-X-C chemokine receptor (CXCR) 3 and 5 and C-C chemokine ligand (CCL) 20 by IL22⁺ILC3 innate lymphoid cells in the lung of postnatal day 4 (P4) ABX-free or ABX-exposed newborn mice. (B) An equal number of ILC3 from P4 WT or Ccr4^−/− newborn mice were adoptively transferred into age-matched ABX-exposed newborn mice, and the ability of ILC3 to traffic to the lungs (homing index) was determined. (C) Survival of P4 WT or Ccr4^−/− newborn mice that received adoptive transfer of WT ILC3 after infection with S. pneumoniae. (D) Representative flow cytometry plots and relative frequencies of distinct subsets of mononuclear phagocytes in the small intestine of P4 newborn mice. (E) The absolute numbers of IL22⁺ILC3 in the lungs of P4 WT or Zbtb46^DTR newborn mice treated with DT (CD11b⁺CD103⁺ DC-depleted) that received adoptive transfer of CD11b⁺CD103⁺ DCs. (F) Survival of P4 WT or Zbtb46^DTR newborn mice treated with DT (CD11b⁺CD103⁺ DC-depleted) that then received adoptive transfer of CD11b⁺CD103⁺ DCs, after infection with S. pneumoniae . (G) ILC3 isolated from lungs of P4 ABX-exposed mice were co-cultured with CD11b⁺CD103⁺ DCs isolated from age-matched ABX-exposed or ABX-free mice and examined for surface expression of various chemokine receptors. A representative flow cytometry plot is shown and (H) relative frequencies of IL-22⁺ILC3 cells expressing CCR4. (I) ILC3 isolated from lungs of P4 ABX-exposed mice were co-cultured either alone or with CD11b⁺CD103⁺ DCs isolated from age-matched ABX-exposed or ABX-free newborn mice. The ability of these ILC3 to migrate in vitro in response to a gradient of the chemokine ligand (CCL) 17 is shown. (J) The absolute numbers of IL22⁺ILC3 in the lungs of P4 Zbtb46^DTR newborn mice either exposed to ABX or no ABX that were then treated with DT (CD11b⁺CD103⁺ DC-depleted) or no DT (no DC depletion) before they were reconstituted with commensal bacteria. Data and plots are representative of three independent experiments. Results are shown as the mean ± s.e.m (Student’s t-test or ANOVA or Wilcoxon signed-rank test,). *P ≤ 0.05; **P ≤ 0.01. Number of individual animals [n] are indicated.