Reciprocal cross-talk between MD-DCs and activated NK/LAK cells.

Cocultures were performed as described briefly in Figure 5. Immunophenotyping of 1:1 ratio cocultures of NK cells/iDCs, NK cells/iDCs with LPS, and NK cells/mDCs were performed by 4-color flow cytometry with the combinations of FITC-, PE-, PerCP-, and APC-conjugated mAbs CD86/CD83/HLA-DR/CD45 and CD16/CD3/CD8/CD56. Left panels show the cellular morphology according to FSC/SSC criteria, allowing the identification of 3 cell subpopulations: DC1 cells, DC2 cells, and CD56⁺CD3⁻ NK/LAK cells. Center panels show a dot plot analysis of CD86/CD83 staining after gating on CD45⁺HLA-DR⁺ cells (not shown). Percentages of DC1 CD86^bright/CD83^bright and DC2 CD86^dim/CD83^dim are reported; these 2 subsets of DCs are identified by color backgating on the FSC/SSC dot blots. Right panels show a dot plot analysis of CD56/CD16 staining after gating on CD56⁺CD3⁻ cells (not shown), whose percentages are reported on the corresponding FSC/SSC dot plots. Percentages of CD56⁺CD16⁺ are reported. This shows a representative experiment performed in 1 patient out of 2. (A) Results of immunophenotyping at day 3. At this early time point, NK/LAK cells turn iDCs into maturing DCs exhibiting a CD86⁺/CD83⁺ phenotype, which is further enhanced with addition of LPS. (B) Results of immunophenotyping at day 7. At this late time point, survival of CD56⁺CD3⁻ is significantly increased in the NK cells/iDCs with LPS.