Fig. 2

In vivo tissue migration and in vitro chemotaxis of activated NK cells. Activated CD45.1+ NK cells were mixed 1:1 with freshly isolated (naïve) CD45.2 NK cells, stained with CFSE and i.v. transferred into C57BL/KaLwRij mice 3 weeks after tumor cell injection. NK cell number was determined after 18 h in BM (two tibias and femurs), spleen and blood by FACS analysis of CD45.1+ or CD45.2+ NK1.1+ cells within donor CFSE+ cells and normalized with the number of input cells (% of input cells). a) Dot plots show the gating strategy for the analysis of IL-15 activated donor NK cells in the spleen of MM-bearing mice. b) CFSE+ cells were enumerated in each organ and frequency of donor cells out of transferred (input) cells is shown as average ± SEM of 2 independent experiments n = 5 mice per groups. Right hand graph: BM homing of activated donor NK cells was normalized by donor cell frequency into spleen. c) Tissue migration of activated NK cells in healthy control (ctrl) and tumor-bearing mice (tum). One-way ANOVA with multiple comparison was performed to compare tissue distribution of activated cells and naïve cells (b) and of activated cells in ctr and tum (c). *p < 0.05; **p < 0.01

d) In vitro chemotaxis assay of activated or control (cells treated with IL-15 10 ng/ml) NK cells in response to medium alone (no chemokine), to CXCL10 (250 ng/ml) or to CXCL12 (200 ng/ml). Results show average ± SEM from 3 independent experiments. One-way ANOVA was performed to compare migration of activated cells versus control cells. *p < 0.05; **p < 0.01