Extended Data Fig. 9: CXCL12 limits the proliferation of NK cells from healthy donors and patients with breast cancer with liver metastases.
From: Hepatic stellate cells suppress NK cell-sustained breast cancer dormancy
a, t-distributed stochastic neighbour embedding (t-SNE) plot showing the relative expression of CXCR4 on different liver cell types based on 8,444 human liver cells previously sequenced55. Each dot represents a single cell, and cells are coloured from lowest (yellow) to highest (purple) expression. b, Histogram of CXCR4 measured by flow cytometry on human NK-92 cells. c, Exogenous CXCL12 increases the number of G0–G1 resting NK-92 cells, but it has no effect on NK cell viability (n = 5 independent experiments; mean ± s.d.; two-tailed nonparametric Kruskal–Wallis test with Dunn’s multiple comparison post-hoc test). d, Schematic of experiments to test the effect of CXCL12 on blood-derived NK cells purified from healthy donors and patients with breast cancer (BC) with liver metastases. NK cells labelled with CellTrace Violet (CTV) were primed with IL-2 and IL-15, and then expanded with IL-2 in the presence of CXCL12 alone or combined with IL-15 until assessed for division profile. e, Representative histogram of the NK cell division profile of a healthy donor. f, Quantification of the division index (that is, the average number of cell divisions a cell has undergone) of blood-derived NK cells from healthy donors (left, n = 6) and patients with breast cancer with liver metastases (right, n = 6) after treatment with CXCL12 alone or combined with IL-15. C1–C3 correspond to different concentrations of recombinant CXCL12 (C1 = 0.02 μg ml−1, C2 = 0.2 μg ml−1 and C3 = 2 μg ml−1). Mean ± s.d.; two-tailed nonparametric Kruskal–Wallis test with Dunn’s multiple comparison post-hoc test. g, Experimental schematic to assess the effects of aHSC-secreted CXCL12 on liver NK cells. Mouse NK cells were treated with conditioned medium (CM) from liver-derived aHSCs in the presence of a function-blocking antibody against CXCL12 or a control IgG, and G0–G1 resting cells were quantified after EdU incorporation. h, Flow cytometry quantification of quiescent Ki67− NK cells in mouse liver milieus (n = 6 no tumour, n = 12 dormancy, n = 9 metastasis; data combine two independent experiments; mean ± s.d.; nonparametric two-tailed Kruskal–Wallis test with Dunn’s multiple comparison post-hoc test). i, Proliferation of CXCR4+ NK cells from metastatic milieus (n = 9 metastasis; mean ± s.d.; nonparametric Mann–Whitney U test).
