Extended Data Fig. 9: Effects of neutrophil depletion or augmentation in mice.
From: Neutrophils escort circulating tumour cells to enable cell cycle progression
a, Plots showing the mean number of neutrophils in the circulation of mice treated with Ly-6G neutralizing antibodies (anti-Ly-6G) (left), or carrying G-CSF overexpressing tumours (right). Error bars, s.e.m.; the number of independent biological replicates (n) is provided in the Source Data; NA, not available; *P < 0.03, **P < 0.0001 by two-sided Student’s t-test. b, Representative images of the primary tumour of NSG-LM2-GFP mice stained for pan cytokeratin (pCK, green), myeloperoxidase (MPO, gold) and DAPI (nuclei, blue) (left). The plots show the mean number of infiltrated neutrophils per field of view within the tumour (right). W, weeks upon tumour development. Error bars, s.e.m.; n = 3; *P < 0.03, **P < 0.0001 by two-sided Student’s t-test. c, Tumour growth curves representing mean tumour volume measurements in the presence or absence of anti-Ly-6G antibodies or G-CSF overexpression. Error bars, s.e.m.; the number of independent biological replicates (n) is provided in the Source Data; ns, not significant by two-sided Student’s t-test. d, Plots showing the mean counts of single CTCs, CTC clusters and CTC-neutrophil clusters in mice. Error bars, s.e.m.; the number of independent biological replicates (n) is provided in the Source Data; ns, not significant; ND, not detected; *P < 0.05 by two-sided Student’s t-test. e, Pie charts displaying the mean percentage of single CTCs (grey), CTC clusters (green) and CTC–neutrophil clusters (gold) in NSG-LM2-GFP and NSG-CDX-BR16-GFP mice treated with anti-Ly-6G antibodies or G-CSF overexpression. W, weeks upon tumour development; the number of independent biological replicates (n) is shown for each condition. f, Plots showing the mean fold change of CTC ratios from NSG-LM2-GFP and NSG-CDX-BR16-GFP mice treated with anti-Ly-6G antibodies or G-CSF overexpression. Error bars, s.e.m.; the number of independent biological replicates (n) is provided in the Source Data; *P = 0.045, **P = 0.01, ***P = 0.004 by two-sided Student’s t-test. g, Representative bioluminescence images of lungs from mice treated with anti-Ly-6G antibodies or G-CSF overexpression (left); the number of independent biological replicates (n) is provided for simplicity directly within the Source Data; W, weeks upon tumour development. The plots show the mean metastatic index of mice treated with anti-Ly-6G antibodies or G-CSF overexpression (right). The number of independent biological replicates (n) is provided for simplicity directly within the Source Data; error bars, s.e.m.; *P < 0.03 **P < 0.01 by two-sided Student’s t-test. h, Kaplan–Meier survival plots showing overall survival rates of mice. The number of independent biological replicates (n) is provided in the Source Data; *P < 0.02 by two-sided log-rank test. i, Schematic of the experiment. NSG, FVB and BALB/c mice were pre-treated with anti-Ly-6G antibodies or control IgG. 4T1-GFP cells or Py2T-GFP cells were then injected into the tail vein to assess metastasis development. j, Plots showing mean normalized bioluminescence signal in the lungs of injected mice. k, Kaplan–Meier survival plot of injected mice. l, Plots showing the mean percentage of Ki67-positive disseminated tumour cells (DTCs) collected from the bone marrow of injected mice. n = 3; error bars, s.e.m.; ns, not significant by two-sided Student’s t-test (j–l). m, Bar graph showing the proportion of patients with breast cancer who were treated with G-CSF, related to their CTC status. n = 42 for no CTCs, n = 23 for CTCs, n = 9 for CTC–neutrophil clusters; P value by two-sided Fisher’s exact test is shown.
