Diurnal metastasis of melanoma cells in lungs. (A) Experimental design to assess the effect of time of day in metastatic invasion of lungs by B16F1Luc melanoma cells. (B) Number of metastatic lesions 14 d after injection of B16F1Luc melanoma cells, as determined by the luminescence of lung explants, shown at bottom. Values are from a representative of two experiments, each with n = 3–5 mice per time point and experiment. (C) Representative H&E-stained sections of lungs from each time point. Bar, 100 µm. (D) Bioluminescence counts reflecting metastatic burden of lungs, 14 d after injection of B16F1Luc cells in the morning (AM, corresponding to ZT4) or in the evening (PM, corresponding to ZT16), in control mice or mice previously depleted of neutrophils. Values are normalized to AM; n = 9 mice from two independent experiments. Data are shown as mean ± SEM. *, P < 0.05; n.s., not significant, as determined by unpaired t test analysis.