Fig. 3
Increased Id2 or Aldh3a1 expression enhances metastatic colonisation of the brain following intracardiac inoculation. a 4T1-Luc or MDA-MB-231 cells with ectopic expression of Id2 or Aldh3a1 or transduced with mCherry vector (Vec) alone. Left panels show RT-qPCR analysis of Id2 and Aldh3a1 expression. Right panel shows Id2 and Aldh3a1 protein levels assessed by immunoblotting. b BALB/c mice inoculated via the left ventricle with 5 × 104 4T1-Luc cells, n = 7 or 8 mice per group. At day 9, whole brains were collected at necropsy and subject to ex-vivo IVIS imaging. Mean ± SEM. *p < 0.05, Kruskal-Wallis test. Right panel shows representative images of mCherry fluorescent tumour cells in brain sections. Scale bar = 100 μm. c 3 × 105 MDA-MB-231-Luc cells were inoculated into the left ventricle of NSG mice, n = 9 to 10 mice per group and the experiment terminated on day 23, with organs subject to ex-vivo IVIS imaging. Shown are quantification of ex-vivo IVIS signals in the lungs (left panel, p = ns) and brains (middle panel, *p < 0.05, **p < 0.01), Kruskal-Wallis test. Right panel shows representative IVIS images of brains. d Sections of fixed and paraffin-embedded brains from c were co-stained for endothelial cell markers endomucin and CD31 (both in blue) and the human nuclear envelope marker lamin A/C (brown). Scale bar = 100 μm. Arrowheads indicate endomucin/CD31 stained vasculature. ns not significant