Fig. 3

LCN2 promotes angiogenesis in BM from lung cancer patients. a Pathway enrichment analysis of lung cancer tumor cells with high versus low LCN2 expression. b Heatmap of cytokine antibody arrays in PC9-BM control and PC9-BM KD cells, as well as A549 control and A549 OE cells. Each group included three biological replicates. The data are presented as log fold changes and were normalized to the corresponding cell line with lower LCN2 expression. *FDR < 0.05, **FDR < 0.01; P values were adjusted via the Benjamini‒Hochberg method. c Western blot analysis of LCN2 expression in PC9-BM control, PC9-BM KD, A549 control, and A549 OE cells. d ELISA quantification of LCN2 levels in PC9-BM control, PC9-BM KD, A549 control, and A549 OE cells (n = 3). The data are presented as the means ± SDs; two-sided t test, ****P < 0.0001. e Quantification of junction and branch points per field (n = 3) of angiogenesis in PC9-BM control, PC9-BM KD, A549 control, and A549 OE cells. The data are presented as the means ± SDs; two-sided t test, * P < 0.05, ** P < 0.01, ***P < 0.001. f Representative immunohistochemistry (IHC) images of CD34 expression. The quantified data are shown as the means ± SDs (n = 3); two-sided t-test, **P < 0.01. Scale bars, 200 μm. g Western blot analysis of phosphorylated JAK2 (p-JAK2), phosphorylated STAT3 (p-STAT3), and VEGF-A. h Western blot analysis of the indicated cells treated with or without 100 ng/mL recombinant human LCN2 (rhLCN2), with or without the STAT3 inhibitor SH4-54 (1 μM). i The tumor inhibition rate and survival analysis were performed by normalizing the tumor burden to that of vehicle-treated BM-bearing mice. LCN2 lipocalin-2, BM brain metastases, IHC immunohistochemistry