Figure 1: ANGPTL4 is expressed in macrophages accumulated in atherosclerotic plaques.

(a) Heat map representation of gene expression from microarray data comparing mouse peritoneal macrophages incubated with or without Ac-LDL (120 μg ml⁻¹) for 24 h. Left panel shows upregulated genes (P<0.05, unpaired t-test; fold change≥1.4) and right panel shows downregulated genes (P<0.05, unpaired t-test; fold change≥2.08) upon Ac-LDL treatment compared with non-treated cells. Samples were analysed in triplicate. (b) qRT-PCR validation of selected genes upregulated in the microarray in mouse peritoneal macrophages treated with or without Ac-LDL for 24 h. qRT-PCR analysis of Angptl4 expression levels in macrophages (c, left) and whole aorta (c, right) from WT and Ldlr^−/− mice fed a WD (left) (n=4 per group), and in human peripheral blood mononuclear cells (d) treated with or without Ac-LDL (120 μg ml⁻¹) for 24 h. Abca1 and Hmgcr genes were used as control genes for cholesterol loading and Cd68 was used as a marker for macrophages. All data represent the mean±s.e.m. and are representative of three experiments in duplicate; *P≤0.05 compared with untreated macrophages (b,d) and WT mice (c) by unpaired t-test. (e) Immunostaining of ANGPTL4 (red) and macrophage marker CD68 (green) and their co-localization in atherosclerotic plaques of Ldlr^−/− mice fed a WD. Scale bar, 400 μm. (f) Immunohistochemistry staining of ANGPTL4 and CD68 in human atherosclerotic plaques. Scale bar, 200 μm. (g) Comparison of ANGPTL4 mRNA expression in NCP and corresponding culprit (CP) human atherosclerotic plaques (n=9). (h) Representative western blot showing comparison of ANGPTL4 expression in NCP and corresponding CP. Lower panel shows densitometry analysis for the 50 kDa bands of the western blots (n=7). *P≤0.05 compared with NCP by unpaired t-test; a.u. arbitrary units. Full scans of westerns blots are provided in Supplementary Fig. 7.