Fig. 1

OxHDL increases LOX-1 expression at the cellular plasma membrane through the activation of LOX-1 and NF-κB in EC. a–d HAEC and HUVEC were exposed to oxHDL (a, b) and HDL (c, d) for 12 and 24 h, and LOX-1 expression was analyzed. Representative images from western blot experiments for detection of LOX-1 in the presence of oxHDL (50 μg/ml) (a) or HDL (50 μg/ml) (c). b and d Densitometric analyses of the experiments shown in a and c, respectively. Protein levels were normalized against tubulin, and data are expressed normalized to 0 time (N = 5). Statistical differences for HAEC and HUVEC samples were assessed by a one-way analysis of variance (ANOVA) (Kruskal–Wallis) followed by Dunn’s post hoc test. *P < 0.05, **P < 0.01, NS: non-significant. Graph bars show the mean ± SD. LOX-1 expression was measured in HAEC and HUVEC in the plasma membrane-rich (e, f) or cytosol-rich (g, h) fractions from ECs exposed to oxHDL (50 μg/ml) and HDL (50 μg/ml) for 24 h. Representative images from western blot experiments performed for detection of LOX-1 in the EC membrane-rich (e) or cytosol-rich (g) fractions, and densitometric analyses of (e) and (g) are shown in (f) and (h), respectively. Protein levels were normalized against Na⁺ pump in the membrane-rich fraction and against tubulin in cytosol-rich fraction. Data are expressed normalized to vehicle condition (N = 3). Statistical differences for HAEC and HUVEC samples were assessed by a one-way analysis of variance (ANOVA) (Kruskal–Wallis) followed by Dunn’s post hoc test. *P < 0.05, **P < 0.01, NS: non-significant. Graphs show the mean ± SD. Ox-HDL-induced LOX-1 mRNA expression was determined by RT-qPCR in HAEC (i) and HUVEC (j). Data are expressed normalized to 0 time (N = 3). Statistical differences for HAEC and HUVEC samples were assessed by a one-way analysis of variance (ANOVA) (Kruskal–Wallis) followed by Dunn’s post hoc test. *P < 0.05, **P < 0.01. Graph bars show the mean ± SD. LOX-1 expression was measured in ECs in the absence (–) or presence (+) of LOX-1 inhibitor κ-carrageenan (250 μg/ml) (k, l), LOX-1 neutralizing anti-LOX (1:50) (m, n), or the NF-κB inhibitor SC-3060 (5 μM) (o, p) (N = 4). All inhibitors were added 1 h before and maintained throughout the treatment. Protein levels were normalized against Na⁺ pump, and data are expressed normalized to vehicle condition. Statistical differences were assessed by a two-way analysis of variance (ANOVA) followed by Tukey post hoc test. **P < 0.01. Graph bars show the mean ± SD