Fig. 3

Expression of NGAL in calcifying VICs isolated from men and women donors undergoing AV replacement. Data are presented as scatter dot plots, mean ± SEM (shown in red colour). Cultured VICs were HP-challenged for 2, 4 and 8 days. Supernatants and protein cell lysates were harvested at the stated time points. Side-by-side sex comparison of the secreted NGAL (sNGAL) measured by ELISA in cell supernatants from calcifying male (n = 6) and female-derived VICs (n = 6). sNGAL was normalized to the total yield of protein cell monolayer (pg/µg) (A). Side-by-side sex comparison of the intracellular NGAL in arbitrary units (AU) upon normalization to the stain free (B). Fold-change analyses of the sNGAL expression in male-derived VICs (n = 6) (C) or female-derived VICs (n = 6) (D). Fold-change analyses of the intracellular NGAL in calcifying male-derived VICs (n=6) (E) or female-derived VICs (n = 6) (F). Band densitometries were normalized to stain free and are plotted as a fold-change relative to Control condition. Control condition within each sex was used as calibrator condition for statistical comparisons in C−F. AU arbitrary units, HP pro-calcifying high phosphate media conditioning (2.6 mM Pi), Ctrl control, NGAL Neutrophil Gelatinase-Associated Lipocalin. *p < 0.05; **p < 0.01; ****p < 0.0001