Fig. 1

Effect of 4-PBA on secretion of rFVII-160R and rFVIIwt. Cells stably expressing rFVII-160R or rFVIIwt were treated with 0 (untreated), 5, 7.5 or 10 mM 4-PBA for 48 h. a–d FVII:Ag levels in conditioned medium (secreted) or cell lysates (intracellular) were examined by ELISA. To adjust for variations in cell number, the FVII:Ag levels were adjusted to the corresponding total protein content in the respective wells. a FVII:Ag levels in conditioned medium or cell lysates from untreated cells expressing rFVIIwt (black bars) or rFVII-160R (white bars). Values are reported as mean ± SD from at least three independent experiments performed in duplicates (unpaired t-test, ****p ≤ 0.0001). b Ratio between secreted/intracellular FVII:Ag levels in untreated cells. Values are reported as mean ± SD from at least three independent experiments performed in duplicates (unpaired t-test, ****p ≤ 0.0001). c, d FVII:Ag levels in the conditioned medium or cell lysates from cells expressing rFVII-160R (c) or rFVIIwt (d), treated with 4-PBA were examined by ELISA, and a ratio between secreted/intracellular FVII:Ag was calculated. Values are expressed relative to untreated cells and are reported as mean ± SD from at least three independent experiments performed in duplicates (one-way ANOVA, ****p ≤ 0.0001, ***p ≤ 0.001, **p ≤ 0.01). F7 mRNA levels from cells expressing rFVII-160R (e) or rFVIIwt (f), treated with 4-PBA were analysed by Taqman qRT-PCR. Quantitation was performed using the comparative CT method with 18S as the endogenous control gene and untreated cells as the calibrator. Bars represent mean fold-change in expression compared to untreated cells. Values are reported as mean ± SD from at least three individual experiments with four biological parallels (one-way ANOVA ****p ≤ 0.0001)