Fig 4.

PN-1 binding to HUVECs at 4°C. (A to C) Cells were incubated at 4°C with recombinant PN-1 variants (1 μg/ml) in the presence or not of heparin (Hep), chondroitin sulfate (CS), or dermatan sulfate (DS) (A and C) or with supernatant of resting or activated platelets (B). Whole-cell lysates were submitted to immunoblotting with a polyclonal anti-PN-1 antibody and an anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) antibody as a protein loading control. Results are shown with representative immunoblots from at least 3 independent experiments (A, B, and C). (D) Densitometric quantification, expressed as the mean intensity (PN-1/GAPDH ratio) relative to control (WT binding), determined for each immunoblot. NS, not significant; **, P < 0.01; ***, P < 0.0001 versus WT or the variants without heparin.