Figure 6.

Effects of plasma serine proteases on the dynamic mass redistribution of confluent endothelial monolayers. A 384 well biosensor microplate was precoated with 0.2 mg/ml gelatin (37 °C, 20 min, PBS), then the wells were washed with PBS, and confluent monolayers of HUVECs were grown on the surface (24 h). The microplate was placed into the Epic BT reader, and after the stabilization of the baseline, serine proteases rMASP-3 (1 μM), FXII (2 μM), rC1s (1 μM), rMASP-2 (0.2, 0.6 or 2 μM) and kallikrein (0.2, 0.6 or 2 μM), rC1r (0.1, 0.3 or 1 μM) or culture medium alone was added. Proteases rMASP-2, kallikrein and rC1r were also applied in complex with their inhibitor, C1-INH (using a three-fold molar excess of C1-INH). Cell treatments were added directly to the culture medium, and the wavelength shift was monitored for 60 min. Wavelength shift curves were normalized to the medium control. The figures show representative graphs of at least 3 independent measurements.