Figure 5
Effect of free and circulating microparticle-bound activated protein C on endothelial permeability. Endothelial cells seeded onto Transwell plates were incubated with 20 nmol/L free- activated protein C ( APC) or with 1.4 × 105 circulating microparticles (MP), for three hours and permeability was assessed using Evans-Blue-Bovine serum albumin. MP were obtained from Day 3 of rhAPC treatment. Specificity of APC and protease activated receptor 1 (PAR1) was investigated by inclusion of protein C (PCAb) antibody (10 μg/mL) or PAR1 specific antibody (ATAP2 at 20 μg/mL) for 30 minutes before incubation with APC or MPs. All experiments were performed as duplicates; n = 3. **P < 0.006 denote statistical significance in relation to untreated baseline; ♦♦ P < 0.006 denote significance between APC or, microparticle-associated APC (MP-APC) and blockade as assessed by a Mann-Whitney U test with Bonferroni correction.