FIGURE 7.

Endothelial permeability to lipoproteins in vitro and aortic lesion VE-Cadherin expression in vivo. HUVECs were seeded onto gelatin-coated Transwell devices (upper chamber) before exposure to vehicle, α-thrombin (10 nM), FXIa (30 nM), FXI (30 nM), or FXIa or thrombin in the presence of PPACK for 3 h. (A) Absorbance of labeled BSA measured at 650 nm every 10 min from the lower chamber media. (B) Alexa Fluor 488-conjugated acLDL was added in 0.3% BSA to the upper chamber and media from the lower chamber was collected at 24 h after incubation. Fluorescence was measured at 495/519 Ex/Em and acLDL concentration (ng/ml) was calculated based on standard curve. (C) Cross-sections of aortic sinus were obtained and lesion endothelial barrier integrity was determined by VE-Cadherin (red) and CD31 (green) staining together with nuclei (blue). Scale bar = 200 μm. White arrows indicate regions with disrupted VE-Cadherin staining pattern, White boxes highlight regions chosen for zoomed images. Data were analyzed using one-way anova with Tukey’s post hoc tests to compare treatment groups. *p < .05 thrombin vs. vehicle. ^#p < .05 FXIa vs. vehicle. **p < .005 FXIa vs. vehicle