Fig. 4

NRG-1 reduces heme-induced endothelial cell permeability. hCMEC/D3 cells were grown on the inner surface of gelatin-coated transwell inserts, then incubated with 30 μM heme, 10 ng/ml IL-1β, 1 μg/ml lipopolysaccharides (LPS), and 100 ng/ml NRG-1 as indicated for 24 h. Permeability was measured by adding 10 μM sodium fluorescein to the upper chamber. After incubation for 0.5 h, the sample from the lower compartment was collected and 100 μl of sample was measured for fluorescence at excitation 485 nm and emission 535 nm. Incubation with heme, LPS, and IL-1β resulted in increased permeability of the endothelial monolayer by 1.93-fold, 1.92-fold, and 2.33-fold, respectively. The presence of NRG-1 during the incubation period reduced endothelial permeability down to 1.32-fold, 1.50-fold, and 1.8-fold, respectively