Fig. 2.

Effects of washed red blood cells (RBCs) on human pulmonary artery endothelial cell (HPAEC) inflammatory activation. HPAECs were challenged with vehicle, indicated dilutions of fresh (RBC-F) and old (RBC-O) RBCs, 200 ng/mL LPS or 0.2 ng/mL TNF-α, followed by incubation for 16–20 h. A and B: the conditioned medium was analyzed for soluble ICAM-1 (sICAM-1) (A) and IL-8 (B) using ELISA assay. C: Western blot analysis of ICAM-1 and VCAM-1 expression by HPAECs treated with RBC-F (Rf) and RBC-O (Ro) at indicated dilutions or LPS. Membrane reprobing for β-tubulin was used as normalization control. Bar graphs depict the quantitative densitometry analysis of Western blot data; n = 3. *P < 0.05 vs. nonstimulated control. #P < 0.005.