Chronic hypoxia in vivo increases von Willebrand factor expression in lung endothelial cells

The basis of hypoxia‐induced lung inflammation is not understood. To address this understanding, we considered the role of the endothelial (EC) platelet and leukocyte receptor, von Willebrand factor (vWf). We exposed mice (Swiss Webster) to room air, or 12% O2 for 24 hours. Then, we anesthetized the mice (ketamine: 3ml/kg and xylazine: 1ml/kg, i.p.), opened the chest, injected intra‐cardiac heparin and covered the lungs with chilled buffer (4°C). Through a pulmonary artery cannula we cleared blood in lung vessels by buffer infusion. Then, we infused chilled collagenase (1000 units/ml) and trypsin (3ml), and collected the effluent through a left atrial cannula. After filtering the sample, we treated it with paraformaldehyde (4%) and Triton X‐100 (0.1%). Then, using Alexa 488‐conjugated, specific and non‐specific mAbs (1μg/100 μl) we stained the samples respectively, to detect vWf and to rule out non‐specific ligation. Specific Alexa fluorescence (normalized units) as detected by flow cytometry on 5×10 4 cells/lung, defined EC. In animals exposed to normoxia, EC fluorescence was 4.2±0.3 units/10 3 cells (mean±SD; n=3). By contrast, in hypoxia EC fluorescence increased >5 times baseline to 21.2±3.5 units/10 3 cells (n=3, p<0.05). Our findings indicate that hypoxia activated vWf expression in lung EC. In hypoxia, vWf‐induced EC signaling might underlie lung inflammation (Support: HL54157, HL36024).