Fig. 1

Neutrophil elastase (NE) impedes tubule formation and decreases angiopoietin (ANGPT) expression, whereas inflammatory factors differentially modulate ANGPT expression, in human umbilical vein endothelial cells (HUVECs). A tubule formation assay was performed as described in the Methods section. Recombinant human NE was added at concentrations of 100, 250, 500, and 1000 ng/ml (HUVECs exposed only to medium served as the control) to determine tubule formation (a), the percentage of covered area (b), total tube length (c), and total numbers of tubes (d). e and g Total RNA was prepared from HUVECs exposed to various concentrations of NE for 24 h to determine the expression of ANGPT1 and ANGPT2. f and h ANGPT-1 and ANGPT-2 immunocytochemistry was performed on fixed HUVECs as described in the Methods sections. i–l ANGPT1 and ANGPT2 mRNA expression was determined by real-time quantitative reverse transcription-polymerase chain reaction in HUVECs collected 0.5, 1, 3, 6, 9, and 12 h after treatment with lipopolysaccharide ([LPS] 2 μg/ml) and tumor necrosis factor alpha ([TNF-α] 100 ng/ml). 18S was used as the internal control. Data represent means ± SEMs (n = 2–3/group performed in triplicate). *p < 0.05, **p < 0.01, ***p < 0.001 vs. control