American Society of Hematology

Fig. 8.

$Fig. 8. The tie-1 protease is membrane-associated, activated specifically by TNF, and inhibited by BB-24. HUVECs incubated in the presence or absence of TNF were harvested, and cell fractionation was performed as described in Materials and Methods. (A) Cytosol (a) or membrane (b) fractions prepared from TNF-treated HUVECs were incubated with 500 μg/mL tie-1a peptide for 1 hour at 37°C. (B) HUVEC membrane fractions from untreated (a) or TNF-treated (b and c) cells were incubated with 500 μg/mL tie-1a peptide in the presence (c) or absence (b) of BB-24 for 1 hour at 37°C. Samples quenched with EDTA/EGTA were separated by HPLC and peptide elution followed by absorbance at 350 nm. The control full-length tie-1a and truncated tie-1b peptides are shown in the control chromatogram (d). The arrows indicate the peaks that correspond to peptide fragments with the same size and mobility as the truncated control peptide tie-1b.$

The tie-1 protease is membrane-associated, activated specifically by TNF, and inhibited by BB-24. HUVECs incubated in the presence or absence of TNF were harvested, and cell fractionation was performed as described in Materials and Methods. (A) Cytosol (a) or membrane (b) fractions prepared from TNF-treated HUVECs were incubated with 500 μg/mL tie-1a peptide for 1 hour at 37°C. (B) HUVEC membrane fractions from untreated (a) or TNF-treated (b and c) cells were incubated with 500 μg/mL tie-1a peptide in the presence (c) or absence (b) of BB-24 for 1 hour at 37°C. Samples quenched with EDTA/EGTA were separated by HPLC and peptide elution followed by absorbance at 350 nm. The control full-length tie-1a and truncated tie-1b peptides are shown in the control chromatogram (d). The arrows indicate the peaks that correspond to peptide fragments with the same size and mobility as the truncated control peptide tie-1b.

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