Effect of Various Inhibitors on PS Externalization and Caspase-3–Like In Vitro Activity in Constitutive and Fas/APO-1–Triggered Apoptosis
| Treatment . | PS Exposure (% annexin V-FITC fluorescence) . | DEVD-AMC Cleavage (pmol/min/106 cells) . |
|---|---|---|
| Medium alone | 22.5 ± 5.4 | 20.2 ± 4.8 |
| + zVAD-cmk (10 μmol/L) | 5.0 ± 3.2 | 10.7 ± 2.2 |
| + DPI (10 μmol/L) | 35.2 ± 7.4 | 36.2 ± 12.5 |
| + ZB4MoAb (1 μg/mL) | 20.4 ± 4.1 | 22.5 ± 4.8 |
| Anti-Fas MoAb (250 ng/mL) | 25.4 ± 6.2 | 35.8 ± 6.1 |
| + zVAD-cmk (10 μmol/L) | 5.5 ± 2.5 | 5.1 ± 2.8 |
| + DPI (10 μmol/L) | 55.2 ± 10.5 | 51.8 ± 11.2 |
| + ZB4MoAb (1 μg/mL) | 13.6 ± 5.3 | 12.4 ± 4.9 |
| Treatment . | PS Exposure (% annexin V-FITC fluorescence) . | DEVD-AMC Cleavage (pmol/min/106 cells) . |
|---|---|---|
| Medium alone | 22.5 ± 5.4 | 20.2 ± 4.8 |
| + zVAD-cmk (10 μmol/L) | 5.0 ± 3.2 | 10.7 ± 2.2 |
| + DPI (10 μmol/L) | 35.2 ± 7.4 | 36.2 ± 12.5 |
| + ZB4MoAb (1 μg/mL) | 20.4 ± 4.1 | 22.5 ± 4.8 |
| Anti-Fas MoAb (250 ng/mL) | 25.4 ± 6.2 | 35.8 ± 6.1 |
| + zVAD-cmk (10 μmol/L) | 5.5 ± 2.5 | 5.1 ± 2.8 |
| + DPI (10 μmol/L) | 55.2 ± 10.5 | 51.8 ± 11.2 |
| + ZB4MoAb (1 μg/mL) | 13.6 ± 5.3 | 12.4 ± 4.9 |
Freshly isolated neutrophils were maintained in culture with anti-Fas MoAb (clone CH-11) or medium alone in the presence or absence of the inhibitors DPI and zVAD-cmk and harvested after 6 hours (anti-Fas–treated) and 12 hours (medium alone), respectively. The antagonistic anti-Fas MoAb ZB4 was added 1 hour before the addition of the CH-11 MoAb. PS exposure and DEVD-AMC cleavage was determined as described in Materials and Methods. Data are presented as mean ± SD (n = 3).