Apoptosis and caspase activity in control and BTHS neutrophils
. | Control, n = 5 . | . | . | BTHS, n = 5 . | . | . | ||||
|---|---|---|---|---|---|---|---|---|---|---|
| Neutrophil preparation . | Annexin-V+, %* . | Apoptotic morphology, %* . | Caspase activity, RFU† . | Annexin-V+, %* . | Apoptotic morphology, %* . | Caspase activity, RFU† . | ||||
| Fresh | 3 ± 2 | < 2 | 0 ± 13 | 56 ± 3 | < 2 | 0 ± 15‡ | ||||
| Overnight cultured | 75 ± 8 | 86 ± 12 | 527 ± 70 | 98 ± 2 | 92 ± 6 | NT | ||||
| Overnight + G-CSF | 52 ± 12 | 46 ± 17 | NT | 78§ | 50§ | NT | ||||
. | Control, n = 5 . | . | . | BTHS, n = 5 . | . | . | ||||
|---|---|---|---|---|---|---|---|---|---|---|
| Neutrophil preparation . | Annexin-V+, %* . | Apoptotic morphology, %* . | Caspase activity, RFU† . | Annexin-V+, %* . | Apoptotic morphology, %* . | Caspase activity, RFU† . | ||||
| Fresh | 3 ± 2 | < 2 | 0 ± 13 | 56 ± 3 | < 2 | 0 ± 15‡ | ||||
| Overnight cultured | 75 ± 8 | 86 ± 12 | 527 ± 70 | 98 ± 2 | 92 ± 6 | NT | ||||
| Overnight + G-CSF | 52 ± 12 | 46 ± 17 | NT | 78§ | 50§ | NT | ||||
RFU indicates relative fluorescence unit (mean ± SD); and NT, not tested.
Fresh neutrophils or neutrophils cultured overnight with or without G-CSF (100 ng/mL) were labeled with annexin-V-FITC or stained with Giemsa solution, and apoptosis was assessed. Values represent mean ± SD of annexin-V+ cells or cells with apoptotic morphology in percentage of total number of neutrophils.
Overall caspase proteolytic activity in cell lysates prepared from freshly isolated or overnight cultured control neutrophils and fresh neutrophils of 3 BTHS patients, measured by the cleavage of the fluorogenic caspase substrate DEVD-AMC. The amount of released AMC was used as a measure of caspase activity.
n = 3.
n = 2.