Extracellular calcium is necessary for the soluble HLA class I–mediated apoptosis
| . | Nil . | sHLA-I . | SHLA-I + EGTA . | sHLA-I + DMSO . | sHLA-I + verapamil . |
|---|---|---|---|---|---|
| Cl.1.25 CD8bright | 1.0 | 38.0 | 15.0 | 37.0 | 12.0 |
| Cl.10.50 CD8dull | 3.0 | 5.0 | 3.0 | 3.0 | 2.1 |
| NK-2 | 2.0 | 45.3 | 17.4 | 44.2 | 10.5 |
| NK-3 | 5.6 | 35.5 | 15.3 | 36.0 | 12.6 |
| . | Nil . | sHLA-I . | SHLA-I + EGTA . | sHLA-I + DMSO . | sHLA-I + verapamil . |
|---|---|---|---|---|---|
| Cl.1.25 CD8bright | 1.0 | 38.0 | 15.0 | 37.0 | 12.0 |
| Cl.10.50 CD8dull | 3.0 | 5.0 | 3.0 | 3.0 | 2.1 |
| NK-2 | 2.0 | 45.3 | 17.4 | 44.2 | 10.5 |
| NK-3 | 5.6 | 35.5 | 15.3 | 36.0 | 12.6 |
A total of 5 × 104 cells (from natural killer [NK] cell clones Cl.1.25, Cl.10.50, NK-2 bulk population, 90% CD8+; NK-3 bulk population, 80% CD8+) were cultured for 48 hours in medium alone (nil) or with 4 μg/mL soluble HLA class-I (sHLA-I), either in the absence or in the presence of the extracellular calcium chelator EGTA (2 mM) or of the L-type calcium channel blocker verapamil (10 μM) diluted in dimethyl sulfoxide (DMSO) or with the solvent DMSO alone. Apoptosis was evaluated by staining with fluorescein isothiocyanate–annexin V. Samples were run on a FACSort (Becton Dickinson), and results are expressed as the percentage of annexin V+ PI− cells.