Effects of tet removal and PKC activators on DNA fragmentation
| Cells | Tet (1 μg/ml) | Drug (10 nM) | Cell numbers | DNA Fragmentation | ||
|---|---|---|---|---|---|---|
| Viable | Dead | % Dead | ||||
| ×10−4 | % cells | |||||
| LNα 17 adh | + | Vhcl. | 379 ± 31 | 4.08 ± 0.51 | 1.07 ± 0.08 | 0.00 |
| LNα 17 adh | −3 days | Vhcl. | 347 ± 29 | 3.48 ± 0.28 | 0.99 ± 0.06 | 0.13 ± 0.07 |
| LNα 17 adh | −4 days | Vhcl. | 353 ± 33 | 3.93 ± 0.20 | 1.10 ± 0.14 | 0.00 |
| LNα 17 adh | −5 days | Vhcl. | 338 ± 38 | 3.66 ± 0.29 | 1.07 ± 0.18 | 0.00 |
| LNα 17 adh | −7 days | Vhcl. | 313 ± 27 | 3.68 ± 0.38 | 1.16 ± 0.09 | 0.13 ± 0.13 |
| LNCaP adh | Vhcl. | 267 ± 22 | 3.08 ± 0.31 | 1.14 ± 0.09 | 0.00 | |
| LNCaP adh | TPA, 2 days | 45.8 ± 5.0 | 4.63 ± 0.42 | 9.18 ± 1.0 | 16.6 ± 1.3 | |
| LNCaP det | TPA, 2 days | 11.2 ± 1.8 | 45.2 ± 3.2 | 80.1 ± 7.4 | 88.4 ± 5.8 | |
| LNCaP adh | TPA, 3 days | 16.3 ± 2.3 | 3.15 ± 0.45 | 16.2 ± 1.1 | 15.9 ± 1.4 | |
| LNCaP det | TPA, 3 days | 2.74 ± 0.49 | 40.9 ± 5.4 | 93.7 ± 8.8 | 81.0 ± 5.6 | |
| LNα 17 adh | + | Bryo, 2 days | 366 ± 30 | 4.45 ± 0.48 | 1.20 ± 0.11 | 0.00 |
| LNα 17 adh | −11 days | Bryo, 2 days | 42.4 ± 3.7 | 5.95 ± 0.43 | 12.3 ± 0.93 | 17.3 ± 1.8 |
| LNα 17 det | −11 days | Bryo, 2 days | 10.7 ± 1.3 | 49.1 ± 4.7 | 82.1 ± 6.6 | 91.5 ± 5.8 |
| LNα 17 adh | −12 days | Bryo, 3 days | 11.5 ± 2.1 | 2.31 ± 0.34 | 16.7 ± 1.7 | 16.3 ± 1.1 |
| LNα 17 det | −12 days | Bryo, 3 days | 2.95 ± 0.36 | 42.6 ± 4.9 | 93.5 ± 9.0 | 82.7 ± 5.2 |
LNα 17 cells were isolated and maintained in medium containing 1 μg/ml tet, and tet either remained in the medium until harvest (+) or was removed from the medium for the indicated number of days before harvest (− days). Untransfected LNCaP cells were maintained in the absence of tet. Cells (750,000) were plated in 25-cm2 flasks, with three flasks per condition. TPA (final, 10 nM), bryostatin 1 (Bryo; final, 10 nM), or ethanol vehicle (Vhcl.; final, 0.006%) was added to the medium 3 days after plating, and cells were harvested 2 or 3 days after drug addition or 2 days after vehicle addition. Cells that excluded (Viable) or were permeable to trypan blue (Dead) were counted, and the percentage of viable plus dead cells that were dead was calculated (% Dead). The percentage of cells with fragmented DNA (DNA frag) was determined as described in the text. Detached cells (det) were included in the table only when the fraction of detached cells was greater than 0.1% of the total number of cells in adherent (adh) plus detached fractions.