Effects of freezing on cell viability and mechanisms of cell death in a human prostate cancer cell line

The prostate cancer cell line PC-3 was used as an in vitro model to study t he effects of temperature on the molecular mechanisms underlying cell death . Confluent PC-3 cultures were exposed to temperatures ranging from 37 degr ees C to -80 degrees C and allowed to recover for 2 days. Detached (dead) a nd adherent (living) cells were then isolated and assayed for DNA fragmenta tion using agarose gel electrophoresis, Cells exposed to temperatures above -5 degrees C died through apoptosis, as evidenced by nonrandom DNA fragmen tation, whereas cells exposed to temperatures below -15 degrees C died thro ugh necrosis, as revealed by random DNA fragmentation. An apoptotic inhibit or, IDN-1529, completely blocked DNA fragmentation in cells exposed to temp eratures above -5 degrees C but below 37 degrees C, Cell viability was then assessed with the noninvasive metabolic indicator dye Alamar Blue. Remarka bly, IDN-1529 inhibited cell death in PC-3 cells exposed to temperatures ra nging from -10 degrees to -75 degrees C, even at temperatures at which necr osis appeared to be the primary cause of death. It is concluded that: (1) P C-3 cells can die by either apoptosis or necrosis, and the mode of death is determined by the treatment temperature; and (2) apoptotic inhibitors prot ect against the death of PC-3 cells even even with regimens that appear to induce necrosis exclusively. These observations have direct implications fo r the cryogenic treatment of cancer, especially that of the prostate.