A population of PC12 cells that is initiating apoptosis can be rescued by nerve growth factor

Programmed cell death, or apoptosis, occurs asynchronously in neuronal cell s. To overcome this asynchrony, rat pheochromocytoma (PC12) cells were sepa rated at different stages of apoptosis on the basis of cell density. Live c ells that exhibited no apoptotic features floated to the top of density gra dients. The most dense cells showed extensive loss of cytochrome c from mit ochondria, caspase activation, chromatin condensation, and DNA fragmentatio n. These cells were committed to apoptosis and could not be rescued by recu lturing in with nerve growth factor (NGF). Cells of intermediate density di splayed no DNA fragmentation, but had begun to show cytochrome c loss, casp ase activation, and chromatin condensation. This population displayed upreg ulation of the prodeath factor, c-Jun, and downregulation of prosurvival ki nase, Akt. Importantly, apoptosis was reversible by NGF in this population. These studies suggest that increased cell density correlates with an initi al step in the apoptosis mechanism that precedes irreversible commitment to suicide.