Characterization of embryonic cortical neuron death in prolonged cell suspension

Cell transplantation may be an effective therapeutic strategy for many neurodegenerative diseases. However, difficulty in obtaining a sufficient amount of donor cells and low graft survival are two major limiting factors. Dissociation of cells from tissues or culture is an inevitable step for cell transplantation, and cell viability in suspension may influence the outcome of the cell therapy. To this end, we asked whether the suspension time of freshly dissociated neurons in vitro affects their viability. Following 4-24h cell suspension, primary cortical neurons underwent cell death. Interestingly, the neurons exhibited only marginal caspase-3 immunoreactivity with very few sub-G1 apoptotic cell proportions in flow cytometry. In addition, the suppression of caspase-3 or Bax action failed to prevent cell death of primary cortical neurons, indicating minimal apoptotic cell death. On the other hand, there was a marked increase in the TdT-mediated dUTP nick end labeling-positive and propidium iodide-labeled necrotic cells (∼50%) with enhanced poly [ADP-ribose] polymerase-1 activity. Therefore, prevention against necrosis rather than apoptosis may be required for optimal benefits in cell transplantation.