The role of nerve growth factor in vitro in cell resistance to 6-hydroxydopamine toxicity

Cell resistance to the catecholaminergic neurotoxin 6-hydroxydopamine was studied in various cell lines: human neuroblastoma lines SK-N-MC. SK-N-SH, and SK-N-SH-SY5Y; and non-neuroblastoma lines CHO-K1, S-180, C-6, and L-M, the latter three of which synthesize nerve growth factor. Cells were treated one day after seeding for 1 h with 6OHDA. Cytotoxicity of the drug was quantified as the percent live cells, determined by the trypan blue exclusion test, 24 h after treatment. At 100 μg/ml, 6OHDA lethal toxicity was confined mainly to neuroblastoma cells. However, drug specificity was dependent not only on cell type, but also on cell density and presence of NGF. Thus, the non-neuroblastoma cell strain S-180-A, which produces less NGF than its parent line S-180, lost resistance to toxicity at low cell density, and even at high density was less resistant than SK-N-MC neuroblastoma cells. Moreover, when mouse β-NGF (500 BU/ml) was administered to human neuroblastoma clones SY5Y and IN one day after seeding for 24 h before drug treatment, the cell survival rate increased significantly, although only SY5Y cells were protected by a lower concentration (1 BU/ml) of exogenous NGF. Finally, cell line S-180 became susceptible to 6OHDA killing when incubated one hour with high titer anti-mouse β-NGF immediately prior to drug treatment, whereas cell line C-6 did not. NGF was therefore proposed to have an important, though not determinative, role in cell resistance to 6OHDA toxicity.