Hydrogen peroxide-induced neurotoxicity in cultured cortical cells grown in serum-free and serum-containing media.
To compare different culture conditions for neuroprotection assays in cultured cortic neurons, we evaluated cell viability after H2O2 exposure in cells cultured with standard N2 and with the enriched B-27 developed by GIBCO, both serum-free supplements. The following additives/associations were compared: N2 (+N2), B-27 (+B-27), 10% FBS (+FBS), 1% FBS in combination with N2 (FBS/N2) or N2 supplement preceded by an 1 hour precoating with 10% FBS (N2 + precoated). Our data demonstrated that B-27 is as efficient as 10% FBS to support neuronal growth for more than a week. As shown by phase-contrast optics cells grown in N2 started degenerating within 24-48 hours although measurable absorbance was seen with MTT. The pre-coating procedure failed to modify substantially cell viability as compared with N2 alone. Dose-response curves for H2O2 to induce neuronal apoptosis were almost identical for B-27 and serum supplemented samples. Catalase (100 U/ml) or vitamin E (200 microM) prevented cell death in both culture conditions. Our results indicate that DMEM/B-27 provides a serum-free cell culture environment that allows neurons to grow with optimal cell viability, comparable to that obtained with serum. We conclude that this culture condition reveals as a useful tool to test the efficacy of neuroprotectants when a serum free medium is required.
Ricart
K C
KC
Laboratorio de Neurociencias, Centro de Investigaciones Médicas Albert Einstein-Fundación CIMAE, Buenos Aires, Argentina.
Fiszman
M L
ML
D000818:Animals; D001769:Blood; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D003470:Culture Media / Q000494:pharmacology*; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D009498:Neurotoxins / Q000494:pharmacology*; D016877:Oxidants / Q000494:pharmacology*; D018384:Oxidative Stress
D003470:Culture Media; D016895:Culture Media, Serum-Free; D009498:Neurotoxins; D016877:Oxidants; D006861:Hydrogen Peroxide