Estradiol attenuation of beta-amyloid-induced toxicity: a comparison o.
17Beta-estradiol (betaE2) has been shown to attenuate the toxicity of beta-amyloid peptides (A beta) in neuronal cultures with the effective concentration of betaE2 ranging from low nM to high microM. This study compares the effective neuroprotective concentration of betaE2 against both A beta-mediated toxicity in a human neuroblastoma cell line, SK-N-SH using cellular reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) as an endpoint to the effective betaE2 concentration obtained using a calcein acetoxymethyl ester (calcein AM) viability assay. The minimum betaE2 concentration required for protection varied 1000-fold between the two viability assays with 1 nM betaE2 conferring significant protection in the calcein AM assay but 1 microM betaE2 required for significant protection in the MTT assay Interestingly, the maximal inhibition of MTT reduction occured at sub-toxic A beta concentrations and did not correlate with other markers of cellular viability including calcein fluorescence, dye exclusion (propidium iodide or trypan blue), cellular ATP levels, or reduction of another tetrazolium dye, 5-(3-carboxymethoxyphenyl)-2-(4,5-dimethylthiazolyl)-3-(4-sulfophenyl) tetrazolium (MTS). By contrast, there was no difference between the MTT and calcein AM assays with respect to H2O2 toxicity or the neuroprotective effectiveness of 10 nM betaE2 against H2O2 toxicity. These results indicate that low concentrations of betaE2 can attenuate A beta and H2O2 toxicity in a human neuroblastoma cell line. Further, these results suggest that the MTT assay is not an appropriate assay for the determination of betaE2-mediated attenuation of A beta toxicity.
Green
P S
PS
Center for the Neurobiology of Aging, University of Florida, Gainesville, USA.
Perez
E J
EJ
Calloway
T
T
Simpkins
J W
JW
D016229:Amyloid beta-Peptides / Q000633:toxicity*; D002470:Cell Survival / Q000187:drug effects; D004396:Coloring Agents; D004958:Estradiol / Q000494:pharmacology*; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D009447:Neuroblastoma; D009474:Neurons / Q000166:cytology / Q000187:drug effects; D018696:Neuroprotective Agents / Q000494:pharmacology; D016877:Oxidants / Q000633:toxicity; D010446:Peptide Fragments / Q000633:toxicity*; D013778:Tetrazolium Salts; D013844:Thiazoles; D014407:Tumor Cells, Cultured
D016229:Amyloid beta-Peptides; D004396:Coloring Agents; D005452:Fluoresceins; D005456:Fluorescent Dyes; D018696:Neuroprotective Agents; D016877:Oxidants; D010446:Peptide Fragments; D013778:Tetrazolium Salts; D013844:Thiazoles; C074636:amyloid beta-protein (25-35); C085925:calcein AM; D004958:Estradiol; D006861:Hydrogen Peroxide; C022616:thiazolyl blue