Comparison of the amyloid pore forming properties of rat and human Alzheimer’s beta-amyloid peptide 1-42: Calcium imaging data

The data here consists of calcium imaging of human neuroblastoma SH-SY5Y cells treated with the calcium-sensitive dye Fluo-4AM and then incubated with nanomolar concentrations of either human or rat Alzheimer’s β-amyloid peptide Aβ1-42. These data are both of a qualitative (fluorescence micrographs) and semi-quantitative nature (estimation of intracellular calcium concentrations of cells probed by Aβ1-42 peptides vs. control untreated cells). Since rat Aβ1-42 differs from its human counterpart at only three amino acid positions, this comparative study is a good assessment of the specificity of the amyloid pore forming assay. The interpretation of this dataset is presented in the accompanying study “Broad neutralization of calcium-permeable amyloid pore channels with a chimeric Alzheimer/Parkinson peptide targeting brain gangliosides” [1].


a b s t r a c t
The data here consists of calcium imaging of human neuroblastoma SH-SY5Y cells treated with the calcium-sensitive dye Fluo-4AM and then incubated with nanomolar concentrations of either human or rat Alzheimer's β-amyloid peptide Aβ1-42. These data are both of a qualitative (fluorescence micrographs) and semiquantitative nature (estimation of intracellular calcium concentrations of cells probed by Aβ1-42 peptides vs. control untreated cells). Since rat Aβ1-42 differs from its human counterpart at only three amino acid positions, this comparative study is a good assessment of the specificity of the amyloid pore forming assay. The interpretation of this dataset is presented in the accompanying study "Broad neutralization of calcium-permeable amyloid pore channels with a chimeric Alzheimer/Parkinson peptide targeting brain gangliosides" [1]. &

Value of the data
The data provides a comparative study of Ca 2 þ fluxes induced by human and rat forms of Aβ1-42 peptide.
The comparison of the Ca 2 þ fluxes induced by human and rat Aβ1-42 peptides may be used as an internal reference to validate this assay for studying amyloid pore formation induced by any amyloid protein in living neural cells.
Researchers interested in testing amyloid pore formation and inhibitors of amyloid pores might carefully choose positive and negative controls with calibrated compounds.

Materials
SH-SY5Y cells were obtained from ATCC. Dulbecco's Modified Eagle Medium: Nutrient Mixture F12 (DMEM/F12), HBSS, glutamine and penicillin/streptomycin were furnished by Gibco. Fluo-4AM was from Invitrogen. Human and rat Aβ1-42 peptides were purchased from rPeptide. The purity of these peptides was 495% as assessed by HPLC.

Calcium measurements
SH-SY5Y cells were plated (45.000 cells/dish) in 35 mm culture dishes and grown during 72 h at 37°C. They were loaded with 5 mM Fluo-4AM for 30 min in the dark as previously described [1,5]. The calcium fluxes were estimated by measuring the variation of cell fluorescence intensity after the injection of either rat or human Aβ1-42 (220 nM) into the recording chamber directly above an upright microscope objective (BX51W Olympus) equipped with an illuminator system MT20 module. Fluorescence emission at 525 nm was imaged by a digital camera CDD (Hamanatsu ORCA-ER) after fluorescence excitation at 490 nm. Time-lapse images (1 frame/10 s) were collected using the CellR Software (Olympus). Fluorescence intensity were measured from region of interest (ROI) centered on individual cells. Signals were expressed as fluorescence after treatment (F t60 ) divided by the fluorescence before treatment (F 0 ) and multiplied by 100. The results were averaged and the fluorescence of control is subtracted of each value. The experiments were performed at 30°C during 60 min. In the   [1,12]. SH-SY5Y cells treated with the calcium dye Fluo-4AM were incubated with human Aβ1-42 (blue histogram, n¼ 87) or rat Aβ1-42 (red histogram, n¼120) and calcium-dependent fluorescence was recorded after 60 min of incubation with the indicated peptides (lower panel). Results are expressed as mean 7 SEM. Statistical significance between human and rat Aβ1-42 (p ¼2.75 Â 10 À 11 ) was evaluated by using Student's t-test.
pseudocolor representations of cells, warmer colors correspond to higher fluorescence and thus to higher Ca 2 þ levels.

Statistical analysis
Quantitative data are expressed as mean 7S.E.M. and the statistical significance was assessed with the Student's t-test.