Retinoic acid evoked-differentiation of neuroblastoma cells predominates over growth factor stimulation: an automated image capture and quantitation approach to neuritogenesis.
To facilitate the characterization of compounds that have positive growth factor mimetic effects on neuritogenesis, we have implemented a high-throughput functional assay which measures, in a multiparametric manner, the proliferation and differentiation characteristics of cells in a microtiter plate. Conditions were established using chronic incubation of SH-SY5Y human neuroblastoma cells with retinoic acid (RA) and/or nerve growth factor (NGF) in which discernible alterations in proliferation, growth, and differentiation of cells were induced. SH-SY5Y cells were fixed and labeled by immunocytochemistry, and an automated image acquisition and analysis package on Cellomics ArrayScanII was utilized to quantify the effects of these treatments on cell characteristics. NGF and retinoic acid were found to increase multiple parameters of SH-SY5Y differentiation, including an increased proportion of cells having neurites and increased extent of branching. However, marked differences in the effects of these compounds on SH-SY5Y growth and differentiation were also detected: whereas NGF increased cell number, RA treatment decreased cell number, and RA but not NGF caused significant elongation of neurites. This study quantifies and characterizes the effects of differentiating and proliferating agents on a human-derived neuroblastoma cell line. The high-content, rapid-throughput nature of this assay makes it ideal for functional identification and characterization of compounds regulating cell behavior.
Simpson
P B
PB
Department of Biochemistry and Molecular Biology, Neuroscience Research Centre, Merck Sharp & Dohme Research Laboratories, Harlow, Essex, United Kingdom. Peter_Simpsom@merck.com
Bacha
J I
JI
Palfreyman
E L
EL
Woollacott
A J
AJ
McKernan
R M
RM
Kerby
J
J
D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D005453:Fluorescence; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D007124:Immunoenzyme Techniques; D009414:Nerve Growth Factors / Q000494:pharmacology*; D016501:Neurites / Q000473:pathology; D009447:Neuroblastoma / Q000378:metabolism / Q000473:pathology*; D014212:Tretinoin / Q000494:pharmacology*
D009414:Nerve Growth Factors; D014212:Tretinoin