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Nicole J. Van Bergen, John P. M. Wood, Glyn Chidlow, Ian A. Trounce, Robert J. Casson, Won-Kyu Ju, Robert N. Weinreb, Jonathan G. Crowston; Recharacterization of the RGC-5 Retinal Ganglion Cell Line. Invest. Ophthalmol. Vis. Sci. 2009;50(9):4267-4272. doi: 10.1167/iovs.09-3484.
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purpose. The transformed RGC-5 retinal ganglion cell line is used widely in glaucoma research. Increased resistance to glutamate was noted in published literature and led to the recharacterization of the RGC-5 cell line.
methods. Characterization of the RGC-5 cell line was performed by sequencing of a region of the nuclear Thy1 gene and mitochondrial DNA sequencing of a region of the d-loop and tRNAPhe gene. Marker expression was examined in undifferentiated cells, and cells differentiated with 50 μg/mL succinyl concanavalin A (S Con A) for 3 days. Glutamate sensitivity was examined in undifferentiated and S Con A differentiated cells by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay after 24-hours of glutamate treatment.
results. RGC-5 cells were found to be of mouse (Mus musculus), not rat (Rattus norvegicus), origin by mitochondrial and nuclear DNA analyses. RGC-5 DNA sequenced in a second laboratory was subsequently found to be of M. musculus origin. Cells stained positively for the neuronal markers β-tubulin and PGP9.5 and for the microtubule-associated protein tau, but not for known markers of ganglion cells such as neurofilaments or Thy1.2, suggesting that they likely represented a lineage of mouse neuronal precursor cells. Differentiation with S Con A did not increase RGC-5 sensitivity to glutamate excitotoxicity or increase the expression of retinal or ganglion cell marker proteins.
conclusions. Investigators using cells designated as RGC-5 should confirm the species to be of rat origin and retinal-specific marker expression before considering their use as retinal ganglion-like cells.
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