Recharacterization of the RGC-5 retinal ganglion cell line.
OBJECTIVE
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 tRNA(Phe) gene. Marker expression was examined in undifferentiated cells, and cells differentiated with 50 microg/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 beta-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.
Van Bergen
Nicole J
NJ
Department of Ophthalmology, Centre for Eye Research Australia, University of Melbourne, Royal Victorian Eye and Ear Hospital, East Melbourne, Victoria, Australia.
Wood
John P M
JP
Chidlow
Glyn
G
Trounce
Ian A
IA
Casson
Robert J
RJ
Ju
Won-Kyu
WK
Weinreb
Robert N
RN
Crowston
Jonathan G
JG
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002461:Cell Line, Transformed; D003208:Concanavalin A / Q000494:pharmacology; D019084:Fluorescent Antibody Technique, Indirect; D018698:Glutamic Acid / Q000494:pharmacology; D051379:Mice; D051381:Rats; D012165:Retinal Ganglion Cells / Q000166:cytology* / Q000378:metabolism
D015415:Biomarkers; D003208:Concanavalin A; D018698:Glutamic Acid; C019558:succinylconcanavalin A