Light induces programmed cell death by activating multiple independent proteases in a cone photoreceptor cell line.
OBJECTIVE
Although the apoptotic death of photoreceptor cells in retinal degenerative disorders is well documented, the molecular mechanism is not understood. The objective of this study was to determine the molecular events leading to the death of photoreceptor cells.


METHODS
An assay was developed wherein 661W cells, a cone photoreceptor cell line, were stressed with light and percentage of surviving cells was determined. The degree of cell death was established using the MTT assay. Western blot analysis was used to confirm the activation of multiple proteases. Amounts of retinaldehydes were determined by extraction and HPLC.


RESULTS
661W cells were more susceptible to light stress only in the presence of the chromophore 9-cis retinal for 4 hours. On exposure to light, 9-cis retinal was converted to all-trans retinal, which was found to be toxic to cells in the presence of light. However, all-trans retinol, which is the product of action by the enzyme retinol dehydrogenase on all-trans retinal, was not toxic. The sensitivity to light increased with serum deprivation. Light stress activated caspases, calpain 2, and cathepsin D independently and led to the demise of the cell. The mitochondria-dependent apoptotic pathway was also activated after the truncation of Bid, the pre-proapoptotic protein. Truncation of Bid led to the release of cytochrome c from the mitochondria and the activation of caspase 9.


CONCLUSIONS
The activation of multiple proteases by light-induced stress is a relevant finding for studies conducted to investigate the use of pharmaceutical agents to retard or cure the loss of cone photoreceptors observed in age-related macular degeneration and other degenerative retinal diseases.
Kanan
Yogita
Y
Departments of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
Moiseyev
Gennadiy
G
Agarwal
Neeraj
N
Ma
Jian-Xing
JX
Al-Ubaidi
Muayyad R
MR
D000818:Animals; D017209:Apoptosis / Q000528:radiation effects*; D015153:Blotting, Western; D002154:Calpain / Q000378:metabolism; D020169:Caspases / Q000378:metabolism; D002402:Cathepsin D / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival; D002851:Chromatography, High Pressure Liquid; D004224:Diterpenes; D004789:Enzyme Activation; D008027:Light; D051379:Mice; D010447:Peptide Hydrolases / Q000378:metabolism*; D017949:Retinal Cone Photoreceptor Cells / Q000201:enzymology* / Q000473:pathology / Q000528:radiation effects*; D012172:Retinaldehyde / Q000378:metabolism; D014801:Vitamin A / Q000378:metabolism
D004224:Diterpenes; D014801:Vitamin A; C031390:9-cis-retinal; D010447:Peptide Hydrolases; D002154:Calpain; D020169:Caspases; D002402:Cathepsin D; D012172:Retinaldehyde