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N. N. Osborne, J. Dan, L. Guang-Yu, G. Lascaratos, H. J. Mortiboys, S. Jackson; Light Can Cause Apoptosis of Cells in Culture by a Mitochondrial Dependent Mechanism. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1556. doi: https://doi.org/.
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To provide support for the idea that retinal ganglion cells are susceptible to light injury under certain circumstances because of the many mitochondria in their axons within the globe.
Five day old primary rat retinal cultures were exposed to light (400-760nm, intensity 1000 lux) or dark in the same incubator for 48 hours. BJhTERT immortalised fibroblasts (Bioscience Clontech, Germany) in culture were depleted of mtDNA with ethidium bromide (5µg/ml). The depletion of mtDNA from these so called rho0 cells caused an almost complete loss of mitochondrial respiratory chain activity and rendered the cells auxotrophic for pyruvate and uridine present in the growth medium. Sub-confluent cultures of BJhTERT and rhoO cells were exposed to light for 4 days or maintained in the dark in the same incubator. Cell culture medium was sometimes modified by adding selected antioxidants or reducing the serum concentration. Oxidative status of the different cell cultures was assessed by means of a MTT assay and measurement of mitochondrial dehydrogenase activity. Cultures were stained for the presence of TUNEL-positive nuclei as an indicator of apoptosis and stained for reactive oxygen species (ROS) using 2’,7’-dihydroethidium. Primary retinal cultures were immunostained to determine numbers GABA neurones.
Oxidative status and mitochondrial dehydrogenase activity in primary retinal cultures (-40±5%) and BJhTERT cells (-13±3%) was reduced significantly by light. Primary retinal cultures exposed to light had fewer GABA-positive neurones (-42±6%) and a 3-5 fold increase in TUNEL-positive cell nuclei in light-exposed primary retinal and BJhTERT cultures than when maintained in the dark. ROS staining was also clearly increased in cultures exposed to light. The numbers of TUNEL-positive neurones caused by light rose significantly in primary retinal cultures with reduced serum. Light did not generate ROS or induce cell death of rhoO cells. The light-induced toxic effect on BJhTERT cells and GABA-positive neurones in primary retinal cultures was significantly blunted by vitamin E and α-lipoic acid.
Light stimulates retinal neurones or BJhTERT cells to die in culture by affecting their mitochondria. This influence is blunted in the presence of α-lipoic acid or vitamin E. Cells (rhoO cells) lacking normal mitochondrial function are unaffected by a light insult. Light may therefore be a risk factor to ganglion cells within the retina in certain circumstances because of their many mitochondria.
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