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JANET Ya-An CHANG, Laurie Sheldon, Michael L. Ko, Gladys Y P Ko; Circadian regulation of mitochondrial dynamics in retinal photoreceptors. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3011.
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© ARVO (1962-2015); The Authors (2016-present)
We previously demonstrated that the circadian oscillators in avian photoreceptors regulate photoreceptor physiology and metabolism. In this study, we further determined the circadian oscillations of mitochondrial dynamics in cultured 661W cells.
The 661W cells were cultured in Dulbecco’s Modified Eagle’s Medium supplemented with 10% fetal Bovine serum, 1% glutamax and 1% penicillin-streptomycin antibiotics at 37°C in 5% CO2. Incubators were equipped with lights and timers for the circadian entrainment to 12:12 h light-dark (LD) cycles in vitro. After 5 days in cultures under LD entrainment, 661W cells were collected at various Zeitgeber time points for further analyses. After cultured 4 days in LD, some cultured 661W cells were kept in complete darkness (DD) for 2 days, and on the second day of DD, these cells were collected at various circadian time points for further analyses. Chicken embryos from embryonic day 11 were entrained in LD cycles for 7 days in ovo, and retina tissues were collected at various time points for Western blots. Markers for mitochondrial dynamics, mitophagy, and oxidative stress were examined. One-way ANOVA were used for statistical analyses across the time points
: In 661w cells, the protein level of mitofusin 2 (MFN2) was higher during the day and lower at night when cells were under LD, but MFN2 became non-rhythmic in DD. The expression of dynamin related protein 1 (DRP1) was higher at night. The PTEN-induced putative kinase 1 (PINK1), a mitophagy marker, was rhythmic under both LD and DD. Both MFN2 and DRP1 displayed diurnal rhythms in the whole chicken retina, but their rhythmic expressions were antiphase from each other.
Even though 661W cells are a mammalian immortal cone-derived cell-line, our results showed that 661W cells can be entrained in cyclic lights. While mitochondrial fission and fusion dynamics is apparent to be light-driven, the mitochondrial response to stress might be under the circadian control in 661W cells.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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