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Brian Choi, Alireza Shabanzadeh, Philippe D'Onofrio, Paulo D Koeberle; Inhibition of primary cilia resorption reduces retinal ganglion cell degeneration after axotomy. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4413. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
It is unknown whether the primary cilia of RGCs (retinal ganglion cells) regulate cell cycle re-entry that is exhibited during apoptosis. Aurora A kinase and HDAC6 (histone deacetylase 6) are involved in cilia resorption and highly expressed in rapidly dividing cells. This study investigates the hypothesis that inhibiting the pathways that control cilia resorption can prevent apoptotic cell cycle re-entry in RGCs after optic nerve transection.
Adult Sprague-Dawley rats received intraorbital optic nerve transections. At 3 and 8 days postaxotomy, animals received intraocular injections of an Aurora A kinase inhibitor (N=6) or Tubastatin A (TBA; N=4), an inhibitor of HDAC6, delivered at 10mM or 20 mM concentrations respectively. At 14 days postaxotomy RGCs were imaged by immunofluorescence directed against RBPMS (RNA Binding Protein with Multiple Splicing) and survival was quantified from fixed, flat-mounted retinas. Data was analyzed using a one-way ANOVA followed by Tukey’s post-hoc test in order to identify statistically significant differences between control and experimental groups. To examine the effects of cilia disruption in RGCs, Ift88 (intraflagellar transport) siRNAs were injected into the vitreous chamber or into the transected optic nerve stump. After 4 days of siRNA treatment, cell cycle entry in RGCs was assayed in fixed retinas using a Ki67 antibody.
At 14 days postaxotomy, retinas treated with Aurora A kinase inhibitor or TBA had a significantly higher RGC density compared to controls (p<0.001). Ift88 siRNA treated RGCs showed nuclear expression of Ki67 whereas normal (untreated) or control retinas, treated with a scrambled siRNA sequence did not.
Our findings suggest that the primary cilia may play a role in keeping RGCs in a post mitotic state. Blocking Aurora A and HDAC6 activity appears to halt the signalling cascades that lead to apoptotic cell cycle re-entry, via cilia resorption. These findings support a novel role of the primary cilium in regulating the survival of RGCs after injury.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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