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J. M. Harder, Q. Ding, L. Gan, R. T. Libby; BH3-Only Proteins in RGC Death. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2766.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal ganglion cells (RGCs) die by a BAX-dependent mechanism in DBA/2J glaucoma and after axonal injury in mice, injuries thought to be associated with neurotrophic deprivation (NTD). To begin to determine the cell death signaling pathway(s) active in glaucomatous RGCs, we investigated which of the molecules capable of activating BAX-the BH3-only proteins-are required for RGC death.
We examined the expression of the BH3-only proteins most likely involved in NTD-induced death, BIM, BAD, and HRK, by immunohistochemistry in retinas from glaucomatous DBA/2J mice or after controlled optic nerve crush (CONC). To test if RGC death after axonal injury requires these molecules, CONC was performed on wildtype, Bim-/-, Bad-/-, and Hrk-/- mice, as well as, double and triple mutant mice. We quantified RGC loss by counting total RGC layer neurons and activated-CASPASE 3 positive cells at multiple time points after CONC. RGC layer neuron counts showed that deficiency in these genes did not prevent the naturally occurring developmental death of RGCs.
After CONC and in glaucomatous DBA/2J mice, RGCs expressed BIM and HRK. Müller cells constitutively express BAD and RGCs may express BAD as well. BIM deficiency delayed loss of RGC layer neurons after CONC at 7 days (genotype, percentage of cells remaining compared to unmanipulated control ± SEM, number of retinas: +/+, 72 ± 1.9, n=4, P<0.001; -/-, 94 ± 2.8, n=4, P=0.17) and 10 days (+/+, 64 ± 2.2, n=4, P<0.001; -/-, 91 ± 2.4, n=4, P=0.03). However it is important to note, in Bim-/- mice CONC-induced cell death continued at least through 21 days (-/-, 72 ± 2.1, n=4, P<0.001). Deficiency in HRK or BAD did not protect RGCs after CONC. Triple mutant (Bim-/-Hrk-/-Bad-/-) mice exhibited similar cell death after CONC as Bim-/- mice.
Several of the BH3-only proteins that activate BAX are present in RGCs in DBA/2J glaucoma and after CONC. However, deficiency in three BH3-only proteins often associated with NTD did not protect RGCs after axonal injury. These data suggest a role for other BH3-only proteins in RGC death and/or that cell death pathways may be triggered in injured RGCs that are not associated with NTD.
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