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Hyungsik Lim, Denis Sharoukhov; Microtubule deficit predisposes the retinal ganglion cell axons to atrophy in DBA/2J. Invest. Ophthalmol. Vis. Sci. 2018;59(9):3027.
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© ARVO (1962-2015); The Authors (2016-present)
Glaucoma is characterized by progressive loss of the retinal ganglion cells (RGCs) and the RGC axon is the primary site of degeneration. Here we test a long-standing notion in the field that axonal microtubules (MT) cytoskeleton is disrupted prior to the loss of RGC axons in the course of glaucomatous progression (‘the MT hypothesis’).
Label-free nonlinear optical microscopy was employed, namely second-harmonic generation (SHG) microscopy, whose signal arises from uniformly polarized MTs. The retinal nerve fibers in the fresh wholemounts were imaged using DBA/2J mice as an experimental model of glaucoma and DBA/2J-Gpnmb+ as a non-glaucomatous control. Animals were examined at ages from 5 to 16 months (male and female, N=52 and 17, respectively). The axonal morphology and the integrity of MT cytoskeleton were quantified by bias-free morphometry and statistical analysis.
The previously known pathologies of glaucoma, e.g., sectorial degeneration of RGC axons, could be observed by SHG imaging. In addition, we found that MTs can be lost within intact axons. The mean MT density was significantly lower in DBA/2J than in non-glaucomatous control DBA/2J-Gpnmb+. The loss of axonal MTs was DBA/2J-specific and initiates at an age that roughly coincides with the onset of IOP elevation. MT disruption occurred in a sectorial manner, which was spatially correlated with the loss of RGC axons. The loss of MT density was dependent on age and gender. The decay rate of MT density was approximately 70% higher than that of RGC axons.
Supporting evidence is presented for the MT hypothesis. Collectively, our results indicate that the breakdown of MTs is pathology of glaucoma. The spatio-temporal relationship of MT deficiency and axonal atrophy strongly suggests that cytoskeletal breakdown predisposes the RGC axons to degeneration. Based on a new finding that MT density is highly variable and spatially discrete, a new model of RGC degeneration is proposed. Our study validates SHG technique for elucidating the role and mechanism of MT deficiency in the pathogenesis of glaucoma.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
(a) Mosaics of DBA/2J and DBA/2J-Gpnmb+ at 7 to 11 months. (b) The volume of the retinal nerves. (c) and (d) The integrated SHG intensity and the thickness of the DBA/2J (9 mo), respectively, and (e) vs angle (red and black, respectively).
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