April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Activation of Unfolded Protein Response in the Trabecular Meshwork of Transgenic Mice Expressing Tyr437His Mutant of Human Myocilin
Author Affiliations & Notes
  • G. S. Zode
    Department of Pediatrics, Howard Hughes Medical Institute,, Iowa City, Iowa
  • D. Y. Nishimura
    Department of Pediatrics, Howard Hughes Medical Institute,, Iowa City, Iowa
  • C. C. Searby
    Department of Pediatrics, Howard Hughes Medical Institute,, Iowa City, Iowa
  • E. M. Stone
    Department of Pediatrics, Howard Hughes Medical Institute,, Iowa City, Iowa
  • A. V. Dumitrescu
    Department of Ophthalmology and Visual Sciences, University of Iowa Carver College of Medicine, Iowa City, Iowa
  • V. C. Sheffiled
    Department of Pediatrics, Howard Hughes Medical Institute,, Iowa City, Iowa
  • M. H. Kuehn
    Department of Ophthalmology and Visual Sciences, University of Iowa Carver College of Medicine, Iowa City, Iowa
  • Footnotes
    Commercial Relationships  G.S. Zode, None; D.Y. Nishimura, None; C.C. Searby, None; E.M. Stone, None; A.V. Dumitrescu, None; V.C. Sheffiled, None; M.H. Kuehn, None.
  • Footnotes
    Support  HHMI
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 5835. doi:
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      G. S. Zode, D. Y. Nishimura, C. C. Searby, E. M. Stone, A. V. Dumitrescu, V. C. Sheffiled, M. H. Kuehn; Activation of Unfolded Protein Response in the Trabecular Meshwork of Transgenic Mice Expressing Tyr437His Mutant of Human Myocilin. Invest. Ophthalmol. Vis. Sci. 2010;51(13):5835.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : Myocilin mutations have been associated with congenital glaucoma and 2-4% of primary open angle glaucoma (POAG) cases, but the pathogenic mechanisms remain largely unknown. The objective of this study was to examine the glaucoma phenotype of the Tyr437His mutant of human myocilin in transgenic (Tg) mice.

Methods: : Tg mice were generated by expressing the Tyr437His mutant form of human myocilin under the control of the CMV promoter. Myocilin expression in the anterior chamber of Tg mice was examined by RT-PCR, immunohistochemistry and western blot analysis. Intraocular pressure (IOP) was measured using a rebound tonometer. Whole mount retina and PPD staining of optic nerve was used to examine retinal ganglion cells (RGC) loss and axonal damage respectively. Western blot analysis and immunostaining of the anterior chamber was performed to examine the unfolded protein response (UPR) in the trabecular meshwork (TM) of Tg mice. Myocilin and XBP-1 plasmids were co-transfected to examine ER stress in TM cells.

Results: : Tg mice expressed higher levels of myocilin in the irodocorneal angle than wild type (wt) littermates. Starting at 3-month age, Tg mice exhibited significantly higher intraocular pressure (12 mmHg in wt vs 16 mmHg in Tg, n=40). In addition, Tg mice lost ~ 30% RGCs by 6 month age and demonstrated progressive axonal damage. Interestingly, expression of mutant myocilin in the anterior chamber of Tg mice induced ER stress and activated UPR, detected by induction of GRP78, GRP94, IRE œ, and crystallins. Immunostaining of anterior chamber sections demonstrated elevated levels of GRP78 in the trabecular meshwork of Tg mice. At a later stage, the ER stress in the anterior chamber of Tg mice switched from UPR to pro-apoptotic response as demonstrated by upregulation of chop and processing of pro-caspase-12 in Tg mice. In TM-5 cells, expression Tyr437His mutant of human myocilin and not wild type human myocilin induced and associated with GRP78 and spliced XBP-1, indicating activation of UPR by expression of Tyr437His mutant of human myocilin.

Conclusions: : Our observations indicate that Tg mice exhibit pathological changes similar to those in glaucoma patients. In addition, expression of Tyr437His mutant of human myocilin in the TM leads to ER stress and activation of UPR. These events may compromise TM function leading to outflow deficiency in glaucoma patients with myocilin mutation.

Keywords: trabecular meshwork • retinal degenerations: cell biology • anterior chamber 
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