April 2010
Volume 51, Issue 13
ARVO Annual Meeting Abstract  |   April 2010
Cellular and Biochemical Analyses of Alpha 2 Collagen VIII Mutations Causing Fuchs Endothelial Dystrophy
Author Affiliations & Notes
  • C. Kelliher
    Ophthalmology, Wilmer Eye Institute, Baltimore, Maryland
  • C. Engler
    Ophthalmology, Wilmer Eye Institute, Baltimore, Maryland
  • N. Vij
    Johns Hopkins, Baltimore, Maryland
  • C. Speck
    Ophthalmology, Wilmer Eye Institute, Baltimore, Maryland
  • S. Mazur
    Johns Hopkins, Baltimore, Maryland
  • A. S. Jun
    Ophthalmology, Wilmer Eye Institute, Baltimore, Maryland
  • Footnotes
    Commercial Relationships  C. Kelliher, None; C. Engler, None; N. Vij, None; C. Speck, None; S. Mazur, None; A.S. Jun, None.
  • Footnotes
    Support  Eye Bank Association of America Richard Lindstrom Research Grant
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 2845. doi:https://doi.org/
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      C. Kelliher, C. Engler, N. Vij, C. Speck, S. Mazur, A. S. Jun; Cellular and Biochemical Analyses of Alpha 2 Collagen VIII Mutations Causing Fuchs Endothelial Dystrophy. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2845. doi: https://doi.org/.

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

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To create a novel model of Fuchs' Endothelial Dystrophy (FED) using both collagen mimetic peptides & a cellular model of the disease.


Collagen VIII, specifically the α2 chain of collagen VIII, has been implicated in the pathogenesis of FED.A model of FED was produced by transiently transfecting CHO-K1 cells with wildtype and mutant collagen VIII constructs. The production of collagen VIII by these cells was then assessed using Western blotting. Transfection efficiency was measured by measuring neomycin phosphotransferase II (NPTII) levels; NPTII was present on the same plasmid as the collagen VIII constructs and mediated antibiotic resistance. The mortality rates of the transfected cells was assesed using flow cytometry.Folding studies on collagens are complicated by the length of collagen proteins. Recently shortened peptides [termed collagen mimetic peptides (CMP)] have been used instead of full length proteins to model collagen diseases. Collagen VIII CMPs were created and circular dichroism (CD) spectroscopy was used to assess the folding of the wildtype and mutant CMPs.


Production of collagen VIII by the transfected cells was confirmed by Western blotting. However, it was noted that significantly more trimeric collagen VIII was produced by the cells transfected with the mutant genotype constructs [see attached Figure]. There was no difference in the transfection efficiency of the wildtype and mutant constructs as measured by the cellular NPTII production. There was no observable difference in cell mortality between the wildtype and mutant-transfected cells, which is consistent with the long clinical course of the disease.Results of CD comparing folding of wild-type and mutant alleles will be presented.


This study provides a new insight into the pathogenic role of aberrant collagen VIII in Fuchs' Endothelial Dystrophy.  

Keywords: cornea: endothelium • cornea: basic science • proteins encoded by disease genes 

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