May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Generation and Characterization of Chimeric Complement Factor H Transgenic Mice With an "At-Risk" Mutation for Age-Related Macular Degeneration
Author Affiliations & Notes
  • R. Ufret-Vincenty
    Ophthalmology, UT Southwestern Medical Center, Dallas, Texas
  • A. McMahon
    Ophthalmology, UT Southwestern Medical Center, Dallas, Texas
  • P. Chen
    Ophthalmology, UT Southwestern Medical Center, Dallas, Texas
  • J. Y. Niederkorn
    Ophthalmology, UT Southwestern Medical Center, Dallas, Texas
  • W. Kedzierski
    Ophthalmology, UT Southwestern Medical Center, Dallas, Texas
  • Footnotes
    Commercial Relationships  R. Ufret-Vincenty, None; A. McMahon, None; P. Chen, None; J.Y. Niederkorn, None; W. Kedzierski, None.
  • Footnotes
    Support  Unrestricted Grant from Research to Prevent Blindness, Inc
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 210. doi:https://doi.org/
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      R. Ufret-Vincenty, A. McMahon, P. Chen, J. Y. Niederkorn, W. Kedzierski; Generation and Characterization of Chimeric Complement Factor H Transgenic Mice With an "At-Risk" Mutation for Age-Related Macular Degeneration. Invest. Ophthalmol. Vis. Sci. 2008;49(13):210. doi: https://doi.org/.

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

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Abstract

Purpose: : Complement factor H (Cfh) is a key regulator of the alternative complement pathway. A single nucleotide polymorphism that leads to a histidine instead of a tyrosine at amino acid position 402 of Cfh (Y402H) increases the risk for age-related macular degeneration (AMD) up to 7-fold. Our goal is to generate transgenic mice expressing either the tyrosine or the histidine variants of Cfh in order to: (a) identify the molecular basis for the susceptibility to AMD observed in human H-CFH carriers, and (b) generate a relevant model of AMD that would allow the dissection of the role of the different arms of the immune system in the disease.

Methods: : CFH consists of 20 short consensus repeats (SCR). Y402H localizes to SCR7. We have generated two transgenic constructs which consist of the human SCR6-8 sequence flanked by the mouse SCR1-5 and SCR9-20 sequences. One of the constructs codes for a tyrosine (Y-Cfh) at amino acid position 402, and the second one codes for a histidine (H-Cfh) at this position. The ApoE promoter was used to induce liver expression of the chimeric Cfh molecules and secretion into the circulation. The coding sequences were isolated and injected into C57Bl/6 fertilized eggs.

Results: : Sequencing validated the accuracy of our constructs. Genotyping allowed us to identify 7 transgenic founders for the H-Cfh transgene and 10 for the Y-Cfh transgene. A nuclease protection assay was used to determine levels of chimeric vs. native mRNA. We then used a western blot to determine the level of Cfh protein expression in the serum.

Keywords: age-related macular degeneration • inflammation • pathobiology 
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