June 2015
Volume 56, Issue 7
Free
ARVO Annual Meeting Abstract  |   June 2015
Senescence-associated choroidal endothelial cell stiffening contributes to choriocapillaris dysfunction seen in AMD
Author Affiliations & Notes
  • Andrea Cabrera
    Bioengineering, Univerity of California, Riverside, Riverside, CA
  • Arun Bhaskaran
    Bioengineering, Univerity of California, Riverside, Riverside, CA
  • Jun Xu
    Physics and Astronomy, University of California, Riverside, Riverside, CA
  • Xiao Yang
    Bioengineering, Univerity of California, Riverside, Riverside, CA
  • Harry Scott
    Bioengineering, Univerity of California, Riverside, Riverside, CA
  • Umar Mohideen
    Physics and Astronomy, University of California, Riverside, Riverside, CA
  • Kaustabh Ghosh
    Bioengineering, Univerity of California, Riverside, Riverside, CA
  • Footnotes
    Commercial Relationships Andrea Cabrera, None; Arun Bhaskaran, None; Jun Xu, None; Xiao Yang, None; Harry Scott, None; Umar Mohideen, None; Kaustabh Ghosh, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2015, Vol.56, 4215. doi:
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      Andrea Cabrera, Arun Bhaskaran, Jun Xu, Xiao Yang, Harry Scott, Umar Mohideen, Kaustabh Ghosh; Senescence-associated choroidal endothelial cell stiffening contributes to choriocapillaris dysfunction seen in AMD. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4215.

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

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Abstract

Purpose: Despite the common prevalence of age-related macular degeneration (AMD), only 10-15% of individuals with advanced ‘wet’ AMD benefit from current therapies. Thus, there is an unmet need to prevent AMD progression at the more prevalent early ‘dry’ stage. Success in this pursuit will, however, depend on our ability to fully uncover the mechanisms that mediate AMD progression. Sub-retinal inflammation, characterized by leukocyte accumulation and complement activation, is a key hallmark of early AMD. Since aging has been shown to correlate with stiffening and inflammation of non-retinal vessels, here we explored the hypothesis that aging/senescence also leads to choriocapillaris stiffening and stiffness-dependent choroidal endothelial inflammation.

Methods: Stiffness of normal and senescent chorioretinal ECs (RF/6A) and EC-derived matrix were measured by atomic force microscopy (AFM). To examine leukocyte-EC adhesion, fluorescently-labeled U937 monocytes were added to ECs for 30 min, followed by rinsing, fixation, and counting of adherent monocytes. Complement-induced choroidal EC dysfunction was assessed by treating cells with 10% complement-competent normal human serum (NHS) for 2 hours and measuring C5b-9 (membrane attack complex; MAC) deposition by flow cytometry, cell lysis by trypan blue exclusion assay, and monocyte-EC adhesion. To inhibit cell stiffness, ECs were treated with Y27632, a pharmacological inhibitor of Rho/ROCK-dependent cell stiffness (tension).

Results: Our AFM force indentation measurements reveal that cellular senescence leads to significant stiffening of both choroidal ECs and their secreted matrix. Further, although senescent ECs exhibit lower levels of surface-deposited complement MAC, they undergo greater complement-mediated lysis. Additionally, senescent ECs exhibit stronger monocyte adhesion when compared with normal cells, which is further exacerbated by cell surface-deposited MAC. Remarkably, inhibition of cell stiffness (by Y27632) alone reversed all of these inflammatory effects exhibited by senescent ECs.

Conclusions: Thus, these findings implicate age-induced choriocapillaris stiffening in complement-induced EC dysfunction and sub-retinal leukocyte accumulation seen in early AMD. Work is currently underway to examine the mechanical control of choriocapillaris dysfunction in the rhesus macaques model of AMD pathogenesis.

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