April 2014
Volume 55, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2014
Role of claudin-19 and claudin-3 on the barrier function of human retinal pigment epithelium (RPE)
Author Affiliations & Notes
  • Shaomin Peng
    Surgery/Ophthalmology, Yale University, New Haven, CT
    Ophthalmology, 2nd Hospital of Harbin Medical University, Harbin, China
  • Peter Yu Cheng Zhao
    Surgery/Ophthalmology, Yale University, New Haven, CT
  • Ron A Adelman
    Ophthalmology, Yale University, New Haven, CT
  • Lawrence J Rizzolo
    Surgery/Ophthalmology, Yale University, New Haven, CT
  • Footnotes
    Commercial Relationships Shaomin Peng, None; Peter Zhao, None; Ron Adelman, None; Lawrence Rizzolo, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 2964. doi:https://doi.org/
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      Shaomin Peng, Peter Yu Cheng Zhao, Ron A Adelman, Lawrence J Rizzolo; Role of claudin-19 and claudin-3 on the barrier function of human retinal pigment epithelium (RPE). Invest. Ophthalmol. Vis. Sci. 2014;55(13):2964. doi: https://doi.org/.

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

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Abstract

Purpose: Human RPE uses principally claudin-3 and claudin-19 to form its tight junctions. The ion selectivity of these claudins is not well known. Although newly confluent ARPE-19 expresses many tight junction proteins, the expression of claudins is low and non-uniform. Consequently the tight junctions are rudimentary and non-functional. This circumstance gives a unique opportunity to characterize individual claudins in a background that essentially lacks other claudins, but contains other tight junction proteins.

Methods: ARPE-19 cells were seeded on Snapwell filters and infected with adenoviral vectors that express either GFP, claudin-3 or claudin-19. The expression of claudin and various RPE genes or proteins was monitored by quantitative RT-PCR, immunoblotting, and confocal immunofluorescence microscopy. An Ussing chamber was used to measure the transepithelial electrical resistance (TER), transepithelial electrical potential (TEP), sodium dilution potential, and sodium-potassium bi-ionic potential. From these data, we estimated the permeability of sodium, potassium, and chloride. To examine cell proliferation and migration, the cultures were scratched and subsequently examined using phalloidin and Ki-67 to monitor filamentous actin and cell proliferation, respectively.

Results: Over-expression of either claudin increased the expression of bestrophin 1, PEDF and TRPC4 more than 2-fold. However, only claudin-19 increased the expression of the EGF receptor. Each claudin converted the morphology of cells from squamous to cuboidal. Cell proliferation and the presence of actin stress fibers were reduced. Wounds were healed principally by cell spreading. Each claudin increased the TER and decreased the permeability of sodium, potassium, and chloride. Both claudins were slightly cation-specific, and claudin-3 had a slight preference for sodium over potassium relative to claudin-19.

Conclusions: The study demonstrates that besides regulating paracellular permeability, claudin-3 and claudin-19 regulate gene expression. Both up-regulate the expression of several RPE-specific genes, but only claudin-19 up-regulates the EGF receptor. Claudin-3 and claudin-19 formed general permeability barriers with only slight ion-selectivity relative to the reports of other claudins. The functions of claudin-3 and claudin-19 appear to be redundant except for their regulation of gene expression.

Keywords: 701 retinal pigment epithelium • 446 cell adhesions/cell junctions  
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