April 2014
Volume 55, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2014
RPE tight junctions are regulated by endothelial cells
Author Affiliations & Notes
  • Ignacio Benedicto
    Ophtalmology, Weill Cornell Medical College, New York, NY
    Margaret Dyson Vision Research Institute, Weill Cornell Medical College, New York, NY
  • Guillermo Lehmann-Mantaras
    Ophtalmology, Weill Cornell Medical College, New York, NY
    Margaret Dyson Vision Research Institute, Weill Cornell Medical College, New York, NY
  • Shahin Rafii
    Genetic Medicine, Weill Cornell Medical College, New York, NY
    Ansary Stem Cell Institute, Weill Cornell Medical College, New York, NY
  • Enrique Rodriguez-Boulan
    Ophtalmology, Weill Cornell Medical College, New York, NY
    Margaret Dyson Vision Research Institute, Weill Cornell Medical College, New York, NY
  • Footnotes
    Commercial Relationships Ignacio Benedicto, None; Guillermo Lehmann-Mantaras, None; Shahin Rafii, None; Enrique Rodriguez-Boulan, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 2979. doi:
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    • Get Citation

      Ignacio Benedicto, Guillermo Lehmann-Mantaras, Shahin Rafii, Enrique Rodriguez-Boulan; RPE tight junctions are regulated by endothelial cells. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2979.

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

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Abstract

Purpose: The retinal pigment epithelium (RPE) separates the neural retina from the fenestrated choroid endothelium. RPE tight junctions (TJs) are key for the correct maintenance of the outer blood-retinal barrier. It has been shown that RPE TJs are affected by factors secreted by the neural retina. However, it is not known whether RPE TJs are also regulated by choroid endothelial cells. Recent studies have demonstrated that endothelial cells play key instructive roles in the differentiation and maintenance of parenchymal cells in several body organs. Thus, we studied whether endothelial cells regulate RPE TJ function.

Methods: We performed co-culture experiments on Transwell inserts using human fetal RPE (hfRPE) and human umbilical vein endothelial cells expressing the adenoviral protein E4 (E4-HUVECs). E4-HUVECs are able to survive in the absence of endothelial factors or serum. hfRPE were seeded on top of the filter and E4-HUVECs were seeded either on the other side of the filter or on the bottom chamber of the insert. Transepithelial electrical resistance (TER) was measured at different time points in the absence or presence of E4-HUVECs. As controls, several cell lines were used instead of E4-HUVECs. Alternatively, co-cultures were carried out with E4-HUVECs that had been previously exposed to polarized hfRPE conditioned medium. Expression levels of the TJ-associated proteins claudin-1, -2, -3, -9, -10b, -12, -15, -16, -19 and occludin were analyzed in hfRPE at different time points by real time PCR.

Results: hfRPE co-cultured with E4-HUVECs presented a significantly increased TER both in the presence or absence of serum. This effect was was not detected when other cell types were used instead of endothelial cells. The increase in TER was faster when E4-HUVECs had been previously primed with hfRPE. We also observed that the presence of endothelial cells induced a specific decrease in hfRPE claudin-2 mRNA levels.

Conclusions: Endothelial cells altered both TER and claudin-2 expression in hfRPE in vitro, suggesting that choroidal endothelium may modulate RPE TJ function. In addition, the fact that the effect on TER was faster when endothelial cells were previously primed with hfRPE suggests a bidirectional communication model in which endothelial cells receive a signal from RPE which in turn allows them to modulate RPE tight junctions. This model fits well with the observed synchronized development of RPE and choroidal endothelium.

Keywords: 701 retinal pigment epithelium • 452 choroid  
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