Expression and function of connexin 43 protein in mouse and human retinal pigment epithelial cells as hemichannels and gap junction proteins

Shin ichi Akanuma, Hideyuki Higashi, Souhei Maruyama, Koji Murakami, Masanori Tachikawa, Yoshiyuki Kubo, Ken ichi Hosoya

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)

Abstract

The changes in the transport function of the outer blood-retinal barrier (BRB), formed by retinal pigment epithelial (RPE) cells, under pathological conditions need to be understood to normalize the retinal homeostasis in retinal diseases. Connexin 43 (Cx43) is known to be one of the basic units of gap junctions and hemichannels, which are opened by changes in extracellular conditions. The purpose of this study was to clarify the expression of Cx43 in RPE cells of the retina and Cx43 contribution to compound transport functions in RPE cells. Immunohistochemistry using guinea pig-derived polyclonal anti-Cx43 antibodies indicated that Cx43 is localized at the apical and intercellular membrane of mouse RPE cells. In addition, the immunoprecipitation study using the anti-Cx43 antibodies suggested that Cx43 at the intercellular membrane is associated with gap and adherent junctions in mouse RPE cells. The intercellular transfer after scrape loading of Lucifer Yellow (457 g/mol) among a human RPE cell line, ARPE-19 cells, was greater than that of fluorescein isothiocyanate-dextran (∼3000 g/mol). This Lucifer Yellow transfer was significantly inhibited by carbenoxolone, a connexin inhibitor, suggesting that connexins take part in compound transfer via gap junctions. In addition, Lucifer Yellow uptake by ARPE-19 cells in the absence of extracellular Ca2+, which is a condition of hemichannel opening, was increased compared with that under normal conditions. This uptake of Lucifer Yellow in the absence of extracellular Ca2+ was significantly reduced in the presence of hemichannel inhibitors and Cx43-gene silencing conditions. This study suggests the involvement of Cx43 in dye transfer via gap junctions among RPE cells and hemichannel-mediated compound transport between the neural retina and RPE cells.

Original languageEnglish
Pages (from-to)128-137
Number of pages10
JournalExperimental Eye Research
Volume168
DOIs
Publication statusPublished - 2018 Mar

Keywords

  • Connexin 43
  • Gap junction
  • Hemichannel
  • Outer blood-retinal barrier
  • Retinal pigment epithelial cells

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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