Epidermal growth factor protects the apical junctional complexes from hydrogen peroxide in bile duct epithelium

Srikar R. Guntaka, Geetha Samak, Ankur Seth, Nicholas F La Russo, Radhakrishna Rao

Research output: Contribution to journalArticle

20 Scopus citations

Abstract

The tight junctions of bile duct epithelium form a barrier between the toxic bile and liver parenchyma. Disruption of tight junctions appears to have a crucial role in the pathogenesis of various liver diseases. In this study, we investigated the disruptive effect of hydrogen peroxide and the protective effect of epidermal growth factor (EGF) on the tight junctions and adherens junctions in the bile duct epithelium. Oxidative stress in NRC-1 and Mz-ChA-1 cell monolayers was induced by administration of hydrogen peroxide. Barrier function was evaluated by measuring electrical resistance and inulin permeability. Integrity of tight junctions, adherens junctions and the actin cytoskeleton was determined by imunofluorescence microscopy. Role of signaling molecules was determined by evaluating the effect of specific inhibitors. Hydrogen peroxide caused a rapid disruption of tight junctions and adherens junctions leading to barrier dysfunction without altering the cell viability. Hydrogen peroxide rapidly increased the levels of p-MLC (myosin light chain) and c-Src(pY418). ML-7 and PP2 (MLCK and Src kinase inhibitors) attenuated hydrogen peroxide-induced barrier dysfunction, tight junction disruption and reorganization of actin cytoskeleton. Pretreatment of cell monolayers with EGF ameliorated hydrogen peroxide-induced tight junction disruption and barrier dysfunction. The protective effect of EGF was abrogated by ET-18-OCH 3 and the Ro-32-0432 (PLCγ and PKC inhibitors). Hydrogen peroxide increased tyrosine phosphorylation of ZO-1, claudin-3, E-cadherin and Β-catenin, and pretreatment of cells with EGF attenuated tyrosine phosphorylation of these proteins. These results demonstrate that hydrogen peroxide disrupts tight junctions, adherens junctions and the actin cytoskeleton by an MLCK and Src kinase-dependent mechanism in the bile duct epithelium. EGF prevents hydrogen peroxide-induced tight junction disruption by a PLCγ and PKC-dependent mechanism.

Original languageEnglish (US)
Pages (from-to)1396-1409
Number of pages14
JournalLaboratory Investigation
Volume91
Issue number9
DOIs
StatePublished - Sep 2011

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Keywords

  • adherens junctions
  • cholangiocyte
  • EGF
  • oxidative stress
  • protein kinase
  • tight junction

ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Cell Biology
  • Molecular Biology

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