DOI

Tumor necrosis factor alpha (TNFα) has been shown to impair the intestinal barrier, inducing and maintaining inflammatory states of the intestine. The aim of the current study was to analyze functional, molecular and regulatory effects of TNFα in a newly established non-transformed jejunal enterocyte model, namely IPEC-J2 monolayers. Incubation with 1000 U/mL TNFα induced a marked decrease in transepithelial electrical resistance (TEER), and an increase in permeability for the paracellular flux marker [3 H]-D-mannitol compared to controls. Immunoblots revealed a significant decrease in tight junction (TJ) proteins occludin, claudin-1 and claudin-3. Moreover, a dose-dependent increase in the TNF receptor (TNFR)-1 was detected, explaining the exponential nature of pro-inflammatory effects, while TNFR-2 remained unchanged. Recovery experiments revealed reversible effects after the removal of the cytokine, excluding apoptosis as a reason for the observed changes. Furthermore, TNFα signaling could be inhibited by the specific myosin light chain kinase (MLCK) blocker ML-7. Results of confocal laser scanning immunofluorescence microscopy were in accordance with all quantitative changes. This study explains the self-enhancing effects of TNFα mediated by MLCK, leading to a differential regulation of TJ proteins resulting in barrier impairment in the intestinal epithelium.

Original languageEnglish
Article number8746
Number of pages19
JournalInternational Journal of Molecular Sciences
Volume22
Issue number16
DOIs
StatePublished - 14 Aug 2021

    Scopus subject areas

  • Molecular Biology
  • Spectroscopy
  • Catalysis
  • Inorganic Chemistry
  • Computer Science Applications
  • Physical and Theoretical Chemistry
  • Organic Chemistry

    Research areas

  • Claudins, Epithelial barrier, IPEC-J2, ML-7, Tight junction, TNFR-1, TNFα, TNF alpha, PERMEABILITY, KINASE, TIGHT JUNCTION BARRIER, FOLLICLE-ASSOCIATED EPITHELIUM, PATHOGENESIS, IN-VITRO, CLAUDIN-2, epithelial barrier, CELL LINE, DYSFUNCTION, claudins, NF-KAPPA-B, tight junction

ID: 86120672