Publikationsdatenbank

TNFa induces epithelial barrier perturbation by a self-enhancing effect on TNFR-1 in porcine jejunal epithelial cells (2021)

Art

Poster

Autoren

Droessler, Linda (WE 2)

Cornelius, Valeria (WE 2)

Markov, Alexander (WE 2)

Amasheh, Salah (WE 2)

Kongress

100th annual meeting of the German Physiological Society

Frankfurt am Main, 30.09. – 02.10.2021

Quelle

100th annual meeting of the German Physiological Society : book of abstracts : 30.09.– 02.10.2021 Frankfurt am Main — DPG, Deutsche Physiologische Gesellschaft (Hrsg.)

[S.l.], 2021 — S. 450

Sprache

Englisch

Verweise

URL (Volltext): https://www.dpg2021.de/wp-content/uploads/DPG2021_Abstract-Book.pdf

Kontakt

Institut für Veterinär-Physiologie

Oertzenweg 19 b
14163 Berlin
+49 30 838 62600
physiologie@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Introduction: The pro-inflammatory cytokine Tumor necrosis factor alpha (TNFα) has been described to increase the intestinal permeability by affecting tight junctions (TJ). In our study, the non-transformed porcine jejunal epithelial cell line IPEC-J2 was employed to analyze these effects in detail [1].

Methods: IPEC-J2 cells were grown on cell culture inserts until confluency. Recombinant human TNFα (1000 U/mL) was added to the basolateral compartment of the permeable supports, and transepithelial electrical resistance (TEER) was recorded for 48 hours. For signaling analyses, the specific Myosin light chain kinase blocker ML-7 was employed. Subsequently, tight junction proteins and the TNFα receptor (TNFR) were further analyzed by
immunohistochemistry and immunoblotting. Statistical analysis was performed using one-way ANOVA for normally distributed data, Kruskal-Wallis test for not normally distributed data, and Dunnett’s test for correction of multiple testing. In signaling experiments, Tukey Kramer method was used to ensure pairwise comparisons.

Results: An incubation with TNFα led to a decrease in TEER after 48 hours, while controls remained unchanged. Western blot analysis of TJ proteins revealed a marked decrease of claudin-1, claudin-3 and occludin, after incubation with TNFα. Furthermore, an incubation with the cytokine induced a dose-dependent increase of the expression of its own specific receptor TNFR-1. Confocal microscopy confirmed all observations. Blocker experiments revealed that ML-7 prevented the TNFα-induced decrease of resistance and claudin expression as well as the increase of TNFR-1.

Conclusion: TNFα induced an increase of TNFR-1 and a decrease of claudin-1, -3, and occludin in IPEC-J2, resulting in a barrier perturbation of monolayers mediated via MLCK. This mechanism contributes to explain the exponential nature of TNFα effects on epithelial integrity in inflammatory processes.