Publikationsdatenbank

Nanomolar ouabain stimulates epithelial barrier formation in IPEC-J2 cells (2019)

Art

Poster

Autoren

Fedorova, A.

Cornelius, V. (WE 2)

Okorokova, L.

Krivoi, I.

Markov, A. (WE 2)

Amasheh, S. (WE 2)

Kongress

98th meeting of the German Physiological Society

Ulm, 30.09. – 02.10.2019

Quelle

Acta physiologica Scandinavica

Bandzählung: 227

Heftzählung: S719

Seiten: 130

ISSN: 0001-6772

Sprache

Englisch

Verweise

URL (Volltext): https://onlinelibrary.wiley.com/doi/epdf/10.1111/apha.13384

Kontakt

Institut für Veterinär-Physiologie

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

Abstract / Zusammenfassung

Question:
In addition to its classical function in ion translocation, the Na,K-ATPase has been reported as an important signaling molecule in neuronal, cardiac, vascular and epithelial tissues (Matchkov, Krivoi, 2016). Ouabain, specific ligand of the Na,K-ATPase, serves as the main tool in such investigations. Micromolar concentrations of ouabain have been shown to disturb the molecular organization of tight junctions in epithelial cells and to decrease transepithelial resistance (TER), but nanomolar concentrations have been reported to induce an opposite effect, including an increase in TER (Larre et al., 2014). In the present work, we analyzed the effect of nanomolar ouabain concentration on TER and claudin expression using the porcine jejunal epithelial cell line IPEC-J2.

Methods:
IPEC-J2 cells were seeded and cultured in medium with 10 nM ouabain compared to controls. Starting from day 10, TER values were determined in cell culture inserts with an EVOM volt-ohmmeter. Structural changes of IPEC-J2 cells with ouabain treatment were identified and compared with control cells via macroscopic inspection. In order to characterize claudin expression, proteins were extracted and used for Western blotting, using antibodies raised against tight junction proteins claudin-1, -3 and -8. Statistical analysis was done using the unpaired Student’s t-test. Values below p=0.05 were considered to be statistically significant with denotation as *p<0.05, **p<0.01 and ***p<0.001.

Results and Conclusion:
Incubation of IPEC-J2 monolayers with 10 nM ouabain induced structural differences, such as the presence of more round cells compared with controls without ouabain treatment. Moreover, monolayers treated with 10 nM ouabain showed higher TER, and Western blots revealed a higher claudin-1 expression compared with controls, but levels of claudin-3 and -8 were not changed. Thus, these findings show that ouabain at nanomolar concentrations is able to effectively influence the process of epithelium barrier formation itself.

Acknowledgments:
The study was supported by the Russian Science Foundation grant no. 18-15-00043, and the Deutsche Forschungsgemeinschaft, grant no. DFG AM 141/11-1.