gms | German Medical Science

Deutscher Kongress für Orthopädie und Unfallchirurgie (DKOU 2015)

20.10. - 23.10.2015, Berlin

Site-specific effects of LPS on the alveolar epithelial-endothelial capillary barrier

Meeting Abstract

  • presenting/speaker Sabine Fuchs - Experimentelle Unfallchirurgie, Klinik für Unfallchirurgie, Kiel, Germany
  • Janga Harshavardhan - Experimentelle Unfallchirurgie, Klinik für Unfallchirurgie, Kiel, Germany
  • Larissa Ueckert - Experimentelle Unfallchirurgie, Klinik für Unfallchirurgie, Kiel, Germany
  • Stefanie Fitschen-Oestern - Universitätsklinikum Schleswig-Holstein, Campus Kiel, Klinik für Unfallchirurgie, Kiel, Germany
  • Nadine Steubesand - Experimentelle Unfallchirurgie, Klinik für Unfallchirurgie, Kiel, Germany
  • Andreas Seekamp - Universitätsklinikum Schleswig-Holstein, Campus Kiel, Klinik für Unfallchirurgie, Kiel, Germany

Deutscher Kongress für Orthopädie und Unfallchirurgie (DKOU 2015). Berlin, 20.-23.10.2015. Düsseldorf: German Medical Science GMS Publishing House; 2015. DocPO15-1118

doi: 10.3205/15dkou615, urn:nbn:de:0183-15dkou6152

Veröffentlicht: 5. Oktober 2015

© 2015 Fuchs et al.
Dieser Artikel ist ein Open-Access-Artikel und steht unter den Lizenzbedingungen der Creative Commons Attribution 4.0 License (Namensnennung). Lizenz-Angaben siehe http://creativecommons.org/licenses/by/4.0/.


Gliederung

Text

Objectives: Alveolar epithelial cells mediate gaseous exchange in the lung and their dysfunction during acute lung injury/acute respiratory distress syndrome (ALI/ARDS) often leads to worse outcomes, especially when Lipopolysaccharides (LPS), bacterial endotoxins, occur as additional challengers for the alveolar-capillary barrier. In consequence, pro-inflammatory signalling pathways via the activation of nuclear factor-kappa B and mitogen activated protein kinase are induced. The aim of this study was to analyze the effect of LPS on the blood-air barrier using a defined cell culture model in order to reveal the site- specific effects of LPS on the alveolar barrier in a co-culture model in vitro.

Methods: NCI-H441 cells resembling type-II like epithelial cells were seeded on the apical surface of the transwell filter®. For co-cultures OEC (outgrowth endothelial cells) isolated from the peripheral blood were used as a source of endothelial cells and seeded on the basolateral surface of the transwell® to mimic the alveolar capillary barrier. The monocultures (MC) of H441 and the cocultures were cultured for 5 days and treated with10 or 20 microg/ml LPS for 48h from the apical or basolateral compartment and the transepithelial electrical resistance (TEER) was measured using a volt ohm meter (EVOM). We investigated the changes in the barrier properties using TEER measurement, sodium fluorescein transport and immunostaining; pro-inflammatory cytokines (IL6, IL8, IL1-beta) were evaluated by ELISA and qPCR. The role of OEC in barrier break down was determined by treating the H441 MC with conditioned medium from LPS treated OEC.

Results and Conclusion: In cocultures treated with LPS from the basolateral compartment we noticed a significant reduction of TEER, increased permeability and induction of pro-inflammatory cytokines expression, whereas apical treatment did not affect the barrier. No significant change was noticed in H441 MC upon treatment with LPS. However, LPS stimulation of OEC resulted in an increased expression of pro-inflammatory cytokines. Conditioned medium from LPS treated OEC induced the reduction of TEER in H441MC, similar to the effect on the barrier of the H441/OEC cocultures with LPS treatment from basolateral site. Thus, we assume that the epithelial barrier dysfunction is due to soluble factors and pro-inflammatory cytokines secreted by endothelial cells under LPS treatment. This study emphasises the importance of epithelial-endothelial cross talk in alveolar integrity in ALI/ARDS.