Feed‑back loops integrating RELA, SOX18 and FAK mediate the break‑down of the lymph‑endothelial barrier that is triggered by 12(S)‑HETE

Engleitner,Stefanie; Milovanovic,Daniela; Kirisits,Kerstin; Brenner,Stefan; Hong,Junli; Ropek,Nathalie; Huttary,Nicole; Rehak,Judith; Nguyen,Chi   Huu; Bago‑Horvath,Zsuzsanna; Knasmüller,Siegfried; De Martin,Rainer; Jäger,Walter; Krupitza,Georg

doi:10.3892/ijo.2020.4985

Feed‑back loops integrating RELA, SOX18 and FAK mediate the break‑down of the lymph‑endothelial barrier that is triggered by 12(S)‑HETE

Authors:
- Stefanie Engleitner
- Daniela Milovanovic
- Kerstin Kirisits
- Stefan Brenner
- Junli Hong
- Nathalie Ropek
- Nicole Huttary
- Judith Rehak
- Chi Huu Nguyen
- Zsuzsanna Bago‑Horvath
- Siegfried Knasmüller
- Rainer De Martin
- Walter Jäger
- Georg Krupitza
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Affiliations: Department of Pathology, Medical University of Vienna, A‑1090 Vienna, Austria, Department of Clinical Pharmacy and Diagnostics, Faculty of Life Sciences, University of Vienna, A‑1090 Vienna, Austria, Institute of Cancer Research, Department of Internal Medicine 1, Medical University of Vienna, A‑1090 Vienna, Austria, Department of Vascular Biology and Thrombosis Research, Centre of Biomolecular Medicine and Pharmacology, Medical University of Vienna, A‑1090 Vienna, Austria
Published online on: February 13, 2020 https://doi.org/10.3892/ijo.2020.4985
Pages: 1034-1044

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Abstract

Metastatic cancer cells cross endothelial barriers and travel through the blood or lymphatic fluid to pre‑metastatic niches, leading to their colonisation. ‘S’ stereoisomer 12S‑hydroxy‑5Z,8Z,10E,14Z‑eicosatetraenoic acid [12(S)‑HETE] is secreted by a variety of cancer cell types and has been indicated to open up these barriers. In the present study, another aspect of the endothelial unlocking mechanism was elucidated. This was achieved by investigating 12(S)‑HETE‑treated lymph endothelial cells (LECs) with regard to their expression and mutual interaction with v‑rel avian reticuloendotheliosis viral oncogene homolog A (RELA), intercellular adhesion molecule 1, SRY‑box transcription factor 18 (SOX18), prospero homeobox 1 (PROX1) and focal adhesion kinase (FAK). These key players of LEC retraction, which is a prerequisite for cancer cell transit into vasculature, were analysed using western blot analysis, reverse transcription‑quantitative PCR and transfection with small interfering (si)RNA. The silencing of a combination of these signalling and executing molecules using siRNA, or pharmacological inhibition with defactinib and Bay11‑7082, extended the mono‑culture experiments to co‑culture settings using HCT116 colon cancer cell spheroids that were placed on top of LEC monolayers to measure their retraction using the validated ‘circular chemorepellent‑induced defect’ assay. 12(S)‑HETE was indicated to induce the upregulation of the RELA/SOX18 feedback loop causing the subsequent phosphorylation of FAK, which fed back to RELA/SOX18. Therefore, 12(S)‑HETE was demonstrated to be associated with circuits involving RELA, SOX18 and FAK, which transduced signals causing the retraction of LECs. The FAK‑inhibitor defactinib and the NF‑κB inhibitor Bay11‑7082 attenuated LEC retraction additively, which was similar to the suppression of FAK and PROX1 (the target of SOX18) by the transfection of respective siRNAs. FAK is an effector molecule at the distal end of a pro‑metastatic signalling cascade. Therefore, targeting the endothelial‑specific activity of FAK through the pathway demonstrated herein may provide a potential therapeutic method to combat cancer dissemination via vascular routes.

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