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The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis

Clinical success of Toll-Like receptor-4 (TLR-4) antagonists in sepsis therapy has thus far been lacking. As inhibition of a receptor can only be useful if the receptor is active, stratification of patients with active TLR-4 would be desirable. Our aim was to establish an assay to quantify phosphory...

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Autores principales: Thon, Patrick, Rump, Katharina, Knorr, Annika, Dyck, Birte, Ziehe, Dominik, Unterberg, Matthias, Nowak, Hartmuth, Bergmann, Lars, Wolf, Alexander, Bazzi, Maha, Orlowski, Jennifer, Peters, Marcus, Zarbock, Alexander, Brenner, Thorsten, Adamzik, Michael, Rahmel, Tim, Koos, Björn
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9563554/
https://www.ncbi.nlm.nih.gov/pubmed/36230982
http://dx.doi.org/10.3390/cells11193020
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author Thon, Patrick
Rump, Katharina
Knorr, Annika
Dyck, Birte
Ziehe, Dominik
Unterberg, Matthias
Nowak, Hartmuth
Bergmann, Lars
Wolf, Alexander
Bazzi, Maha
Orlowski, Jennifer
Peters, Marcus
Zarbock, Alexander
Brenner, Thorsten
Adamzik, Michael
Rahmel, Tim
Koos, Björn
author_facet Thon, Patrick
Rump, Katharina
Knorr, Annika
Dyck, Birte
Ziehe, Dominik
Unterberg, Matthias
Nowak, Hartmuth
Bergmann, Lars
Wolf, Alexander
Bazzi, Maha
Orlowski, Jennifer
Peters, Marcus
Zarbock, Alexander
Brenner, Thorsten
Adamzik, Michael
Rahmel, Tim
Koos, Björn
author_sort Thon, Patrick
collection PubMed
description Clinical success of Toll-Like receptor-4 (TLR-4) antagonists in sepsis therapy has thus far been lacking. As inhibition of a receptor can only be useful if the receptor is active, stratification of patients with active TLR-4 would be desirable. Our aim was to establish an assay to quantify phosphorylated TLR-4 using the proximity ligation assay (PLA). HEK293 TLR4/MD2/CD14 as well as THP-1 cells were stimulated with LPS and the activation of TLR-4 was measured using the PLA. Furthermore, peripheral blood mononuclear cells (PBMCs) from 25 sepsis patients were used to show the feasibility of this assay in clinical material. Activation of TLR-4 in these samples was compared to the PBMCs of 11 healthy individuals. We could show a transient activation of TLR-4 in both cell lines. Five min after the LPS stimulation, the signal increased 6.7-fold in the HEK293 cells and 4.3-fold in the THP-1 cells. The assay also worked well in the PBMCs of septic patients. Phosphorylation of TLR-4 at study inclusion was 2.9 times higher in septic patients compared to healthy volunteers. To conclude, we established a diagnostic assay that is able to quantify the phosphorylation of TLR-4 in cell culture and in clinical samples of sepsis patients. This makes large-scale stratification of sepsis patients for their TLR-4 activation status possible.
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spelling pubmed-95635542022-10-15 The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis Thon, Patrick Rump, Katharina Knorr, Annika Dyck, Birte Ziehe, Dominik Unterberg, Matthias Nowak, Hartmuth Bergmann, Lars Wolf, Alexander Bazzi, Maha Orlowski, Jennifer Peters, Marcus Zarbock, Alexander Brenner, Thorsten Adamzik, Michael Rahmel, Tim Koos, Björn Cells Article Clinical success of Toll-Like receptor-4 (TLR-4) antagonists in sepsis therapy has thus far been lacking. As inhibition of a receptor can only be useful if the receptor is active, stratification of patients with active TLR-4 would be desirable. Our aim was to establish an assay to quantify phosphorylated TLR-4 using the proximity ligation assay (PLA). HEK293 TLR4/MD2/CD14 as well as THP-1 cells were stimulated with LPS and the activation of TLR-4 was measured using the PLA. Furthermore, peripheral blood mononuclear cells (PBMCs) from 25 sepsis patients were used to show the feasibility of this assay in clinical material. Activation of TLR-4 in these samples was compared to the PBMCs of 11 healthy individuals. We could show a transient activation of TLR-4 in both cell lines. Five min after the LPS stimulation, the signal increased 6.7-fold in the HEK293 cells and 4.3-fold in the THP-1 cells. The assay also worked well in the PBMCs of septic patients. Phosphorylation of TLR-4 at study inclusion was 2.9 times higher in septic patients compared to healthy volunteers. To conclude, we established a diagnostic assay that is able to quantify the phosphorylation of TLR-4 in cell culture and in clinical samples of sepsis patients. This makes large-scale stratification of sepsis patients for their TLR-4 activation status possible. MDPI 2022-09-27 /pmc/articles/PMC9563554/ /pubmed/36230982 http://dx.doi.org/10.3390/cells11193020 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Thon, Patrick
Rump, Katharina
Knorr, Annika
Dyck, Birte
Ziehe, Dominik
Unterberg, Matthias
Nowak, Hartmuth
Bergmann, Lars
Wolf, Alexander
Bazzi, Maha
Orlowski, Jennifer
Peters, Marcus
Zarbock, Alexander
Brenner, Thorsten
Adamzik, Michael
Rahmel, Tim
Koos, Björn
The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis
title The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis
title_full The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis
title_fullStr The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis
title_full_unstemmed The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis
title_short The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis
title_sort distinctive activation of toll-like receptor 4 in human samples with sepsis
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9563554/
https://www.ncbi.nlm.nih.gov/pubmed/36230982
http://dx.doi.org/10.3390/cells11193020
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