The Distinctive Activation of Toll-Like Receptor 4 in Human Samples with Sepsis.
Patrick ThonKatharina RumpAnnika KnorrBirte DyckDominik ZieheMatthias UnterbergHartmuth NowakLars BergmannAlexander WolfMaha BazziJennifer OrlowskiMarcus PetersAlexander ZarbockThorsten BrennerMichael AdamzikTim RahmelBjörn KoosPublished in: Cells (2022)
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.
Keyphrases
- toll like receptor
- inflammatory response
- end stage renal disease
- immune response
- nuclear factor
- newly diagnosed
- ejection fraction
- chronic kidney disease
- acute kidney injury
- intensive care unit
- induced apoptosis
- stem cells
- peritoneal dialysis
- high throughput
- cell death
- endothelial cells
- patient reported outcomes
- signaling pathway
- oxidative stress
- cell proliferation
- bone marrow
- endoplasmic reticulum stress
- patient reported
- binding protein
- anti inflammatory
- subarachnoid hemorrhage