Background Cytokine production by monocytes might contribute to pathogenesis of AS.
Objectives The aim of the study was to analyze cytokine production of monocytes in response to different toll-like-receptor (TLR) ligands. Therefore, whole peripheral blood (PB) and isolated peripheral blood mononuclear cells (PBMNC) were stimulated with different TLR ligands and cytokine production was determined by intracellular cytokine staining and flow cytometry.
Methods PB and PBMNC of 19 AS patients and 12 healthy controls (Co) were stimulated for 5 hours with toll-like-receptor (TLR)-ligands (TLR-4: lipopolysaccharide (LPS), TLR2/6: fibroblast-stimulating lipopeptide (FSL-1), TLR1/2: PAM3CSK4. After stimulation cells were stained for CD68 to identify monocytes and intracellularly for TNF-a, IL-1b, IL-6, p40 (IL12/23), IL-10 and IL-1ra. Cytokine production of CD68+ monocytes was determined by flow cytometry.
In addition we quantified the LPS binding protein (LBP) concentration in the serum of 19 AS patients and 18 Co by enzyme linked immunosorbent assay (ELISA) measurement.
Results In PB stimulated with LPS the frequency of IL-6-producing (57.99±11.56 vs. 76.63±9.395% respectively; p=0.0006) and IL-10-producing (0.9845±1.588 vs. 1.4060±0.938% respectively; p=0.0385) CD68+ monocytes was significantly reduced in AS patients compared to Co. No significant difference was observed for IL-1b, TNF-a and IL-1ra.
Moreover, PB stimulation with other TLR-ligands did not result in significant differences between AS patients and controls. In contrast to PB, stimulation of isolated PBMC with LPS or other TLR ligands, showed no differences in cytokine expression.
These results suggested that serum factors, such as LPS binding protein, might affect LPS-induced cytokine expression in PB. Quantification of LBP revealed a significant increase in AS compared to Co (107.033±65.246 vs. 35.174±18.880 µg/ml respectively; p=0.0006).
Conclusions The LPS-induced IL-6 production by monocytes in PB is impaired. High serum concentration of LBP might contribute to this impairment, as plasma levels of this protein exhibiting LPS neutralizing capacity is strongly enhanced in AS patients.
Disclosure of Interest None Declared
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