Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 Jul 16;17(1):179.
doi: 10.1186/s13075-015-0694-2.

In Vivo Pre-Activation of Monocytes in Patients With Axial Spondyloarthritis

Affiliations
Free PMC article

In Vivo Pre-Activation of Monocytes in Patients With Axial Spondyloarthritis

Kristina Conrad et al. Arthritis Res Ther. .
Free PMC article

Abstract

Introduction: Innate immune responses, including monocyte functions, seem to play an important role in the pathogenesis of axial spondyloarthritis (axSpA). Therefore, we characterized the phenotype and functional state of monocytes of patients with axSpA.

Methods: Fifty-seven patients with axSpA, 11 patients with rheumatoid arthritis (RA), and 29 healthy controls were included in the study. We determined the percentage of classic, intermediate, and non-classic monocytes according to CD14 and CD16 expression and the expression of Toll-like receptor (TLR) 1, 2, and 4 in whole blood by flow cytometry. The percentage of monocytes producing interleukin (IL)-1beta, IL-6, tumor necrosis factor alpha (TNFα), IL-12/23p40, and IL-1 receptor antagonist (IL-1ra) was detected by flow cytometry after stimulation of whole blood without and with different TLR and nucleotide-binding oligomerization domain ligands-i.e., lipopolysaccharide (LPS), fibroblast-stimulating lipopeptid-1, PAM3CSK4, and muramyl dipeptide (MDP)-for 5 h. IL-10 production was measured after 18 h of stimulation in supernatants by enzyme-linked immunosorbent assay.

Results: In patients with axSpA but not patients with RA, we found higher frequencies of classic monocytes than in controls (median of 90.4% versus 80.4%, P < 0.05), higher frequencies of monocytes spontaneously producing IL-1beta and IL-1ra (P < 0.05), and a higher percentage of monocytes producing IL-1beta after MDP stimulation (P < 0.05). Elevated cytokine production was confined to axSpA patients under conventional therapy (non-steroidal anti-inflammatory drugs) and not found in patients under TNFα inhibitor treatment. The LPS-induced production of IL-6 and IL-10 was lower in axSpA patients compared with controls (P < 0.05). Monocytic TLR expression was unaffected in patients with axSpA.

Conclusion: Enhanced spontaneous and MDP-induced cytokine secretion by monocytes suggests in vivo pre-activation of monocytes in axSpA patients under conventional therapy which is reverted under TNF inhibitor treatment.

Figures

Fig. 1
Fig. 1
Elevated percentage of classic monocytes in patients with axial spondyloarthritis (axSpA). a The percentage of classic CD14++CD16 (M1), intermediate CD14++CD16+ (M2), and non-classic CD14+CD16+ (M3) monocytes was determined by fluorescence-activated cell sorting in whole blood of controls, patients with axSpA, and patients with rheumatoid arthritis (RA). The percentage of classic, intermediate, and non-classic monocytes was determined among monocytes which were gated according to forward (FSC) and side (SSC) scatter (R1) and HLA-DR expression (R2). b The percentage of classic, intermediate, and non-classic monocytes (individual measurements and median) is shown from measurements in 12 controls (C), 20 patients with axSpA (SpA), and 12 patients with RA (RA). *P < 0.05, **P < 0.01 (Kruskal-Wallis test and Dunn’s post test). c The percentage of the respective populations in axSpA patients separated into patients under conventional treatment (SpA CT) (n = 12) and patients receiving biologics (i.e., tumor necrosis factor inhibitors) (SpA BT) (n = 8) compared with controls (c). *P < 0.05, **P < 0.01 (Kruskal-Wallis test and Dunn’s post test). FITC fluorescein isothiocyanate
Fig. 2
Fig. 2
Elevated spontaneous and muramyl dipeptide (MDP)-induced proinflammatory cytokine production by monocytes from patients with axial spondyloarthritis (axSpA). Cytokine production by monocytes after stimulation of whole blood without (unstimulated) or with MDP, lipopolysaccharide (LPS), fibroblast-stimulating lipopeptid-1 (FSL), and PAM3CSK4 (PAM) for 5 h with brefeldin A added for the last 3 h. Cytokine production was analysed after intracellular staining by fluorescence-activated cell sorting. Monocytes were identified according to CD68 expression. a A representative example of cytokine production of CD68+ monocytes of a patient with axSpA in whole blood without stimulation and stimulation with the respective stimuli is shown. b The percentage of cytokine-positive cells among CD68+ monocytes (individual measurement and median) in response to the different stimuli in controls (C) (n = 26), patients with axSpA (SpA) (n = 42), and patients with rheumatoid arthritis (RA) (n = 8) is given. *P < 0.05; **P < 0.01 (Kruskal-Wallis test and Dunn’s post test). IL interleukin, TNFα tumour necrosis factor alpha
Fig. 3
Fig. 3
a Elevated spontaneous production of interleukin-1 receptor antagonist (IL-1ra) by monocytes of patients with axial spondyloarthritis (axSpA) and rheumatoid arthritis (RA). The percentage of IL-1ra-producing monocytes among CD68+ monocytes was determined as in Fig. 2. The percentage of cytokine-positive cells (individual measurement and median) in response to the different stimuli in controls (C) (n = 26), patients with axSpA (SpA) (n = 42), and patients with RA (RA) (n = 8) is given. **P < 0.01 (Kruskal-Wallis test and Dunn’s post test). Whole blood of patients with axSpA (SpA) (n = 26) and controls (C) (n = 13) was stimulated without or with indicated stimulators for 18 h. b IL-10 was measured in the supernatant by enzyme-linked immunosorbent assay. *P < 0.05 (Mann–Whitney U test). FSL fibroblast-stimulating lipopeptid-1, LPS lipopolysaccharide, MDP muramyl dipeptide, PAM PAM3CSK4, w/o without
Fig. 4
Fig. 4
Disturbed cytokine responses are found predominately in axial spondyloarthritis patients under conventional treatment. Subanalysis of monocytic cytokine production (Fig. 3) in patients stratified according to treatment into a group of patients receiving conventional treatment spondyloarthritis (CT) (n = 20) and patients receiving biological treatment (i.e., TNF inhibitor treatment) (BT) (n = 12) compared with healthy controls (C) (n = 17). *P < 0.05; **P < 0.01 (Kruskal-Wallis test and Dunn’s post test). FSL fibroblast-stimulating lipopeptid-1, IL interleukin, LPS lipopolysaccharide, MDP muramyl dipeptide, TNFα tumour necrosis factor alpha
Fig. 5
Fig. 5
No difference in Toll-like receptor (TLR) 1, 2, and 4 expression on CD14+ monocytes between axial spondyloarthritis patients and controls. The expression of TLR-1, TLR-2, and TLR-4 on monocytes was determined in healthy controls (C) (n = 16), patients with axial spondyloarthritis (SpA) (n = 22), and patients with rheumatoid arthritis (RA) (n = 9) by surface staining and fluorescence-activated cell sorting and is given as the mean fluorescence intensity (MFI). Monocytes were identified according to CD14 expression

Similar articles

See all similar articles

Cited by 8 articles

See all "Cited by" articles

References

    1. Braun J, Sieper J. Ankylosing spondylitis. Lancet. 2007;369:1379–90. doi: 10.1016/S0140-6736(07)60635-7. - DOI - PubMed
    1. May E, Dorris ML, Satumtira N, Iqbal I, Rehman MI, Lightfoot E, et al. CD8 alpha beta T cells are not essential to the pathogenesis of arthritis or colitis in HLA-B27 transgenic rats. J Immunol. 2003;170:1099–105. doi: 10.4049/jimmunol.170.2.1099. - DOI - PubMed
    1. Colbert RA, DeLay ML, Klenk EI, Layh-Schmitt G. From HLA-B27 to spondyloarthritis: a journey through the ER. Immunol Rev. 2010;233:181–202. doi: 10.1111/j.0105-2896.2009.00865.x. - DOI - PMC - PubMed
    1. Freudenberg MA, Galanos C. Bacterial lipopolysaccharides: structure, metabolism and mechanisms of action. Int Rev Immunol. 1990;6:207–21. doi: 10.3109/08830189009056632. - DOI - PubMed
    1. Beutler B, Kruys V. Lipopolysaccharide signal transduction, regulation of tumor necrosis factor biosynthesis, and signaling by tumor necrosis factor itself. J Cardiovasc Pharmacol. 1995;25:S1–8. doi: 10.1097/00005344-199500252-00002. - DOI - PubMed

Publication types

Feedback