Differences in serum and synovial CD4+ T cells and cytokine profiles to stratify patients with inflammatory osteoarthritis and rheumatoid arthritis

Abstract

Background: The aim was to investigate CD4⁺T-cell subsets, immune cells and their cytokine profiles in blood and synovial compartments in rheumatoid arthritis (RA) and inflammatory osteoarthritis (OA) to define specific immune signatures.

Methods: Peripheral blood, synovial fluid (SF) and synovial membranes (SM) of RA and OA patients were analyzed. CD4⁺T-cell subset frequencies were determined by flow cytometry, and cytokine concentrations in serum and SF were measured by ELISA.

Results: In peripheral blood, OA patients had altered frequencies of regulatory T-cell subsets, and higher frequencies of Th17 and of Th1/17 cells than RA patients. In the synovial compartment of OA patients, conventional Th17 cells were largely excluded, while Th1/17 cells were enriched and more frequent than in RA patients. Conversely, in the synovial compartment of RA patients, regulatory T cells and Tfh cells were enriched and more frequent then in OA patients. IL-17 and Blys were increased both in serum and SF of RA patients, and correlated with autoantibodies and disease activity. Notably, Blys levels were already significantly elevated in RA patients with low disease activity score in 28 joints (DAS28) and without autoantibody positivity.

Conclusions: Although patients with inflammatory OA have immune activation in the synovial compartment, they display different T-cell subset frequencies and cytokine profiles. Soluble mediators such as Blys might help to discriminate mild clinical forms of RA from inflammatory OA particularly at the onset of the disease.

Keywords: Blys; Cytokines; Inflammatory osteoarthritis; Rheumatoid arthritis; T helper subsets.

Fig. 1

Patients with osteoarthritis (OA) manifest qualitative and quantitative differences in the immune cell infiltrates in the synovial compartment as compared to patients with rheumatoid arthritis (RA). Frequencies (a) and cumulative statistical analysis (b) of immune cell populations (monocytes, CD14⁺, B cells CD19⁺, natural killer cells, CD56⁺, CD3⁺ CD4⁺ T cells and CD3⁺ CD8⁺ T cells) in peripheral blood (a, b) from healthy donors (HD, n = 25) patients with OA (n = 11) and patients with RA (n = 24). Frequencies (c) and cumulative statistical analysis (d) of immune cell populations in synovial fluid (SF) from patients with OA (n = 6) and patients with RA (n = 8): *p ≤ 0.05, **p ≤ 0.005, ***p ≤ 0.0005, Mann-Whitney unpaired two-tailed t test. Mean value ± SEM are reported. ns not significant, PBMC peripheral blood mononuclear cells, MC mononuclear cells

Fig. 2

CD4⁺ T cell subset frequencies in blood and synovial fluid distinguish patients with rheumatoid arthritis (RA) from patients with osteoarthritis (OA). Frequency of CD4⁺ T helper subsets (a, c) and T regulatory subsets (b, d) identified according to phenotypic markers in blood and synovial fluid from healthy donors (HD) (open diamonds), patients with OA (open circles) and patients with RA (closed circles): *p ≤ 0.05, **p ≤ 0.005, Mann-Whitney unpaired two-tailed t test. Mean value ± SEM are reported. Treg T regulatory cells, Th T helper cells

Fig. 3

Tissue distribution of T-helper (Th) and T-regulatory (Treg) subsets in patients with rheumatoid arthritis (RA) and patients with osteoarthritis (OA). Correlation between Th1 (a), Th1/17 (b), Th17 (c), Tfh (d), Tr1 (e) and Treg (f) subsets in paired samples of blood and synovial fluid (SF) isolated from the same patients with OA and the same patients with RA on the same day (values from the same patients are connected by lines): *p ≤ 0.05, **p ≤ 0.005, ***p ≤ 0.0005, Wilcoxon paired two-tailed t test

Fig. 4

Cytokine levels in serum and synovial fluid discriminate between patients with osteoarthritis (OA) and rheumatoid arthritis (RA). Blys (a), IL-10 (b), IL-17A (c) and IL-21 (d) levels in serum from healthy donors (HD) (n = 25), patients with OA (n = 11) and patients with RA (n = 24) and in synovial fluid (SF) from patients with OA (n = 10) and patients with RA (n = 8) patients: *p ≤ 0.05, ** p ≤ 0.005, ***p ≤ 0.0005, Mann-Whitney unpaired two-tailed t test. Mean value ± SEM are reported. ns not significant

Fig. 5

Cytokine levels in serum and synovial fluid (SF) correlate with disease activity and anti-citrullinated protein antibody (ACPA) positivity. Correlation between serum and synovial fluid levels of Blys (a, b), IL-17 (c, d) and IL-10 (e, f) in patients with rheumatoid arthritis (RA) stratified according to ACPA positivity (a, c, e) or to the disease activity score in 28 joints (DAS28) (b, d, f), and cytokine levels in serum and synovial fluid from patients with osteoarthritis (OA). Cytokine concentrations were determined by ELISA: *p ≤ 0.05, **p ≤ 0.005, ***p ≤ 0.0005, two-tailed t test. Mean value ± SEM are reported. ns not significant