Background Thymic stromal lymphopoietin (TSLP) is well known for its potent activation of myeloid dendritic cells (mDCs) resulting in Th2-mediated immune responses. TSLP signals cells via the IL-7 receptor-alpha chain (IL-7Rα), shared with IL-7, together with the TSLP receptor (TSLPR) subunit. Recently, we have demonstrated that prevention of TSLPR signalling strongly reduces Th17-driven experimental arthritis and immunopathology. Furthermore, we have shown that administration of TSLP enhances severity of inflammation and joint destruction in collagen induced arthritis.
Objectives To determine the levels of TSLP and numbers of TSLPR-expressing mDCs in joints of rheumatoid arthritis (RA) patients as compared to peripheral blood (PB) and the capacity of TSLP to induce mDC-dependent T-cell activation.
Methods TSLP was measured in synovial fluid (SF) of patients with RA (n=50) and osteoarthritis (OA, n=24) by ELISA. CD1c+ mDC numbers and TSLPR expression on these cells were assessed by FACS analysis in paired samples of SF and PB from RA patients (n=9). CD1c+ mDCs, isolated from PB as well as SF of RA patients (n=6), were stimulated with TSLP for 20 hours and cytokine production was measured by multiplex immunoassay (measuring 51 cytokines). Washed TSLP-activated CD1c+ mDCs from PB (n=11) and SF (n=5) were added to autologous CD4 T cells in the absence of additional stimuli, cultured for 6 days and subsequently proliferation was measured. Additionally, T-cell cytokine production was measured (by ELISA) upon restimulation with ionomycin/PMA.
Results TSLP levels in SF of RA patients were significantly increased compared to OA patients (mean 297 vs. 80 pg/ml, resp., p<0.01). mDCs numbers from SF were significantly increased compared to PB (5.0% vs. 0.6%, resp., p<0.01) and expressed increased levels of TSLPR (MFI 24 vs. 15, resp., p<0.01).
TSLP significantly stimulated production of chemokines TARC and MIP1α by mDCs from PB and SF (TARC: PB from 1 to 42 pg/ml, p<0.05 and SF from 26 to 186 pg/ml, p<0.05; MIP1α: PB from 1268 to 5486 pg/ml, p<0.05 and SF from 2776 to 3733 pg/ml, p<0.05). Upon incubation with TSLP, TSLPR-expressing mDCs from PB potently stimulated proliferation of autologous CD4 T cells as compared to unstimulated mDCs (ratio T cell:DC 5:1, from 1503 to 16036 cpm, p<0.01). However, TSLP-mDCs from SF had a strongly increased capacity to activate CD4 T cells (ratio T cell:DC 5:1, from 26395 to 57387 cpm, p<0.05). Enhanced proliferation was associated with increased production of IFNγ (ratio T cell:DC 5:1, PB from 179 to 655 pg/ml, p<0.01 and SF from 601 to 1867 pg/ml, p<0.05), IL-17 (PB from 39 to 353 pg/ml, p<0.05 and SF from 363 to 1382 pg/ml, p<0.05), and IL-4 (PB from 17 to 246 pg/ml, p<0.01 and SF from 193 to 775 pg/ml, n.s.).
Conclusions Our data indicate that increased intra-articular TSLP concentrations in RA potently activate TSLPR-expressing mDCs from SF to secrete enhanced levels of proinflammatory mediators causing T cell chemotaxis and to potently increase arthritogenic T cell activation. This suggests that TSLP and TSLPR-expressing mDCs could both play an essential role in the immunopathology of RA.
Disclosure of Interest None Declared