The abnormal expression of CCR4 and CCR6 on Tregs in rheumatoid arthritis

The study aims to investigate the frequency of CD4+CD25+Foxp3+CD127- T regulatory cells (Tregs) and the expression of CCR4, CCR6 and/or other chemokine receptors on Tregs in peripheral blood (PB) in patients with rheumatoid arthritis, as well as in PB, draining lymph nodes (dLNs), lungs and spleens in collagen-induced arthritis (CIA) mice. We also study the possible role of CCR4 and CCR6 abnormal expression on Tregs in RA patients and the underlying mechanisms. The numbers of Tregs and chemokine receptors expression profile on Tregs in PB from RA patients and healthy controls were investigated by flow cytometry (FACS) using three- or four-color intracellular staining. DBA/1 Foxp3gfp reporter mice were immunized with collagen II (CII) emulsified with CFA. At day 60 after CII immunization, mice were sacrificed and Foxp3 (GFP) expression in PB, dLNs, Lungs and spleens was examined by FACS. The numbers of Tregs in PB were significantly lower in RA patients than in healthy controls (1.21±0.43% vs 3.50±0.98%, P<0.05). The levels of chemokine receptor CCR4 or CCR6 expression on Tregs in PB were higher in active RA patients than in healthy controls (91.13±2.98% vs 79.45±4.72%, P<0.05; or 67.33±7.53% vs 42.73±5.60%, P<0.05). The levels of CCR4 or CCR6 expression on Tregs in active RA patients were positively correlated to DAS28 scores (r=0.42, P<0.03; or r=0.58, P<0.02). Similarly, the numbers of CCR6 expression on GFP+ cells in the spleens, dLNs, lungs and blood of CIA were all increased than those of normal mice (P<0.01). Frequency of CCR4 expression on GFP+ cells in dLNs of CIA was somehow higher but slightly lower in the spleens of CIA compared to normal mice without significant differences (P>0.05). Frequency of CCR5 expression on GFP+ cells in the spleens and dLNs of CIA were both increased than those of normal mice, but there were no significant differences (P>0.05). CCR7 or CCR9 expression on Tregs from spleen and dLN of either normal or CIA mice was undetectable. Although the frequency of CD4+Foxp3+Tregs in peripheral blood was decreased in active rheumatoid arthritis patients, the levels of chemokine receptors such as CCR4 and CCR6 among the Tregs were increased, implicating that Tregs in active RA have obtained the ability migrating to inflammatory joints and may reflect the feedback regulation of the body to local inflammation. Furthermore, CCR4 and CCR6 expressed on Tregs may be related to the activity and severity of RA.