Background Regulatory B cells (Bregs), defined as IL-10 producing B cells, have been well characterized in mice. It has been shown that their transfer can prevent and cure collagen-induced arthritis. In human, very little is known about Bregs in rheumatoid arthritis. Several B cells subsets such as transitional B cells (CD24HiCD38Hi), CD24HiCD27+ and CD5+ B cells were suggested to be precursors of Bregs.
Objectives In the present study, we aimed to compare Bregs generated in culture between RA patients and controls, to correlate Bregs with RA characteristics and with extracellular staining of the different B cell subsets.
Methods To be included, patients should respond to ACR/EULAR 2010 criteria, have corticosteroid doses below 10 mg per day, stable for more than a week and should not have received rituximab in the previous year. Extra-cellular staining for CD5, CD19, CD24, CD38 and CD27 was performed on fresh PBMCs isolated by ficoll and analyzed by flow cytometer. Bregs were generated using frozen PBMCs cultured for 24 hours with CpG ODN B2006, 4 hours with ionomycine / PMA and 2 hours with brefeldine A. Intra-cellular staining of IL-10 was performed after culture.
Results Thirty-one healthy subjects and 103 patients with RA were included and extracellular staining of their PBMCs was performed. Bregs were generated from PBMCs of 16 healthy subjects and 42 patients with RA. CD24HiCD38Hi B cells were lower in RA than in healthy controls (6.5 [2.8-10.5] vs 8.7 [7.2-12.1]% of CD19+, respectively; p=0.03) as well as CD5+ B cells (10.6 [6.7-18.1] vs 16.7 [10.9-23.9], respectively; p=0.005). After adjustment for gender and age; CD24HiCD38Hi tended to be lower in RA subjects (p=0.06) and CD5+ B cells remained significantly lower in this group (p=0.01). None of the different B cell subsets was correlated with IL10 producing B cells. Similar percentage of Bregs were generated in healthy subjects and in RA (medians [IQR]: 7.4 [3.2-14.8] and 5.8 [2.8-9.4]% of CD19+ cells, respectively; p=0.30). However, patients with RA duration under 5 years had significantly less Bregs than healthy controls (2.9 [1.2-5.3] vs 7.4 [3.2-14.8]% B cells; p=0.04). Indeed, Bregs were correlated with RA duration (spearman r=0.32; p=0.04) and patients with a disease duration above 5 years had twice more Bregs than patients with shorter disease duration (6.8 [4.1-11.1] vs 2.9 [1.2-5.3]% of CD19+ cells; p=0.02). Bregs were inversely correlated with swollen joint count (SJC), tender joint count (TDJ), visual analogic scale (VAS) and DAS28 (r=-0.43; p=0.005; r=-0.30; p=0.05; r=-0.40; p=0.01 and r=-0.42; p=0.006, respectively) in all cohort but this was more pronounced in patients with recent RA (spearman r for TJC, SJC and DAS28 were respectively -0.77; -0.68 and -0.84 with p-values of 0.006; 0.02 and 0.001). In recent RA patients, there was also an inverse correlation between Bregs and CRP (Pearson r² for logCRP : 0.69; p=0.002).
Conclusions A lack of Bregs was observed in patients with recent RA. Bregs increased with RA duration and seem to be protective since Bregs inversely correlated with DAS28 especially in patients with RA duration under 5 years.
Acknowledgements To the French Society of Rheumatology and Pfizer (France) for their financial support
Disclosure of Interest None Declared