Marginal-Zone B-Cells Are Main Producers of IgM in Humans, and Are Reduced in Patients With Autoimmune Vasculitis

Abstract

In mice, B1 and marginal zone (MZ) B-cells play an important role in prevention of autoimmunity through production of regulatory cytokines and natural antibodies. There is limited knowledge about the human counterparts of these cells. We therefore investigated functions of MZ-like B-cells and the frequency of circulating MZ-like and B1-like B-cells in healthy controls (HC), as well as in patients with autoimmune vasculitis to learn more about the role of these cells in autoimmune disease. After stimulation with CpG oligodeoxynucleotides (ODN) of class B in vitro, MZ-like B-cells were the main producers of IgM whereas switched memory B-cells primarily produced IgG and IgA. TNF and IL-10 were produced by both MZ-like and switched memory B-cells. Neither antibody nor TNF/IL-10 production by the B-cell subsets differed between patients and HC. Patients with autoimmune vasculitis, irrespective of disease activity, had lower percentage and absolute numbers of circulating MZ-like B-cells, and lower absolute numbers of B1-like B-cells. The percentage of B1-like B-cells was reduced during active disease. These findings remained significant when the analysis was confined to active treatment-naïve patients (disease onset).Our results suggest that human innate-like B-cells might have a physiological role in prevention of autoimmunity.

Keywords: B1 B-cells; IgM; T-cell-independent antibodies; anti-neutrophil cytoplasmic autantibody (ANCA) associated vasculitis; autoimmunity; innate-like B-cells; marginal-zone (MZ) B-cells; natural antibodies.

Figure 1

Antibody and cytokine production in vitro by various B-cell subsets. (A) Stimulation of B-cells from healthy controls (HC) with CpG-B oligodeoxynucleotides (ODN) showed that marginal zone (MZ)-like B-cells are the main producers of IgM, whereas switched memory (SwMe) B-cells primarily produces IgG and IgA. (B) Spontaneous Ig production was generally scarce among the different B-cell subsets, but MZ-like B-cells produced IgM spontaneously. (C) B-cells from patients in remission behaved very similar to those from HC concerning antibody production after CpG-B ODN stimulation, with no significant differences between the groups. (D) MZ-like and SwMe B-cells produced most TNF and IL-10 after CpG-B ODN stimulation, but SwMe B-cells from patients produced TNF to a lower extent than SwMe B-cells from HC. There were, however, no differences in the TNF/IL-10 ratio among the various B-cell subsets in patients and HC. Kruskal-Wallis test followed by Dunn's multiple comparison test was used to compare more than two groups with independent observations (A,B), and Mann-Whitney U test was applied in comparisons between two groups with independent observations (C,D). Bars indicate median and inter-quartile range. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Figure 2

Levels of marginal zone (MZ)-like and B1-like B-cells in the blood circulation. (A) Patients with active disease and patients in remission exhibited a reduced percentage of MZ-like B-cells within the B-cell population, (B) as well as lower absolute numbers of these cells in the circulation compared with healthy controls. (C,D) Percentage of B1-like B-cells was only reduced during active disease, whereas absolute numbers were lower in both patient groups. Gray triangles represent 14 treatment-naïve patients with active disease. Kruskal-Wallis test followed by Dunn's multiple comparison test was used to compare more than two groups with independent observations (A–D). Bars indicate median and inter-quartile range. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.001.

Figure 3

Innate-like B-cells and ANCA levels in active treatment-naïve patients. (A,B) Proportions of marginal-zone (MZ)-like B-cells, (C,D) but not B1-like B-cells, correlated negatively with PR3-ANCA and MPO-ANCA levels in active treatment-naïve patients. Spearman's rank correlation coefficient (r_s) was calculated. P < 0.05 is statistically significant.