Primary B-cell deficiencies reveal a link between human IL-17-producing CD4 T-cell homeostasis and B-cell differentiation

Abstract

IL-17 is a pro-inflammatory cytokine implicated in autoimmune and inflammatory conditions. The development/survival of IL-17-producing CD4 T cells (Th17) share critical cues with B-cell differentiation and the circulating follicular T helper subset was recently shown to be enriched in Th17 cells able to help B-cell differentiation. We investigated a putative link between Th17-cell homeostasis and B cells by studying the Th17-cell compartment in primary B-cell immunodeficiencies. Common Variable Immunodeficiency Disorders (CVID), defined by defects in B-cell differentiation into plasma and memory B cells, are frequently associated with autoimmune and inflammatory manifestations but we found no relationship between these and Th17-cell frequency. In fact, CVID patients showed a decrease in Th17-cell frequency in parallel with the expansion of activated non-differentiated B cells (CD21(low)CD38(low)). Moreover, Congenital Agammaglobulinemia patients, lacking B cells due to impaired early B-cell development, had a severe reduction of circulating Th17 cells. Finally, we found a direct correlation in healthy individuals between circulating Th17-cell frequency and both switched-memory B cells and serum BAFF levels, a crucial cytokine for B-cell survival. Overall, our data support a relationship between Th17-cell homeostasis and B-cell maturation, with implications for the understanding of the pathogenesis of inflammatory/autoimmune diseases and the physiology of B-cell depleting therapies.

Figure 1. Th17 cells in patients with CVID.

(A) Correlation between Th17 frequency and frequency of switched-memory B cells (left), transitional B cells (middle) or CD21^low B cells (right), in CVID patients. Switched-memory B cells were defined as CD27⁺ IgD⁻ cells, transitional B cells as CD38^high IgM^high cells and CD21^low B cells were defined as CD21^low CD38^low cells within gated B cells (CD19⁺) after surface staining of whole blood samples. IL-17 expression was assessed at the single-cell level by intracellular staining following short-term stimulation of PBMC with PMA and ionomycin. (B) Th17 frequency in CVID individuals stratified according to their clinical manifestations, namely autoimmunity, lymphoid proliferation, splenomegaly, and adenopathies. (C) Th17 frequency in healthy controls and CVID patients grouped according to EUROclass. (D) Correlations between frequencies of activated CD4 T cells, defined by concurrent expression of HLA-DR and CD38, and of Th17 cells in CVID and healthy individuals. Each symbol represents one individual. Bars represent mean. Data were compared using Mann-Whitney test, and P values are shown. Correlation significance was assessed using Spearman coefficient test, and r and P values are shown.

Figure 2. Decreased frequency of Th17 cells in individuals lacking B cells.

(A) Representative dot-plots of the analysis of IFN-γ and IL-17 production by CD4 T cells, determined by intracellular staining, for a healthy individual (left), a CVID patient (middle) and a Congenital Agammaglobulinemia (Agamma) patient (right). Numbers inside dot-plots represent the proportion of cells expressing the markers. (B) Frequency of Th17 cells (left) and CXCR5⁺ CD4 T cells (right) in healthy individuals and Congenital Agammaglobulinemia patients (Agamma). Each symbol represents one individual. Bars represent mean. Data were compared using Mann-Whitney test, and P values are shown.

Figure 3. Direct correlation between the frequencies of circulating Th17 cells and switched-memory B cells in healthy individuals.

Correlation between the frequencies of switched-memory (CD27⁺IgD⁻) B cells and Th17 cells in healthy individuals. Each symbol represents one healthy individual. Correlation significance was assessed using Spearman coefficient test, and r and P values are shown.

Figure 4. Negative correlation between the frequency of Th17 cells and serum BAFF levels in healthy subjects.

(A) Correlation between the frequency of Th17 cells and serum levels of the cytokine BAFF, as determined by ELISA, in healthy individuals. (B) Analyses of the serum levels of BAFF in healthy individuals, CVID and Congenital Agammaglobulinemia patients. Each symbol represents one individual. Data were compared using Mann-Whitney test, and P values are shown. (C) The same correlation described in (A) for CVID patients and Congenital Agammaglobulinemia patients. Each symbol represents one individual. Correlation significance was determined using Spearman coefficient test, and r and P values are shown.