Persistent expression of autoantibodies in SLE patients in remission

Abstract

A majority of the antibodies expressed by nascent B cells in healthy humans are self-reactive, but most of these antibodies are removed from the repertoire during B cell development. In contrast, untreated systemic lupus erythematosus (SLE) patients fail to remove many of the self-reactive and polyreactive antibodies from the naive repertoire. Here, we report that SLE patients in clinical remission continue to produce elevated numbers of self-reactive and polyreactive antibodies in the mature naive B cell compartment, but the number of B cells expressing these antibodies is lower than in patients with active disease. Our finding that abnormal levels of self-reactive mature naive B cells persist in the majority of patients in clinical remission suggests that early checkpoint abnormalities are an integral feature of SLE.

Figure 1.

Ig heavy and light chain gene features. The absolute number of sequences analyzed is indicated in the center of each pie chart. (A) V_H and J_H repertoire. (B) IgH CDR3⁺ charges and length. Pie charts show percent of IgH CDR3s with zero, one, two, or three positively charged amino acid (aa) residues. Bar graphs show frequency of IgH CDR3s with ≤9 aa (white bars), 10–14 aa (light gray bars), 15–19 aa (dark gray bars), and 20 aa (black bars). (C) Igκ V and J gene family usage. (D) Igλ V and J gene family usage. p-values indicated below the charts or below horizontal lines are in comparison with healthy controls or data obtained from the same SLE patient during active disease, respectively (references , , and 7). Values for controls (GO, JB, and JH) and untreated SLE patients (SLE100, SLE101, and SLE122) in this and other figures were previously published and are shown here for comparison (references , , and 7).

Figure 2.

Autoreactive antibodies. (A) Pie charts summarize self-reactivity as measured by HEp-2 cell ELISA and HEp-2 cell immunofluorescence assay (Tables S1–S6, Fig. S2, and not depicted) with the number of tested antibodies indicated in the center of each pie chart. Graphs show HEp-2 cell ELISA results. Red lines show the low positive serum control and horizontal lines indicate cutoff OD₄₀₅ levels for positive reactivity. (B) Polyreactivity was measured by ELISA with single-stranded DNA, double-stranded DNA, insulin, and LPS. The frequency of polyreactive clones is summarized in the pie charts with the number of tested antibodies indicated in the pie chart centers. Graphs show polyreactivity ELISA results. Dotted lines show ED38⁺ control (references and 29) and horizontal lines indicate cutoff OD₄₀₅ levels for reactivity. p-values indicated below the pie charts or below the horizontal lines are in comparison with healthy controls or data obtained from the same SLE patient with active disease, respectively (references , , and 7).

Figure 3.

Ig gene usage and antibody self-reactivity of mature naive B cells from SLE patients in clinical remission. (A) Jκ genes were grouped according to their 5′ (Jκ1) and 3′ (Jκ3/4/5) position, and the proportion of self-reactive (black) versus nonself-reactive antibodies (white) is indicated as a bar graph for each group with the ratio of self-reactive versus nonself-reactive (S/NS) antibodies indicated below. p-values are in comparison to antibodies of the corresponding Jκ group from healthy controls. (B) Vκ genes were divided into four groups according to their 5′ to 3′ orientation in the Igκ locus (VκA–VκD). The proportion of self-reactive antibodies in each Vκ group for healthy controls (references and 7), SLE patients before treatment (reference 2), or SLE patients in clinical remission is indicated.