Gut microbiota-specific IgA+ B cells traffic to the CNS in active multiple sclerosis

Abstract

Changes in gut microbiota composition and a diverse role of B cells have recently been implicated in multiple sclerosis (MS), a central nervous system (CNS) autoimmune disease. Immunoglobulin A (IgA) is a key regulator at the mucosal interface. However, whether gut microbiota shape IgA responses and what role IgA⁺ cells have in neuroinflammation are unknown. Here, we identify IgA-bound taxa in MS and show that IgA-producing cells specific for MS-associated taxa traffic to the inflamed CNS, resulting in a strong, compartmentalized IgA enrichment in active MS and other neuroinflammatory diseases. Unlike previously characterized polyreactive anti-commensal IgA responses, CNS IgA cross-reacts with surface structures on specific bacterial strains but not with brain tissue. These findings establish gut microbiota-specific IgA⁺ cells as a systemic mediator in MS and suggest a critical role of mucosal B cells during active neuroinflammation with broad implications for IgA as an informative biomarker and IgA-producing cells as an immune subset to harness for therapeutic interventions.

Fig. 1.. Identification of IgA-coated bacteria enriched in multiple sclerosis patients.

A, Overview of IgA-based cell sorting of fecal bacteria combined with 16S rRNA gene sequencing (IgA-SEQ) in multiple sclerosis (MS) (n=36) and healthy controls (HC) (n=31). B, Gating strategy for flow-cytometry based sorting of IgA-bound (IgA+) and -unbound (IgA-) bacteria. C, Relative counts of significant operational taxonomic units (OTUs) specific to the IgA+ and IgA- fractions, non-differential OTUs and global OTUs. Note significant differential enrichment of IgA-bound OTUs in MS compared to controls as shown by an elevated global microbiota coating index (GMCI). Fisher’s exact test. D, Phylogenetic tree based on IgA-SEQ of significantly different OTUs between IgA+ and IgA- in MS or HC. Circles represent OTUs overabundant in the respective IgA status. E, Volcano plots of significantly differential OTUs between IgA+ and IgA- in MS. Note enrichment of pro-inflammatory taxa in the IgA-bound fraction. F, Heatmap of average relative abundance (log base 10) of total, IgA+ and IgA- fractions of fecal bacteria and IgA coating index (ICI) scores from HC and MS for differential OTUs between IgA+ and IgA- in HC and MS, respectively. DESeq2, Benjamini-Hochberg correction for multiple comparisons.

Fig. 2.. Enrichment of IgA-producing cells during active CNS inflammation in MS and viral encephalitis.

A, Serum and CSF IgA and IgG levels during MS relapse (serum: n=13, CSF: n=11) and remission (serum: n=11, CSF: n=9); unpaired Student’s t-test. IgA and IgG index during relapse (n=6) compared to remission (n=11); unpaired Student’s t-test with Welch’s correction. CSF IgA in active and inactive MS in comparison to another neuroinflammatory condition (neurosarcoidosis: n=12), neurodegeneration (motor neuron diseases, MND: n=10) and healthy controls (HC: n=10); One-way ANOVA with Tukey’s multiple comparison test; Indicated are mean ± SEM. B, Spatial distribution and lineage of IgA+ B cells in active MS lesions. CD20 (cyan arrowheads), IgA (yellow arrowheads). C, Spatial transcriptomics of IGHA- to IGHG-expressing cells in subcortical perivascular versus meningeal and parenchymal infiltrates in active MS; two-tailed Mann-Whitney test. D, IGHA- and IGHG-expressing cells in inflamed meningeal aggregates (left), the majority of which co-express the plasma cell marker PRDM1 (yellow arrowheads). Presence of IGHG- and IGHA-expressing cells (co-expressed with PRDM1) in inflamed perivascular infiltrates (right). Note differential IL10 expression in immune cell subsets with increased expression in IGHA-expressing B cells (yellow arrowheads; orange arrowheads marks IL10-/IGHA+ cells), and lower/absent expression in T cells (CD3E) and macrophages (CD163) (blue arrowheads). E, F, Qualitative (E) and quantitative (F) assessment of IGHA- and IGHG-expressing B cells in inflamed viral encephalitis tissue (Borna disease virus 1, BoDV-1); note recruitment of IGHA- and IGHG-expressing B cells in perivascular spaces with relative enrichment of IGHA- over IGHG-expressing cells in parenchymal infiltrates. For IHC (B) and ISH (C-F) experiments, representative images from individual tissue sections are shown (ctrl, n = 5; MS, n = 12; BoDV-1, n= 5), meninges and perivascular white matter lesions are from the same tissue sections. Inflammatory lesion staging was carried out by MOG and CD68 IHC.

Figure 3.. IgA-producing cells traffic across the blood-brain barrier in active MS.

A-B, B cell receptor repertoire analysis from sorted plasma cell subsets (A) or single cells (B) from peripheral blood (PB, red) and cerebrospinal fluid (CSF, blue) (n=3). PC=plasma cells. Edges (lines): clonal connections between B cell subsets. PB-CSF connections: blue=clonal connections between IgA subsets, red=all other clonal connections. Node size and edge thickness are relative to the number of Ig-VH clusters and connections, respectively. C, Phylogenetic tree of CDR3 sequences from an IgA cluster shared between CSF and blood (patient bBCR2, (A)). Dots indicate amino acids identical to germline segments; color-shaded amino acids indicate differences from germline. D, CSF IgA B cell (clonal) frequencies. Shown are % CSF IgA of all CSF Ig clusters (left), % CSF-PB clonal connections of all CSF IgA (middle), % CSF-PB clonal connections of all CSF-PC Ig clonal connections. E, CSF IgA B cell receptor gene usage.

Figure 4.. Microbial specificity of cerebrospinal fluid IgA B cells.

A, Strategy to characterize gut bacteria-reactivity of cerebrospinal fluid (CSF) IgA and recombinant monoclonal antibodies (mAbs) derived from single CSF IgA B cells from MS patients. B, CSF IgA-reactivity against autologous gut microbiota (remission, n=7; relapse, n=10). Fisher’s exact test. C, CSF mAb-reactivity against autologous gut microbiota (n=5) during acute inflammation. D, CSF mAb binding profile. Strains used are type-strains derived from ATCC or from an Eggerthellaceae collection. MOG=myelin oligodendrocyte glycoprotein, dsDNA=double stranded deoxyribonucleic acid. Dark blue=gram- strains, light blue=gram+ strains. E, Epitope mapping of mAb-1. Antibody reactivity against E. coli K235-derived lipopolysaccharide (LPS) and epitope identification using a glycan array. Shown are the repeating units of the capsular polysaccharides from Streptococcus pneumonia serotypes 4 and 9 with the common 2-N-acetyl mannosamine highlighted in red. F, Schematic illustrates working model of IL10-producing gut microbiota-specific IgA B cell trafficking during acute neuroinflammation in active MS.