IgA-Producing B Cells Exert Regulatory Function through Granzyme B in Kidney Allograft Tolerance

KEY POINTS: Tolerant patients of kidney graft exhibit higher IgA production than other kidney transplanted patients without modification for IgG. IgA-expressing B cells from tolerant patients expressed granzyme B and exert regulatory functions through granzyme B production. BACKGROUND: Tolerant kidney transplant recipients without immunosuppression have a high frequency of circulating granzyme B (GZMB)-expressing regulatory B cells (Breg). Because the precursors of these Bregs remain unknown and IgA-secreting B cells have been shown to have regulatory properties in different situations, we investigated the association between IgA- and GZMB-expressing B cells in a case-control study. METHODS: Forty-five healthy volunteers and 31 kidney transplant recipients with either stable graft function under immunosuppression ( n =10), antibody-mediated rejection ( n =7), or tolerant patients (TOL; n =14) were included. Serum immunoglobulin concentrations as glycosylation were measured by ELISA; forms of IgA were examined by Western blot. Bregs were analyzed by single-cell RNA sequencing and multiparameter spectral flow cytometry. Their function was assessed in cocultured with T cells. RESULTS: Serum IgA concentration was elevated in TOL compared with other transplanted patients, especially the noninflammatory IgA1 subclass, without changes in IgA size or glycosylation. Single-cell transcriptomic analysis revealed higher IgA gene expression in GZMB expressing B cells and conversely higher GZMB gene expression in IgA-expressing B cells. Phenotyping analysis by spectral multiparameter flow cytometry showed that IgA+ B cells were memory B cells, and that IgA+ B cells from TOL tended to express more GZMB compared with antibody-mediated rejection patients. In functional assays, IgA+ B cells exhibited high regulatory functions that were partially prevented by the presence of GZMB inhibitor. CONCLUSIONS: These data show a strong association between IgA+ and GZMB+ Bregs, particularly in tolerant kidney transplant recipients with higher GZMB and IgA expression and greater suppressive properties. CLINICAL TRIAL REGISTRY NAME AND REGISTRATION NUMBER: NCT02900040 .