Rituximab counteracts loss of tolerance in membranous nephropathy patients through NK-mediated Treg induction

BackgroundMembranous nephropathy (MN) is a chronic, autoimmune kidney disease characterized by an autoimmune response against podocyte-specific antigens, compromising renal functions. Few studies attempted to mechanistically describe how rituximab influences Th cells and whether this impacts on clinical outcome. MethodsTh cell profile of MN patients was characterized using peripheral blood mononuclear cell samples from a multi-centre, prospective clinical trial. Th cell subsets (Treg, Th17 and Th1) were measured by flow cytometry at baseline and after rituximab treatment during a two years follow-up. In vitro studies on fresh peripheral blood samples and isolated cell populations were performed to investigate the mechanisms through which rituximab induces Treg cells. FindingsBefore rituximab treatment, MN patients had a Th17/Treg imbalance when compared to age-matched healthy donors (p=0.004). Responders had significantly higher proportions of Treg cells post-rituximab than non-responders (p=0.037). In vitro, rituximab treatment induced Treg cells when using peripheral blood samples from MN patients, however the induction was lost when isolated T cells were used instead of whole blood (p=0.02). Rituximab did not directly induce Treg cells but required a specific cellular environment composed by at least B and Natural Killer (NK) cells as observed by treating pooled T, B and NK cells. Finally, using complementation assays, we demonstrated that rituximab-mediated Treg induction relied on cytokines, mainly Transforming growth factor {beta} (TGF{beta}), secreted by activated NK cells. InterpretationRituximab induces Treg through cytokines produced by NK cells probably during the mechanism of B cell depletion by antibody-dependent cell-mediated cytotoxicity. This mechanism could potentiate the likelihood of remission in MN patients by counteracting loss of immune tolerance.