Neutrophil Gelatinase–Associated Lipocalin Protects from ANCA-Induced GN by Inhibiting TH17 Immunity

Adrian Schreiber; Anthony Rousselle; Jan Klocke; Sebastian Bachmann; Suncica Popovic; Julia Bontscho; Kai M. Schmidt-Ott; Volker Siffrin; Uwe Jerke; Muhammad Imtiaz Ashraf; Ulf Panzer; Ralph Kettritz

doi:10.1681/ASN.2019090879

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Visual Abstract

Significance Statement

Neutrophil gelatinase–associated lipocalin (NGAL) is produced by injured renal cells and by neutrophils that are central to ANCA-associated vasculitis. The authors show that circulating and urinary NGAL is not only a marker for ANCA-induced necrotizing crescentic GN, but also that neutrophil NGAL is involved mechanistically in ANCA-associated vasculitis. They demonstrate that ANCA-activated neutrophils release NGAL, and that chimeric mice deficient in neutrophil-derived NGAL develop accelerated myeloperoxidase-ANCA–induced crescentic GN, with increased renal CD4⁺ T cells—particularly T helper 17 (T_H17) cells—acting as mediators of the accelerated phenotype. They also demonstrated that iron siderophore–loaded NGAL suppresses T_H17 polarization. Their findings indicate that bone marrow–derived NGAL, presumably from neutrophils, protects from ANCA-induced necrotizing and crescentic GN by downregulating T_H17 immunity.

Abstract

Background Neutrophil gelatinase–associated lipocalin (NGAL) is a diagnostic marker of intrinsic kidney injury produced by damaged renal cells and by neutrophils. ANCA-associated vasculitis features necrotizing crescentic GN (NCGN), and ANCA-activated neutrophils contribute to NCGN. Whether NGAL plays a mechanistic role in ANCA-associated vasculitis is unknown.

Methods We measured NGAL in patients with ANCA-associated vasculitis and mice with anti-myeloperoxidase (anti-MPO) antibody–induced NCGN. We compared kidney histology, neutrophil functions, T cell proliferation and polarization, renal infiltrating cells, and cytokines in wild-type and NGAL-deficient chimeric mice with anti-MPO antibody–induced NCGN. To assess the role of T_H17 immunity, we transplanted irradiated MPO-immunized MPO-deficient mice with bone marrow from either wild-type or NGAL-deficient mice; we also transplanted irradiated MPO-immunized MPO/IL-17A double-deficient mice with bone marrow from either IL-17A–deficient or NGAL/IL-17A double-deficient mice.

Results Mice and patients with active ANCA-associated vasculitis demonstrated strongly increased serum and urinary NGAL levels. ANCA-stimulated neutrophils released NGAL. Mice with NGAL-deficient bone marrow developed worsened MPO-ANCA–induced NCGN. Intrinsic neutrophil functions were similar in NGAL-deficient and wild-type neutrophils, whereas T cell immunity was increased in chimeric mice with NGAL-deficient neutrophils with more renal infiltrating T_H17 cells. NGAL-expressing neutrophils and CD3⁺ T cells were in close proximity in kidney and spleen. CD4⁺ T cells showed no intrinsic difference in proliferation and polarization in vitro, whereas iron siderophore–loaded NGAL suppressed T_H17 polarization. We found significantly attenuated NCGN in IL-17A–deficient chimeras compared with MPO-deficient mice receiving wild-type bone marrow, as well as in NGAL/IL-17A–deficient chimeras compared with NGAL-deficient chimeras.

Conclusions Our findings support that bone marrow–derived, presumably neutrophil, NGAL protects from ANCA-induced NCGN by downregulating T_H17 immunity.

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