Clinical and experimental data indicate that anti-neutrophil cytoplasmic autoantibodies (ANCAsAnti-neutrophil cytoplasmic autoantibodies (ANCA) are specific for proteins in the cytoplasm of neutrophils and monocytes. The major target antigens in patients with vasculitis and glomerulonephritis are myeloperoxidase (MPO) and proteinase 3 (PR3). ANCAs occur in greater than 80% of patients with active untreated Wegener's granulomatosis, microscopic polyangiitis, and pauci-immune crescentic glomerulonephritis.1 There is compelling clinical and experimental evidence that ANCA IgG causes ANCA-associated vasculitis and glomerulonephritis. The strongest clinical evidence for causation is the observation that a newborn child developed glomerulonephritis and pulmonary hemorrhage shortly after delivery from a mother with MPO-ANCA-associated microscopic polyangiitis, apparently caused by transplacental transfer of ANCA IgG.2,3 Two compelling animal models of ANCA vasculitis and glomerulonephritis have been described by two different research groups.4,5 Xiao and colleagues 4 induced glomerulonephritis and systemic vasculitis by intravenous injection of either anti-MPO IgG or anti-MPO splenocytes derived from MPO knockout mice immunized with murine MPO. Induction of glomerulonephritis by anti-MPO IgG in this model is enhanced by cytokines 6 and requires neutrophils. 7 Little and colleagues 5 immunized rats with human MPO, resulting in the production of antibodies that cross reacted with rat MPO and caused vasculitis and glomerulonephritis. The pathogenic effects of these anti-MPO antibodies were augmented by cytokines.Numerous in vitro studies indicate that ANCA IgG can activate neutrophils and cause them to release proinflammatory factors. The expression of ANCA antigens (MPO and PR3) at the surface of neutrophils where they are accessible to interact with ANCA IgG is enhanced by proinflammatory cytokines, such as tu-

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Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) constitutes life-threatening autoimmune diseases affecting every organ, including the kidneys, where they cause necrotizing crescentic glomerulonephritis. ANCA activates neutrophils and activated neutrophils damage the endothelium, leading to vascular inflammation and necrosis. Better understanding of neutrophil-mediated AAV disease mechanisms may reveal novel treatment strategies. Here we report that ANCA induces neutrophil extracellular traps (NETs) via receptor-interacting protein kinase (RIPK) 1/3- and mixed-lineage kinase domain-like (MLKL)-dependent necroptosis. NETs from ANCA-stimulated neutrophils caused endothelial cell (EC) damage in vitro. This effect was prevented by () pharmacologic inhibition of RIPK1 or () enzymatic NET degradation. The alternative complement pathway (AP) was recently implicated in AAV, and C5a inhibition is currently being tested in clinical studies. We observed that NETs provided a scaffold for AP activation that in turn contributed to EC damage. We further established the in vivo relevance of NETs and the requirement of RIPK1/3/MLKL-dependent necroptosis, specifically in the bone marrow-derived compartment, for disease induction using murine AAV models and in human kidney biopsies. In summary, we identified a mechanistic link between ANCA-induced neutrophil activation, necroptosis, NETs, the AP, and endothelial damage. RIPK1 inhibitors are currently being evaluated in clinical trials and exhibit a novel therapeutic strategy in AAV.

Clonal hematopoiesis of indeterminate potential (CHIP) occurs in an age-related manner and associates with an increased risk of hematologic cancer, atherosclerotic disease, and shorter overall survival. Little is known about the cell of origin, repartition patterns of clonal mutations within the hematopoietic differentiation tree, and its dynamics under evolutionary pressure. Using targeted sequencing, CHIP was identified in 121 out of 437 elderly individuals (27.7%). Variant allele frequencies (VAFs) of 91 mutations were studied in six peripheral blood cell fractions. VAFs were significantly higher in monocytes, granulocytes, and NK-cells compared to B- or T cells. In all cases with available bone marrow material, mutations could be identified in LinCD34CD38 HSCs with subsequent expansion to myeloid primed progenitors. In 22 patients with solid cancer receiving (radio-)chemotherapy, longitudinal study of 32 mutations at 121 time points identified relative VAF changes of at least 50% in 13/32 mutations. VAFs of DNMT3A, were stable in 12/13 cases (P < .001). Cancer patients with a clonal mutation other than DNMT3A required more often red blood cell transfusions and dose reductions. Our results provide novel insights into cellular distribution of clonal mutations, their dynamics under chemotherapy, and advocate for systematic analyses for CHIP in cancer patients.