Thrombomodulin (TM), which is predominantly expressed on the endothelium, plays an important role in maintaining vascular homeostasis by regulating the coagulation system. Intravascular injury and inflammation are complicated physiological processes that are induced by injured endothelium-mediated pro-coagulant signaling, necrotic endothelialand blood cell-derived damage-associated molecular patterns (DAMPs), and DAMPmediated inflammation. During the hypercoagulable state after endothelial injury, TM is released into the intravascular space by proteolytic cleavage of the endothelium component. Recombinant TM (rTM) is clinically applied to patients with disseminated intravascular coagulation, resulting in protection from tissue injury. Recent studies have revealed that rTM functions as an inflammatory regulator beyond hemostasis through various molecular mechanisms. More specifically, rTM neutralizes DAMPs, including histones and high mobility group box 1 (HMGB1), suppresses excessive activation of the complement system, physiologically protects the endothelium, and influences both innate and acquired immunity. Neutrophil extracellular traps (NETs) promote immunothrombosis by orchestrating platelets to enclose infectious invaders as part of the innate immune system, but excessive immunothrombosis can cause intravascular injury. However, rTM can directly and indirectly regulate NET formation. Furthermore, rTM interacts with mediators of acquired immunity to resolve vascular inflammation. So far, rTM has shown good efficacy in suppressing inflammation in various experimental models, including thrombotic microangiopathy, sterile inflammatory disorders, autoimmune diseases, and sepsis. Thus, rTM has the potential to become a novel tool to regulate intravascular injury via pleiotropic effects.

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Objective Antineutrophil cytoplasmic antibody (ANCA)–associated vasculitis (AAV) is pathologically characterized by focal fibrinoid necrosis, in which ANCA‐mediated neutrophil extracellular trap (NET) formation and subsequent endothelial cell necrosis occur. Cyclophilin D (CypD) plays an important role in mediation of cell necrosis and inflammation via the opening of mitochondrial permeability transition pores. This study was undertaken to examine the role of CypD in AAV pathogenesis. Methods We assessed the role and mechanism of CypD in ANCA‐stimulated neutrophils in vitro by immunostaining and electron microscopy observation. We performed a comprehensive RNA‐sequencing analysis on ANCA‐treated murine neutrophils. To investigate the role of CypD in vivo, we assessed disease features in CypD‐knockout mice and wild‐type mice using 2 different murine AAV models: anti‐myeloperoxidase IgG transfer–induced AAV and spontaneous AAV. Results In vitro experiments showed that pharmacologic and genetic inhibition of CypD suppressed ANCA‐induced NET formation via the suppression of reactive oxygen species and cytochrome c release from the mitochondria. RNA‐sequencing analyses in ANCA‐treated murine neutrophils revealed the involvement of inflammatory responses, with CypD deficiency reducing ANCA‐induced alterations in gene expression. Furthermore, analyses of upstream regulators revealed the relevance of intracellular calcium (CypD activator) and cyclosporin (CypD inhibitor) in ANCA stimulation, indicating that the CypD‐dependent opening of mitochondrial permeability transition pores is associated with ANCA‐induced neutrophil activation and NETosis. In both AAV mouse models, the genetic deletion of CypD ameliorated crescentic glomerulonephritis via the inhibition of CypD‐dependent neutrophil and endothelial necrosis. Conclusion CypD targeting is a novel and specific therapeutic strategy for AAV via the resolution of necrotizing vasculitis.

Neutrophil extracellular trap (NET) formation contributes to immune defense and is a distinct form of cell death. Excessive NET formation is found in patients with anti–neutrophil cytoplasmic antibody–associated (ANCA-associated) vasculitis (AAV), contributing to disease progression. The clearance of dead cells by macrophages, a process known as efferocytosis, is regulated by the CD47-mediated “don’t eat me” signal. Hence, we hypothesized that pathogenic NETs in AAV escape from efferocytosis via the CD47 signaling pathway, resulting in the development of necrotizing vasculitis. Immunostaining for CD47 in human renal tissues revealed high CD47 expression in crescentic glomerular lesions of patients with AAV. In ex vivo studies, ANCA-induced netting neutrophils increased the expression of CD47 with the reduction of efferocytosis. After efferocytosis, macrophages displayed proinflammatory phenotypes. The blockade of CD47 in spontaneous crescentic glomerulonephritis-forming/Kinjoh (SCG/Kj) mice ameliorated renal disease and reduced myeloperoxidase-ANCA (MPO-ANCA) titers with a reduction in NET formation. Thus, CD47 blockade would protect against developing glomerulonephritis in AAV via restored efferocytosis of ANCA-induced NETs.