Objectives: Chronic primary vasculitis describes a group of complex and rare diseases that are characterized by blood vessel inflammation. Classification of vasculitis subtypes is based predominantly on the size of the involved vessels and clinical phenotype. There is a recognized need to improve classification, especially for small-to-medium sized vessel vasculitides, that, ideally, is based on the underlying biology with a view to informing treatment. Methods: We performed RNA-Seq on blood samples from children (n = 41) and from adults (n = 11) with small-to-medium sized vessel vasculitis, and used unsupervised hierarchical clustering of gene expression patterns in combination with clinical metadata to define disease subtypes. Results: Differential gene expression at the time of diagnosis separated patients into two primary endotypes that differed in the expression of ~3,800 genes in children, and ~1,600 genes in adults. These endotypes were also present during disease flares, and both adult and pediatric endotypes could be discriminated based on the expression of just 20 differentially expressed genes. Endotypes were associated with distinct biological processes, namely neutrophil degranulation and T cell receptor signaling. Conclusions: Phenotypically similar subsets of small-to-medium sized vessel vasculitis may have different mechanistic drivers involving innate vs. adaptive immune processes. Discovery of these differentiating immune features provides a mechanistic-based alternative for subclassification of vasculitis.

Chronic granulomatous disease (CGD) is caused by mutations in genes that encode the NADPH-oxidase and result in a failure of phagocytic cells to produce reactive oxygen species (ROS) via this enzyme system. Patients with CGD are highly susceptible to infections and often suffer from inflammatory disorders; the latter occurs in the absence of infection and correlates with the spontaneous production of inflammatory cytokines. This clinical feature suggests that NADPH-oxidase-derived ROS are not required for, or may even suppress, inflammatory processes. Experimental evidence, however, implies that ROS are in fact required for inflammatory cytokine production. By using a myeloid cell line devoid of a functional NADPH-oxidase and primary CGD cells, we analyzed intracellular oxidants, signs of oxidative stress, and inflammatory cytokine production. Herein, we demonstrate that phagocytes lacking a functional NADPH-oxidase, namely primary CGD phagocytes and a gp91phox-deficient cell line, display elevated levels of ROS derived from mitochondria. Accordingly, these cells, despite lacking the major source of cellular ROS, display clear signs of oxidative stress, including an induced expression of antioxidants and altered oxidation of cell surface thiols. These observed changes in redox state were not due to abnormalities in mitochondrial mass or membrane integrity. Finally, we demonstrate that increased mitochondrial ROS enhanced phosphorylation of ERK1/2, and induced production of IL8, findings that correlate with previous observations of increased MAPK activation and inflammatory cytokine production in CGD cells. Our data show that elevated baseline levels of mitochondria-derived oxidants lead to the counter-intuitive observation that CGD phagocytes are under oxidative stress and have enhanced MAPK signaling, which may contribute to the elevated basal production of inflammatory cytokines and the sterile inflammatory manifestations in CGD.

Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a small vessel vasculitis in adults and children that commonly affects the kidneys. Although the frequent antigenic, and presumed pathogenic, targets of ANCA in AAV are proteinase-3 (PR3) and myeloperoxidase (MPO), ANCA against lysosome associated membrane protein-2 (LAMP-2), a lesser known ANCA antigen that is expressed on the glomerular endothelium, are present in some adults with AAV-associated renal disease. LAMP-2-ANCA has not been assessed in children with chronic systemic vasculitis, and, if present, would be a potentially valuable biomarker given that treatment decisions for these pediatric patients at diagnosis are largely informed by kidney function.

Background: The receptor for advanced glycation end products (RAGE) is a multiligand receptor involved in a number of processes and disorders. While it is known that RAGE-signaling can contribute to toxic liver damage and fibrosis, its role in acute inflammatory liver injury and septic multiorgan failure is yet undefined. We examined RAGE in lipopolysaccharide (LPS) induced acute liver injury of D-galN sensitized mice as a classical model for tumor necrosis factor alpha (TNF-α) dependent inflammatory organ damage. Methods: Mice (Rage-/- and C57BL/6) were intraperitoneally injected with D-galN (300 mg/kg) and LPS (10 μg/kg). Animals were monitored clinically, and cytokines, damage associated molecular pattern molecules (DAMPs) as well as liver enzymes were determined in serum. Liver histology, hepatic cytokines as well as RAGE mRNA expression were analyzed. Cellular activation and functionality were evaluated by flow cytometry both in bone marrow- and liver-derived cells. Results: Genetic deficiency of RAGE significantly reduced the mortality of mice exposed to LPS/D-galN. Hepatocyte damage markers were reduced in Rage-/- mice, and liver histopathology was less severe. Rage-/- mice produced less pro-inflammatory cytokines and DAMPs in serum and liver. While immune cell functions appeared normal, TNF-α production by hepatocytes was reduced in Rage-/- mice. Conclusions: We found that RAGE deletion attenuated the expression of pro-inflammatory cytokines and DAMPs in hepatocytes without affecting cellular immune functions in the LPS/D-galN model of murine liver injury. Our data highlight the importance of tissue-specific RAGE-signaling also in acute inflammatory liver stress contributing to sepsis and multiorgan failure.