Profile of blood cells and inflammatory mediators in periodic fever, aphthous stomatitis, pharyngitis and adenitis (PFAPA) syndrome

<p>Abstract</p> <p>Background</p> <p>This study aimed to profile levels of blood cells and serum cytokines during afebrile and febrile phases of periodic fever, aphthous <b>s</b>tomatitis, pharyngitis and adenitis (PFAPA) syndrome to advance pathophysiological understanding of this pediatric disease.</p> <p>Methods</p> <p>A cohort of patients with a median age of 4.9 years experiencing 'typical PFAPA' episodes participated in this study. Blood cells and serum cytokines were analyzed by CBC analysis and multiplex ELISA.</p> <p>Results</p> <p>Oscillations in the concentration of blood cells during the afebrile and febrile phases of typical PFAPA syndrome were observed; novel findings include increased monocytes and decreased eosinophils during a febrile episode and increased thrombocytes in the afebrile interval. Relatively modest levels of pro-inflammatory cytokines were present in sera. IFNγ-induced cytokine IP10/CXCL10 was increased after the onset of fever while T cell-associated cytokines IL7 and IL17 were suppressed during afebrile and febrile periods.</p> <p>Conclusions</p> <p>Identification of dysregulated blood cells and serum cytokines is an initial step towards the identification of biomarkers of PFAPA disease and/or players in disease pathogenesis. Future investigations are required to conclusively discern which mediators are associated specifically with PFAPA syndrome.</p

Galectin-3 type-C self-association on neutrophil surfaces; The carbohydrate recognition domain regulates cell function

Galectin-3 is an endogenous β-galactoside-binding lectin comprising a carbohydrate recognition domain (CRD) linked to a collagen-like N-domain. Both domains are required for galectin-3 to induce cellular effects; a C-terminal fragment of galectin-3, galectin-3C, containing the CRD but lacking the N-domain, binds cell surface glycoconjugates but does not induce cellular effects since cross-linking promoted by the N-domain is thought to be required. Instead, galectin-3C is proposed to antagonize the effects of galectin-3 by competing for binding sites. The aim of this study was to investigate the effects of galectin-3C on galectin-3 interactions with human neutrophils. Recombinant galectin-3C inhibited galectin-3-induced production of reactive oxygen species in primed neutrophils. Surprisingly, this inhibition was not due to competitive inhibition of galectin-3 binding to the cells. In contrast, galectin-3C potentiated galectin-3 binding, in line with emerging evidence that galectin-3 can aggregate not only through the N-domain but also through the CRD. The cell surface interaction between galectin-3C and galectin-3 was corroborated by colocalization of fluorescently labeled galectin-3 and galectin-3C. Galectin-3C can be generated in vivo through cleavage of galectin-3 by proteases. Indeed, in circulation, galectin-3 and galectin-3C were both attached to the cell surface of neutrophils, which displayed great capacity to bind additional galectin-3 and galectin-3C. In conclusion, galectin-3C enhances galectin-3 binding to neutrophils by nonactivating type-C self-association, in parallel to inhibiting neutrophil activation by galectin-3 (induced by type-N self-association). This implicates type-C self-association as a termination system for galectin-3-induced cell activation, with the purpose of avoiding oxidant-dependent tissue damage

Dietary Polyunsaturated Fatty Acids Promote Neutrophil Accumulation in the Spleen by Altering Chemotaxis and Delaying Cell Death

Neutrophils are the most abundant circulating leukocytes in humans and are essential for the defense against invading pathogens. Like many other cells of an organism, neutrophils can be highly influenced by the diet. We have previously described that mice fed a high-fat diet rich in polyunsaturated fatty acids (HFD-P) present a higher frequency of neutrophils in bone marrow than mice fed a high-fat diet rich in saturated fatty acids (HFD-S). Interestingly, such an increase correlated with improved survival against bacterium-induced sepsis. In this study, we aimed to investigate the effects of dietary polyunsaturated and saturated fatty acids on neutrophil homeostasis. We found that HFD-P specifically induced the accumulation of neutrophils in the marginal pools of the spleen and liver. The accumulation of neutrophils in the spleen was a result of a dual effect of polyunsaturated fatty acids on neutrophil homeostasis. First, polyunsaturated fatty acids enhanced the recruitment of neutrophils from the circulation into the spleen via chemokine secretion. Second, they delayed neutrophil cell death in the spleen. Interestingly, these effects were not observed in mice fed a diet rich in saturated fatty acids, suggesting that the type of fat rather than the amount of fat mediates the alterations in neutrophil homeostasis. In conclusion, our results show that dietary polyunsaturated fatty acids have a strong modulatory effect on neutrophil homeostasis that may have future clinical applications

Elevated Mitochondrial Reactive Oxygen Species and Cellular Redox Imbalance in Human NADPH-Oxidase-Deficient Phagocytes

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

Mutational Tuning of Galectin-3 Specificity and Biological Function*

Galectins are defined by a conserved β-galactoside binding site that has been linked to many of their important functions in e.g. cell adhesion, signaling, and intracellular trafficking. Weak adjacent sites may enhance or decrease affinity for natural β-galactoside-containing glycoconjugates, but little is known about the biological role of this modulation of affinity (fine specificity). We have now produced 10 mutants of human galectin-3, with changes in these adjacent sites that have altered carbohydrate-binding fine specificity but that retain the basic β-galactoside binding activity as shown by glycan-array binding and a solution-based fluorescence anisotropy assay. Each mutant was also tested in two biological assays to provide a correlation between fine specificity and function. Galectin-3 R186S, which has selectively lost affinity for LacNAc, a disaccharide moiety commonly found on glycoprotein glycans, has lost the ability to activate neutrophil leukocytes and intracellular targeting into vesicles. K176L has increased affinity for β-galactosides substituted with GlcNAcβ1–3, as found in poly-N-acetyllactosaminoglycans, and increased potency to activate neutrophil leukocytes even though it has lost other aspects of galectin-3 fine specificity. G182A has altered carbohydrate-binding fine specificity and altered intracellular targeting into vesicles, a possible link to the intracellular galectin-3-mediated anti-apoptotic effect known to be lost by this mutant. Finally, the mutants have helped to define the differences in fine specificity shown by Xenopus, mouse, and human galectin-3 and, as such, the evidence for adaptive change during evolution