bCoinfection with malaria and nontyphoidal Salmonella serotypes (NTS) can cause life-threatening bacteremia in humans. Coinfection with malaria is a recognized risk factor for invasive NTS, suggesting that malaria impairs intestinal barrier function. Here, we investigated mechanisms and strategies for prevention of coinfection pathology in a mouse model. Our findings reveal that malarial-parasite-infected mice, like humans, develop L-arginine deficiency, which is associated with intestinal mastocytosis, elevated levels of histamine, and enhanced intestinal permeability. Prevention or reversal of L-arginine deficiency blunts mastocytosis in ileal villi as well as bacterial translocation, measured as numbers of mesenteric lymph node CFU of noninvasive Escherichia coli Nissle and Salmonella enterica serotype Typhimurium, the latter of which is naturally invasive in mice. Dietary supplementation of malarial-parasite-infected mice with L-arginine or L-citrulline reduced levels of ileal transcripts encoding interleukin-4 (IL-4), a key mediator of intestinal mastocytosis and macromolecular permeability. Supplementation with L-citrulline also enhanced epithelial adherens and tight junctions in the ilea of coinfected mice. These data suggest that increasing Larginine bioavailability via oral supplementation can ameliorate malaria-induced intestinal pathology, providing a basis for testing nutritional interventions to reduce malaria-associated mortality in humans. Half of the global population is at risk for malaria, which results in nearly 1 million deaths annually, 86% of which are of children (1). The majority of cases are in sub-Saharan Africa, where there is a high prevalence of coinfection with nontyphoidal Salmonella serotypes (NTS) during the rainy season (2-5). While infections with NTS are normally self-limiting in immunocompetent children, coinfection with malaria can predispose to the development of deadly NTS bacteremia (6-9). During malaria infection, sequestration of parasitized red blood cells (RBCs) and capillary blockage are prominent in intestinal villi (10) and are associated with ischemia, malabsorption, and increased gastrointestinal (GI) permeability (11,12). These phenomena are not restricted to severe malaria; up to 50% of Nigerian children with uncomplicated malaria have GI disturbances (13). The mechanisms that underlie malaria-associated GI pathology and enhance the risk of bacteremia are incompletely understood (14), although recent data indicate that malaria-induced heme oxygenase-1 (HO-1) contributes to impaired resistance to NTS by reducing the production of reactive oxygen species (15). Beyond these findings, knowledge is limited and therapeutic options for coinfection are few in the face of high antibiotic resistance in areas of malaria endemicity (16). To support the development of novel therapeutic interventions for invasive bacterial disease, we developed a murine model of coinfection with Plasmodium yoelii and the NTS strain Salmonella enterica serotype Typhimurium ATCC 14028. In this...

Nimishakavi S, Besprozvannaya M, Raymond WW, Craik CS, Gruenert DC, Caughey GH. Activity and inhibition of prostasin and matriptase on apical and basolateral surfaces of human airway epithelial cells.

Tryptic serine proteases of bronchial epithelium regulate ion flux, barrier integrity, and allergic inflammation. Inhibition of some of these proteases is a strategy to improve mucociliary function in cystic fibrosis and asthmatic inflammation. Several inhibitors have been tested in pre-clinical animal models and humans. We hypothesized that these inhibitors inactivate a variety of airway protease targets, potentially with bystander effects. To establish relative potencies and modes of action, we compared inactivation of human prostasin, matriptase, airway trypsin-like protease (HAT), and β-tryptase by nafamostat, camostat, bis(5-amidino-2-benzimidazolyl)methane (BABIM), aprotinin, and benzamidine. Nafamostat achieved complete, nearly stoichiometric and very slowly reversible inhibition of matriptase and tryptase, but inhibited prostasin less potently and was weakest versus HAT. The IC50 of nafamostat’s leaving group, 6-amidino-2-naphthol, was >104-fold higher than that of nafamostat itself, consistent with suicide rather than product inhibition as mechanisms of prolonged inactivation. Stoichiometric release of 6-amidino-2-naphthol allowed highly sensitive fluorometric estimation of active-site concentration in preparations of matriptase and tryptase. Camostat inactivated all enzymes but was less potent overall and weakest towards matriptase, which, however was strongly inhibited by BABIM. Aprotinin exhibited nearly stoichiometric inhibition of prostasin and matriptase, but was much weaker towards HAT and was completely ineffective versus tryptase. Benzamidine was universally weak. Thus, each inhibitor profile was distinct. Nafamostat, camostat and aprotinin markedly reduced tryptic activity on the apical surface of cystic fibrosis airway epithelial monolayers, suggesting prostasin as the major source of such activity and supporting strategies targeting prostasin for inactivation.