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Shiga toxin producing Escherichia coli (STEC) are foodborne pathogens that release Shiga toxin (Stx), virulence factor responsible for the development of Hemolytic Uremic Syndrome (HUS). Stx causes endothelial cell damage, which leads to platelets deposition and thrombi formation within the microvasculature. It has been described that Stx activates blood cells and induces the shedding of proinflammatory and prothrombotic microvesicles (MVs) containing the toxin. In this sense, it has been postulated that MVs containing Stx2 (MVs-Stx2+) can contribute to the physiopathology of HUS, allowing Stx2 to reach the target organs while evading the immune system. In this work, we propose that circulating MVs-Stx2+ can be a potential biomarker for the diagnosis and prognosis of STEC infections and HUS progression. We developed a rat HUS model by the intraperitoneal injection of a sublethal dose of Stx2 and observed: decrease in body weight, increase of creatinine and urea levels, decrease of creatinine clearance and histological renal damages. After characterization of renal damages, we investigated circulating total MVs and MVs-Stx2+ by flow cytometry at different times after Stx2 injection. Additionally, we evaluated the correlation of biochemical parameters such as creatinine and urea in plasma with MVs-Stx2+. As a result, we found a significant circulation of MVs-Stx2+ at 72 and 96 h after Stx2 injection, nevertheless no correlation with creatinine and urea plasma levels were detected. Our results suggest that MVs-Stx2+ may be an additional biomarker for the characterization and diagnosis of HUS progression. A further analysis is required in order to validate MVs-Stx2+ as biomarker of the disease. [Display omitted] • A rat model of HUS was established with pure Stx2. • Total circulating microvescicles were increased in Stx2 treated rats. • Circulating MVs containing Stx2 (MVs-Stx2+) were detected in Stx2 treated rats by flow cytometry. • Biochemical parameters of disease do not correlate with MVs-Stx2+ amount. • Detection of circulating MVs-Stx2+ in patients may be useful for diagnosis and predictive for severity of disease. [ABSTRACT FROM AUTHOR]

Shiga toxin producing Escherichia coli (STEC) are bacterial pathogens involved in food-borne diseases. Shiga toxin (Stx) is the main virulence factor of STEC and is responsible for systemic complications including Hemolytic Uremic Syndrome (HUS). It has been previously demonstrated that Shiga toxin type 2 (Stx2) induces pregnancy loss in rats in early stage of pregnancy. The main purpose of this study was to determine if an active immunization prevents Stx2 mediated pregnancy loss and confers passive protective immunity to the offspring. For that purpose Sprague Dawley female rats were immunized with the chimera based on the enzyme lumazine synthase from Brucella spp . (BLS) and the B subunit of Shiga toxin 2 (Stx2B) named BLS-Stx2B. After immunization females were mated with males. At day 8 of gestation, dams were challenged intraperitoneally with a sublethal and abortifacient dose of Stx2. The immunization induced high anti-Stx2B-specific antibody titers in sera and most important, prevented pregnancy loss. Pups born and breastfeed by immunized dams had high anti-Stx2B-specific antibody titers in sera. Cross-fostering experiments indicated that passive protective immunity against Stx2 was transmitted through lactation. These results indicate that immunization of adult female rats with BLS-Stx2B prevents Stx2-induced pregnancy loss and confers anti Stx2 protective immunity to the offspring. [ABSTRACT FROM AUTHOR]

Typical hemolytic uremic syndrome (HUS) is responsible for acute and chronic renal failure in children younger than 5 years old in Argentina. Renal damages have been associated with Shiga toxin type 1 and/or 2 (Stx1, Stx2) produced by Escherichia coli O157:H7, although strains expressing Stx2 are highly prevalent in Argentina. Human glomerular endothelial cells (HGEC) and proximal tubule epithelial cells are very Stx-sensitive since they express high levels of Stx receptor (Gb3). Nowadays, there is no available therapy to protect patients from acute toxin-mediated cellular injury. New strategies have been developed based on the Gb3 biosynthesis inhibition through blocking the enzyme glucosylceramide (GL1) synthase. We assayed the action of a GL1 inhibitor (Miglustat: MG), on the prevention of the renal damage induced by Stx2. HGEC primary cultures and HK-2 cell line were pre-treated with MG and then incubated with Stx2. HK- 2 and HGEC express Gb3 and MG was able to decrease the levels of this receptor. As a consequence, both types of cells were protected from Stx2 cytotoxicity and morphology damage. MG was able to avoid Stx2 effects in human renal cells and could be a feasible strategy to protect kidney tissues from the cytotoxic effects of Stx2 in vivo . [ABSTRACT FROM AUTHOR]

Abstract: We have previously demonstrated that bacterial DNA induces neutrophil activation through a CpG- and TLR9-independent but MyD88-dependent-pathway. In this study we determined that GM-CSF enhances the activation of neutrophils by bacterial DNA. Granulocyte-macrophage colony-stimulating factor increased IL-8 and IL-1β secretion, and CD11b-upregulation induced by single-stranded bacterial DNA. It also enhanced neutrophil IL-8 production induced by double-stranded bacterial DNA, methylated single-stranded DNA, plasmid DNA, and phosphorothioated-CpG and non-CpG-oligodeoxynucleotides. Together these observations indicated that GM-CSF enhances neutrophil responses triggered by bacterial DNA in a CpG-independent fashion. We also found that GM-CSF enhanced the activation of the MAPKs p38 and ERK1/2 induced by bacterial DNA. Moreover, the pharmacological inhibition of these pathways significantly diminished GM-CSF ability to increase neutrophil activation by bacterial DNA. Finally, we observed that GM-CSF was unable to increase the activation of MyD88−/− neutrophils by bacterial DNA. Our findings suggest that GM-CSF modulates the CpG-independent, MyD88-dependent neutrophil response to bacterial DNA, by increasing the activation of the MAPKs p38 and ERK1/2. [Copyright &y& Elsevier]

Abstract: Dendritic cells (DCs) are the only antigen-presenting cell capable of activating naïve T lymphocytes, and hence they play a crucial role in the induction of adaptive immunity. Immature DCs sample and process antigens, and efficiently sense a large variety of signals from the surrounding environment. Upon activation, they become capable to activate naïve T cells and to direct the differentiation and polarization of effector T lymphocytes. It is becoming increasingly clear that different signals are able to determine distinct programs of DC differentiation and different forms of immunity and tolerance. In the past few years many advances have been made in addressing the action exerted by pathogen-associated molecular patterns (PAMPs), cytokines, chemokines, and other less characterized stress molecules on the activity of DCs. In this review we focus on the multiplicity of innate signals able to modulate the functional profile of DCs. [Copyright &y& Elsevier]