Use of VacA as a Vaccine Antigen.

One of the major toxins secreted by H. pylori is the Vacuolating cytotoxin A (VacA) named after its ability to induce the formation of "vacuole"-like membrane vesicles in the cytoplasm of gastric cells. VacA has been associated with the disruption of mitochondrial functions, stimulation of apoptosis, blockade of T cell proliferation and promotion of regulatory T cells, thereby making it a promising vaccine target. Immunity to bacterial virulence factors is well known to protect humans against bacterial infections; hence, detoxified VacA has been evaluated as a vaccine antigen. Our short review summarizes the pre-clinical and clinical data that have been published on the use of VacA in the development of the H. pylori vaccine

Novel high-throughput fluorescence-based assay for the identification of nematocidal compounds that target the blood-feeding pathway

Hookworm infections cause a neglected tropical disease (NTD) affecting ~740 million people worldwide, principally those living in disadvantaged communities. Infections can cause high morbidity due to their impact on nutrient uptake and their need to feed on host blood, resulting in a loss of iron and protein, which can lead to severe anaemia and impaired cognitive development in children. Currently, only one drug, albendazole is efficient to treat hookworm infection and the scientific community fears the rise of resistant strains. As part of on-going efforts to control hookworm infections and its associated morbidities, new drugs are urgently needed. We focused on targeting the blood-feeding pathway, which is essential to the parasite survival and reproduction, using the laboratory hookworm model Nippostrongylus brasiliensis (a nematode of rodents with a similar life cycle to hookworms). We established an in vitro-drug screening assay based on a fluorescent-based measurement of parasite viability during blood-feeding to identify novel therapeutic targets. A first screen of a library of 2654 natural compounds identified four that caused decreased worm viability in a blood-feeding-dependent manner. This new screening assay has significant potential to accelerate the discovery of new drugs against hookworms

How to get rid of a pathobiontic bacterium from the stomach mucosa : role of inflammatory monocytes and IL-22 in the vaccine-induced reduction of helicobacter infection

Helicobacter pylori is a bacterium colonizing the human stomach. To prevent or cure this potentially detrimental infection, vaccination might be a suitable alternative to antibiotic therapies. Recently, a study has demonstrated that a vaccine efficiently prevented H pylori infection in human. However, the mechanisms leading to protection remain elusive. In mice, the vaccine-induced protective response relies on CD4+ T cells and especially on Thl7 response. Nevertheless, the factors mediating the reduction of H pylori infection are not fully characterized. Hence, the aim of my thesis was to characterize the factors associated with the Thl7 response. In the context of the vaccine-induced reduction of Helicobacter infection, I first focused on the role of inflammatory monocytes. I showed that CDllb+Ly6CLOW inflammatory monocytes accumulated in the stomach of vaccinated mice in association with the reduction of Helicobacter infection. Remarkably, the depletion of inflammatory monocytes delayed the vaccine-induced protective response. Concerning the role of these cells, I demonstrated that inflammatory monocytes extracted from the stomach of vaccinated mice produced iNOS and killed H pylori in vitro. In a next step, I evaluated the role of IL-22 during the vaccine-induced response. IL-22, which is linked to the Thl7 response, increases innate defense mechanisms of epithelial cells. I demonstrated that IL-22 produced by antigen- specific Thl7 was increased in the stomach of vaccinated mice during the protective response. Interestingly, neutralization of IL-22 was associated with an impaired vaccine-induced protective response. Then, I demonstrated that IL-22 induced antimicrobial peptides (AMPs) secretion by epithelial cells. These AMPs killed H pylori in vitro. In conclusion, I showed that both inflammatory monocytes and IL-22 participated to the vaccine induced reduction of Helicobacter infection. In addition, I demonstrated that the epithelium along with inflammation induced by Thl7 response is a critical factor mediating reduction of Helicobacter infection

Immune responses to Helicobacter pylori infection.

Helicobacter pylori (H. pylori) infection is one of the most common infections in human beings worldwide. H. pylori express lipopolysaccharides and flagellin that do not activate efficiently Toll-like receptors and express dedicated effectors, such as γ-glutamyl transpeptidase, vacuolating cytotoxin (vacA), arginase, that actively induce tolerogenic signals. In this perspective, H. pylori can be considered as a commensal bacteria belonging to the stomach microbiota. However, when present in the stomach, H. pylori reduce the overall diversity of the gastric microbiota and promote gastric inflammation by inducing Nod1-dependent pro-inflammatory program and by activating neutrophils through the production of a neutrophil activating protein. The maintenance of a chronic inflammation in the gastric mucosa and the direct action of virulence factors (vacA and cytotoxin-associated gene A) confer pro-carcinogenic activities to H. pylori. Hence, H. pylori cannot be considered as symbiotic bacteria but rather as part of the pathobiont. The development of a H. pylori vaccine will bring health benefits for individuals infected with antibiotic resistant H. pylori strains and population of underdeveloped countries

IL-22-induced antimicrobial peptides are key determinants of mucosal vaccine-induced protection against H. pylori in mice

International audienceDespite the recent description of the mucosal vaccine-induced reduction of Helicobacter pylori natural infection in a phase 3 clinical trial, the absence of immune correlates of protection slows the final development of the vaccine. In this study, we evaluated the role of interleukin (IL)-22 in mucosal vaccine-induced protection. Gastric IL-22 levels were increased in mice intranasally immunized with urease þ cholera toxin and challenged with H. felis, as compared with controls. Flow cytometry analysis showed that a peak of CD4 þ IL-22 þ IL-17 þ Tcells infiltrating the gastric mucosa occurred in immunized mice in contrast to control mice. The inhibition of the IL-22 biological activity prevented the vaccine-induced reduction of H. pylori infection. Remarkably, anti-microbial peptides (AMPs) extracted from the stomachs of vaccinated mice, but not from the stomachs of non-immunized or immunized mice, injected with anti-IL-22 antibodies efficiently killed H. pylori in vitro. Finally, H. pylori infection in vaccinated RegIIIb-deficient mice was not reduced as efficiently as in wild-type mice. These results demonstrate that IL-22 has a critical role in vaccine-induced protection, by promoting the expression of AMPs, such as RegIIIb, capable of killing Helicobacter. Therefore, it can be concluded that urease-specific memory Th17/Th22 cells could constitute immune correlates of vaccine protection in humans

Role of Inflammatory Monocytes in Vaccine-Induced Reduction of Helicobacter felis Infection.

Despite the proven ability of immunization to reduce Helicobacter infection in mouse models, the precise mechanism of protection has remained elusive. In this study, we evaluated the role of inflammatory monocytes in the vaccine-induced reduction of Helicobacter felis infection. We first showed by using flow cytometric analysis that Ly6C(low) major histocompatibility complex class II-positive chemokine receptor type 2 (CCR2)-positive CD64(+) inflammatory monocytes accumulate in the stomach mucosa during the vaccine-induced reduction of H. felis infection. To determine whether inflammatory monocytes played a role in the protection, these cells were depleted with anti-CCR2 depleting antibodies. Indeed, depletion of inflammatory monocytes was associated with an impaired vaccine-induced reduction of H. felis infection on day 5 postinfection. To determine whether inflammatory monocytes had a direct or indirect role, we studied their antimicrobial activities. We observed that inflammatory monocytes produced tumor necrosis factor alpha and inducible nitric oxide synthase (iNOS), two major antimicrobial factors. Lastly, by using a Helicobacter in vitro killing assay, we showed that mouse inflammatory monocytes and activated human monocytes killed H. pylori in an iNOS-dependent manner. Collectively, these data show that inflammatory monocytes play a direct role in the immunization-induced reduction of H. felis infection from the gastric mucosa

Macrophages promote epithelial repair through hepatocyte growth factor secretion.

Macrophages play a critical role in intestinal wound repair. However, the mechanisms of macrophage-assisted wound repair remain poorly understood. We aimed to characterize more clearly the repair activities of murine and human macrophages. Murine macrophages were differentiated from bone marrow cells and human macrophages from monocytes isolated from peripheral blood mononuclear cells of healthy donors (HD) or Crohn's disease (CD) patients or isolated from the intestinal mucosa of HD. In-vitro models were used to study the repair activities of macrophages. We found that murine and human macrophages were both able to promote epithelial repair in vitro. This function was mainly cell contact-independent and relied upon the production of soluble factors such as the hepatocyte growth factor (HGF). Indeed, HGF-silenced macrophages were less capable of promoting epithelial repair than control macrophages. Remarkably, macrophages from CD patients produced less HGF than their HD counterparts (HGF level: 84âeuro0/00±âeuro0/0027âeuro0/00pg/mg of protein and 45âeuro0/00±âeuro0/0034âeuro0/00pg/mg of protein, respectively, for HD and CD macrophages, Pâeuro0/00<âeuro0/000·009) and were deficient in promoting epithelial repair (repairing activity: 90·1âeuro0/00±âeuro0/004·6 and 75·8âeuro0/00±âeuro0/008·3, respectively, for HD and CD macrophages, Pâeuro0/00<âeuro0/000·0005). In conclusion, we provide evidence that macrophages act on wounded epithelial cells to promote epithelial repair through the secretion of HGF. The deficiency of CD macrophages to secrete HGF and to promote epithelial repair might contribute to the impaired intestinal mucosal healing in CD patients

IL-22-induced antimicrobial peptides are key determinants of mucosal vaccine-induced protection against H. pylori in mice.

Despite the recent description of the mucosal vaccine-induced reduction of Helicobacter pylori natural infection in a phase 3 clinical trial, the absence of immune correlates of protection slows the final development of the vaccine. In this study, we evaluated the role of interleukin (IL)-22 in mucosal vaccine-induced protection. Gastric IL-22 levels were increased in mice intranasally immunized with urease+cholera toxin and challenged with H. felis, as compared with controls. Flow cytometry analysis showed that a peak of CD4(+)IL-22(+)IL-17(+) T cells infiltrating the gastric mucosa occurred in immunized mice in contrast to control mice. The inhibition of the IL-22 biological activity prevented the vaccine-induced reduction of H. pylori infection. Remarkably, anti-microbial peptides (AMPs) extracted from the stomachs of vaccinated mice, but not from the stomachs of non-immunized or immunized mice, injected with anti-IL-22 antibodies efficiently killed H. pylori in vitro. Finally, H. pylori infection in vaccinated RegIIIβ-deficient mice was not reduced as efficiently as in wild-type mice. These results demonstrate that IL-22 has a critical role in vaccine-induced protection, by promoting the expression of AMPs, such as RegIIIβ, capable of killing Helicobacter. Therefore, it can be concluded that urease-specific memory Th17/Th22 cells could constitute immune correlates of vaccine protection in humans

La Charité-sur-Loire (Nièvre): évolution du chenal, des ponts et des autres structures (pêcheries, moulins)

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