Effect of Saccharomyces cerevisiae var. Boulardii and b-galactomannan oligosaccharide on porcine intestinal epithelial and dendritic cells challenged in vitro with Escherichia coli F4 (K88)

Probiotic and prebiotics, often called “immune-enhancing” feed additives, are believed to deal with pathogens, preventing the need of an immune response and reducing tissue damage. In this study, we investigated if a recently developed b-galactomannan (bGM) had a similar protective role compared to Saccharomyces cerevisiae var. Boulardii (Scb), a proven probiotic, in the context of enterotoxigenic Escherichia coli (ETEC) infection. ETEC causes inflammation, diarrhea and intestinal damage in piglets, resulting in large economic loses worldwide. We observed that Scb and bGM products inhibited in vitro adhesion of ETEC on cell surface of porcine intestinal IPI-2I cells. Our data showed that Scb and bGM decreased the mRNA ETEC-induced gene expression of pro-inflammatory cytokines TNF-a, IL-6, GM-CSF and chemokines CCL2, CCL20 and CXCL8 on intestinal IPI-2I. Furthermore, we investigated the putative immunomodulatory role of Scb and bGM on porcine monocyte-derived dendritic cells (DCs) per se and under infection conditions. We observed a slight up-regulation of mRNA for TNF-a and CCR7 receptor after coincubation of DC with Scb and bGM. However, no differences were found in DC activation upon ETEC infection and Scb or bGM co-culture. Therefore, our results indicate that, similar to probiotic Scb, prebiotic bGM may protect intestinal epithelial cells against intestinal pathogens. Finally, although these products may modulate DC activation, their effect under ETEC challenge conditions remains to be elucidated.This work was supported by grants from the French National Institute for Agricultural Research (INRA, France) and grant AGL 2009-11936 of the Ministerio de Ciencia e Innovación (MICIIN, Spain). We gratefully acknowledge Dr I. Badiola (CReSA) for providing ETEC GN1034 strain and Dr J. Domínguez-Juncal (Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria, INIA, Madrid, Spain) for kindly providing antibodies for pig cell surface markers.info:eu-repo/semantics/publishedVersio

Sustainable approaches for drug repurposing in rare diseases: recommendations from the IRDiRC Task Force

Drug repurposing represents a real opportunity to address unmet needs and improve the lives of rare disease patients. It is often presented as a faster, safer and cheaper path for bringing drugs into new indications. However, several economic, regulatory and scientific barriers can impede the successful repurposing of drugs for rare diseases. The International Rare Diseases Research Consortium (IRDiRC) set up the Task Force on Sustainable Models in Drug Repurposing with the objective of identifying key factors for achieving sustainable repurposing approaches in rare diseases. In order to help inform expert opinion, the Task Force investigated six cases of medicinal products repurposed into new rare indications and four cases of ongoing development programs. A questionnaire addressing the major steps of the repurposing approach was developed by the Task Force and sent to contact points of the organizations. In addition, interviews were conducted with the relevant organization representatives to conduct a deeper dive into the sustainability of the repurposing approach for each of the selected cases. Based on the collective experience of the members of the Task Force and the output from the questionnaires/interviews, we have identified ten key factors that should be considered by those embarking on repurposing projects. These factors include the identification of unmet patient needs and partnership with patients, collection of evidence concerning disease prevalence, patient numbers, drug pharmacology and disease etiology, drug industrial property status, off-label or compounding use, data from past clinical studies and needs for extended non-clinical and clinical studies. The development of a collaborative funding framework and early discussion with regulators and payers are additional factors to implement early in the development of sustainable drug repurposing projects

Research on rare diseases:ten years of progress and challenges at IRDiRC

The International Rare Diseases Research Consortium (IRDiRC) is a global collaborative initiative launched in 2011, aimed at tackling rare diseases through research. Here, we summarize IRDiRC’s vision and goals and highlight achievements and prospects after its first decade.</p

Expression and immunogenicity of the mycobacterial Ag85B/ESAT-6 antigens produced in transgenic plants by elastin-like peptide fusion strategy.

International audienceThis study explored a novel system combining plant-based production and the elastin-like peptide (ELP) fusion strategy to produce vaccinal antigens against tuberculosis. Transgenic tobacco plants expressing the mycobacterial antigens Ag85B and ESAT-6 fused to ELP (TBAg-ELP) were generated. Purified TBAg-ELP was obtained by the highly efficient, cost-effective, inverse transition cycling (ICT) method and tested in mice. Furthermore, safety and immunogenicity of the crude tobacco leaf extracts were assessed in piglets. Antibodies recognizing mycobacterial antigens were produced in mice and piglets. A T-cell immune response able to recognize the native mycobacterial antigens was detected in mice. These findings showed that the native Ag85B and ESAT-6 mycobacterial B- and T-cell epitopes were conserved in the plant-expressed TBAg-ELP. This study presents the first results of an efficient plant-expression system, relying on the elastin-like peptide fusion strategy, to produce a safe and immunogenic mycobacterial Ag85B-ESAT-6 fusion protein as a potential vaccine candidate against tuberculosis

Analysis of immuno-modulatory and anti-infectious properties of three Saccharomyces cerevisiae yeast strains in pig

L’objectif de cette étude est d’évaluer si trois souches de levure Saccharomycescerevisiae (Lv01, Lv02, Lv03) exercent in vitro des propriétés immuno-modulatrices suite àl’exposition de cellules épithéliales intestinales porcines à Escherichia coli entérotoxinogène(ETEC). Ensuite, nous avons évalué si ces propriétés sont associées à une protection des porceletscontre une infection colibacillaire au post-sevrage et à une stimulation de l’immunité maternellepassive chez la truie.In vitro, nous avons montré que Lv01 exerce une activité anti-inflammatoire suite àl’exposition de cellules épithéliales intestinales porcines à ETEC. Cette activité réside dans 1) lasécrétion des facteurs solubles et est associée 2) à la diminution de la phosphorylation des protéinesERK1/2, p38 et 3) à l’agglutination des ETEC par Lv01. Cependant, Lv01 ne prévient pasl’altération de la barrière épithéliale induite par ETEC.In vivo, à la différence d’expérimentations antérieures, les levures Lv01 et Lv02 neprotègent pas les porcelets contre une colibacillose expérimentale au post-sevrage. Par ailleurs, lestrois souches de levure stimulent l’accroissement des IgG et IgA dans le colostrum et le lait destruies suggérant la transmission d’une meilleure protection immunitaire aux porcelets.The objective of this work is to investigate whether three Saccharomyces cerevisiaeyeast strains (Lv01, Lv02, Lv03) induce in vitro immuno-modulatory properties in pig intestinalepithelial cells exposed to enterotoxigenic Escherichia coli (ETEC). Thereafter, we assessedwhether these properties are associated to a piglet protection against post-weaning colibacillosisand to a stimulation of maternal passive immunity in sows.In vitro, we have shown that Lv01 induces an anti-inflammatory activity in pig intestinalepithelial cells exposed to ETEC. This activity is induced by 1) secreted soluble factors and isassociated with 2) a decrease of both ERK1/2 and p38 phosphorylation and 3) an agglutination ofETEC by Lv01. However, Lv01 does not prevent the alteration of epithelial barrier integrityinduced by ETEC.In vivo, we have shown that in contrast to previous data, Lv01 and Lv02 do not protectweaned piglets against an experimental ETEC infection. Elsewhere, the three yeast strainsstimulate the immunoglobulin levels (IgG, IgA) in colostrum and milk of sows suggesting thetransfer of a better immune protection to piglets

<em>Saccharomyces cerevisiae[/i modulates immune gene expression and inhibits ETEC-induced ERK1/2 and p38 phosphorylation in porcine intestinal epithelial cell lines

International audienc

Nod2 activates NF-kB in CD4+ T cells but its expression is dispensable for T cell-induced colitis.

Although the etiology of Crohn's disease (CD) remains elusive this disease is characterized by T cell activation that leads to chronic inflammation and mucosal damage. A potential role for maladaptation between the intestinal microbiota and the mucosal immune response is suggested by the fact that mutations in the pattern recognition receptor Nod2 are associated with higher risks for developing CD. Although Nod2 deletion in CD4(+) T cells has been shown to impair the induction of colitis in the murine T cell transfer model, the analysis of T cell intrinsic Nod2 function in T cell differentiation and T cell-mediated immunity is inconsistent between several studies. In addition, the role of T cell intrinsic Nod2 in regulatory T cell (Treg) development and function during colitis remain to be analyzed. In this study, we show that Nod2 expression is higher in activated/memory CD4(+) T cells and its expression was inducible after T cell receptor (TCR) ligation. Nod2 stimulation with muramyl dipeptide (MDP) led to a nuclear accumulation of c-Rel NF-kB subunit. Although functionally active in CD4(+) T cells, the deletion of Nod2 did not impair the induction and the prevention of colitis in the T cell transfer model. Moreover, Nod2 deletion did not affect the development of Foxp3(+) Treg cells in the spleen of recipient mice and Nod2 deficient CD4 T cells expressing the OVA specific transgenic TCR were able to differentiate in Foxp3(+) Treg cells after OVA feeding. In vitro, CD25(+) Nod2 deficient T cells suppressed T cell proliferation as well as wild type counter parts and T cell stimulation with MDP did not affect the proliferation and the cytokine secretion of T cells. In conclusion, our data indicate that Nod2 is functional in murine CD4(+) T cells but its expression is dispensable for the T cell regulation of colitis

Saccharomyces cerevisiae modulates immune gene expressions and inhibits ETEC-mediated ERK1/2 and p38 signaling pathways in intestinal epithelial cells

Background: Enterotoxigenic Escherichia coli (ETEC) infections result in large economic losses in the swine industry worldwide. ETEC infections cause pro-inflammatory responses in intestinal epithelial cells and subsequent diarrhea in pigs, leading to reduced growth rate and mortality. Administration of probiotics as feed additives displayed health benefits against intestinal infections. Saccharomyces cerevisiae (Sc) is non-commensal and non-pathogenic yeast used as probiotic in gastrointestinal diseases. However, the immuno-modulatory effects of Sc in differentiated porcine intestinal epithelial cells exposed to ETEC were not investigated. Methodology/Principal Findings: We reported that the yeast Sc (strain CNCM I-3856) modulates transcript and protein expressions involved in inflammation, recruitment and activation of immune cells in differentiated porcine intestinal epithelial IPEC-1 cells. We demonstrated that viable Sc inhibits the ETEC-induced expression of pro-inflammatory transcripts (IL-6, IL-8, CCL20, CXCL2, CXCL10) and proteins (IL-6, IL-8). This inhibition was associated to a decrease of ERK1/2 and p38 MAPK phosphorylation, an agglutination of ETEC by Sc and an increase of the anti-inflammatory PPAR-γ nuclear receptor mRNA level. In addition, Sc up-regulates the mRNA levels of both IL-12p35 and CCL25. However, measurement of transepithelial electrical resistance displayed that Sc failed to maintain the barrier integrity in monolayer exposed to ETEC suggesting that Sc does not inhibit ETEC enterotoxin activity. Conclusions: Sc (strain CNCM I-3856) displays multiple immuno-modulatory effects at the molecular level in IPEC-1 cells suggesting that Sc may influence intestinal inflammatory reaction

CCL28 involvement in mucosal tissues protection as a chemokine and as an antibacterial peptide.

International audienceCCL28 chemokine is expressed by epithelial cells of various mucosal tissues. This chemokine binds to CCR3 and CCR10 receptors and plays an essential role in the IgA antibody secreting cells (IgA-ASC) homing to mucosal surfaces and to lactating mammary gland as well. In addition, CCL28 has been shown to exert a potent antimicrobial activity against both Gram-negative and Gram-positive bacteria and fungi. Using the pig model, we investigated the expression of both CCR10 and CCR3 receptors in a large panel of mucosal tissues. RT-PCR analysis revealed the expression of CCR3 and CCR10 mRNA in salivary glands, nasal mucosae, Peyer's patches, small and large intestine, suggesting the presence of leucocytes expressing these receptors within these tissues. CCR10 mRNA was observed in sow mammary gland at late gestation with an increasing level during lactation. Recombinant porcine CCL28 protein was produced and mass spectrometry analysis revealed antimicrobial chemokines features such as a high pI value (10.2) and a C-terminal highly positively-charged region. Using a viable count assay, we showed that CCL28 displayed antimicrobial activity against enteric pathogens and was effective in killing Salmonella serotypes Dublin and Choleraesuis, enteroinvasive Escherichia coli K88 and non-pathogenic E. Coli K12. The potent antimicrobial function of CCL28 combined with its wide distribution in mucosal tissues and secretions suggest that this protein plays an important role in innate immune protection of the epithelial surfaces