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Human CD81 and CD9 are members of the tetraspanin family of proteins characterized by a canonical structure of four transmembrane domains and two extracellular loop domains. Tetraspanins are known as molecular facilitators, which assemble and organize cell surface receptors and partner molecules forming clusters known as tetraspanin-enriched microdomains. They have been implicated to play various biological roles including an involvement in infections with microbial pathogens. Here, we demonstrate an important role of CD81 for the invasion of epithelial cells by Salmonella enterica. We show that overexpression of CD81 in HepG2 cells enhances invasion of various typhoidal and non-typhoidal Salmonella serovars. Deletion of CD81 by CRISPR/Cas9 in intestinal epithelial cells (C2BBe1 and HT29-MTX-E12) reduces S. Typhimurium invasion. In addition, the effect of human CD81 is species-specific as only human but not rat CD81 facilitates Salmonella invasion. Finally, immunofluorescence microscopy and proximity ligation assay revealed that both human tetraspanins CD81 and CD9 are recruited to the entry site of S. Typhimurium during invasion but not during adhesion to the host cell surface. Overall, we demonstrate that the human tetraspanin CD81 facilitates Salmonella invasion into epithelial host cells.
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Monosodium glutamate (MSG) is commonly used as a flavor-enhancing agent in foods, and studies have demonstrated its toxic effects in animal models. Black garlic is known for its antioxidant and anti-inflammatory properties; however, there is a lack of studies on the potential hepatoprotective effect of black garlic ethanol extract (BGE) against MSG-induced hepatotoxicity in rats. The aim of this study was to investigate the hepatoprotective effects of ethanol extract of black garlic against MSG-induced liver damage in animal model. Twenty-five male Wistar rats were randomly assigned to five groups (n=5): negative control, MSG only, and MSG with three different doses of BGE. The MSG only and MSG with BGE groups were orally administered with 8 mg/kg MSG daily. After MSG treatment, the MSG with BGE groups received BGE orally at daily doses of 200, 400, or 600 mg/kg body weight for 16 consecutive days. Subsequently, the levels of serum liver enzymes aspartate aminotransferase (AST), alanine aminotransferase (ALT), interferon-gamma (IFN-γ), and cyclooxygenase-2 (COX-2) were measured. Our data indicated that the group treated with 200 mg/kg BGE had significant lower levels of AST and ALT significantly compared to the MSG-only group. The MSG-treated group had higher levels of the inflammatory markers COX-2 and IFN-γ, which were lowered by administration of 200 mg/kg BGE. In contrast, higher doses of BGE led to greater levels of COX-2 and IFN-γ compared to those in the MSG-only group. This study suggested that BGE might have hepatoprotective effects at low dose, potentially mitigating MSG-induced liver damage. However, the higher dose of black garlic extract did not alleviate inflammation, as shown by the higher levels of COX-2 and IFN-γ.
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Enfermedad Hepática Inducida por Sustancias y Drogas , Ajo , Extractos Vegetales , Ratas Wistar , Glutamato de Sodio , Animales , Ajo/química , Extractos Vegetales/farmacología , Extractos Vegetales/uso terapéutico , Ratas , Enfermedad Hepática Inducida por Sustancias y Drogas/prevención & control , Enfermedad Hepática Inducida por Sustancias y Drogas/tratamiento farmacológico , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Masculino , Modelos Animales de Enfermedad , Alanina Transaminasa/sangre , Alanina Transaminasa/metabolismo , Aspartato Aminotransferasas/sangre , Aspartato Aminotransferasas/metabolismo , Hígado/efectos de los fármacos , Hígado/patología , Hígado/metabolismo , Interferón gamma/metabolismo , Ciclooxigenasa 2/metabolismoRESUMEN
Fasting has been practiced with different time span in different areas of the world and for various reasons. One of the types of fasting regimens is Ramadan intermittent fasting (RIF), which is described as intermittent dry fasting and known as the most commonly practiced form of religious fasting. Different studies have shown its effects on body composition parameters and mental health, fatigue and quality of life (QoL). Elucidating the relationship of RIF on biological parameters would also be of importance to show its mechanism. Therefore, we evaluated several biological mediators related to mental health, such as ß-nerve growth factor (ß-NGF), brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and insulin-like growth factor-1 (IGF-1), interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α), and matrix-metalloproteinase-9 (MMP-9). This study consisted of fasting (FG; n = 25) and non-fasting group (NFG; n = 25). Four different time points were assessed for FG: one week before (T1), mid (T2), last days (T3), and one week after (T4) RIF. T1 and T3 were the assessment time points for NFG. Biological mediators were determined from serum samples by using Human Magnetic Luminex and enzyme-linked immunosorbent assay. Furthermore, we then performed correlation analyses between biological mediators and our previously published clinical parameters including body composition and mental health parameters at all time points. Significant alterations were shown in FG for ß-NGF (T2vsT3, p < 0.05; T2vsT4, p < 0.05), GDNF (T1vsT4, p < 0.05; T2vsT4, p < 0.05), IL-8 (T2vsT3, p < 0.05; T3vsT4, p < 0.05), TNF-α (T1vsT3, p < 0.05; T1vsT4, p < 0.001; T2vsT4, p < 0.001), and MMP-9 (T1vsT4, p < 0.01). There were no statistically significant differences between FG and NFG in all biological mediators at T1 and T3. Correlation analysis showed that MMP-9 levels had negative correlation with body mass index (BMI) at T3. At T3 BDNF levels had negative correlation with Epworth Sleepiness Scale (ESS) as one of measured QoL parameters. ß-NGF, GDNF, TNF-α, and MMP-9 had positive correlation with some of body composition and mental health parameters. Findings demonstrate that RIF altered different biological mediators could give benefit to health. Its benefit is mediated by the alteration of biological mediators.
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Autophagy plays an important role in recognizing and protecting cells from invading intracellular pathogens such as Salmonella. In this work, we investigated the role of p38MAPK/MK2 in modulating the host cell susceptibility to Salmonella infection. Inhibition of p38MAPK or MK2 led to a significant increase of bacterial counts in Salmonella infected mouse embryonic fibroblasts (MEFs), as well as in MK2-deficient (Mk2-/-) cells. Furthermore, western blot analysis showed that Mk2-/- cells have lower level of LC3 lipidation, which is the indicator of general autophagy compared to Mk2-rescued cells. In Mk2-/- cells, we also observed lower activated TANK-binding kinase-1 phosphorylation on Ser172 and p62/SQTM1-Ser403 phosphorylation, which are important to promote the translocation of p62 to ubiquitinated microbes and required for efficient autophagy of bacteria. Furthermore, immunofluorescence analysis revealed reduced colocalization of Salmonella with LC3 and p62 in MEFs. Inhibition of autophagy with bafilomycin A1 showed increased bacterial counts in treated cells compared to control cell. Overall, these results indicate that p38MAPK/MK2-mediated protein phosphorylation modulates the host cell susceptibility to Salmonella infection by affecting the autophagy pathways.
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Infecciones por Salmonella , Proteínas Quinasas p38 Activadas por Mitógenos , Animales , Ratones , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Fibroblastos/metabolismo , AutofagiaRESUMEN
Infectious disease is widely considered to be a major driver of evolution. A preponderance of signatures of balancing selection at blood group-related genes is thought to be driven by inherent trade-offs in susceptibility to disease. B4galnt2 is subject to long-term balancing selection in house mice, where two divergent allele classes direct alternative tissue-specific expression of a glycosyltransferase in the intestine versus blood vessels. The blood vessel allele class leads to prolonged bleeding times similar to von Willebrand disease in humans, yet has been maintained for millions of years. Based on in vivo functional studies in inbred lab strains, it is hypothesized that the cost of prolonged bleeding times may be offset by an evolutionary trade-off involving susceptibility to a yet unknown pathogen(s). To identify candidate pathogens for which resistance could be mediated by B4galnt2 genotype, we here employed a novel "pathometagenomic" approach in a wild mouse population, which combines bacterial 16S rRNA gene-based community profiling with histopathology of gut tissue. Through subsequent isolation, genome sequencing and controlled experiments in lab mice, we show that the presence of the blood vessel allele is associated with resistance to a newly identified subspecies of Morganella morganii, a clinically important opportunistic pathogen. Given the increasing importance of zoonotic events, the approach outlined here may find useful application in the detection of emerging diseases in wild animal populations.
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Antígenos de Grupos Sanguíneos , Microbioma Gastrointestinal , Humanos , Ratones , Animales , Morganella , ARN Ribosómico 16S , GenotipoRESUMEN
Histo-blood group antigens in the intestinal mucosa play important roles in host-microbe interactions and modulate the susceptibility to enteric pathogens. The B4galnt2 gene, expressed in the GI tract of most mammals, including humans, encodes a beta-1,4-N-acetylgalactosaminyltransferase enzyme which catalyzes the last step in the biosynthesis of the Sd(a) and Cad blood group antigens by adding an N-acetylgalactosamine (GalNAc) residue to the precursor molecules. In our study, we found that loss of B4galnt2 expression is associated with increased susceptibility to Citrobacter rodentium infection, a murine model pathogen for human enteropathogenic Escherichia coli. We observed increased histopathological changes upon C. rodentium infection in mice lacking B4galnt2 compared to B4galnt2-expressing wild-type mice. In addition, wild-type mice cleared the C. rodentium infection faster than B4galnt2-/- knockout mice. It is known that C. rodentium uses its type 1 fimbriae adhesive subunit to bind specifically to D-mannose residues on mucosal cells. Flow cytometry analysis of intestinal epithelial cells showed the absence of GalNAc-modified glycans but an increase in mannosylated glycans in B4galnt2-deficient mice compared to B4galnt2-sufficient mice. Adhesion assays using intestinal epithelial organoid-derived monolayers revealed higher C. rodentium adherence to cells lacking B4galnt2 expression compared to wild-type cells which in turn was reduced in the absence of type I fimbriae. In summary, we show that B4galnt2 expression modulates the susceptibility to C. rodentium infection, which is partly mediated by fimbriae-mannose interaction.
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Ramadan fasting (RF) is a type of diurnal intermittent fasting. Previous studies reported the benefits of RF in healthy subjects on mood and health related to quality of life (QoL). Cortisol and brain-derived neurotrophic factor (BDNF) have been shown to play a role in mood, body composition parameters, and health-related QoL. This study aimed at elucidating the mechanism of the benefit of RF, particularly cortisol and BNDF and their association with mood and QoL. Insulin growth factor-1 (IGF-1), interleukin (IL)-8, matrix metalloproteinase (MMP)-9, and myoglobin were determined. Thirty-four healthy men and women were recruited. Serum from peripheral venous blood samples was collected at five time points: 1 week before RF (T1); mid of RF (T2), last days of RF (T3), 1 week after RF (T4), and 1 month after RF (T5). The amounts of biological mediators in the serum samples were determined by enzyme-linked immunosorbent assay (ELISA) and Luminex assays. BDNF and cortisol significantly decreased at T3 (p < 0.05) and T4 (p < 0.001) compared to T1, respectively. It seems the benefits of RF for mood-related symptoms are mediated by different biological mediators, particularly cortisol and BDNF.
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The glycosylation profile of the gastrointestinal tract is an important factor mediating host-microbe interactions. Variation in these glycan structures is often mediated by blood group-related glycosyltransferases, and can lead to wide-ranging differences in susceptibility to both infectious- as well as chronic disease. In this review, we focus on the interplay between host glycosylation, the intestinal microbiota and susceptibility to gastrointestinal pathogens based on studies of two exemplary blood group-related glycosyltransferases that are conserved between mice and humans, namely FUT2 and B4GALNT2. We highlight that differences in susceptibility can arise due to both changes in direct interactions, such as bacterial adhesion, as well as indirect effects mediated by the intestinal microbiota. Although a large body of experimental work exists for direct interactions between host and pathogen, determining the more complex and variable mechanisms underlying three-way interactions involving the intestinal microbiota will be the subject of much-needed future research.
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Antígenos de Grupos Sanguíneos , Enfermedades Transmisibles , Fucosiltransferasas , Microbioma Gastrointestinal , N-Acetilgalactosaminiltransferasas , Animales , Fucosiltransferasas/genética , Tracto Gastrointestinal , Humanos , Ratones , N-Acetilgalactosaminiltransferasas/genética , Galactósido 2-alfa-L-FucosiltransferasaRESUMEN
By crossing septin7-floxed mice with Lyz2-Cre mice carrying the Cre recombinase inserted in the Lysozyme-M (Lyz2) gene locus we aimed the specific deletion of septin7 in myeloid cells, such as monocytes, macrophages and granulocytes. Septin7 flox/flox :Lyz2-Cre mice show no alterations in the myeloid compartment. Septin7-deleted macrophages (BMDMs) were isolated and analyzed. The lack of Septin7 expression was confirmed and a constitutive double-nucleation was detected in Septin7-deficient BMDMs indicating a defect in macrophage cytokinesis. However, phagocytic function of macrophages as judged by uptake of labelled E. coli particles and LPS-stimulated macrophage activation as judged by induction of TNF mRNA expression and TNF secretion were not compromised. In addition to myeloid cells, Lyz2-Cre is also active in type II pneumocytes (AT2 cells). We monitored lung adenocarcinoma formation in these mice by crossing them with the conditional knock-in Kras-LSL-G12D allele. Interestingly, we found that control mice without septin7 depletion die after 3-5 weeks, while the Septin7-deficient animals survived 11 weeks or even longer. Control mice sacrificed in the age of 4 weeks display a bronchiolo-alveolar hyperplasia with multiple adenomas, whereas the Septin7-deficient animals of the same age are normal or show only a weak multifocal brochiolo-alveolar hyperplasia. Our findings indicate an essential role of Septin7 in macrophage cytokinesis but not in macrophage function. Furthermore, septin7 seems absolutely essential for oncogenic Kras-driven lung tumorigenesis making it a potential target for anti-tumor interventions.
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Proteoglycans (PGs) are glycoconjugates which are predominately expressed on cell surfaces and consist of glycosaminoglycans (GAGs) linked to a core protein. An initial step of GAGs assembly is governed by the ß-D-xylosyltransferase enzymes encoded in mammals by the XylT1/XylT2 genes. PGs are essential for the interaction of a cell with other cells as well as with the extracellular matrix. A number of studies highlighted a role of PGs in bacterial adhesion, invasion, and immune response. In this work, we investigated a role of PGs in Salmonella enterica serovar Typhimurium (S. Typhimurium) infection of epithelial cells. Gentamicin protection and chloroquine resistance assays were applied to assess invasion and replication of S. Typhimurium in wild-type and xylosyltransferase-deficient (ΔXylT2) Chinese hamster ovary (CHO) cells lacking PGs. We found that S. Typhimurium adheres to and invades CHO WT and CHO ΔXylT2 cells at comparable levels. However, 24 h after infection, proteoglycan-deficient CHO ΔXylT2 cells are significantly less colonized by S. Typhimurium compared to CHO WT cells. This proteoglycan-dependent phenotype could be rescued by addition of PGs to the cell culture medium, as well as by complementation of the XylT2 gene. Chloroquine resistance assay and immunostaining revealed that in the absence of PGs, significantly less bacteria are associated with Salmonella-containing vacuoles (SCVs) due to a re-distribution of endocytosed gentamicin. Inhibition of endo-lysosomal fusion by a specific inhibitor of phosphatidylinositol phosphate kinase PIKfyve significantly increased S. Typhimurium burden in CHO ΔXylT2 cells demonstrating an important role of PGs for PIKfyve dependent vesicle fusion which is modulated by Salmonella to establish infection. Overall, our results demonstrate that PGs influence survival of intracellular Salmonella in epithelial cells via modulation of PIKfyve-dependent endo-lysosomal fusion.
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Lisosomas/fisiología , Proteoglicanos/metabolismo , Salmonella typhimurium/efectos de los fármacos , Salmonella typhimurium/patogenicidad , Animales , Células CHO , Membrana Celular , Cloroquina/farmacología , Cricetulus , Endocitosis/efectos de los fármacos , Endocitosis/fisiología , Células Epiteliales , Gentamicinas/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Proteoglicanos/deficiencia , Salmonella typhimurium/crecimiento & desarrollo , SobrevidaRESUMEN
Mycobacterium avium subspecies paratuberculosis (MAP) causes Johne's disease (JD), an incurable chronic intestinal bowel disease in ruminants. JD occurs worldwide and causes enormous economic burden in dairy industry. Research on JD pathobiology is hampered by its complexity which cannot completely be mimicked by small animal models. As a model the mouse allows dissecting some pathogenicity features of MAP. However, for unknown reasons MAP exhibits reduced growth in granulomas of infected mice compared to other Mycobacterium avium subspecies. Here, we characterized immune reactions of MAP-infected C57BL/6 mice. After infection, mice appeared fully immunocompetent. A strong antigen-specific T cell response was elicited indicated by IFNγ production of splenic T cells re-stimulated with MAP antigens. Function of splenic dendritic cells and proliferation of adoptively transferred antigen-specific CD4+ T cells was unaltered. Isolated splenic myeloid cells from infected mice revealed that MAP resides in CD11b+ macrophages. Importantly, sorted CD11b+CD11c- cells expressed high level of type 2 nitric oxide synthase (NOS2) but only low levels of pro- and anti-inflammatory cytokines. Correspondingly, MAP-infected MAC2 expressing myeloid cells in spleen and liver granuloma displayed strong expression of NOS2. In livers of infected Nos2-/-mice higher bacterial loads, more granuloma and larger areas of tissue damage were observed 5 weeks post infection compared to wild type mice. In vitro, MAP was sensitive to NO released by a NO-donor. Thus, a strong T cell response and concomitant NOS2/NO activity appears to control MAP infection, but allows development of chronicity and pathogen persistence. A similar mechanism might explain persistence of MAP in ruminants.
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Citocinas/inmunología , Óxido Nítrico Sintasa de Tipo II/inmunología , Paratuberculosis/inmunología , Animales , Femenino , Inmunidad Celular , Hígado/microbiología , Hígado/patología , Macrófagos/inmunología , Macrófagos/microbiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mycobacterium avium subsp. paratuberculosis , Óxido Nítrico Sintasa de Tipo II/genética , Bazo/microbiología , Bazo/patología , Linfocitos T/inmunologíaRESUMEN
Expression of ABO and Lewis histo-blood group antigens by the gastrointestinal epithelium is governed by an α-1,2-fucosyltransferase enzyme encoded by the Fut2 gene. Alterations in mucin glycosylation have been associated with susceptibility to various bacterial and viral infections. Salmonella enterica serovar Typhimurium is a food-borne pathogen and a major cause of gastroenteritis. In order to determine the role of Fut2-dependent glycans in Salmonella-triggered intestinal inflammation, Fut2+/+ and Fut2-/- mice were orally infected with S. Typhimurium and bacterial colonization and intestinal inflammation were analyzed. Bacterial load in the intestine of Fut2-/- mice was significantly lower compared to Fut2+/+ mice. Analysis of histopathological changes revealed significantly lower levels of intestinal inflammation in Fut2-/- mice compared to Fut2+/+ mice and measurement of lipocalin-2 level in feces corroborated histopathological findings. Salmonella express fimbriae that assist in adherence of bacteria to host cells thereby facilitating their invasion. The std fimbrial operon of S. Typhimurium encodes the π-class Std fimbriae which bind terminal α(1,2)-fucose residues. An isogenic mutant of S. Typhimurium lacking Std fimbriae colonized Fut2+/+ and Fut2-/- mice to similar levels and resulted in similar intestinal inflammation. In vitro adhesion assays revealed that bacteria possessing Std fimbriae adhered significantly more to fucosylated cell lines or primary epithelial cells in comparison to cells lacking α(1,2)-fucose. Overall, these results indicate that Salmonella-triggered intestinal inflammation and colonization are dependent on Std-fucose interaction.
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Fimbrias Bacterianas/metabolismo , Fucosa/metabolismo , Salmonella typhimurium/patogenicidad , Animales , Adhesión Bacteriana , Colitis/etiología , Colitis/metabolismo , Colitis/microbiología , Femenino , Proteínas Fimbrias/genética , Proteínas Fimbrias/metabolismo , Fimbrias Bacterianas/genética , Fucosiltransferasas/deficiencia , Fucosiltransferasas/genética , Fucosiltransferasas/metabolismo , Interacciones Microbiota-Huesped , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiología , Mucosa Intestinal/patología , Masculino , Ratones , Ratones Endogámicos CBA , Ratones Noqueados , Operón , Salmonelosis Animal/etiología , Salmonelosis Animal/metabolismo , Salmonelosis Animal/microbiología , Salmonella typhimurium/genética , Salmonella typhimurium/fisiología , Galactósido 2-alfa-L-FucosiltransferasaRESUMEN
BACKGROUND: Salmonella enterica serovar Infantis (S. Infantis) is one of the ubiquitous serovars of the bacterial pathogen S. enterica and recently has been emerging in many countries worldwide. Nonetheless, not much is known about its epidemiology, host adaptation, and virulence. METHODS: Epidemiological and molecular approaches were used together with tissue-culture and mouse models to conduct phenotypic comparison with the model S. enterica serovar Typhimurium. RESULTS: We show that S. Infantis is more frequently associated with infections in infants <2 years old and prone to cause significantly less invasive infections than serovar Typhimurium. Moreover, although S. Infantis adheres better to host cells and highly colonizes mouse intestines soon after infection, it is significantly less invasive and induces much lower inflammation and disease in vivo than S. Typhimurium. These differences were associated with lower expression of Salmonella pathogenicity island (SPI) 1 genes in S. Infantis than in S. Typhimurium. CONCLUSIONS: Our results demonstrate previously unknown differences in the epidemiology, virulence pathway expression, and pathogenicity between two highly abundant Salmonella serovars and suggest that native variation in the expression of the SPI-1 regulon is likely to contribute to epidemiological and virulence variation between genetically similar nontyphoidal Salmonella serovars.
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Proteínas Bacterianas/genética , Expresión Génica , Salmonelosis Animal/epidemiología , Salmonella typhimurium/patogenicidad , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Células CACO-2 , Niño , Preescolar , Modelos Animales de Enfermedad , Femenino , Regulación Bacteriana de la Expresión Génica , Células HeLa , Humanos , Lactante , Recién Nacido , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Fenotipo , ARN Bacteriano/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Regulón , Salmonelosis Animal/microbiología , Virulencia/genética , Adulto JovenRESUMEN
Countries with a high incidence of helminth infections are characterized by high morbidity and mortality to infections with intracellular pathogens such as Salmonella. Some patients with Salmonella-Schistosoma co-infections develop a so-called "chronic septicemic salmonellosis," with prolonged fever and enlargement of the liver and spleen. These effects are most likely due to the overall immunoregulatory activities of schistosomes such as induction of Tregs, Bregs, alternatively activated macrophages, and degradation of antibodies. However, detailed underlying mechanisms are not very well investigated. Here, we show that intraperitoneal application of live Schistosoma mansoni eggs prior to infection with Salmonella Typhimurium in mice leads to an impairment of IFN-γ and IL-17 responses together with a higher bacterial load compared to Salmonella infection alone. S. mansoni eggs were found in granulomas in the visceral peritoneum attached to the colon. Immunohistological staining revealed IPSE/alpha-1, a glycoprotein secreted from live schistosome eggs, and recruited basophils around the eggs. Noteworthy, IPSE/alpha-1 is known to trigger IL-4 and IL-13 release from basophils which in turn is known to suppress Th1/Th17 responses. Therefore, our data support a mechanism of how schistosomes impair a protective immune response against Salmonella infection and increase our understanding of helminth-bacterial co-infections.
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Basófilos/inmunología , Proteínas del Huevo/metabolismo , Granuloma/patología , Proteínas del Helminto/metabolismo , Peritoneo/patología , Infecciones por Salmonella/inmunología , Salmonella typhimurium/fisiología , Schistosoma mansoni/fisiología , Esquistosomiasis mansoni/inmunología , Células TH1/inmunología , Células Th17/inmunología , Animales , Carga Bacteriana , Células Cultivadas , Coinfección , Citocinas/metabolismo , Huevos , Humanos , Inmunomodulación , Ratones , Ratones Endogámicos C57BLRESUMEN
The immune system is tightly controlled by regulatory processes that allow for the elimination of invading pathogens, while limiting immunopathological damage to the host. In the present study, we found that conditional deletion of the cell surface receptor Toso on B cells unexpectedly resulted in impaired proinflammatory T cell responses, which led to impaired immune protection in an acute viral infection model and was associated with reduced immunopathological tissue damage in a chronic inflammatory context. Toso exhibited its B cell-inherent immunoregulatory function by negatively controlling the pool of IL-10-competent B1 and B2 B cells, which were characterized by a high degree of self-reactivity and were shown to mediate immunosuppressive activity on inflammatory T cell responses in vivo. Our results indicate that Toso is involved in the differentiation/maintenance of regulatory B cells by fine-tuning B cell receptor activation thresholds. Furthermore, we showed that during influenza A-induced pulmonary inflammation, the application of Toso-specific antibodies selectively induced IL-10-competent B cells at the site of inflammation and resulted in decreased proinflammatory cytokine production by lung T cells. These findings suggest that Toso may serve as a novel therapeutic target to dampen pathogenic T cell responses via the modulation of IL-10-competent regulatory B cells.
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Linfocitos B Reguladores/inmunología , Diferenciación Celular/inmunología , Virus de la Influenza A , Proteínas de la Membrana/inmunología , Infecciones por Orthomyxoviridae/inmunología , Neumonía Viral/inmunología , Linfocitos T/inmunología , Animales , Linfocitos B Reguladores/patología , Diferenciación Celular/genética , Perros , Interleucina-10/genética , Interleucina-10/inmunología , Células de Riñón Canino Madin Darby , Proteínas de la Membrana/genética , Ratones , Ratones Noqueados , Infecciones por Orthomyxoviridae/genética , Infecciones por Orthomyxoviridae/patología , Neumonía Viral/genética , Neumonía Viral/patología , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/inmunología , Linfocitos T/patologíaRESUMEN
The FUT2 gene encodes an α-1,2-fucosyltransferase responsible for the expression of ABO histo-blood-group antigens on mucosal surfaces and bodily secretions. Individuals who carry at least one functional allele are known as "secretors," whereas those homozygous for loss-of-function mutations are known as "non-secretors." Non-secretor individuals are more susceptible to chronic inflammatory disorders such as Crohn's Disease, which may be mediated by alterations in the microbiota. Here, we investigated the dynamics of microbial community assembly with respect to genotype using a Fut2-deficient mouse model, taking the genotype of the maternal lineage over two generations into account. We found strong differences in community assembly of microbial communities over time, depending on the Fut2 genotype of the host and that of their progenitors. By applying network analyses, we further identified patterns of specialization and stabilization over time, which are influenced by the host and parental genotype during the process of community development. We also show genotype- and breeding-dependent patterns of community susceptibility to disturbance in a novel in silico approach integrating ecological- and network analysis. Our results indicate that it may be important to investigate the influence of Fut2 genotype in a familial context in order to fully understand its role in the etiology of chronic inflammatory disorders.
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Mycobacterium avium ssp. paratuberculosis (MAP) is the causative agent of Johne's disease (JD), a chronic inflammatory bowel disease of cattle characterized by intermittent to chronic diarrhea. In addition, MAP has been isolated from Crohn's disease (CD) patients. The impact of MAP on severity of clinical symptoms in JD as well as its role in CD are yet unknown. We have previously shown that MAP is able to colonize inflamed enteric tissue and to exacerbate the inflammatory tissue response (Suwandi et al., 2014). In the present study, we analyzed how repeated MAP administration influences the course of dextran sulfate sodium (DSS)-induced colitis. In comparison to mice exposed to DSS or MAP only, repeated exposure of DSS-treated mice to MAP (DSS/MAP) revealed a significantly enhanced clinical score, reduction of colon length as well as severe CD4+ T cell infiltration into the colonic lamina propria. Functional analysis identified a critical role of CD4+ T cells in the MAP-induced disease exacerbation. Additionally, altered immune responses were observed when closely related mycobacteria species such as M. avium ssp. avium and M. avium ssp. hominissuis were administered. These data reveal the specific ability of MAP to aggravate intestinal inflammation and clinical symptoms. Overall, this phenotype is compatible with similar disease promoting capabilites of MAP in JD and CD.
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Linfocitos T CD4-Positivos/inmunología , Colitis/patología , Mycobacterium avium subsp. paratuberculosis/inmunología , Mycobacterium avium subsp. paratuberculosis/patogenicidad , Paratuberculosis/patología , Animales , Colitis/inducido químicamente , Colon/patología , Enfermedad de Crohn/patología , Sulfato de Dextran/administración & dosificación , Sulfato de Dextran/toxicidad , Humanos , RatonesRESUMEN
Type I interferons (IFN-I), such as IFN-α and IFN-ß are important messengers in the host response against bacterial infections. Knowledge about the role of IFN-I in infections by nontuberculous mycobacteria (NTM) is limited. Here we show that macrophages infected with pathogens of the Mycobacterium avium complex produced significantly lower amounts of IFN-ß than macrophages infected with the opportunistic pathogen M. smegmatis. To dissect the molecular mechanisms of this phenomenon, we focused on the obligate pathogen Mycobacterium avium ssp paratuberculosis (MAP) and the opportunistic M. smegmatis. Viability of both bacteria was required for induction of IFN-ß in macrophages. Both bacteria induced IFN-ß via the cGAS-STING-TBK1-IRF3/7-pathway of IFN-ß activation. Stronger phosphorylation of TBK1 and higher amounts of extracellular bacterial DNA in the macrophage cytosol were found in M. smegmatis infected macrophages than in MAP infected macrophages. After intraperitoneal infection of mice, a strong Ifnb induction by M. smegmatis correlated with clearance of the bacteria. In contrast, MAP only induced weak Ifnb expression which correlated with bacterial persistence and increased number of granulomas in the liver. In mice lacking the type I interferon receptor we observed improved survival of M. smegmatis while survival of MAP was similar to that in wildtype mice. On the other hand, treatment of MAP infected wildtype mice with the IFN-I inducer poly(I:C) or recombinant IFN-ß impaired the survival of MAP. This indicates an essential role of IFN-I in clearing infections by MAP and M. smegmatis. The expression level of IFN-I is decisive for transient versus persistent NTM infection.
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Factor 3 Regulador del Interferón/metabolismo , Factor 7 Regulador del Interferón/metabolismo , Interferón beta/metabolismo , Infecciones por Mycobacterium no Tuberculosas/metabolismo , Mycobacterium avium subsp. paratuberculosis/fisiología , Mycobacterium smegmatis/fisiología , Nucleotidiltransferasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Animales , Femenino , Interacciones Huésped-Patógeno , Humanos , Factor 3 Regulador del Interferón/genética , Factor 7 Regulador del Interferón/genética , Interferón beta/genética , Macrófagos/microbiología , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Infecciones por Mycobacterium no Tuberculosas/genética , Infecciones por Mycobacterium no Tuberculosas/microbiología , Mycobacterium avium subsp. paratuberculosis/genética , Mycobacterium smegmatis/genética , Nucleotidiltransferasas/genética , Fosforilación , Proteínas Serina-Treonina Quinasas/genética , Transducción de SeñalRESUMEN
Glycans play important roles in host-microbe interactions. Tissue-specific expression patterns of the blood group glycosyltransferase ß-1,4-N-acetylgalactosaminyltransferase 2 (B4galnt2) are variable in wild mouse populations, and loss of B4galnt2 expression is associated with altered intestinal microbiota. We hypothesized that variation in B4galnt2 expression alters susceptibility to intestinal pathogens. To test this, we challenged mice genetically engineered to express different B4galnt2 tissue-specific patterns with a Salmonella Typhimurium infection model. We found B4galnt2 intestinal expression was strongly associated with bacterial community composition and increased Salmonella susceptibility as evidenced by increased intestinal inflammatory cytokines and infiltrating immune cells. Fecal transfer experiments demonstrated a crucial role of the B4galnt2-dependent microbiota in conferring susceptibility to intestinal inflammation, while epithelial B4galnt2 expression facilitated epithelial invasion of S. Typhimurium. These data support a critical role for B4galnt2 in gastrointestinal infections. We speculate that B4galnt2-specific differences in host susceptibility to intestinal pathogens underlie the strong signatures of balancing selection observed at the B4galnt2 locus in wild mouse populations.
Asunto(s)
Microbioma Gastrointestinal/genética , Predisposición Genética a la Enfermedad/genética , Mucosa Intestinal/microbiología , N-Acetilgalactosaminiltransferasas/biosíntesis , Salmonelosis Animal/genética , Animales , Ensayo de Inmunoadsorción Enzimática , Interacciones Huésped-Parásitos/fisiología , Inmunohistoquímica , Hibridación Fluorescente in Situ , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , N-Acetilgalactosaminiltransferasas/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Salmonelosis Animal/microbiología , Salmonella typhimurium , TransfecciónRESUMEN
Mycobacterium avium subspecies paratuberculosis (MAP) causes Johne's disease, a chronic granulomatous enteritis in ruminants. Furthermore, infections of humans with MAP have been reported and a possible association with Crohn's disease and diabetes type I is currently discussed. MAP owns large sequence polymorphisms (LSPs) that were exclusively found in this mycobacteria species. The relevance of these LSPs in the pathobiology of MAP is still unclear. The mptD gene (MAP3733c) of MAP belongs to a small group of functionally uncharacterized genes, which are not present in any other sequenced mycobacteria species. mptD is part of a predicted operon (mptABCDEF), encoding a putative ATP binding cassette-transporter, located on the MAP-specific LSP14. In the present study, we generated an mptD knockout strain (MAPΔmptD) by specialized transduction. In order to investigate the potential role of mptD in the host, we performed infection experiments with macrophages. By this, we observed a significantly reduced cell number of MAPΔmptD early after infection, indicating that the mutant was hampered with respect to adaptation to the early macrophage environment. This important role of mptD was supported in mouse infection experiments where MAPΔmptD was significantly attenuated after peritoneal challenge. Metabolic profiling was performed to determine the cause for the reduced virulence and identified profound metabolic disorders especially in the lipid metabolism of MAPΔmptD. Overall our data revealed the mptD gene to be an important factor for the metabolic adaptation of MAP required for persistence in the host.