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1.
Helicobacter ; 28(5): e13001, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37334992

RESUMO

BACKGROUND: It has been documented that Helicobacter hepaticus produces a nickel-containing hydrogen-oxidizing hydrogenase enzyme, which is necessary for hydrogen-supported amino acid uptake. Although H. hepaticus infection has been shown to promote liver inflammation and fibrosis in BALB/c mice, the impact of hydrogenase on the progression of liver fibrosis induced by H. hepaticus has not been explored. MATERIALS AND METHODS: BALB/c mice were inoculated with hydrogenase mutant (ΔHyaB) or wild type (WT) H. hepaticus 3B1 for 12 and 24 weeks. H. hepaticus colonization, hepatic histopathology, serum biochemistry, expression of inflammatory cytokines, and oxidative stress signaling pathways were detected. RESULTS: We found that ΔHyaB had no influence on the colonization of H. hepaticus in the liver of mice at 12 and 24 weeks post infection (WPI). However, mice infected by ΔHyaB strains developed significantly alleviated liver inflammation and fibrosis compared with WT infection. Moreover, ΔHyaB infection remarkably increased the expression of hepatic GSH, SOD, and GSH-Px, and decreased the liver levels of MDA, ALT, and AST compared to WT H. hepaticus infected group from 12 to 24 WPI. Furthermore, mRNA levels of Il-6, Tnf-α, iNos, Hmox-1, and α-SMA were significantly decreased with an increase of Nfe2l2 in the liver of mice infected by ΔHyaB strains. In addition, ΔHyaB H. hepaticus restored the activation of the Nrf2/HO-1 signaling pathway, which is inhibited by H. hepaticus infection. CONCLUSIONS: These data demonstrated that H. hepaticus hydrogenase promoted liver inflammation and fibrosis development mediated by oxidative stress in male BALB/c mice.


Assuntos
Infecções por Helicobacter , Helicobacter pylori , Hidrogenase , Masculino , Animais , Camundongos , Helicobacter hepaticus/genética , Hidrogenase/genética , Hidrogenase/metabolismo , Camundongos Endogâmicos BALB C , Infecções por Helicobacter/patologia , Helicobacter pylori/metabolismo , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Fígado/patologia , Fibrose , Estresse Oxidativo , Hidrogênio/metabolismo
2.
Curr Opin Microbiol ; 65: 145-155, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34883389

RESUMO

Pathobionts are members of the gut microbiota with the capacity to cause disease when there is malfunctioning intestinal homeostasis. These organisms are thought to be major contributors to the pathogenesis of inflammatory bowel disease (IBD), a group of chronic inflammatory disorders driven by dysregulated responses towards the microbiota. Over two decades have passed since the discovery of Helicobacter hepaticus, a mouse pathobiont which causes colitis in the context of immune deficiency. During this time, we have developed a detailed understanding of the cellular players and cytokine networks which drive H. hepaticus immunopathology. However, we are just beginning to understand the microbial factors that enable H. hepaticus to interact with the host and influence colonic health and disease. Here we review key H. hepaticus-host interactions, their relevance to other exemplar pathobionts and how when maladapted they drive colitis. Further understanding of these pathways may offer new therapeutic approaches for IBD.


Assuntos
Colite , Doenças Inflamatórias Intestinais , Animais , Helicobacter hepaticus/genética , Doenças Inflamatórias Intestinais/genética , Intestinos , Camundongos
3.
Helicobacter ; 25(2): e12677, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31881556

RESUMO

BACKGROUND: It has been documented that Helicobacter hepaticus (H hepaticus) infection is linked to chronic hepatitis and liver cancer. However, our understanding of the molecular mechanisms underlying progression of the H hepaticus-induced hepatic inflammation to cellular hepatocarcinoma is still limited. MATERIALS AND METHODS: In our study, male BALB/c mice were infected by H hepaticus for 8, 12, 16, 20, and 24 weeks. Histopathology, H hepaticus colonization dynamics, select signaling pathways, and expression of key inflammatory cytokines in the liver were examined. RESULTS: We found that H hepaticus was detectible in feces of mice at 7 days postinfection (DPI) by PCR, but it was not detected in the livers by PCR until 8 weeks postinfection (WPI). In addition, abundance of colonic and hepatic H hepaticus was progressively increased over the infection duration. H hepaticus-induced hepatic inflammation and fibrosis were aggravated over the infection duration, and necrosis or cirrhosis developed in the infected liver at 24 WPI H hepaticus infection increased levels of alanine aminotransferase and aspartate aminotransferase. Moreover, mRNA levels of Il-6 and Tnf-α were significantly elevated in the livers of H hepaticus-infected mice compared to uninfected control from 8 WPI to 24 WPI. Furthermore, Stat3, nuclear factor-κB (p65), and MAPK (Erk1/2 and p38) were activated by H hepaticus infection. CONCLUSIONS: These data demonstrated that male BALB/c mice can be used as a new mouse model of H hepaticus-induced liver diseases and that the H hepaticus-induced liver injury is triggered by NF-κB, Jak-Stat, and MAPK signaling pathways.


Assuntos
Fibrose/microbiologia , Helicobacter hepaticus , Hepatite Crônica/microbiologia , Sistema de Sinalização das MAP Quinases , NF-kappa B/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Citocinas/biossíntese , Fezes/microbiologia , Infecções por Helicobacter/patologia , Helicobacter hepaticus/genética , Helicobacter hepaticus/isolamento & purificação , Fígado/microbiologia , Fígado/patologia , Neoplasias Hepáticas/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Modelos Animais , Transdução de Sinais , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
4.
Nature ; 554(7692): 373-377, 2018 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-29414937

RESUMO

Both microbial and host genetic factors contribute to the pathogenesis of autoimmune diseases. There is accumulating evidence that microbial species that potentiate chronic inflammation, as in inflammatory bowel disease, often also colonize healthy individuals. These microorganisms, including the Helicobacter species, can induce pathogenic T cells and are collectively referred to as pathobionts. However, how such T cells are constrained in healthy individuals is not yet understood. Here we report that host tolerance to a potentially pathogenic bacterium, Helicobacter hepaticus, is mediated by the induction of RORγt+FOXP3+ regulatory T (iTreg) cells that selectively restrain pro-inflammatory T helper 17 (TH17) cells and whose function is dependent on the transcription factor c-MAF. Whereas colonization of wild-type mice by H. hepaticus promoted differentiation of RORγt-expressing microorganism-specific iTreg cells in the large intestine, in disease-susceptible IL-10-deficient mice, there was instead expansion of colitogenic TH17 cells. Inactivation of c-MAF in the Treg cell compartment impaired differentiation and function, including IL-10 production, of bacteria-specific iTreg cells, and resulted in the accumulation of H. hepaticus-specific inflammatory TH17 cells and spontaneous colitis. By contrast, RORγt inactivation in Treg cells had only a minor effect on the bacteria-specific Treg and TH17 cell balance, and did not result in inflammation. Our results suggest that pathobiont-dependent inflammatory bowel disease is driven by microbiota-reactive T cells that have escaped this c-MAF-dependent mechanism of iTreg-TH17 homeostasis.


Assuntos
Colite/imunologia , Colite/microbiologia , Helicobacter hepaticus/imunologia , Tolerância Imunológica , Intestinos/imunologia , Intestinos/microbiologia , Proteínas Proto-Oncogênicas c-maf/metabolismo , Linfócitos T Reguladores/imunologia , Animais , Bioengenharia , Colite/patologia , Feminino , Fatores de Transcrição Forkhead/metabolismo , Helicobacter hepaticus/genética , Helicobacter hepaticus/patogenicidade , Homeostase , Interações Hospedeiro-Patógeno , Interleucina-10/biossíntese , Interleucina-10/deficiência , Interleucina-10/imunologia , Masculino , Camundongos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/antagonistas & inibidores , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Proteínas Proto-Oncogênicas c-maf/deficiência , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/metabolismo , Células Th17/citologia , Células Th17/imunologia
5.
Cell Microbiol ; 19(7)2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28111881

RESUMO

Multiple pathogenic Gram-negative bacteria produce the cytolethal distending toxin (CDT) with activity of DNase I; CDT can induce DNA double-strand breaks (DSBs), G2/M cell cycle arrest, and apoptosis in cultured mammalian cells. However, the link of CDT to in vivo tumorigenesis is not fully understood. In this study, 129/SvEv Rag2-/- mice were gavaged with wild-type Helicobacter hepatics 3B1(Hh) and its isogenic cdtB mutant HhcdtBm7, followed by infection for 10 and 20 weeks (WPI). HhCDT deficiency did not affect cecal colonization levels of HhcdtBm7, but attenuated severity of cecal pathology in HhcdtBm7-infected mice. Of importance, preneoplasic dysplasia was progressed to cancer from 10 to 20 WPI in the Hh-infected mice but not in the HhcdtBm7-infected mice. In addition, the loss of HhCDT significantly dampened transcriptional upregulation of cecal Tnfα and Il-6, but elevated Il-10 mRNA levels when compared to Hh at 10 WPI. Furthermore, the presence of HhCDT increased numbers of lower bowel intestinal γH2AX-positive epithelial cells (a marker of DSBs) at both 10 and 20 WPI and augmented phospho-Stat3 foci+ intestinal crypts (activation of Stat3) at 20 WPI. Our findings suggest that CDT promoted Hh carcinogenesis by enhancing DSBs and activation of the Tnfα/Il-6-Stat3 signaling pathway.


Assuntos
Toxinas Bacterianas/metabolismo , Carcinogênese/patologia , Quebras de DNA de Cadeia Dupla , Helicobacter hepaticus/patogenicidade , Interleucina-6/metabolismo , Intestinos/patologia , Fator de Transcrição STAT3/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Apoptose , Toxinas Bacterianas/genética , Ceco/microbiologia , Ceco/patologia , Feminino , Pontos de Checagem da Fase G2 do Ciclo Celular , Helicobacter hepaticus/genética , Histonas/metabolismo , Interleucina-10/genética , Masculino , Camundongos , Camundongos Transgênicos , Neoplasias/microbiologia , Neoplasias/patologia , RNA Mensageiro/biossíntese , Transdução de Sinais/fisiologia
6.
Helicobacter ; 19(4): 280-8, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24853076

RESUMO

BACKGROUND: Helicobacter pylori cholesterol-α-glucosyltransferase (cgt) is essential for survival of H. pylori in mice. Enterohepatic H. hepaticus, the cause of colonic and hepatocellular carcinoma in susceptible mouse strains, contains an ortholog of the H. pylori cgt. However, the role of cgt in the pathogenesis of H. hepaticus has not been investigated. MATERIALS AND METHODS: Two cgt-deficient isogenic mutants of wild-type H. hepaticus (WT) 3B1 were generated and used to inoculate male A/JCr mice. Cecal and hepatic colonization levels of the mutants and WT 3B1 as well as select inflammation-associated cytokines were measured by qPCR at 4 months postinoculation. RESULTS: Both mutants were undetectable in the cecum of any inoculated mice (10 per mutant) but were detected in two livers (one for each mutant); by contrast, 9 and 7 of 10 mice inoculated with WT 3B1 were qPCR positive in the ceca and livers, respectively. The mice inoculated with the mutants developed significantly less severe hepatic inflammation (p < .05) and also produced significantly lower hepatic mRNA levels of proinflammatory cytokines Ifn-γ (p < .01) and Tnf-α (p ≤ .02) as well as anti-inflammatory factors Il10 and Foxp3 compared with the WT 3B1-inoculated mice. Additionally, the WT 3B1-inoculated mice developed significantly higher Th1-associated IgG2a (p < .0001) and Th2-associated IgG1 responses (p < .0001) to H. hepaticus infection than mice dosed with isogenic cgt mutants. CONCLUSION: Our data indicate that the cholesterol-α-glucosyltransferase is required for establishing colonization of the intestine and liver and therefore plays a critical role in the pathogenesis of H. hepaticus.


Assuntos
Colesterol/metabolismo , Glucosiltransferases/metabolismo , Infecções por Helicobacter/microbiologia , Helicobacter hepaticus/enzimologia , Helicobacter hepaticus/crescimento & desenvolvimento , Fatores de Virulência/metabolismo , Animais , Ceco/microbiologia , Citocinas/biossíntese , Modelos Animais de Doenças , Deleção de Genes , Glucosiltransferases/genética , Helicobacter hepaticus/genética , Fígado/microbiologia , Masculino , Camundongos Endogâmicos A , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Virulência/genética
7.
Singapore Med J ; 54(8): 451-7, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24005452

RESUMO

INTRODUCTION: Helicobacter (H.) hepaticus infection causes chronic active hepatitis and induces hepatocellular tumours in A/JCr mice, but evidence of this in humans is scarce. This study aimed to demonstrate the correlation between H. hepaticus and human primary hepatocellular carcinoma (HCC). METHODS: The sera of 50 patients with primary HCC were tested for the presence of anti-H. pylori and anti-H. hepaticus immunoglobulin G (IgG) antibodies. The liver tissues of patients who tested positive for serum antibody were analysed for H. hepaticus-specific 16S rRNA, H. hepaticus cdtB, H. pylori cagA, H. pylori vacA and H. pylori ureC genes using polymerase chain reaction. RESULTS: After the anti-H. pylori antibodies in the serum samples were absorbed by H. pylori antigen, the anti-H. hepaticus IgG serum antibody detection rate was 50.0% in patients with primary HCC. This was significantly higher (p < 0.001) than the detection rate in the benign liver tumour (7.7%) and normal liver tissue (6.3%) groups. Of the 25 primary HCC samples that tested positive for anti-H. hepaticus IgG serum antibody, the H. hepaticus-specific 16S rRNA gene was detected in nine (36.0%) samples. Sequencing showed that the polymerase chain reaction-amplified product exhibited 95.5%-100% homology to the H. hepaticus-specific 16S rRNA gene. Among these nine primary HCC tissue samples, the H. hepaticus cdtB gene was detected in four (44.4%) samples, while no such expression was observed in the benign liver tumour or normal liver tissue groups. CONCLUSION: The present study identified the presence of H. hepaticus infection in patients with primary HCC using serological and molecular biological detection, suggesting that H. hepaticus infection may be involved in the progression of HCC.


Assuntos
Carcinoma Hepatocelular/microbiologia , Infecções por Helicobacter/microbiologia , Helicobacter hepaticus/isolamento & purificação , Neoplasias Hepáticas/microbiologia , Adulto , Idoso , DNA Bacteriano/genética , Feminino , Infecções por Helicobacter/genética , Helicobacter hepaticus/genética , Helicobacter pylori/genética , Helicobacter pylori/isolamento & purificação , Humanos , Imunoglobulina G/sangue , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase
8.
Cell Microbiol ; 15(6): 992-1011, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23278999

RESUMO

The enterohepatic Epsilonproteobacterium Helicobacter hepaticus persistently colonizes the intestine of mice and causes chronic inflammatory symptoms in susceptible mouse strains. The bacterial factors causing intestinal inflammation are poorly characterized. A large genomic pathogenicity island, HHGI1, which encodes components of a type VI secretion system (T6SS), was previously shown to contribute to the colitogenic potential of H. hepaticus. We have now characterized the T6SS components Hcp, VgrG1, VgrG2 and VgrG3, encoded on HHGI1, including the potential impact of the T6SS on intestinal inflammation in a mouse T-cell transfer model. The H. hepaticus T6SS components were expressed during the infection and secreted in a T6SS-dependent manner, when the bacteria were cultured either in the presence or in the absence of mouse intestinal epithelial cells. Mutants deficient in VgrG1 displayed a significantly lower colitogenic potential in T-cell-transferred C57BL/6 Rag2(-/-) mice, despite an unaltered ability to colonize mice persistently. Intestinal microbiota analyses demonstrated only minor changes in mice infected with wild-typeH. hepaticus as compared with mice infected with VgrG1-deficient isogenic bacteria. In addition, competitive assays between both wild-type and T6SS-deficient H. hepaticus, and between wild-type H. hepaticus and Campylobacter jejuni or Enterobacteriaceae species did not show an effect of the T6SS on interbacterial competitiveness. Therefore, we suggest that microbiota alterations did not play a major role in the changes of pro-inflammatory potential mediated by the T6SS. Cellular innate pro-inflammatory responses were increased by the secreted T6SS proteins VgrG1 and VgrG2. We therefore concluded that the type VI secretion component VgrG1 can modulate and specifically exacerbate the innate pro-inflammatory effect of the chronic H. hepaticus infection.


Assuntos
Proteínas de Bactérias/metabolismo , Sistemas de Secreção Bacterianos/fisiologia , Colite/microbiologia , Infecções por Helicobacter/fisiopatologia , Helicobacter hepaticus/fisiologia , Helicobacter hepaticus/patogenicidade , Animais , Proteínas de Bactérias/fisiologia , Campylobacter jejuni/fisiologia , Células Cultivadas , Colite/metabolismo , Colite/fisiopatologia , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Enterobacteriaceae/fisiologia , Infecções por Helicobacter/metabolismo , Helicobacter hepaticus/genética , Técnicas In Vitro , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Mucosa Intestinal/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação/genética
9.
Microbiology (Reading) ; 159(Pt 1): 136-146, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23139401

RESUMO

Helicobacter hepaticus open reading frame HH0352 was identified as a nickel-responsive regulator NikR. The gene was disrupted by insertion of an erythromycin resistance cassette. The H. hepaticus nikR mutant had five- to sixfold higher urease activity and at least twofold greater hydrogenase activity than the wild-type strain. However, the urease apo-protein levels were similar in both the wild-type and the mutant, suggesting the increase in urease activity in the mutant was due to enhanced Ni-maturation of the urease. Compared with the wild-type strain, the nikR strain had increased cytoplasmic nickel levels. Transcription of nikABDE (putative inner membrane Ni transport system) and hh0418 (putative outer membrane Ni transporter) was nickel- and NikR-repressed. Electrophoretic mobility shift assays (EMSAs) revealed that purified HhNikR could bind to the nikABDE promoter (P(nikA)), but not to the urease or the hydrogenase promoter; NikR-P(nikA) binding was enhanced in the presence of nickel. Also, qRT-PCR and EMSAs indicated that neither nikR nor the exbB-exbD-tonB were under the control of the NikR regulator, in contrast with their Helicobacter pylori homologues. Taken together, our results suggest that HhNikR modulates urease and hydrogenase activities by repressing the nickel transport/nickel internalization systems in H. hepaticus, without direct regulation of the Ni-enzyme genes (the latter is the case for H. pylori). Finally, the nikR strain had a two- to threefold lower growth yield than the parent, suggesting that the regulatory protein might play additional roles in the mouse liver pathogen.


Assuntos
Regulação Bacteriana da Expressão Gênica , Helicobacter hepaticus/enzimologia , Hidrogenase/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Níquel/metabolismo , Proteínas Repressoras/metabolismo , Urease/metabolismo , Sequência de Aminoácidos , Sequência de Bases , DNA Bacteriano/metabolismo , Ensaio de Desvio de Mobilidade Eletroforética , Perfilação da Expressão Gênica , Técnicas de Inativação de Genes , Helicobacter hepaticus/genética , Helicobacter hepaticus/crescimento & desenvolvimento , Dados de Sequência Molecular , Mutagênese Insercional , Regiões Promotoras Genéticas , Ligação Proteica , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Repressoras/genética
10.
Inflamm Bowel Dis ; 18(5): 943-54, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22238116

RESUMO

BACKGROUND: Colitis susceptibility in Il10(-/-) mice depends on genetic background and microbiota composition. A major genetic locus mediating colitis susceptibility, Cdcs1, was transferred from susceptible C3Bir-Il10(-/-) to resistant B6-Il10(-/-) mice, resulting in susceptible congenic BC-R3-Il10(-/-) mice. The aim of this study was to determine the impact of microbiota on this differential colitis susceptibility using a Helicobacter hepaticus infection model. METHODS: Parental C3Bir-Il10(-/-) , B6-Il10(-/-) , and congenic BC-R3-Il10(-/-) mice were inoculated with H. hepaticus and analyzed for inflammation. In parental Il10(-/-) mice, microbiota composition was determined by terminal restriction fragment length polymorphism (T-RFLP) and quantitative polymerase chain reaction (qPCR). RESULTS: Most severe inflammation was observed in C3Bir-Il10(-/-) in the cecum, in BC-R3-Il10(-/-) in cecum and colon, and, unexpectedly, in B6-Il10(-/-) in the colon. C3Bir-Il10(-/-) and BC-R3-Il10(-/-) secreted significantly more interferon-gamma (IFNγ) and interleukin (IL)17 than B6-Il10(-/-) . T-RFLP analyses in C3Bir-Il10(-/-) and B6-Il10(-/-) mice revealed 1) a significant impact of H. hepaticus infection on species richness and diversity, and 2) strain differences in microbiota composition only after H. hepaticus infection. qPCR revealed higher numbers of Clostridia leptum and Bacteroides spp. in the cecum of infected C3Bir-Il10(-/-) mice, and Lactobacillus spp. in B6-Il10(-/-) mice. CONCLUSIONS: Cdcs1 modifies the response to H. hepaticus infection. However, this infection alone does not reflect the original response to a complex colitogenic biota. H. hepaticus-induced inflammation altered intestinal microbiota in a mouse strain-specific manner. Bacteroides spp. became more abundant in susceptible C3Bir-Il10(-/-) , lactobacilli in B6-Il10(-/-) mice. Therefore, both host immune response and differential compositional changes of microbiota play a role in strain-specific colitis susceptibility in Il10(-/-) mice.


Assuntos
Colite/etiologia , Suscetibilidade a Doenças , Trato Gastrointestinal/microbiologia , Interleucina-10/fisiologia , Metagenoma/fisiologia , Animais , Colite/patologia , Colo/microbiologia , Citocinas/metabolismo , DNA Bacteriano/genética , Ensaio de Imunoadsorção Enzimática , Infecções por Helicobacter/genética , Infecções por Helicobacter/microbiologia , Helicobacter hepaticus/genética , Inflamação/etiologia , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Polimorfismo de Fragmento de Restrição , RNA Ribossômico 16S/genética , Reação em Cadeia da Polimerase em Tempo Real , Especificidade da Espécie
11.
Mucosal Immunol ; 4(1): 22-30, 2011 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-20944559

RESUMO

Pioneering work in the 1990s first linked a novel microaerobic bacterium, Helicobacter hepaticus, with chronic active hepatitis and inflammatory bowel disease in several murine models. Targeted H. hepaticus infection experiments subsequently demonstrated its ability to induce colitis, colorectal cancer, and extraintestinal diseases in a number of mouse strains with defects in immune function and/or regulation. H. hepaticus is now widely utilized as a model system to dissect how intestinal microbiota interact with the host to produce both inflammatory and tolerogenic responses. This model has been used to make important advances in understanding factors that regulate both acquired and innate immune response within the intestine. Further, it has been an effective tool to help define the function of regulatory T cells, including their ability to directly inhibit the innate inflammatory response to gut microbiota. The complete genomic sequence of H. hepaticus has advanced the identification of several virulence factors and aided in the elucidation of H. hepaticus pathogenesis. Delineating targets of H. hepaticus virulence factors could facilitate novel approaches to treating microbially induced lower bowel inflammatory diseases.


Assuntos
Neoplasias Colorretais , Modelos Animais de Doenças , Infecções por Helicobacter , Helicobacter hepaticus , Interações Hospedeiro-Patógeno , Doenças Inflamatórias Intestinais , Trato Gastrointestinal Inferior , Animais , Neoplasias Colorretais/microbiologia , Neoplasias Colorretais/patologia , Infecções por Helicobacter/imunologia , Infecções por Helicobacter/microbiologia , Infecções por Helicobacter/patologia , Helicobacter hepaticus/genética , Helicobacter hepaticus/imunologia , Helicobacter hepaticus/patogenicidade , Doenças Inflamatórias Intestinais/imunologia , Doenças Inflamatórias Intestinais/microbiologia , Doenças Inflamatórias Intestinais/patologia , Trato Gastrointestinal Inferior/imunologia , Trato Gastrointestinal Inferior/microbiologia , Trato Gastrointestinal Inferior/patologia , Camundongos , Úlcera Péptica/microbiologia , Linfócitos T/imunologia , Fatores de Virulência
12.
Glycobiology ; 20(9): 1077-88, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20466652

RESUMO

Lewis x (Le(x)) and sialyl Lewis x (SLe(x))-containing glycans play important roles in numerous physiological and pathological processes. The key enzyme for the final step formation of these Lewis antigens is alpha1-3-fucosyltransferase. Here we report molecular cloning and functional expression of a novel Helicobacter hepaticus alpha1-3-fucosyltransferase (HhFT1) which shows activity towards both non-sialylated and sialylated Type II oligosaccharide acceptor substrates. It is a promising catalyst for enzymatic and chemoenzymatic synthesis of Le(x), sialyl Le(x) and their derivatives. Unlike all other alpha1-3/4-fucosyltransferases characterized so far which belong to Carbohydrate Active Enzyme (CAZy, http://www.cazy.org/) glycosyltransferase family GT10, the HhFT1 shares protein sequence homology with alpha1-2-fucosyltransferases and belongs to CAZy glycosyltransferase family GT11. The HhFT1 is thus the first alpha1-3-fucosyltransferase identified in the GT11 family.


Assuntos
Proteínas de Bactérias/genética , Fucosiltransferases/genética , Glicosiltransferases/genética , Helicobacter hepaticus/genética , Sequência de Aminoácidos , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Sequência de Bases , Clonagem Molecular , Fucosiltransferases/isolamento & purificação , Fucosiltransferases/metabolismo , Glicosiltransferases/metabolismo , Modelos Biológicos , Dados de Sequência Molecular , Família Multigênica , Filogenia , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos
13.
Cell Host Microbe ; 7(4): 265-276, 2010 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-20413095

RESUMO

The gastrointestinal tract harbors a diverse microbiota that has coevolved with mammalian hosts. Though most associations are symbiotic or commensal, some resident bacteria (termed pathobionts) have the potential to cause disease. Bacterial type VI secretion systems (T6SSs) are one mechanism for forging host-microbial interactions. Here we reveal a protective role for the T6SS of Helicobacter hepaticus, a Gram-negative bacterium of the intestinal microbiota. H. hepaticus mutants with a defective T6SS display increased numbers within intestinal epithelial cells (IECs) and during intestinal colonization. Remarkably, the T6SS directs an anti-inflammatory gene expression profile in IECs, and CD4+ T cells from mice colonized with T6SS mutants produce increased interleukin-17 in response to IECs presenting H. hepaticus antigens. Thus, the H. hepaticus T6SS limits colonization and intestinal inflammation, promoting a balanced relationship with the host. We propose that disruption of such balances contributes to human disorders such as inflammatory bowel disease and colon cancer.


Assuntos
Proteínas de Bactérias/fisiologia , Helicobacter hepaticus/imunologia , Helicobacter hepaticus/patogenicidade , Interações Hospedeiro-Patógeno , Substâncias Macromoleculares/metabolismo , Proteínas de Membrana/fisiologia , Animais , Proteínas de Bactérias/genética , Linfócitos T CD4-Positivos/microbiologia , Contagem de Colônia Microbiana , DNA Bacteriano/química , DNA Bacteriano/genética , Células Epiteliais/microbiologia , Perfilação da Expressão Gênica , Técnicas de Inativação de Genes , Helicobacter hepaticus/genética , Humanos , Evasão da Resposta Imune , Interleucina-17/biossíntese , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Modelos Biológicos , Dados de Sequência Molecular , Análise de Sequência de DNA
14.
Helicobacter ; 14(6): 545-51, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19889072

RESUMO

BACKGROUND: Since the discovery of Helicobacter pylori, various enterohepatic Helicobacter spices have been detected in the guts of humans and animals. Some enterohepatic Helicobacters have been associated with inflammatory bowel disease or liver disease in mice. However the association of these bacteria with human diseases remains unknown. MATERIALS AND METHODS: We collected 126 bile samples from patients with cholelithiasis, cholecystitis, gallbladder polyp, and other nonbiliary diseases. Samples were screened for the presence of enterohepatic Helicobacter spp. using cultures, nested PCR, or in situ hybridization. We tested for antibodies to H. pylori and H. hepaticus by Western blot analysis. RESULTS: Attempts at cultivation were unsuccessful. However, H. hepaticus was detected in bile samples with nested PCR whereas H. bilis was not. Helicobacter hepaticus in the bile was confirmed by in situ hybridization, but H. hepaticus from bile samples was coccoid in appearance. We detected immunoglobulin G antibodies to H. hepaticus in bile samples by Western blotting. Helicobacter hepaticus was detected in 40 (32%) of total 126 samples as H. hepaticus positive if at least one of the three methods with nested PCR, in situ, or Western blotting. Patients with cholelithiasis (41%) and cholecystitis with gastric cancer (36%) had significantly higher (p = .029) prevalence of H. hepaticus infection than samples from patients with other diseases. CONCLUSION: Helicobacter hepaticus may closely associate with diseases of the liver and biliary tract in humans.


Assuntos
Bile/microbiologia , Doenças Biliares/microbiologia , Infecções por Helicobacter/microbiologia , Helicobacter hepaticus/isolamento & purificação , Idoso , Anticorpos Antibacterianos/análise , Bile/imunologia , Doenças Biliares/imunologia , Feminino , Infecções por Helicobacter/imunologia , Helicobacter hepaticus/genética , Helicobacter hepaticus/imunologia , Humanos , Masculino , Pessoa de Meia-Idade
15.
Nan Fang Yi Ke Da Xue Xue Bao ; 29(6): 1212-5, 2009 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-19726364

RESUMO

OBJECTIVE: To clone the gene encoding methyl-accepting chemotaxis signal transduction protein (MCSTP) of Helicobacter hepaticus and analyze the gene structures using bioinformatics methods. METHODS: With the specific primer of Helicobacter hepaticus MCSTP c1977, MCSTP gene was amplified by PCR from the genomic DNA of Helicobacter hepaticus and ligated to the prokaryotic expression vector pET22b(+). After sequencing, the sequence homology and structural feature of MCSTP gene were analyzed by bioinformatics method. RESULTS: A 99% similarity was identified between MCSTP gene cloned and its counterpart in standard Helicobacter hepaticus strain ATCC51449 genome DNA published by GenBank, with only a replacement of A by T at 1160 bp. A low homology was found in the MCSTP genes between Helicobacter hepaticus, Campylobacter jejuni and Helicobacter pylori by bioinformatics analysis, suggesting the specificity of MCSTP gene in Helicobacter hepaticus among the microbes. CONCLUSION: The prokaryotic expression plasmid pET22b(+)/MCSTP is constructed successfully, and the bioinformatics analysis provided evidences and clues for further study of the biological functions and pathogenic mechanism of MCSTP.


Assuntos
Proteínas de Bactérias/metabolismo , Helicobacter hepaticus/genética , Proteínas de Membrana/metabolismo , Proteínas de Bactérias/genética , Clonagem Molecular , Biologia Computacional/métodos , Vetores Genéticos/genética , Helicobacter hepaticus/isolamento & purificação , Helicobacter hepaticus/metabolismo , Proteínas de Membrana/genética , Proteínas Quimiotáticas Aceptoras de Metil , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais
16.
Helicobacter ; 14(1): 72-9, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19191900

RESUMO

BACKGROUND: Gastrointestinal infections with pathogenic Helicobacter species are commonly treated with combination therapies, which often include amoxicillin. Although this treatment is effective for eradication of Helicobacter pylori, the few existing reports are less clear about antibiotic susceptibility of other Helicobacter species. In this study we have determined the susceptibility of gastric and enterohepatic Helicobacter species to amoxicillin, and have investigated the mechanism of amoxicillin resistance in Helicobacter hepaticus. MATERIALS AND METHODS: The minimal inhibitory concentration (MIC) of antimicrobial compounds was determined by E-test and agar/broth dilution assays. The hefA gene of H. hepaticus was inactivated by insertion of a chloramphenicol resistance gene. Transcription was measured by quantitative real-time polymerase chain reaction. RESULTS: Three gastric Helicobacter species (H. pylori, H. mustelae, and H. acinonychis) were susceptible to amoxicillin (MIC < 0.25 mg/L). In contrast, three enterohepatic Helicobacter species (H. rappini, H. bilis, and H. hepaticus) were resistant to amoxicillin (MIC of 8, 16, and 6-64 mg/L, respectively). There was no detectable beta-lactamase activity in H. hepaticus, and inhibition of beta-lactamases did not change the MIC of amoxicillin of H. hepaticus. A H. hepaticus hefA (hh0224) mutant, encoding a TolC-component of a putative efflux system, resulted in loss of amoxicillin resistance (MIC 0.25 mg/L), and also resulted in increased sensitivity to bile acids. Finally, transcription of the hefA gene was not responsive to amoxicillin, but induced by bile acids. CONCLUSIONS: Rodents are frequently colonized by a variety of enterohepatic Helicobacter species, and this may affect their global health status and intestinal inflammatory responses. Animal facilities should have treatment strategies for Helicobacter infections, and hence resistance of enterohepatic Helicobacter species to amoxicillin should be considered when designing eradication programs.


Assuntos
Amoxicilina/farmacologia , Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Helicobacter hepaticus/genética , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Animais , Regulação Bacteriana da Expressão Gênica , Helicobacter hepaticus/efeitos dos fármacos , Helicobacter hepaticus/metabolismo , Humanos , Testes de Sensibilidade Microbiana , Mutação
17.
J Bacteriol ; 190(19): 6398-408, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18689480

RESUMO

The enterohepatic Helicobacter species Helicobacter hepaticus colonizes the murine intestinal and hepatobiliary tract and is associated with chronic intestinal inflammation, gall stone formation, hepatitis, and hepatocellular carcinoma. Thus far, the role of H. hepaticus motility and flagella in intestinal colonization is unknown. In other, closely related bacteria, late flagellar genes are mainly regulated by the sigma factor FliA (sigma(28)). We investigated the function of the H. hepaticus FliA in gene regulation, flagellar biosynthesis, motility, and murine colonization. Competitive microarray analysis of the wild type versus an isogenic fliA mutant revealed that 11 genes were significantly more highly expressed in wild-type bacteria and 2 genes were significantly more highly expressed in the fliA mutant. Most of these were flagellar genes, but four novel FliA-regulated genes of unknown function were identified. H. hepaticus possesses two identical copies of the gene encoding the FliA-dependent major flagellin subunit FlaA (open reading frames HH1364 and HH1653). We characterized the phenotypes of mutants in which fliA or one or both copies of the flaA gene were knocked out. flaA_1 flaA_2 double mutants and fliA mutants did not synthesize detectable amounts of FlaA and possessed severely truncated flagella. Also, both mutants were nonmotile and unable to colonize mice. Mutants with either flaA gene knocked out produced flagella morphologically similar to those of wild-type bacteria and expressed FlaA and FlaB. flaA_1 mutants which had flagella but displayed reduced motility did not colonize mice, indicating that motility is required for intestinal colonization by H. hepaticus and that the presence of flagella alone is not sufficient.


Assuntos
Proteínas de Bactérias/fisiologia , Infecções por Helicobacter/microbiologia , Helicobacter hepaticus/fisiologia , Fator sigma/fisiologia , Animais , Proteínas de Bactérias/genética , Sequência de Bases , Western Blotting , Flagelina/genética , Helicobacter hepaticus/genética , Helicobacter hepaticus/ultraestrutura , Interações Hospedeiro-Patógeno , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica de Transmissão , Dados de Sequência Molecular , Mutação , Análise de Sequência com Séries de Oligonucleotídeos , Regiões Promotoras Genéticas/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência do Ácido Nucleico , Fator sigma/genética
18.
Microbes Infect ; 10(7): 726-33, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18538610

RESUMO

Helicobacter hepaticus strain 3B1 (H. hepaticus) contains a genomic island of approximately 71 kb, HHGI1, with some of the common features shared among known bacterial pathogenicity islands. In this study, we characterized the pathogenic potential of HHGI1 by infecting B6.129-IL10 tm1Cgn (IL10-/-) mice with an isogenic mutant (namely HhPAId1) lacking 19 predicted genes within HHGI1. In contrast to H. hepaticus (P<0.001), HhPAId1 did not cause typhlocolitis and hyperplasia in IL10-/- mice. Colonization levels of HhPAId1 were significantly higher in the cecum (P<0.007) and similar in the colon (P=0.27) when compared to H. hepaticus by 13 or 16 weeks post inoculation (WPI). The magnitude of the Th1-associated IgG2c response against HhPAId1 was less than that against H. hepaticus (P<0.004). There was no significant difference in Th2-associated IgG1 responses against these two strains. Cecal and colonic mRNA levels of proinflammatory cytokines IFN-gamma, TNF-alpha and IL-17a in the HhPAId1-infected mice were significantly lower than those in the H. hepaticus-infected mice (P<0.05) at 13 WPI. These results demonstrate that genes in the HHGI1 contribute to the pathogenicity of H. hepaticus, at least in part via up-regulation of proinflammatory mediators IFN-gamma, TNF-alpha and IL-17a.


Assuntos
Colite/microbiologia , Ilhas Genômicas , Infecções por Helicobacter/microbiologia , Helicobacter hepaticus/patogenicidade , Animais , Anticorpos Antibacterianos/sangue , Ceco/microbiologia , Colo/microbiologia , Citocinas/biossíntese , Deleção de Genes , Genes Bacterianos , Helicobacter hepaticus/genética , Imunoglobulina G/sangue , Interleucina-10/deficiência , Mucosa Intestinal/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Virulência
19.
Microb Pathog ; 45(1): 18-24, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18486436

RESUMO

Urease activity contributes to bacterial survival in the acidic environment of the stomach and is essential for persistent infection by known gastric helicobacters such as the human pathogen Helicobacter pylori. Several enterohepatic Helicobacter species (EHS) that primarily infect the less acidic intestine also have very active urease enzymes. The importance of urease and its contribution to pathogenesis for these EHS are poorly understood. In this study, we generated a urease-deficient, isogenic mutant (HhureNT9) of Helicobacter hepaticus 3B1 (Hh 3B1), an EHS that possesses a urease gene cluster similar to that of H. pylori. Lack of urease activity did not affect the level of cecal colonization by HhureNT9 compared to Hh 3B1 in male A/JCr mice (P=0.48) at 4 months post-inoculation (MPI). In contrast, there was no HhureNT9 detected in the livers of any infected mice, whereas all livers from the Hh 3B1-infected mice were PCR-positive for Hh 3B1. The mice infected with HhureNT9 developed significantly less severe hepatitis (P=0.017) and also produced significantly lower hepatic mRNA levels of proinflammatory cytokines IFN-gamma (P=0.0007) and TNF-alpha (P<0.0001) compared to the Hh 3B1-infected mice. The Hh 3B1-infected mice developed significantly higher total IgG, Th1-associated IgG2a and Th2-associated IgG1 responses to infection. These results indicate that H. hepaticus urease activity plays a crucial role in hepatic disease but is not required for cecal colonization by H. hepaticus.


Assuntos
Proteínas de Bactérias/metabolismo , Infecções por Helicobacter/microbiologia , Helicobacter hepaticus/enzimologia , Hepatite/microbiologia , Intestinos/microbiologia , Urease/metabolismo , Animais , Anticorpos Antibacterianos/sangue , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Citocinas/genética , Citocinas/imunologia , Helicobacter/enzimologia , Infecções por Helicobacter/imunologia , Infecções por Helicobacter/patologia , Helicobacter hepaticus/genética , Helicobacter hepaticus/imunologia , Helicobacter hepaticus/patogenicidade , Hepatite/imunologia , Hepatite/patologia , Humanos , Imunoglobulina G/sangue , Intestinos/imunologia , Intestinos/patologia , Masculino , Camundongos , Camundongos Endogâmicos A , Transcrição Gênica , Urease/genética , Urease/imunologia
20.
Nan Fang Yi Ke Da Xue Xue Bao ; 28(5): 843-5, 2008 May.
Artigo em Chinês | MEDLINE | ID: mdl-18504216

RESUMO

Helicobacter hepaticus is nongastric helicobacter that can reside in the hepatobiliary and intestinal systems of many animal hosts, leading to proliferative hepatitis, hepatocellular carcinoma, typhlitis, and colonitis. In this study, the intestinal mucosa was isolated from BALB/c mice to prepare tissue homogenate and spread onto selective C jejuni blood agar plates for incubation in the presence of trimethoprim, vancomycin, and polymyxin at 37 degrees Celsius; under microaerobic conditions in vented jars containing 5% O2, 10%CO2, and 85% N2. The bacteria were identified morphologically and biochemically. Gene sequence analysis of the 16s rRNA confirmed the presence of Helicobacter hepaticus, and the success in isolating this bacteria may have significant implications for studies of nongastric helicobacter.


Assuntos
Helicobacter hepaticus/isolamento & purificação , Intestinos/microbiologia , Animais , China , DNA Bacteriano/análise , DNA Bacteriano/genética , Helicobacter hepaticus/genética , Helicobacter hepaticus/ultraestrutura , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Varredura , RNA Ribossômico 16S/genética
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