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1.
Helicobacter ; 27(3): e12891, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35384141

RESUMO

For decades, traditional in vitro and in vivo models used for the study of Helicobacter pylori infection have relied heavily on the use of gastric cancer cell lines and rodents. Major challenges faced by these methods have been the inability to study cancer initiation in already cancerous cell lines, and the difficulty in translating results obtained in animal models due to genetic differences. These challenges have prevented a thorough understanding of the pathogenesis of disease and slowed the development of cancer therapies and a suitable vaccine against the pathogen. In recent years, the development of gastric organoids has provided great advantages over the traditional in vivo and in vitro models due to their similarities to the human stomach in vivo, their ease of use, and the capacity for long-term culture. This review discusses the advantages and limitations of existing in vivo and in vitro models of H. pylori infection, and how gastric organoids have been applied to study H. pylori pathogenesis, with a focus on how the pathogen interacts with the gastric epithelium, inflammatory processes, epithelial repair, and cancer initiation. The potential applications of organoids to address more complex questions on the role of hormones, vaccine-induced immunity are also discussed.


Assuntos
Infecções por Helicobacter , Helicobacter pylori , Neoplasias Gástricas , Animais , Mucosa Gástrica/patologia , Infecções por Helicobacter/patologia , Helicobacter pylori/metabolismo , Humanos , Inflamação , Organoides , Estômago/patologia , Neoplasias Gástricas/metabolismo
2.
Int J Mol Sci ; 23(5)2022 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-35269931

RESUMO

Gastric cancer represents a significant disease burden worldwide. The factors that initiate cancer are not well understood. Chronic inflammation such as that triggered by H. pylori infection is the most significant cause of gastric cancer. In recent years, organoid cultures developed from human and animal adult stem cells have facilitated great advances in our understanding of gastric homeostasis. Organoid models are now being exploited to investigate the role of host genetics and bacterial factors on proliferation and DNA damage in gastric stem cells. The impact of a chronic inflammatory state on gastric stem cells and the stroma has been less well addressed. This review discusses what we have learned from the use of organoid models to investigate cancer initiation, and highlights questions on the contribution of the microbiota, chronic inflammatory milieu, and stromal cells that can now be addressed by more complex coculture models.


Assuntos
Infecções por Helicobacter , Helicobacter pylori , Neoplasias Gástricas , Animais , Mucosa Gástrica/microbiologia , Infecções por Helicobacter/microbiologia , Homeostase , Inflamação/complicações , Organoides , Neoplasias Gástricas/genética
3.
J Biol Chem ; 293(14): 5079-5089, 2018 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-29449370

RESUMO

In response to the widespread emergence of antibiotic-resistant microbes, new therapeutic agents are required for many human pathogens. A non-mammalian polysaccharide, poly-N-acetyl-d-glucosamine (PNAG), is produced by bacteria, fungi, and protozoan parasites. Antibodies that bind to PNAG and its deacetylated form (dPNAG) exhibit promising in vitro and in vivo activities against many microbes. A human IgG1 mAb (F598) that binds both PNAG and dPNAG has opsonic and protective activities against multiple microbial pathogens and is undergoing preclinical and clinical assessments as a broad-spectrum antimicrobial therapy. Here, to understand how F598 targets PNAG, we determined crystal structures of the unliganded F598 antigen-binding fragment (Fab) and its complexes with N-acetyl-d-glucosamine (GlcNAc) and a PNAG oligosaccharide. We found that F598 recognizes PNAG through a large groove-shaped binding site that traverses the entire light- and heavy-chain interface and accommodates at least five GlcNAc residues. The Fab-GlcNAc complex revealed a deep binding pocket in which the monosaccharide and a core GlcNAc of the oligosaccharide were almost identically positioned, suggesting an anchored binding mechanism of PNAG by F598. The Fab used in our structural analyses retained binding to PNAG on the surface of an antibiotic-resistant, biofilm-forming strain of Staphylococcus aureus Additionally, a model of intact F598 binding to two pentasaccharide epitopes indicates that the Fab arms can span at least 40 GlcNAc residues on an extended PNAG chain. Our findings unravel the structural basis for F598 binding to PNAG on microbial surfaces and biofilms.


Assuntos
Anticorpos Monoclonais/imunologia , Imunoglobulina G/imunologia , Polissacarídeos Bacterianos/imunologia , Anticorpos Monoclonais/química , Biofilmes , Configuração de Carboidratos , Cristalografia por Raios X , Humanos , Fragmentos Fab das Imunoglobulinas/química , Fragmentos Fab das Imunoglobulinas/imunologia , Imunoglobulina G/química , Modelos Moleculares , Polissacarídeos Bacterianos/química , Conformação Proteica , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/imunologia , Staphylococcus aureus/fisiologia
4.
Adv Exp Med Biol ; 1149: 257-275, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31016627

RESUMO

Helicobacter pylori is a highly-adapted gastrointestinal pathogen of humans and the immunology of this chronic infection is extremely complex. Despite the availability of antibiotic therapy, the global incidence of H. pylori infection remains high, particularly in low to middle-income nations. Failure of therapy and the spread of antibiotic resistance among the bacteria are significant problems and provide impetus for the development of new therapies and vaccines to treat or prevent gastric ulcer, and gastric carcinoma. The expansion of knowledge on gastric conventional and regulatory T cell responses, and the role of TH17 in chronic gastritis from studies in mouse models and patients have provided valuable insights into how gastritis is initiated and maintained. The development of human challenge models for testing candidate vaccines has meant a unique opportunity to study acute infection, but the field of vaccine development has not progressed as rapidly as anticipated. One clear lesson learned from previous studies is that we need a better understanding of the immune suppressive mechanisms in vivo to be able to design vaccine strategies. There is still an urgent need to identify practical surrogate markers of protection that could be deployed in future field vaccine trials. Important developments in our understanding of the chronic inflammatory response, progress and problems arising from human studies, and an outlook for the future of clinical vaccine trials will be discussed.


Assuntos
Vacinas Bacterianas , Gastrite , Infecções por Helicobacter , Helicobacter pylori , Animais , Vacinas Bacterianas/imunologia , Gastrite/microbiologia , Gastrite/prevenção & controle , Infecções por Helicobacter/imunologia , Infecções por Helicobacter/prevenção & controle , Helicobacter pylori/imunologia , Humanos
5.
Immunology ; 150(2): 172-183, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27676456

RESUMO

There is a current lack of effective mucosal vaccines against major gastroenteric pathogens and particularly against Helicobacter pylori, which causes a chronic infection that can lead to peptic ulcers and gastric cancer in a subpopulation of infected individuals. Mucosal CD4+ T-cell responses have been shown to be essential for vaccine-induced protection against H. pylori infection. The current study addresses the influence of the adjuvant and site of mucosal immunization on early CD4+ T-cell priming to H. pylori antigens. The vaccine formulation consisted of H. pylori lysate antigens and mucosal adjuvants, cholera toxin (CT) or a detoxified double-mutant heat-labile enterotoxin from Escherichia coli (dmLT), which were administered by either the sublingual or intragastric route. We report that in vitro, adjuvants CT and dmLT induce up-regulation of pro-inflammatory gene expression in purified dendritic cells and enhance the H. pylori-specific CD4+ T-cell response including interleukin-17A (IL-17A), interferon-γ (IFN-γ) and tumour necrosis factor-α (TNF-α) secretion. In vivo, sublingual immunization led to an increased frequency of IL-17A+ , IFN-γ+ and TNF-α+ secreting CD4+ T cells in the cervical lymph nodes compared with in the mesenteric lymph nodes after intragastric immunization. Subsequently, IL-17A+ cells were visualized in the stomach of sublingually immunized and challenged mice. In summary, our results suggest that addition of an adjuvant to the vaccine clearly activated dendritic cells, which in turn, enhanced CD4+ T-cell cytokines IL-17A, IFN-γ and TNF-α responses, particularly in the cervical lymph nodes after sublingual vaccination.


Assuntos
Antígenos de Bactérias/imunologia , Vacinas Bacterianas/imunologia , Linfócitos T CD4-Positivos/imunologia , Células Dendríticas/imunologia , Infecções por Helicobacter/imunologia , Helicobacter pylori/imunologia , Imunidade nas Mucosas , Adjuvantes Imunológicos/administração & dosagem , Administração Sublingual , Animais , Toxinas Bacterianas/administração & dosagem , Células Cultivadas , Toxina da Cólera/administração & dosagem , Citocinas/metabolismo , Enterotoxinas/administração & dosagem , Proteínas de Escherichia coli/administração & dosagem , Feminino , Mediadores da Inflamação/metabolismo , Intubação Gastrointestinal , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL
6.
Biology (Basel) ; 13(1)2024 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-38248478

RESUMO

As research on parasitic helminths has entered the post-genomic era, research efforts have turned to deciphering the function of genes in the public databases of genome sequences. It is hoped that, by understanding the role of parasite genes in maintaining their parasitic lifestyle, critical insights can be gained to develop new intervention and control strategies. Methods to manipulate and transform parasitic worms are now developed to a point where it has become possible to gain a comprehensive understanding of the molecular mechanisms underlying host-parasite interplay, and here, we summarise and discuss the advances that have been made in schistosome transgenesis over the past 25 years. The ability to genetically manipulate schistosomes holds promise in finding new ways to control schistosomiasis, which ultimately may lead to the eradication of this debilitating disease.

7.
Sci Rep ; 14(1): 8587, 2024 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-38615147

RESUMO

Helicobacter pylori infects approximately half the human population and has an unusual infective niche of the human stomach. Helicobacter pylori is a major cause of gastritis and has been classified as a group 1 carcinogen by the WHO. Treatment involves triple or quadruple antibiotic therapy, but antibiotic resistance is becoming increasingly prevalent. Helicobacter pylori expresses certain blood group related antigens (Lewis system) as a part of its lipopolysaccharide (LPS), which is thought to assist in immune evasion. Additionally, H. pylori LPS participates in adhesion to host cells alongside several adhesion proteins. This study profiled the carbohydrates of H. pylori reference strains (SS1 and 26695) using monoclonal antibodies (mAbs) and lectins, identifying interactions between two carbohydrate-targeting mAbs and multiple lectins. Atomic force microscopy (AFM) scans were used to probe lectin and antibody interactions with the bacterial surfaces. The selected mAb and lectins displayed an increased adhesive force over the surface of the curved H. pylori rods. Furthermore, this study demonstrates the ability of anti-carbohydrate antibodies to reduce the adhesion of H. pylori 26695 to human gastric adenocarcinoma cells via AFM. Targeting bacterial carbohydrates to disrupt crucial adhesion and immune evasion mechanisms represents a promising strategy for combating H. pylori infection.


Assuntos
Antígenos de Grupos Sanguíneos , Infecções por Helicobacter , Helicobacter pylori , Humanos , Lipopolissacarídeos , Polissacarídeos , Anticorpos Monoclonais , Lectinas
8.
Vaccines (Basel) ; 12(4)2024 Apr 12.
Artigo em Inglês | MEDLINE | ID: mdl-38675791

RESUMO

Nanoparticles show great promise as a platform for developing vaccines for the prevention of infectious disease. We have been investigating a method whereby nanocapsules can be formulated from protein, such that the final capsules contain only the cross-linked protein itself. Such nanocapsules are made using a silica templating system and can be customised in terms of size and porosity. Here we compare the construction and characteristics of nanocapsules from four different proteins: one a model protein (ovalbumin) and three from infectious disease pathogens, namely the influenza virus, Helicobacter pylori and HIV. Two of the nanocapsules were assessed further. We confirm that nanocapsules constructed from the urease A subunit of H. pylori can reduce subsequent infection in a vaccinated mouse model. Further, we show that capsules constructed from the HIV gp120 protein can be taken up by dendritic cells in tissue culture and can be recognised by antibodies raised against the virus. These results point to the utility of this method in constructing protein-only nanocapsules from proteins of varying sizes and isoelectric points.

9.
Infect Immun ; 81(2): 470-7, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23208607

RESUMO

Bacterial infections are a common and serious complication of type 2 diabetes (T2D). The prevalence of melioidosis, an emerging tropical infection caused by the Gram-negative bacterium Burkholderia pseudomallei, is increased in people with T2D. This is the first study to compare murine models of T2D and melioidosis. Susceptibility and disease progression following infection with B. pseudomallei were compared in our diet-induced polygenic mouse model and a leptin receptor-deficient monogenic model of T2D. The metabolic profile of mice with diet-induced diabetes, including body weight, blood glucose, cholesterol, triglycerides, insulin resistance, and baseline levels of inflammation, closely resembled that of clinical T2D. Following subcutaneous infection with B. pseudomallei, bacterial loads at 24 and 72 h postinfection in the blood, spleen, liver, lungs, and subcutaneous adipose tissue (SAT) at the site of infection were compared in parallel with the expression of inflammatory cytokines and tissue histology. As early as 24 h postinfection, the expression of inflammatory (interleukin-1ß [IL-1ß], tumor necrosis factor alpha [TNF-α], and IL-6) and T(H)1 (IL-12 and gamma interferon [IFN-γ]) cytokines was impaired in diabetic mice compared to nondiabetic littermates. Early differences in cytokine expression were associated with excessive infiltration of polymorphonuclear neutrophils (PMN) in diabetic mice compared to nondiabetic littermates. This was accompanied by bacteremia, hematogenous dissemination of bacteria to the lungs, and uncontrolled bacterial growth in the spleens of diabetic mice by 72 h postinfection. The findings from our novel model of T2D and melioidosis comorbidity support the role of impaired early immune pathways in the increased susceptibility of individuals with T2D to bacterial infections.


Assuntos
Diabetes Mellitus Tipo 2/imunologia , Diabetes Mellitus Tipo 2/microbiologia , Melioidose/imunologia , Melioidose/microbiologia , Animais , Burkholderia pseudomallei/imunologia , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Inflamação/imunologia , Inflamação/microbiologia , Interferon gama/imunologia , Interleucina-12/imunologia , Interleucina-1beta/imunologia , Interleucina-6/imunologia , Masculino , Melioidose/metabolismo , Metaboloma/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/imunologia , Neutrófilos/microbiologia , Fator de Necrose Tumoral alfa/imunologia
10.
Infect Immun ; 81(10): 3880-93, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23918779

RESUMO

The natural immune response to Helicobacter pylori neither clears infection nor prevents reinfection. However, the ability of secretory antibodies to influence the course of H. pylori infection has not been determined. We compared the natural progression of H. pylori infection in wild-type C57BL/6 mice with that in mice lacking the polymeric immunoglobulin receptor (pIgR) that is essential for the secretion of polymeric antibody across mucosal surfaces. H. pylori SS1-infected wild-type and pIgR knockout (KO) mice were sampled longitudinally for gastrointestinal bacterial load, antibody response, and histological changes. The gastric bacterial loads of wild-type and pIgR KO mice remained constant and comparable at up to 3 months postinfection (mpi) despite SS1-reactive secretory IgA in the intestinal contents of wild-type mice at that time. Conversely, abundant duodenal colonization of pIgR KO animals contrasted with the near-total eradication of H. pylori from the intestine of wild-type animals by 3 mpi. H. pylori was cultured only from the duodenum of those animals in which colonization in the distal gastric antrum was of sufficient density for immunohistological detection. By 6 mpi, the gastric load of H. pylori in wild-type mice was significantly lower than in pIgR KO animals. While there was no corresponding difference between the two mouse strains in gastric pathology results at 6 mpi, reductions in gastric bacterial load correlated with increased gastric inflammation together with an intestinal secretory antibody response in wild-type mice. Together, these results suggest that naturally produced secretory antibodies can modulate the progress of H. pylori infection, particularly in the duodenum.


Assuntos
Anticorpos Antibacterianos/metabolismo , Infecções por Helicobacter/imunologia , Helicobacter pylori , Imunidade nas Mucosas/fisiologia , Mucosa Intestinal/metabolismo , Animais , Western Blotting , Regulação da Expressão Gênica/imunologia , Imunoglobulina A/sangue , Imunoglobulina A/metabolismo , Imunoglobulina G/sangue , Imunoglobulina G/metabolismo , Mucosa Intestinal/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Imunoglobulina Polimérica/genética , Receptores de Imunoglobulina Polimérica/metabolismo
11.
J Immunol ; 184(5): 2237-42, 2010 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-20164434

RESUMO

Given the central role of intestinal dendritic cells (DCs) in the regulation of gut immune responses, it is not surprising that several bacterial pathogens have evolved strategies to prevent or bypass recognition by DCs. In this article, we will review recent findings on the interaction between intestinal DCs and prototypical bacterial pathogens, such as Salmonella, Yersinia, or Helicobacter. We will discuss the different approaches with which these pathogens seek to evade DC recognition and subsequent T cell activation. These diverse strategies span to include mounting irrelevant immune responses, inhibition of Ag presentation by DCs, and stretch as far as to manipulate the Th1/Th2 balance of CD4(+) T cells in the bacteria's favor.


Assuntos
Células Dendríticas/imunologia , Helicobacter pylori/imunologia , Intestinos/imunologia , Salmonella typhimurium/imunologia , Yersinia enterocolitica/imunologia , Animais , Infecções Bacterianas/imunologia , Infecções Bacterianas/microbiologia , Helicobacter pylori/fisiologia , Interações Hospedeiro-Patógeno/imunologia , Humanos , Intestinos/microbiologia , Modelos Imunológicos , Salmonella typhimurium/fisiologia , Yersinia enterocolitica/fisiologia
12.
Clin Exp Pharmacol Physiol ; 39(3): 218-26, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22211630

RESUMO

  NADPH oxidases (Nox) are reactive oxygen species (ROS)-generating enzymes that play important physiological roles in host defence and redox signalling. However, Nox activity is upregulated in the vascular wall during atherosclerosis and contributes to plaque formation by promoting oxidative stress and inflammation.   The bacterium Chlamydia pneumoniae has been detected in vascular smooth muscle cells (VSMC) of human atheroma. We hypothesized that C. pneumoniae infection of VSMC causes Nox activation, which initially limits infection but ultimately causes oxidative stress, activation of pro-inflammatory pathways and an atherogenic phenotype.   Chlamydia pneumoniae infection of mouse cultured VSMC significantly increased ROS production by twofold but did not upregulate mRNA expression of Nox1 or Nox4. Chlamydia pneumoniae did increase Nox2 mRNA levels significantly by threefold, but this did not translate to elevated Nox2 protein expression.   The Nox inhibitor gp91ds-tat had no effect on C. pneumoniae-induced ROS production. In contrast, apocynin significantly reduced ROS levels by 75% in C. pneumoniae-infected VSMC, an effect most likely attributable to its direct anti-oxidant action.   Although apocynin had no effect on C. pneumoniae-induced expression of inflammatory markers, bacteria recovered from apocynin-treated VSMC displayed a higher degree of infectivity in HEp-2 cells.   In conclusion, C. pneumoniae infection increases ROS production in VSMC independently of Nox activity. Although elevated ROS production appears to serve a protective role by limiting the spread of infection, we speculate that this response will be detrimental over the long term by causing oxidative stress and a smouldering inflammatory response by maintaining C. pneumoniae persistence within the cell.


Assuntos
Infecções por Chlamydophila/genética , Chlamydophila pneumoniae/patogenicidade , Imunofenotipagem , Mediadores da Inflamação/fisiologia , Miócitos de Músculo Liso/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Infecções por Chlamydophila/metabolismo , Infecções por Chlamydophila/patologia , Humanos , Camundongos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/microbiologia , Miócitos de Músculo Liso/microbiologia
13.
Front Immunol ; 13: 868225, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35634347

RESUMO

Helicobacter pylori is an important human pathogen that infects half the human population and can lead to significant clinical outcomes such as acute and chronic gastritis, duodenal ulcer, and gastric adenocarcinoma. To establish infection, H. pylori employs several mechanisms to overcome the innate and adaptive immune systems. H. pylori can modulate interleukin (IL) secretion and innate immune cell function by the action of several virulence factors such as VacA, CagA and the type IV secretion system. Additionally, H. pylori can modulate local dendritic cells (DC) negatively impacting the function of these cells, reducing the secretion of immune signaling molecules, and influencing the differentiation of CD4+ T helper cells causing a bias to Th1 type cells. Furthermore, the lipopolysaccharide (LPS) of H. pylori displays a high degree of phase variation and contains human blood group carbohydrate determinants such as the Lewis system antigens, which are proposed to be involved in molecular mimicry of the host. Lastly, the H. pylori group of outer membrane proteins such as BabA play an important role in attachment and interaction with host Lewis and other carbohydrate antigens. This review examines the various mechanisms that H. pylori utilises to evade the innate immune system as well as discussing how the structure of the H. pylori LPS plays a role in immune evasion.


Assuntos
Infecções por Helicobacter , Helicobacter pylori , Humanos , Evasão da Resposta Imune , Lipopolissacarídeos , Fatores de Virulência/metabolismo
14.
Vaccine ; 40(40): 5769-5780, 2022 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-36064671

RESUMO

Vaccines are very effective in providing protection against many infectious diseases. However, it has proven difficult to develop highly efficacious vaccines against some pathogens and so there is a continuing need to improve vaccine technologies. The first successful and widely used vaccines were based on attenuated pathogens (e.g., laboratory passaged Pasteurella multocida to vaccinate against fowl cholera) or closely related non-pathogenic organisms (e.g., cowpox to vaccinate against smallpox). Subsequently, live vaccines, either attenuated pathogens or non-pathogenic microorganisms modified to deliver heterologous antigens, have been successfully used to induce protective immune responses against many pathogens. Unlike conventional killed and subunit vaccines, live vaccines can deliver antigens to mucosal surfaces in a similar manner and context as the natural infection and hence can often produce a more appropriate and protective immune response. Despite these advantages, there is still a need to improve the immunogenicity of some live vaccines. The efficacy of injectable killed and subunit vaccines is usually enhanced using adjuvants such mineral salts, oils, and saponin, but such adjuvants cannot be used with live vaccines. Instead, live vaccines can be engineered to produce immunomodulatory molecules that can stimulate the immune system to induce more robust and long-lasting adaptive immune responses. This review focuses on research that has been undertaken to engineer live vaccines to produce immunomodulatory molecules that act as adjuvants to increase immunogenicity. Adjuvant strategies with varying mechanisms of action (inflammatory, antibody-mediated, cell-mediated) and delivery modes (oral, intramuscular, intranasal) have been investigated, with varying degrees of success. The goal of such research is to define adjuvant strategies that can be adapted to enhance live vaccine efficacy by triggering strong innate and adaptive immune responses and produce vaccines against a wider range of pathogens.


Assuntos
Infecções por Pasteurella , Pasteurella multocida , Vacinas , Adjuvantes Imunológicos , Humanos , Vacinas Atenuadas , Vacinas de Subunidades Antigênicas
15.
Nanoscale ; 14(48): 17940-17954, 2022 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-36349848

RESUMO

Carbon nanodots (C-dots) have attracted much attention for their use in the fields of bioimaging, drug delivery, and sensing due to their excellent fluorescent and photoluminescent properties, photostability, biocompatibility, and amenability to surface modification. Herein, we report a nanocomposite formulation of C-dots (<5 nm) encapsulated in lipid-based lyotropic liquid crystalline nanoparticles (∼250 nm) via either passive diffusion or electrostatic mechanisms. The physicochemical properties of the nanocomposite formulation including particle size, surface charge, internal cubic nanostructures, and pH-dependent fluorescent properties were characterised. Upon loading of C-dots into lipid nanoparticles, the highly ordered inverse bicontinuous cubic mesophase existed in the internal phase of the nanoparticles, demonstrated by synchrotron small angle X-ray scattering, molecular dynamic simulation and cryogenic transmission electron microscopy. The pH-dependent fluorescent property of the C-dots was modified via electrostatic interaction between the C-dots and cationic lipid nanoparticles, which further enhanced the brightness of C-dots through self-quenching prevention. The cytotoxicity and cellular uptake efficiency of the developed nanocomposites were also examined in an epithelial gastric adenocarcinoma cell line (AGS) and a macrophage cell line (stimulated THP-1). Compared to free C-dots, the uptake and cell imaging potential of the C-dot nanocomposites was significantly improved, by several orders of magnitude as demonstrated by cytoplasmic fluorescent intensities using confocal microscopy. Loading C-dots into mesoporous lipid nanocarriers presents a new way of modifying C-dot physicochemical and fluorescent properties, alternative to direct chemical surface modification, and advances the bioimaging potential of C-dots by enhancing cellular uptake efficiency and converging C-dot light emission.


Assuntos
Carbono , Nanocompostos , Carbono/química , Sistemas de Liberação de Medicamentos/métodos , Tamanho da Partícula , Lipídeos
16.
Infect Immun ; 79(10): 4186-92, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-21807907

RESUMO

While gastric adenocarcinoma is the most serious consequence of Helicobacter pylori infection, not all infected persons develop this pathology. Individuals most at risk of this cancer are those in whom the bacteria colonize the acid-secreting region of the stomach and subsequently develop severe inflammation in the gastric corpus. It has been reported anecdotally that male mice become infected with greater numbers of H. pylori bacteria than female mice. While investigating this phenomenon, we found that increased H. pylori infection densities in male mice were not related to antibody production, and this phenomenon was not normalized by gonadectomy. However, the gastric pH in male 129/Sv mice was significantly elevated compared with that in female mice. Differences in colonization were evident within 1 day postinfection and significantly arose due to colonization of the gastric corpus region in male mice. This provided a potential model for comparing the effect of corpus colonization on the development of gastritis. This was explored using two models of H. pylori-induced inflammation, namely, 2-month infections of Muc1(-/-) mice and 6-month infections of wild-type 129/Sv mice. While H. pylori infection of female mice induced a severe, corpus-predominant atrophic gastritis, to our surprise, male mice developed minimal inflammation despite being colonized with significantly more H. pylori bacteria than female controls. Thus, colonization of the gastric corpus in male mice was associated with a loss of inflammation in that region. The suppression of inflammation concomitant with infection of the gastric corpus in male mice demonstrates a powerful localized suppression of inflammation induced at sites of H. pylori colonization.


Assuntos
Mucosa Gástrica/patologia , Gastrite Atrófica/patologia , Infecções por Helicobacter/complicações , Helicobacter pylori/patogenicidade , Inflamação/patologia , Estômago/patologia , Animais , Modelos Animais de Doenças , Feminino , Mucosa Gástrica/microbiologia , Gastrite Atrófica/imunologia , Gastrite Atrófica/microbiologia , Infecções por Helicobacter/microbiologia , Infecções por Helicobacter/patologia , Helicobacter pylori/isolamento & purificação , Humanos , Concentração de Íons de Hidrogênio , Inflamação/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estômago/microbiologia
17.
Gastroenterology ; 139(6): 2005-17, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20801119

RESUMO

BACKGROUND & AIMS: Trefoil factors (TFFs) regulate mucosal repair and suppress tumor formation in the stomach. Tff1 deficiency results in gastric cancer, whereas Tff2 deficiency increases gastric inflammation. TFF2 expression is frequently lost in gastric neoplasms, but the nature of the silencing mechanism and associated impact on tumorigenesis have not been determined. METHODS: We investigated the epigenetic silencing of TFF2 in gastric biopsy specimens from individuals with Helicobacter pylori-positive gastritis, intestinal metaplasia, gastric cancer, and disease-free controls. TFF2 function and methylation were manipulated in gastric cancer cell lines. The effects of Tff2 deficiency on tumor growth were investigated in the gp130(F/F) mouse model of gastric cancer. RESULTS: In human tissue samples, DNA methylation at the TFF2 promoter began at the time of H pylori infection and increased throughout gastric tumor progression. TFF2 methylation levels were inversely correlated with TFF2 messenger RNA levels and could be used to discriminate between disease-free controls, H pylori-infected, and tumor tissues. Genome demethylation restored TFF2 expression in gastric cancer cell lines, so TFF2 silencing requires methylation. In Tff2-deficient gp130(F/F)/Tff2(-/-) mice, proliferation of mucosal cells and release of T helper cell type-1 (Th-1) 1 cytokines increased, whereas expression of gastric tumor suppressor genes and Th-2 cytokines were reduced, compared with gp130(F/F)controls. The fundus of gp130(F/F)/Tff2(-/-) mice displayed glandular atrophy and metaplasia, indicating accelerated preneoplasia. Experimental H pylori infection in wild-type mice reduced antral expression of Tff2 by increased promoter methylation. CONCLUSIONS: TFF2 negatively regulates preneoplastic progression and subsequent tumor development in the stomach, a role that is subverted by promoter methylation during H pylori infection.


Assuntos
Gastrite , Infecções por Helicobacter/genética , Helicobacter pylori , Mucinas/genética , Proteínas Musculares/genética , Peptídeos/genética , Neoplasias Gástricas , Animais , Atrofia , Linhagem Celular Tumoral , Receptor gp130 de Citocina/genética , Metilação de DNA/fisiologia , Epigenômica , Gastrite/genética , Gastrite/microbiologia , Gastrite/patologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Inativação Gênica/fisiologia , Infecções por Helicobacter/patologia , Humanos , Metaplasia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Transdução de Sinais/fisiologia , Neoplasias Gástricas/genética , Neoplasias Gástricas/microbiologia , Neoplasias Gástricas/patologia , Células Th1/fisiologia , Fator Trefoil-2
18.
Eur J Immunol ; 40(10): 2778-90, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-21038469

RESUMO

Helicobacter pylori is recognised as the chief cause of chronic gastritis, ulcers and gastric cancer in humans. With increased incidence of treatment failure and antibiotic resistance, development of prophylactic or therapeutic vaccination is a desirable alternative. Although the results of vaccination studies in animal models have been promising, studies in human volunteers have revealed problems such as 'post-immunisation gastritis' and comparatively poor responses to vaccine antigens. The focus of this study was to compare the gastric and systemic cellular immune responses induced by recombinant attenuated Salmonella Typhimurium-based vaccination in the C57BL/6 model of H. pylori infection. Analysis of lymphocyte populations in the gastric mucosa, blood, spleen, paragastric LN and MLN revealed that the effects of vaccination were largely confined to the parenchymal stomach rather than lymphoid organs. Vaccine-induced protection was correlated with an augmented local recall response in the gastric mucosa, with increased proportions of CD4(+) T cells, neutrophils and reduced proportions of CD4(+) Treg. CD4(+) T cells isolated from the stomachs of vaccinated mice proliferated ex vivo in response to H. pylori antigen, and secreted Th1 cytokines, particularly IFN-γ. This detailed analysis of local gastric immune responses provides insight into the mechanism of vaccine-induced protection.


Assuntos
Vacinas Bacterianas/imunologia , Gastrite/microbiologia , Gastrite/prevenção & controle , Infecções por Helicobacter/imunologia , Infecções por Helicobacter/prevenção & controle , Helicobacter pylori/imunologia , Vacinação/métodos , Animais , Vacinas Bacterianas/administração & dosagem , Linfócitos T CD4-Positivos/imunologia , Proliferação de Células , Feminino , Citometria de Fluxo , Mucosa Gástrica/imunologia , Mucosa Gástrica/microbiologia , Gastrite/imunologia , Infecções por Helicobacter/microbiologia , Histocitoquímica , Memória Imunológica/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Estatísticas não Paramétricas , Células Th1/imunologia , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/imunologia
19.
Infect Immun ; 78(10): 4363-73, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20643854

RESUMO

Taenia ovis is a cestode parasite infecting primarily sheep as intermediate hosts and dogs as definitive hosts. The first highly effective, recombinant vaccine against a parasitic organism was developed against T. ovis infection in sheep. Three separate host-protective antigens (To16, To18, and To45W) have been cloned from the oncosphere of the parasite. We localize these antigens in the oncosphere by using quantitative immunogold labeling and transmission electron microscopy. The three antigens were uniquely associated with penetration gland cells. The cytoplasm and secretory granules of both penetration gland type 1 and type 2 cells exhibited statistically significant levels of staining for each of the three antigens. The intensity of labeling of the penetration gland type 1 cell was approximately three to five times greater (P < 0.01) compared to the level of staining intensity seen in the penetration gland type 2 cell. In activated oncospheres, secretory blebs were found to contain granules with a structure similar to those observed in the penetration gland cells. The granules within the secretory blebs were shown to stain specifically for the presence of each of the three host-protective antigens. The absence of surface location of the T. ovis antigens suggests that the parasite may not be susceptible to vaccine-induced antibody- and complement-mediated attack until some postoncospheral development has occurred after infection of the intermediate host.


Assuntos
Antígenos de Helmintos/imunologia , Taenia/imunologia , Teníase/prevenção & controle , Vacinas , Animais , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Transmissão , Sensibilidade e Especificidade , Taenia/ultraestrutura , Vacinas/imunologia
20.
Mol Cancer ; 8: 22, 2009 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-19317916

RESUMO

BACKGROUND: Helicobacter pylori is a crucial determining factor in the pathogenesis of benign and neoplastic gastric diseases. Cyclooxygenase-2 (Cox-2) is the inducible key enzyme of arachidonic acid metabolism and is a central mediator in inflammation and cancer. Expression of the Cox-2 gene is up-regulated in the gastric mucosa during H. pylori infection but the pathobiological consequences of this enhanced Cox-2 expression are not yet characterized. The aim of this study was to identify novel genes down-stream of Cox-2 in an in vivo model, thereby identifying potential targets for the study of the role of Cox- 2 in H. pylori pathogenesis and the initiation of pre- cancerous changes. RESULTS: Gene expression profiles in the gastric mucosa of mice treated with a specific Cox-2 inhibitor (NS398) or vehicle were analysed at different time points (6, 13 and 19 wk) after H. pylori infection. H. pylori infection affected the expression of 385 genes over the experimental period, including regulators of gastric physiology, proliferation, apoptosis and mucosal defence. Under conditions of Cox-2 inhibition, 160 target genes were regulated as a result of H. pylori infection. The Cox-2 dependent subset included those influencing gastric physiology (Gastrin, Galr1), epithelial barrier function (Tjp1, connexin45, Aqp5), inflammation (Icam1), apoptosis (Clu) and proliferation (Gdf3, Igf2). Treatment with NS398 alone caused differential expression of 140 genes, 97 of which were unique, indicating that these genes are regulated under conditions of basal Cox-2 expression. CONCLUSION: This study has identified a panel of novel Cox-2 dependent genes influenced under both normal and the inflammatory conditions induced by H. pylori infection. These data provide important new links between Cox-2 and inflammatory processes, epithelial repair and integrity.


Assuntos
Ciclo-Oxigenase 2/metabolismo , Perfilação da Expressão Gênica , Infecções por Helicobacter/genética , Helicobacter pylori , Animais , Células Cultivadas , Ciclo-Oxigenase 2/genética , Dinoprostona/genética , Dinoprostona/metabolismo , Mucosa Gástrica/enzimologia , Mucosa Gástrica/metabolismo , Infecções por Helicobacter/enzimologia , Camundongos , Nitrobenzenos/farmacologia , Sulfonamidas/farmacologia , Regulação para Cima
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