Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 84
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Cell Commun Signal ; 19(1): 108, 2021 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-34742300

RESUMO

BACKGROUND: High temperature requirement A (HtrA) is an active serine protease secreted by the group-I carcinogen Helicobacter pylori (H. pylori). The human cell adhesion protein and tumor suppressor E-cadherin (hCdh1) expressed on the surface of gastric epithelial cells was identified as the first HtrA substrate. HtrA-mediated hCdh1 cleavage and subsequent disruption of intercellular adhesions are considered as important steps in H. pylori pathogenesis. In this study, we performed a proteomic profiling of H. pylori HtrA (HpHtrA) to decipher the complex mechanism of H. pylori interference with the epithelial barrier integrity. RESULTS: Using a proteomic approach we identified human desmoglein-2 (hDsg2), neuropilin-1, ephrin-B2, and semaphorin-4D as novel extracellular HpHtrA substrates and confirmed the well characterized target hCdh1. HpHtrA-mediated hDsg2 cleavage was further analyzed by in vitro cleavage assays using recombinant proteins. In infection experiments, we demonstrated hDsg2 shedding from H. pylori-colonized MKN28 and NCI-N87 cells independently of pathogen-induced matrix-metalloproteases or ADAM10 and ADAM17. CONCLUSIONS: Characterizing the substrate specificity of HpHtrA revealed efficient hDsg2 cleavage underlining the importance of HpHtrA in opening intercellular junctions. Video Abstract.

2.
FEMS Microbiol Lett ; 2021 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-34755852

RESUMO

High Temperature Requirement A (HtrA) was identified as a secreted virulence factor in many pathogenic bacteria, including Listeria monocytogenes. Recently, it was discovered that Helicobacter pylori and Campylobacter jejuni HtrAs can directly cleave the human cell adhesion molecule E-cadherin, which facilitates bacterial transmigration. HtrAs also interact with extracellular matrix (ECM) molecules. However, only a limited number of studies have been carried out in this regard. In the present study, the protease and ECM binding properties of L. monocytogenes HtrA (LmHtrA) were studied using native rLmHtrA, catalytically inactive rLmHtrA(S343A) and rLmHtrA lacking the PDZ domain (∆PDZ) to gain more insights into HtrA-ECM molecule interaction. The results show that 1) native rLmHtrA cleaves fibrinogen, fibronectin, plasminogen, and casein in a time and temperature dependent manner, 2) interaction of rLmHtrA with various host proteins was found in the micromolar to nanomolar range, 3) in the absence of PDZ domain, rLmHtrA exhibits no drastic change in binding affinity towards the host molecules when compared with native rLmHtrA, and 4) the PDZ domain plays an important role in the substrate cleavage as rLmHtrA1-394∆PDZ cleaves the substrates only under certain conditions. The proteolysis of various ECM molecules by rLmHtrA possibly highlights the role of HtrA in L. monocytogenes pathogenesis involving ECM degradation.

3.
Toxins (Basel) ; 13(9)2021 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-34564597

RESUMO

BACKGROUND: Helicobacter pylori (Hp) colonizes the human stomach and can induce gastric cancer and mucosa-associated lymphoid tissue (MALT) lymphoma. Clinical observations suggest a role for the Hp virulence factor cytotoxin-associated gene A (CagA) in pathogenesis. The pathogenic activity of CagA is partly regulated by tyrosine phosphorylation of C-terminal Glu-Pro-Ile-Tyr-Ala (EPIYA) motifs in host cells. However, CagA differs considerably in EPIYA motifs, whose functions have been well characterized in epithelial cells. Since CagA is fragmented in immune cells, different CagA variants may exhibit undetected functions in B cells. METHODS: B cells were infected with Hp isolates and isogenic mutants expressing different CagA EPIYA variants. CagA translocation and tyrosine phosphorylation were investigated by Western blotting. Apoptosis was analyzed by flow cytometry and metabolic activity was detected by an MTT assay. RESULTS: Isogenic CagA EPIYA variants are equally well translocated into B cells, followed by tyrosine phosphorylation and cleavage. B cell apoptosis was induced in a CagA-independent manner. However, variants containing at least one EPIYA-C motif affected metabolic activity independently of phosphorylation or multiplication of EPIYA-C motifs. CONCLUSIONS: The diverse structure of CagA regulates B cell physiology, whereas B cell survival is independent of CagA.

4.
Cell Commun Signal ; 18(1): 160, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-33023610

RESUMO

BACKGROUND: Helicobacter pylori (H. pylori) is a gram-negative bacterium that chronically infects approximately 50% of the world's human population. While in most cases the infection remains asymptomatic, 10% of infected individuals develop gastric pathologies and 1-3% progress to gastric cancer. Although H. pylori induces severe inflammatory responses, the host's immune system fails to clear the pathogen and H. pylori can persist in the human stomach for decades. As suppressor of cytokine signaling (SOCS) proteins are important feedback regulators limiting inflammatory responses, we hypothesized that H. pylori could modulate the host's immune responses by inducing SOCS expression. METHODS: The phenotype of human monocyte-derived DCs (moDCs) infected with H. pylori was analyzed by flow cytometry and multiplex technology. SOCS expression levels were monitored by qPCR and signaling studies were conducted by means of Western blot. For functional studies, RNA interference-based silencing of SOCS1-3 and co-cultures with CD4+ T cells were performed. RESULTS: We show that H. pylori positive gastritis patients express significantly higher SOCS3, but not SOCS1 and SOCS2, levels compared to H. pylori negative patients. Moreover, infection of human moDCs with H. pylori rapidly induces SOCS3 expression, which requires the type IV secretion system (T4SS), release of TNFα, and signaling via the MAP kinase p38, but appears to be independent of TLR2, TLR4, MEK1/2 and STAT proteins. Silencing of SOCS3 expression in moDCs prior to H. pylori infection resulted in increased release of both pro- and anti-inflammatory cytokines, upregulation of PD-L1, and decreased T-cell proliferation. CONCLUSIONS: This study shows that H. pylori induces SOCS3 via an autocrine loop involving the T4SS and TNFα and p38 signaling. Moreover, we demonstrate that high levels of SOCS3 in DCs dampen PD-L1 expression on DCs, which in turn drives T-cell proliferation. Video Abstract.


Assuntos
Sistemas de Secreção Bacterianos , Células Dendríticas/metabolismo , Células Dendríticas/microbiologia , Helicobacter pylori/fisiologia , Proteína 3 Supressora da Sinalização de Citocinas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Antígenos de Bactérias/metabolismo , Antígeno B7-H1/metabolismo , Proteínas de Bactérias/metabolismo , Proliferação de Células , Quimiocinas/metabolismo , Retroalimentação Fisiológica , Infecções por Helicobacter/metabolismo , Humanos , Janus Quinases/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Monócitos/metabolismo , Mutação/genética , Fosforilação , Transdução de Sinais , Receptores Toll-Like/metabolismo
5.
Biochemistry ; 59(39): 3772-3781, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-32936629

RESUMO

Naturally occurring membranolytic antimicrobial peptides (AMPs) are rarely cell-type selective and highly potent at the same time. Template-based peptide design can be used to generate AMPs with improved properties de novo. Following this approach, 18 linear peptides were obtained by computationally morphing the natural AMP Aurein 2.2d2 GLFDIVKKVVGALG into the synthetic model AMP KLLKLLKKLLKLLK. Eleven of the 18 chimeric designs inhibited the growth of Staphylococcus aureus, and six peptides were tested and found to be active against one resistant pathogenic strain or more. One of the peptides was broadly active against bacterial and fungal pathogens without exhibiting toxicity to certain human cell lines. Solution nuclear magnetic resonance and molecular dynamics simulation suggested an oblique-oriented membrane insertion mechanism of this helical de novo peptide. Temperature-resolved circular dichroism spectroscopy pointed to conformational flexibility as an essential feature of cell-type selective AMPs.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Sequência de Aminoácidos , Desenho de Fármacos , Células HEK293 , Humanos , Simulação de Dinâmica Molecular , Conformação Proteica em alfa-Hélice , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/crescimento & desenvolvimento
6.
Microorganisms ; 8(9)2020 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-32878302

RESUMO

Persistent infections with the human pathogen Helicobacter pylori (H. pylori) have been closely associated with the induction and progression of a wide range of gastric disorders, including acute and chronic gastritis, ulceration in the stomach and duodenum, mucosa-associated lymphoid tissue (MALT) lymphoma, and gastric adenocarcinoma. The pathogenesis of H. pylori is determined by a complicated network of manifold mechanisms of pathogen-host interactions, which involves a coordinated interplay of H. pylori pathogenicity and virulence factors with host cells. While these molecular and cellular mechanisms have been intensively investigated to date, the knowledge about outer membrane vesicles (OMVs) derived from H. pylori and their implication in bacterial pathogenesis is not well developed. In this review, we summarize the current knowledge on H. pylori-derived OMVs.

7.
Pathog Dis ; 78(7)2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866262

RESUMO

Persistent infections with the bacterial group-I carcinogen Helicobacter pylori (H. pylori) have been associated with a broad range of gastric disorders, including gastritis, ulceration, gastric cancer or mucosa-associated lymphoid tissue (MALT) lymphoma. Pathogenesis of H. pylori requires a balance between immune tolerance and defense. Although H. pylori induces inflammatory responses, the immune system cannot eliminate the pathogen. The detailed molecular mechanisms of how H. pylori interferes with cells of the immune system, in particular infiltrated B cells, are not well investigated. Previously, it was shown that the bacterial effector and oncoprotein cytotoxin-associated gene A (CagA) is delivered into B cells followed by its tyrosine-phosphorylation. To investigate the functional consequences in B cells colonized by CagA-positive H. pylori, we analyzed the global transcriptome of H. pylori-infected Mec-1 cells by RNA sequencing. We found 889 differentially expressed genes (DEGs) and validated JUN, FOSL2, HSPA1B, SRC, CXCR3, TLR-4, TNF-α, CXCL8, CCL2, CCL4, MHC class I and MHC class II molecules by qPCR, western blot, flow cytometry and ELISA assays. The H. pylori-specific mRNA expression signature reveals a downregulation of inflammation- and migration-associated genes, whereas central signal transduction regulators of cell survival and death are upregulated.


Assuntos
Linfócitos B/imunologia , Linfócitos B/metabolismo , Infecções por Helicobacter/genética , Infecções por Helicobacter/microbiologia , Helicobacter pylori/fisiologia , Interações Hospedeiro-Patógeno/genética , Transcriptoma , Biologia Computacional/métodos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Infecções por Helicobacter/complicações , Infecções por Helicobacter/imunologia , Interações Hospedeiro-Patógeno/imunologia , Humanos , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Linfoma de Zona Marginal Tipo Células B/etiologia , Reprodutibilidade dos Testes , Neoplasias Gástricas/etiologia
8.
Sci Rep ; 10(1): 10563, 2020 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-32601479

RESUMO

Helicobacter pylori (H. pylori) secretes the chaperone and serine protease high temperature requirement A (HtrA) that cleaves gastric epithelial cell surface proteins to disrupt the epithelial integrity and barrier function. First inhibitory lead structures have demonstrated the essential role of HtrA in H. pylori physiology and pathogenesis. Comprehensive drug discovery techniques allowing high-throughput screening are now required to develop effective compounds. Here, we designed a novel fluorescence resonance energy transfer (FRET) peptide derived from a gel-based label-free proteomic approach (direct in-gel profiling of protease specificity) as a valuable substrate for H. pylori HtrA. Since serine proteases are often sensitive to metal ions, we investigated the influence of different divalent ions on the activity of HtrA. We identified Zn++ and Cu++ ions as inhibitors of H. pylori HtrA activity, as monitored by in vitro cleavage experiments using casein or E-cadherin as substrates and in the FRET peptide assay. Putative binding sites for Zn++ and Cu++ were then analyzed in thermal shift and microscale thermophoresis assays. The findings of this study will contribute to the development of novel metal ion-dependent protease inhibitors, which might help to fight bacterial infections.


Assuntos
Proteínas de Bactérias/antagonistas & inibidores , Avaliação Pré-Clínica de Medicamentos/métodos , Transferência Ressonante de Energia de Fluorescência/métodos , Proteínas de Bactérias/metabolismo , Caderinas/metabolismo , Cobre/metabolismo , Infecções por Helicobacter/tratamento farmacológico , Infecções por Helicobacter/metabolismo , Helicobacter pylori/metabolismo , Chaperonas Moleculares/metabolismo , Peptídeos/metabolismo , Proteômica/métodos , Serina Endopeptidases/metabolismo , Serina Proteases/metabolismo , Zinco/metabolismo
9.
Int J Mol Sci ; 21(11)2020 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-32486097

RESUMO

Helicobacter pylori (H. pylori) is a stomach pathogen that persistently colonizes the gastric mucosa, often leading to chronic inflammation and gastric pathologies. Although infection with H. pylori is the primary risk factor for gastric cancer, the underlying mechanisms of pathogen persistence and consequential chronic inflammation are still not well understood. Conventional dendritic cells (cDCs), which are among the first immune cells to encounter H. pylori in the gastric lining, and the cytokines and chemokines they secrete, contribute to both acute and chronic inflammation. Therefore, this study aimed to unravel the contributions of specific signaling pathways within human CD1c+ cDCs (cDC2s) to the composition of secreted cytokines and chemokines in H. pylori infection. Here, we show that the type IV secretion system (T4SS) plays only a minor role in H. pylori-induced activation of cDC2s. In contrast, Toll-like receptor 4 (TLR4) signaling drives the secretion of inflammatory mediators, including IL-12 and IL-18, while signaling via TLR10 attenuates the release of IL-1ß and other inflammatory cytokines upon H. pylori infection. The TLR2 pathway significantly blocks the release of CXCL1 and CXCL8, while it promotes the secretion of TNFα and GM-CSF. Taken together, these results highlight how specific TLR-signaling pathways in human cDC2s shape the H. pylori-induced cytokine and chemokine milieu, which plays a pivotal role in the onset of an effective immune response.


Assuntos
Quimiocinas/metabolismo , Citocinas/metabolismo , Células Dendríticas/metabolismo , Células Dendríticas/microbiologia , Receptor 10 Toll-Like/metabolismo , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Antígenos CD1/metabolismo , Mucosa Gástrica/metabolismo , Infecções por Helicobacter/metabolismo , Helicobacter pylori , Humanos , Inflamação , Mediadores da Inflamação/metabolismo , Leucócitos Mononucleares/citologia , Transdução de Sinais , Neoplasias Gástricas/microbiologia
10.
Gut Microbes ; 12(1): 1770017, 2020 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-32584649

RESUMO

Mechanisms of host-pathogen interactions resulting in immunopathological responses upon human Campylobacter jejuni infection are not completely understood, but the recent availability of murine infection models mimicking key features of campylobacteriosis helps solving this dilemma. During a screen for proteases expressed by C. jejuni, we identified a peptidase of the M24 family as a potential novel virulence factor, which was named PepP. The gene is strongly conserved in various Campylobacter species. A constructed deletion mutant ΔpepP of C. jejuni strain 81-176 grew as efficiently compared to isogenic wild-type (WT) or pepP complemented bacteria. To shed light on the potential role of this protease in mediating immunopathological responses in the mammalian host, we perorally challenged microbiota-depleted IL-10-/- mice with these strains. All strains stably colonized the murine gastrointestinal tract with comparably high loads. Remarkably, pepP deficiency was associated with less severe induced malaise, with less distinct apoptotic and innate immune cell responses, but also with more pronounced proliferative/regenerative epithelial cell responses in the large intestine at d6post-infection. Furthermore, pro-inflammatory mediators were lower in the colon, ileum, and mesenteric lymph nodes of mice that had been challenged with the ΔpepP mutant compared to the WT or pepP complemented strains. This also held true for extra-intestinal organs including liver, kidneys, and lungs, and, strikingly, to systemic compartments. Taken together, protease PepP is a novel virulence determinant involved in mediating campylobacteriosis. The finding that apoptosis in the colon is significantly diminished in mice infected with the pepP mutant highlights the epithelial layer as the first and main target of PepP in the intestine.


Assuntos
Infecções por Campylobacter/microbiologia , Campylobacter jejuni , Microbioma Gastrointestinal/fisiologia , Serina Endopeptidases/genética , Animais , Apoptose/fisiologia , Campylobacter jejuni/genética , Campylobacter jejuni/imunologia , Campylobacter jejuni/patogenicidade , Feminino , Doenças Transmitidas por Alimentos/microbiologia , Trato Gastrointestinal/microbiologia , Trato Gastrointestinal/patologia , Interleucina-10/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Serina Endopeptidases/metabolismo , Fatores de Virulência/genética
11.
Nanoscale ; 12(3): 2154-2155, 2020 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-31912840

RESUMO

Correction for 'Nanoparticle binding attenuates the pathobiology of gastric cancer-associated Helicobacter pylori' by Dana Westmeier et al., Nanoscale, 2018, 10, 1453-1463.

12.
BMC Microbiol ; 19(1): 255, 2019 11 14.
Artigo em Inglês | MEDLINE | ID: mdl-31726993

RESUMO

BACKGROUND: High temperature requirement A (HtrA) is a widely expressed chaperone and serine protease in bacteria. HtrA proteases assemble and hydrolyze misfolded proteins to enhance bacterial survival under stress conditions. Listeria monocytogenes (L. monocytogenes) is a foodborne pathogen that induces listeriosis in humans. In previous studies, it was shown that deletion of htrA in the genome of L. monocytogenes increased the susceptibility to cellular stress and attenuated virulence. However, expression and protease activity of listerial HtrA (LmHtrA) were never analyzed in detail. RESULTS: In this study, we cloned LmHtrA wildtype (LmHtrAwt) and generated a proteolytic inactive LmHtrASA mutant. Recombinant LmHtrAwt and LmHtrASA were purified and the proteolytic activity was analyzed in casein zymography and in vitro cleavage assays. LmHtrA activity could be efficiently blocked by a small molecule inhibitor targeting bacterial HtrA proteases. The expression of LmHtrA was enhanced in the stationary growth phase of L. monocytogenes and significantly contributed to bacterial survival at high temperatures. CONCLUSIONS: Our data show that LmHtrA is a highly active caseinolytic protease and provide a deeper insight into the function and mechanism, which could lead to medical and biotechnological applications in the future.


Assuntos
Caseínas/metabolismo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Listeria monocytogenes/crescimento & desenvolvimento , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Microbiologia de Alimentos , Regulação Bacteriana da Expressão Gênica , Proteínas de Choque Térmico/química , Resposta ao Choque Térmico , Listeria monocytogenes/patogenicidade , Viabilidade Microbiana , Dobramento de Proteína , Multimerização Proteica , Proteólise , Regulação para Cima
13.
Toxins (Basel) ; 11(10)2019 10 11.
Artigo em Inglês | MEDLINE | ID: mdl-31614680

RESUMO

Helicobacter pylori (H. pylori) has been identified as a leading cause of gastric cancer, which is one of the most frequent and malignant types of tumor. It is characterized by its rapid progression, distant metastases, and resistance to conventional chemotherapy. A number of receptor tyrosine kinases and non-receptor tyrosine kinases have been implicated in H. pylori-mediated pathogenesis and tumorigenesis. In this review, recent findings of deregulated EGFR, c-Met, JAK, FAK, Src, and c-Abl and their functions in H. pylori pathogenesis are summarized.


Assuntos
Infecções por Helicobacter/enzimologia , Helicobacter pylori , Proteínas Tirosina Quinases/metabolismo , Neoplasias Gástricas/enzimologia , Animais , Infecções por Helicobacter/complicações , Humanos , Neoplasias Gástricas/etiologia
14.
Adv Exp Med Biol ; 1149: 35-56, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31016624

RESUMO

Helicobacter pylori is a very successful Gram-negative pathogen colonizing the stomach of humans worldwide. Infections with this bacterium can generate pathologies ranging from chronic gastritis and peptic ulceration to gastric cancer. The best characterized H. pylori virulence factors that cause direct cell damage include an effector protein encoded by the cytotoxin-associated gene A (CagA), a type IV secretion system (T4SS) encoded in the cag-pathogenicity island (cag PAI), vacuolating cytotoxin A (VacA), γ-glutamyl transpeptidase (GGT), high temperature requirement A (HtrA, a serine protease) and cholesterol glycosyl-transferase (CGT). Since these H. pylori factors are either surface-exposed, secreted or translocated, they can directly interact with host cell molecules and are able to hijack cellular functions. Studies on these bacterial factors have progressed substantially in recent years. Here, we review the current status in the characterization of signaling cascades by these factors in vivo and in vitro, which comprise the disruption of cell-to-cell junctions, induction of membrane rearrangements, cytoskeletal dynamics, proliferative, pro-inflammatory, as well as, pro-apoptotic and anti-apoptotic responses or immune evasion. The impact of these signal transduction modules in the pathogenesis of H. pylori infections is discussed.


Assuntos
Infecções por Helicobacter , Helicobacter pylori , Fatores de Virulência , Proteínas de Bactérias/metabolismo , Ilhas Genômicas , Infecções por Helicobacter/microbiologia , Helicobacter pylori/patogenicidade , Humanos
15.
Cell Commun Signal ; 17(1): 10, 2019 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-30704478

RESUMO

BACKGROUND: Deregulated c-Abl activity has been intensively studied in a variety of solid tumors and leukemia. The class-I carcinogen Helicobacter pylori (Hp) activates the non-receptor tyrosine kinase c-Abl to phosphorylate the oncoprotein cytotoxin-associated gene A (CagA). The role of c-Abl in CagA-dependent pathways is well established; however, the knowledge of CagA-independent c-Abl processes is scarce. METHODS: c-Abl phosphorylation and localization were analyzed by immunostaining and immunofluorescence. Interaction partners were identified by tandem-affinity purification. Cell elongation and migration were analyzed in transwell-filter experiments. Apoptosis and cell survival were examined by FACS analyses and MTT assays. In mice experiments and human biopsies, the involvement of c-Abl in Hp pathogenesis was investigated. RESULTS: Here, we investigated the activity and subcellular localization of c-Abl in vitro and in vivo and unraveled the contribution of c-Abl in CagA-dependent and -independent pathways to gastric Hp pathogenesis. We report a novel mechanism and identified strong c-Abl threonine 735 phosphorylation (pAblT735) mediated by the type-IV secretion system (T4SS) effector D-glycero-ß-D-manno-heptose-1,7-bisphosphate (ßHBP) and protein kinase C (PKC) as a new c-Abl kinase. pAblT735 interacted with 14-3-3 proteins, which caused cytoplasmic retention of c-Abl, where it potentiated Hp-mediated cell elongation and migration. Further, the nuclear exclusion of pAblT735 attenuated caspase-8 and caspase-9-dependent apoptosis. Importantly, in human patients suffering from Hp-mediated gastritis c-Abl expression and pAblT735 phosphorylation were drastically enhanced as compared to type C gastritis patients or healthy individuals. Pharmacological inhibition using the selective c-Abl kinase inhibitor Gleevec confirmed that c-Abl plays an important role in Hp pathogenesis in a murine in vivo model. CONCLUSIONS: In this study, we identified a novel regulatory mechanism in Hp-infected gastric epithelial cells by which Hp determines the subcellular localization of activated c-Abl to control Hp-mediated EMT-like processes while decreasing cell death.


Assuntos
Apoptose , Movimento Celular , Helicobacter pylori/metabolismo , Proteínas Proto-Oncogênicas c-abl/metabolismo , Linhagem Celular Tumoral , Infecções por Helicobacter/metabolismo , Infecções por Helicobacter/microbiologia , Humanos , Modelos Biológicos , Fosforilação , Fosfotreonina/metabolismo , Fosfotirosina/metabolismo , Proteína Quinase C/metabolismo , Transporte Proteico
17.
Cell Microbiol ; 20(6): e12845, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29582532

RESUMO

The HtrA family of chaperones and serine proteases is important for regulating stress responses and controlling protein quality in the periplasm of bacteria. HtrA is also associated with infectious diseases since inactivation of htrA genes results in significantly reduced virulence properties by various bacterial pathogens. These virulence features of HtrA can be attributed to reduced fitness of the bacteria, higher susceptibility to environmental stress and/or diminished secretion of virulence factors. In some Gram-negative and Gram-positive pathogens, HtrA itself can be exposed to the extracellular environment promoting bacterial colonisation and invasion of host tissues. Most of our knowledge on the function of exported HtrAs stems from research on Helicobacter pylori, Campylobacter jejuni, Borrelia burgdorferi, Bacillus anthracis, and Chlamydia species. Here, we discuss recent progress showing that extracellular HtrAs are able to cleave cell-to-cell junction factors including E-cadherin, occludin, and claudin-8, as well as extracellular matrix proteins such as fibronectin, aggrecan, and proteoglycans, disrupting the epithelial barrier and producing substantial host cell damage. We propose that the export of HtrAs is a newly discovered strategy, also applied by additional bacterial pathogens. Consequently, exported HtrA proteases represent highly attractive targets for antibacterial treatment by inhibiting their proteolytic activity or application in vaccine development.


Assuntos
Bactérias/patogenicidade , Proteínas de Bactérias/metabolismo , Interações Hospedeiro-Patógeno , Chaperonas Moleculares/metabolismo , Serina Endopeptidases/metabolismo , Fatores de Virulência/metabolismo , Animais , Humanos , Estresse Fisiológico , Virulência
18.
Nanoscale ; 10(3): 1453-1463, 2018 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-29303193

RESUMO

Enteric bacteria may cause severe diseases, including gastric cancer-associated Helicobacter pylori. Their infection paths overlap with the oro-gastrointestinal uptake route for nanoparticles, increasingly occurring during environmental or consumer/medical exposure. By comprehensive independent analytical methods, such as live cell fluorescence, electron as well as atomic force microscopy and elemental analysis, we show that a wide array of nanoparticles (NPs) but not microparticles form complexes with H. pylori and enteric pathogens without the need for specific functionalization. The NP-assembly that occurred rapidly was not influenced by variations in physiological temperature, though affected by the NPs' physico-chemical characteristics. Improved binding was observed for small NPs with a negative surface charge, whereas binding could be reduced by surface 'stealth' modifications. Employing human gastric epithelial cells and 3D-organoid models of the stomach, we show that NP-coating did not inhibit H. pylori's cellular attachment. However, even the assembly of non-bactericidal silica NPs attenuated H. pylori infection by reducing CagA phosphorylation, cytoskeletal rearrangement, and IL-8 secretion. Here we demonstrate that NP binding to enteric bacteria may impact their pathobiology which could be further exploited to rationally modulate the (patho)biology of microbes by nanomaterials.


Assuntos
Helicobacter pylori/metabolismo , Helicobacter pylori/patogenicidade , Nanopartículas/metabolismo , Neoplasias Gástricas/microbiologia , Aderência Bacteriana , Células Epiteliais/microbiologia , Mucosa Gástrica/citologia , Humanos , Organoides/microbiologia , Dióxido de Silício
19.
NPJ Sci Food ; 2(1): 22, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30882042

RESUMO

Nanotechnology provides the food industry with new ways to modulate various aspects of food. Hence, engineered nanoparticles (NPs) are increasingly added to food and beverage products as functional ingredients. However, the impact of engineered as well as naturally occurring NPs on both commensal and pathogenic microorganisms within the gastrointestinal tract (GI) is not fully understood. Here, well-defined synthetic NPs and bacterial models were used to probe nanoparticle-bacteria interactions, from analytical to in situ to in vitro. NP-bacteria complexation occurred most efficiently for small NPs, independent of their core material or surface charge, but could be reduced by NPs' steric surface modifications. Adsorption to bacteria could also be demonstrated for naturally occurring carbon NPs isolated from beer. Complex formation affected the (patho)biological behavior of both the NPs and bacteria, including their cellular uptake into epithelial cells and phagocytes, pathogenic signaling pathways, and NP-induced cell toxicity. NP-bacteria complex formation was concentration-dependently reduced when the NPs became coated with biomolecule coronas with sequential simulation of first oral uptake and then the GI. However, efficient NP adsorption was restored when the pH was sufficiently low, such as in simulating the conditions of the stomach. Collectively, NP binding to enteric bacteria may impact their (patho)biology, particularly in the stomach. Nanosized-food additives as well as naturally occurring NPs may be exploited to (rationally) shape the microbiome. The information contained in this article should facilitate a "safe by design" strategy for the development and application of engineered NPs as functional foods ingredients.

20.
Cell Host Microbe ; 22(4): 552-560.e5, 2017 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-29024645

RESUMO

The Helicobacter pylori (Hp) type IV secretion system (T4SS) forms needle-like pili, whose binding to the integrin-ß1 receptor results in injection of the CagA oncoprotein. However, the apical surface of epithelial cells is exposed to Hp, whereas integrins are basolateral receptors. Hence, the mechanism of CagA delivery into polarized gastric epithelial cells remains enigmatic. Here, we demonstrate that T4SS pilus formation during infection of polarized cells occurs predominantly at basolateral membranes, and not at apical sites. Hp accomplishes this by secreting another bacterial protein, the serine protease HtrA, which opens cell-to-cell junctions through cleaving epithelial junctional proteins including occludin, claudin-8, and E-cadherin. Using a genetic system expressing a peptide inhibitor, we demonstrate that HtrA activity is necessary for paracellular transmigration of Hp across polarized cell monolayers to reach basolateral membranes and inject CagA. The contribution of this unique signaling cascade to Hp pathogenesis is discussed.


Assuntos
Antígenos de Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Infecções por Helicobacter/microbiologia , Helicobacter pylori/metabolismo , Helicobacter pylori/patogenicidade , Sistemas de Secreção Tipo IV/metabolismo , Linhagem Celular Tumoral , Polaridade Celular , Células Epiteliais/microbiologia , Fímbrias Bacterianas/metabolismo , Helicobacter pylori/genética , Serina Peptidase 1 de Requerimento de Alta Temperatura A/genética , Serina Peptidase 1 de Requerimento de Alta Temperatura A/metabolismo , Humanos , Transdução de Sinais , Migração Transendotelial e Transepitelial
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...