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1.
Cell Death Discov ; 10(1): 120, 2024 Mar 07.
Article in English | MEDLINE | ID: mdl-38453889

ABSTRACT

Gastric cancer's (GC) bad prognosis is usually associated with metastatic spread. Invasive cancer stem cells (CSC) are considered to be the seed of GC metastasis and not all CSCs are able to initiate metastasis. Targeting these aggressive metastasis-initiating CSC (MIC) is thus vital. Leukaemia inhibitory factor (LIF) is hereby used to target Hippo pathway oncogenic members, found to be induced in GC and associated with CSC features. LIF-treated GC cell lines, patient-derived xenograft (PDX) cells and/or CSC tumourspheres underwent transcriptomics, laser microdissection-associated proteomics, 2D and 3D invasion assays and in vivo xenograft in mice blood circulation. LIFR expression was analysed on tissue microarrays from GC patients and in silico from public databases. LIF-treated cells, especially CSC, presented decreased epithelial to mesenchymal transition (EMT) phenotype and invasion capacity in vitro, and lower metastasis initiation ability in vivo. These effects involved both the Hippo and Jak/Stat pathways. Finally, GC's high LIFR expression was associated with better clinical outcomes in patients. LIF treatment could thus represent a targeted anti-CSC strategy to fight against metastatic GC, and LIFR detection in primary tumours could constitute a potential new prognosis marker in this disease.

2.
J Infect Dis ; 229(6): 1688-1701, 2024 Jun 14.
Article in English | MEDLINE | ID: mdl-38416880

ABSTRACT

BACKGROUND: The bacterial genotoxin, cytolethal distending toxin (CDT), causes DNA damage in host cells, a risk factor for carcinogenesis. Previous studies have shown that CDT induces phenotypes reminiscent of epithelial to mesenchymal transition (EMT), a process involved in cancer initiation and progression. METHODS: We investigated different steps of EMT in response to Helicobacter hepaticus CDT and its active CdtB subunit using in vivo and in vitro models. RESULTS: Most of the steps of the EMT process were induced by CDT/CdtB and observed throughout the study in murine and epithelial cell culture models. CdtB induced cell-cell junction disassembly, causing individualization of cells and acquisition of a spindle-like morphology. The key transcriptional regulators of EMT (SNAIL and ZEB1) and some EMT markers were upregulated at both RNA and protein levels in response to CDT/CdtB. CdtB increased the expression and proteolytic activity of matrix metalloproteinases, as well as cell migration. A range of these results were confirmed in Helicobacter hepaticus-infected and xenograft murine models. In addition, colibactin, a genotoxic metabolite produced by Escherichia coli, induced EMT-like effects in cell culture. CONCLUSIONS: Overall, these data show that infection with genotoxin-producing bacteria elicits EMT process activation, supporting their role in tumorigenesis.


Subject(s)
Bacterial Toxins , Cell Differentiation , Epithelial-Mesenchymal Transition , Animals , Epithelial-Mesenchymal Transition/drug effects , Bacterial Toxins/toxicity , Bacterial Toxins/metabolism , Mice , Humans , Cell Differentiation/drug effects , Helicobacter hepaticus , Epithelial Cells/drug effects , Epithelial Cells/metabolism , Epithelial Cells/microbiology , Helicobacter Infections/microbiology , Snail Family Transcription Factors/metabolism , Snail Family Transcription Factors/genetics , Female
3.
Gastric Cancer ; 27(2): 292-307, 2024 03.
Article in English | MEDLINE | ID: mdl-38280128

ABSTRACT

BACKGROUND: Gastric cancer (GC), the fourth leading cause of cancer-related death worldwide, with most deaths caused by advanced and metastatic disease, has limited curative options. Here, we revealed the importance of proprotein convertases (PCs) in the malignant and metastatic potential of GC cells through the regulation of the YAP/TAZ/TEAD pathway and epithelial-to-mesenchymal transition (EMT) in cancer stem cells (CSC). METHODS: The general PCs inhibitor, decanoyl-RVKR-chloromethyl-ketone (CMK), was used to repress PCs activity in CSCs of various GC cell lines. Their tumorigenic properties, drug resistance, YAP/TAZ/TEAD pathway activity, and invasive properties were then investigated in vitro, and their metastatic properties were explored in a mouse xenograft model. The prognostic value of PCs in GC patients was also explored in molecular databases of GC. RESULTS: Inhibition of PCs activity in CSCs in all GC cell lines reduced tumorsphere formation and growth, drug efflux, EMT phenotype, and invasive properties that are associated with repressed YAP/TAZ/TEAD pathway activity in vitro. In vivo, PCs' inhibition in GC cells reduced their metastatic spread. Molecular analysis of tumors from GC patients has highlighted the prognostic value of PCs. CONCLUSIONS: PCs are overexpressed in GC and associated with poor prognosis. PCs are involved in the malignant and metastatic potential of CSCs via the regulation of EMT, the YAP/TAZ/TEAD oncogenic pathway, and their stemness and invasive properties. Their repression represents a new strategy to target CSCs and impair metastatic spreading in GC.


Subject(s)
Stomach Neoplasms , Transcription Factors , Humans , Animals , Mice , Transcription Factors/genetics , YAP-Signaling Proteins , Stomach Neoplasms/pathology , Disease Models, Animal , Proprotein Convertases/metabolism , Neoplastic Stem Cells/metabolism
4.
Gastric Cancer ; 26(2): 234-249, 2023 03.
Article in English | MEDLINE | ID: mdl-36528833

ABSTRACT

BACKGROUND: Cancer stem cells (CSCs) are at the origin of tumour initiation and progression in gastric adenocarcinoma (GC). However, markers of metastasis-initiating cells remain unidentified in GC. In this study, we characterized CD44 variants expressed in GC and evaluated the tumorigenic and metastatic properties of CD44v3+ cells and their clinical significance in GC patients. METHODS: Using GC cell lines and patient-derived xenografts, we evaluated CD44+ and CD44v3+ GC cells molecular signature and their tumorigenic, chemoresistance, invasive and metastatic properties, and expression in patients-derived tissues. RESULTS: CD44v3+ cells, which represented a subpopulation of CD44+ cells, were detected in advanced preneoplastic lesions and presented CSCs chemoresistance and tumorigenic properties in vitro and in vivo. Molecular and functional analyses revealed two subpopulations of gastric CSCs: CD44v3+ CSCs with an epithelial-mesenchymal transition (EMT)-like signature, and CD44+/v3- CSCs with an epithelial-like signature; both were tumorigenic but CD44v3+ cells showed higher invasive and metastatic properties in vivo. CD44v3+ cells detected in the primary tumours of GC patients were associated with a worse prognosis. CONCLUSION: CD44v3 is a marker of a subpopulation of CSCs with metastatic properties in GC. The identification of metastasis-initiating cells in GC represents a major advance for further development of anti-metastatic therapeutic strategies.


Subject(s)
Carcinoma , Stomach Neoplasms , Humans , Stomach Neoplasms/pathology , Neoplastic Stem Cells/metabolism , Carcinoma/pathology , Hyaluronan Receptors , Epithelial-Mesenchymal Transition
5.
Cancers (Basel) ; 14(17)2022 Sep 02.
Article in English | MEDLINE | ID: mdl-36077851

ABSTRACT

BACKGROUND: Gastric cancer, the fifth most common cancer worldwide, is mainly linked to Helicobacter pylori infection. H. pylori induces chronic inflammation of the gastric mucosa associated with high oxidative stress. Our study aimed at assessing the implication of Nrf2, a major regulator of cellular redox homeostasis, in H. pylori-induced gastric carcinogenesis. METHODS: Using three different gastric epithelial cell lines, a non-cancerous (HFE-145) and two different subtypes of gastric cancer (AGS and MKN74), we analyzed the modulation of Nrf2 expression over time. After invalidation of Nrf2 by CRISPR-cas9, we assessed its role in H. pylori-induced epithelial-to-mesenchymal transition (EMT). Finally, we evaluated the expression of Nrf2 and ZEB1, a central EMT transcription factor, in human gastric tissues. RESULTS: We first demonstrated that the Nrf2 signaling pathway is differentially regulated depending on the infection stage. Rapidly and transiently activated, Nrf2 was downregulated 24 h post-infection in a VacA-dependent manner. We then demonstrated that Nrf2 invalidation leads to increased EMT, which is even exacerbated after H. pylori infection. Finally, Nrf2 expression tended to decrease in human patients' gastric mucosa infected with H. pylori. CONCLUSIONS: Our work supports the hypothesis that Nrf2 downregulation upon H. pylori infection participates in EMT, one of the most important events in gastric carcinogenesis.

6.
Gastric Cancer ; 24(1): 133-144, 2021 Jan.
Article in English | MEDLINE | ID: mdl-32940810

ABSTRACT

BACKGROUND: The main cause of gastric cancer is the infection by the bacterium Helicobacter pylori which induces a chronic inflammation and an epithelial-to-mesenchymal transition (EMT) leading to the emergence of cells with cancer stem cell (CSC) properties. However, the underlying mechanisms have not been fully characterized. Moreover, H. pylori modulates the host cell autophagic process, but a few studies have investigated the role of this process in tumoral transformation. The aim of this study was to determine whether H. pylori-induced autophagy has a role in CSC emergence. METHODS: Autophagic flux in response to H. pylori infection was characterized in AGS cell line expressing the tandem-tagged mCherry-GFP-LC3 protein and using a ratiometric flow cytometry analysis. Then, AGS and MKN45 cell lines were treated with bafilomycin or chloroquine, two pharmaceutical well-known inhibitors of autophagy, and different EMT and CSC characteristics were analyzed. RESULTS: First, a co-expression of the gastric CSC marker CD44 and the autophagic marker LC3 in mice and human stomach tissues infected with H. pylori was observed. Then, we demonstrated in vitro that H. pylori was able to activate the autophagy process with a reduced autophagic flux. Finally, infected cells were treated with autophagy inhibitors, which reduced (i) appearance of mesenchymal phenotypes and migration ability related to EMT and (ii) CD44 expression as well as tumorsphere formation capacities reflecting CSC properties. CONCLUSION: In conclusion, all these data show that H. pylori-induced autophagy is implicated in gastric CSC emergence and could represent an interesting therapeutic target.


Subject(s)
Autophagy/physiology , Helicobacter Infections/microbiology , Helicobacter pylori/physiology , Neoplastic Stem Cells/microbiology , Stomach Neoplasms/microbiology , Animals , Cell Line, Tumor , Epithelial-Mesenchymal Transition , Helicobacter Infections/complications , Humans , Hyaluronan Receptors/metabolism , Mice , Microtubule-Associated Proteins/metabolism , Stomach/cytology , Stomach/microbiology
7.
Sci Rep ; 10(1): 14858, 2020 09 09.
Article in English | MEDLINE | ID: mdl-32908188

ABSTRACT

The roles of the inflammatory response and production of a proliferation-inducing ligand (APRIL) cytokine in gastric mucosa-associated lymphoid tissue (MALT) lymphomagenesis induced by Helicobacter species infection are not clearly understood. We characterized the gastric mucosal inflammatory response associated with gastric MALT lymphoma (GML) and identified APRIL-producing cells in two model systems: an APRIL transgenic mouse model of GML induced by Helicobacter infection (Tg-hAPRIL) and human gastric biopsy samples from Helicobacter pylori-infected GML patients. In the mouse model, polarization of T helper 1 (tbet), T helper 2 (gata3), and regulatory T cell (foxp3) responses was evaluated by quantitative PCR. In humans, a significant increase in april gene expression was observed in GML compared to gastritis. APRIL-producing cells were eosinophilic polynuclear cells located within lymphoid infiltrates, and tumoral B lymphocytes were targeted by APRIL. Together, the results of this study demonstrate that the Treg-balanced inflammatory environment is important for gastric lymphomagenesis induced by Helicobacter species, and suggest the pro-tumorigenic potential of APRIL-producing eosinophils.


Subject(s)
B-Lymphocytes/immunology , Eosinophils/immunology , Helicobacter Infections , Lymphoma, B-Cell, Marginal Zone , T-Lymphocytes/immunology , Tumor Necrosis Factor Ligand Superfamily Member 13/immunology , Adult , Animals , Female , Gastric Mucosa/immunology , Gastric Mucosa/pathology , Helicobacter Infections/complications , Helicobacter Infections/immunology , Helicobacter Infections/pathology , Humans , Lymphoma, B-Cell, Marginal Zone/etiology , Lymphoma, B-Cell, Marginal Zone/immunology , Lymphoma, B-Cell, Marginal Zone/pathology , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Middle Aged
8.
Cancers (Basel) ; 12(8)2020 Jul 22.
Article in English | MEDLINE | ID: mdl-32707998

ABSTRACT

Cancer stem cells (CSCs) present chemo-resistance mechanisms contributing to tumour maintenance and recurrence, making their targeting of utmost importance in gastric cancer (GC) therapy. The Hippo pathway has been implicated in gastric CSC properties and was shown to be regulated by leukaemia inhibitory factor receptor (LIFR) and its ligand LIF in breast cancer. This study aimed to determine LIF's effect on CSC properties in GC cell lines and patient-derived xenograft (PDX) cells, which remains unexplored. LIF's treatment effect on CSC markers expression and tumoursphere formation was evaluated. The Hippo kinase inhibitor XMU-MP-1 and/or the JAK1 inhibitor Ruxolitinib were used to determine Hippo and canonical JAK/STAT pathway involvement in gastric CSCs' response to LIF. Results indicate that LIF decreased tumorigenic and chemo-resistant CSCs, in both GC cell lines and PDX cells. In addition, LIF increased activation of LATS1/2 Hippo kinases, thereby decreasing downstream YAP/TAZ nuclear accumulation and TEAD transcriptional activity. LIF's anti-CSC effect was reversed by XMU-MP-1 but not by Ruxolitinib treatment, highlighting the opposite effects of these two pathways downstream LIFR. In conclusion, LIF displays anti-CSC properties in GC, through Hippo kinases activation, and could in fine constitute a new CSCs-targeting strategy to help decrease relapse cases and bad prognosis in GC.

9.
J Food Prot ; 83(12): 2080-2086, 2020 Dec 01.
Article in English | MEDLINE | ID: mdl-32634222

ABSTRACT

ABSTRACT: Arcobacter is considered an emergent foodborne enteropathogen. Despite the high prevalence of this genus in poultry, the occurrence of Arcobacter spp. contamination in Tunisia remains unclear. The objectives of this study were (i) to isolate Arcobacter species (A. butzleri and A. cryaerophilus) by the culture method from different species of raw poultry meat, (ii) to verify the isolates by multiplex PCR (m-PCR) assay and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), and (iii) to determine the antibiotic resistance profiles of the isolates. A total of 250 poultry product samples (149 chicken and 101 turkey) were collected from various supermarkets in Sfax. The samples consisted of breasts, wings, legs, and neck skins. The overall isolation frequency of Arcobacter spp. was 10.4%. Arcobacter spp. were found in 13.42% of the chicken samples and in 5.49% of the turkey samples. All the acquired isolates were subject to detailed confirmation with subsequent species classification using m-PCR and MALDI-TOF MS. A. butzleri was found in 22 samples (84.61%) and A. cryaerophilus in 4 samples (15.38%). Thus, m-PCR and MALDI-TOF MS were able to detect A. butzleri significantly better than the conventional method (χ2 = 49.1 and P < 0.001). Arcobacter was isolated from poultry in every season, at contamination levels of 30.76, 23.07, 19.23, and 26.92% in summer, spring, autumn, and winter, respectively. The disk diffusion method was used to determine the susceptibility of Arcobacter isolates to six antimicrobial drugs. All A. butzleri isolates (n = 24) were significantly resistant to erythromycin (P = 0.0015), ampicillin (P = 0.001), and ciprofloxacin (P = 0.05). All tested A. cryaerophilus strains (n = 4) were susceptible to ampicillin, gentamicin, and amoxicillin-clavulanic acid. Multidrug resistance was observed in 83% of the Arcobacter spp. isolates. Our study detected Arcobacter spp. in Tunisian poultry; because of their multidrug resistance, these species may constitute a public health problem.


Subject(s)
Arcobacter , Animals , Drug Resistance, Microbial , Food Microbiology , Poultry , Tunisia
10.
Cells ; 9(6)2020 06 13.
Article in English | MEDLINE | ID: mdl-32545795

ABSTRACT

Helicobacter pylori infection, the main risk factor for gastric cancer (GC), leads to an epithelial-mesenchymal transition (EMT) of gastric epithelium contributing to gastric cancer stem cell (CSC) emergence. The Hippo pathway effectors yes-associated protein (YAP) and transcriptional co-activator with PDZ binding motif (TAZ) control cancer initiation and progression in many cancers including GC. Here, we investigated the role of TAZ in the early steps of H. pylori-mediated gastric carcinogenesis. TAZ implication in EMT, invasion, and CSC-related tumorigenic properties were evaluated in three gastric epithelial cell lines infected by H. pylori. We showed that H. pylori infection increased TAZ nuclear expression and transcriptional enhancer TEA domain (TEAD) transcription factors transcriptional activity. Nuclear TAZ and zinc finger E-box-binding homeobox 1 (ZEB1) were co-overexpressed in cells harboring a mesenchymal phenotype in vitro, and in areas of regenerative hyperplasia in gastric mucosa of H. pylori-infected patients and experimentally infected mice, as well as at the invasive front of gastric carcinoma. TAZ silencing reduced ZEB1 expression and EMT phenotype, and strongly inhibited invasion and tumorsphere formation induced by H. pylori. In conclusion, TAZ activation in response to H. pylori infection contributes to H. pylori-induced EMT, invasion, and CSC-like tumorigenic properties. TAZ overexpression in H. pylori-induced pre-neoplastic lesions and in GC could therefore constitute a biomarker of early transformation in gastric carcinogenesis.


Subject(s)
Epithelial Cells/metabolism , Gastric Mucosa/pathology , Helicobacter Infections/metabolism , Neoplastic Stem Cells/pathology , Animals , Epithelial Cells/pathology , Epithelial-Mesenchymal Transition/physiology , Helicobacter Infections/genetics , Helicobacter Infections/pathology , Helicobacter pylori/metabolism , Humans , Hyaluronan Receptors/metabolism , Mice , Neoplastic Stem Cells/metabolism , Transcription Factors/metabolism
11.
J Alzheimers Dis ; 73(2): 801-809, 2020.
Article in English | MEDLINE | ID: mdl-31868664

ABSTRACT

Despite extensive research, the origin of Alzheimer's disease (AD) remains unknown. The role of infectious pathogens has recently emerged. Epidemiological studies have shown that Helicobacter pylori infection increases the risk of developing AD. We hypothesized that H. pylori-induced gastritis may be associated with a systemic inflammation and finally neuroinflammation. C57BL/6 mice were infected with H. pylori (n = 15) or Helicobacter felis (n = 13) or left uninfected (n = 9) during 18 months. Gastritis, amyloid deposition, astroglial and microglial cell area, and systemic and brain cytokines were assessed. The infection (H. felis> H. pylori) induced a severe gastritis and an increased neuroinflammation but without brain amyloid deposition or systemic inflammation.


Subject(s)
Alzheimer Disease/complications , Alzheimer Disease/microbiology , Encephalitis/etiology , Gastritis/complications , Gastritis/microbiology , Helicobacter Infections/complications , Helicobacter Infections/microbiology , Helicobacter pylori , Inflammation/complications , Inflammation/microbiology , Animals , Astrocytes/pathology , Brain Chemistry , Cytokines/metabolism , Helicobacter felis , Mice , Mice, Inbred C57BL , Microglia/pathology , Plaque, Amyloid/pathology
12.
Int J Cancer ; 146(8): 2255-2267, 2020 04 15.
Article in English | MEDLINE | ID: mdl-31489619

ABSTRACT

Gastric carcinomas (GC) are heterogeneous tumors, composed of a subpopulation of cluster of differentiation-44 (CD44)+ tumorigenic and chemoresistant cancer stem cells (CSC). YAP1 and TAZ oncoproteins (Y/T) interact with TEA domain family member 1 (TEAD) transcription factors to promote cell survival and proliferation in multiple tissues. Their activity and role in GC remain unclear. This work aimed to analyze Y/T-TEAD activity and molecular signature in gastric CSC, and to assess the effect of verteporfin, a Food and Drug Administration-approved drug preventing Y/T-TEAD interaction, on gastric CSC tumorigenic properties. Y/T-TEAD molecular signature was investigated using bioinformatical (KmPlot database), transcriptomic and immunostaining analyses in patient-derived GC and cell lines. Verteporfin effects on Y/T-TEAD transcriptional activity, CSC proliferation and tumorigenic properties were evaluated using in vitro tumorsphere assays and mouse models of patient-derived GC xenografts. High expressions of YAP1, TAZ, TEAD1, TEAD4 and their target genes were associated with low overall survival in nonmetastatic human GC patients (n = 444). This Y/T-TEAD molecular signature was enriched in CD44+ patient-derived GC cells and in cells resistant to conventional chemotherapy. Verteporfin treatment inhibited Y/T-TEAD transcriptional activity, cell proliferation and CD44 expression, and decreased the pool of tumorsphere-forming CD44+ /aldehyde dehydrogenase (ALDH)high gastric CSC. Finally, verteporfin treatment inhibited GC tumor growth in vivo; the residual tumor cells exhibited reduced expressions of CD44 and ALDH1, and more importantly, they were unable to initiate new tumorspheres in vitro. All these data demonstrate that Y/T-TEAD activity controls gastric CSC tumorigenic properties. The repositioning of verteporfin targeting YAP1/TAZ-TEAD activity could be a promising CSC-based strategy for the treatment of GC.


Subject(s)
Adaptor Proteins, Signal Transducing/genetics , DNA-Binding Proteins/genetics , Neoplastic Stem Cells/drug effects , Nuclear Proteins/genetics , Stomach Neoplasms/drug therapy , Stomach Neoplasms/genetics , Trans-Activators/genetics , Transcription Factors/genetics , Verteporfin/pharmacology , Adaptor Proteins, Signal Transducing/metabolism , Animals , Cell Growth Processes/drug effects , Cell Line, Tumor , DNA-Binding Proteins/metabolism , Drug Resistance, Neoplasm , Humans , Hyaluronan Receptors/biosynthesis , Hyaluronan Receptors/genetics , Male , Mice , Mice, Inbred NOD , Mice, SCID , Molecular Targeted Therapy , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Nuclear Proteins/metabolism , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology , TEA Domain Transcription Factors , Trans-Activators/metabolism , Transcription Factors/metabolism , Transcription, Genetic/drug effects , Transcriptional Coactivator with PDZ-Binding Motif Proteins , Up-Regulation , Xenograft Model Antitumor Assays , YAP-Signaling Proteins
13.
Cell Mol Gastroenterol Hepatol ; 9(2): 257-276, 2020.
Article in English | MEDLINE | ID: mdl-31669263

ABSTRACT

BACKGROUND & AIMS: Gastric carcinoma is related mostly to CagA+-Helicobacter pylori infection, which disrupts the gastric mucosa turnover and elicits an epithelial-mesenchymal transition (EMT) and preneoplastic transdifferentiation. The tumor suppressor Hippo pathway controls stem cell homeostasis; its core, constituted by the large tumor suppressor 2 (LATS2) kinase and its substrate Yes-associated protein 1 (YAP1), was investigated in this context. METHODS: Hippo, EMT, and intestinal metaplasia marker expression were investigated by transcriptomic and immunostaining analyses in human gastric AGS and MKN74 and nongastric immortalized RPE1 and HMLE epithelial cell lines challenged by H pylori, and on gastric tissues of infected patients and mice. LATS2 and YAP1 were silenced using small interfering RNAs. A transcriptional enhanced associated domain (TEAD) reporter assay was used. Cell proliferation and invasion were evaluated. RESULTS: LATS2 and YAP1 appear co-overexpressed in the infected mucosa, especially in gastritis and intestinal metaplasia. H pylori via CagA stimulates LATS2 and YAP1 in a coordinated biphasic pattern, characterized by an early transient YAP1 nuclear accumulation and stimulated YAP1/TEAD transcription, followed by nuclear LATS2 up-regulation leading to YAP1 phosphorylation and targeting for degradation. LATS2 and YAP1 reciprocally positively regulate each other's expression. Loss-of-function experiments showed that LATS2 restricts H pylori-induced EMT marker expression, invasion, and intestinal metaplasia, supporting a role of LATS2 in maintaining the epithelial phenotype of gastric cells and constraining H pylori-induced preneoplastic changes. CONCLUSIONS: H pylori infection engages a number of signaling cascades that alienate mucosa homeostasis, including the Hippo LATS2/YAP1/TEAD pathway. In the host-pathogen conflict, which generates an inflammatory environment and perturbations of the epithelial turnover and differentiation, Hippo signaling appears as a protective pathway, limiting the loss of gastric epithelial cell identity that precedes gastric carcinoma development.


Subject(s)
Epithelial-Mesenchymal Transition/immunology , Gastric Mucosa/pathology , Helicobacter Infections/pathology , Precancerous Conditions/pathology , Protein Serine-Threonine Kinases/metabolism , Tumor Suppressor Proteins/metabolism , Adaptor Proteins, Signal Transducing/metabolism , Adenocarcinoma/genetics , Adenocarcinoma/immunology , Adenocarcinoma/pathology , Aged , Aged, 80 and over , Animals , Cell Cycle Proteins/metabolism , Female , Gastric Mucosa/microbiology , Gene Expression Regulation, Neoplastic/immunology , Helicobacter Infections/genetics , Helicobacter Infections/microbiology , Helicobacter pylori/pathogenicity , Host-Pathogen Interactions/genetics , Humans , Male , Metaplasia/genetics , Metaplasia/microbiology , Metaplasia/pathology , Mice , Precancerous Conditions/genetics , Precancerous Conditions/immunology , Protective Factors , Protein Serine-Threonine Kinases/genetics , Signal Transduction/genetics , Signal Transduction/immunology , Stomach Neoplasms/genetics , Stomach Neoplasms/immunology , Stomach Neoplasms/pathology , Transcription Factors/metabolism , Tumor Suppressor Proteins/genetics , YAP-Signaling Proteins
14.
Cancers (Basel) ; 11(4)2019 Apr 19.
Article in English | MEDLINE | ID: mdl-31010193

ABSTRACT

Gastric cancer is the third leading cause of cancer mortality worldwide. Cancer stem cells (CSC) are at the origin of tumor initiation, chemoresistance, and the formation of metastases. However, there is a lack of mouse models enabling the study of the metastatic process in gastric adenocarcinoma (GC). The aims of this study were to develop original mouse models of patient-derived primary GC orthotopic xenografts (PDOX) allowing the development of distant metastases as preclinical models to study the anti-metastatic efficiency of drugs such as the phosphatidylinositol 3-kinase (PI3K) inhibitor Buparlisib (BKM120). Luciferase-encoding cells generated from primary GC were injected into the stomach wall of immunocompromised mice; gastric tumor and metastases development were followed by bioluminescence imaging. The anti-CSC properties of BKM120 were evaluated on the GC cells' phenotype (CD44 expression) and tumorigenic properties in vitro and in vivo on BKM120-treated mice. After eight weeks, PDOX mice formed tumors in the stomach as well as distant metastases, that were enriched in CSC, in the liver, the lung, and the peritoneal cavity. BKM120 treatment significantly inhibited the CSC properties in vitro and reduced the number of distant metastases in mice. These new preclinical models offer the opportunity to study the anti-metastatic efficiency of new CSC-based therapeutic strategies.

15.
J Clin Microbiol ; 56(9)2018 09.
Article in English | MEDLINE | ID: mdl-29976591

ABSTRACT

A whole-genome sequencing (WGS) approach was conducted in order to identify the molecular determinants associated with antimicrobial resistance in 12 multidrug-resistant Campylobacter jejuni and Campylobacter coli isolates, with a focus on aminoglycoside resistance determinants. Two variants of a new aminoglycoside phosphotransferase gene [aph(2″)-Ii1 and aph(2″)-Ii2 ] putatively associated with gentamicin resistance were found. In addition, the following new genes were identified for the first time in Campylobacter: a lincosamide nucleotidyltransferase gene [lnu(G)], likely associated with lincomycin resistance, and two resistance enzyme genes (spw and apmA) similar to those found in Staphylococcus aureus, which may confer spectinomycin and gentamicin resistance, respectively. A C1192T mutation of the 16S rRNA gene that may be involved in spectinomycin resistance was also found in a C. coli isolate. Genes identified in the present study were located either on the bacterial chromosome or on plasmids that could be transferred naturally. Their role in aminoglycoside resistance remains to be supported by genetic studies. Regarding the other antimicrobial agents studied, i.e., ampicillin, ciprofloxacin, erythromycin, and tetracycline, a perfect correlation between antimicrobial phenotypes and genotypes was found. Overall, our data suggest that WGS analysis is a powerful tool for identifying resistance determinants in Campylobacter and can disclose the full genetic elements associated with resistance, including antimicrobial compounds not tested routinely in antimicrobial susceptibility testing.


Subject(s)
Aminoglycosides/pharmacology , Anti-Bacterial Agents/pharmacology , Campylobacter Infections/microbiology , Campylobacter/genetics , Campylobacter/isolation & purification , Drug Resistance, Multiple, Bacterial/genetics , Genome, Bacterial/genetics , Animals , Bacterial Proteins/genetics , Campylobacter/classification , Campylobacter/drug effects , Campylobacter coli/classification , Campylobacter coli/drug effects , Campylobacter coli/genetics , Campylobacter coli/isolation & purification , Campylobacter jejuni/classification , Campylobacter jejuni/drug effects , Campylobacter jejuni/isolation & purification , DNA, Bacterial/genetics , Humans , Mutation , Phylogeny , Plasmids , RNA, Ribosomal, 16S/genetics , Red Meat/microbiology , Sequence Analysis, DNA
16.
Eur J Cancer ; 84: 193-201, 2017 10.
Article in English | MEDLINE | ID: mdl-28822889

ABSTRACT

Gastric cancer is the third leading cause of cancer-related deaths worldwide and has still a poor prognosis. Therefore, new therapeutic strategies are needed: among them, targeting cancer stem cells (CSCs) could offer new opportunities. The aim of our study was to evaluate the anti-tumoural effect of metformin on gastric cancer in vitro and in vivo and especially, to determine whether this molecule could target the gastric CSCs. Metformin effects were evaluated on the proliferation and tumourigenic properties of the gastric CSCs from patient-derived primary tumour xenografts (PDXs) and cancer cell lines (MKN45, AGS and MKN74) in vitro in conventional 2 dimensional (2D) and in 3 dimensional (3D) culture systems, in which only CSCs are able to form tumourspheres and in mouse xenograft models in vivo. Metformin induced a cell cycle arrest, which decreased cell proliferation in the 2D cultures. In a 3D culture system, metformin decreased the number of tumourspheres, revealing its capacity to target the CSCs. This effect was confirmed by the study of the expression of CSC markers (CD44 and Sox2) and differentiation markers (Kruppel-like factor 4 and MUC5AC), which were decreased or increased in response to metformin, respectively. Finally, in vivo treatment of PDXs with metformin led to a tumour growth delay and decreased the self-renewal ability of the CSCs. These results suggest that the use of metformin could represent an efficient strategy to inhibit tumour growth by targeting gastric CSCs.


Subject(s)
Antineoplastic Agents/pharmacology , Cell Differentiation/drug effects , Cell Proliferation/drug effects , Metformin/pharmacology , Neoplastic Stem Cells/drug effects , Stomach Neoplasms/drug therapy , Animals , Biomarkers, Tumor/metabolism , Cell Cycle Checkpoints/drug effects , Cell Line, Tumor , Cell Self Renewal/drug effects , Dose-Response Relationship, Drug , Humans , Hyaluronan Receptors/metabolism , Kruppel-Like Factor 4 , Kruppel-Like Transcription Factors/metabolism , Mice , Mucin 5AC/metabolism , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Phenotype , SOXB1 Transcription Factors/metabolism , Spheroids, Cellular , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology , Time Factors , Tumor Burden/drug effects , Tumor Cells, Cultured , Xenograft Model Antitumor Assays
17.
Article in English | MEDLINE | ID: mdl-28713773

ABSTRACT

Cytolethal distending toxins (CDTs) are common among pathogenic bacteria of the human and animal microbiota. CDTs exert cytopathic effets, via their active CdtB subunit. No clear description of those cytopathic effects has been reported at the cellular level in the target organs in vivo. In the present study, xenograft mouse models of colon and liver cell lines were set up to study the effects of the CdtB subunit of Helicobacter hepaticus. Conditional transgenic cell lines were established, validated in vitro and then engrafted into immunodeficient mice. After successful engraftment, mice were treated with doxycyclin to induce the expression of transgenes (red fluorescent protein, CdtB, and mutated CdtB). For both engrafted cell lines, results revealed a delayed tumor growth and a reduced tumor weight in CdtB-expressing tumors compared to controls. CdtB-derived tumors showed γ-H2AX foci formation, an increase in apoptosis, senescence, p21 and Ki-67 nuclear antigen expression. No difference in proliferating cells undergoing mitosis (phospho-histone H3) was observed. CdtB intoxication was also associated with an overexpression of cytokeratins in cells at the invasive front of the tumor as well as an increase in ploidy. All these features are hallmarks of endoreplication, as well as aggressiveness in cancer. These effects were dependent on the histidine residue at position 265 of the CdtB, underlying the importance of this residue in CdtB catalytic activity. Taken together, these data indicate that the CdtB triggers senescence and cell endoreplication leading to giant polyploid cells in these xenograft mouse models.


Subject(s)
Aging/drug effects , Bacterial Toxins/pharmacology , Cell Line, Tumor/drug effects , Corneal Dystrophies, Hereditary/metabolism , Endoreduplication/drug effects , Helicobacter hepaticus/metabolism , Intestines/drug effects , Liver/drug effects , Animals , Apoptosis , Bacterial Toxins/metabolism , Cell Cycle/drug effects , Cytoskeleton/drug effects , Doxycycline/pharmacology , Epithelial Cells , HT29 Cells/drug effects , Heterografts , Histones/metabolism , Humans , Ki-67 Antigen/metabolism , Luminescent Proteins , Mice , Microbiota , Red Fluorescent Protein
18.
Article in English | MEDLINE | ID: mdl-28560185

ABSTRACT

Helicobacter pylori infection is considered as an excellent model of chronic inflammation-induced tumor development. Our project focuses on gastric MALT lymphoma (GML) related to H. pylori infection and mediated by the chronic inflammatory process initiated by the infection. Recently, microRNAs (miRNAs) have emerged as a new class of gene regulators, which play key roles in inflammation and carcinogenesis acting as oncogenes or tumor suppressors. Their precise characterization in the development of inflammation and their contribution in regulating host cells responses to infection by H. pylori have been little explored. Our goal was to analyze the changes in miRNAs in a GML mouse model using BALB/c mice thymectomized at day 3 post-birth (d3Tx model) and to clarify their implication in GML pathogenesis. PCR array followed by RT-qPCR identified five miRNAs (miR-21a, miR-135b, miR-142a, miR-150, miR-155) overexpressed in the stomachs of GML-developing d3Tx mice infected by H. pylori. The analysis of their putative targets allowed us to identify TP53INP1, an anti-proliferative and pro-apoptotic protein, as a common target of 4 of the 5 up-regulated miRNAs. We postulate that these miRNAs may act in synergy to promote the development of GML. miR-142a was also overexpressed in mouse sera samples and therefore could serve as a diagnostic marker. In situ hybridization on gastric samples with miR-142a revealed a global up-regulation of this miRNA by the tumor microenvironment at the lymphoma stage. Dysregulation of miR-21a, miR-135b, miR-142a, miR-150, miR-155 could play a critical role in the pathogenesis of GML and might offer potential applications as therapeutic targets and novel biomarkers for this disease.


Subject(s)
Helicobacter Infections/complications , Helicobacter pylori/pathogenicity , Lymphoma, B-Cell, Marginal Zone/immunology , Lymphoma, Non-Hodgkin/immunology , MicroRNAs/biosynthesis , MicroRNAs/genetics , Stomach Neoplasms/immunology , Animals , Apoptosis , Biomarkers , Carcinogenesis , Disease Models, Animal , Gene Expression Regulation , In Situ Hybridization , Inflammation/immunology , Inflammation/microbiology , Lymphoma, B-Cell, Marginal Zone/genetics , Lymphoma, B-Cell, Marginal Zone/metabolism , Lymphoma, Non-Hodgkin/genetics , Lymphoma, Non-Hodgkin/metabolism , Mice , Mice, Inbred BALB C , Nuclear Proteins/metabolism , Oncogenes , Stomach Neoplasms/genetics , Stomach Neoplasms/metabolism
19.
Am J Pathol ; 187(7): 1473-1484, 2017 Jul.
Article in English | MEDLINE | ID: mdl-28460208

ABSTRACT

APRIL is a member of the tumor necrosis factor cytokine family involved in the regulation of B-cell immunity. We present a study of the infection by Helicobacter species of transgenic (Tg) C57BL6 mice, ectopically expressing the human form of APRIL. Wild-type (WT) and APRIL Tg mice were infected with Helicobacter felis and Helicobacter pylori and compared with noninfected animals. Mice were euthanized 18 months after infection, and inflammatory responses and histologic alterations were analyzed. Flow cytometry results revealed that WT-infected mice had less leukocyte infiltration than APRIL Tg-infected mice. In WT-infected mice, infiltrates in gastric tissues were predominantly composed of T cells, mainly CD4+ for H. pylori and CD8+ for H. felis. In APRIL Tg-infected mice, leukocyte infiltrates were composed of B cells with few CD4+ T cells for both species. B cells expressed B surface markers compatible with a marginal zone origin. These results were confirmed by immunohistochemistry. B cells in particular were involved in lymphoepithelial lesions, a hallmark of gastric MALT lymphoma. Monoclonality was observed in a few infiltrates in the presence of lymphoepithelial lesions. These results confirm the importance of APRIL in the development of gastric lymphoid infiltrates induced by Helicobacter species in vivo. We believe that APRIL Tg mice infected by Helicobacter species may represent a novel animal model of gastric lymphomagenesis.


Subject(s)
Helicobacter Infections/microbiology , Helicobacter pylori/immunology , Lymphoma, B-Cell, Marginal Zone/microbiology , Lymphoma, Non-Hodgkin/microbiology , Stomach Neoplasms/microbiology , Animals , B-Lymphocytes/microbiology , B-Lymphocytes/pathology , Bacterial Load , CD4-Positive T-Lymphocytes/microbiology , CD4-Positive T-Lymphocytes/pathology , Disease Models, Animal , Female , Helicobacter Infections/immunology , Helicobacter Infections/pathology , Humans , Immunohistochemistry , Inflammation , Lymphoid Tissue/microbiology , Lymphoid Tissue/pathology , Lymphoma, B-Cell, Marginal Zone/immunology , Lymphoma, B-Cell, Marginal Zone/pathology , Lymphoma, Non-Hodgkin/immunology , Lymphoma, Non-Hodgkin/pathology , Mice , Mice, Inbred C57BL , Mice, Transgenic , Stomach/microbiology , Stomach/pathology , Stomach Neoplasms/immunology , Stomach Neoplasms/pathology , Tumor Necrosis Factor Ligand Superfamily Member 13/genetics , Tumor Necrosis Factor Ligand Superfamily Member 13/immunology
20.
Oncotarget ; 7(49): 80688-80699, 2016 12 06.
Article in English | MEDLINE | ID: mdl-27729612

ABSTRACT

Helicobacter pylori infection is responsible for gastric carcinogenesis but host factors are also implicated. IQGAP1, a scaffolding protein of the adherens junctions interacting with E-cadherin, regulates cellular plasticity and proliferation. In mice, IQGAP1 deficiency leads to gastric hyperplasia. The aim of this study was to elucidate the consequences of IQGAP1 deletion on H. pylori-induced gastric carcinogenesis.Transgenic mice deleted for iqgap1 and WT littermates were infected with Helicobacter sp., and histopathological analyses of the gastric mucosa were performed. IQGAP1 and E-cadherin expression was evaluated in gastric tissues and in gastric epithelial cell lines in response to H. pylori infection. The consequences of IQGAP1 deletion on gastric epithelial cell behaviour and on the acquisition of cancer stem cell (CSC)-like properties were evaluated. After one year of infection, iqgap1+/- mice developed more preneoplastic lesions and up to 8 times more gastro-intestinal neoplasia (GIN) than WT littermates. H. pylori infection induced IQGAP1 and E-cadherin delocalization from cell-cell junctions. In vitro, knock-down of IQGAP1 favoured the acquisition of a mesenchymal phenotype and CSC-like properties induced by H. pylori infection.Our results indicate that alterations in IQGAP1 signalling promote the emergence of CSCs and gastric adenocarcinoma development in the context of an H. pylori infection.


Subject(s)
Adenocarcinoma/microbiology , Gastric Mucosa/microbiology , Helicobacter Infections/microbiology , Helicobacter pylori/pathogenicity , Neoplastic Stem Cells/microbiology , Precancerous Conditions/microbiology , Stomach Neoplasms/microbiology , ras GTPase-Activating Proteins/deficiency , Adenocarcinoma/genetics , Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Animals , Cadherins/metabolism , Cell Line, Tumor , Cell Proliferation , Cell Transformation, Neoplastic/genetics , Cell Transformation, Neoplastic/metabolism , Cell Transformation, Neoplastic/pathology , Disease Models, Animal , Epithelial-Mesenchymal Transition , Female , Gastric Mucosa/metabolism , Gastric Mucosa/pathology , Genetic Predisposition to Disease , Helicobacter Infections/genetics , Helicobacter Infections/metabolism , Helicobacter Infections/pathology , Host-Pathogen Interactions , Hyaluronan Receptors/metabolism , Hyperplasia , Mice, 129 Strain , Mice, Knockout , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Phenotype , Precancerous Conditions/genetics , Precancerous Conditions/metabolism , Precancerous Conditions/pathology , Stomach Neoplasms/genetics , Stomach Neoplasms/metabolism , Stomach Neoplasms/pathology , Time Factors , ras GTPase-Activating Proteins/genetics
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