Helicobacter pylori
            ‐activated gastric fibroblasts induce epithelial‐mesenchymal transition of gastric epithelial cells in vitro in a TGF‐β‐dependent manner

Background/Aims: Transforming growth factor β 1 (TGFβ1) plays a critical role in the epithelial-to-mesenchymal transition (EMT) of renal tubular epithelial cells (TECs) during renal injury, a major cause of acute renal failure, renal fibrosis and obstructive nephropathy. However, the underlying molecular mechanisms remain ill-defined. Here, we addressed this question. Methods: Expression of TGFβ1, Snail, and phosphorylated Stat3 was examined by immunohistochemistry in the kidney after induction of unilateral ureteral obstruction (UUO) in mice. In vitro, primary TECs were purified by flow cytometry, and then challenged with TGFβ1 with/without presence of specific inhibitors for phosphorylation of SMAD3 or Stat3. Protein levels were determined by Western blotting. Results: We detected significant increases in Snail and phosphorylated Stat3, an activated form for Stat3, in the kidney after induction of UUO in mice. In vitro, TGFβ1-challenged primary TECs upregulated Snail, in a SMAD3/Stat3 dependent manner. Conclusion: Our study sheds light on the mechanism underlying the EMT of TECs after renal injury, and suggests Stat3 signaling as a promising innovative therapeutic target for prevention of renal fibrosis.

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Pathological Study on Epithelial-Mesenchymal Transition in Silicotic Lung Lesions in Rat

Veterinary Sciences ◽

10.3390/vetsci6030070 ◽

2019 ◽

Vol 6 (3) ◽

pp. 70 ◽

Cited By ~ 1

Author(s):

Mao Komai ◽

Karin Mihira ◽

Akinori Shimada ◽

Ikumi Miyamoto ◽

Kikumi Ogihara ◽

...

Keyword(s):

Epithelial Cells ◽

Pulmonary Fibrosis ◽

Epithelial Mesenchymal Transition ◽

Crystalline Silicon ◽

Interstitial Fibrosis ◽

Female Rats ◽

Dependent Manner ◽

Type Ii ◽

Mesenchymal Transition ◽

Spindle Cells

Silicosis, caused by the inhalation of crystalline silicon dioxide or silica, is one of the most severe occupational diseases. Persistent inflammation and progressive massive pulmonary fibrosis are the most common histological changes caused by silicosis. Association of epithelial-mesenchymal transition (EMT) of hyperplastic type II epithelial cells with the fibrotic events of pulmonary fibrosis has been suggested in in vitro silica-exposed cultured cell models, patients with idiopathic pulmonary fibrosis, and bleomycin-induced experimental models. Histological features of EMT, however, are not fully described in silicotic lungs in in vivo. The purpose of this study was to demonstrate EMT of hyperplastic type II epithelial cells in the developmental process of progressive massive pulmonary fibrosis in the lungs of rats exposed to silica. F344 female rats were intratracheally instilled with 20 mg of crystalline silica (Min-U-Sil-5), followed by sacrifice at 1, 3, 6, and 12 months after instillation. Fibrosis, characterized by the formation of silicotic nodules, progressive massive fibrosis, and diffuse interstitial fibrosis, was observed in the lungs of the treated rats; the effects of fibrosis intensified in a time-dependent manner. Hyperplasia of the type II epithelial cells, observed in the massive fibrotic lesions, dominated in the lungs of rats at 6 and 12 months after the treatment. Immunohistochemistry of the serial sections of the lung tissues demonstrated positive labeling for cytokeratin, vimentin, and α-smooth muscle actin in spindle cells close to the foci of hyperplasia of type II epithelial cells. Spindle cells, which exhibited features of both epithelial cells and fibroblasts, were also demonstrated with bundles of collagen fibers in the fibrotic lesions, using electron microscopy. Increased expression of TGF-β was shown by Western blotting and immunohistochemistry in the lungs of the treated rats. These findings suggested that enhanced TGF-β expression and EMT of hyperplastic type II epithelial cells are involved in the development process of progressive massive pulmonary fibrosis during silicosis.

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Helicobacter pylori Induces Gastric Epithelial Cell Apoptosis in Association with Increased Fas Receptor Expression

Infection and Immunity ◽

10.1128/iai.67.8.4237-4242.1999 ◽

1999 ◽

Vol 67 (8) ◽

pp. 4237-4242 ◽

Cited By ~ 106

Author(s):

Nicola L. Jones ◽

Andrew S. Day ◽

Hilary A. Jennings ◽

Philip M. Sherman

Keyword(s):

Cell Death ◽

Helicobacter Pylori ◽

Epithelial Cells ◽

Receptor Expression ◽

Gastric Epithelial Cells ◽

Ags Cells ◽

Fas Receptor ◽

Epithelial Cell Apoptosis ◽

H Pylori

ABSTRACT The mechanisms involved in mediating the enhanced gastric epithelial cell apoptosis observed during infection withHelicobacter pylori in vivo are unknown. To determine whether H. pylori directly induces apoptosis of gastric epithelial cells in vitro and to define the role of the Fas-Fas ligand signal transduction cascade, human gastric epithelial cells were infected with H. pylori for up to 72 h under microaerophilic conditions. As assessed by both transmission electron microscopy and fluorescence microscopy, incubation with acagA-positive, cagE-positive, VacA-positive clinical H. pylori isolate stimulated an increase in apoptosis compared to the apoptosis of untreated AGS cells (16.0% ± 2.8% versus 5.9% ± 1.4%, P < 0.05) after 72 h. In contrast, apoptosis was not detected following infection withcagA-negative, cagE-negative, VacA-negative clinical isolates or a Campylobacter jejuni strain. In addition to stimulating apoptosis, infection with H. pylorienhanced Fas receptor expression in AGS cells to a degree comparable to that of treatment with a positive control, gamma interferon (12.5 ng/ml) (148% ± 24% and 167% ± 24% of control, respectively). The enhanced Fas receptor expression was associated with increased sensitivity to Fas-mediated cell death. Ligation of the Fas receptor with an agonistic monoclonal antibody resulted in an increase in apoptosis compared to the apoptosis of cells infected with the bacterium alone (38.5% ± 7.1% versus 16.0% ± 2.8%,P < 0.05). Incubation with neutralizing anti-Fas antibody did not prevent apoptosis of H. pylori-infected cells. Taken together, these findings demonstrate that the gastric pathogen H. pylori stimulates apoptosis of gastric epithelial cells in vitro in association with the enhanced expression of the Fas receptor. These data indicate a role for Fas-mediated signaling in the programmed cell death that occurs in response toH. pylori infection.

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Interleukin-27 Inhibits Epithelial-Mesenchymal Transition in Ovalbumin-Induced Mice Bronchial Epithelial Cells

Journal of Biomaterials and Tissue Engineering ◽

10.1166/jbt.2021.2669 ◽

2021 ◽

Vol 11 (6) ◽

pp. 1129-1137

Author(s):

Yuanyuan Liu ◽

Chao He ◽

Xin Li ◽

Zewen Zhang ◽

Ju Liu ◽

...

Keyword(s):

Epithelial Cells ◽

Epithelial Mesenchymal Transition ◽

Airway Remodeling ◽

Bronchial Epithelial Cell ◽

Phenotypic Marker ◽

Bronchial Epithelial ◽

Mesenchymal Transition ◽

Stat3 Phosphorylation

The epithelial-mesenchymal transition (EMT) of bronchial epithelial cells is a critical mechanism involved in transforming growth factor beta 1 (TGF-β1) induced asthma airway remodeling. Previous study has shown that interleukin 27 (IL-27) attenuates EMT in alveolar epithelial cells, but its effects on the BEAS-2B human bronchial epithelial cell line EMT remain unknown. Herein, we explored the effects of IL-27 on BEAS-2B EMT in vivo and in vitro. In the in vivo experiments, we found that IL-27 nose-drip therapy alleviated airway remodeling, increased the epithelial phenotypic marker epithelial-cadherin (E-cadherin), and decreased the mesenchymal phenotypic marker alpha-smooth muscle actin (α-SMA) compared with the asthmatic control group. We also found that IL-27 suppressed the signal transducer and activator of transcription (STAT3) in the lung tissue of asthmatic mice. in vitro, TGF-β1-induced EMT changes, including downregulation of E-cadherin and upregulation of α-SMA, were suppressed by IL-27 treatment. Additionally, STAT3 phosphorylation was activated by TGF-β1, whereas IL-27 inhibited the activation of TGF-β1 induced STAT3 phosphorylation. Our findings indicated that IL-27 could inhibit airway remodeling by attenuating bronchial epithelial cell EMT in vivo and in vitro. Therefore, IL-27 may be a beneficial therapeutic option targeting asthmatic airway remodeling.

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Helicobacter pylori induces intracellular galectin-8 aggregation around damaged lysosomes within gastric epithelial cells in a host O-glycan-dependent manner

Glycobiology ◽

10.1093/glycob/cwy095 ◽

2018 ◽

Vol 29 (2) ◽

pp. 151-162 ◽

Cited By ~ 6

Author(s):

Fang-Yen Li ◽

I-Chun Weng ◽

Chun-Hung Lin ◽

Mou-Chieh Kao ◽

Ming-Shiang Wu ◽

...

Keyword(s):

Helicobacter Pylori ◽

Epithelial Cells ◽

Rhesus Macaques ◽

Dependent Manner ◽

Gastric Epithelial Cells ◽

Vacuolating Cytotoxin ◽

Infected Cells ◽

H Pylori ◽

Autophagy Activity ◽

Binding Lectin

Abstract Galectin-8, a beta-galactoside-binding lectin, is upregulated in the gastric tissues of rhesus macaques infected with Helicobacter pylori. In this study, we found that H. pylori infection triggers intracellular galectin-8 aggregation in human-derived AGS gastric epithelial cells, and that these aggregates colocalize with lysosomes. Notably, this aggregation is markedly reduced following the attenuation of host O-glycan processing. This indicates that H. pylori infection induces lysosomal damage, which in turn results in the accumulation of cytosolic galectin-8 around damaged lysosomes through the recognition of exposed vacuolar host O-glycans. H. pylori-induced galectin-8 aggregates also colocalize with autophagosomes, and galectin-8 ablation reduces the activation of autophagy by H. pylori. This suggests that galectin-8 aggregates may enhance autophagy activity in infected cells. We also observed that both autophagy and NDP52, an autophagy adapter, contribute to the augmentation of galectin-8 aggregation by H. pylori. Additionally, vacuolating cytotoxin A, a secreted H. pylori cytotoxin, may contribute to the increased galectin-8 aggregation and elevated autophagy response in infected cells. Collectively, these results suggest that H. pylori promotes intracellular galectin-8 aggregation, and that galectin-8 aggregation and autophagy may reciprocally regulate each other during infection.

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196 EPITHELIAL–MESENCHYMAL TRANSITION IN EQUINE AMNIOTIC PROGENITOR CELLS INDUCES CHANGES OF THE CELL GLYCAN PROFILE

Reproduction Fertility and Development ◽

10.1071/rdv26n1ab196 ◽

2014 ◽

Vol 26 (1) ◽

pp. 212

Author(s):

A. Lange-Consiglio ◽

G. Accogli ◽

F. Cremonesi ◽

S. Desantis

Keyword(s):

Epithelial Cells ◽

Progenitor Cells ◽

Binding Sites ◽

Epithelial Mesenchymal Transition ◽

Lectin Binding ◽

Mesenchymal Transition ◽

Glycan Profile ◽

Amniotic Epithelial Cells

Epithelial to mesenchymal transition (EMT) is the process by which epithelial cells dramatically alter their shape and motile behaviour as they differentiate into mesenchymal cells. The EMT and the reverse process, termed mesenchymal–epithelial transition, play central roles in embryogenesis. Gastrulation and neural crest formation are processes governed by EMT in amniotes. It is noteworthy that in placental mammals, the epithelial layer of amnion originates from the trophectoderm and it is continuous with the epiblast. On this basis, it is reasonable to speculate that some amniotic epithelial cells may escape the specification that accompanies gastrulation, and may retain some of the characteristics of epiblastic cells, such as pluripotency, behaving as stem cells that are able to preserve intrinsically the ability to transdifferentiate. Because it seems that malignant cells use the same mechanisms during the formation of tumours in vivo, the amniotic epithelial cells (AEC) could represent a good model to study in vitro this phenomenon that we observed to occur spontaneously in our culture conditions. The aim of this study was to characterise the glycoprotein pattern expressed in fresh or cryopreserved equine AEC, mesenchymal (AMC), and transdifferentiated cells by means of lectin histochemistry. AEC and AMC were cultured until passage (P) 3, while transdifferentiated cells at P1(EMT1) and P2 (EMT2). All cell lines were frozen for 1 month at –196°C in liquid nitrogen. The glycoanalysis was performed with a panel of twelve lectins to detect the glycans terminating with sialic acids (MAL II, SNA, PNA after sialidase digestion (K-s), K-s-DBA), galactose (PNA, RCA120, GSA I-B4,), N-acetylgalactosamine (DBA, HPA, SBA), N-acetylglucosamine (GSA II), fucose (UEA I, LTA), or with internal mannose (Con A). After freezing: 1) AEC exhibited decrease of binding sites for DBA, SBA, HPA, GSA II, and disappearance of GSA I-B4 and UEA I binders; 2) AMC displayed increase of SBA reactivity, decrease of K-s-PNA, HPA, GSA II staining, and absence of GSA I-B4 affinity; 3) EMT1 cells showed the appearance of K-s-DBA staining, the increase of K-s-PNA, RCA120, SBA, GSA I-B4, and UEA I reactivity, the decrease of MAL II, SNA, HPA, GSA II binders, and the disappearance of DBA and LTA binding sites; 4) EMT2 cells revealed the increase of K-s-PNA, GSA I-B4, UEA I affinity, the decrease of MAL II, SNA, RCA120, HPA, GSA II binders, and the lack of DBA, SBA, and LTA reactivity. In conclusion, this study demonstrates that the EMT induces changes in cell surface glycan profile of equine amniotic progenitor cells, and for the first time revealed that freezing modifies the lectin binding pattern of these cells. The observed glycan pattern modification may represent one aspect of the spontaneous complex process of EMT.

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65 Improvement of bovine early embryo development in vitro by coculture with endometrial epithelial cells

Reproduction Fertility and Development ◽

10.1071/rdv31n1ab65 ◽

2019 ◽

Vol 31 (1) ◽

pp. 158

Author(s):

M. Sponchiado ◽

W. F. A. Marei ◽

P. E. J. Bols ◽

M. Binelli ◽

J. L. M. R. Leroy

Keyword(s):

Epithelial Cells ◽

Primary Culture ◽

Epithelial Mesenchymal Transition ◽

Blastocyst Stage ◽

Early Embryo ◽

Phenol Red ◽

Functional Characteristics ◽

Indirect Contact ◽

Mesenchymal Transition

We optimized a bovine endometrial epithelial cell (BEEC) line as a valuable research model for the study of very early embryo-maternal interactions in vitro. In this study, we aimed to (1) characterise the BEEC monolayers along the primary culture and first passages with respect to the expression of epithelial and mesenchymal cell markers and abundance of functional key transcripts; (2) to test whether direct or indirect contact with endometrial cells alter the quality of the embryos in vitro; and (3) to test the specificity of the effect. In Exp. 1, after isolation from slaughterhouse uteri at the early luteal phase, BEEC were cultured in DMEM/F12 phenol red-free medium supplemented with 10% fetal bovine serum (FBS) from primary culture until subculture 3. Fixed samples were immunostained for cytokeratin and vimentin. Transcript abundances for cellular lineage markers (KRT18 and VIM), oestrogen receptor (ESR1), interferon α/beta receptor 1 (IFNAR1), and prostaglandin G/H synthase 1 (PTGS1) and 2 (PTGS2) were evaluated by real-time quantitative PCR. Statistical analyses were carried out by ANOVA and Tukey test. Immunofluorescence data revealed that the BEEC line co-expresses cytokeratin together with a mesenchymal marker (Vimentin). This indicates that these epithelial cells underwent an epithelial-mesenchymal transition in vitro. Gene expression data showed a 6-fold increased (P<0.001) abundance of VIM mRNA from the primary culture to the subculture 1, which remained constant until subculture 3; however, KRT18, ESR1, IFNAR1, PTGS1, and PTGS2 were similar between the passages, suggesting that the cells conserved their functional characteristics. In Exp. 2, groups of 15 morulas (Day 5.5) were cultured in SOF medium supplemented with 5% FBS in the absence (control) or in the presence (co-culture) of BEEC at passage 2, for 48h. Embryos were placed on direct or indirect contact with a BEEC monolayer using a 96-well insert containing 8μm pores. Developmental rates were compared by chi-square test and P-values were adjusted by Tukey’s test. The percentage of embryos that had developed from morula into blastocyst stage on Day 7.5 was significantly higher in the direct and indirect contact co-culture (65%; P<0.05) groups compared with the control (53%) group. Moreover, 63% of the blastocysts were expanded, hatching, or hatched in the co-culture groups, whereas a rate of 46% was found in the control counterparts (P<0.05). In Exp. 3, the same experimental conditions from Exp. 2 were used, but groups of 15 Day 5.5 morulas were cultured in control, or conditioned medium from BEEC (CondBEEC) or bovine fibroblasts (CondFib). Blastocyst development rate on Day 7.5 was higher in the CondBEEC group (71%; P<0.001) compared with the control (54%) and CondFib (50%) groups. In conclusion, based on the markers studied, BEEC monolayers undergo epithelial-mesenchymal transition in vitro but preserve functional characteristics after few passages. The co-culture system improves bovine embryonic development from morula into blastocyst stage. This support is BEEC specific and does not rely on a direct cell-to-embryo contact.

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Wogonoside Inhibits Prostate Cancer Cell Growth and Metastasis via Regulating Wnt/β-Catenin Pathway and Epithelial-Mesenchymal Transition

Pharmacology ◽

10.1159/000502400 ◽

2019 ◽

Vol 104 (5-6) ◽

pp. 312-319 ◽

Cited By ~ 1

Author(s):

Can Wei ◽

Junfeng Jing ◽

Yanbin Zhang ◽

Ling Fang

Keyword(s):

Prostate Cancer ◽

Cell Viability ◽

Epithelial Mesenchymal Transition ◽

Phase Cell ◽

Dependent Manner ◽

Mesenchymal Transition ◽

Pc3 Cells ◽

Gsk 3Β

Background: Wogonoside, an effective component of Scutellaria baicalensis extract, has recently become a hot topic for its newly discovered anticancer efficacy, but the underlying pharmacological mechanism is still unclear. In this study, we tested the inhibitory effects of wogonoside in human prostate cancer PC3 cells in vitro and vivo. Methods: The effects of wogonoside on cell viability, cycle progression, invasion, migration, and apoptosis were assessed in vitro. The levels of proteins in related signaling pathways were detected by western blotting assay. Finally, nude mouse tumorigenicity assay was conducted to detect the anticancer effect of wogonoside in vivo. Results: Wogonoside inhibited cell viability, invasive and migratory ability in a time- and dose-dependent manner. Flow cytometry indicated that wogonoside could induce cell apoptosis and S phase cell-cycle arrest. Mechanically, wogonoside suppressed the Wnt/β-catenin signaling pathway, and the level of p-glycogen synthase kinase-3β (GSK-3β; Ser9) was inhibited by wogonoside. The epithelial-mesenchymal transition (EMT) process was also reversed in PC3 cell line after wogonoside treatment. In vivo experiments showed that wogonoside inhibited tumor growth in xenograft mouse models. Conclusion: These findings revealed that wogonoside could suppress Wnt/β-catenin pathway and reversing the EMT process in PC3 cells. GSK-3β acts as a tumor suppressor in prostate cancer. Wogonoside may serve as an effective agent for treating prostate cancer.

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