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Sample records for epithelial cell invasion

  1. Nerve Invasion by Epithelial Cells in Benign Breast Diseases

    Directory of Open Access Journals (Sweden)

    Yu-Jan Chan

    2009-03-01

    Full Text Available Nerve invasion by glandular epithelial cells in a lesion is usually regarded as invasive carcinoma. However, some benign conditions in the pancreas, prostate, breast and other organs may show involvement of nerve bundles by benign epithelial cells. We report an 18-year-old female with nerve invasion in benign breast disease. The lesion in her right breast revealed fibrocystic changes with ductal hyperplasia and stromal sclerosis. Perineural and intraneural involvement by bland-looking small ducts lined by 2 layers of cells including an outer layer of myoepithelial cells were found, suggestive of benign nerve invasion. There was no evidence of malignant cells in any of the sections. The patient remains well after 31 months of follow-up. About 44 cases of nerve invasion in benign breast diseases have been reported in the literature. It is necessary to carefully evaluate nerve involvement in breast lesions to avoid over-diagnosis and inappropriate operation.

  2. Invasion of Human Oral Epithelial Cells by Prevotella intermedia

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    Dorn, Brian R.; Leung, K.-P.; Progulske-Fox, Ann

    1998-01-01

    Invasion of oral epithelial cells by pathogenic oral bacteria may represent an important virulence factor in the progression of periodontal disease. Here we report that a clinical isolate of Prevotella intermedia, strain 17, was found to invade a human oral epithelial cell line (KB), whereas P. intermedia 27, another clinical isolate, and P. intermedia 25611, the type strain, were not found to invade the cell line. Invasion was quantified by the recovery of viable bacteria following a standard antibiotic protection assay and observed by electron microscopy. Cytochalasin D, cycloheximide, monodansylcadaverine, and low temperature (4°C) inhibited the internalization of P. intermedia 17. Antibodies raised against P. intermedia type C fimbriae and against whole cells inhibited invasion, but the anti-type-C-fimbria antibody inhibited invasion to a greater extent than the anti-whole-cell antibody. This work provides evidence that at least one strain of P. intermedia can invade an oral epithelial cell line and that the type C fimbriae and a cytoskeletal rearrangement are required for this invasion. PMID:9826397

  3. The genetic and molecular basis of bacterial invasion of epithelial cells

    African Journals Online (AJOL)

    Invasion of epithelial cells was demonstrated to be triggered by invasion plasmid antigens B, C, and D ( IpaB, IpaC and IpaD ) which is accomplished by intracellular spread gene icsA. The invasion of epithelial cells by some individual species of bacteria were also reviewed.Yersinia enterocolitica invasiveness was shown ...

  4. The Inside Story of Shigella Invasion of Intestinal Epithelial Cells

    Science.gov (United States)

    Carayol, Nathalie; Tran Van Nhieu, Guy

    2013-01-01

    As opposed to other invasive pathogens that reside into host cells in a parasitic mode, Shigella, the causative agent of bacillary dysentery, invades the colonic mucosa but does not penetrate further to survive into deeper tissues. Instead, Shigella invades, replicates, and disseminates within the colonic mucosa. Bacterial invasion and spreading in intestinal epithelium lead to the elicitation of inflammatory responses responsible for the tissue destruction and shedding in the environment for further infection of other hosts. In this article, we highlight specific features of the Shigella arsenal of virulence determinants injected by a type III secretion apparatus (T3SA) that point to the targeting of intestinal epithelial cells as a discrete route of invasion during the initial event of the infectious process. PMID:24086068

  5. Helicobacter pylori induces cell migration and invasion through casein kinase 2 in gastric epithelial cells.

    Science.gov (United States)

    Lee, Yeo Song; Lee, Do Yeon; Yu, Da Yeon; Kim, Shin; Lee, Yong Chan

    2014-12-01

    Chronic infection with Helicobacter pylori (H. pylori) is causally linked with gastric carcinogenesis. Virulent H. pylori strains deliver bacterial CagA into gastric epithelial cells. Induction of high motility and an elongated phenotype is considered to be CagA-dependent process. Casein kinase 2 plays a critical role in carcinogenesis through signaling pathways related to the epithelial mesenchymal transition. This study was aimed to investigate the effect of H. pylori infection on the casein kinase 2-mediated migration and invasion in gastric epithelial cells. AGS or MKN28 cells as human gastric epithelial cells and H. pylori strains Hp60190 (ATCC 49503, CagA(+)) and Hp8822 (CagA(-)) were used. Cells were infected with H. pylori at multiplicity of infection of 100 : 1 for various times. We measured in vitro kinase assay to examine casein kinase 2 activity and performed immunofluorescent staining to observe E-cadherin complex. We also examined β-catenin transactivation through promoter assay and MMP7 expression by real-time PCR and ELISA. H. pylori upregulates casein kinase 2 activity and inhibition of casein kinase 2 in H. pylori-infected cells profoundly suppressed cell invasiveness and motility. We confirmed that casein kinase 2 mediates membranous α-catenin depletion through dissociation of the α-/β-catenin complex in H. pylori-infected cells. We also found that H. pylori induces β-catenin nuclear translocation and increases MMP7 expressions mediated through casein kinase 2. We show for the first time that CagA(+) H. pylori upregulates cellular invasiveness and motility through casein kinase 2. The demonstration of a mechanistic interplay between H. pylori and casein kinase 2 provides important insights into the role of CagA(+) H. pylori in the gastric cancer invasion and metastasis. © 2014 John Wiley & Sons Ltd.

  6. Hakai reduces cell-substratum adhesion and increases epithelial cell invasion

    International Nuclear Information System (INIS)

    Rodríguez-Rigueiro, Teresa; Valladares-Ayerbes, Manuel; Haz-Conde, Mar; Aparicio, Luis A; Figueroa, Angélica

    2011-01-01

    The dynamic regulation of cell-cell adhesions is crucial for developmental processes, including tissue formation, differentiation and motility. Adherens junctions are important components of the junctional complex between cells and are necessary for maintaining cell homeostasis and normal tissue architecture. E-cadherin is the prototype and best-characterized protein member of adherens junctions in mammalian epithelial cells. Regarded as a tumour suppressor, E-cadherin loss is associated with poor prognosis in carcinoma. The E3 ubiquitin-ligase Hakai was the first reported posttranslational regulator of the E-cadherin complex. Hakai specifically targetted E-cadherin for internalization and degradation and thereby lowered epithelial cell-cell contact. Hakai was also implicated in controlling proliferation, and promoted cancer-related gene expression by increasing the binding of RNA-binding protein PSF to RNAs encoding oncogenic proteins. We sought to investigate the possible implication of Hakai in cell-substratum adhesions and invasion in epithelial cells. Parental MDCK cells and MDCK cells stably overexpressing Hakai were used to analyse cell-substratum adhesion and invasion capabilities. Western blot and immunofluoresecence analyses were performed to assess the roles of Paxillin, FAK and Vinculin in cell-substratum adhesion. The role of the proteasome in controlling cell-substratum adhesion was studied using two proteasome inhibitors, lactacystin and MG132. To study the molecular mechanisms controlling Paxillin expression, MDCK cells expressing E-cadherin shRNA in a tetracycline-inducible manner was employed. Here, we present evidence that implicate Hakai in reducing cell-substratum adhesion and increasing epithelial cell invasion, two hallmark features of cancer progression and metastasis. Paxillin, an important protein component of the cell-matrix adhesion, was completely absent from focal adhesions and focal contacts in Hakai-overexpressing MDCK cells. The

  7. Myoferlin depletion in breast cancer cells promotes mesenchymal to epithelial shape change and stalls invasion.

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    Ruth Li

    Full Text Available Myoferlin (MYOF is a mammalian ferlin protein with homology to ancestral Fer-1, a nematode protein that regulates spermatic membrane fusion, which underlies the amoeboid-like movements of its sperm. Studies in muscle and endothelial cells have reported on the role of myoferlin in membrane repair, endocytosis, myoblast fusion, and the proper expression of various plasma membrane receptors. In this study, using an in vitro human breast cancer cell model, we demonstrate that myoferlin is abundantly expressed in invasive breast tumor cells. Depletion of MYOF using lentiviral-driven shRNA expression revealed that MDA-MB-231 cells reverted to an epithelial morphology, suggesting at least some features of mesenchymal to epithelial transition (MET. These observations were confirmed by the down-regulation of some mesenchymal cell markers (e.g., fibronectin and vimentin and coordinate up-regulation of the E-cadherin epithelial marker. Cell invasion assays using Boyden chambers showed that loss of MYOF led to a significant diminution in invasion through Matrigel or type I collagen, while cell migration was unaffected. PCR array and screening of serum-free culture supernatants from shRNA(MYOF transduced MDA-MB-231 cells indicated a significant reduction in the steady-state levels of several matrix metalloproteinases. These data when considered in toto suggest a novel role of MYOF in breast tumor cell invasion and a potential reversion to an epithelial phenotype upon loss of MYOF.

  8. Reactive oxygen species induced by Streptococcus pyogenes invasion trigger apoptotic cell death in infected epithelial cells.

    Science.gov (United States)

    Aikawa, Chihiro; Nozawa, Takashi; Maruyama, Fumito; Tsumoto, Kohei; Hamada, Shigeyuki; Nakagawa, Ichiro

    2010-06-01

    Streptococcus pyogenes (group A streptococcus, GAS), one of the most common pathogens of humans, attaches and invades into human pharyngeal or skin epithelial cells. We have previously reported that induction of apoptosis is associated with GAS invasion, which induces mitochondrial dysfunction and apoptotic cell death. We demonstrate here that GAS-induced apoptosis is mediated by reactive oxygen species (ROS) production. Both the induction of apoptosis and ROS production markedly increased upon invasion of wild-type GAS strain JRS4 into HeLa cells; however, the apoptotic response was not observed in fibronectin-binding protein F1-disrupted mutant SAM1-infected cells. In Bcl-2-overexpressing HeLa cells (HBD98-2-4), the induction of apoptosis, ROS production and mitochondrial dysfunction were significantly suppressed, whereas the numbers of invaded GAS was not different between HeLa (mock cells) and the HeLa HBD98-2-4 cells. Whereas Rac1 activation occurred during GAS invasion, ROS production in GAS-infected cells was clearly inhibited by transfection with the Rac1 mutants (L37 or V12L37), but not by the dominant active mutant (V12L61) or by the dominant negative mutant (N17). These observations indicate that GAS invasion triggers ROS production through Rac1 activation and generated ROS induced mitochondrial dysfunction leading to cellular apoptosis.

  9. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    Energy Technology Data Exchange (ETDEWEB)

    Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing, E-mail: caijingmmm@hotmail.com; Wang, Zehua, E-mail: zehuawang@163.net

    2015-09-10

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs.

  10. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    International Nuclear Information System (INIS)

    Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing; Wang, Zehua

    2015-01-01

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs

  11. FRK inhibits breast cancer cell migration and invasion by suppressing epithelial-mesenchymal transition.

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    Ogunbolude, Yetunde; Dai, Chenlu; Bagu, Edward T; Goel, Raghuveera Kumar; Miah, Sayem; MacAusland-Berg, Joshua; Ng, Chi Ying; Chibbar, Rajni; Napper, Scott; Raptis, Leda; Vizeacoumar, Frederick; Vizeacoumar, Franco; Bonham, Keith; Lukong, Kiven Erique

    2017-12-22

    The human fyn-related kinase (FRK) is a non-receptor tyrosine kinase known to have tumor suppressor activity in breast cancer cells. However, its mechanism of action has not been fully characterized. We generated FRK-stable MDA-MB-231 breast cancer cell lines and analyzed the effect on cell proliferation, migration, and invasiveness. We also used kinome analysis to identify potential FRK-regulated signaling pathways. We employed both immunoblotting and RT-PCR to identify/validate FRK-regulated targets (proteins and genes) in these cells. Finally, we interrogated the TCGA and GENT gene expression databases to determine the correlation between the expression of FRK and epithelial/mesenchymal markers. We observed that FRK overexpression suppressed cell proliferation, migration, and invasiveness, inhibited various JAK/STAT, MAPK and Akt signaling pathways, and suppressed the expression of some STAT3 target genes. Also, FRK overexpression increased the expression of epithelial markers including E-cadherin mRNA and down-regulated the transcript levels of vimentin, fibronectin, and slug. Finally, we observed an inverse correlation between FRK expression and mesenchymal markers in a large cohort of breast cancer cells. Our data, therefore, suggests that FRK represses cell proliferation, migration and invasiveness by suppressing epithelial to mesenchymal transition.

  12. Macropinosomes are Key Players in Early Shigella Invasion and Vacuolar Escape in Epithelial Cells.

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    Allon Weiner

    2016-05-01

    Full Text Available Intracellular pathogens include all viruses, many bacteria and parasites capable of invading and surviving within host cells. Key to survival is the subversion of host cell pathways by the pathogen for the purpose of propagation and evading the immune system. The intracellular bacterium Shigella flexneri, the causative agent of bacillary dysentery, invades host cells in a vacuole that is subsequently ruptured to allow growth of the pathogen within the host cytoplasm. S. flexneri invasion has been classically described as a macropinocytosis-like process, however the underlying details and the role of macropinosomes in the intracellular bacterial lifestyle have remained elusive. We applied dynamic imaging and advanced large volume correlative light electron microscopy (CLEM to study the highly transient events of S. flexneri's early invasion into host epithelial cells and elucidate some of its fundamental features. First, we demonstrate a clear distinction between two compartments formed during the first step of invasion: the bacterial containing vacuole and surrounding macropinosomes, often considered identical. Next, we report a functional link between macropinosomes and the process of vacuolar rupture, demonstrating that rupture timing is dependent on the availability of macropinosomes as well as the activity of the small GTPase Rab11 recruited directly to macropinosomes. We go on to reveal that the bacterial containing vacuole and macropinosomes come into direct contact at the onset of vacuolar rupture. Finally, we demonstrate that S. flexneri does not subvert pre-existing host endocytic vesicles during the invasion steps leading to vacuolar rupture, and propose that macropinosomes are the major compartment involved in these events. These results provide the basis for a new model of the early steps of S. flexneri epithelial cell invasion, establishing a different view of the enigmatic process of cytoplasmic access by invasive bacterial

  13. Dielectric spectroscopy for non-invasive monitoring of epithelial cell differentiation within three-dimensional scaffolds

    International Nuclear Information System (INIS)

    Daoud, Jamal; Tabrizian, Maryam; Asami, Koji; Rosenberg, Lawrence

    2012-01-01

    In this study, we introduce a cellular differentiation cellular model based on dielectric spectroscopy that characterizes epithelial differentiation processes. Non-invasive cellular monitoring was achieved within a three-dimensional microenvironment consisting of a cell-containing collagen I gel seeded onto microfabricated scaffolds. In this proof-of-concept investigation, Madin–Darby canine kidney cells were cultured within microfabricated, geometrically controlled scaffolds and allowed us to differentiate to hollow cyst-like structures. This transformation within the three-dimensional environment is monitored and characterized through dielectric spectroscopy while maintaining cell culture in vitro. (paper)

  14. How Shigella Utilizes Ca(2+) Jagged Edge Signals during Invasion of Epithelial Cells.

    Science.gov (United States)

    Bonnet, Mariette; Tran Van Nhieu, Guy

    2016-01-01

    Shigella, the causative agent of bacillary dysentery invades intestinal epithelial cells using a type III secretion system (T3SS). Through the injection of type III effectors, Shigella manipulates the actin cytoskeleton to induce its internalization in epithelial cells. At early invasion stages, Shigella induces atypical Ca(2+) responses confined at entry sites allowing local cytoskeletal remodeling for bacteria engulfment. Global Ca(2+) increase in the cell triggers the opening of connexin hemichannels at the plasma membrane that releases ATP in the extracellular milieu, favoring Shigella invasion and spreading through purinergic receptor signaling. During intracellular replication, Shigella regulates inflammatory and death pathways to disseminate within the epithelium. At later stages of infection, Shigella downregulates hemichannel opening and the release of extracellular ATP to dampen inflammatory signals. To avoid premature cell death, Shigella activates cell survival by upregulating the PI3K/Akt pathway and downregulating the levels of p53. Furthermore, Shigella interferes with pro-apoptotic caspases, and orients infected cells toward a slow necrotic cell death linked to mitochondrial Ca(2+) overload. In this review, we will focus on the role of Ca(2+) responses and their regulation by Shigella during the different stages of bacterial infection.

  15. How Shigella Utilizes Ca2+ Jagged Edge Signals during Invasion of Epithelial Cells

    Science.gov (United States)

    Bonnet, Mariette; Tran Van Nhieu, Guy

    2016-01-01

    Shigella, the causative agent of bacillary dysentery invades intestinal epithelial cells using a type III secretion system (T3SS). Through the injection of type III effectors, Shigella manipulates the actin cytoskeleton to induce its internalization in epithelial cells. At early invasion stages, Shigella induces atypical Ca2+ responses confined at entry sites allowing local cytoskeletal remodeling for bacteria engulfment. Global Ca2+ increase in the cell triggers the opening of connexin hemichannels at the plasma membrane that releases ATP in the extracellular milieu, favoring Shigella invasion and spreading through purinergic receptor signaling. During intracellular replication, Shigella regulates inflammatory and death pathways to disseminate within the epithelium. At later stages of infection, Shigella downregulates hemichannel opening and the release of extracellular ATP to dampen inflammatory signals. To avoid premature cell death, Shigella activates cell survival by upregulating the PI3K/Akt pathway and downregulating the levels of p53. Furthermore, Shigella interferes with pro-apoptotic caspases, and orients infected cells toward a slow necrotic cell death linked to mitochondrial Ca2+ overload. In this review, we will focus on the role of Ca2+ responses and their regulation by Shigella during the different stages of bacterial infection. PMID:26904514

  16. How Shigella utilizes Ca2+ jagged edge signals during invasion of epithelial cells

    Directory of Open Access Journals (Sweden)

    Mariette eBonnet

    2016-02-01

    Full Text Available Shigella, the causative agent of bacillary dysentery invades intestinal epithelial cells using a type III secretion system (T3SS. Through the injection of type III effectors, Shigella manipulates the actin cytoskeleton to induce its internalization in epithelial cells. At early invasion stages, Shigella induces atypical Ca2+ responses confined at entry sites allowing local cytoskeletal remodeling for bacteria engulfment. Global Ca2+ increase in the cell triggers the opening of connexin hemichannels at the plasma membrane that releases ATP in the extracellular milieu, favoring Shigella invasion and spreading through purinergic receptor signaling. During intracellular replication, Shigella regulates inflammatory and death pathways to disseminate within the epithelium. At later stages of infection, Shigella downregulates hemichannel opening and the release of extracellular ATP to dampen inflammatory signals. To avoid premature cell death, Shigella activates cell survival by upregulating the PI3K/Akt pathway and downregulating the levels of p53. Furthermore, Shigella interferes with pro-apoptotic caspases, and orients infected cells towards a slow necrotic cell death linked to mitochondrial Ca2+ overload. In this review, we will focus on the role of Ca2+ responses and their regulation by Shigella during the different stages of bacterial infection.

  17. Adipocytes Promote B16BL6 Melanoma Cell Invasion and the Epithelial-to-Mesenchymal Transition

    OpenAIRE

    Kushiro, Kyoko; Chu, Randy A.; Verma, Akanksha; Núñez, Nomelí P.

    2011-01-01

    Metastatic melanoma is one of the most deadly and evasive types of cancer. On average, cancer patients with metastatic melanoma survive only 6–9 months after diagnosis. Epidemiological and animal studies suggest that obesity increases the metastatic ability of malignant melanoma, though the mechanism is not known. In the present studies, we assessed the ability of 3T3L1 adipocytes to modulate B16BL6 melanoma cell invasion and the Epithelial-to-Mesenchymal Transition (EMT). For this purpose, w...

  18. Lactoferrin affects the adherence and invasion of Streptococcus dysgalactiae ssp. dysgalactiae in mammary epithelial cells.

    Science.gov (United States)

    O'Halloran, Fiona; Beecher, Christine; Chaurin, Valerie; Sweeney, Torres; Giblin, Linda

    2016-06-01

    Streptococcus dysgalactiae ssp. dysgalactiae is an important causative agent of bovine mastitis worldwide. Lactoferrin is an innate immune protein that is associated with many functions including immunomodulatory, antiproliferative, and antimicrobial properties. This study aimed to investigate the interactions between lactoferrin and a clinical bovine mastitis isolate, Strep. dysgalactiae ssp. dysgalactiae DPC5345. Initially a deliberate in vivo bovine intramammary challenge was performed with Strep. dysgalactiae DPC5345. Results demonstrated a significant difference in lactoferrin mRNA levels in milk cells between the control and infused quarters 7h postinfusion. Milk lactoferrin levels in the Strep. dysgalactiae DPC5345 infused quarters were significantly increased compared with control quarters at 48h postinfusion. In vitro studies demonstrated that lactoferrin had a bacteriostatic effect on the growth of Strep. dysgalactiae DPC5345 and significantly decreased the ability of the bacteria to internalize into HC-11 mammary epithelial cells. Confocal microscopy images of HC-11 cells exposed to Strep. dysgalactiae and lactoferrin further supported this effect by demonstrating reduced invasion of bacteria to HC-11 cells. The combined data suggest that a bovine immune response to Strep. dysgalactiae infection includes a significant increase in lactoferrin expression in vivo, and based on in vitro data, lactoferrin limits mammary cell invasion of this pathogen by binding to the bacteria and preventing its adherence. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  19. MUC1 enhances invasiveness of pancreatic cancer cells by inducing epithelial to mesenchymal transition.

    Science.gov (United States)

    Roy, L D; Sahraei, M; Subramani, D B; Besmer, D; Nath, S; Tinder, T L; Bajaj, E; Shanmugam, K; Lee, Y Y; Hwang, S I L; Gendler, S J; Mukherjee, P

    2011-03-24

    Increased motility and invasiveness of pancreatic cancer cells are associated with epithelial to mesenchymal transition (EMT). Snai1 and Slug are zinc-finger transcription factors that trigger this process by repressing E-cadherin and enhancing vimentin and N-cadherin protein expression. However, the mechanisms that regulate this activation in pancreatic tumors remain elusive. MUC1, a transmembrane mucin glycoprotein, is associated with the most invasive forms of pancreatic ductal adenocarcinomas (PDA). In this study, we show that over expression of MUC1 in pancreatic cancer cells triggers the molecular process of EMT, which translates to increased invasiveness and metastasis. EMT was significantly reduced when MUC1 was genetically deleted in a mouse model of PDA or when all seven tyrosines in the cytoplasmic tail of MUC1 were mutated to phenylalanine (mutated MUC1 CT). Using proteomics, RT-PCR and western blotting, we revealed a significant increase in vimentin, Slug and Snail expression with repression of E-Cadherin in MUC1-expressing cells compared with cells expressing the mutated MUC1 CT. In the cells that carried the mutated MUC1 CT, MUC1 failed to co-immunoprecipitate with β-catenin and translocate to the nucleus, thereby blocking transcription of the genes associated with EMT and metastasis. Thus, functional tyrosines are critical in stimulating the interactions between MUC1 and β-catenin and their nuclear translocation to initiate the process of EMT. This study signifies the oncogenic role of MUC1 CT and is the first to identify a direct role of the MUC1 in initiating EMT during pancreatic cancer. The data may have implications in future design of MUC1-targeted therapies for pancreatic cancer.

  20. IL1β-mediated Stromal COX-2 signaling mediates proliferation and invasiveness of colonic epithelial cancer cells

    International Nuclear Information System (INIS)

    Zhu, Yingting; Zhu, Min; Lance, Peter

    2012-01-01

    COX-2 is a major inflammatory mediator implicated in colorectal inflammation and cancer. However, the exact origin and role of COX-2 on colorectal inflammation and carcinogenesis are still not well defined. Recently, we reported that COX-2 and iNOS signalings interact in colonic CCD18Co fibroblasts. In this article, we investigated whether activation of COX-2 signaling by IL1β in primary colonic fibroblasts obtained from normal and cancer patients play a critical role in regulation of proliferation and invasiveness of human colonic epithelial cancer cells. Our results demonstrated that COX-2 level was significantly higher in cancer associated fibroblasts than that in normal fibroblasts with or without stimulation of IL-1β, a powerful stimulator of COX-2. Using in vitro assays for estimating proliferative and invasive potential, we discovered that the proliferation and invasiveness of the epithelial cancer cells were much greater when the cells were co-cultured with cancer associated fibroblasts than with normal fibroblasts, with or without stimulation of IL1β. Further analysis indicated that the major COX-2 product, prostaglandin E 2 , directly enhanced proliferation and invasiveness of the epithelial cancer cells in the absence of fibroblasts. Moreover, a selective COX-2 inhibitor, NS-398, blocked the proliferative and invasive effect of both normal and cancer associate fibroblasts on the epithelial cancer cells, with or without stimulation of IL-1β. Those results indicate that activation of COX-2 signaling in the fibroblasts plays a major role in promoting proliferation and invasiveness of the epithelial cancer cells. In this process, PKC is involved in the activation of COX-2 signaling induced by IL-1β in the fibroblasts.

  1. Effects of vitamin D and its metabolites on cell viability and Staphylococcus aureus invasion in bovine mammary epithelial cells

    DEFF Research Database (Denmark)

    Yue, Yuan; Hymøller, Lone; Jensen, Søren Krogh

    2017-01-01

    Vitamin D has been found have various biological effects that may be potent in preventing bovine mastitis. Two forms of vitamin D, vitamin D2 (D2) and vitamin D3 (D3), can be hydroxylated to functional metabolites in cattle. The objectives of the present study were to investigate the effects of D2...... and D3 compounds on bovine mammary epithelial cell proliferation and Staphylococcus aureus (S. aureus) invasion.. Results showed that 1,25-dihydroxyvitamin D2 have an anti-proliferation activity comparable to 1,25-dihydroxyvitamin D3, while D2 and 25-hydroxyvitamin D2 (25(OH)D2) was slightly more potent...

  2. Regulation of the actin cytoskeleton in Helicobacter pylori-induced migration and invasive growth of gastric epithelial cells

    Directory of Open Access Journals (Sweden)

    Rieder Gabriele

    2011-11-01

    Full Text Available Abstract Dynamic rearrangement of the actin cytoskeleton is a significant hallmark of Helicobacter pylori (H. pylori infected gastric epithelial cells leading to cell migration and invasive growth. Considering the cellular mechanisms, the type IV secretion system (T4SS and the effector protein cytotoxin-associated gene A (CagA of H. pylori are well-studied initiators of distinct signal transduction pathways in host cells targeting kinases, adaptor proteins, GTPases, actin binding and other proteins involved in the regulation of the actin lattice. In this review, we summarize recent findings of how H. pylori functionally interacts with the complex signaling network that controls the actin cytoskeleton of motile and invasive gastric epithelial cells.

  3. Identification of H-Ras-Specific Motif for the Activation of Invasive Signaling Program in Human Breast Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Hae-Young Yong

    2011-02-01

    Full Text Available Increased expression and/or activation of H-Ras are often associated with tumor aggressiveness in breast cancer. Previously, we showed that H-Ras, but not N-Ras, induces MCF10A human breast epithelial cell invasion and migration, whereas both H-Ras and N-Ras induce cell proliferation and phenotypic transformation. In an attempt to determine the sequence requirement directing the divergent phenotype induced by H-Ras and N-Ras with a focus on the induction of human breast cell invasion, we investigated the structural and functional relationships between H-Ras and N-Ras using domain-swap and site-directed mutagenesis approaches. Here, we report that the hypervariable region (HVR, consisting of amino acids 166 to 189 in H-Ras, determines the invasive/migratory signaling program as shown by the exchange of invasive phenotype by swapping HVR sequences between H-Ras and N-Ras. We also demonstrate that the H-Ras-specific additional palmitoylation site at Cys184 is not responsible for the signaling events that distinguish between H-Ras and N-Ras. Importantly, this work identifies the C-terminal HVR, especially the flexible linker domain with two consecutive proline residues Pro173 and Pro174, as a critical domain that contributes to activation of H-Ras and its invasive potential in human breast epithelial cells. The present study sheds light on the structural basis for the Ras isoform-specific invasive program of breast epithelial cells, providing information for the development of agents that specifically target invasion-related H-Ras pathways in human cancer.

  4. AGE-modified basement membrane cooperates with Endo180 to promote epithelial cell invasiveness and decrease prostate cancer survival

    DEFF Research Database (Denmark)

    Rodriguez-Teja, Mercedes; Gronau, Julian H; Breit, Claudia

    2015-01-01

    Biomechanical strain imposed by age-related thickening of the basal lamina and augmented tissue stiffness in the prostate gland coincides with increased cancer risk. Here we hypothesized that the structural alterations in the basal lamina associated with age can induce mechanotransduction pathways...... in prostate epithelial cells (PECs) to promote invasiveness and cancer progression. To demonstrate this, we developed a 3D model of PEC acini in which thickening and stiffening of basal lamina matrix was induced by advanced glycation end-product (AGE)-dependent non-enzymatic crosslinking of its major......(Δ) (Ex2-6/) (Δ) (Ex2-6) mice, with constitutively exposed CTLD2 and decreased survival of men with early (non-invasive) prostate cancer with high epithelial Endo180 expression and levels of AGE. These findings indicate that AGE-dependent modification of the basal lamina induces invasive behaviour...

  5. CD147, CD44, and the epidermal growth factor receptor (EGFR) signaling pathway cooperate to regulate breast epithelial cell invasiveness.

    Science.gov (United States)

    Grass, G Daniel; Tolliver, Lauren B; Bratoeva, Momka; Toole, Bryan P

    2013-09-06

    The immunoglobulin superfamily glycoprotein CD147 (emmprin; basigin) is associated with an invasive phenotype in various types of cancers, including malignant breast cancer. We showed recently that up-regulation of CD147 in non-transformed, non-invasive breast epithelial cells is sufficient to induce an invasive phenotype characterized by membrane type-1 matrix metalloproteinase (MT1-MMP)-dependent invadopodia activity (Grass, G. D., Bratoeva, M., and Toole, B. P. (2012) Regulation of invadopodia formation and activity by CD147. J. Cell Sci. 125, 777-788). Here we found that CD147 induces breast epithelial cell invasiveness by promoting epidermal growth factor receptor (EGFR)-Ras-ERK signaling in a manner dependent on hyaluronan-CD44 interaction. Furthermore, CD147 promotes assembly of signaling complexes containing CD147, CD44, and EGFR in lipid raftlike domains. We also found that oncogenic Ras regulates CD147 expression, hyaluronan synthesis, and formation of CD147-CD44-EGFR complexes, thus forming a positive feedback loop that may amplify invasiveness. Last, we showed that malignant breast cancer cells are heterogeneous in their expression of surface-associated CD147 and that high levels of membrane CD147 correlate with cell surface EGFR and CD44 levels, activated EGFR and ERK1, and activated invadopodia. Future studies should evaluate CD147 as a potential therapeutic target and disease stratification marker in breast cancer.

  6. CD147, CD44, and the Epidermal Growth Factor Receptor (EGFR) Signaling Pathway Cooperate to Regulate Breast Epithelial Cell Invasiveness*

    Science.gov (United States)

    Grass, G. Daniel; Tolliver, Lauren B.; Bratoeva, Momka; Toole, Bryan P.

    2013-01-01

    The immunoglobulin superfamily glycoprotein CD147 (emmprin; basigin) is associated with an invasive phenotype in various types of cancers, including malignant breast cancer. We showed recently that up-regulation of CD147 in non-transformed, non-invasive breast epithelial cells is sufficient to induce an invasive phenotype characterized by membrane type-1 matrix metalloproteinase (MT1-MMP)-dependent invadopodia activity (Grass, G. D., Bratoeva, M., and Toole, B. P. (2012) Regulation of invadopodia formation and activity by CD147. J. Cell Sci. 125, 777–788). Here we found that CD147 induces breast epithelial cell invasiveness by promoting epidermal growth factor receptor (EGFR)-Ras-ERK signaling in a manner dependent on hyaluronan-CD44 interaction. Furthermore, CD147 promotes assembly of signaling complexes containing CD147, CD44, and EGFR in lipid raftlike domains. We also found that oncogenic Ras regulates CD147 expression, hyaluronan synthesis, and formation of CD147-CD44-EGFR complexes, thus forming a positive feedback loop that may amplify invasiveness. Last, we showed that malignant breast cancer cells are heterogeneous in their expression of surface-associated CD147 and that high levels of membrane CD147 correlate with cell surface EGFR and CD44 levels, activated EGFR and ERK1, and activated invadopodia. Future studies should evaluate CD147 as a potential therapeutic target and disease stratification marker in breast cancer. PMID:23888049

  7. Recombinant invasive Lactococcus lactis can transfer DNA vaccines either directly to dendritic cells or across an epithelial cell monolayer.

    Science.gov (United States)

    de Azevedo, Marcela; Meijerink, Marjolein; Taverne, Nico; Pereira, Vanessa Bastos; LeBlanc, Jean Guy; Azevedo, Vasco; Miyoshi, Anderson; Langella, Philippe; Wells, Jerry M; Chatel, Jean-Marc

    2015-09-11

    Lactococcus lactis (L. lactis), a generally regarded as safe (GRAS) bacterium has recently been investigated as a mucosal delivery vehicle for DNA vaccines. Because of its GRAS status, L. lactis represents an attractive alternative to attenuated pathogens. Previous studies showed that eukaryotic expression plasmids could be delivered into intestinal epithelial cells (IECs) by L. lactis, or recombinant invasive strains of L. lactis, leading to heterologous protein expression. Although expression of antigens in IECs might lead to vaccine responses, it would be of interest to know whether uptake of L. lactis DNA vaccines by dendritic cells (DCs) could lead to antigen expression as they are unique in their ability to induce antigen-specific T cell responses. To test this, we incubated mouse bone marrow-derived DCs (BMDCs) with invasive L. lactis strains expressing either Staphylococcus aureus Fibronectin Binding Protein A (LL-FnBPA+), or Listeria monocytogenes mutated Internalin A (LL-mInlA+), both strains carrying a plasmid DNA vaccine (pValac) encoding for the cow milk allergen β-lactoglobulin (BLG). We demonstrated that they can transfect BMDCs, inducing the secretion of the pro-inflammatory cytokine IL-12. We also measured the capacity of strains to invade a polarized monolayer of IECs, mimicking the situation encountered in the gastrointestinal tract. Gentamycin survival assay in these cells showed that LL-mInlA+ is 100 times more invasive than L. lactis. The cross-talk between differentiated IECs, BMDCs and bacteria was also evaluated using an in vitro transwell co-culture model. Co-incubation of strains in this model showed that DCs incubated with LL-mInlA+ containing pValac:BLG could express significant levels of BLG. These results suggest that DCs could sample bacteria containing the DNA vaccine across the epithelial barrier and express the antigen. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  8. Ionizing Radiation Promotes Migration and Invasion of Cancer Cells Through Transforming Growth Factor-Beta-Mediated Epithelial-Mesenchymal Transition

    Energy Technology Data Exchange (ETDEWEB)

    Zhou Yongchun [Department of Radiation Oncology, Xijing Hospital Fourth Military Medical University, Xi' an (China); Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China); Liu Junye; Li Jing; Zhang Jie [Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China); Xu Yuqiao [Department of Pathology, Xijing Hospital Fourth Military Medical University, Xi' an (China); Zhang Huawei; Qiu Lianbo; Ding Guirong [Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China); Su Xiaoming [Department of Radiation Oncology, 306th Hospital of PLA, Beijing (China); Mei Shi [Department of Radiation Oncology, Xijing Hospital Fourth Military Medical University, Xi' an (China); Guo Guozhen, E-mail: guozhenguo@hotmail.com [Department of Radiation Medicine, College of Preventive Medicine, Xijing Hospital Fourth Military Medical University, Xi' an (China)

    2011-12-01

    Purpose: To examine whether ionizing radiation enhances the migratory and invasive abilities of cancer cells through transforming growth factor (TGF-{beta})-mediated epithelial-mesenchymal transition (EMT). Methods and Materials: Six cancer cell lines originating from different human organs were irradiated by {sup 60}Co {gamma}-ray at a total dose of 2 Gy, and the changes associated with EMT, including morphology, EMT markers, migration and invasion, were observed by microscope, Western blot, immunofluorescence, scratch assay, and transwell chamber assay, respectively. Then the protein levels of TGF-{beta} in these cancer cells were detected by enzyme-linked immunosorbent assay, and the role of TGF-{beta} signaling pathway in the effect of ionizing radiation on EMT was investigate by using the specific inhibitor SB431542. Results: After irradiation with {gamma}-ray at a total dose of 2 Gy, cancer cells presented the mesenchymal phenotype, and compared with the sham-irradiation group the expression of epithelial markers was decreased and of mesenchymal markers was increased, the migratory and invasive capabilities were strengthened, and the protein levels of TGF-{beta} were enhanced. Furthermore, events associated with EMT induced by IR in A549 could be reversed through inhibition of TGF-{beta} signaling. Conclusions: These results suggest that EMT mediated by TGF-{beta} plays a critical role in IR-induced enhancing of migratory and invasive capabilities in cancer cells.

  9. Oligomannose-Rich Membranes of Dying Intestinal Epithelial Cells Promote Host Colonization by Adherent-Invasive E. coli

    Directory of Open Access Journals (Sweden)

    Tetiana Dumych

    2018-04-01

    Full Text Available A novel mechanism is revealed by which clinical isolates of adherent-invasive Escherichia coli (AIEC penetrate into the epithelial cell layer, replicate, and establish biofilms in Crohn's disease. AIEC uses the FimH fimbrial adhesin to bind to oligomannose glycans on the surface of host cells. Oligomannose glycans exposed on early apoptotic cells are the preferred binding targets of AIEC, so apoptotic cells serve as potential entry points for bacteria into the epithelial cell layer. Thereafter, the bacteria propagate laterally in the epithelial intercellular spaces. We demonstrate oligomannosylation at two distinct sites of a glycoprotein receptor for AIEC, carcinoembryonic antigen related cell adhesion molecule 6 (CEACAM6 or CD66c, on human intestinal epithelia. After bacterial binding, FimH interacts with CEACAM6, which then clusters. The presence of the highest-affinity epitope for FimH, oligomannose-5, on CEACAM6 is demonstrated using LC-MS/MS. As mannose-dependent infections are abundant, this mechanism might also be used by other adherent-invasive pathogens.

  10. Inhibition of Staphylococcus aureus invasion into bovine mammary epithelial cells by contact with live Lactobacillus casei.

    Science.gov (United States)

    Bouchard, Damien S; Rault, Lucie; Berkova, Nadia; Le Loir, Yves; Even, Sergine

    2013-02-01

    Staphylococcus aureus is a major pathogen that is responsible for mastitis in dairy herds. S. aureus mastitis is difficult to treat and prone to recurrence despite antibiotic treatment. The ability of S. aureus to invade bovine mammary epithelial cells (bMEC) is evoked to explain this chronicity. One sustainable alternative to treat or prevent mastitis is the use of lactic acid bacteria (LAB) as mammary probiotics. In this study, we tested the ability of Lactobacillus casei strains to prevent invasion of bMEC by two S. aureus bovine strains, RF122 and Newbould305, which reproducibly induce acute and moderate mastitis, respectively. L. casei strains affected adhesion and/or internalization of S. aureus in a strain-dependent manner. Interestingly, L. casei CIRM-BIA 667 reduced S. aureus Newbould305 and RF122 internalization by 60 to 80%, and this inhibition was confirmed for two other L. casei strains, including one isolated from bovine teat canal. The protective effect occurred without affecting bMEC morphology and viability. Once internalized, the fate of S. aureus was not affected by L. casei. It should be noted that L. casei was internalized at a low rate but survived in bMEC cells with a better efficiency than that of S. aureus RF122. Inhibition of S. aureus adhesion was maintained with heat-killed L. casei, whereas contact between live L. casei and S. aureus or bMEC was required to prevent S. aureus internalization. This first study of the antagonism of LAB toward S. aureus in a mammary context opens avenues for the development of novel control strategies against this major pathogen.

  11. Inhibition of Staphylococcus aureus Invasion into Bovine Mammary Epithelial Cells by Contact with Live Lactobacillus casei

    Science.gov (United States)

    Bouchard, Damien S.; Rault, Lucie; Berkova, Nadia; Le Loir, Yves

    2013-01-01

    Staphylococcus aureus is a major pathogen that is responsible for mastitis in dairy herds. S. aureus mastitis is difficult to treat and prone to recurrence despite antibiotic treatment. The ability of S. aureus to invade bovine mammary epithelial cells (bMEC) is evoked to explain this chronicity. One sustainable alternative to treat or prevent mastitis is the use of lactic acid bacteria (LAB) as mammary probiotics. In this study, we tested the ability of Lactobacillus casei strains to prevent invasion of bMEC by two S. aureus bovine strains, RF122 and Newbould305, which reproducibly induce acute and moderate mastitis, respectively. L. casei strains affected adhesion and/or internalization of S. aureus in a strain-dependent manner. Interestingly, L. casei CIRM-BIA 667 reduced S. aureus Newbould305 and RF122 internalization by 60 to 80%, and this inhibition was confirmed for two other L. casei strains, including one isolated from bovine teat canal. The protective effect occurred without affecting bMEC morphology and viability. Once internalized, the fate of S. aureus was not affected by L. casei. It should be noted that L. casei was internalized at a low rate but survived in bMEC cells with a better efficiency than that of S. aureus RF122. Inhibition of S. aureus adhesion was maintained with heat-killed L. casei, whereas contact between live L. casei and S. aureus or bMEC was required to prevent S. aureus internalization. This first study of the antagonism of LAB toward S. aureus in a mammary context opens avenues for the development of novel control strategies against this major pathogen. PMID:23183972

  12. RSK is a principal effector of the RAS-ERK pathway for eliciting a coordinate promotile/invasive gene program and phenotype in epithelial cells

    DEFF Research Database (Denmark)

    Doehn, Ulrik; Hauge, Camilla; Frank, Scott R

    2009-01-01

    The RAS-stimulated RAF-MEK-ERK pathway confers epithelial cells with critical motile and invasive capacities during development, tissue regeneration, and carcinoma progression, often via promoting the epithelial-mesenchymal transition (EMT). Many mechanisms by which ERK exerts this control remain...... elusive. We demonstrate that the ERK-activated kinase RSK is necessary to induce mesenchymal motility and invasive capacities in nontransformed epithelial and carcinoma cells. RSK is sufficient to induce certain motile responses. Expression profiling analysis revealed that a primary role of RSK...... to stimulate motility and invasion. These findings uncover a mechanism whereby the RAS-ERK pathway controls epithelial cell motility by identifying RSK as a key effector, from which emanate multiple highly coordinate transcription-dependent mechanisms for stimulation of motility and invasive properties....

  13. Naringenin decreases invasiveness and metastasis by inhibiting TGF-β-induced epithelial to mesenchymal transition in pancreatic cancer cells.

    Directory of Open Access Journals (Sweden)

    Changjie Lou

    Full Text Available Epithelial to mesenchymal transition (EMT promotes cellular motility, invasiveness and metastasis during embryonic development and tumorigenesis. Transforming growth factor-β (TGF-β signaling pathway is a key regulator of EMT. A lot of evidences suggest that this process is Smad3-dependent. Herein we showed that exposure of aspc-1 and panc-1 pancreatic cancer cells to TGF-β1 resulted in characteristic morphological alterations of EMT, and enhancement of cell motility and gemcitabine (Gem resistance along with an up-regulation of EMT markers genes such as vimentin, N-cadherin, MMP2 and MMP9. Naringenin (Nar down-regulated EMT markers expression in both mRNA and protein levels by inhibiting TGF-β1/Smad3 signal pathway in the pancreatic cancer cells. Consequently, Nar suppressed the cells migration and invasion and reversed their resistance to Gem.

  14. Mesenchymal stem cells promote cell invasion and migration and autophagy-induced epithelial-mesenchymal transition in A549 lung adenocarcinoma cells.

    Science.gov (United States)

    Luo, Dan; Hu, Shiyuan; Tang, Chunlan; Liu, Guoxiang

    2018-03-01

    Mesenchymal stem cells (MSCs) are recruited into the tumour microenvironment and promote tumour growth and metastasis. Tumour microenvironment-induced autophagy is considered to suppress primary tumour formation by impairing migration and invasion. Whether these recruited MSCs regulate tumour autophagy and whether autophagy affects tumour growth are controversial. Our data showed that MSCs promote autophagy activation, reactive oxygen species production, and epithelial-mesenchymal transition (EMT) as well as increased migration and invasion in A549 cells. Decreased expression of E-cadherin and increased expression of vimentin and Snail were observed in A549 cells cocultured with MSCs. Conversely, MSC coculture-mediated autophagy positively promoted tumour EMT. Autophagy inhibition suppressed MSC coculture-mediated EMT and reduced A549 cell migration and invasion slightly. Furthermore, the migratory and invasive abilities of A549 cells were additional increased when autophagy was further enhanced by rapamycin treatment. Taken together, this work suggests that microenvironments containing MSCs can promote autophagy activation for enhancing EMT; MSCs also increase the migratory and invasive abilities of A549 lung adenocarcinoma cells. Mesenchymal stem cell-containing microenvironments and MSC-induced autophagy signalling may be potential targets for blocking lung cancer cell migration and invasion. Copyright © 2018 John Wiley & Sons, Ltd.

  15. Stromal COX-2 signaling activated by deoxycholic acid mediates proliferation and invasiveness of colorectal epithelial cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Yingting, E-mail: yitizhu@yahoo.com [Arizona Cancer Center, The University of Arizona, Tucson, AZ 85724 (United States); Tissue Tech Inc., Miami, FL 33173 (United States); Zhu, Min; Lance, Peter [Arizona Cancer Center, The University of Arizona, Tucson, AZ 85724 (United States)

    2012-08-31

    Highlights: Black-Right-Pointing-Pointer Human colonic cancer associated fibroblasts are major sources of COX-2 and PGE{sub 2}. Black-Right-Pointing-Pointer The fibroblasts interact with human colonic epithelial cancer cells. Black-Right-Pointing-Pointer Activation of COX-2 signaling in the fibroblasts affects behavior of the epithelia. Black-Right-Pointing-Pointer Protein Kinase C controls the activation of COX-2 signaling. -- Abstract: COX-2 is a major regulator implicated in colonic cancer. However, how COX-2 signaling affects colonic carcinogenesis at cellular level is not clear. In this article, we investigated whether activation of COX-2 signaling by deoxycholic acid (DCA) in primary human normal and cancer associated fibroblasts play a significant role in regulation of proliferation and invasiveness of colonic epithelial cancer cells. Our results demonstrated while COX-2 signaling can be activated by DCA in both normal and cancer associated fibroblasts, the level of activation of COX-2 signaling is significantly greater in cancer associated fibroblasts than that in normal fibroblasts. In addition, we discovered that the proliferative and invasive potential of colonic epithelial cancer cells were much greater when the cells were co-cultured with cancer associated fibroblasts pre-treated with DCA than with normal fibroblasts pre-treated with DCA. Moreover, COX-2 siRNA attenuated the proliferative and invasive effect of both normal and cancer associate fibroblasts pre-treated with DCA on the colonic cancer cells. Further studies indicated that the activation of COX-2 signaling by DCA is through protein kinase C signaling. We speculate that activation of COX-2 signaling especially in cancer associated fibroblasts promotes progression of colonic cancer.

  16. Stromal COX-2 signaling activated by deoxycholic acid mediates proliferation and invasiveness of colorectal epithelial cancer cells

    International Nuclear Information System (INIS)

    Zhu, Yingting; Zhu, Min; Lance, Peter

    2012-01-01

    Highlights: ► Human colonic cancer associated fibroblasts are major sources of COX-2 and PGE 2 . ► The fibroblasts interact with human colonic epithelial cancer cells. ► Activation of COX-2 signaling in the fibroblasts affects behavior of the epithelia. ► Protein Kinase C controls the activation of COX-2 signaling. -- Abstract: COX-2 is a major regulator implicated in colonic cancer. However, how COX-2 signaling affects colonic carcinogenesis at cellular level is not clear. In this article, we investigated whether activation of COX-2 signaling by deoxycholic acid (DCA) in primary human normal and cancer associated fibroblasts play a significant role in regulation of proliferation and invasiveness of colonic epithelial cancer cells. Our results demonstrated while COX-2 signaling can be activated by DCA in both normal and cancer associated fibroblasts, the level of activation of COX-2 signaling is significantly greater in cancer associated fibroblasts than that in normal fibroblasts. In addition, we discovered that the proliferative and invasive potential of colonic epithelial cancer cells were much greater when the cells were co-cultured with cancer associated fibroblasts pre-treated with DCA than with normal fibroblasts pre-treated with DCA. Moreover, COX-2 siRNA attenuated the proliferative and invasive effect of both normal and cancer associate fibroblasts pre-treated with DCA on the colonic cancer cells. Further studies indicated that the activation of COX-2 signaling by DCA is through protein kinase C signaling. We speculate that activation of COX-2 signaling especially in cancer associated fibroblasts promotes progression of colonic cancer.

  17. Inhibition of Staphylococcus aureus Invasion into Bovine Mammary Epithelial Cells by Contact with Live Lactobacillus casei

    OpenAIRE

    Bouchard, Damien S.; Rault, Lucie; Berkova, Nadia; Le Loir, Yves; Even, Sergine

    2013-01-01

    Staphylococcus aureus is a major pathogen that is responsible for mastitis in dairy herds. S. aureus mastitis is difficult to treat and prone to recurrence despite antibiotic treatment. The ability of S. aureus to invade bovine mammary epithelial cells (bMEC) is evoked to explain this chronicity. One sustainable alternative to treat or prevent mastitis is the use of lactic acid bacteria (LAB) as mammary probiotics. In this study, we tested the ability of Lactobacillus casei strains to prevent...

  18. Identification of CsrC and Characterization of Its Role in Epithelial Cell Invasion in Salmonella enterica Serovar Typhimurium

    OpenAIRE

    Fortune, Doreen R.; Suyemoto, Mitsu; Altier, Craig

    2006-01-01

    The csr regulatory system of Salmonella regulates the expression of the genes of Salmonella pathogenicity island 1 (SPI1) required for the invasion of epithelial cells. This system consists of the posttranscriptional regulator CsrA and an untranslated regulatory RNA, CsrB, that opposes the action of CsrA. Here we identify and characterize the role of a second regulatory RNA, CsrC, whose ortholog was discovered previously in Escherichia coli. We show that a mutant of csrC has only mild defects...

  19. α-Solanine Inhibits Invasion of Human Prostate Cancer Cell by Suppressing Epithelial-Mesenchymal Transition and MMPs Expression

    Directory of Open Access Journals (Sweden)

    Kun-Hung Shen

    2014-08-01

    Full Text Available α-Solanine, a naturally occurring steroidal glycoalkaloid found in nightshade (Solanum nigrum Linn., was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in suppression of cancer cell metastasis by α-solanine remains unclear. This study investigates the suppression mechanism of α-solanine on motility of the human prostate cancer cell PC-3. Results show that α-solanine reduces the viability of PC-3 cells. When treated with non-toxic doses of α-solanine, cell invasion is markedly suppressed by α-solanine. α-Solanine also significantly elevates epithelial marker E-cadherin expression, while it concomitantly decreases mesenchymal marker vimentin expression, suggesting it suppresses epithelial-mesenchymal transition (EMT. α-Solanine reduces the mRNA level of matrix metalloproteinase-2 (MMP-2, MMP-9 and extracellular inducer of matrix metalloproteinase (EMMPRIN, but increases the expression of reversion-inducing cysteine-rich protein with kazal motifs (RECK, and tissue inhibitor of metalloproteinase-1 (TIMP-1 and TIMP-2. Immunoblotting assays indicate α-solanine is effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K, Akt and ERK. Moreover, α-solanine downregulates oncogenic microRNA-21 (miR-21 and upregulates tumor suppressor miR-138 expression. Taken together, the results suggest that inhibition of PC-3 cell invasion by α-solanine may be, at least in part, through blocking EMT and MMPs expression. α-Solanine also reduces ERK and PI3K/Akt signaling pathways and regulates expression of miR-21 and miR-138. These findings suggest an attractive therapeutic potential of α-solanine for suppressing invasion of prostate cancer cell.

  20. A core invasiveness gene signature reflects epithelial-to-mesenchymal transition but not metastatic potential in breast cancer cell lines and tissue samples.

    Directory of Open Access Journals (Sweden)

    Melike Marsan

    Full Text Available INTRODUCTION: Metastases remain the primary cause of cancer-related death. The acquisition of invasive tumour cell behaviour is thought to be a cornerstone of the metastatic cascade. Therefore, gene signatures related to invasiveness could aid in stratifying patients according to their prognostic profile. In the present study we aimed at identifying an invasiveness gene signature and investigated its biological relevance in breast cancer. METHODS & RESULTS: We collected a set of published gene signatures related to cell motility and invasion. Using this collection, we identified 16 genes that were represented at a higher frequency than observed by coincidence, hereafter named the core invasiveness gene signature. Principal component analysis showed that these overrepresented genes were able to segregate invasive and non-invasive breast cancer cell lines, outperforming sets of 16 randomly selected genes (all P<0.001. When applied onto additional data sets, the expression of the core invasiveness gene signature was significantly elevated in cell lines forced to undergo epithelial-mesenchymal transition. The link between core invasiveness gene expression and epithelial-mesenchymal transition was also confirmed in a dataset consisting of 2420 human breast cancer samples. Univariate and multivariate Cox regression analysis demonstrated that CIG expression is not associated with a shorter distant metastasis free survival interval (HR = 0.956, 95%C.I. = 0.896-1.019, P = 0.186. DISCUSSION: These data demonstrate that we have identified a set of core invasiveness genes, the expression of which is associated with epithelial-mesenchymal transition in breast cancer cell lines and in human tissue samples. Despite the connection between epithelial-mesenchymal transition and invasive tumour cell behaviour, we were unable to demonstrate a link between the core invasiveness gene signature and enhanced metastatic potential.

  1. The IpaC carboxyterminal effector domain mediates Src-dependent actin polymerization during Shigella invasion of epithelial cells.

    Directory of Open Access Journals (Sweden)

    Joëlle Mounier

    2009-01-01

    Full Text Available Shigella, the causative agent of bacillary dysentery, invades epithelial cells by locally reorganizing the actin cytoskeleton. Shigella invasion requires actin polymerization dependent on the Src tyrosine kinase and a functional bacterial type III secretion (T3S apparatus. Using dynamic as well as immunofluorescence microscopy, we show that the T3S translocon component IpaC allows the recruitment of the Src kinase required for actin polymerization at bacterial entry sites during the initial stages of Shigella entry. Src recruitment occurred at bacterial-cell contact sites independent of actin polymerization at the onset of the invasive process and was still observed in Shigella strains mutated for translocated T3S effectors of invasion. A Shigella strain with a polar mutation that expressed low levels of the translocator components IpaB and IpaC was fully proficient for Src recruitment and bacterial invasion. In contrast, a Shigella strain mutated in the IpaC carboxyterminal effector domain that was proficient for T3S effector translocation did not induce Src recruitment. Consistent with a direct role for IpaC in Src activation, cell incubation with the IpaC last 72 carboxyterminal residues fused to the Iota toxin Ia (IaC component that translocates into the cell cytosol upon binding to the Ib component led to Src-dependent ruffle formation. Strikingly, IaC also induced actin structures resembling bacterial entry foci that were enriched in activated Src and were inhibited by the Src inhibitor PP2. These results indicate that the IpaC effector domain determines Src-dependent actin polymerization and ruffle formation during bacterial invasion.

  2. Host epithelial cell invasion by Campylobacter jejuni: trigger or zipper mechanism?

    Directory of Open Access Journals (Sweden)

    Tadhg eÓ Cróinín

    2012-03-01

    Full Text Available Campylobacter jejuni, a spiral-shaped Gram-negative pathogen, is a highly frequent cause of gastrointestinal foodborne illness in humans worldwide. Clinical outcome of C. jejuni infections ranges from mild to severe diarrheal disease, and some other complications including reactive arthritis and Guillain–Barré syndrome. This review article highlights various C. jejuni pathogenicity factors, host cell determinants and proposed signaling mechanisms involved in human host cell invasion and their potential role in the development of C. jejuni-mediated disease. A model is presented which outlines the various important interactions of C. jejuni with the intestinal epithelium, and we discuss the pro’s and con’s for the zipper over the trigger mechanism of invasion. Future work should clarify the contradictory role of some previously identified factors, and should identify and characterize novel virulence determinants, which are crucial to provide fresh insights into the diversity of strategies employed by this pathogen to cause disease.

  3. Effect of Twist, Snail and YB-1 gene expression in cervical cancer tissue on cell invasion and epithelial-mesenchymal transition

    Directory of Open Access Journals (Sweden)

    Xin-Qin Kang1

    2017-05-01

    Full Text Available Objective: To study the effect of Twist, Snail and YB-1 gene expression in cervical cancer tissue on cell invasion and epithelial-mesenchymal transition. Methods: Cervical cancer tissue samples and tissue samples adjacent to carcinoma were collected from 138 patients with radical operation for cervical cancer, fluorescence quantitative PCR method was used to detect the mRNA expression of Twist, Snail and YB-1 genes, cell invasion-related genes and epithelial-mesenchymal transition marker genes, the Pearson test was used to analyze the correlation of Twist, Snail and YB-1 gene mRNA expression in cervical cancer tissue with cell invasion and epithelial-mesenchymal transition. Results: Twist, Snail and YB-1 gene mRNA expression in cervical cancer tissue were higher than those in tissue adjacent to carcinoma, the invasion genes STAT3, YAP1, TUG1, FoxM1 and Rab11 mRNA expression were higher than those in tissue adjacent to carcinoma, and the epithelial-mesenchymal transition markers E-cadherin and β-catenin gene mRNA expression were lower than those in tissue adjacent to carcinoma while vimentin gene mRNA expression was higher than that in tissue adjacent to carcinoma. Pearson test showed that Twist, Snail and YB-1 gene mRNA expression in cervical cancer tissue were directly correlated with cell invasion and epithelial-mesenchymal transition. Conclusion: Twist, Snail and YB-1 genes are highly expressed in cervical cancer tissue, and their abnormal expression directly leads to the increased tumor cell invasion activity and the aggravated epithelial-mesenchymal transition.

  4. Kefir-isolated bacteria and yeasts inhibit Shigella flexneri invasion and modulate pro-inflammatory response on intestinal epithelial cells.

    Science.gov (United States)

    Bolla, P A; Abraham, A G; Pérez, P F; de Los Angeles Serradell, M

    2016-02-01

    The aim of this work was to evaluate the ability of a kefir-isolated microbial mixture containing three bacterial and two yeast strains (MM) to protect intestinal epithelial cells against Shigella flexneri invasion, as well as to analyse the effect on pro-inflammatory response elicited by this pathogen. A significant decrease in S. flexneri strain 72 invasion was observed on both HT-29 and Caco-2 cells pre-incubated with MM. Pre-incubation with the individual strains Saccharomyces cerevisiae CIDCA 8112 or Lactococcus lactis subsp. lactis CIDCA 8221 also reduced the internalisation of S. flexneri into HT-29 cells although in a lesser extent than MM. Interestingly, Lactobacillus plantarum CIDCA 83114 exerted a protective effect on the invasion of Caco-2 and HT-29 cells by S. flexneri. Regarding the pro-inflammatory response on HT-29 cells, S. flexneri infection induced a significant activation of the expression of interleukin 8 (IL-8), chemokine (C-C motif) ligand 20 (CCL20) and tumour necrosis factor alpha (TNF-α) encoding genes (P<0.05), whereas incubation of cells with MM did not induce the expression of any of the mediators assessed. Interestingly, pre-incubation of HT-29 monolayer with MM produced an inhibition of S. flexneri-induced IL-8, CCL20 and TNF-α mRNA expression. In order to gain insight on the effect of MM (or the individual strains) on this pro-inflammatory response, a series of experiments using a HT-29-NF-κB-hrGFP reporter system were performed. Pre-incubation of HT-29-NF-κB-hrGFP cells with MM significantly dampened Shigella-induced activation. Our results showed that the contribution of yeast strain Kluyveromyces marxianus CIDCA 8154 seems to be crucial in the observed effect. In conclusion, results presented in this study demonstrate that pre-treatment with a microbial mixture containing bacteria and yeasts isolated from kefir, resulted in inhibition of S. flexneri internalisation into human intestinal epithelial cells, along with the

  5. Semaphorin 3 C drives epithelial-to-mesenchymal transition, invasiveness, and stem-like characteristics in prostate cells.

    Science.gov (United States)

    Tam, Kevin J; Hui, Daniel H F; Lee, Wilson W; Dong, Mingshu; Tombe, Tabitha; Jiao, Ivy Z F; Khosravi, Shahram; Takeuchi, Ario; Peacock, James W; Ivanova, Larissa; Moskalev, Igor; Gleave, Martin E; Buttyan, Ralph; Cox, Michael E; Ong, Christopher J

    2017-09-13

    Prostate cancer (PCa) is among the most commonly-occurring cancers worldwide and a leader in cancer-related deaths. Local non-invasive PCa is highly treatable but limited treatment options exist for those with locally-advanced and metastatic forms of the disease underscoring the need to identify mechanisms mediating PCa progression. The semaphorins are a large grouping of membrane-associated or secreted signalling proteins whose normal roles reside in embryogenesis and neuronal development. In this context, semaphorins help establish chemotactic gradients and direct cell movement. Various semaphorin family members have been found to be up- and down-regulated in a number of cancers. One family member, Semaphorin 3 C (SEMA3C), has been implicated in prostate, breast, ovarian, gastric, lung, and pancreatic cancer as well as glioblastoma. Given SEMA3C's roles in development and its augmented expression in PCa, we hypothesized that SEMA3C promotes epithelial-to-mesenchymal transition (EMT) and stem-like phenotypes in prostate cells. In the present study we show that ectopic expression of SEMA3C in RWPE-1 promotes the upregulation of EMT and stem markers, heightened sphere-formation, and cell plasticity. In addition, we show that SEMA3C promotes migration and invasion in vitro and cell dissemination in vivo.

  6. Phagocytosis and Epithelial Cell Invasion by Crohn’s Disease-Associated Adherent-Invasive Escherichia coli Are Inhibited by the Anti-inflammatory Drug 6-Mercaptopurine

    Directory of Open Access Journals (Sweden)

    Federica Migliore

    2018-05-01

    Full Text Available Adherent-invasive Escherichia coli (AIEC strains are overrepresented in the dysbiotic microbiota of Crohn’s disease (CD patients, and contribute to the onset of the chronic inflammation typical of the disease. However, the effects of anti-inflammatory drugs used for CD treatment on AIEC virulence have not yet been investigated. In this report, we show that exposure of AIEC LF82 strain to amino-6-mercaptopurine (6-MP riboside, one of the most widely used anti-inflammatory drugs in CD, impairs its ability to adhere to, and consequently to invade, human epithelial cells. Notably, phagocytosis of LF82 treated with 6-MP by human macrophages is also reduced, suggesting that 6-MP affects AIEC cell surface determinants involved both in interaction with epithelial cells and in uptake by macrophages. Since a main target of 6-MP in bacterial cells is the inhibition of the important signal molecule c-di-GMP, we also tested whether perturbations in cAMP, another major signaling pathway in E. coli, might have similar effects on interactions with human cells. To this aim, we grew LF82 in the presence of glucose, which leads to inhibition of cAMP synthesis. Growth in glucose-supplemented medium resulted in a reduction in AIEC adhesion to epithelial cells and uptake by macrophages. Consistent with these results, both 6-MP and glucose can affect expression of cell adhesion-related genes, such as the csg genes, encoding thin aggregative fimbriae (curli. In addition, glucose strongly inhibits expression of the fim operon, encoding type 1 pili, a known AIEC determinant for adhesion to human cells. To further investigate whether 6-MP can indeed inhibit c-di-GMP signaling in AIEC, we performed biofilm and motility assays and determination of extracellular polysaccharides. 6-MP clearly affected biofilm formation and cellulose production, but also, unexpectedly, reduced cell motility, itself an important virulence factor for AIEC. Our results provide strong evidence

  7. MicroRNA-122 triggers mesenchymal-epithelial transition and suppresses hepatocellular carcinoma cell motility and invasion by targeting RhoA.

    Directory of Open Access Journals (Sweden)

    Sheng-Chun Wang

    Full Text Available The loss of microRNA-122 (miR-122 expression is strongly associated with increased invasion and metastasis, and poor prognosis of hepatocellular carcinoma (HCC, however, the underlying mechanisms remain poorly understood. In the present study, we observed that miR-122 over-expression in HCC cell lines Sk-hep-1 and Bel-7402 triggered the mesenchymal-epithelial transition (MET, as demonstrated by epithelial-like morphological changes, up-regulated epithelial proteins (E-cadherin, ZO-1, α-catenin, occludin, BVES, and MST4, and down-regulated mesenchymal proteins (vimentin and fibronectin. The over-expression of miRNA-122 also caused cytoskeleton disruption, RhoA/Rock pathway inactivation, enhanced cell adhesion, and suppression of migration and invasion of Sk-hep-1 and Bel-7402 cells, whereas, these effects could be reversed through miR-122 inhibition. Additional studies demonstrated that the inhibition of wild-type RhoA function induced MET and inhibited cell migration and invasion, while RhoA over-expression reversed miR-122-induced MET and inhibition of migration and invasion of HCC cells, suggesting that miR-122 induced MET and suppressed the migration and invasion of HCC cells by targeting RhoA. Moreover, our results demonstrated that HNF4α up-regulated its target gene miR-122 that subsequently induced MET and inhibited cell migration and invasion, whereas miR-122 inhibition reversed these HNF4α-induced phenotypes. These results revealed functional and mechanistic links among the tumor suppressors HNF4α, miR-122, and RhoA in EMT and invasive and metastatic phenotypes of HCC. Taken together, our study provides the first evidence that the HNF4α/miR-122/RhoA axis negatively regulates EMT and the migration and invasion of HCC cells.

  8. Gefitinib inhibits invasive phenotype and epithelial-mesenchymal transition in drug-resistant NSCLC cells with MET amplification.

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    Silvia La Monica

    Full Text Available Despite the initial response, all patients with epidermal growth factor receptor (EGFR-mutant non-small cell lung cancer (NSCLC eventually develop acquired resistance to EGFR tyrosine kinase inhibitors (TKIs. The EGFR-T790M secondary mutation is responsible for half of acquired resistance cases, while MET amplification has been associated with acquired resistance in about 5-15% of NSCLCs. Clinical findings indicate the retained addiction of resistant tumors on EGFR signaling. Therefore, we evaluated the molecular mechanisms supporting the therapeutic potential of gefitinib maintenance in the HCC827 GR5 NSCLC cell line harbouring MET amplification as acquired resistance mechanism. We demonstrated that resistant cells can proliferate and survive regardless of the presence of gefitinib, whereas the absence of the drug significantly enhanced cell migration and invasion. Moreover, the continuous exposure to gefitinib prevented the epithelial-mesenchymal transition (EMT with increased E-cadherin expression and down-regulation of vimentin and N-cadherin. Importantly, the inhibition of cellular migration was correlated with the suppression of EGFR-dependent Src, STAT5 and p38 signaling as assessed by a specific kinase array, western blot analysis and silencing functional studies. On the contrary, the lack of effect of gefitinib on EGFR phosphorylation in the H1975 cells (EGFR-T790M correlated with the absence of effects on cell migration and invasion. In conclusion, our findings suggest that certain EGFR-mutated patients may still benefit from a second-line therapy including gefitinib based on the specific mechanism underlying tumor cell resistance.

  9. Titanium dioxide nanoparticles induce an adaptive inflammatory response and invasion and proliferation of lung epithelial cells in chorioallantoic membrane

    International Nuclear Information System (INIS)

    Medina-Reyes, Estefany I.; Déciga-Alcaraz, Alejandro; Freyre-Fonseca, Verónica; Delgado-Buenrostro, Norma L.; Flores-Flores, José O.; Gutiérrez-López, Gustavo F.; Sánchez-Pérez, Yesennia; García-Cuéllar, Claudia M.

    2015-01-01

    Titanium dioxide nanoparticles (TiO 2 NPs) studies have been performed using relatively high NPs concentration under acute exposure and limited studies have compared shape effects. We hypothesized that midterm exposure to low TiO 2 NPs concentration in lung epithelial cells induces carcinogenic characteristics modulated partially by NPs shape. To test our hypothesis we synthesized NPs shaped as belts (TiO 2 -B) using TiO 2 spheres (TiO 2 -SP) purchased from Sigma Aldrich Co. Then, lung epithelial A549 cells were low-exposed (10 µg/cm 2 ) to both shapes during 7 days and internalization, cytokine release and invasive potential were determined. Results showed greater TiO 2 -B effect on agglomerates size, cell size and granularity than TiO 2 -SP. Agglomerates size in cell culture medium was 310 nm and 454 nm for TiO 2 -SP and TiO 2 -B, respectively; TiO 2 -SP and TiO 2 -B induced 23% and 70% cell size decrease, respectively, whilst TiO 2 -SP and TiO 2 -B induced 7 and 14-fold of granularity increase. NO x production was down-regulated (31%) by TiO 2 -SP and up-regulated (70%) by TiO 2 -B. Both NPs induced a transient cytokine release (IL-2, IL-6, IL-8, IL-4, IFN-γ, and TNF-α) after 4 days, but cytokines returned to basal levels in TiO 2 -SP exposed cells while TiO 2 -B induced a down-regulation after 7 days. Midterm exposure to both shapes of NPs induced capability to degrade cellular extracellular matrix components from chorioallantoic membrane and Ki-67 marker showed that TiO 2 -B had higher proliferative potential than TiO 2 -SP. We conclude that midterm exposure to low NPs concentration of NPs has an impact in the acquisition of new characteristics of exposed cells and NPs shape influences cellular outcome. - Graphical abstract: (A) Lung epithelial cells were low exposed (below 10 µg/cm 2 ) to titanium dioxide nanoparticles (TiO 2 -NPs) shaped as spheres (TiO 2 -SP) and belts (TiO 2 -B) for midterm (7 continuous days) separately. (B) Then, cells from each cell

  10. Titanium dioxide nanoparticles induce an adaptive inflammatory response and invasion and proliferation of lung epithelial cells in chorioallantoic membrane

    Energy Technology Data Exchange (ETDEWEB)

    Medina-Reyes, Estefany I.; Déciga-Alcaraz, Alejandro [Unidad de Biomedicina, Facultad de Estudios Superiores Iztacala, Universidad Nacional Autónoma de México, CP 54059 Estado de México (Mexico); Freyre-Fonseca, Verónica [Unidad de Biomedicina, Facultad de Estudios Superiores Iztacala, Universidad Nacional Autónoma de México, CP 54059 Estado de México (Mexico); Doctorado en Ciencias en Alimentos, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, CP 11340 México, DF (Mexico); Delgado-Buenrostro, Norma L. [Unidad de Biomedicina, Facultad de Estudios Superiores Iztacala, Universidad Nacional Autónoma de México, CP 54059 Estado de México (Mexico); Flores-Flores, José O. [Centro de Ciencias Aplicadas y Desarrollo Tecnológico, Universidad Nacional Autónoma de México, Circuito Exterior S/N, Ciudad Universitaria AP 70-186, CP 04510 México, DF (Mexico); Gutiérrez-López, Gustavo F. [Escuela Nacional de Ciencias Biológicas del Instituto Politécnico Nacional, CP 11340 México, DF (Mexico); Sánchez-Pérez, Yesennia; García-Cuéllar, Claudia M. [Instituto Nacional de Cancerología, Subdirección de Investigación Básica, San Fernando 22, Tlalpan, CP 14080 México, DF (Mexico); and others

    2015-01-15

    Titanium dioxide nanoparticles (TiO{sub 2} NPs) studies have been performed using relatively high NPs concentration under acute exposure and limited studies have compared shape effects. We hypothesized that midterm exposure to low TiO{sub 2} NPs concentration in lung epithelial cells induces carcinogenic characteristics modulated partially by NPs shape. To test our hypothesis we synthesized NPs shaped as belts (TiO{sub 2}-B) using TiO{sub 2} spheres (TiO{sub 2}-SP) purchased from Sigma Aldrich Co. Then, lung epithelial A549 cells were low-exposed (10 µg/cm{sup 2}) to both shapes during 7 days and internalization, cytokine release and invasive potential were determined. Results showed greater TiO{sub 2}-B effect on agglomerates size, cell size and granularity than TiO{sub 2}-SP. Agglomerates size in cell culture medium was 310 nm and 454 nm for TiO{sub 2}-SP and TiO{sub 2}-B, respectively; TiO{sub 2}-SP and TiO{sub 2}-B induced 23% and 70% cell size decrease, respectively, whilst TiO{sub 2}-SP and TiO{sub 2}-B induced 7 and 14-fold of granularity increase. NO{sub x} production was down-regulated (31%) by TiO{sub 2}-SP and up-regulated (70%) by TiO{sub 2}-B. Both NPs induced a transient cytokine release (IL-2, IL-6, IL-8, IL-4, IFN-γ, and TNF-α) after 4 days, but cytokines returned to basal levels in TiO{sub 2}-SP exposed cells while TiO{sub 2}-B induced a down-regulation after 7 days. Midterm exposure to both shapes of NPs induced capability to degrade cellular extracellular matrix components from chorioallantoic membrane and Ki-67 marker showed that TiO{sub 2}-B had higher proliferative potential than TiO{sub 2}-SP. We conclude that midterm exposure to low NPs concentration of NPs has an impact in the acquisition of new characteristics of exposed cells and NPs shape influences cellular outcome. - Graphical abstract: (A) Lung epithelial cells were low exposed (below 10 µg/cm{sup 2}) to titanium dioxide nanoparticles (TiO{sub 2}-NPs) shaped as spheres (TiO{sub 2

  11. MicroRNA-145 Inhibits Cell Migration and Invasion and Regulates Epithelial-Mesenchymal Transition (EMT) by Targeting Connective Tissue Growth Factor (CTGF) in Esophageal Squamous Cell Carcinoma.

    Science.gov (United States)

    Han, Qiang; Zhang, Hua-Yong; Zhong, Bei-Long; Wang, Xiao-Jing; Zhang, Bing; Chen, Hua

    2016-10-23

    BACKGROUND This study investigated the mechanism of miR-145 in targeting connective tissue growth factor (CTGF), which affects the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of ESCC cells. MATERIAL AND METHODS A total of 50 ESCC tissues and their corresponding normal adjacent esophageal tissue samples were collected. Then, miR-145 expression in both ESCC clinical specimens and cell lines was detected using quantitative real-time PCR. CTGF protein was detected using immunohistochemistry. Dual luciferase reporter gene assay was employed to assess the effect of miR-145 on the 3'UTR luciferase activity of CTGF. Eca109 cells were transfected with miR-145 mimics and CTGF siRNA, respectively, and changes in cellular proliferation, migration, and invasion were detected via MTT assay, wound-healing assay, and Transwell assay, respectively. Western blotting assay was used to detect the expression of marker genes related to EMT. RESULTS MiR-145 was significantly down-regulated in ESCC tissues and cell lines compared with normal tissues and cell lines (Ptissues was than in normal adjacent esophageal tissues (Ptissues and cell lines, while the protein expression of CTGF exhibited the opposite trend. MiR-145 inhibited the proliferation, migration, invasiveness, and the EMT process of ESCC cells through targeted regulation of CTGF expression.

  12. Targeting Epithelial-Mesenchymal Transition for Identification of Inhibitors for Pancreatic Cancer Cell Invasion and Tumor Spheres Formation.

    Directory of Open Access Journals (Sweden)

    Kishore Polireddy

    Full Text Available Pancreatic cancer has an enrichment of stem-like cancer cells (CSCs that contribute to chemoresistant tumors prone to metastasis and recurrence. Drug screening assays based on cytotoxicity cannot identify specific CSC inhibitors, because CSCs comprise only a small portion of cancer cell population, and it is difficult to propagate stable CSC populations in vitro for high-throughput screening (HTS assays. Based on the important role of cancer cell epithelial-to-mesenchymal transition (EMT in promoting CSCs, we hypothesized that inhibition of EMT can be a useful strategy for inhibiting CSCs, and therefore a feasible approach for HTS can be built for identification of CSC inhibitors, based on assays detecting EMT inhibition.An immunofluorescent assay was established and optimized for HTS to identify compounds that enhance E-cadherin expression, as a hallmark of inhibition of EMT. Four chemical libraries containing 41,472 compounds were screened in PANC-1 pancreatic cancer cell line. Positive hits were validated for EMT and CSC inhibition in vitro using sphere formation assay, western blotting, immune fluorescence, and scratch assay.Initial hits were refined to 73 compounds with a secondary screening, among which 17 exhibited concentration dependent induction of E-cadherin expression. Six compounds were selected for further study which belonged to 2 different chemical structural clusters. A novel compound 1-(benzylsulfonyl indoline (BSI, Compound #38 significantly inhibited pancreatic cancer cell migration and invasion. BSI inhibited histone deacetylase, increased histone 4 acetylation preferably, resulting in E-cadherin up-regulation. BSI effectively inhibited tumor spheres formation. Six more analogues of BSI were tested for anti-migration and anti-CSC activities.This study demonstrated a feasible approach for discovery of agents targeting EMT and CSCs using HTS, and identified a class of novel chemicals that could be developed as anti-EMT and

  13. Fisetin inhibits human melanoma cell invasion through promotion of mesenchymal to epithelial transition and by targeting MAPK and NFκB signaling pathways.

    Directory of Open Access Journals (Sweden)

    Harish Chandra Pal

    Full Text Available Malignant melanoma is responsible for approximately 75% of skin cancer-related deaths. BRAF plays an important role in regulating the mitogen-activated protein kinase (MAPK signaling cascade in melanoma with activating mutations in the serine/threonine kinase BRAF occurring in 60-70% of malignant melanomas. The BRAF-MEK-ERK (MAPK pathway is a key regulator of melanoma cell invasion. In addition, activation of NFκB via the MAPK pathway is regulated through MEK-induced activation of IKK. These pathways are potential targets for prevention and treatment of melanoma. In this study, we investigated the effect of fisetin, a phytochemical present in fruits and vegetables, on melanoma cell invasion and epithelial-mesenchymal transition, and delineated the underlying molecular mechanism. Treatment of multiple human malignant melanoma cell lines with fisetin (5-20 µM resulted in inhibition of cell invasion. BRAF mutated melanoma cells were more sensitive to fisetin treatment, and this was associated with a decrease in the phosphorylation of MEK1/2 and ERK1/2. In addition, fisetin inhibited the activation of IKK leading to a reduction in the activation of the NFκB signaling pathway. Treatment of cells with an inhibitor of MEK1/2 (PD98059 or of NFκB (caffeic acid phenethyl ester also reduced melanoma cell invasion. Furthermore, treatment of fisetin promoted mesenchymal to epithelial transition in melanoma cells, which was associated with a decrease in mesenchymal markers (N-cadherin, vimentin, snail and fibronectin and an increase in epithelial markers (E-cadherin and desmoglein. Employing three dimensional skin equivalents consisting of A375 cells admixed with normal human keratinocytes embedded onto a collagen-constricted fibroblast matrix, we found that treatment of fisetin reduced the invasive potential of melanoma cells into the dermis and increased the expression of E-cadherin with a concomitant decrease in vimentin. These results indicate that

  14. Fisetin Inhibits Human Melanoma Cell Invasion through Promotion of Mesenchymal to Epithelial Transition and by Targeting MAPK and NFκB Signaling Pathways

    Science.gov (United States)

    Pal, Harish Chandra; Sharma, Samriti; Strickland, Leah Ray; Katiyar, Santosh K.; Ballestas, Mary E.; Athar, Mohammad; Elmets, Craig A.; Afaq, Farrukh

    2014-01-01

    Malignant melanoma is responsible for approximately 75% of skin cancer-related deaths. BRAF plays an important role in regulating the mitogen-activated protein kinase (MAPK) signaling cascade in melanoma with activating mutations in the serine/threonine kinase BRAF occurring in 60–70% of malignant melanomas. The BRAF-MEK-ERK (MAPK) pathway is a key regulator of melanoma cell invasion. In addition, activation of NFκB via the MAPK pathway is regulated through MEK-induced activation of IKK. These pathways are potential targets for prevention and treatment of melanoma. In this study, we investigated the effect of fisetin, a phytochemical present in fruits and vegetables, on melanoma cell invasion and epithelial-mesenchymal transition, and delineated the underlying molecular mechanism. Treatment of multiple human malignant melanoma cell lines with fisetin (5–20 µM) resulted in inhibition of cell invasion. BRAF mutated melanoma cells were more sensitive to fisetin treatment, and this was associated with a decrease in the phosphorylation of MEK1/2 and ERK1/2. In addition, fisetin inhibited the activation of IKK leading to a reduction in the activation of the NFκB signaling pathway. Treatment of cells with an inhibitor of MEK1/2 (PD98059) or of NFκB (caffeic acid phenethyl ester) also reduced melanoma cell invasion. Furthermore, treatment of fisetin promoted mesenchymal to epithelial transition in melanoma cells, which was associated with a decrease in mesenchymal markers (N-cadherin, vimentin, snail and fibronectin) and an increase in epithelial markers (E-cadherin and desmoglein). Employing three dimensional skin equivalents consisting of A375 cells admixed with normal human keratinocytes embedded onto a collagen-constricted fibroblast matrix, we found that treatment of fisetin reduced the invasive potential of melanoma cells into the dermis and increased the expression of E-cadherin with a concomitant decrease in vimentin. These results indicate that fisetin

  15. Forkhead Box Protein C2 Promotes Epithelial-Mesenchymal Transition, Migration and Invasion in Cisplatin-Resistant Human Ovarian Cancer Cell Line (SKOV3/CDDP

    Directory of Open Access Journals (Sweden)

    Chanjuan Li

    2016-08-01

    Full Text Available Background/Aims: Forkhead Box Protein C2 (FOXC2 has been reported to be overexpressed in a variety of human cancers. However, it is unclear whether FOXC2 regulates epithelial-mesenchymal transition (EMT in CDDP-resistant ovarian cancer cells. The aim of this study is to investigate the effects of FOXC2 on EMT and invasive characteristics of CDDP-resistant ovarian cancer cells and the underlying molecular mechanism. Methods: MTT, Western blot, scratch wound healing, matrigel transwell invasion, attachment and detachment assays were performed to detect half maximal inhibitory concentration (IC50 of CDDP, expression of EMT-related proteins and invasive characteristics in CDDP-resistant ovarian cancer cell line (SKOV3/CDDP and its parental cell line (SKOV3. Small hairpin RNA (shRNA was used to knockdown FOXC2 and analyze the effect of FOXC2 knockdown on EMT and invasive characteristics of SKOV3/CDDP cells. Also, the effect of FOXC2 upregulation on EMT and invasive characteristics of SKOV3 cells was analyzed. Furthermore, the molecular mechanism underlying FOXC2-regulating EMT in ovarian cancer cells was determined. Results: Compared with parental SKOV3 cell line, SKOV3/CDDP showed higher IC50 of CDDP (43.26μM (PConclusions: Taken together, these data demonstrate that FOXC2 may be a promoter of EMT phenotype in CDDP-resistant ovarian cancer cells and a potential therapeutic target for the treatment of advanced ovarian cancer.

  16. Leukocyte-associated immunoglobulin-like receptor-1 expressed in epithelial ovarian cancer cells and involved in cell proliferation and invasion

    International Nuclear Information System (INIS)

    Cao, Qizhi; Fu, Aili; Yang, Shude; He, Xiaoli; Wang, Yue; Zhang, Xiaoshu; Zhou, Jiadi; Luan, Xiying; Yu, Wenzheng; Xue, Jiangnan

    2015-01-01

    Previous studies have shown that leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1) is expressed on most types of hamatopoietic cells and negatively regulate immune response, but the roles of LAIR-1 in tumor of the non-hematopoietic lineage have not been determined. Despite advances in therapy of epithelial ovarian cancer (EOC), many questions relating to EOC pathogenesis remain unanswered. The aim of this study was to investigate the clinical significance of LAIR-1 expression in EOC and explore the possible association between LAIR-1 and cancer. In this study, a tissue microarray containing 78 ovarian cancer cases was stained following a standard immunohistochemical protocol for LAIR-1 and the correlation of LAIR-1 expression with clinicopathologic features was assessed. LAIR-1 was detected to express in tumor cells of ovarian cancer tissues (73.1%) and EOC cell lines COC1 and HO8910, not in normal ovarian tissues. In addition, LAIR-1 expression correlates significantly with tumor grade (p = 0.004). Furthermore, down-regulation of LAIR-1 in HO8910 cells increased cell proliferation, colony formation and cell invasion. These data suggest that LAIR-1 has a relevant impact on EOC progression and may be helpful for a better understanding of molecular pathogenesis of cancer. - Highlights: • LAIR-1 is expressed in epithelial ovarian cancer cells. • LAIR-1 expression correlates significantly with tumor grade. • Down-regulation of LAIR-1 expression increased cell proliferation and invasion. • LAIR-1 may be a novel candidate for cancer diagnosis and therapy

  17. Propolin C Inhibited Migration and Invasion via Suppression of EGFR-Mediated Epithelial-to-Mesenchymal Transition in Human Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Jih-Tung Pai

    2018-01-01

    Full Text Available Controlling lung cancer cell migration and invasion via epithelial-to-mesenchymal transition (EMT through the regulation of epidermal growth factor receptor (EGFR signaling pathway has been demonstrated. Searching biological active phytochemicals to repress EGFR-regulated EMT might prevent lung cancer progression. Propolis has been used as folk medicine in many countries and possesses anti-inflammatory, antioxidant, and anticancer activities. In this study, the antimigration and anti-invasion activities of propolin C, a c-prenylflavanone from Taiwanese propolis, were investigated on EGFR-regulated EMT signaling pathway. Cell migration and invasion activities were dose-dependently suppressed by noncytotoxic concentration of propolin C. Downregulations of vimentin and snail as well as upregulation of E-cadherin expressions were through the inhibition of EGFR-mediated phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt and extracellular signal-regulated kinase (ERK signaling pathway in propolin C-treated cells. In addition, EGF-induced migration and invasion were suppressed by propolin C-treated A549 lung cancer cells. No significant differences in E-cadherin expression were observed in EGF-stimulated cells. Interestingly, EGF-induced expressions of vimentin, snail, and slug were suppressed through the inhibition of PI3K/Akt and ERK signaling pathway in propolin C-treated cells. Inhibition of cell migration and invasion by propolin C was through the inhibition of EGF/EGFR-mediated signaling pathway, followed by EMT suppression in lung cancer.

  18. Legionella dumoffii Tex-KL Mutated in an Operon Homologous to traC-traD is Defective in Epithelial Cell Invasion.

    Science.gov (United States)

    Qin, Tian; Ken-Ichiro, Iida; Ren, Hong Yu; Zhou, Hai Jian; Yoshida, Shin-Ichi

    2016-06-01

    To understand the mechanism of invasion by Legionella dumoffii. The L. dumoffii strain Tex-KL was mutated using the Tn903 derivative, Tn903dIIlacZ. After screening 799 transposon insertion mutants, we isolated one defective mutant. We then constructed the gene-disrupted mutant, KL16, and studied its invasion of and intracellular growth in HeLa and A549 cells, and in A/J mice survival experiments. The structure of traC-traD operon was analyzed by RT-PCR. The transposon insertion was in a gene homologous to Salmonella typhi traC, which is required for the assembly of F pilin into the mature F pilus structure and for conjugal DNA transmission. Results from RT-PCR suggested that the traC-traD region formed an operon. We found that when the traC gene was disrupted, invasion and intracellular growth of L. dumoffii Tex-KL were impaired in human epithelial cells. When mice were infected by intranasal inoculation with a traC deficient mutant, their survival significantly increased when compared to mice infected with the wild-type strain.. Our results indicated that the traC-traD operon is required for the invasion and intracellular growth abilities of L. dumoffii Tex-KL in epithelial cells. Copyright © 2016 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  19. Upregulation of CD147 promotes cell invasion, epithelial-to-mesenchymal transition and activates MAPK/ERK signaling pathway in colorectal cancer

    OpenAIRE

    Xu, Tao; Zhou, Mingliang; Peng, Lipan; Kong, Shuai; Miao, Ruizheng; Shi, Yulong; Sheng, Hongguang; Li, Leping

    2014-01-01

    Colorectal cancer (CRC) is one of the most common cancers in the world. CD147, a transmembrane protein, has been reported to be correlated with various cancers. In this study, we aimed to investigate the mechanism of CD147 in regulating drug resistance, cell invasion and epithelial-to-mesenchymal transition (EMT) in CRC cells. qRT-PCR and western blotting were used to evaluated the expression of CD147 in 40 CRC cases and 4 cell lines. Increased expression of CD147 at both mRNA and protein lev...

  20. Regulation of DMBT1 via NOD2 and TLR4 in intestinal epithelial cells modulates bacterial recognition and invasion

    DEFF Research Database (Denmark)

    Rosenstiel, Philip; Sina, Christian; End, Caroline

    2007-01-01

    Mucosal epithelial cell layers are constantly exposed to a complex resident microflora. Deleted in malignant brain tumors 1 (DMBT1) belongs to the group of secreted scavenger receptor cysteine-rich proteins and is considered to be involved in host defense by pathogen binding. This report describe...

  1. HMGB1 is negatively correlated with the development of endometrial carcinoma and prevents cancer cell invasion and metastasis by inhibiting the process of epithelial-to-mesenchymal transition

    Directory of Open Access Journals (Sweden)

    Luan XR

    2017-03-01

    Full Text Available Xiaorong Luan,1,2 Chunjing Ma,2 Ping Wang,2 Fenglan Lou1 1Nursing College, Shandong University, 2Qilu Hospital of Shandong University, Jinan, People’s Republic of China Abstract: High-mobility group box protein 1 (HMGB1, a nuclear protein that plays a significant role in DNA architecture and transcription, was correlated with the progression of some types of cancer. However, the role of HMGB1 in endometrial cancer cell invasion and metastasis remains unexplored. HMGB1 expression was initially assessed by immunohistochemistry and reverse transcription-quantitative polymerase chain reaction (RT-qPCR in normal endometrial tissue and endometrial carcinoma tissue. High expressions of HMGB1 protein were detected in normal endometrial tissues; however, in endometrial cancer tissues, the expressions of HMGB1 were found to be very weak. Furthermore, HMGB1 expressions were negatively correlated with advanced stage and lymph node metastasis in endometrial cancer. Then by RT-qPCR, Western blot and immunocytochemistry, HMGB1 was also detected in primary cultured endometrial cells and four kinds of endometrial cancer cell lines (Ishikawa, HEC-1A, HEC-1B and KLE. We found that the expression of HMGB1 was much higher in normal endometrial cells than in endometrial cancer cells, and reduced expression levels of HMGB1 were observed especially in the highly metastatic cell lines. Using lentivirus transfection, HMGB1 small hairpin RNA was constructed, and this infected the lowly invasive endometrial cancer cell lines, Ishikawa and HEC-1B. HMGB1 knockdown significantly enhanced the proliferation, invasion and metastasis of endometrial cancer cells and induced the process of epithelial-to-mesenchymal transition. These results can contribute to the development of a new potential therapeutic target for endometrial cancer. Keywords: HMGB1, endometrial cancer, invasion, metastasis, epithelial-to-mesenchymal transition

  2. Upregulation of CD147 promotes cell invasion, epithelial-to-mesenchymal transition and activates MAPK/ERK signaling pathway in colorectal cancer.

    Science.gov (United States)

    Xu, Tao; Zhou, Mingliang; Peng, Lipan; Kong, Shuai; Miao, Ruizheng; Shi, Yulong; Sheng, Hongguang; Li, Leping

    2014-01-01

    Colorectal cancer (CRC) is one of the most common cancers in the world. CD147, a transmembrane protein, has been reported to be correlated with various cancers. In this study, we aimed to investigate the mechanism of CD147 in regulating drug resistance, cell invasion and epithelial-to-mesenchymal transition (EMT) in CRC cells. qRT-PCR and western blotting were used to evaluated the expression of CD147 in 40 CRC cases and 4 cell lines. Increased expression of CD147 at both mRNA and protein levels was found in CRC samples, and the level of CD147 was correlated with lymph node metastasis. CD147 overexpression increased the 5-Fluorouracil (5-FU) resistance, enhanced the invasion and EMT of CRC cells by regulating EMT markers and MMPs. Adverse results were obtained in CD147 knockdown CRC cell line. Further investigation revealed that CD147 activated MAPK/ERK pathway, ERK inhibitor U0126 suppressed the CD147-induced cell invasion, migration and MMP-2, MMP-9 expression. Taken together, our study indicates that CD147 promotes the 5-FU resistance, and MAPK/ERK signaling pathway is involved in CD147-promoted invasion and EMT of CRC cells.

  3. PRRX2 as a novel TGF-β-induced factor enhances invasion and migration in mammary epithelial cell and correlates with poor prognosis in breast cancer.

    Science.gov (United States)

    Juang, Yu-Lin; Jeng, Yung-Ming; Chen, Chi-Long; Lien, Huang-Chun

    2016-12-01

    TGF-β and cancer progression share a multifaceted relationship. Despite the knowledge of TGF-β biology in the development of cancer, several factors that mediate the cancer-promoting role of TGF-β continue to be identified. This study aimed to identify and characterise novel factors potentially related to TGF-β-mediated tumour aggression in breast cells. We treated the human mammary epithelial cell line MCF10A with TGF-β and identified TGF-β-dependent upregulation of PRRX2, the gene encoding paired-related homeobox 2 transcription factor. Overexpression of PRRX2 enhanced migration, invasion and anchorage-independent growth of MCF10A cells and induced partial epithelial mesenchymal transition (EMT), as determined by partial fibroblastoid morphology of cells, upregulation of EMT markers and partially disrupted acinar structure in a three-dimensional culture. We further identified PLAT, the gene encoding tissue-type plasminogen activator (tPA), as the highest differentially expressed gene in PRRX2-overexpressing MCF10A cells, and demonstrated direct binding and transactivation of the PLAT promoter by PRRX2. Furthermore, PLAT knockdown inhibited PRRX2-mediated enhanced migration and invasion, suggesting that tPA may mediate PRRX2-induced migration and invasion. Finally, the significant correlation of PRRX2 expression with poor survival in 118 primary breast tumour samples (P = 0.027) and the increased PRRX2 expression in metaplastic breast carcinoma samples, which is pathogenetically related to EMT, validated the biological importance of PRRX2-enhanced migration and invasion and PRRX2-induced EMT. Thus, our data suggest that upregulation of PRRX2 may be a mechanism contributing to TGF-β-induced invasion and EMT in breast cancer. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  4. Osteopontin Promotes Invasion, Migration and Epithelial-Mesenchymal Transition of Human Endometrial Carcinoma Cell HEC-1A Through AKT and ERK1/2 Signaling.

    Science.gov (United States)

    Li, Yinghua; Xie, Yunpeng; Cui, Dan; Ma, Yanni; Sui, Linlin; Zhu, Chenyang; Kong, Hui; Kong, Ying

    2015-01-01

    Osteopontin (OPN) is an Extracellular Matrix (ECM) molecule and is involved in many physiologic and pathologic processes, including cell adhesion, angiogenesis and tumor metastasis. OPN is a well-known multifunctional factor involved in various aspects of cancer progression, including endometrial cancer. In this study, we examined the significance of OPN in endometrial cancer. The proliferation, migration and invasion ability of HEC-1A cells were detected by Cell Counting Kit-8 (CCK-8), Wound scratch assay and transwell. Western blots were employed to detect the expression of Matrix metalloproteinase-2 (MMP-2) and epithelial-mesenchymal transition (EMT)-related factors in HEC-1A cells treated with rhOPN. rhOPN promotes cell proliferation, migration and invasion in HEC-1A cells. rhOPN influenced EMT-related factors and MMP-2 expression in HEC-1A cells. rhOPN promoted HEC-1A cells migration, invasion and EMT through protein kinase B (PKB/AKT) and Extracellular regulated protein kinases (ERK1/2) signaling pathway. These results may open up a novel therapeutic strategy for endometrial cancer: namely, rhOPN have important roles in controlling growth of endometrial of cancer cells and suggest a novel target pathway for treatment of this cancer. © 2015 The Author(s) Published by S. Karger AG, Basel.

  5. Osteopontin Promotes Invasion, Migration and Epithelial-Mesenchymal Transition of Human Endometrial Carcinoma Cell HEC-1A Through AKT and ERK1/2 Signaling

    Directory of Open Access Journals (Sweden)

    Yinghua Li

    2015-10-01

    Full Text Available Background/Aims: Osteopontin (OPN is an Extracellular Matrix (ECM molecule and is involved in many physiologic and pathologic processes, including cell adhesion, angiogenesis and tumor metastasis. OPN is a well-known multifunctional factor involved in various aspects of cancer progression, including endometrial cancer. In this study, we examined the significance of OPN in endometrial cancer. Methods: The proliferation, migration and invasion ability of HEC-1A cells were detected by Cell Counting Kit-8 (CCK-8, Wound scratch assay and transwell. Western blots were employed to detect the expression of Matrix metalloproteinase-2 (MMP-2 and epithelial-mesenchymal transition (EMT-related factors in HEC-1A cells treated with rhOPN. Results: rhOPN promotes cell proliferation, migration and invasion in HEC-1A cells. rhOPN influenced EMT-related factors and MMP-2 expression in HEC-1A cells. rhOPN promoted HEC-1A cells migration, invasion and EMT through protein kinase B (PKB/AKT and Extracellular regulated protein kinases (ERK1/2 signaling pathway. Conclusions: These results may open up a novel therapeutic strategy for endometrial cancer: namely, rhOPN have important roles in controlling growth of endometrial of cancer cells and suggest a novel target pathway for treatment of this cancer.

  6. Patterning bacterial communities on epithelial cells.

    Directory of Open Access Journals (Sweden)

    Mohammed Dwidar

    Full Text Available Micropatterning of bacteria using aqueous two phase system (ATPS enables the localized culture and formation of physically separated bacterial communities on human epithelial cell sheets. This method was used to compare the effects of Escherichia coli strain MG1655 and an isogenic invasive counterpart that expresses the invasin (inv gene from Yersinia pseudotuberculosis on the underlying epithelial cell layer. Large portions of the cell layer beneath the invasive strain were killed or detached while the non-invasive E. coli had no apparent effect on the epithelial cell layer over a 24 h observation period. In addition, simultaneous testing of the localized effects of three different bacterial species; E. coli MG1655, Shigella boydii KACC 10792 and Pseudomonas sp DSM 50906 on an epithelial cell layer is also demonstrated. The paper further shows the ability to use a bacterial predator, Bdellovibriobacteriovorus HD 100, to selectively remove the E. coli, S. boydii and P. sp communities from this bacteria-patterned epithelial cell layer. Importantly, predation and removal of the P. Sp was critical for maintaining viability of the underlying epithelial cells. Although this paper focuses on a few specific cell types, the technique should be broadly applicable to understand a variety of bacteria-epithelial cell interactions.

  7. Chimaphilin inhibits human osteosarcoma cell invasion and metastasis through suppressing the TGF-β1-induced epithelial-to-mesenchymal transition markers via PI-3K/Akt, ERK1/2, and Smad signaling pathways.

    Science.gov (United States)

    Dong, Feng; Liu, Tingting; Jin, Hao; Wang, Wenbo

    2018-01-01

    Epithelial-to-mesenchymal transition is a cellular process associated with cancer invasion and metastasis. However, the antimetastatic effects of chimaphilin remain elusive. In this study, we attempted to investigate the potential use of chimaphilin as an inhibitor of TGF-β1-induced epithelial-to-mesenchymal transition in U2OS cells. We found that TGF-β1 induced epithelial-to-mesenchymal transition to promote U2OS cell invasion and metastasis. Western blotting demonstrated that chimaphilin inhibited U2OS cell invasion and migration, increased the expression of the epithelial phenotype marker E-cadherin, repressed the expression of the mesenchymal phenotype marker vimentin, as well as decreased the level of epithelial-to-mesenchymal-inducing transcription factors Snail1 and Slug during the initiation of TGF-β1-induced epithelial-to-mesenchymal transition. In this study, we revealed that chimaphilin up-regulated the E-cadherin expression level and inhibited the production of vimentin, Snail1, and Slug in TGF-β1-induced U2OS cells by blocking PI-3K/Akt and ERK 1/2 signaling pathway. Additionally, the TGF-β1-mediated phosphorylated levels of Smad2/3 were inhibited by chimaphilin pretreatment. Above all, we conclude that chimaphilin represents an effective inhibitor of the metastatic potential of U2OS cells through suppression of TGF-β1-induced epithelial-to-mesenchymal transition.

  8. InvS Coordinates Expression of PrgH and FimZ and Is Required for Invasion of Epithelial Cells by Salmonella enterica serovar Typhimurium

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lu; Cai, Xia; Wu, Shuyan; Bomjan, Rajdeep; Nakayasu, Ernesto S.; Händler, Kristian; Hinton, Jay C. D.; Zhou, Daoguo; DiRita, Victor J.

    2017-04-24

    ABSTRACT

    Deep sequencing has revolutionized our understanding of the bacterial RNA world and has facilitated the identification of 280 small RNAs (sRNAs) inSalmonella. Despite the suspicions that sRNAs may play important roles inSalmonellapathogenesis, the functions of most sRNAs remain unknown. To advance our understanding of RNA biology inSalmonellavirulence, we searched for sRNAs required for bacterial invasion into nonphagocytic cells. After screening 75 sRNAs, we discovered that the ablation of InvS caused a significant decrease ofSalmonellainvasion into epithelial cells. A proteomic analysis showed that InvS modulated the levels of several type III secretedSalmonellaproteins. The level of PrgH, a type III secretion apparatus protein, was significantly lower in the absence of InvS, consistent with the known roles of PrgH in effector secretion and bacterial invasion. We discovered that InvS modulatesfimZexpression and hence flagellar gene expression and motility. We propose that InvS coordinates the increase of PrgH and decrease in FimZ that promote efficientSalmonellainvasion into nonphagocytic cells.

    IMPORTANCESalmonellosis continues to be the most common foodborne infection reported by the CDC in the United States. Central toSalmonellapathogenesis is the ability to invade nonphagocytic cells and to replicate inside host cells. Invasion genes are known to be regulated by protein transcriptional networks, but little is known

  9. Epithelial Cell Cultures

    Directory of Open Access Journals (Sweden)

    Imran S. Chaudhry

    2011-01-01

    Full Text Available The biological effects of only a finite number of tobacco toxins have been studied. Here, we describe exposure of cultures of human bronchial epithelial cells to low concentrations of tobacco carcinogens: nickel sulphate, benzo(bfluoranthene, N-nitrosodiethylamine, and 4-(methylnitrosamino-1-(3-pyridyl-1-butanone (NNK. After a 24-hour exposure, EGFR was expressed in cell membrane and cytoplasm, BCL-2 was expressed only in the irregular nuclei of large atypical cells, MKI67 was expressed in nuclei with no staining in larger cells, cytoplasmic BIRC5 with stronger nuclear staining was seen in large atypical cells, and nuclear TP53 was strongly expressed in all cells. After only a 24-hour exposure, cells exhibited atypical nuclear and cytoplasmic features. After a 48-hour exposure, EGFR staining was localized to the nucleus, BCL-2 was slightly decreased in intensity, BIRC5 was localized to the cytoplasm, and TP53 staining was increased in small and large cells. BCL2L1 was expressed in both the cytoplasm and nuclei of cells at 24- and 48-hour exposures. We illustrate that short-termexposure of a bronchial epithelial cell line to smoking-equivalent concentrations of tobacco carcinogens alters the expression of key proliferation regulatory genes, EGFR, BCL-2, BCL2L1, BIRC5, TP53, and MKI67, similar to that reported in biopsy specimens of pulmonary epithelium described to be preneoplastic lesions.

  10. PKCδ-mediated phosphorylation of BAG3 at Ser187 site induces epithelial-mesenchymal transition and enhances invasiveness in thyroid cancer FRO cells.

    Science.gov (United States)

    Li, N; Du, Z-X; Zong, Z-H; Liu, B-Q; Li, C; Zhang, Q; Wang, H-Q

    2013-09-19

    Protein kinase C delta (PKCδ) is a serine (Ser)/threonine kinase, which regulates numerous cellular processes, including proliferation, differentiation, migration and apoptosis. In the current study, Chinese hamster ovary cells were transfected with either a constitutively activated PKCδ or a dominant negative PKCδ, phosphoprotein enrichment, two-dimensional difference gel electrophoresis and mass spectrometry was combined to globally identified candidates of PKCδ cascade. We found that Bcl-2 associated athanogene 3 (BAG3) was one of the targets of PKCδ cascade, and BAG3 interacted with PKCδ in vivo. In addition, we clarified that BAG3 was phosphorylate at Ser187 site in a PKCδ-dependent manner in vivo. BAG3 has been implicated in multiple cellular functions, including proliferation, differentiation, apoptosis, migration, invasion, macroautophagy and so on. We generated wild-type (WT)-, Ser187Ala (S187A)- or Ser187Asp (S187D)-BAG3 stably expressing FRO cells, and noticed that phosphorylation state of BAG3 influenced FRO morphology. Finally, for the first time, we showed that BAG3 was implicated in epithelial-mesenchymal transition (EMT) procedure, and phosphorylation state at Ser187 site had a critical role in EMT regulation by BAG3. Collectively, the current study indicates that BAG3 is a novel substrate of PKCδ, and PKCδ-mediated phosphorylation of BAG3 is implicated in EMT and invasiveness of thyroid cancer cells.

  11. miR-1271 inhibits migration, invasion and epithelial-mesenchymal transition by targeting ZEB1 and TWIST1 in pancreatic cancer cells

    International Nuclear Information System (INIS)

    Liu, Huaize; Wang, Han; Liu, Xiaoxiao; Yu, Tingting

    2016-01-01

    Pancreatic cancer (PC) remains one of the most lethal types of cancer in adults. The purpose of this study was to determine the role of miR-1271 in regulation of epithelial mesenchymal transition (EMT) and metastasis of pancreatic cancer cells. miR-1271 was identified to be significantly down-regulated in PC tissues by miRNA array. Also, an increase of EMT-regulators ZEB1 and TWIST1 expression level is accompanied by a decrease of miR-1271. We showed that expression of miR-1271 was significantly down-regulated in PC tissues as compared with that in normal tissues. In addition, our results showed that miR-1271 expression levels were decreased while ZEB1 and TWIST1 expression levels were increased in detected PC cell lines. Moreover, ectopic expression of miR-1271 suppressed and antagomiR-1271 promoted proliferation, migration, and invasion in SW1990 and PANC-1 cells. Bioinformatics coupled with luciferase and Western blot assays also revealed that miR-1271 inhibited expression of ZEB1 and TWIST1, which are master regulators of tumor metastasis. Our study first indicates that miR-1271 functions as a suppressor in regulating of pancreatic cancer EMT by targeting ZEB1 and TWIST1, and it promise as a therapeutic target and prognostic marker for metastatic pancreatic cancer. - Highlights: • miR-1271 is downregulated in pancreatic cancer tissues and cell lines. • miR-1271 regulates cell metastasis ability and EMT marker expression. . • miR-1271 directly targets ZEB1 and TWIST1. • ZEB1 and TWIST1 are functionally related to the effects of miR-1271.

  12. miR-1271 inhibits migration, invasion and epithelial-mesenchymal transition by targeting ZEB1 and TWIST1 in pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Huaize [Department of Developmental Genetics, Nanjing Medical University, Nanjing 210029 (China); Wang, Han [The First Clinical Medical College of Nanjing Medical University, Nanjing 210029 (China); Liu, Xiaoxiao [Department of Biotechnology, Nanjing Medical University, Nanjing 210029 (China); Yu, Tingting, E-mail: tingting@njmu.edu.cn [Department of Developmental Genetics, Nanjing Medical University, Nanjing 210029 (China)

    2016-04-01

    Pancreatic cancer (PC) remains one of the most lethal types of cancer in adults. The purpose of this study was to determine the role of miR-1271 in regulation of epithelial mesenchymal transition (EMT) and metastasis of pancreatic cancer cells. miR-1271 was identified to be significantly down-regulated in PC tissues by miRNA array. Also, an increase of EMT-regulators ZEB1 and TWIST1 expression level is accompanied by a decrease of miR-1271. We showed that expression of miR-1271 was significantly down-regulated in PC tissues as compared with that in normal tissues. In addition, our results showed that miR-1271 expression levels were decreased while ZEB1 and TWIST1 expression levels were increased in detected PC cell lines. Moreover, ectopic expression of miR-1271 suppressed and antagomiR-1271 promoted proliferation, migration, and invasion in SW1990 and PANC-1 cells. Bioinformatics coupled with luciferase and Western blot assays also revealed that miR-1271 inhibited expression of ZEB1 and TWIST1, which are master regulators of tumor metastasis. Our study first indicates that miR-1271 functions as a suppressor in regulating of pancreatic cancer EMT by targeting ZEB1 and TWIST1, and it promise as a therapeutic target and prognostic marker for metastatic pancreatic cancer. - Highlights: • miR-1271 is downregulated in pancreatic cancer tissues and cell lines. • miR-1271 regulates cell metastasis ability and EMT marker expression. . • miR-1271 directly targets ZEB1 and TWIST1. • ZEB1 and TWIST1 are functionally related to the effects of miR-1271.

  13. Endothelial Induced EMT in Breast Epithelial Cells with Stem Cell Properties

    OpenAIRE

    Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J. R.; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A.; Petersen, Ole William; Magnusson, Magnus K.; Gudjonsson, Thorarinn

    2011-01-01

    Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell proper...

  14. Restriction endonucleases from invasive Neisseria gonorrhoeae cause double-strand breaks and distort mitosis in epithelial cells during infection.

    Directory of Open Access Journals (Sweden)

    Linda Weyler

    Full Text Available The host epithelium is both a barrier against, and the target for microbial infections. Maintaining regulated cell growth ensures an intact protective layer towards microbial-induced cellular damage. Neisseria gonorrhoeae infections disrupt host cell cycle regulation machinery and the infection causes DNA double strand breaks that delay progression through the G2/M phase. We show that intracellular gonococci upregulate and release restriction endonucleases that enter the nucleus and damage human chromosomal DNA. Bacterial lysates containing restriction endonucleases were able to fragment genomic DNA as detected by PFGE. Lysates were also microinjected into the cytoplasm of cells in interphase and after 20 h, DNA double strand breaks were identified by 53BP1 staining. In addition, by using live-cell microscopy and NHS-ester stained live gonococci we visualized the subcellular location of the bacteria upon mitosis. Infected cells show dysregulation of the spindle assembly checkpoint proteins MAD1 and MAD2, impaired and prolonged M-phase, nuclear swelling, micronuclei formation and chromosomal instability. These data highlight basic molecular functions of how gonococcal infections affect host cell cycle regulation, cause DNA double strand breaks and predispose cellular malignancies.

  15. Restriction endonucleases from invasive Neisseria gonorrhoeae cause double-strand breaks and distort mitosis in epithelial cells during infection.

    Science.gov (United States)

    Weyler, Linda; Engelbrecht, Mattias; Mata Forsberg, Manuel; Brehwens, Karl; Vare, Daniel; Vielfort, Katarina; Wojcik, Andrzej; Aro, Helena

    2014-01-01

    The host epithelium is both a barrier against, and the target for microbial infections. Maintaining regulated cell growth ensures an intact protective layer towards microbial-induced cellular damage. Neisseria gonorrhoeae infections disrupt host cell cycle regulation machinery and the infection causes DNA double strand breaks that delay progression through the G2/M phase. We show that intracellular gonococci upregulate and release restriction endonucleases that enter the nucleus and damage human chromosomal DNA. Bacterial lysates containing restriction endonucleases were able to fragment genomic DNA as detected by PFGE. Lysates were also microinjected into the cytoplasm of cells in interphase and after 20 h, DNA double strand breaks were identified by 53BP1 staining. In addition, by using live-cell microscopy and NHS-ester stained live gonococci we visualized the subcellular location of the bacteria upon mitosis. Infected cells show dysregulation of the spindle assembly checkpoint proteins MAD1 and MAD2, impaired and prolonged M-phase, nuclear swelling, micronuclei formation and chromosomal instability. These data highlight basic molecular functions of how gonococcal infections affect host cell cycle regulation, cause DNA double strand breaks and predispose cellular malignancies.

  16. High Glucose Promotes Tumor Invasion and Increases Metastasis-Associated Protein Expression in Human Lung Epithelial Cells by Upregulating Heme Oxygenase-1 via Reactive Oxygen Species or the TGF-β1/PI3K/Akt Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Xiaowen Kang

    2015-02-01

    Full Text Available Background: Growing evidence indicates that heme oxygenase-1 (HO-1 is up-regulated in malignancies and subsequently alters tumor aggressiveness and various cancer-related factors, such as high glucose (HG levels. HO-1 expression can be induced when glucose concentrations are above 25 mM; however, the role of HO-1 in lung cancer patients with diabetes remains unknown. Therefore, in this study we investigated the promotion of tumor cell invasion and the expression of metastasis-associated proteins by inducing the up-regulation of HO-1 expression by HG treatment in A549 human lung epithelial cells. Methods: The expression of HO-1and metastasis-associated protein expression was explored by western blot analysis. HO-1 enzymatic activity, reactive oxygen species (ROS production and TGF-β1 production were examined by ELISA. Invasiveness was analyzed using a Transwell chamber. Results: HG treatment of A549 cells induced an increase in HO-1 expression, which was mediated by the HG-induced generation of reactive oxygen species (ROS and transforming growth factor-β1 (TGF-β1 in a concentration- and time-dependent manner. Following the increase in HO-1 expression, the enzymatic activity of HO-1 also increased in HG-treated cells. Pretreatment with N-acetyl-L-cysteine (NAC or with phosphatidylinositol 3-kinase (PI3K/Akt inhibitors attenuated the HG-induced increase in HO-1 expression. HG treatment of A549 cells enhanced the invasion potential of these cells, as shown with a Transwell assay, and increased metastasis-associated protein expression. However, HO-1 siRNA transfection significantly decreased these capabilities. Conclusion: this study is the first to demonstrate that HG treatment of A549 human lung epithelial cells promotes tumor cell invasion and increases metastasis-associated protein expression by up-regulating HO-1 expression via ROS or the TGF-β1/PI3K/Akt signaling pathway.

  17. Stromal-epithelial interactions in aging and cancer: Senescent fibroblasts alter epithelial cell differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Parrinello, Simona; Coppe, Jean-Philippe; Krtolica, Ana; Campisi, Judith

    2004-07-14

    Cellular senescence suppresses cancer by arresting cells at risk for malignant tumorigenesis. However, senescent cells also secrete molecules that can stimulate premalignant cells to proliferate and form tumors, suggesting the senescence response is antagonistically pleiotropic. We show that premalignant mammary epithelial cells exposed to senescent human fibroblasts in mice irreversibly lose differentiated properties, become invasive and undergo full malignant transformation. Moreover, using cultured mouse or human fibroblasts and non-malignant breast epithelial cells, we show that senescent fibroblasts disrupt epithelial alveolar morphogenesis, functional differentiation, and branching morphogenesis. Further, we identify MMP-3 as the major factor responsible for the effects of senescent fibroblasts on branching morphogenesis. Our findings support the idea that senescent cells contribute to age-related pathology, including cancer, and describe a new property of senescent fibroblasts--the ability to alter epithelial differentiation--that might also explain the loss of tissue function and organization that is a hallmark of aging.

  18. Epithelial cell polarity, stem cells and cancer

    DEFF Research Database (Denmark)

    Martin-Belmonte, Fernando; Perez-Moreno, Mirna

    2011-01-01

    , deregulation of adhesion and polarity proteins can cause misoriented cell divisions and increased self-renewal of adult epithelial stem cells. In this Review, we highlight some advances in the understanding of how loss of epithelial cell polarity contributes to tumorigenesis.......After years of extensive scientific discovery much has been learned about the networks that regulate epithelial homeostasis. Loss of expression or functional activity of cell adhesion and cell polarity proteins (including the PAR, crumbs (CRB) and scribble (SCRIB) complexes) is intricately related...

  19. Buccal Epithelial Cells as Potential Non- invasive Materials for the Monitoring of Mitochondrial Disturbances to Track Huntington‘s Disease Progression. A Pilot Study

    Czech Academy of Sciences Publication Activity Database

    Rodinová, M.; Trefilová, E.; Lišková, Irena; Klempíř, J.; Ellederová, Zdeňka; Vidinská, Daniela; Motlík, Jan; Zeman, J.; Hansíková, H.

    2015-01-01

    Roč. 78, Suppl 2 (2015), s. 49-54 ISSN 1210-7859. [Conference on Animal Models for neurodegenerative Diseases /3./. Liblice, 08.11.2015-10.11.2015] R&D Projects: GA MŠk ED2.1.00/03.0124 Institutional support: RVO:67985904 Keywords : Huntington’s disease * mitochondrial respiratory chain complex I and IV * buccal epithelial cells Subject RIV: FH - Neurology Impact factor: 0.209, year: 2015

  20. Osteopontin Promotes Invasion, Migration and Epithelial-Mesenchymal Transition of Human Endometrial Carcinoma Cell HEC-1A Through AKT and ERK1/2 Signaling

    OpenAIRE

    Yinghua Li; Yunpeng Xie; Dan Cui; Yanni Ma; Linlin Sui; Chenyang Zhu; Hui Kong; Ying Kong

    2015-01-01

    Background/Aims: Osteopontin (OPN) is an Extracellular Matrix (ECM) molecule and is involved in many physiologic and pathologic processes, including cell adhesion, angiogenesis and tumor metastasis. OPN is a well-known multifunctional factor involved in various aspects of cancer progression, including endometrial cancer. In this study, we examined the significance of OPN in endometrial cancer. Methods: The proliferation, migration and invasion ability of HEC-1A cells were detected by Cell Cou...

  1. Adhesion to the host cell surface is sufficient to mediate Listeria monocytogenes entry into epithelial cells

    Science.gov (United States)

    Ortega, Fabian E.; Rengarajan, Michelle; Chavez, Natalie; Radhakrishnan, Prathima; Gloerich, Martijn; Bianchini, Julie; Siemers, Kathleen; Luckett, William S.; Lauer, Peter; Nelson, W. James; Theriot, Julie A.

    2017-01-01

    The intestinal epithelium is the first physiological barrier breached by the Gram-positive facultative pathogen Listeria monocytogenes during an in vivo infection. Listeria monocytogenes binds to the epithelial host cell receptor E-cadherin, which mediates a physical link between the bacterium and filamentous actin (F-actin). However, the importance of anchoring the bacterium to F-actin through E-cadherin for bacterial invasion has not been tested directly in epithelial cells. Here we demonstrate that depleting αE-catenin, which indirectly links E-cadherin to F-actin, did not decrease L. monocytogenes invasion of epithelial cells in tissue culture. Instead, invasion increased due to increased bacterial adhesion to epithelial monolayers with compromised cell–cell junctions. Furthermore, expression of a mutant E-cadherin lacking the intracellular domain was sufficient for efficient L. monocytogenes invasion of epithelial cells. Importantly, direct biotin-mediated binding of bacteria to surface lipids in the plasma membrane of host epithelial cells was sufficient for uptake. Our results indicate that the only requirement for L. monocytogenes invasion of epithelial cells is adhesion to the host cell surface, and that E-cadherin–mediated coupling of the bacterium to F-actin is not required. PMID:28877987

  2. Effect of hGC-MSCs from human gastric cancer tissue on cell proliferation, invasion and epithelial-mesenchymal transition in tumor tissue of gastric cancer tumor-bearing mice.

    Science.gov (United States)

    Song, Lin; Zhou, Xin; Jia, Hong-Jun; Du, Mei; Zhang, Jin-Ling; Li, Liang

    2016-08-01

    To study the effect of hGC-MSCs from human gastric cancer tissue on cell proliferation, invasion and epithelial-mesenchymal transition in tumor tissue of gastric cancer tumor-bearing mice. BABL/c nude mice were selected as experimental animals and gastric cancer tumor-bearing mice model were established by subcutaneous injection of gastric cancer cells, randomly divided into different intervention groups. hGC-MSCs group were given different amounts of gastric cancer cells for subcutaneous injection, PBS group was given equal volume of PBS for subcutaneous injection. Then tumor tissue volume were determined, tumor-bearing mice were killed and tumor tissues were collected, mRNA expression of proliferation, invasion, EMT-related molecules were determined. 4, 8, 12, 16, 20 d after intervention, tumor tissue volume of hGC-MSCs group were significantly higher than those of PBS group and the more the number of hGC-MSCs, the higher the tumor tissue volume; mRNA contents of Ki-67, PCNA, Bcl-2, MMP-2, MMP-7, MMP-9, MMP-14, N-cadherin, vimentin, Snail and Twist in tumor tissue of hGC-MSCs group were higher than those of PBS group, and mRNA contents of Bax, TIMP1, TIMP2 and E-cadherin were lower than those of PBS group. hGC-MSCs from human gastric cancer tissue can promote the tumor growth in gastric cancer tumor-bearing mice, and the molecular mechanism includes promoting cell proliferation, invasion and epithelial-mesenchymal transition. Copyright © 2016 Hainan Medical College. Production and hosting by Elsevier B.V. All rights reserved.

  3. An mDia2/ROCK signaling axis regulates invasive egress from epithelial ovarian cancer spheroids.

    Science.gov (United States)

    Pettee, Krista M; Dvorak, Kaitlyn M; Nestor-Kalinoski, Andrea L; Eisenmann, Kathryn M

    2014-01-01

    Multi-cellular spheroids are enriched in ascites of epithelial ovarian cancer (OvCa) patients. They represent an invasive and chemoresistant cellular population fundamental to metastatic dissemination. The molecular mechanisms triggering single cell invasive egress from spheroids remain enigmatic. mDia formins are Rho GTPase effectors that are key regulators of F-actin cytoskeletal dynamics. We hypothesized that mDia2-driven F-actin dynamics promote single cell invasive transitions in clinically relevant three-dimensional (3D) OvCa spheroids. The current study is a dissection of the contribution of the F-actin assembly factor mDia2 formin in invasive transitions and using a clinically relevant ovarian cancer spheroid model. We show that RhoA-directed mDia2 activity is required for tight spheroid organization, and enrichment of mDia2 in the invasive cellular protrusions of collagen-embedded OVCA429 spheroids. Depleting mDia2 in ES-2 spheroids enhanced invasive dissemination of single amoeboid-shaped cells. This contrasts with spheroids treated with control siRNA, where a mesenchymal invasion program predominated. Inhibition of another RhoA effector, ROCK, had no impact on ES-2 spheroid formation but dramatically inhibited spheroid invasion through induction of a highly elongated morphology. Concurrent inhibition of ROCK and mDia2 blocked single cell invasion from ES-2 spheroids more effectively than inhibition of either protein alone, indicating that invasive egress of amoeboid cells from mDia2-depleted spheroids is ROCK-dependent. Our findings indicate that multiple GTPase effectors must be suppressed in order to fully block invasive egress from ovarian cancer spheroids. Furthermore, tightly regulated interplay between ROCK and mDia2 signaling pathways dictates the invasive capacities and the type of invasion program utilized by motile spheroid-derived ovarian cancer cells. As loss of the gene encoding mDia2, DRF3, has been linked to cancer progression and

  4. An mDia2/ROCK signaling axis regulates invasive egress from epithelial ovarian cancer spheroids.

    Directory of Open Access Journals (Sweden)

    Krista M Pettee

    Full Text Available Multi-cellular spheroids are enriched in ascites of epithelial ovarian cancer (OvCa patients. They represent an invasive and chemoresistant cellular population fundamental to metastatic dissemination. The molecular mechanisms triggering single cell invasive egress from spheroids remain enigmatic. mDia formins are Rho GTPase effectors that are key regulators of F-actin cytoskeletal dynamics. We hypothesized that mDia2-driven F-actin dynamics promote single cell invasive transitions in clinically relevant three-dimensional (3D OvCa spheroids. The current study is a dissection of the contribution of the F-actin assembly factor mDia2 formin in invasive transitions and using a clinically relevant ovarian cancer spheroid model. We show that RhoA-directed mDia2 activity is required for tight spheroid organization, and enrichment of mDia2 in the invasive cellular protrusions of collagen-embedded OVCA429 spheroids. Depleting mDia2 in ES-2 spheroids enhanced invasive dissemination of single amoeboid-shaped cells. This contrasts with spheroids treated with control siRNA, where a mesenchymal invasion program predominated. Inhibition of another RhoA effector, ROCK, had no impact on ES-2 spheroid formation but dramatically inhibited spheroid invasion through induction of a highly elongated morphology. Concurrent inhibition of ROCK and mDia2 blocked single cell invasion from ES-2 spheroids more effectively than inhibition of either protein alone, indicating that invasive egress of amoeboid cells from mDia2-depleted spheroids is ROCK-dependent. Our findings indicate that multiple GTPase effectors must be suppressed in order to fully block invasive egress from ovarian cancer spheroids. Furthermore, tightly regulated interplay between ROCK and mDia2 signaling pathways dictates the invasive capacities and the type of invasion program utilized by motile spheroid-derived ovarian cancer cells. As loss of the gene encoding mDia2, DRF3, has been linked to cancer

  5. Epithelial cells as alternative human biomatrices for comet assay.

    Science.gov (United States)

    Rojas, Emilio; Lorenzo, Yolanda; Haug, Kristiane; Nicolaissen, Bjørn; Valverde, Mahara

    2014-01-01

    The comet assay is a valuable experimental tool aimed at mapping DNA damage in human cells in vivo for environmental and occupational monitoring, as well as for therapeutic purposes, such as storage prior to transplant, during tissue engineering, and in experimental ex vivo assays. Furthermore, due to its great versatility, the comet assay allows to explore the use of alternative cell types to assess DNA damage, such as epithelial cells. Epithelial cells, as specialized components of many organs, have the potential to serve as biomatrices that can be used to evaluate genotoxicity and may also serve as early effect biomarkers. Furthermore, 80% of solid cancers are of epithelial origin, which points to the importance of studying DNA damage in these tissues. Indeed, studies including comet assay in epithelial cells have either clear clinical applications (lens and corneal epithelial cells) or examine genotoxicity within human biomonitoring and in vitro studies. We here review improvements in determining DNA damage using the comet assay by employing lens, corneal, tear duct, buccal, and nasal epithelial cells. For some of these tissues invasive sampling procedures are needed. Desquamated epithelial cells must be obtained and dissociated prior to examination using the comet assay, and such procedures may induce varying amounts of DNA damage. Buccal epithelial cells require lysis enriched with proteinase K to obtain free nucleosomes. Over a 30 year period, the comet assay in epithelial cells has been little employed, however its use indicates that it could be an extraordinary tool not only for risk assessment, but also for diagnosis, prognosis of treatments and diseases.

  6. Epithelial invasion outcompetes hypha development during Candida albicans infection as revealed by an image-based systems biology approach.

    Science.gov (United States)

    Mech, Franziska; Wilson, Duncan; Lehnert, Teresa; Hube, Bernhard; Thilo Figge, Marc

    2014-02-01

    Candida albicans is the most common opportunistic fungal pathogen of the human mucosal flora, frequently causing infections. The fungus is responsible for invasive infections in immunocompromised patients that can lead to sepsis. The yeast to hypha transition and invasion of host-tissue represent major determinants in the switch from benign colonizer to invasive pathogen. A comprehensive understanding of the infection process requires analyses at the quantitative level. Utilizing fluorescence microscopy with differential staining, we obtained images of C. albicans undergoing epithelial invasion during a time course of 6 h. An image-based systems biology approach, combining image analysis and mathematical modeling, was applied to quantify the kinetics of hyphae development, hyphal elongation, and epithelial invasion. The automated image analysis facilitates high-throughput screening and provided quantities that allow for the time-resolved characterization of the morphological and invasive state of fungal cells. The interpretation of these data was supported by two mathematical models, a kinetic growth model and a kinetic transition model, that were developed using differential equations. The kinetic growth model describes the increase in hyphal length and revealed that hyphae undergo mass invasion of epithelial cells following primary hypha formation. We also provide evidence that epithelial cells stimulate the production of secondary hyphae by C. albicans. Based on the kinetic transition model, the route of invasion was quantified in the state space of non-invasive and invasive fungal cells depending on their number of hyphae. This analysis revealed that the initiation of hyphae formation represents an ultimate commitment to invasive growth and suggests that in vivo, the yeast to hypha transition must be under exquisitely tight negative regulation to avoid the transition from commensal to pathogen invading the epithelium. © 2013 International Society for

  7. PTEN Loss in E-Cadherin-Deficient Mouse Mammary Epithelial Cells Rescues Apoptosis and Results in Development of Classical Invasive Lobular Carcinoma

    Directory of Open Access Journals (Sweden)

    Mirjam C. Boelens

    2016-08-01

    Full Text Available Invasive lobular carcinoma (ILC is an aggressive breast cancer subtype with poor response to chemotherapy. Besides loss of E-cadherin, a hallmark of ILC, genetic inactivation of PTEN is frequently observed in patients. Through concomitant Cre-mediated inactivation of E-cadherin and PTEN in mammary epithelium, we generated a mouse model of classical ILC (CLC, the main histological ILC subtype. While loss of E-cadherin induced cell dissemination and apoptosis, additional PTEN inactivation promoted cell survival and rapid formation of invasive mammary tumors that recapitulate the histological and molecular features, estrogen receptor (ER status, growth kinetics, metastatic behavior, and tumor microenvironment of human CLC. Combined inactivation of E-cadherin and PTEN is sufficient to cause CLC development. These CLCs showed significant tumor regression upon BEZ235-mediated inhibition of PI3K signaling. In summary, this mouse model provides important insights into CLC development and suggests inhibition of phosphatidylinositol 3-kinase (PI3K signaling as a potential therapeutic strategy for targeting CLC.

  8. PTEN Loss in E-Cadherin-Deficient Mouse Mammary Epithelial Cells Rescues Apoptosis and Results in Development of Classical Invasive Lobular Carcinoma.

    Science.gov (United States)

    Boelens, Mirjam C; Nethe, Micha; Klarenbeek, Sjoerd; de Ruiter, Julian R; Schut, Eva; Bonzanni, Nicola; Zeeman, Amber L; Wientjens, Ellen; van der Burg, Eline; Wessels, Lodewyk; van Amerongen, Renée; Jonkers, Jos

    2016-08-23

    Invasive lobular carcinoma (ILC) is an aggressive breast cancer subtype with poor response to chemotherapy. Besides loss of E-cadherin, a hallmark of ILC, genetic inactivation of PTEN is frequently observed in patients. Through concomitant Cre-mediated inactivation of E-cadherin and PTEN in mammary epithelium, we generated a mouse model of classical ILC (CLC), the main histological ILC subtype. While loss of E-cadherin induced cell dissemination and apoptosis, additional PTEN inactivation promoted cell survival and rapid formation of invasive mammary tumors that recapitulate the histological and molecular features, estrogen receptor (ER) status, growth kinetics, metastatic behavior, and tumor microenvironment of human CLC. Combined inactivation of E-cadherin and PTEN is sufficient to cause CLC development. These CLCs showed significant tumor regression upon BEZ235-mediated inhibition of PI3K signaling. In summary, this mouse model provides important insights into CLC development and suggests inhibition of phosphatidylinositol 3-kinase (PI3K) signaling as a potential therapeutic strategy for targeting CLC. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  9. MUC-1-ESA+ progenitor cells in normal benign and malignant human breast epithelial cells

    OpenAIRE

    Lu, Xinquan; Li, Huixiang; Xu, Kejia; Nesland, Jahn M.; Suo, Zhenhe

    2009-01-01

    The existence of mammary epithelial stem/progenitor cells has been demonstrated in MUC-1-/ ESA+ subpopulations of breast epithelial cells. However, knowledge about the expression and localization in benign and malignant breast lesions is unknown. Using a double-staining immunohistochemistry method, we investigated MUC-1-/ESA+ cells in 10 normal breast tissues, 49 cases with fibrocystic disease, 40 fibroadenomas, 36 invasive ductal carcinomas and the breast cancer ce...

  10. An epithelial to mesenchymal transition programme does not usually drive the phenotype of invasive lobular carcinomas.

    Science.gov (United States)

    McCart Reed, Amy E; Kutasovic, Jamie R; Vargas, Ana C; Jayanthan, Janani; Al-Murrani, Amel; Reid, Lynne E; Chambers, Rachael; Da Silva, Leonard; Melville, Lewis; Evans, Elizabeth; Porter, Alan; Papadimos, David; Thompson, Erik W; Lakhani, Sunil R; Simpson, Peter T

    2016-03-01

    Epithelial to mesenchymal transition (EMT) is a cellular phenotype switching phenomenon which occurs during normal development and is proposed to promote tumour cell invasive capabilities during tumour progression. Invasive lobular carcinoma (ILC) is a histological special type of breast cancer with a peculiar aetiology - the tumour cells display an invasive growth pattern, with detached, single cells or single files of cells, and a canonical feature is the loss of E-cadherin expression. These characteristics are indicative of an EMT or at the very least that they represent some plasticity between phenotypes. While some gene expression profiling data support this view, the tumour cells remain epithelial and limited immunohistochemistry data suggest that EMT markers may not feature prominently in ILC. We assessed the expression of a panel of EMT markers (fibronectin, vimentin, N-cadherin, smooth muscle actin, osteonectin, Snail, Twist) in 148 ILCs and performed a meta-analysis of publically available molecular data from 154 ILCs. Three out of 148 (2%) ILCs demonstrated an early and coordinated alteration of multiple EMT markers (down-regulation of E-cadherin, nuclear TWIST, and up-regulation of vimentin, osteonectin, and smooth muscle actin). However, the data overall do not support a role for EMT in defining the phenotypic peculiarities of the majority of ILCs. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  11. 1,25(OH)2D3 attenuates TGF-β1/β2-induced increased migration and invasion via inhibiting epithelial-mesenchymal transition in colon cancer cells.

    Science.gov (United States)

    Chen, Shanwen; Zhu, Jing; Zuo, Shuai; Ma, Ju; Zhang, Junling; Chen, Guowei; Wang, Xin; Pan, Yisheng; Liu, Yucun; Wang, Pengyuan

    1,25-Dihydroxyvitamin D3 (1,25(OH)2D3) has been reported to inhibit proliferation and migration of multiple types of cancer cells. However, the mechanism underlying its anti-metastasis effect is not fully illustrated. In this study, the effect of 1,25(OH)2D3 on TGF-β1/β2-induced epithelial-mesenchymal transition (EMT) is tested in colon cancer cells. The results suggest that 1,25(OH)2D3 inhibited TGF-β1/β2-induced increased invasion and migration of in SW-480 and HT-29 cells. 1,25(OH)2D3 also inhibited the cadherin switch in SW-480 and HT-29 cells. TGF-β1/β2-induced increased expression of EMT-related transcription factors was also inhibited by 1,25(OH)2D3. 1,25(OH)2D3 also inhibited the secretion of MMP-2 and MMP-9 and increased expression of F-actin induced by TGF-β1/β2 in SW-480 cells. Taken together, this study suggests that the suppression of EMT might be one of the mechanisms underlying the anti-metastasis effect of 1,25(OH)2D3 in colon cancer cells. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Epithelial Cells in Urine: MedlinePlus Lab Test Information

    Science.gov (United States)

    ... page: https://medlineplus.gov/labtests/epithelialcellsinurine.html Epithelial Cells in Urine To use the sharing features on ... page, please enable JavaScript. What is an Epithelial Cells in Urine Test? Epithelial cells are a type ...

  13. Roles of the outer membrane protein AsmA of Salmonella enterica in the control of marRAB expression and invasion of epithelial cells.

    Science.gov (United States)

    Prieto, Ana I; Hernández, Sara B; Cota, Ignacio; Pucciarelli, M Graciela; Orlov, Yuri; Ramos-Morales, Francisco; García-del Portillo, Francisco; Casadesús, Josep

    2009-06-01

    A genetic screen for suppressors of bile sensitivity in DNA adenine methylase (dam) mutants of Salmonella enterica serovar Typhimurium yielded insertions in an uncharacterized locus homologous to the Escherichia coli asmA gene. Disruption of asmA suppressed bile sensitivity also in phoP and wec mutants of S. enterica and increased the MIC of sodium deoxycholate for the parental strain ATCC 14028. Increased levels of marA mRNA were found in asmA, asmA dam, asmA phoP, and asmA wec strains of S. enterica, suggesting that lack of AsmA activates expression of the marRAB operon. Hence, asmA mutations may enhance bile resistance by inducing gene expression changes in the marRAB-controlled Mar regulon. In silico analysis of AsmA structure predicted the existence of one transmembrane domain. Biochemical analysis of subcellular fractions revealed that the asmA gene of S. enterica encodes a protein of approximately 70 kDa located in the outer membrane. Because AsmA is unrelated to known transport and/or efflux systems, we propose that activation of marRAB in asmA mutants may be a consequence of envelope reorganization. Competitive infection of BALB/c mice with asmA(+) and asmA isogenic strains indicated that lack of AsmA attenuates Salmonella virulence by the oral route but not by the intraperitoneal route. Furthermore, asmA mutants showed a reduced ability to invade epithelial cells in vitro.

  14. Thymic epithelial cells. I. Expression of strong suppressive (veto) activity in mouse thymic epithelial cell cultures

    DEFF Research Database (Denmark)

    Claesson, Mogens Helweg; Ropke, C

    1990-01-01

    We show that thymic epithelial cells grown under serum-free conditions in a chemically defined culture medium can act as veto cells in vitro. The veto activity of thymic epithelial cells results in inactivation of specific alloreactive cytotoxic T-cell precursors at the clonal level. It is conclu......We show that thymic epithelial cells grown under serum-free conditions in a chemically defined culture medium can act as veto cells in vitro. The veto activity of thymic epithelial cells results in inactivation of specific alloreactive cytotoxic T-cell precursors at the clonal level...

  15. Human glomerular epithelial cell proteoglycans

    International Nuclear Information System (INIS)

    Thomas, G.J.; Jenner, L.; Mason, R.M.; Davies, M.

    1990-01-01

    Proteoglycans synthesized by cultures of human glomerular epithelial cells have been isolated and characterized. Three types of heparan sulfate were detected. Heparan sulfate proteoglycan I (HSPG-I; Kav 6B 0.04) was found in the cell layer and medium and accounted for 12% of the total proteoglycans synthesized. HSPG-II (Kav 6B 0.25) accounted for 18% of the proteoglycans and was located in the medium and cell layer. A third population (9% of the proteoglycan population), heparan sulfate glycosaminoglycan (HS-GAG; Kav 6B 0.4-0.8), had properties consistent with single glycosaminoglycan chains or their fragments and was found only in the cell layer. HSPG-I and HSPG-II from the cell layer had hydrophobic properties; they were released from the cell layer by mild trypsin treatment. HS-GAG lacked these properties, consisted of low-molecular-mass heparan sulfate oligosaccharides, and were intracellular. HSPG-I and -II released to the medium lacked hydrophobic properties. The cells also produced three distinct types of chondroitin sulfates. The major species, chondroitin sulfate proteoglycan I (CSPG-I) eluted in the excluded volume of a Sepharose CL-6B column, accounted for 30% of the proteoglycans detected, and was found in both the cell layer and medium. Cell layer CSPG-I bound to octyl-Sepharose. It was released from the cell layer by mild trypsin treatment. CSPG-II (Kav 6B 0.1-0.23) accounted for 10% of the total 35S-labeled macromolecules and was found predominantly in the culture medium. A small amount of CS-GAG (Kav 6B 0.25-0.6) is present in the cell extract and like HS-GAG is intracellular. Pulse-chase experiments indicated that HSPG-I and -II and CSPG-I and -II are lost from the cell layer either by direct release into the medium or by internalization where they are metabolized to single glycosaminoglycan chains and subsequently to inorganic sulfate

  16. Basolateral BMP signaling in polarized epithelial cells.

    Directory of Open Access Journals (Sweden)

    Masao Saitoh

    Full Text Available Bone morphogenetic proteins (BMPs regulate various biological processes, mostly mediated by cells of mesenchymal origin. However, the roles of BMPs in epithelial cells are poorly understood. Here, we demonstrate that, in polarized epithelial cells, BMP signals are transmitted from BMP receptor complexes exclusively localized at the basolateral surface of the cell membrane. In addition, basolateral stimulation with BMP increased expression of components of tight junctions and enhanced the transepithelial resistance (TER, counteracting reduction of TER by treatment with TGF-β or an anti-tumor drug. We conclude that BMPs maintain epithelial polarity via intracellular signaling from basolaterally localized BMP receptors.

  17. DNA repair in human bronchial epithelial cells

    International Nuclear Information System (INIS)

    Fornace, A.J. Jr.; Lechner, J.F.; Grafstrom, R.C.; Harris, C.C.

    1982-01-01

    The purpose of this investigation was to compare the response of human cell types (bronchial epithelial cells and fibroblasts and skin fibroblasts) to various DNA damaging agents. Repair of DNA single strand breaks (SSB) induced by 5 krads of X-ray was similar for all cell types; approximately 90% of the DNA SSB were rejoined within one hour. During excision repair of DNA damage from u.v.-radiation, the frequencies of DNA SSB as estimated by the alkaline elution technique, were similar in all cell types. Repair replication as measured by BND cellulose chromatography was also similar in epithelial and fibroblastic cells after u.v.-irradiation. Similar levels of SSB were also observed in epithelial and fibroblastic cells after exposure to chemical carcinogens: 7,12-dimethylbenz[a]anthracene; benzo[a]pyrene diol epoxide (BPDE); or N-methyl-N-nitro-N-nitrosoguanidine. Significant repair replication of BPDE-induced DNA damage was detected in both bronchial epithelial and fibroblastic cells, although the level in fibroblasts was approximately 40% of that in epithelial cells. The pulmonary carcinogen asbestos did not damage DNA. DNA-protein crosslinks induced by formaldehyde were rapidly removed in bronchial cells. Further, epithelial and fibroblastic cells, which were incubated with formaldehyde and the polymerase inhibitor combination of cytosine arabinoside and hydroxyurea, accumulated DNA SSB at approximately equal frequencies. These results should provide a useful background for further investigations of the response of human bronchial cells to various DNA damaging agents

  18. Serratia marcescens is injurious to intestinal epithelial cells.

    Science.gov (United States)

    Ochieng, John B; Boisen, Nadia; Lindsay, Brianna; Santiago, Araceli; Ouma, Collins; Ombok, Maurice; Fields, Barry; Stine, O Colin; Nataro, James P

    2014-01-01

    Diarrhea causes substantial morbidity and mortality in children in low-income countries. Although numerous pathogens cause diarrhea, the etiology of many episodes remains unknown. Serratia marcescens is incriminated in hospital-associated infections, and HIV/AIDS associated diarrhea. We have recently found that Serratia spp. may be found more commonly in the stools of patients with diarrhea than in asymptomatic control children. We therefore investigated the possible enteric pathogenicity of S. marcescens in vitro employing a polarized human colonic epithelial cell (T84) monolayer. Infected monolayers were assayed for bacterial invasion, transepithelial electrical resistance (TEER), cytotoxicity, interleukin-8 (IL-8) release and morphological changes by scanning electron microscopy. We observed significantly greater epithelial cell invasion by S. marcescens compared to Escherichia coli strain HS (p = 0.0038 respectively). Cell invasion was accompanied by reduction in TEER and secretion of IL-8. Lactate dehydrogenase (LDH) extracellular concentration rapidly increased within a few hours of exposure of the monolayer to S. marcescens. Scanning electron microscopy of S. marcescens-infected monolayers demonstrated destruction of microvilli and vacuolization. Our results suggest that S. marcescens interacts with intestinal epithelial cells in culture and induces dramatic alterations similar to those produced by known enteric pathogens.

  19. Mesenchymal Stromal Cell-Derived Interleukin-6 Promotes Epithelial-Mesenchymal Transition and Acquisition of Epithelial Stem-Like Cell Properties in Ameloblastoma Epithelial Cells.

    Science.gov (United States)

    Jiang, Chunmiao; Zhang, Qunzhou; Shanti, Rabie M; Shi, Shihong; Chang, Ting-Han; Carrasco, Lee; Alawi, Faizan; Le, Anh D

    2017-09-01

    Epithelial-mesenchymal transition (EMT), a biological process associated with cancer stem-like or cancer-initiating cell formation, contributes to the invasiveness, metastasis, drug resistance, and recurrence of the malignant tumors; it remains to be determined whether similar processes contribute to the pathogenesis and progression of ameloblastoma (AM), a benign but locally invasive odontogenic neoplasm. Here, we demonstrated that EMT- and stem cell-related genes were expressed in the epithelial islands of the most common histologic variant subtype, the follicular AM. Our results revealed elevated interleukin (IL)-6 signals that were differentially expressed in the stromal compartment of the follicular AM. To explore the stromal effect on tumor pathogenesis, we isolated and characterized both mesenchymal stromal cells (AM-MSCs) and epithelial cells (AM-EpiCs) from follicular AM and demonstrated that, in in vitro culture, AM-MSCs secreted a significantly higher level of IL-6 as compared to the counterpart AM-EpiCs. Furthermore, both in vitro and in vivo studies revealed that exogenous and AM-MSC-derived IL-6 induced the expression of EMT- and stem cell-related genes in AM-EpiCs, whereas such effects were significantly abrogated either by a specific inhibitor of STAT3 or ERK1/2, or by knockdown of Slug gene expression. These findings suggest that AM-MSC-derived IL-6 promotes tumor-stem like cell formation by inducing EMT process in AM-EpiCs through STAT3 and ERK1/2-mediated signaling pathways, implying a role in the etiology and progression of the benign but locally invasive neoplasm. Stem Cells 2017;35:2083-2094. © 2017 AlphaMed Press.

  20. Membrane lipidome of an epithelial cell line

    DEFF Research Database (Denmark)

    Sampaio, Julio L; Gerl, Mathias J; Klose, Christian

    2011-01-01

    Tissue differentiation is an important process that involves major cellular membrane remodeling. We used Madin-Darby canine kidney cells as a model for epithelium formation and investigated the remodeling of the total cell membrane lipidome during the transition from a nonpolarized morphology...... to an epithelial morphology and vice versa. To achieve this, we developed a shotgun-based lipidomics workflow that enabled the absolute quantification of mammalian membrane lipidomes with minimal sample processing from low sample amounts. Epithelial morphogenesis was accompanied by a major shift from sphingomyelin...... to generate an apical membrane domain that serves as a protective barrier for the epithelial sheet....

  1. PTEN Loss in E-Cadherin-Deficient Mouse Mammary Epithelial Cells Rescues Apoptosis and Results in Development of Classical Invasive Lobular Carcinoma

    NARCIS (Netherlands)

    Boelens, M.C.; Nethe, M.; Klarenbeek, S.; de Ruiter, J.R.; Schut, E.; Bonzanni, N.; Zeeman, A.L.; Wientjens, E.; van der Burg, E.; Wessels, L.; van Amerongen, R.; Jonkers, J.

    2016-01-01

    Invasive lobular carcinoma (ILC) is an aggressive breast cancer subtype with poor response to chemotherapy. Besides loss of E-cadherin, a hallmark of ILC, genetic inactivation of PTEN is frequently observed in patients. Through concomitant Cre-mediated inactivation of E-cadherin and PTEN in mammary

  2. Chemo-elastic modeling of invasive carcinoma development accompanied by oncogenic epithelial-mesenchymal transition

    Science.gov (United States)

    Bratsun, D. A.; Krasnyakov, I. V.; Pismen, L.

    2017-09-01

    We present a further development of a multiscale chemo-mechanical model of carcinoma growth in the epithelium tissue proposed earlier. The epithelium is represented by an elastic 2D array of polygonal cells, each with its own gene regulation dynamics. The model allows the simulation of evolution of multiple cells interacting via the chemical signaling or mechanically induced strain. The algorithm takes into account the division and intercalation of cells. The latter is most important since, first of all, carcinoma cells lose cell-cell adhesion and polarity via the oncogenic variant of the epithelial-mesenchymal transition (EMT) at which cells gain migratory and invasive properties. This process is mediated by E-cadherin repression and requires the differentiation of tumor cells with respect to the edge of the tumor that means that front cells should be most mobile. Taking into account this suggestion, we present the results of simulations demonstrating different patterns of carcinoma invasion. The comparison of our results with recent experimental observations is given and discussed.

  3. A human breast cell model of pre-invasive to invasive transition

    Energy Technology Data Exchange (ETDEWEB)

    Bissell, Mina J; Rizki, Aylin; Weaver, Valerie M.; Lee, Sun-Young; Rozenberg, Gabriela I.; Chin, Koei; Myers, Connie A.; Bascom, Jamie L.; Mott, Joni D.; Semeiks, Jeremy R.; Grate, Leslie R.; Mian, I. Saira; Borowsky, Alexander D.; Jensen, Roy A.; Idowu, Michael O.; Chen, Fanqing; Chen, David J.; Petersen, Ole W.; Gray, Joe W.; Bissell, Mina J.

    2008-03-10

    A crucial step in human breast cancer progression is the acquisition of invasiveness. There is a distinct lack of human cell culture models to study the transition from pre-invasive to invasive phenotype as it may occur 'spontaneously' in vivo. To delineate molecular alterations important for this transition, we isolated human breast epithelial cell lines that showed partial loss of tissue polarity in three-dimensional reconstituted-basement membrane cultures. These cells remained non-invasive; however, unlike their non-malignant counterparts, they exhibited a high propensity to acquire invasiveness through basement membrane in culture. The genomic aberrations and gene expression profiles of the cells in this model showed a high degree of similarity to primary breast tumor profiles. The xenograft tumors formed by the cell lines in three different microenvironments in nude mice displayed metaplastic phenotypes, including squamous and basal characteristics, with invasive cells exhibiting features of higher grade tumors. To find functionally significant changes in transition from pre-invasive to invasive phenotype, we performed attribute profile clustering analysis on the list of genes differentially expressed between pre-invasive and invasive cells. We found integral membrane proteins, transcription factors, kinases, transport molecules, and chemokines to be highly represented. In addition, expression of matrix metalloproteinases MMP-9,-13,-15,-17 was up regulated in the invasive cells. Using siRNA based approaches, we found these MMPs to be required for the invasive phenotype. This model provides a new tool for dissection of mechanisms by which pre-invasive breast cells could acquire invasiveness in a metaplastic context.

  4. Squamous cell carcinoma - invasive (image)

    Science.gov (United States)

    This irregular red nodule is an invasive squamous cell carcinoma (a form of skin cancer). Initial appearance, shown here, may be very similar to a noncancerous growth called a keratoacanthoma. Squamous cell cancers ...

  5. [Epithelial cadherins and associated molecules in invasive lobular breast cancer].

    Science.gov (United States)

    Brilliant, Yu M; Brilliant, A A; Sazonov, S V

    to estimate the expression of cell adhesion molecules E- and P-cadherin, as well as that of cadherin-catenin complexes in invasive lobular breast cancer (BC) cells. 250 cases of postoperative material from patients diagnosed with invasive lobular BC were studied. The expressions of cell adhesion molecules E-cadherin, P-cadherin, β-catenin, p120 catenin, and vimentin were determined by immunohistochemical assay in all cases. The examined cases were divided into molecular biological subtypes, based on the evaluation of estrogen receptors (ER), progesterone receptors (PR), HER-2/neu, and Ki-67 proliferative index. The membrane expression of E-cadherin on the tumor cells was found to be preserved in 93%; the cytoplasmic expression of β-catenin and p120-catenin appeared in 60 and 72% of cases, respectively. The expression of P-cadherin was detected in 82% of cases. The coexpression of E- and P-cadherin was noted in 90% of all the examined cases. There was a correlation between the expression of E- and P-cadherins (V=0.34; pcancer and its metastasis.

  6. Coronavirus infection of polarized epithelial cells

    NARCIS (Netherlands)

    Rossen, J W; Horzinek, M C; Rottier, P J

    1995-01-01

    Epithelial cells are the first host cells to be infected by incoming c oronaviruses. Recent observations in vitro show that coronaviruses are released from a specific side of these polarized cells, and this polarized release might be important for the spread of the infection in vivo. Mechanisms for

  7. Paracytosis of Haemophilus influenzae through cell layers of NCI-H292 lung epithelial cells

    NARCIS (Netherlands)

    van Schilfgaarde, M.; van Alphen, L.; Eijk, P.; Everts, V.; Dankert, J.

    1995-01-01

    Haemophilus influenzae penetrates the respiratory epithelium during carriage and invasive disease, including respiratory tract infections. We developed an in vitro model system consisting of lung epithelial NCI-H292 cells on permeable supports to study the passage of H. influenzae through lung

  8. Heterogeneity of limbal basal epithelial progenitor cells.

    Science.gov (United States)

    Hayashida, Yasutaka; Li, Wei; Chen, Ying-Ting; He, Hua; Chen, Szu-yu; Kheirkah, Ahmad; Zhu, Ying-Tien; Matsumoto, Yukihiro; Tseng, Scheffer C G

    2010-11-01

    Although corneal epithelial stem cells (SCs) are located at the limbus between the cornea and the conjunctiva, not all limbal basal epithelial cells are SCs. Using 2 dispase digestions to remove different amounts of limbal basal epithelial cells for cross-sections, flat mounts, and cytospin preparations, double immunostaining to pancytokeratins (PCK) and vimentin (Vim) identified 3 p63+ epithelial progenitors such as PCK-/Vim+, PCK/Vim, and PCK-/Vim+ and 1 p63+ mesenchymal cell, PCK-/Vim+. PCK-/Vim- progenitors had the smallest cell size were 10-20 times more enriched on collagen I-coated dishes in the 5-minute rapid adherent fraction that contained the highest percentage of p63+ cells but the lowest percentage of cytokeratin12+ cells, and gave rise to high Ki67 labeling and vivid clonal growth. In contrast, PCK+/Vim+ and PCK+/Vim- progenitors were found more in the slow-adherent fraction and yielded poor clonal growth. PCK/Vim progenitors and clusters of PCK-/Vim+ mesenchymal cells, which were neither melanocytes nor Langerhans cells, were located in the limbal basal region. Therefore, differential expression of PCK and Vim helps identify small PCK-/Vim- cells as the most likely candidate for SCs among a hierarchy of heterogeneous limbal basal progenitors, and their close association with PCK-/Vim+ presumed "niche" cells.

  9. Suppression of epithelial ovarian cancer invasion into the omentum by 1α,25-dihydroxyvitamin D3 and its receptor.

    Science.gov (United States)

    Lungchukiet, Panida; Sun, Yuefeng; Kasiappan, Ravi; Quarni, Waise; Nicosia, Santo V; Zhang, Xiaohong; Bai, Wenlong

    2015-04-01

    Epithelial ovarian cancer (EOC) is the leading cause of gynecological cancer death in women, mainly because it has spread to intraperitoneal tissues such as the omentum in the peritoneal cavity by the time of diagnosis. In the present study, we established in vitro assays, ex vivo omental organ culture system and syngeneic animal tumor models using wild type (WT) and vitamin D receptor (VDR) null mice to investigate the effects of 1α,25-dihydroxyvitamin D3 (1,25D3) and VDR on EOC invasion. Treatment of human EOC cells with 1,25D3 suppressed their migration and invasion in monolayer scratch and transwell assays and ability to colonize the omentum in the ex vivo system, supporting a role for epithelial VDR in interfering with EOC invasion. Furthermore, VDR knockdown in OVCAR3 cells increased their ability to colonize the omentum in the ex vivo system in the absence of 1,25D3, showing a potential ligand-independent suppression of EOC invasion by epithelial VDR. In syngeneic models, ID8 tumors exhibited an increased ability to colonize omenta of VDR null over that of WT mice; pre-treatment of WT, not VDR null, mice with EB1089 reduced ID8 colonization, revealing a role for stromal VDR in suppressing EOC invasion. These studies are the first to demonstrate a role for epithelial and stromal VDR in mediating the activity of 1,25D3 as well as a 1,25D3-independent action of the VDR in suppressing EOC invasion. The data suggest that VDR-based drug discovery may lead to the development of new intervention strategies to improve the survival of patients with EOC at advanced stages. This article is part of a Special Issue entitled "Vitamin D Workshop". Copyright © 2014 Elsevier Ltd. All rights reserved.

  10. TGFβ loss activates ADAMTS-1-mediated EGF-dependent invasion in a model of esophageal cell invasion

    Energy Technology Data Exchange (ETDEWEB)

    Le Bras, Grégoire F.; Taylor, Chase; Koumangoye, Rainelli B. [Department of Surgery, Vanderbilt University, Nashville, TN (United States); Revetta, Frank [Department of Pathology, Vanderbilt University, Nashville, TN (United States); Loomans, Holli A. [Department of Cancer Biology, Vanderbilt University, Nashville, TN (United States); Andl, Claudia D., E-mail: claudia.andl@vanderbilt.edu [Department of Surgery, Vanderbilt University, Nashville, TN (United States); Department of Cancer Biology, Vanderbilt University, Nashville, TN (United States); Department of Vanderbilt Ingram Cancer Center, Vanderbilt University, Nashville, TN (United States)

    2015-01-01

    The TGFβ signaling pathway is essential to epithelial homeostasis and is often inhibited during progression of esophageal squamous cell carcinoma. Recently, an important role for TGFβ signaling has been described in the crosstalk between epithelial and stromal cells regulating squamous tumor cell invasion in mouse models of head-and-neck squamous cell carcinoma (HNSCC). Loss of TGFβ signaling, in either compartment, leads to HNSCC however, the mechanisms involved are not well understood. Using organotypic reconstruct cultures (OTC) to model the interaction between epithelial and stromal cells that occur in dysplastic lesions, we show that loss of TGFβ signaling promotes an invasive phenotype in both fibroblast and epithelial compartments. Employing immortalized esophageal keratinocytes established to reproduce common mutations of esophageal squamous cell carcinoma, we show that treatment of OTC with inhibitors of TGFβ signaling (A83-01 or SB431542) enhances invasion of epithelial cells into a fibroblast-embedded Matrigel/collagen I matrix. Invasion induced by A83-01 is independent of proliferation but relies on protease activity and expression of ADAMTS-1 and can be altered by matrix density. This invasion was associated with increased expression of pro-inflammatory cytokines, IL1 and EGFR ligands HB-EGF and TGFα. Altering EGF signaling prevented or induced epithelial cell invasion in this model. Loss of expression of the TGFβ target gene ROBO1 suggested that chemorepulsion may regulate keratinocyte invasion. Taken together, our data show increased invasion through inhibition of TGFβ signaling altered epithelial-fibroblasts interactions, repressing markers of activated fibroblasts, and altering integrin-fibronectin interactions. These results suggest that inhibition of TGFβ signaling modulates an array of pathways that combined promote multiple aspects of tumor invasion. - Highlights: • Chemical inhibition of TGFβ signaling advances collective invasion

  11. Fungal invasion of normally non-phagocytic host cells.

    Directory of Open Access Journals (Sweden)

    Scott G Filler

    2006-12-01

    Full Text Available Many fungi that cause invasive disease invade host epithelial cells during mucosal and respiratory infection, and subsequently invade endothelial cells during hematogenous infection. Most fungi invade these normally non-phagocytic host cells by inducing their own uptake. Candida albicans hyphae interact with endothelial cells in vitro by binding to N-cadherin on the endothelial cell surface. This binding induces rearrangement of endothelial cell microfilaments, which results in the endocytosis of the organism. The capsule of Cryptococcus neoformans is composed of glucuronoxylomannan, which binds specifically to brain endothelial cells, and appears to mediate both adherence and induction of endocytosis. The mechanisms by which other fungal pathogens induce their own uptake are largely unknown. Some angioinvasive fungi, such as Aspergillus species and the Zygomycetes, invade endothelial cells from the abluminal surface during the initiation of invasive disease, and subsequently invade the luminal surface of endothelial cells during hematogenous dissemination. Invasion of normally non-phagocytic host cells has different consequences, depending on the type of invading fungus. Aspergillus fumigatus blocks apoptosis of pulmonary epithelial cells, whereas Paracoccidioides brasiliensis induces apoptosis of epithelial cells. This review summarizes the mechanisms by which diverse fungal pathogens invade normally non-phagocytic host cells and discusses gaps in our knowledge that provide opportunities for future research.

  12. Replication of cultured lung epithelial cells

    International Nuclear Information System (INIS)

    Guzowski, D.; Bienkowski, R.

    1986-01-01

    The authors have investigated the conditions necessary to support replication of lung type 2 epithelial cells in culture. Cells were isolated from mature fetal rabbit lungs (29d gestation) and cultured on feeder layers of mitotically inactivated 3T3 fibroblasts. The epithelial nature of the cells was demonstrated by indirect immunofluorescent staining for keratin and by polyacid dichrome stain. Ultrastructural examination during the first week showed that the cells contained myofilaments, microvilli and lamellar bodies (markers for type 2 cells). The following changes were observed after the first week: increase in cell size; loss of lamellar bodies and appearance of multivesicular bodies; increase in rough endoplasmic reticulum and golgi; increase in tonafilaments and well-defined junctions. General cell morphology was good for up to 10 wk. Cells cultured on plastic surface degenerated after 1 wk. Cell replication was assayed by autoradiography of cultures exposed to ( 3 H)-thymidine and by direct cell counts. The cells did not replicate during the first week; however, between 2-10 wk the cells incorporated the label and went through approximately 6 population doublings. They have demonstrated that lung alveolar epithelial cells can replicate in culture if they are maintained on an appropriate substrate. The coincidence of ability to replicate and loss of markers for differentiation may reflect the dichotomy between growth and differentiation commonly observed in developing systems

  13. Interaction with Epithelial Cells Modifies Airway Macrophage Response to Ozone

    Science.gov (United States)

    The initial innate immune response to ozone (03) in the lung is orchestrated by structural cells, such as epithelial cells, and resident immune cells, such as airway macrophages (Macs). We developed an epithelial cell-Mac coculture model to investigate how epithelial cell-derived...

  14. Epithelial cell-cell junctions and plasma membrane domains

    NARCIS (Netherlands)

    Giepmans, Ben N. G.; van Ijzendoorn, Sven C. D.

    Epithelial cells form a barrier against the environment, but are also required for the regulated exchange of molecules between an organism and its surroundings. Epithelial cells are characterised by a remarkable polarization of their plasma membrane, evidenced by the appearance of structurally,

  15. Non-Saccharomyces yeasts protect against epithelial cell barrier disruption induced by Salmonella enterica subsp. enterica serovar Typhimurium.

    Science.gov (United States)

    Smith, I M; Baker, A; Arneborg, N; Jespersen, L

    2015-11-01

    The human gastrointestinal epithelium makes up the largest barrier separating the body from the external environment. Whereas invasive pathogens cause epithelial barrier disruption, probiotic micro-organisms modulate tight junction regulation and improve epithelial barrier function. In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability of four nonpathogenic yeast species to modulate transepithelial electrical resistance (TER) across a monolayer of differentiated human colonocytes (Caco-2 cells). Further, we assessed yeast modulation of a Salmonella Typhimurium-induced epithelial cell barrier function insult. Our findings demonstrate distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study demonstrates distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Further, our data demonstrate significant yeast-mediated modulation of Salmonella Typhimurium-induced epithelial cell barrier disruption and identify Kluyveromyces marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study is the first to demonstrate significant non-Saccharomyces yeast

  16. Altered expression of epithelial cell surface glycoconjugates and intermediate filaments at the margins of mucosal wounds

    DEFF Research Database (Denmark)

    Dabelsteen, Erik; Grøn, B.; Mandel, U.

    1998-01-01

    Alterations in cell to cell adhesion are necessary to enable the type of cell movements that are associated with epithelial wound healing and malignant invasion. Several studies of transformed cells have related epithelial cell movement to changes in the cell surface expression of the carbohydrate......-T antigen. The changes induced by wounding in the expression of collagen IV, laminin gamma2-chain (laminin-5), and laminin alpha5-chain were similar to those found in skin wounds and served to define the region of epithelial movement. This region was found to show a marked increase in staining for both...... epithelium, a pattern of expression similar to K16, which was also strongly upregulated in both the outgrowth and the adjacent nonwounded epithelium. These findings provide further support for an influence of such carbohydrate structures on the migratory behavior of epithelial cells....

  17. Implantation of Induced Pluripotent Stem Cell-Derived Tracheal Epithelial Cells.

    Science.gov (United States)

    Ikeda, Masakazu; Imaizumi, Mitsuyoshi; Yoshie, Susumu; Nakamura, Ryosuke; Otsuki, Koshi; Murono, Shigeyuki; Omori, Koichi

    2017-07-01

    Compared with using autologous tissue, the use of artificial materials in the regeneration of tracheal defects is minimally invasive. However, this technique requires early epithelialization on the inner side of the artificial trachea. After differentiation from induced pluripotent stem cells (iPSCs), tracheal epithelial tissues may be used to produce artificial tracheas. Herein, we aimed to demonstrate that after differentiation from fluorescent protein-labeled iPSCs, tracheal epithelial tissues survived in nude rats with tracheal defects. Red fluorescent tdTomato protein was electroporated into mouse iPSCs to produce tdTomato-labeled iPSCs. Embryoid bodies derived from these iPSCs were then cultured in differentiation medium supplemented with growth factors, followed by culture on air-liquid interfaces for further differentiation into tracheal epithelium. The cells were implanted with artificial tracheas into nude rats with tracheal defects on day 26 of cultivation. On day 7 after implantation, the tracheas were exposed and examined histologically. Tracheal epithelial tissue derived from tdTomato-labeled iPSCs survived in the tracheal defects. Moreover, immunochemical analyses showed that differentiated tissues had epithelial structures similar to those of proximal tracheal tissues. After differentiation from iPSCs, tracheal epithelial tissues survived in rat bodies, warranting the use of iPSCs for epithelial regeneration in tracheal defects.

  18. The human airway epithelial basal cell transcriptome.

    Directory of Open Access Journals (Sweden)

    Neil R Hackett

    2011-05-01

    Full Text Available The human airway epithelium consists of 4 major cell types: ciliated, secretory, columnar and basal cells. During natural turnover and in response to injury, the airway basal cells function as stem/progenitor cells for the other airway cell types. The objective of this study is to better understand human airway epithelial basal cell biology by defining the gene expression signature of this cell population.Bronchial brushing was used to obtain airway epithelium from healthy nonsmokers. Microarrays were used to assess the transcriptome of basal cells purified from the airway epithelium in comparison to the transcriptome of the differentiated airway epithelium. This analysis identified the "human airway basal cell signature" as 1,161 unique genes with >5-fold higher expression level in basal cells compared to differentiated epithelium. The basal cell signature was suppressed when the basal cells differentiated into a ciliated airway epithelium in vitro. The basal cell signature displayed overlap with genes expressed in basal-like cells from other human tissues and with that of murine airway basal cells. Consistent with self-modulation as well as signaling to other airway cell types, the human airway basal cell signature was characterized by genes encoding extracellular matrix components, growth factors and growth factor receptors, including genes related to the EGF and VEGF pathways. Interestingly, while the basal cell signature overlaps that of basal-like cells of other organs, the human airway basal cell signature has features not previously associated with this cell type, including a unique pattern of genes encoding extracellular matrix components, G protein-coupled receptors, neuroactive ligands and receptors, and ion channels.The human airway epithelial basal cell signature identified in the present study provides novel insights into the molecular phenotype and biology of the stem/progenitor cells of the human airway epithelium.

  19. Collective cell migration: Implications for wound healing and cancer invasion

    Directory of Open Access Journals (Sweden)

    Li Li

    2013-07-01

    Full Text Available During embryonic morphogenesis, wound repair and cancer invasion, cells often migrate collectively via tight cell-cell junctions, a process named collective migration. During such migration, cells move as coherent groups, large cell sheets, strands or tubes rather than individually. One unexpected finding regarding collective cell migration is that being a "multicellular structure" enables cells to better respond to chemical and physical cues, when compared with isolated cells. This is important because epithelial cells heal wounds via the migration of large sheets of cells with tight intercellular connections. Recent studies have gained some mechanistic insights that will benefit the clinical understanding of wound healing in general. In this review, we will briefly introduce the role of collective cell migration in wound healing, regeneration and cancer invasion and discuss its underlying mechanisms as well as implications for wound healing.

  20. Ezrin mediates c-Myc actions in prostate cancer cell invasion

    DEFF Research Database (Denmark)

    Chuan, Yin Choy; Iglesias Gato, Diego; Fernandez-Perez, L

    2010-01-01

    The forced overexpression of c-Myc in mouse prostate and in normal human prostate epithelial cells results in tumor transformation with an invasive phenotype. How c-Myc regulates cell invasion is poorly understood. In this study, we have investigated the interplay of c-Myc and androgens in the re...

  1. Reexcision Perineural Invasion and Epithelial Sheath Neuroma Possibly on a Spectrum of Postinjury Reactive Hyperplasia Mediated by IL-6.

    Science.gov (United States)

    Wang, James Y; Nuovo, Gerard; Kline, Mitchell; Magro, Cynthia M

    2017-01-01

    Epithelial sheath neuroma is a rarely recognized but established entity in the medical literature. First described in 2000 by Requena et al, there have only been 7 published cases to date, mostly in female patients and presenting as symptomatic solitary lesions on the back without a known history of trauma. In 2006, Beer et al described and reviewed a dozen cases in which epithelial sheath neuroma-like features were seen in the advent of a surgical procedure, which was termed "re-excision perineural invasion" and attributed to possible eccrine duct implantation during surgery. Our case is a 66-year-old male patient who underwent an excision of a melanocytic neoplasm in which a reactive epithelial sheath neuroma was incidentally discovered in the excision specimen, adjacent to the biopsy site cicatrix. Histologically, there was benign cutaneous nerve hyperplasia with a proliferation of squamous epithelium in intimate apposition to the nerve bundles in the superficial dermis. We postulate that the process active in the formation of re-excision perineural invasion is the same as in epithelial sheath neuroma and that minor trauma not appreciable on histologic examination is responsible in the latter entity. We performed IL-6 staining and documented that IL-6 was upregulated at the interface of the nerve and reactive epithelium, but was absent in nerves distant from the site of surgery, suggesting that IL-6 may be essential to the lesion's development. The recognition of reactive epithelial sheath neuroma including the subcategory of re-excision perineural invasion is crucial for the dermatopathologist to prevent mislabeling this reactive entity as a perineural squamous cell carcinoma, which has clinical consequences for the patient such as wider re-excision and radiation treatment. Additionally, we have identified a potential pathophysiologic basis for this lesion.

  2. Clinicopathological study of rare invasive epithelial tumors of breast: An institutional study

    Directory of Open Access Journals (Sweden)

    Karthik Kasireddy

    2016-01-01

    Full Text Available Introduction: Invasive breast cancer (BC is the most common carcinoma in women. It accounts for 22% of all female cancers. Most tumors are derived from mammary duct epithelium, and up to 75% of BCs are ductal carcinomas. The second most common tumor is invasive lobular carcinoma. However, there are many variants which are less common but well defined by the World Health Organization classification. They comprise <10% of breast tumors. Their clinical behavior differs greatly. Hence, it is important to know their main histomorphological features to make the best treatment of choice and to foresee prognosis. Aims and Objectives: To study the histomorphological features, incidence, and clinical features of rare invasive epithelial tumors of the breast. Materials and Methods: This study was done in the department of pathology, Sri Devaraj Urs Medical College, Kolar. All the neoplastic breast lesions over a period of 5 years (July 2010-September 2015 are included in the study. Clinical features and other details (estrogen receptor/progesterone receptor, human epidermal receptor-2, lymph nodes are obtained from the department (surgery records. Specimens are received and preserved in 10% formalin and are subjected to routine histopathological processing. Hematoxylin and eosin sections are studied, and a morphological diagnosis is given. All rare invasive epithelial breast tumors will be reviewed meticulously. Results and Conclusion: A total number of invasive epithelial tumors of breast were 105. The most common presenting symptom was breast lump. Rare invasive epithelial breast tumors account to 28.5%. The age range from 15 to 70 years. Most common, rare invasive epithelial tumor in our study is medullary carcinoma. Hence, it is imperative to always maintain a Hawks vigil during microscopic diagnosis to know prognosis of the condition and to facilitate early and prompt treatment to the patient.

  3. Human Mammary Luminal Epithelial Cells Contain Progenitors to Myoepithelial Cells

    Energy Technology Data Exchange (ETDEWEB)

    Pechoux, Christine; Gudjonsson, Thorarinn; Ronnov-Jessen, Lone; Bissell, Mina J; Petersen, Ole

    1999-02-01

    The origin of the epithelial and myoepithelial cells in the human breast has not been delineated. In this study we have addressed whether luminal epithelial cells and myoepithelial cells are vertically connected, i.e., whether one is the precursor for the other. We used a primary culture assay allowing preservation of basic phenotypic traits of luminal epithelial and myoepithelial cells in culture. The two cell types were then separated immunomagnetically using antibodies directed against lineage-specific cell surface antigens into at best 100% purity. The cellular identity was ascertained by cytochemistry, immunoblotting, and 2-D gel electrophoresis. Luminal epithelial cells were identified by strong expression of cytokeratins 18 and 19 while myoepithelial cells were recognized by expression of vimentin and {alpha}-smooth muscle actin. We used a previously devised culture medium (CDM4) that allows vigorous expansion of proliferative myoepithelial cells and also devised a medium (CDM6) that allowed sufficient expansion of differentiated luminal epithelial cells based on addition of hepatocyte growth factor/scatter factor. The two different culture media supported each lineage for at least five passages without signs of interconversion. We used parallel cultures where we switched culture media, thus testing the ability of each lineage to convert to the other. Whereas the myoepithelial lineage showed no signs of interconversion, a subset of luminal epithelial cells, gradually, but distinctly, converted to myoepithelial cells. We propose that in the mature human breast, it is the luminal epithelial cell compartment that gives rise to myoepithelial cells rather than the other way around.

  4. Development of Thymic Epithelial Cells

    DEFF Research Database (Denmark)

    Ulyanchenko, Svetlana; Vaidya, Harsh J.; O'Neill, Kathy E.

    2016-01-01

    The thymus is the primary lymphoid organ in which the T cell repertoire is generated. The complex cellularity of this organ is uniquely designed to facilitate T cell development: defects in thymus development or function can cause immunodeficiencies ranging from the absence of T cell-mediated imm......The thymus is the primary lymphoid organ in which the T cell repertoire is generated. The complex cellularity of this organ is uniquely designed to facilitate T cell development: defects in thymus development or function can cause immunodeficiencies ranging from the absence of T cell......-mediated immunity to broad-spectrum autoimmune disease. Peak thymus size and output occurs early in life, after which the thymus undergoes a natural process of involution. This results in the progressive loss of functional thymus tissue and correspondingly in decreased production of new naïve T cells with age...... - contributing to the diminished capacity of the aged immune system to adequately respond to new antigenic challenge. Age-related thymic involutions, together with the thymic involutions associated with cytotoxic therapies (e.g., radio- or chemotherapy), have raised interest in development of clinically useful...

  5. Traction forces exerted by epithelial cell sheets

    International Nuclear Information System (INIS)

    Saez, A; Anon, E; Ghibaudo, M; Di Meglio, J-M; Hersen, P; Ladoux, B; Du Roure, O; Silberzan, P; Buguin, A

    2010-01-01

    Whereas the adhesion and migration of individual cells have been well described in terms of physical forces, the mechanics of multicellular assemblies is still poorly understood. Here, we study the behavior of epithelial cells cultured on microfabricated substrates designed to measure cell-to-substrate interactions. These substrates are covered by a dense array of flexible micropillars whose deflection enables us to measure traction forces. They are obtained by lithography and soft replica molding. The pillar deflection is measured by video microscopy and images are analyzed with home-made multiple particle tracking software. First, we have characterized the temporal and spatial distributions of traction forces of cellular assemblies of various sizes. The mechanical force balance within epithelial cell sheets shows that the forces exerted by neighboring cells strongly depend on their relative position in the monolayer: the largest deformations are always localized at the edge of the islands of cells in the active areas of cell protrusions. The average traction stress rapidly decreases from its maximum value at the edge but remains much larger than the inherent noise due to the force resolution of our pillar tracking software, indicating an important mechanical activity inside epithelial cell islands. Moreover, these traction forces vary linearly with the rigidity of the substrate over about two decades, suggesting that cells exert a given amount of deformation rather than a force. Finally, we engineer micropatterned substrates supporting pillars with anisotropic stiffness. On such substrates cellular growth is aligned with respect to the stiffest direction in correlation with the magnitude of the applied traction forces.

  6. Epithelial Cell Adherence Mediated by the Enterotoxigenic Escherichia coli Tia Protein

    OpenAIRE

    Mammarappallil, Joseph G.; Elsinghorst, Eric A.

    2000-01-01

    In vitro studies have shown that enterotoxigenic Escherichia coli (ETEC) strains are capable of invading cultured epithelial cells derived from the human ileum and colon. Two separate invasion loci (tia and tib) have previously been isolated from the classical ETEC strain H10407. The tia locus has been shown to direct the synthesis of Tia, a 25-kDa outer membrane protein. Tia is sufficient to confer the adherence and invasion phenotypes on laboratory stains of E. coli, suggesting that this pr...

  7. Transcriptional landscape of glomerular parietal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Sina A Gharib

    Full Text Available Very little is known about the function of glomerular parietal epithelial cells (PECs. In this study, we performed genome-wide expression analysis on PEC-enriched capsulated vs. PEC-deprived decapsulated rat glomeruli to determine the transcriptional state of PECs under normal conditions. We identified hundreds of differentially expressed genes that mapped to distinct biologic modules including development, tight junction, ion transport, and metabolic processes. Since developmental programs were highly enriched in PECs, we characterized several of their candidate members at the protein level. Collectively, our findings confirm that PECs are multifaceted cells and help define their diverse functional repertoire.

  8. Lactobacillus Decelerates Cervical Epithelial Cell Cycle Progression

    Science.gov (United States)

    Vielfort, Katarina; Weyler, Linda; Söderholm, Niklas; Engelbrecht, Mattias; Löfmark, Sonja; Aro, Helena

    2013-01-01

    We investigated cell cycle progression in epithelial cervical ME-180 cells during colonization of three different Lactobacillus species utilizing live cell microscopy, bromodeoxyuridine incorporation assays, and flow cytometry. The colonization of these ME-180 cells by L. rhamnosus and L. reuteri, originating from human gastric epithelia and saliva, respectively, was shown to reduce cell cycle progression and to cause host cells to accumulate in the G1 phase of the cell cycle. The G1 phase accumulation in L. rhamnosus-colonized cells was accompanied by the up-regulation and nuclear accumulation of p21. By contrast, the vaginal isolate L. crispatus did not affect cell cycle progression. Furthermore, both the supernatants from the lactic acid-producing L. rhamnosus colonies and lactic acid added to cell culture media were able to reduce the proliferation of ME-180 cells. In this study, we reveal the diversity of the Lactobacillus species to affect host cell cycle progression and demonstrate that L. rhamnosus and L. reuteri exert anti-proliferative effects on human cervical carcinoma cells. PMID:23675492

  9. Lactobacillus decelerates cervical epithelial cell cycle progression.

    Directory of Open Access Journals (Sweden)

    Katarina Vielfort

    Full Text Available We investigated cell cycle progression in epithelial cervical ME-180 cells during colonization of three different Lactobacillus species utilizing live cell microscopy, bromodeoxyuridine incorporation assays, and flow cytometry. The colonization of these ME-180 cells by L. rhamnosus and L. reuteri, originating from human gastric epithelia and saliva, respectively, was shown to reduce cell cycle progression and to cause host cells to accumulate in the G1 phase of the cell cycle. The G1 phase accumulation in L. rhamnosus-colonized cells was accompanied by the up-regulation and nuclear accumulation of p21. By contrast, the vaginal isolate L. crispatus did not affect cell cycle progression. Furthermore, both the supernatants from the lactic acid-producing L. rhamnosus colonies and lactic acid added to cell culture media were able to reduce the proliferation of ME-180 cells. In this study, we reveal the diversity of the Lactobacillus species to affect host cell cycle progression and demonstrate that L. rhamnosus and L. reuteri exert anti-proliferative effects on human cervical carcinoma cells.

  10. Endothelial induced EMT in breast epithelial cells with stem cell properties.

    Directory of Open Access Journals (Sweden)

    Valgardur Sigurdsson

    Full Text Available Epithelial to mesenchymal transition (EMT is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad to N-Cadherin (N-Cad and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44(high/CD24(low ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer.

  11. Endothelial induced EMT in breast epithelial cells with stem cell properties.

    Science.gov (United States)

    Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J R; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A; Petersen, Ole William; Magnusson, Magnus K; Gudjonsson, Thorarinn

    2011-01-01

    Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M) derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad) to N-Cadherin (N-Cad) and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44(high)/CD24(low) ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer.

  12. Chromosome aberration induction in human diploid fibroblast and epithelial cells

    International Nuclear Information System (INIS)

    Scott, D.

    1986-01-01

    The relative sensitivity of cultured human fibroblasts and epithelial cells to radiation-induced chromosomal aberrations was investigated. Lung fibroblast and kidney epithelial cells from the same fetus were compared, as were skin fibroblasts and epithelial keratinocytes from the same foreskin sample. After exposure of proliferating fetal cells to 1.5 Gy X-rays there was a very similar aberration yield in the fibroblasts and epithelial cells. Observations of either little or no difference in chromosomal sensitivity between human fibroblasts and epithelial cells give added confidence that quantitative cytogenetic data obtained from cultured fibroblasts are relevant to the question of sensitivity of epithelial cells which are the predominant cell type in human cancers. (author)

  13. The diagnosis and management of pre-invasive breast disease: Flat epithelial atypia – classification, pathologic features and clinical significance

    International Nuclear Information System (INIS)

    Schnitt, Stuart J

    2003-01-01

    Flat epithelial atypia is a descriptive term that encompasses lesions of the breast terminal duct lobular units in which variably dilated acini are lined by one to several layers of epithelial cells, which are usually columnar in shape and which display low-grade cytologic atypia. Observational studies have suggested that at least some of these lesions may represent either a precursor of ductal carcinoma in situ (DCIS) or the earliest morphological manifestation of DCIS. In contrast, the limited available clinical follow-up data suggest that the risk of both local recurrence and progression of these lesions to invasive cancer is extremely low, supporting the notion that categorizing such lesions as 'clinging carcinoma' and managing them as if they were fully developed DCIS will result in overtreatment of many patients. Additional studies are needed to better understand the biological nature and clinical significance of these lesions

  14. Breast fibroblasts modulate epithelial cell proliferation in three-dimensional in vitro co-culture

    International Nuclear Information System (INIS)

    Sadlonova, Andrea; Novak, Zdenek; Johnson, Martin R; Bowe, Damon B; Gault, Sandra R; Page, Grier P; Thottassery, Jaideep V; Welch, Danny R; Frost, Andra R

    2005-01-01

    Stromal fibroblasts associated with in situ and invasive breast carcinoma differ phenotypically from fibroblasts associated with normal breast epithelium, and these alterations in carcinoma-associated fibroblasts (CAF) may promote breast carcinogenesis and cancer progression. A better understanding of the changes that occur in fibroblasts during carcinogenesis and their influence on epithelial cell growth and behavior could lead to novel strategies for the prevention and treatment of breast cancer. To this end, the effect of CAF and normal breast-associated fibroblasts (NAF) on the growth of epithelial cells representative of pre-neoplastic breast disease was assessed. NAF and CAF were grown with the nontumorigenic MCF10A epithelial cells and their more transformed, tumorigenic derivative, MCF10AT cells, in direct three-dimensional co-cultures on basement membrane material. The proliferation and apoptosis of MCF10A cells and MCF10AT cells were assessed by 5-bromo-2'-deoxyuridine labeling and TUNEL assay, respectively. Additionally, NAF and CAF were compared for expression of insulin-like growth factor II as a potential mediator of their effects on epithelial cell growth, by ELISA and by quantitative, real-time PCR. In relatively low numbers, both NAF and CAF suppressed proliferation of MCF10A cells. However, only NAF and not CAF significantly inhibited proliferation of the more transformed MCF10AT cells. The degree of growth inhibition varied among NAF or CAF from different individuals. In greater numbers, NAF and CAF have less inhibitory effect on epithelial cell growth. The rate of epithelial cell apoptosis was not affected by NAF or CAF. Mean insulin-like growth factor II levels were not significantly different in NAF versus CAF and did not correlate with the fibroblast effect on epithelial cell proliferation. Both NAF and CAF have the ability to inhibit the growth of pre-cancerous breast epithelial cells. NAF have greater inhibitory capacity than CAF

  15. MiR-203 controls proliferation, migration and invasive potential of prostate cancer cell lines

    DEFF Research Database (Denmark)

    Viticchiè, Giuditta; Lena, Anna Maria; Latina, Alessia

    2011-01-01

    Prostate cancers show a slow progression from a local lesion (primary tumor) to a metastatic and hormone-resistant phenotype. After an initial step of hyperplasia, in a high percentage of cases a neoplastic transformation event occurs that, less frequently, is followed by epithelial to mesenchymal...... cell lines compared to normal epithelial prostatic cells. Overexpression of miR-203 in brain or bone metastatic prostate cell lines (DU145 and PC3) is sufficient to induce a mesenchymal to epithelial transition with inhibition of cell proliferation, migration and invasiveness. We have identified CKAP2...

  16. Acinetobacter baumannii and A. pittii clinical isolates lack adherence and cytotoxicity to lung epithelial cells in vitro.

    Science.gov (United States)

    Lázaro-Díez, María; Navascués-Lejarza, Teresa; Remuzgo-Martínez, Sara; Navas, Jesús; Icardo, José Manuel; Acosta, Felix; Martínez-Martínez, Luis; Ramos-Vivas, José

    2016-09-01

    The molecular and genetic basis of Acinetobacter baumannii and Acinetobacter pittii virulence remains poorly understood, and there is still lack of knowledge in host cell response to these bacteria. In this study, we have used eleven clinical Acinetobacter strains (A. baumannii n = 5; A. pittii n = 6) to unravel bacterial adherence, invasion and cytotoxicity to human lung epithelial cells. Our results showed that adherence to epithelial cells by Acinetobacter strains is scarce and cellular invasion was not truly detected. In addition, all Acinetobacter strains failed to induce any cytotoxic effect on A549 cells. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  17. Expression of a fms-related oncogene in carcinogen-induced neoplastic epithelial cells

    International Nuclear Information System (INIS)

    Walker, C.; Nettesheim, P.; Barrett, J.C.; Gilmer, T.M.

    1987-01-01

    Following carcinogen exposure in vitro, normal rat tracheal epithelial cells are transformed in a multistage process in which the cultured cells become immortal and ultimately, neoplastic. Five cell lines derived from tumors produced by neoplastically transformed rat tracheal epithelial cells were examined for the expression of 11 cellular oncogenes previously implicated in pulmonary or epithelial carcinogenesis. RNA homologous to fms was expressed at a level 5-19 times higher than normal tracheal epithelial cells in three of five of the tumor-derived lines. All three lines expressing high levels of fms-related RNA gave rise to invasive tumors of epithelial origin when injected into nude mice. Increased expression of the fms-related mRNA was not due to gene amplification, and no gene rearrangement was detected by Southern analyses. RNA blot analysis using a 3' v-fms probe detected a 9.5-kilobase message in the three tumor-derived lines, whereas both normal rat aveolar macrophages and the human choriocarcinoma line BeWo expressed a fms transcript of ≅ 4 kilobases. The authors conclude from these data that the gene expressed as a 9.5-kilobase transcript in these neoplastic epithelial cells is a member of a fms-related gene family but may be distinct from the gene that encodes the macrophage colony-stimulating factor (CSF-1) receptor

  18. Establishment and evaluation of a stable cattle type II alveolar epithelial cell line.

    Directory of Open Access Journals (Sweden)

    Feng Su

    Full Text Available Macrophages and dendritic cells are recognized as key players in the defense against mycobacterial infection. Recent research has confirmed that alveolar epithelial cells (AECs also play important roles against mycobacterium infections. Thus, establishing a stable cattle AEC line for future endogenous immune research on bacterial invasion is necessary. In the present study, we first purified and immortalized type II AECs (AEC II cells by transfecting them with a plasmid containing the human telomerase reverse trancriptase gene. We then tested whether or not the immortalized cells retained the basic physiological properties of primary AECs by reverse-transcription polymerase chain reaction and Western blot. Finally, we tested the secretion capacity of immortalized AEC II cells upon stimulation by bacterial invasion. The cattle type II alveolar epithelial cell line (HTERT-AEC II that we established retained lung epithelial cell characteristics: the cells were positive for surfactants A and B, and they secreted tumor necrosis factor-α and interleukin-6 in response to bacterial invasion. Thus, the cell line we established is a potential tool for research on the relationship between AECs and Mycobacterium tuberculosis.

  19. Nuclear microscopy of rat colon epithelial cells

    Science.gov (United States)

    Ren, M.; Rajendran, Reshmi; Ng, Mary; Udalagama, Chammika; Rodrigues, Anna E.; Watt, Frank; Jenner, Andrew Michael

    2011-10-01

    Using Nuclear microscopy, we have investigated iron distributions in the colons of Sprague Dawley rats, in order to elucidate heme uptake. Four groups of five Sprague Dawley rats (mean weight 180 g) were fed different purified diets containing either heme diet (2.5% w/w hemoglobin), high fat diet (HFD) (18% w/w fat, 1% w/w cholesterol), 'western' diet (combination of hemoglobin 2.5% and 18% fat, 1% cholesterol) or control diet (7% w/w fat). After 4 weeks, animals were sacrificed by exsanguination after anaesthesia. Thin sections of frozen colon tissue were taken, freeze dried and scanned using nuclear microscopy utilising the techniques PIXE, RBS and STIM. The new data acquisition system (IonDaq) developed in CIBA was used to obtain high resolution images and line scans were used to map the iron distributions across the colon boundaries. The nuclear microscope results indicate that when HFD is given in addition to heme, the iron content of the epithelial cells that line the colon decreases, and the zinc in the smooth muscle wall increases. This implies that the level of heme and fat in diet has an important role in colon health, possibly by influencing epithelial cells directly or changing luminal composition such as bacterial flora or levels of metabolites and cytotoxins.

  20. Nuclear microscopy of rat colon epithelial cells

    International Nuclear Information System (INIS)

    Ren, M.; Rajendran, Reshmi; Ng, Mary; Udalagama, Chammika; Rodrigues, Anna E.; Watt, Frank; Jenner, Andrew Michael

    2011-01-01

    Using Nuclear microscopy, we have investigated iron distributions in the colons of Sprague Dawley rats, in order to elucidate heme uptake. Four groups of five Sprague Dawley rats (mean weight 180 g) were fed different purified diets containing either heme diet (2.5% w/w hemoglobin), high fat diet (HFD) (18% w/w fat, 1% w/w cholesterol), 'western' diet (combination of hemoglobin 2.5% and 18% fat, 1% cholesterol) or control diet (7% w/w fat). After 4 weeks, animals were sacrificed by exsanguination after anaesthesia. Thin sections of frozen colon tissue were taken, freeze dried and scanned using nuclear microscopy utilising the techniques PIXE, RBS and STIM. The new data acquisition system (IonDaq) developed in CIBA was used to obtain high resolution images and line scans were used to map the iron distributions across the colon boundaries. The nuclear microscope results indicate that when HFD is given in addition to heme, the iron content of the epithelial cells that line the colon decreases, and the zinc in the smooth muscle wall increases. This implies that the level of heme and fat in diet has an important role in colon health, possibly by influencing epithelial cells directly or changing luminal composition such as bacterial flora or levels of metabolites and cytotoxins.

  1. Nuclear microscopy of rat colon epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Ren, M., E-mail: phyrenmq@nus.edu.sg [Centre for Ion Beam Applications (CIBA), Department of Physics, National University of Singapore, Singapore 117542 (Singapore); Rajendran, Reshmi [Lab of Molecular Imaging, Singapore Bioimaging Consotium, 11 Biopolis Way, 02-02 Helios, Singapore 138667 (Singapore); Ng, Mary [Department of Pharmacology, National University of Singapore (Singapore); Udalagama, Chammika; Rodrigues, Anna E.; Watt, Frank [Centre for Ion Beam Applications (CIBA), Department of Physics, National University of Singapore, Singapore 117542 (Singapore); Jenner, Andrew Michael [Illawara Health and Medical Research Institute (IHMRI), University of Wollongong, NSW 2522 (Australia)

    2011-10-15

    Using Nuclear microscopy, we have investigated iron distributions in the colons of Sprague Dawley rats, in order to elucidate heme uptake. Four groups of five Sprague Dawley rats (mean weight 180 g) were fed different purified diets containing either heme diet (2.5% w/w hemoglobin), high fat diet (HFD) (18% w/w fat, 1% w/w cholesterol), 'western' diet (combination of hemoglobin 2.5% and 18% fat, 1% cholesterol) or control diet (7% w/w fat). After 4 weeks, animals were sacrificed by exsanguination after anaesthesia. Thin sections of frozen colon tissue were taken, freeze dried and scanned using nuclear microscopy utilising the techniques PIXE, RBS and STIM. The new data acquisition system (IonDaq) developed in CIBA was used to obtain high resolution images and line scans were used to map the iron distributions across the colon boundaries. The nuclear microscope results indicate that when HFD is given in addition to heme, the iron content of the epithelial cells that line the colon decreases, and the zinc in the smooth muscle wall increases. This implies that the level of heme and fat in diet has an important role in colon health, possibly by influencing epithelial cells directly or changing luminal composition such as bacterial flora or levels of metabolites and cytotoxins.

  2. Cellular Plasticity of Epithelial Cells-Cause of Metastasis

    National Research Council Canada - National Science Library

    Sukumar, Saraswati

    2005-01-01

    .... We present a novel concept that cancer epithelial cells, possibly of stem cell origin, have inherent cellular plasticity and can differentiate into endothelial cells and form microvessels that serve...

  3. Heterologously expressed Staphylococcus aureus fibronectin-binding proteins are sufficient for invasion of host cells

    NARCIS (Netherlands)

    Sinha, B; Francois, P; Que, Y A; Hussain, M; Heilmann, C; Moreillon, P; Lew, D; Krause, K H; Peters, Georg; Herrmann, M

    2000-01-01

    Staphylococcus aureus invasion of mammalian cells, including epithelial, endothelial, and fibroblastic cells, critically depends on fibronectin bridging between S. aureus fibronectin-binding proteins (FnBPs) and the host fibronectin receptor integrin alpha(5)beta(1) (B. Sinha et al., Cell.

  4. Susceptibility of Primary Human Choroid Plexus Epithelial Cells and Meningeal Cells to Infection by JC Virus.

    Science.gov (United States)

    O'Hara, Bethany A; Gee, Gretchen V; Atwood, Walter J; Haley, Sheila A

    2018-04-15

    -cerebrospinal fluid barrier, the choroid plexus and leptomeninges are also poised to play roles in virus invasion of brain parenchyma, where infection of macroglial cells leads to the development of progressive multifocal leukoencephalopathy, a severely debilitating and often fatal infection. In this paper we show for the first time that primary choroid plexus epithelial cells and meningeal cells are infected by JCPyV, lending support to the association of JCPyV with meningoencephalopathies. These data also suggest that JCPyV could use these cells as reservoirs for the subsequent invasion of brain parenchyma. Copyright © 2018 American Society for Microbiology.

  5. Silk Film Topography Directs Collective Epithelial Cell Migration

    Science.gov (United States)

    Rosenblatt, Mark I.

    2012-01-01

    The following study provides new insight into how surface topography dictates directed collective epithelial cell sheet growth through the guidance of individual cell movement. Collective cell behavior of migrating human corneal limbal-epithelial cell sheets were studied on highly biocompatible flat and micro-patterned silk film surfaces. The silk film edge topography guided the migratory direction of individual cells making up the collective epithelial sheet, which resulted in a 75% increase in total culture elongation. This was due to a 3-fold decrease in cell sheet migration rate efficiency for movement perpendicular to the topography edge. Individual cell migration direction is preferred in the parallel approach to the edge topography where localization of cytoskeletal proteins to the topography’s edge region is reduced, which results in the directed growth of the collective epithelial sheet. Findings indicate customized biomaterial surfaces may be created to direct both the migration rate and direction of tissue epithelialization. PMID:23185573

  6. Turbulent Dynamics of Epithelial Cell Cultures

    Science.gov (United States)

    Blanch-Mercader, C.; Yashunsky, V.; Garcia, S.; Duclos, G.; Giomi, L.; Silberzan, P.

    2018-05-01

    We investigate the large length and long time scales collective flows and structural rearrangements within in vitro human bronchial epithelial cell (HBEC) cultures. Activity-driven collective flows result in ensembles of vortices randomly positioned in space. By analyzing a large population of vortices, we show that their area follows an exponential law with a constant mean value and their rotational frequency is size independent, both being characteristic features of the chaotic dynamics of active nematic suspensions. Indeed, we find that HBECs self-organize in nematic domains of several cell lengths. Nematic defects are found at the interface between domains with a total number that remains constant due to the dynamical balance of nucleation and annihilation events. The mean velocity fields in the vicinity of defects are well described by a hydrodynamic theory of extensile active nematics.

  7. Invasive cancer cells and metastasis

    Science.gov (United States)

    Mierke, Claudia Tanja

    2013-12-01

    The physics of cancer is a relatively new emerging field of cancer research. In the last decade it has become a focus of biophysical research as well as becoming a novel focus for classical cancer research. This special section of Physical Biology focusing on invasive cancer cells and metastasis (physical oncology) will give greater insight into the different subfields where physical approaches are being applied to cancer research. This focus on the physical aspects of cancer is necessary because novel approaches in the field of genomics and proteomics have not altered the field of cancer research dramatically, due to the fact that few breakthroughs have been made. It is still not understood why some primary tumors metastasize and thus have a worse outcome compared to others that do not metastasize. As biophysicists, we and others suggest that the mechanical properties of the cancer cells, which possess the ability to transmigrate, are quite different compared to non-metastatic and non-invasive cancer cells. Furthermore, we hypothesize that these cancer cells undergo a selection process within the primary tumor that enables them to weaken their cell-cell adhesions and to alter their cell-matrix adhesions in order to be able to cross the outermost boundary of the primary tumor, as well as the surrounding basement membrane, and to invade the connective tissue. This prerequisite may also help the cancer cells to enter blood or lymph vessels, get transported with the vessel flow and form secondary tumors either within the vessel, directly on the endothelium, or in a different organ after crossing the endothelial lining a second time. This special section begins with a paper by Mark F Coughlin and Jeffrey J Fredberg on the changes in cytoskeletal dynamics and nonlinear rheology due to the metastatic capability of cancer cells from different cancer tissue types such as skin, bladder, prostate and kidney [1]. The hypothesis was that the metastatic outcome is impacted by

  8. Non-Saccharomyces yeasts protect against epithelial cell barrier disruption induced by Salmonella enterica subsp. enterica serovar Typhimurium

    DEFF Research Database (Denmark)

    Smith, Ida Mosbech; Baker, A; Arneborg, Nils

    2015-01-01

    distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase....... In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability......). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. SIGNIFICANCE AND IMPACT...

  9. Molecular Characterization of Gastric Epithelial Cells Using Flow Cytometry

    Directory of Open Access Journals (Sweden)

    Kevin A. Bockerstett

    2018-04-01

    Full Text Available The ability to analyze individual epithelial cells in the gastric mucosa would provide important insight into gastric disease, including chronic gastritis and progression to gastric cancer. However, the successful isolation of viable gastric epithelial cells (parietal cells, neck cells, chief cells, and foveolar cells from gastric glands has been limited due to difficulties in tissue processing. Furthermore, analysis and interpretation of gastric epithelial cell flow cytometry data has been difficult due to the varying sizes and light scatter properties of the different epithelial cells, high levels of autofluorescence, and poor cell viability. These studies were designed to develop a reliable method for isolating viable single cells from the corpus of stomachs and to optimize analyses examining epithelial cells from healthy and diseased stomach tissue by flow cytometry. We performed a two stage enzymatic digestion in which collagenase released individual gastric glands from the stromal tissue of the corpus, followed by a Dispase II digestion that dispersed these glands into greater than 1 × 106 viable single cells per gastric corpus. Single cell suspensions were comprised of all major cell lineages found in the normal gastric glands. A method describing light scatter, size exclusion, doublet discrimination, viability staining, and fluorescently-conjugated antibodies and lectins was used to analyze individual epithelial cells and immune cells. This technique was capable of identifying parietal cells and revealed that gastric epithelial cells in the chronically inflamed mucosa significantly upregulated major histocompatibility complexes (MHC I and II but not CD80 or CD86, which are costimulatory molecules involved in T cell activation. These studies describe a method for isolating viable single cells and a detailed description of flow cytometric analysis of cells from healthy and diseased stomachs. These studies begin to identify effects of

  10. induced acute cytotoxicity in human cervical epithelial carcinoma cells

    African Journals Online (AJOL)

    Molecular basis of arsenite (As +3 )-induced acute cytotoxicity in human cervical epithelial carcinoma cells. ... Libyan Journal of Medicine ... Methods: After performing cytotoxic assays on a human epithelial carcinoma cell line, expression analysis was done by quantitative polymerase chain reaction, western blotting, and ...

  11. Globoside accelerates the differentiation of dental epithelial cells into ameloblasts

    Institute of Scientific and Technical Information of China (English)

    Takashi Nakamura; Yuta Chiba; Masahiro Naruse; Kan Saito; Hidemitsu Harada; Satoshi Fukumoto

    2016-01-01

    Tooth crown morphogenesis is tightly regulated by the proliferation and differentiation of dental epithelial cells. Globoside (Gb4), a globo-series glycosphingolipid, is highly expressed during embryogenesis as well as organogenesis, including tooth development. We previously reported that Gb4 is dominantly expressed in the neutral lipid fraction of dental epithelial cells. However, because its functional role in tooth development remains unknown, we investigated the involvement of Gb4 in dental epithelial cell differentiation. The expression of Gb4 was detected in ameloblasts of postnatal mouse molars and incisors. A cell culture analysis using HAT-7 cells, a rat-derived dental epithelial cell line, revealed that Gb4 did not promote dental epithelial cell proliferation. Interestingly, exogenous administration of Gb4 enhanced the gene expression of enamel extracellular matrix proteins such as ameloblastin, amelogenin, and enamelin in dental epithelial cells as well as in developing tooth germs. Gb4 also induced the expression of TrkB, one of the key receptors required for ameloblast induction in dental epithelial cells. In contrast, Gb4 downregulated the expression of p75, a receptor for neurotrophins (including neurotrophin-4) and a marker of undifferentiated dental epithelial cells. In addition, we found that exogenous administration of Gb4 to dental epithelial cells stimulated the extracellular signal-regulated kinase and p38 mitogen-activated protein kinase signalling pathways. Furthermore, Gb4 induced the expression of epiprofin and Runx2, the positive regulators for ameloblastin gene transcription. Thus, our results suggest that Gb4 contributes to promoting the differentiation of dental epithelial cells into ameloblasts.

  12. Oral epithelial stem cells – implications in normal development and cancer metastasis

    Science.gov (United States)

    Papagerakis, Silvana; Pannone, Giuseppe; Zheng, Li; About, Imad; Taqi, Nawar; Nguyen, Nghia P.T.; Matossian, Margarite; McAlpin, Blake; Santoro, Angela; McHugh, Jonathan; Prince, Mark E.; Papagerakis, Petros

    2014-01-01

    Oral mucosa is continuously exposed to environmental forces and has to be constantly renewed. Accordingly, the oral mucosa epithelium contains a large reservoir of epithelial stem cells necessary for tissue homeostasis. Despite considerable scientific advances in stem cell behavior in a number of tissues, fewer studies have been devoted to the stem cells in the oral epithelium. Most of oral mucosa stem cells studies are focused on identifying cancer stem cells (CSC) in oral squamous cell carcinomas (OSCCs) among other head and neck cancers. OSCCs are the most prevalent epithelial tumors of the head and neck region, marked by their aggressiveness and invasiveness. Due to their highly tumorigenic properties, it has been suggested that CSC may be the critical population of cancer cells in the development of OSCC metastasis. This review presents a brief overview of epithelium stem cells with implications in oral health, and the clinical implications of the CSC concept in OSCC metastatic dissemination. PMID:24803391

  13. E. coli Nissle 1917 Affects Salmonella adhesion to porcine intestinal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Peter Schierack

    Full Text Available BACKGROUND: The probiotic Escherichia coli strain Nissle 1917 (EcN has been shown to interfere in a human in vitro model with the invasion of several bacterial pathogens into epithelial cells, but the underlying molecular mechanisms are not known. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the inhibitory effects of EcN on Salmonella Typhimurium invasion of porcine intestinal epithelial cells, focusing on EcN effects on the various stages of Salmonella infection including intracellular and extracellular Salmonella growth rates, virulence gene regulation, and adhesion. We show that EcN affects the initial Salmonella invasion steps by modulating Salmonella virulence gene regulation and Salmonella SiiE-mediated adhesion, but not extra- and intracellular Salmonella growth. However, the inhibitory activity of EcN against Salmonella invasion always correlated with EcN adhesion capacities. EcN mutants defective in the expression of F1C fimbriae and flagellae were less adherent and less inhibitory toward Salmonella invasion. Another E. coli strain expressing F1C fimbriae was also adherent to IPEC-J2 cells, and was similarly inhibitory against Salmonella invasion like EcN. CONCLUSIONS: We propose that EcN affects Salmonella adhesion through secretory components. This mechanism appears to be common to many E. coli strains, with strong adherence being a prerequisite for an effective reduction of SiiE-mediated Salmonella adhesion.

  14. Identification of molecular pathways facilitating glioma cell invasion in situ.

    Directory of Open Access Journals (Sweden)

    Ido Nevo

    Full Text Available Gliomas are mostly incurable secondary to their diffuse infiltrative nature. Thus, specific therapeutic targeting of invasive glioma cells is an attractive concept. As cells exit the tumor mass and infiltrate brain parenchyma, they closely interact with a changing micro-environmental landscape that sustains tumor cell invasion. In this study, we used a unique microarray profiling approach on a human glioma stem cell (GSC xenograft model to explore gene expression changes in situ in Invading Glioma Cells (IGCs compared to tumor core, as well as changes in host cells residing within the infiltrated microenvironment relative to the unaffected cortex. IGCs were found to have reduced expression of genes within the extracellular matrix compartment, and genes involved in cell adhesion, cell polarity and epithelial to mesenchymal transition (EMT processes. The infiltrated microenvironment showed activation of wound repair and tissue remodeling networks. We confirmed by protein analysis the downregulation of EMT and polarity related genes such as CD44 and PARD3 in IGCs, and EFNB3, a tissue-remodeling agent enriched at the infiltrated microenvironment. OLIG2, a proliferation regulator and glioma progenitor cell marker upregulated in IGCs was found to function in enhancing migration and stemness of GSCs. Overall, our results unveiled a more comprehensive picture of the complex and dynamic cell autonomous and tumor-host interactive pathways of glioma invasion than has been previously demonstrated. This suggests targeting of multiple pathways at the junction of invading tumor and microenvironment as a viable option for glioma therapy.

  15. Wnt is necessary for mesenchymal to epithelial transition in colorectal cancer cells.

    Science.gov (United States)

    Schwab, Renate H M; Amin, Nancy; Flanagan, Dustin J; Johanson, Timothy M; Phesse, Toby J; Vincan, Elizabeth

    2018-03-01

    Metastasis underlies most colorectal cancer mortality. Cancer cells spread through the body as single cells or small clusters of cells that have an invasive, mesenchymal, nonproliferative phenotype. At the secondary site, they revert to a proliferative "tumor constructing" epithelial phenotype to rebuild a tumor. We previously developed a unique in vitro three-dimensional model, called LIM1863-Mph, which faithfully recapitulates these reversible transitions that underpin colorectal cancer metastasis. Wnt signaling plays a key role in these transitions and is initiated by the coupling of extracellular Wnt to Frizzled (FZD). Using the LIM1863-Mph model system we demonstrated that the Wnt receptor FZD7 is necessary for mesenchymal to epithelial transition (MET). Here we investigate the role of Wnt in MET. Wnt secretion is dependent on palmitoylation by Porcupine (PORC). A PORC inhibitor (IWP2) that prevents Wnt secretion, blocked the epithelial transition of mesenchymal LIM1863-Mph cells. Wnt gene array analysis identified several Wnts that are upregulated in epithelial compared with mesenchymal LIM1863-Mph cells, suggesting these ligands in MET. Wnt2B was the most abundant differentially expressed Wnt gene. Indeed, recombinant Wnt2B could overcome the IWP2-mediated block in epithelial transition of mesenchymal LIM1863-Mph cells. Wnt2B co-operates with Frizzled7 to mediate MET in colorectal cancer. Developmental Dynamics 247:521-530, 2018. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  16. Cytomatrix synthesis in MDCK epithelial cells

    International Nuclear Information System (INIS)

    Mitchell, J.J.; Low, R.B.; Woodcock-Mitchell, J.L.

    1990-01-01

    Detailed information regarding the synthesis rates of individual protein components is important in understanding the assembly and dynamics of the cytoskeletal matrix of eukaryotic cells. As an approach to this topic, the dual isotope technique of Clark and Zak, was employed to measure fractional synthesis rates (FSRs) in growing and quiescent cultures of MDCK epithelial cells. Cell protein was labeled to equilibrium with [14C]leucine over several days and then pulse-labeled for 4 hours with [3H]leucine. FSRs (as percent per hour) were calculated from the 3H/14C ratio of cell extracts or individual proteins separated by two-dimensional polyacrylamide gel electrophoresis and the 3H/14C ratio of free leucine in the medium. Synthesis of total cell protein rose from approximately 1.4%/hour in quiescent cells to 3.5%/hour in the growing cultures. The latter rate was sufficient to account for the rate of protein accumulation and a low level of turnover in the growing cultures. The FSR of the buffered-Triton soluble extract was higher and the cytoskeletal FSR significantly lower than that for total protein in quiescent monolayers. This difference, however, was not observed in growing cultures. A distinct pattern of differences was seen in the FSRs of individual cytoskeletal proteins in the quiescent cultures. Vimentin synthesis was significantly lower than that of the keratins and the keratin FSRs were not obviously matched in pairwise fashion. Unexpectedly, the FSRs of alpha- and beta-tubulin diverged in quiescent cells with alpha-tubulin turnover exceeding beta-tubulin. Likewise, components of the microfilament lattice showed unequal fractional synthesis rates, myosin and alpha-actinin being faster than actin. In addition, the FSR for globular actin exceeded that of the cytoskeletal associated form

  17. TGF-β1 induced epithelial to mesenchymal transition (EMT in human bronchial epithelial cells is enhanced by IL-1β but not abrogated by corticosteroids

    Directory of Open Access Journals (Sweden)

    Zuraw Bruce L

    2009-10-01

    Full Text Available Abstract Background Chronic persistent asthma is characterized by ongoing airway inflammation and airway remodeling. The processes leading to airway remodeling are poorly understood, and there is increasing evidence that even aggressive anti-inflammatory therapy does not completely prevent this process. We sought to investigate whether TGFβ1 stimulates bronchial epithelial cells to undergo transition to a mesenchymal phenotype, and whether this transition can be abrogated by corticosteroid treatment or enhanced by the pro-inflammatory cytokine IL-1β. Methods BEAS-2B and primary normal human bronchial epithelial cells were stimulated with TGFβ1 and expression of epithelial and mesenchymal markers assessed by quantitative real-time PCR, immunoblotting, immunofluorescence microscopy and zymography. In some cases the epithelial cells were also incubated with corticosteroids or IL-1β. Results were analyzed using non-parametric statistical tests. Results Treatment of BEAS-2B or primary human bronchial epithelial cells with TGFβ1 significantly reduced the expression level of the epithelial adherence junction protein E-cadherin. TGFβ1 then markedly induced mesenchymal marker proteins such as collagen I, tenascin C, fibronectin and α-smooth muscle actin mRNA in a dose dependant manner. The process of mesenchymal transition was accompanied by a morphological change towards a more spindle shaped fibroblast cell type with a more motile and invasive phenotype. Corticosteroid pre-treatment did not significantly alter the TGFβ1 induced transition but IL-1β enhanced the transition. Conclusion Our results indicate, that TGFβ1 can induce mesenchymal transition in the bronchial epithelial cell line and primary cells. Since asthma has been strongly associated with increased expression of TGFβ1 in the airway, epithelial to mesenchymal transition may contribute to the contractile and fibrotic remodeling process that accompanies chronic asthma.

  18. Canine tracheal epithelial cells are more sensitive than rat tracheal epithelial cells to transforming growth factor beta induced growth inhibition

    International Nuclear Information System (INIS)

    Hubbs, A.F.; Hahn, F.F.; Kelly, G.; Thomassen, D.G.

    1988-01-01

    Transforming growth factor beta (TGFβ) markedly inhibited growth of canine tracheal epithelial (CTE) cells. Reduced responsiveness to TGFβ-induced growth inhibition accompanied neoplastic progression of these cells from primary to transformed to neoplastic. This was similar to the relationship between neoplastic progression and increased resistance to TGFβ-induced growth inhibition seen for rat tracheal epithelial (RTE) cells. The canine cells were more sensitive than rat cells to TGFβ-induced growth inhibition at all stages in the neoplastic process. (author)

  19. Transplantation of retinal pigment epithelial cells - a possible future treatment for age-related macular degeneration

    DEFF Research Database (Denmark)

    Wiencke, Anne Katrine

    2001-01-01

    ophthalmology, age-related macular degeneration, transplantation, retinal pigment epithelial cells, treatment......ophthalmology, age-related macular degeneration, transplantation, retinal pigment epithelial cells, treatment...

  20. Transplantation of retinal pigment epithelial cells - a possible future treatment for age-related macular degeneration

    DEFF Research Database (Denmark)

    Wiencke, Anne Katrine

    2001-01-01

    ophthalmology, age-related macular degeneration, retinal pigment epithelial cells, transplantation, treatment......ophthalmology, age-related macular degeneration, retinal pigment epithelial cells, transplantation, treatment...

  1. Overexpression of microRNA-194 suppresses the epithelial-mesenchymal transition in targeting stem cell transcription factor Sox3 in endometrial carcinoma stem cells.

    Science.gov (United States)

    Gong, Baolan; Yue, Yan; Wang, Renxiao; Zhang, Yi; Jin, Quanfang; Zhou, Xi

    2017-06-01

    The epithelial-mesenchymal transition is the key process driving cancer metastasis. MicroRNA-194 inhibits epithelial-mesenchymal transition in several cancers and its downregulation indicates a poor prognosis in human endometrial carcinoma. Self-renewal factor Sox3 induces epithelial-mesenchymal transition at gastrulation and is also involved epithelial-mesenchymal transition in several cancers. We intended to determine the roles of Sox3 in inducing epithelial-mesenchymal transition in endometrial cancer stem cells and the possible role of microRNA-194 in controlling Sox3 expression. Firstly, we found that Sox3 and microRNA-194 expressions were associated with the status of endometrial cancer stem cells in a panel of endometrial carcinoma tissue, the CD133+ cell was higher in tumorsphere than in differentiated cells, and overexpression of microRNA-194 would decrease CD133+ cell expression. Silencing of Sox3 in endometrial cancer stem cell upregulated the epithelial marker E-cadherin, downregulated the mesenchymal marker vimentin, and significantly reduced cell invasion in vitro; overexpression of Sox3 reversed these phenotypes. Furthermore, we discovered that the expression of Sox3 was suppressed by microRNA-194 through direct binding to the Sox3 3'-untranslated region. Ectopic expression of microRNA-194 in endometrial cancer stem cells induced a mesenchymal-epithelial transition by restoring E-cadherin expression, decreasing vimentin expression, and inhibiting cell invasion in vitro. Moreover, overexpression of microRNA-194 inhibited endometrial cancer stem cell invasion or metastasis in vivo by injection of adenovirus microRNA-194. These findings demonstrate the novel mechanism by which Sox3 contributes to endometrial cancer stem cell invasion and suggest that repression of Sox3 by microRNA-194 may have therapeutic potential to suppress endometrial carcinoma metastasis. The cancer stem cell marker, CD133, might be the surface marker of endometrial cancer stem

  2. Early Trypanosoma cruzi Infection Reprograms Human Epithelial Cells

    Directory of Open Access Journals (Sweden)

    María Laura Chiribao

    2014-01-01

    Full Text Available Trypanosoma cruzi, the causative agent of Chagas disease, has the peculiarity, when compared with other intracellular parasites, that it is able to invade almost any type of cell. This property makes Chagas a complex parasitic disease in terms of prophylaxis and therapeutics. The identification of key host cellular factors that play a role in the T. cruzi invasion is important for the understanding of disease pathogenesis. In Chagas disease, most of the focus is on the response of macrophages and cardiomyocytes, since they are responsible for host defenses and cardiac lesions, respectively. In the present work, we studied the early response to infection of T. cruzi in human epithelial cells, which constitute the first barrier for establishment of infection. These studies identified up to 1700 significantly altered genes regulated by the immediate infection. The global analysis indicates that cells are literally reprogrammed by T. cruzi, which affects cellular stress responses (neutrophil chemotaxis, DNA damage response, a great number of transcription factors (including the majority of NFκB family members, and host metabolism (cholesterol, fatty acids, and phospholipids. These results raise the possibility that early host cell reprogramming is exploited by the parasite to establish the initial infection and posterior systemic dissemination.

  3. No junctional communication between epithelial cells in hydra

    DEFF Research Database (Denmark)

    de Laat, S W; Tertoolen, L G; Grimmelikhuijzen, C J

    1980-01-01

    junctions between epithelial cells of hydra. However, until now, there has been no report published on whether these junctions enable the epithelial cells to exchange molecules of small molecular weight, as has been described in other organisms. Therefore we decided to investigate the communicative...... properties of the junctional membranes by electrophysiological methods and by intracellular-dye iontophoresis. We report here that no electrotonic coupling is detectable between epithelial cells of Hydra attenuata in: (1) intact animals, (2) head-regenerating animals, (3) cell re-aggregates, and (4) hydra...

  4. Epithelial-mesenchymal transition in breast epithelial cells treated with cadmium and the role of Snail.

    Science.gov (United States)

    Wei, Zhengxi; Shan, Zhongguo; Shaikh, Zahir A

    2018-04-01

    Epidemiological and experimental studies have implicated cadmium (Cd) with breast cancer. In breast epithelial MCF10A and MDA-MB-231 cells, Cd has been shown to promote cell growth. The present study examined whether Cd also promotes epithelial-mesenchymal transition (EMT), a hallmark of cancer progression. Human breast epithelial cells consisting of non-cancerous MCF10A, non-metastatic HCC 1937 and HCC 38, and metastatic MDA-MB-231 were treated with 1 or 3 μM Cd for 4 weeks. The MCF10A epithelial cells switched to a more mesenchymal-like morphology, which was accompanied by a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal markers N-cadherin and vimentin. In both non-metastatic HCC 1937 and HCC 38 cells, treatment with Cd decreased the epithelial marker claudin-1. In addition, E-cadherin also decreased in the HCC 1937 cells. Even the mesenchymal-like MDA-MB-231 cells exhibited an increase in the mesenchymal marker vimentin. These changes indicated that prolonged treatment with Cd resulted in EMT in both normal and cancer-derived breast epithelial cells. Furthermore, both the MCF10A and MDA-MB-231 cells labeled with Zcad, a dual sensor for tracking EMT, demonstrated a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal marker ZEB-1. Treatment of cells with Cd significantly increased the level of Snail, a transcription factor involved in the regulation of EMT. However, the Cd-induced Snail expression was completely abolished by actinomycin D. Luciferase reporter assay indicated that the expression of Snail was regulated by Cd at the promotor level. Snail was essential for Cd-induced promotion of EMT in the MDA-MB-231 cells, as knockdown of Snail expression blocked Cd-induced cell migration. Together, these results indicate that Cd promotes EMT in breast epithelial cells and does so by modulating the transcription of Snail. Copyright © 2018 Elsevier Inc. All rights reserved.

  5. Proteinase-activated receptor 4 stimulation-induced epithelial-mesenchymal transition in alveolar epithelial cells

    Directory of Open Access Journals (Sweden)

    Araki Hiromasa

    2007-04-01

    Full Text Available Abstract Background Proteinase-activated receptors (PARs; PAR1–4 that can be activated by serine proteinases such as thrombin and neutrophil catepsin G are known to contribute to the pathogenesis of various pulmonary diseases including fibrosis. Among these PARs, especially PAR4, a newly identified subtype, is highly expressed in the lung. Here, we examined whether PAR4 stimulation plays a role in the formation of fibrotic response in the lung, through alveolar epithelial-mesenchymal transition (EMT which contributes to the increase in myofibroblast population. Methods EMT was assessed by measuring the changes in each specific cell markers, E-cadherin for epithelial cell, α-smooth muscle actin (α-SMA for myofibroblast, using primary cultured mouse alveolar epithelial cells and human lung carcinoma-derived alveolar epithelial cell line (A549 cells. Results Stimulation of PAR with thrombin (1 U/ml or a synthetic PAR4 agonist peptide (AYPGKF-NH2, 100 μM for 72 h induced morphological changes from cobblestone-like structure to elongated shape in primary cultured alveolar epithelial cells and A549 cells. In immunocytochemical analyses of these cells, such PAR4 stimulation decreased E-cadherin-like immunoreactivity and increased α-SMA-like immunoreactivity, as observed with a typical EMT-inducer, tumor growth factor-β (TGF-β. Western blot analyses of PAR4-stimulated A549 cells also showed similar changes in expression of these EMT-related marker proteins. Such PAR4-mediated changes were attenuated by inhibitors of epidermal growth factor receptor (EGFR kinase and Src. PAR4-mediated morphological changes in primary cultured alveolar epithelial cells were reduced in the presence of these inhibitors. PAR4 stimulation increased tyrosine phosphorylated EGFR or tyrosine phosphorylated Src level in A549 cells, and the former response being inhibited by Src inhibitor. Conclusion PAR4 stimulation of alveolar epithelial cells induced epithelial

  6. Type 3 innate lymphoid cells maintain intestinal epithelial stem cells after tissue damage

    NARCIS (Netherlands)

    P. Aparicio-Domingo (Patricia); M. Romera Hernández (Mónica); J.J. Karrich (Julien J.); F.H.J. Cornelissen (Ferry); N. Papazian (Natalie); D.J. Lindenbergh-Kortleve (Dicky); J.A. Butler (James A.); L. Boon (Louis); M. Coles (Mark); J.N. Samsom (Janneke); T. Cupedo (Tom)

    2015-01-01

    textabstractDisruption of the intestinal epithelial barrier allows bacterial translocation and predisposes to destructive inflammation. To ensure proper barrier composition, crypt-residing stem cells continuously proliferate and replenish all intestinal epithelial cells within days. As a consequence

  7. Gremlin Activates the Smad Pathway Linked to Epithelial Mesenchymal Transdifferentiation in Cultured Tubular Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Raquel Rodrigues-Diez

    2014-01-01

    Full Text Available Gremlin is a developmental gene upregulated in human chronic kidney disease and in renal cells in response to transforming growth factor-β (TGF-β. Epithelial mesenchymal transition (EMT is one process involved in renal fibrosis. In tubular epithelial cells we have recently described that Gremlin induces EMT and acts as a downstream TGF-β mediator. Our aim was to investigate whether Gremlin participates in EMT by the regulation of the Smad pathway. Stimulation of human tubular epithelial cells (HK2 with Gremlin caused an early activation of the Smad signaling pathway (Smad 2/3 phosphorylation, nuclear translocation, and Smad-dependent gene transcription. The blockade of TGF-β, by a neutralizing antibody against active TGF-β, did not modify Gremlin-induced early Smad activation. These data show that Gremlin directly, by a TGF-β independent process, activates the Smad pathway. In tubular epithelial cells long-term incubation with Gremlin increased TGF-β production and caused a sustained Smad activation and a phenotype conversion into myofibroblasts-like cells. Smad 7 overexpression, which blocks Smad 2/3 activation, diminished EMT changes observed in Gremlin-transfected tubuloepithelial cells. TGF-β neutralization also diminished Gremlin-induced EMT changes. In conclusion, we propose that Gremlin could participate in renal fibrosis by inducing EMT in tubular epithelial cells through activation of Smad pathway and induction of TGF-β.

  8. Sodium selectivity of Reissner's membrane epithelial cells

    Directory of Open Access Journals (Sweden)

    Kim Kyunghee X

    2011-02-01

    Full Text Available Abstract Background Sodium absorption by Reissner's membrane is thought to contribute to the homeostasis of the volume of cochlear endolymph. It was previously shown that the absorptive transepithelial current was blocked by amiloride and benzamil. The most commonly-observed target of these drugs is the epithelial sodium channel (ENaC, which is composed of the three subunits α-,β- and γ-ENaC. However, other less-selective cation channels have also been observed to be sensitive to benzamil and amiloride. The aim of this study was to determine whether Reissner's membrane epithelial cells could support parasensory K+ absorption via amiloride- and benzamil-sensitive electrogenic pathways. Results We determined the molecular and functional expression of candidate cation channels with gene array (GEO GSE6196, RT-PCR, and whole-cell patch clamp. Transcript expression analysis of Reissner's membrane detected no amiloride-sensitive acid-sensing ion channels (ASIC1a, ASIC2a, ASIC2b nor amiloride-sensitive cyclic-nucleotide gated channels (CNGA1, CNGA2, CNGA4, CNGB3. By contrast, α-,β- and γ-ENaC were all previously reported as present in Reissner's membrane. The selectivity of the benzamil-sensitive cation currents was observed in whole-cell patch clamp recordings under Cl--free conditions where cations were the only permeant species. The currents were carried by Na+ but not K+, and the permeability of Li+ was greater than that of Na+ in Reissner's membrane. Complete replacement of bath Na+ with the inpermeable cation NMDG+ led to the same inward current as with benzamil in a Na+ bath. Conclusions These results are consistent with the amiloride/benzamil-sensitive absorptive flux of Reissner's membrane mediated by a highly Na+-selective channel that has several key characteristics in common with αβγ-ENaC. The amiloride-sensitive pathway therefore absorbs only Na+ in this epithelium and does not provide a parasensory K+ efflux route from scala

  9. Whole-Genome Sequencing of Invasion-Resistant Cells Identifies Laminin α2 as a Host Factor for Bacterial Invasion

    DEFF Research Database (Denmark)

    van Wijk, Xander M.; Döhrmann, Simon; Hallstrom, Bjorn

    2017-01-01

    cells. Whole-genome sequencing and transcriptome sequencing (RNA-Seq) uncovered a deletion in the gene encoding the laminin subunit α2 (Lama2) that eliminated much of domain L4a. Silencing of the long Lama2 isoform in wild-type cells strongly reduced bacterial invasion, whereas transfection with human...... LAMA2 cDNA significantly enhanced invasion in pgsA745 cells. The addition of exogenous laminin-α2β1γ1/laminin-α2β2γ1 strongly increased bacterial invasion in CHO cells, as well as in human alveolar basal epithelial and human brain microvascular endothelial cells. Thus, the L4a domain in laminin α2...

  10. Single nucleotide polymorphisms in the TP53 region and susceptibility to invasive epithelial ovarian cancer

    DEFF Research Database (Denmark)

    Schildkraut, Joellen M; Goode, Ellen L; Clyde, Merlise A

    2009-01-01

    The p53 protein is critical for multiple cellular functions including cell growth and DNA repair. We assessed whether polymorphisms in the region encoding TP53 were associated with risk of invasive ovarian cancer. The study population includes a total of 5,206 invasive ovarian cancer cases (2,829...

  11. Pancreatic stellate cells promote epithelial-mesenchymal transition in pancreatic cancer cells

    International Nuclear Information System (INIS)

    Kikuta, Kazuhiro; Masamune, Atsushi; Watanabe, Takashi; Ariga, Hiroyuki; Itoh, Hiromichi; Hamada, Shin; Satoh, Kennichi; Egawa, Shinichi; Unno, Michiaki; Shimosegawa, Tooru

    2010-01-01

    Research highlights: → Recent studies have shown that pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. → Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and scattered, fibroblast-like appearance. → PSCs decreased the expression of epithelial markers but increased that of mesenchymal markers, along with increased migration. → This study suggests epithelial-mesenchymal transition as a novel mechanism by which PSCs contribute to the aggressive behavior of pancreatic cancer cells. -- Abstract: The interaction between pancreatic cancer cells and pancreatic stellate cells (PSCs), a major profibrogenic cell type in the pancreas, is receiving increasing attention. There is accumulating evidence that PSCs promote the progression of pancreatic cancer by increasing cancer cell proliferation and invasion as well as by protecting them from radiation- and gemcitabine-induced apoptosis. Because epithelial-mesenchymal transition (EMT) plays a critical role in the progression of pancreatic cancer, we hypothesized that PSCs promote EMT in pancreatic cancer cells. Panc-1 and SUIT-2 pancreatic cancer cells were indirectly co-cultured with human PSCs isolated from patients undergoing operation for pancreatic cancer. The expression of epithelial and mesenchymal markers was examined by real-time PCR and immunofluorescent staining. The migration of pancreatic cancer cells was examined by scratch and two-chamber assays. Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and a scattered, fibroblast-like appearance. The expression of E-cadherin, cytokeratin 19, and membrane-associated β-catenin was decreased, whereas vimentin and Snail (Snai-1) expression was increased more in cancer cells co-cultured with PSCs than in mono-cultured cells. The migration of pancreatic cancer cells was increased by co-culture with PSCs. The PSC-induced decrease of E-cadherin expression was not altered by treatment with anti

  12. Andrographolide suppresses epithelial mesenchymal transition by inhibition of MAPK signalling pathway in lens epithelial cells.

    Science.gov (United States)

    Kayastha, Forum; Johar, Kaid; Gajjar, Devarshi; Arora, Anshul; Madhu, Hardik; Ganatra, Darshini; Vasavada, Abhay

    2015-06-01

    Epithelial mesenchymal transition (EMT) of lens epithelial cells (LECs) may contribute to the development of posterior capsular opacification (PCO), which leads to visual impairment. Andrographolide has been shown to have therapeutic potential against various cancers. However, its effect on human LECs is still unknown. The purpose of this study is to evaluate the effect of andrographolide on EMT induced by growth factors in the fetal human lens epithelial cell line (FHL 124). Initially the LECs were treated with growth factors (TGF-beta 2 and bFGF) to induce EMT. Subsequently these EMT-induced cells were treated with andrographolide at 100 and 500 nM concentrations for 24 h. Our results showed that FHL 124 cells treated with growth factors had a significant decrease in protein and m-RNA levels of epithelial markers pax6 and E-Cadherin. After administering andrographolide, these levels significantly increased. It was noticed that EMT markers alpha-SMA, fibronectin and collagen IV significantly decreased after treatment with andrographolide when compared to the other group. Treatment with andrographolide significantly inhibited phosphorylation of ERK and JNK. Cell cycle analysis showed that andrographolide did not arrest cells at G0/G1 or G2/M at tested concentrations. Our findings suggest that andrographolide helps sustain epithelial characteristics by modulating EMT markers and inhibiting the mitogen-activated protein kinase (MAPK) signalling pathway in LECs. Hence it can prove to be useful in curbing EMT-mediated PCO.

  13. Regulated Mucin Secretion from Airway Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Kenneth Bruce Adler

    2013-09-01

    Full Text Available Secretory epithelial cells of the proximal airways synthesize and secrete gel-forming polymeric mucins. The secreted mucins adsorb water to form mucus that is propelled by neighboring ciliated cells, providing a mobile barrier which removes inhaled particles and pathogens from the lungs. Several features of the intracellular trafficking of mucins make the airway secretory cell an interesting comparator for the cell biology of regulated exocytosis. Polymeric mucins are exceedingly large molecules (up to 3x10^6 D per monomer whose folding and initial polymerization in the ER requires the protein disulfide isomerase Agr2. In the Golgi, mucins further polymerize to form chains and possibly branched networks comprising more than 20 monomers. The large size of mucin polymers imposes constraints on their packaging into transport vesicles along the secretory pathway. Sugar side chains account for >70% of the mass of mucins, and their attachment to the protein core by O-glycosylation occurs in the Golgi. Mature polymeric mucins are stored in large secretory granules ~1 um in diameter. These are translocated to the apical membrane to be positioned for exocytosis by cooperative interactions among MARCKS, cysteine string protein (CSP, HSP70 and the cytoskeleton. Mucin granules undergo exocytic fusion with the plasma membrane at a low basal rate and a high stimulated rate. Both rates are mediated by a regulated exocytic mechanism as indicated by phenotypes in both basal and stimulated secretion in mice lacking Munc13-2, a sensor of the second messengers calcium and diacylglycerol (DAG. Basal secretion is induced by low levels of activation of P2Y2 purinergic and A3 adenosine receptors by extracellular ATP released in paracrine fashion and its metabolite adenosine. Stimulated secretion is induced by high levels of the same ligands, and possibly by inflammatory mediators as well. Activated receptors are coupled to phospholipase C by Gq, resulting in the

  14. CCAAT/enhancer binding protein beta (C/EBPβ) isoform balance as a regulator of epithelial-mesenchymal transition in mouse mammary epithelial cells

    International Nuclear Information System (INIS)

    Miura, Yuka; Hagiwara, Natsumi; Radisky, Derek C.; Hirai, Yohei

    2014-01-01

    Activation of the epithelial-mesenchymal transition (EMT) program promotes cell invasion and metastasis, and is reversed through mesenchymal-epithelial transition (MET) after formation of distant metastases. Here, we show that an imbalance of gene products encoded by the transcriptional factor C/EBPβ, LAP (liver-enriched activating protein) and LIP (liver-enriched inhibitory protein), can regulate both EMT- and MET-like phenotypic changes in mouse mammary epithelial cells. By using tetracycline repressive LIP expression constructs, we found that SCp2 cells, a clonal epithelial line of COMMA1-D cells, expressed EMT markers, lost the ability to undergo alveolar-like morphogenesis in 3D Matrigel, and acquired properties of benign adenoma cells. Conversely, we found that inducible expression of LAP in SCg6 cells, a clonal fibroblastic line of COMMA1-D cells, began to express epithelial keratins with suppression of proliferation. The overexpression of the C/EBPβ gene products in these COMMA1-D derivatives was suppressed by long-term cultivation on tissue culture plastic, but gene expression was maintained in cells grown on Matrigel or exposed to proteasome inhibitors. Thus, imbalances of C/EBPβ gene products in mouse mammary epithelial cells, which are affected by contact with basement membrane, are defined as a potential regulator of metastatic potential. - Highlights: • We created a temporal imbalance of C/EBPβ gene products in the mammary model cells. • The temporal up-regulation of LIP protein induced EMT-like cell behaviors. • The temporal up-regulation of LAP protein induced MET-like cell behaviors. • Excess amount of C/EBPβ gene products were eliminated by proteasomal-degradation. • Basement membrane components attenuated proteasome-triggered protein elimination

  15. URG11 Regulates Prostate Cancer Cell Proliferation, Migration, and Invasion

    Directory of Open Access Journals (Sweden)

    Bin Pan

    2018-01-01

    Full Text Available Upregulated gene 11 (URG11, a new gene upregulated by hepatitis B virus X protein, is involved in the development and progression of several tumors, including liver, stomach, lung, and colon cancers. However, the role of URG11 in prostate cancer remains yet to be elucidated. By determined expression in human prostate cancer tissues, URG11 was found significantly upregulated and positively correlated with the severity of prostate cancer, compared with that in benign prostatic hyperplasia tissues. Further, the mRNA and protein levels of URG11 were significantly upregulated in human prostate cancer cell lines (DU145, PC3, and LNCaP, compared with human prostate epithelial cell line (RWPE-1. Moreover, by the application of siRNA against URG11, the proliferation, migration, and invasion of prostate cancer cells were markedly inhibited. Genetic knockdown of URG11 also induced cell cycle arrest at G1/S phase, induced apoptosis, and decreased the expression level of β-catenin in prostate cancer cells. Overexpression of URG11 promoted the expression of β-catenin, the growth, the migration, and invasion ability of prostate cancer cells. Taken together, this study reveals that URG11 is critical for the proliferation, migration, and invasion in prostate cancer cells, providing the evidence of URG11 to be a novel potential therapeutic target of prostate cancer.

  16. Cell volume regulation in epithelial physiology and cancer

    DEFF Research Database (Denmark)

    Pedersen, Stine Helene Falsig; Hoffmann, Else Kay; Novak, Ivana

    2013-01-01

    expression of ion transporters and channels is now recognized as one of the hallmarks of cancer, it is timely to consider this especially for epithelia. Epithelial cells are highly proliferative and epithelial cancers, carcinomas, account for about 90% of all cancers. In this review we will focus on ion...... such as cancer, transepithelial and cell volume regulatory ion transport are dys-regulated. Furthermore, epithelial architecture and coordinated ion transport function are lost, cell survival/death balance is altered, and new interactions with the stroma arise, all contributing to drug resistance. Since altered...... transporters and channels with key physiological functions in epithelia and known roles in the development of cancer in these tissues. Their roles in cell survival, cell cycle progression, and development of drug resistance in epithelial cancers will be discussed....

  17. Chronic inorganic arsenic exposure in vitro induces a cancer cell phenotype in human peripheral lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Person, Rachel J.; Olive Ngalame, Ntube N.; Makia, Ngome L.; Bell, Matthew W.; Waalkes, Michael P.; Tokar, Erik J., E-mail: tokare@niehs.nih.gov

    2015-07-01

    Inorganic arsenic is a human lung carcinogen. We studied the ability of chronic inorganic arsenic (2 μM; as sodium arsenite) exposure to induce a cancer phenotype in the immortalized, non-tumorigenic human lung peripheral epithelial cell line, HPL-1D. After 38 weeks of continuous arsenic exposure, secreted matrix metalloproteinase-2 (MMP2) activity increased to over 200% of control, levels linked to arsenic-induced cancer phenotypes in other cell lines. The invasive capacity of these chronic arsenic-treated lung epithelial (CATLE) cells increased to 320% of control and colony formation increased to 280% of control. CATLE cells showed enhanced proliferation in serum-free media indicative of autonomous growth. Compared to control cells, CATLE cells showed reduced protein expression of the tumor suppressor gene PTEN (decreased to 26% of control) and the putative tumor suppressor gene SLC38A3 (14% of control). Morphological evidence of epithelial-to-mesenchymal transition (EMT) occurred in CATLE cells together with appropriate changes in expression of the EMT markers vimentin (VIM; increased to 300% of control) and e-cadherin (CDH1; decreased to 16% of control). EMT is common in carcinogenic transformation of epithelial cells. CATLE cells showed increased KRAS (291%), ERK1/2 (274%), phosphorylated ERK (p-ERK; 152%), and phosphorylated AKT1 (p-AKT1; 170%) protein expression. Increased transcript expression of metallothioneins, MT1A and MT2A and the stress response genes HMOX1 (690%) and HIF1A (247%) occurred in CATLE cells possibly in adaptation to chronic arsenic exposure. Thus, arsenic induced multiple cancer cell characteristics in human peripheral lung epithelial cells. This model may be useful to assess mechanisms of arsenic-induced lung cancer. - Highlights: • Chronic arsenic exposure transforms a human peripheral lung epithelia cell line. • Cells acquire characteristics in common with human lung adenocarcinoma cells. • These transformed cells provide a

  18. CUX1/Wnt signaling regulates Epithelial Mesenchymal Transition in EBV infected epithelial cells

    International Nuclear Information System (INIS)

    Malizia, Andrea P.; Lacey, Noreen; Walls, Dermot; Egan, Jim J.; Doran, Peter P.

    2009-01-01

    Idiopathic pulmonary fibrosis (IPF) is a refractory and lethal interstitial lung disease characterized by alveolar epithelial cells apoptosis, fibroblast proliferation and extra-cellular matrix protein deposition. EBV, localised to alveolar epithelial cells of pulmonary fibrosis patients is associated with a poor prognosis. A strategy based on microarray-differential gene expression analysis to identify molecular drivers of EBV-associated lung fibrosis was utilized. Alveolar epithelial cells were infected with EBV to identify genes whose expression was altered following TGFβ1-mediated lytic phase. EBV lytic reactivation by TGFβ1 drives a selective alteration in CUX1 variant (a) (NCBI accession number NM 1 81552) expression, inducing activation of non-canonical Wnt pathway mediators, implicating it in Epithelial Mesenchymal Transition (EMT), the molecular event underpinning scar production in tissue fibrosis. The role of EBV in EMT can be attenuated by antiviral strategies and inhibition of Wnt signaling by using All-Trans Retinoic Acids (ATRA). Activation of non-canonical Wnt signaling pathway by EBV in epithelial cells suggests a novel mechanism of EMT via CUX1 signaling. These data present a framework for further description of the link between infectious agents and fibrosis, a significant disease burden.

  19. CUX1/Wnt signaling regulates Epithelial Mesenchymal Transition in EBV infected epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Malizia, Andrea P.; Lacey, Noreen [Clinical Research Centre, School of Medicine and Medical Science, University College Dublin. 21, Nelson Street. Dublin, 7. Ireland (Ireland); Walls, Dermot [School of Biotechnology, Dublin City University. Dublin, 9. Ireland (Ireland); Egan, Jim J. [Advanced Lung Disease and Lung Transplant Program, Mater Misericordiae University Hospital. 44, Eccles Street. Dublin, 7. Ireland (Ireland); Doran, Peter P., E-mail: peter.doran@ucd.ie [Clinical Research Centre, School of Medicine and Medical Science, University College Dublin. 21, Nelson Street. Dublin, 7. Ireland (Ireland)

    2009-07-01

    Idiopathic pulmonary fibrosis (IPF) is a refractory and lethal interstitial lung disease characterized by alveolar epithelial cells apoptosis, fibroblast proliferation and extra-cellular matrix protein deposition. EBV, localised to alveolar epithelial cells of pulmonary fibrosis patients is associated with a poor prognosis. A strategy based on microarray-differential gene expression analysis to identify molecular drivers of EBV-associated lung fibrosis was utilized. Alveolar epithelial cells were infected with EBV to identify genes whose expression was altered following TGF{beta}1-mediated lytic phase. EBV lytic reactivation by TGF{beta}1 drives a selective alteration in CUX1 variant (a) (NCBI accession number NM{sub 1}81552) expression, inducing activation of non-canonical Wnt pathway mediators, implicating it in Epithelial Mesenchymal Transition (EMT), the molecular event underpinning scar production in tissue fibrosis. The role of EBV in EMT can be attenuated by antiviral strategies and inhibition of Wnt signaling by using All-Trans Retinoic Acids (ATRA). Activation of non-canonical Wnt signaling pathway by EBV in epithelial cells suggests a novel mechanism of EMT via CUX1 signaling. These data present a framework for further description of the link between infectious agents and fibrosis, a significant disease burden.

  20. Epithelial–mesenchymal transition during oncogenic transformation induced by hexavalent chromium involves reactive oxygen species-dependent mechanism in lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Ding, Song-Ze, E-mail: dingsongze@hotmail.com [Department of Internal Medicine, Henan Provincial People’s Hospital, Zhengzhou University, Wei-Wu Road, Zhengzhou, Henan 450000 (China); Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States); Yang, Yu-Xiu; Li, Xiu-Ling [Department of Internal Medicine, Henan Provincial People’s Hospital, Zhengzhou University, Wei-Wu Road, Zhengzhou, Henan 450000 (China); Michelli-Rivera, Audrey [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States); Han, Shuang-Yin [Department of Internal Medicine, Henan Provincial People’s Hospital, Zhengzhou University, Wei-Wu Road, Zhengzhou, Henan 450000 (China); Wang, Lei; Pratheeshkumar, Poyil; Wang, Xin; Lu, Jian; Yin, Yuan-Qin; Budhraja, Amit; Hitron, Andrew J. [Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States)

    2013-05-15

    Hexavalent chromium [Cr(VI)] is an important human carcinogen associated with pulmonary diseases and lung cancer. Exposure to Cr(VI) induces DNA damage, cell morphological change and malignant transformation in human lung epithelial cells. Despite extensive studies, the molecular mechanisms remain elusive, it is also not known if Cr(VI)-induced transformation might accompany with invasive properties to facilitate metastasis. We aimed to study Cr(VI)-induced epithelial–mesenchymal transition (EMT) and invasion during oncogenic transformation in lung epithelial cells. The results showed that Cr(VI) at low doses represses E-cadherin mRNA and protein expression, enhances mesenchymal marker vimentin expression and transforms the epithelial cell into fibroblastoid morphology. Cr(VI) also increases cell invasion and promotes colony formation. Further studies indicated that Cr(VI) uses multiple mechanisms to repress E-cadherin expression, including activation of E-cadherin repressors such as Slug, ZEB1, KLF8 and enhancement the binding of HDAC1 in E-cadherin gene promoter, but DNA methylation is not responsible for the loss of E-cadherin. Catalase reduces Cr(VI)-induced E-cadherin and vimentin protein expression, attenuates cell invasion in matrigel and colony formation on soft agar. These results demonstrate that exposure to a common human carcinogen, Cr(VI), induces EMT and invasion during oncogenic transformation in lung epithelial cells and implicate in cancer metastasis and prevention. - Graphical abstract: Epithelial–mesenchymal transition during oncogenic transformation induced by hexavalent chromium involves reactive oxygen species-dependent mechanisms in lung epithelial cells. - Highlights: • We study if Cr(VI) might induce EMT and invasion in epithelial cells. • Cr(VI) induces EMT by altering E-cadherin and vimentin expression. • It also increases cell invasion and promotes oncogenic transformation. • Catalase reduces Cr(VI)-induced EMT, invasion and

  1. Two-layer tissue engineered urethra using oral epithelial and muscle derived cells.

    Science.gov (United States)

    Mikami, Hiroshi; Kuwahara, Go; Nakamura, Nobuyuki; Yamato, Masayuki; Tanaka, Masatoshi; Kodama, Shohta

    2012-05-01

    We fabricated novel tissue engineered urethral grafts using autologously harvested oral cells. We report their viability in a canine model. Oral tissues were harvested by punch biopsy and divided into mucosal and muscle sections. Epithelial cells from mucosal sections were cultured as epithelial cell sheets. Simultaneously muscle derived cells were seeded on collagen mesh matrices to form muscle cell sheets. At 2 weeks the sheets were joined and tubularized to form 2-layer tissue engineered urethras, which were autologously grafted to surgically induced urethral defects in 10 dogs in the experimental group. Tissue engineered grafts were not applied to the induced urethral defect in control dogs. The dogs were followed 12 weeks postoperatively. Urethrogram and histological examination were done to evaluate the grafting outcome. We successfully fabricated 2-layer tissue engineered urethras in vitro and transplanted them in dogs in the experimental group. The 12-week complication-free rate was significantly higher in the experimental group than in controls. Urethrogram confirmed urethral patency without stricture in the complication-free group at 12 weeks. Histologically urethras in the transplant group showed a stratified epithelial layer overlying well differentiated submucosa. In contrast, urethras in controls showed severe fibrosis without epithelial layer formation. Two-layer tissue engineered urethras were engineered using cells harvested by minimally invasive oral punch biopsy. Results suggest that this technique can encourage regeneration of a functional urethra. Copyright © 2012 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  2. Kaempferol modulates the metastasis of human non-small cell lung cancer cells by inhibiting epithelial-mesenchymal transition

    Directory of Open Access Journals (Sweden)

    Meng Hang

    2015-06-01

    Full Text Available The present study was done to determine whether kaempferol, a natural polyphenol of the flavonoid family, affects Epithelial-Mesenchymal Transition (EMT in non-small cell lung cancer cells. Kaempferol not only inhibited cancer cell proliferation and migration in a dose-dependent manner but also modulated the expression of EMT-related proteins E-cadherin and vimentin which are indispensible to cellular motility, invasiveness and metastasis. These results indicate that kaempferol suppresses non-small cell lung cancer migration by modulating the expression of EMT proteins. Therefore, kaempferol may be useful as a potential anticancer agent for non-small cell lung cancer.

  3. Extracellular Molecules Involved in Cancer Cell Invasion

    International Nuclear Information System (INIS)

    Stivarou, Theodora; Patsavoudi, Evangelia

    2015-01-01

    Nowadays it is perfectly clear that understanding and eradicating cancer cell invasion and metastasis represent the crucial, definitive points in cancer therapeutics. During the last two decades there has been a great interest in the understanding of the extracellular molecular mechanisms involved in cancer cell invasion. In this review, we highlight the findings concerning these processes, focusing in particular on extracellular molecules, including extracellular matrix proteins and their receptors, growth factors and their receptors, matrix metalloproteinases and extracellular chaperones. We report the molecular mechanisms underlying the important contribution of this pool of molecules to the complex, multi-step phenomenon of cancer cell invasion

  4. Extracellular Molecules Involved in Cancer Cell Invasion

    Directory of Open Access Journals (Sweden)

    Theodora Stivarou

    2015-01-01

    Full Text Available Nowadays it is perfectly clear that understanding and eradicating cancer cell invasion and metastasis represent the crucial, definitive points in cancer therapeutics. During the last two decades there has been a great interest in the understanding of the extracellular molecular mechanisms involved in cancer cell invasion. In this review, we highlight the findings concerning these processes, focusing in particular on extracellular molecules, including extracellular matrix proteins and their receptors, growth factors and their receptors, matrix metalloproteinases and extracellular chaperones. We report the molecular mechanisms underlying the important contribution of this pool of molecules to the complex, multi-step phenomenon of cancer cell invasion.

  5. Molecular Signature for Lymphatic Invasion Associated with Survival of Epithelial Ovarian Cancer.

    Science.gov (United States)

    Paik, E Sun; Choi, Hyun Jin; Kim, Tae-Joong; Lee, Jeong-Won; Kim, Byoung-Gie; Bae, Duk-Soo; Choi, Chel Hun

    2018-04-01

    We aimed to develop molecular classifier that can predict lymphatic invasion and their clinical significance in epithelial ovarian cancer (EOC) patients. We analyzed gene expression (mRNA, methylated DNA) in data from The Cancer Genome Atlas. To identify molecular signatures for lymphatic invasion, we found differentially expressed genes. The performance of classifier was validated by receiver operating characteristics analysis, logistic regression, linear discriminant analysis (LDA), and support vector machine (SVM). We assessed prognostic role of classifier using random survival forest (RSF) model and pathway deregulation score (PDS). For external validation,we analyzed microarray data from 26 EOC samples of Samsung Medical Center and curatedOvarianData database. We identified 21 mRNAs, and seven methylated DNAs from primary EOC tissues that predicted lymphatic invasion and created prognostic models. The classifier predicted lymphatic invasion well, which was validated by logistic regression, LDA, and SVM algorithm (C-index of 0.90, 0.71, and 0.74 for mRNA and C-index of 0.64, 0.68, and 0.69 for DNA methylation). Using RSF model, incorporating molecular data with clinical variables improved prediction of progression-free survival compared with using only clinical variables (p < 0.001 and p=0.008). Similarly, PDS enabled us to classify patients into high-risk and low-risk group, which resulted in survival difference in mRNA profiles (log-rank p-value=0.011). In external validation, gene signature was well correlated with prediction of lymphatic invasion and patients' survival. Molecular signature model predicting lymphatic invasion was well performed and also associated with survival of EOC patients.

  6. Different Effects of BORIS/CTCFL on Stemness Gene Expression, Sphere Formation and Cell Survival in Epithelial Cancer Stem Cells.

    Directory of Open Access Journals (Sweden)

    Loredana Alberti

    Full Text Available Cancer stem cells are cancer cells characterized by stem cell properties and represent a small population of tumor cells that drives tumor development, progression, metastasis and drug resistance. To date, the molecular mechanisms that generate and regulate cancer stem cells are not well defined. BORIS (Brother of Regulator of Imprinted Sites or CTCFL (CTCF-like is a DNA-binding protein that is expressed in normal tissues only in germ cells and is re-activated in tumors. Recent evidences have highlighted the correlation of BORIS/CTCFL expression with poor overall survival of different cancer patients. We have previously shown an association of BORIS-expressing cells with stemness gene expression in embryonic cancer cells. Here, we studied the role of BORIS in epithelial tumor cells. Using BORIS-molecular beacon that was already validated, we were able to show the presence of BORIS mRNA in cancer stem cell-enriched populations (side population and spheres of cervical, colon and breast tumor cells. BORIS silencing studies showed a decrease of sphere formation capacity in breast and colon tumor cells. Importantly, BORIS-silencing led to down-regulation of hTERT, stem cell (NANOG, OCT4, SOX2 and BMI1 and cancer stem cell markers (ABCG2, CD44 and ALDH1 genes. Conversely, BORIS-induction led to up-regulation of the same genes. These phenotypes were observed in cervical, colon and invasive breast tumor cells. However, a completely different behavior was observed in the non-invasive breast tumor cells (MCF7. Indeed, these cells acquired an epithelial mesenchymal transition phenotype after BORIS silencing. Our results demonstrate that BORIS is associated with cancer stem cell-enriched populations of several epithelial tumor cells and the different phenotypes depend on the origin of tumor cells.

  7. Epimorphin Functions as a Key Morphoregulator for Mammary Epithelial Cells

    Energy Technology Data Exchange (ETDEWEB)

    Hirai, H.; Lochter, A.; Galosy, S.; Koshida, S.; Niwa, S.; Bissell, M.J.

    1997-10-13

    Hepatocyte growth factor (HGF) and EGF have been reported to promote branching morphogenesis of mammary epithelial cells. We now show that it is epimorphin that is primarily responsible for this phenomenon. In vivo, epimorphin was detected in the stromal compartment but not in lumenal epithelial cells of the mammary gland; in culture, however, a subpopulation of mammary epithelial cells produced significant amounts of epimorphin. When epimorphin-expressing epithelial cell clones were cultured in collagen gels they displayed branching morphogenesis in the presence of HGF, EGF, keratinocyte growth factor, or fibroblast growth factor, a process that was inhibited by anti-epimorphin but not anti-HGF antibodies. The branch length, however, was roughly proportional to the ability of the factors to induce growth. Accordingly, epimorphin-negative epithelial cells simply grew in a cluster in response to the growth factors and failed to branch. When recombinant epimorphin was added to these collagen gels, epimorphin-negative cells underwent branching morphogenesis. The mode of action of epimorphin on morphogenesis of the gland, however, was dependent on how it was presented to the mammary cells. If epimorphin was overexpressed in epimorphin-negative epithelial cells under regulation of an inducible promoter or was allowed to coat the surface of each epithelial cell in a nonpolar fashion, the cells formed globular, alveoli-like structures with a large central lumen instead of branching ducts. This process was enhanced also by addition of HGF, EGF, or other growth factors and was inhibited by epimorphin antibodies. These results suggest that epimorphin is the primary morphogen in the mammary gland but that growth factors are necessary to achieve the appropriate cell numbers for the resulting morphogenesis to be visualized.

  8. Rabbit uterine epithelial cells: Co-culture with spermatozoa

    International Nuclear Information System (INIS)

    Boice, M.L.

    1988-01-01

    A primary culture of rabbit uterine epithelial cells was established and their effects on sperm function were examined in vitro. Epithelial cells were isolated from uteri of estrous rabbits and cultured on floating collagen gels in phenol red-free medium supplemented with 5% fetal bovine serum. Light microscopy and keratin staining showed that the epithelial cell population established in culture had morphological characteristics similar to that seen in the intact endometrium. Cells were cultured with 3 H-leucine and uptake of label by cells and its incorporation into cellular and secretory proteins determined. When compared to cells cultured for 24-48 h, incorporation of label into cellular protein was lower at 72-96 h, but secretion increased. Estradiol 17-β did not affect label uptake or incorporation, but did enhance proliferation of cells as judged by total DNA content of the cell population. Analysis of proteins in media by sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorography suggested that epithelial and stromal cells synthesis proteins that may be secretory in nature during 72-96 h culture. Twenty-nine to thirty-one h after initiation of epithelial cultures, 1-2 x 10 6 sperm were co-incubated with cells and sperm viability, motility, loss of acrosome and fertilizing ability determined

  9. Variation in NF-κB Signaling Pathways and Survival in Invasive Epithelial Ovarian Cancer

    Science.gov (United States)

    Block, Matthew S.; Charbonneau, Bridget; Vierkant, Robert A.; Fogarty, Zachary; Bamlet, William R.; Pharoah, Paul D.P.; Chenevix-Trench, Georgia; Rossing, Mary Anne; Cramer, Daniel; Pearce, Celeste Leigh; Schildkraut, Joellen; Menon, Usha; Kjaer, Susanne K.; Levine, Douglas A.; Gronwald, Jacek; Culver, Hoda Anton; Whittemore, Alice S.; Karlan, Beth Y.; Lambrechts, Diether; Wentzensen, Nicolas; Kupryjanczyk, Jolanta; Chang-Claude, Jenny; Bandera, Elisa V.; Hogdall, Estrid; Heitz, Florian; Kaye, Stanley B.; Fasching, Peter A.; Campbell, Ian; Goodman, Marc T.; Pejovic, Tanja; Bean, Yukie T.; Hays, Laura E.; Lurie, Galina; Eccles, Diana; Hein, Alexander; Beckmann, Matthias W.; Ekici, Arif B.; Paul, James; Brown, Robert; Flanagan, James M.; Harter, Philipp; du Bois, Andreas; Schwaab, Ira; Hogdall, Claus K.; Lundvall, Lene; Olson, Sara H.; Orlow, Irene; Paddock, Lisa E.; Rudolph, Anja; Eilber, Ursula; Dansonka-Mieszkowska, Agnieszka; Rzepecka, Iwona K.; Ziolkowska-Seta, Izabela; Brinton, Louise A.; Yang, Hannah; Garcia-Closas, Montserrat; Despierre, Evelyn; Lambrechts, Sandrina; Vergote, Ignace; Walsh, Christine S.; Lester, Jenny; Sieh, Weiva; McGuire, Valerie; Rothstein, Joseph H.; Ziogas, Argyrios; Lubiński, Jan; Cybulski, Cezary; Menkiszak, Janusz; Jensen, Allan; Gayther, Simon A.; Ramus, Susan J.; Gentry-Maharaj, Aleksandra; Berchuck, Andrew; Wu, Anna H.; Pike, Malcolm C.; Van Den Berg, David; Terry, Kathryn L.; Vitonis, Allison F.; Ramirez, Starr M.; Rider, David N.; Knutson, Keith L.; Sellers, Thomas A.; Phelan, Catherine M.; Doherty, Jennifer A.; Johnatty, Sharon E.; deFazio, Anna; Song, Honglin; Tyrer, Jonathan; Kalli, Kimberly R.; Fridley, Brooke L.; Cunningham, Julie M.; Goode, Ellen L.

    2014-01-01

    Survival in epithelial ovarian cancer (EOC) is influenced by the host immune response, yet the key genetic determinants of inflammation and immunity that impact prognosis are not known. The nuclear factor-kappa B (NF-κB) transcription factor family plays an important role in many immune and inflammatory responses, including the response to cancer. We studied common inherited variation in 210 genes in the NF-κB family in 10,084 patients with invasive EOC (5,248 high grade serous, 1,452 endometrioid, 795 clear cell, and 661 mucinous) from the Ovarian Cancer Association Consortium. Associations between genotype and overall survival were assessed using Cox regression for all patients and by major histology, adjusting for known prognostic factors and correcting for multiple testing (threshold for statistical significance—p < 2.5×10−5). Results were statistically significant when assessed for patients of a single histology. Key associations were with CARD11 (caspase recruitment domain family, member 11) rs41324349 in patients with mucinous EOC (HR 1.82, 95% CI 1.41–2.35, p=4.13×10−6) and TNFRSF13B (tumor necrosis factor receptor superfamily, member 13B) rs7501462 in patients with endometrioid EOC (HR 0.68, 95% CI 0.56–0.82, p=2.33×10−5). Other associations of note included TRAF2 (TNF receptor-associated factor 2) rs17250239 in patients with high-grade serous EOC (HR 0.84, 95% CI 0.77–0.92, p=6.49×10−5) and PLCG1 (phospholipase C, gamma 1) rs11696662 in patients with clear cell EOC (HR 0.43, 95% CI 0.26–0.73, p=4.56×10−4). These associations highlight the potential importance of genes associated with host inflammation and immunity in modulating clinical outcomes in distinct EOC histologies. PMID:24740199

  10. Tracking and Functional Characterization of Epithelial-Mesenchymal Transition and Mesenchymal Tumor Cells During Prostate Cancer Metastasis

    Science.gov (United States)

    Ruscetti, Marcus; Quach, Bill; Dadashian, Eman L.; Mulholland, David J.; Wu, Hong

    2015-01-01

    The epithelial-mesenchymal transition (EMT) has been postulated as a mechanism by which cancer cells acquire the invasive and stem-like traits necessary for distant metastasis. However, direct in vivo evidence for the role of EMT in the formation of cancer stem-like cells (CSC) and the metastatic cascade remains lacking. Here we report the first isolation and characterization of mesenchymal and EMT tumor cells, which harbor both epithelial and mesenchymal characteristics, in an autochthonous murine model of prostate cancer. By crossing the established Pb-Cre+/−;PtenL/L;KrasG12D/+ prostate cancer model with a vimentin-GFP reporter strain, generating CPKV mice, we were able to isolate epithelial, EMT and mesenchymal cancer cells based on expression of vimentin and EpCAM. CPKV mice (but not mice with Pten deletion alone) exhibited expansion of cells with EMT (EpCAM+/Vim-GFP+) and mesenchymal (EpCAM−/Vim-GFP+) characteristics at the primary tumor site and in circulation. These EMT and mesenchymal tumor cells displayed enhanced stemness and invasive character compared to epithelial tumor cells. Moreover, they displayed an enriched tumor-initiating capacity and could regenerate epithelial glandular structures in vivo, indicative of epithelia-mesenchyme plasticity. Interestingly, while mesenchymal tumor cells could persist in circulation and survive in the lung following intravenous injection, only epithelial and EMT tumor cells could form macrometastases. Our work extends the evidence that mesenchymal and epithelial states in cancer cells contribute differentially to their capacities for tumor initiation and metastatic seeding, respectively, and that EMT tumor cells exist with plasticity that can contribute to multiple stages of the metastatic cascade. PMID:25948589

  11. Rat glomerular epithelial cells in culture. Parietal or visceral epithelial origin

    International Nuclear Information System (INIS)

    Norgaard, J.O.

    1987-01-01

    Isolated glomeruli from rats were explanted under standard culture conditions and outgrowths were studied by light and electron microscopy in order to identify the cells. Rat glomerular samples contained 20 to 30% structurally well-preserved encapsulated glomeruli which had a large rate of attachment to the substrate and very constantly gave rise to cellular outgrowth. In order to label cells from which outgrowth originated the glomerular incorporation of [ 3 H]thymidine was studied in the preattachment phase. By light and electron microscope autoradiograph it was demonstrated that label was located only over visceral and parietal epithelial cells during the first 3 days of culture. Incorporation of [ 3 H]thymidine was seen in mesangial cells after 5 days, i.e., after the glomeruli had attached to the culture vessels and the initial outgrowth had appeared. Consequently the first cells to grow out were of epithelial origin. Glomeruli were then incubated with [ 3 H]thymidine for the first 2 1/2 days of culture in order to label the epithelial cells, then were allowed to attach to the substrate and induce cell outgrowth. By light microscope autoradiography performed with the outgrowths in situ two types of cells with labeled nuclei were seen: (a) a small, polyhedral ciliated cell which grew in colonies where the cells were joined by junctional complexes (type I), and (b) a second very large, often multinucleated cell (type II). Based on the structural resemblance with their counterparts in situ and on comparisons with positively identified visceral epithelial cells in outgrowths from other species it is suggested that type I cells are derived from the parietal epithelium of Bowman's capsule and type II cells from the visceral epithelium

  12. Fibronectin Modulates Cell Adhesion and Signaling to Promote Single Cell Migration of Highly Invasive Oral Squamous Cell Carcinoma

    Science.gov (United States)

    Ramos, Grasieli de Oliveira; Bernardi, Lisiane; Lauxen, Isabel; Sant’Ana Filho, Manoel; Horwitz, Alan Rick; Lamers, Marcelo Lazzaron

    2016-01-01

    Cell migration is regulated by adhesion to the extracellular matrix (ECM) through integrins and activation of small RhoGTPases, such as RhoA and Rac1, resulting in changes to actomyosin organization. During invasion, epithelial-derived tumor cells switch from laminin-enriched basal membrane to collagen and fibronectin-enriched connective tissue. How this switch affects the tumor migration is still unclear. We tested the hypothesis that ECM dictates the invasiveness of Oral Squamous Cell Carcinoma (OSCC). We analyzed the migratory properties of two OSCC lines, a low invasive cell line with high e-cadherin levels (Linv/HE-cad) or a highly invasive cell line with low e-cadherin levels (Hinv/LE-cad), plated on different ECM components. Compared to laminin, fibronectin induced non-directional collective migration and decreased RhoA activity in Linv/HE-cad OSCC. For Hinv/LE-cad OSCC, fibronectin increased Rac1 activity and induced smaller adhesions, resulting in a fast single cell migration in both 2D and 3D environments. Consistent with these observations, human OSCC biopsies exhibited similar changes in cell-ECM adhesion distribution at the invasive front of the tumor, where cells encounter fibronectin. Our results indicate that ECM composition might induce a switch from collective to single cell migration according to tumor invasiveness due to changes in cell-ECM adhesion and the resulting signaling pathways that alter actomyosin organization. PMID:26978651

  13. Fibronectin Modulates Cell Adhesion and Signaling to Promote Single Cell Migration of Highly Invasive Oral Squamous Cell Carcinoma.

    Directory of Open Access Journals (Sweden)

    Grasieli de Oliveira Ramos

    Full Text Available Cell migration is regulated by adhesion to the extracellular matrix (ECM through integrins and activation of small RhoGTPases, such as RhoA and Rac1, resulting in changes to actomyosin organization. During invasion, epithelial-derived tumor cells switch from laminin-enriched basal membrane to collagen and fibronectin-enriched connective tissue. How this switch affects the tumor migration is still unclear. We tested the hypothesis that ECM dictates the invasiveness of Oral Squamous Cell Carcinoma (OSCC. We analyzed the migratory properties of two OSCC lines, a low invasive cell line with high e-cadherin levels (Linv/HE-cad or a highly invasive cell line with low e-cadherin levels (Hinv/LE-cad, plated on different ECM components. Compared to laminin, fibronectin induced non-directional collective migration and decreased RhoA activity in Linv/HE-cad OSCC. For Hinv/LE-cad OSCC, fibronectin increased Rac1 activity and induced smaller adhesions, resulting in a fast single cell migration in both 2D and 3D environments. Consistent with these observations, human OSCC biopsies exhibited similar changes in cell-ECM adhesion distribution at the invasive front of the tumor, where cells encounter fibronectin. Our results indicate that ECM composition might induce a switch from collective to single cell migration according to tumor invasiveness due to changes in cell-ECM adhesion and the resulting signaling pathways that alter actomyosin organization.

  14. DA-6034 Induces [Ca(2+)]i Increase in Epithelial Cells.

    Science.gov (United States)

    Yang, Yu-Mi; Park, Soonhong; Ji, Hyewon; Kim, Tae-Im; Kim, Eung Kweon; Kang, Kyung Koo; Shin, Dong Min

    2014-04-01

    DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca(2+) signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca(2+) signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca(2+)-activated Cl(-) channels (CaCCs) and increased intracellular calcium concentrations ([Ca(2+)]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca(2+)]i in mouse salivary gland cells and human corneal epithelial cells. [Ca(2+)]i increase of DA-6034 was dependent on the Ca(2+) entry from extracellular and Ca(2+) release from internal Ca(2+) stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca(2+) stores. These results suggest that DA-6034 induces Ca(2+) signaling via extracellular Ca(2+) entry and RyRs-sensitive Ca(2+) release from internal Ca(2+) stores in epithelial cells.

  15. Plasmodium falciparum ookinetes require mosquito midgut chondroitin sulfate proteoglycans for cell invasion.

    NARCIS (Netherlands)

    Dinglasan, R.R.; Alaganan, A.; Ghosh, A.K.; Saito, A.; Kuppevelt, A.H.M.S.M. van; Jacobs-Lorena, M.

    2007-01-01

    Malaria transmission entails development of the Plasmodium parasite in its insect vector, the Anopheles mosquito. Parasite invasion of the mosquito midgut is the critical first step and involves adhesion to host epithelial cell ligands. Partial evidence suggests that midgut oligosaccharides are

  16. From cells to tissue: A continuum model of epithelial mechanics

    Science.gov (United States)

    Ishihara, Shuji; Marcq, Philippe; Sugimura, Kaoru

    2017-08-01

    A two-dimensional continuum model of epithelial tissue mechanics was formulated using cellular-level mechanical ingredients and cell morphogenetic processes, including cellular shape changes and cellular rearrangements. This model incorporates stress and deformation tensors, which can be compared with experimental data. Focusing on the interplay between cell shape changes and cell rearrangements, we elucidated dynamical behavior underlying passive relaxation, active contraction-elongation, and tissue shear flow, including a mechanism for contraction-elongation, whereby tissue flows perpendicularly to the axis of cell elongation. This study provides an integrated scheme for the understanding of the orchestration of morphogenetic processes in individual cells to achieve epithelial tissue morphogenesis.

  17. Micronucleus Assay in Exfoliated Buccal Epithelial Cells Using Liquid Based Cytology Preparations in Building Construction Workers.

    Science.gov (United States)

    Arul, P; Smitha, Shetty; Masilamani, Suresh; Akshatha, C

    2018-01-01

    Cytogenetic damage in exfoliated buccal epithelial cells due to environmental and occupational exposure is often monitored by micronucleus (MN) assay using liquid based cytology (LBC) preparations. This study was performed to evaluate MN in exfoliated buccal epithelial cells of building construction workers using LBC preparations. LBC preparations of exfoliated buccal epithelial cells from 100 subjects [50 building construction workers (cases) and 50 administrative staffs (controls)] was evaluated by May-Grunwald Giemsa, Hematoxylin and Eosin and Papanicolaou stains. Student's t test was used for statistical analysis and a P value of 5 years) and smokers and non-smokers of cases (P=0.001). However, there were meaningful differences regarding mean frequencies of MN between smokers, non-smokers, those with alcohol consumption or not in cases and controls using various stains (P=0.001). There was an increased risk of cytogenetic damage in building construction workers. However, evaluation of MN of exfoliated buccal epithelial cells in building construction workers serve as a minimally invasive biomarker for cytogenetic damage. LBC preparations can be applied for MN assay as it improves the quality of smears and cell morphology, decreases the confounding factors and reduces false positive results.

  18. Protein kinase Cδ signaling downstream of the EGF receptor mediates migration and invasiveness of prostate cancer cells

    International Nuclear Information System (INIS)

    Kharait, Sourabh; Dhir, Rajiv; Lauffenburger, Douglas; Wells, Alan

    2006-01-01

    Tumor progression to the invasive phenotype occurs secondary to upregulated signaling from growth factor receptors that drive key cellular responses like proliferation, migration, and invasion. We hypothesized that Protein kinase Cδ (PKCδ)-mediated transcellular contractility is required for migration and invasion of prostate tumor cells. Two invasive human prostate cancer cell lines, DU145 cells overexpressing wildtype human EGFR (DU145WT) and PC3 cells, were studied. PKCδ is overexpressed in these cells relative to normal prostate epithelial cells, and is activated downstream of EGFR leading to cell motility via modulation of myosin light chain activity. Abrogation of PKCδ using Rottlerin and specific siRNA significantly decreased migration and invasion of both cell lines in vitro. Both PKCδ and phosphorylated PKCδ protein levels were higher in human prostate cancer tissue relative to normal donor prostate as assessed by Western blotting and immunohistochemistry. Thus, we conclude that PKCδ inhibition can limit migration and invasion of prostate cancer cells

  19. Human airway epithelial cells investigated by atomic force microscopy: A hint to cystic fibrosis epithelial pathology

    Energy Technology Data Exchange (ETDEWEB)

    Lasalvia, Maria [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Castellani, Stefano [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy); D’Antonio, Palma [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Perna, Giuseppe [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Carbone, Annalucia [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy); Colia, Anna Laura; Maffione, Angela Bruna [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Capozzi, Vito [Department of Clinical and Experimental Medicine, University of Foggia, Foggia (Italy); Istituto Nazionale di Fisica Nucleare, Sezione di Bari, Bari (Italy); Conese, Massimo, E-mail: massimo.conese@unifg.it [Department of Medical and Surgical Sciences, University of Foggia, Foggia (Italy)

    2016-10-15

    The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption. - Highlights: • CF bronchial epithelial (CFBE) cells show a disorganized actin cytoskeleton. • CFBE cells present high roughness and low rigidity in

  20. Human airway epithelial cells investigated by atomic force microscopy: A hint to cystic fibrosis epithelial pathology

    International Nuclear Information System (INIS)

    Lasalvia, Maria; Castellani, Stefano; D’Antonio, Palma; Perna, Giuseppe; Carbone, Annalucia; Colia, Anna Laura; Maffione, Angela Bruna; Capozzi, Vito; Conese, Massimo

    2016-01-01

    The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption. - Highlights: • CF bronchial epithelial (CFBE) cells show a disorganized actin cytoskeleton. • CFBE cells present high roughness and low rigidity in the

  1. Relationship between circulating tumor cells and epithelial to mesenchymal transition in early breast cancer

    International Nuclear Information System (INIS)

    Mego, M.; Cierna, Z.; Janega, P.; Karaba, M.; Minarik, G.; Benca, J.; Sedlácková, T.; Sieberova, G.; Gronesova, P.; Manasova, D.; Pindak, D.; Sufliarsky, J.; Danihel, L.; Reuben, JM; Mardiak, J.

    2015-01-01

    Circulating tumor cells (CTCs) play a crucial role in tumor dissemination and are an independent survival predictor in breast cancer (BC) patients. Epithelial to mesenchymal transition (EMT) is involved in cancer invasion and metastasis. The aim of this study was to assess correlation between CTCs and expression of EMT transcription factors TWIST1 and SLUG in breast tumor tissue. This study included 102 early BC patients treated by primary surgery. Peripheral blood mononuclear cells (PBMC) were depleted of hematopoietic cells using RossetteSep™ negative selection kit. RNA extracted from CD45-depleted PBMC was interrogated for expression of EMT (TWIST1, SNAIL1, SLUG, FOXC2 and ZEB1) and epithelial (KRT19) gene transcripts by qRT-PCR. Expression of TWIST1 and SLUG in surgical specimens was evaluated by immunohistochemistry and quantified by multiplicative score. CTCs were detected in 24.5 % patients. CTCs exhibiting only epithelial markers were present in 8.8 % patients, whereas CTCs with only EMT markers were observed in 12.8 % of pts and CTCs co-expressing both markers were detected in 2.9 % pts. We observed lack of correlation between CTCs and expression of TWIST1 and SLUG in breast cancer cells or cancer associated stroma. Lack of correlation was observed for epithelial CTCs as well as for CTCs with EMT. In this translational study, we showed a lack of association between CTCs and expression of EMT-inducing transcription factors, TWIST1 and SLUG, in breast tumor tissue. Despite the fact that EMT is involved in cancer invasion and metastasis our results suggest, that expression of EMT proteins in unselected tumor tissue is not surrogate marker of CTCs with either mesenchymal or epithelial features

  2. Probiotics promote endocytic allergen degradation in gut epithelial cells

    International Nuclear Information System (INIS)

    Song, Chun-Hua; Liu, Zhi-Qiang; Huang, Shelly; Zheng, Peng-Yuan; Yang, Ping-Chang

    2012-01-01

    Highlights: ► Knockdown of A20 compromised the epithelial barrier function. ► The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. ► Antigens transported across A20-deficient HT-29 monolayers conserved antigenicity. ► Probiotic proteins increased the expression of A20 in HT-29 cells. -- Abstract: Background and aims: Epithelial barrier dysfunction plays a critical role in the pathogenesis of allergic diseases; the mechanism is to be further understood. The ubiquitin E3 ligase A20 (A20) plays a role in the endocytic protein degradation in the cells. This study aims to elucidate the role of A20 in the maintenance of gut epithelial barrier function. Methods: Gut epithelial cell line, HT-29 cell, was cultured into monolayers to evaluate the barrier function in transwells. RNA interference was employed to knock down the A20 gene in HT-29 cells to test the role of A20 in the maintenance of epithelial barrier function. Probiotic derived proteins were extracted from the culture supernatants using to enhance the expression of A20 in HT-29 cells. Results: The results showed that the knockdown of A20 compromised the epithelial barrier function in HT-29 monolayers, mainly increased the intracellular permeability. The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Allergens collected from the transwell basal chambers of A20-deficient HT-29 monolayers still conserved functional antigenicity. Treating with probiotic derived proteins increased the expression of A20 in HT-29 cells and promote the barrier function. Conclusion: A20 plays an important role in the maintenance of epithelial barrier function as shown by HT-29 monolayer. Probiotic derived protein increases the expression of A20 and promote the HT-29 monolayer barrier function.

  3. Probiotics promote endocytic allergen degradation in gut epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Song, Chun-Hua [Department of Epidemiology and Biostatistics, College of Public Health, Zhengzhou University, Zhengzhou (China); Liu, Zhi-Qiang [Department of Gastroenterology, The Second Hospital, Zhengzhou University, Zhengzhou (China); Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada); Huang, Shelly [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada); Zheng, Peng-Yuan, E-mail: medp7123@126.com [Department of Gastroenterology, The Second Hospital, Zhengzhou University, Zhengzhou (China); Yang, Ping-Chang, E-mail: yangp@mcmaster.ca [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON (Canada)

    2012-09-14

    Highlights: Black-Right-Pointing-Pointer Knockdown of A20 compromised the epithelial barrier function. Black-Right-Pointing-Pointer The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Black-Right-Pointing-Pointer Antigens transported across A20-deficient HT-29 monolayers conserved antigenicity. Black-Right-Pointing-Pointer Probiotic proteins increased the expression of A20 in HT-29 cells. -- Abstract: Background and aims: Epithelial barrier dysfunction plays a critical role in the pathogenesis of allergic diseases; the mechanism is to be further understood. The ubiquitin E3 ligase A20 (A20) plays a role in the endocytic protein degradation in the cells. This study aims to elucidate the role of A20 in the maintenance of gut epithelial barrier function. Methods: Gut epithelial cell line, HT-29 cell, was cultured into monolayers to evaluate the barrier function in transwells. RNA interference was employed to knock down the A20 gene in HT-29 cells to test the role of A20 in the maintenance of epithelial barrier function. Probiotic derived proteins were extracted from the culture supernatants using to enhance the expression of A20 in HT-29 cells. Results: The results showed that the knockdown of A20 compromised the epithelial barrier function in HT-29 monolayers, mainly increased the intracellular permeability. The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Allergens collected from the transwell basal chambers of A20-deficient HT-29 monolayers still conserved functional antigenicity. Treating with probiotic derived proteins increased the expression of A20 in HT-29 cells and promote the barrier function. Conclusion: A20 plays an important role in the maintenance of epithelial barrier function as shown by HT-29 monolayer. Probiotic derived protein increases the expression of A20 and promote the HT-29 monolayer barrier function.

  4. Lingual Epithelial Stem Cells and Organoid Culture of Them

    Directory of Open Access Journals (Sweden)

    Hiroko Hisha

    2016-01-01

    Full Text Available As tongue cancer is one of the major malignant cancers in the world, understanding the mechanism of maintenance of lingual epithelial tissue, which is known to be the origin of tongue cancer, is unquestionably important. However, the actual stem cells that are responsible for the long-term maintenance of the lingual epithelium have not been identified. Moreover, a simple and convenient culture method for lingual epithelial stem cells has not yet been established. Recently, we have shown that Bmi1-positive cells, residing at the second or third layer of the epithelial cell layer at the base of the interpapillary pit (IPP, were slow-cycling and could supply keratinized epithelial cells for over one year, indicating that Bmi1-positive cells are long-term lingual epithelial stem cells. In addition, we have developed a novel lingual epithelium organoid culture system using a three-dimensional matrix and growth factors. Here, we discuss current progress in the identification of lingual stem cells and future applications of the lingual culture system for studying the regulatory mechanisms of the lingual epithelium and for regenerative medicine.

  5. Cell signaling during Trypanosoma cruzi invasion

    Directory of Open Access Journals (Sweden)

    Fernando Yukio Maeda

    2012-11-01

    Full Text Available Cell signaling is an essential requirement for mammalian cell invasion by Trypanosoma cruzi. Depending on the parasite strain and the parasite developmental form, distinct signaling pathways may be induced. In this short review, we focus on the data coming from studies with metacyclic trypomastigotes (MT generated in vitro and tissue culture-derived trypomastigotes (TCT, used as counterparts of insect-borne and bloodstream parasites respectively. During invasion of host cells by MT or TCT, intracellular Ca2+ mobilization and host cell lysosomal exocytosis are triggered. Invasion mediated by MT surface molecule gp82 requires the activation of mammalian target of rapamycin (mTOR, phosphatidylinositol 3-kinase (PI3K and protein kinase C (PKC in the host cell, associated with Ca2+-dependent disruption of the actin cytoskeleton. In MT, protein tyrosine kinase (PTK, PI3K, phospholipase C (PLC and PKC appear to be activated. TCT invasion, on the other hand, does not rely on mTOR activation, rather on target cell PI3K, and may involve the host cell autophagy for parasite internalization. Enzymes, such oligopeptidase B and the major T. cruzi cysteine proteinase cruzipain, have been shown to generate molecules that induce target cell Ca2+ signal. In addition, TCT may trigger host cell responses mediated by TGF-β receptor or integrin family member. Further investigations are needed for a more complete and detailed picture of T. cruzi invasion.

  6. The regenerative potential of parietal epithelial cells in adult mice

    NARCIS (Netherlands)

    Berger, K.; Schulte, K.; Boor, P.; Kuppe, C.; Kuppevelt, T.H. van; Floege, J.; Smeets, B.; Moeller, M.J.

    2014-01-01

    Previously, we showed that some podocytes in juvenile mice are recruited from cells lining Bowman's capsule, suggesting that parietal epithelial cells (PECs) are a progenitor cell population for podocytes. To investigate whether PECs also replenish podocytes in adult mice, PECs were genetically

  7. stg fimbrial operon from S. Typhi STH2370 contributes to association and cell disruption of epithelial and macrophage-like cells.

    Science.gov (United States)

    Berrocal, Liliana; Fuentes, Juan A; Trombert, A Nicole; Jofré, Matías R; Villagra, Nicolás A; Valenzuela, Luis M; Mora, Guido C

    2015-07-07

    Salmonella enterica serovar Typhi (S. Typhi) stg operon, encoding a chaperone/usher fimbria (CU), contributes to an increased adherence to human epithelial cells. However, one report suggests that the presence of the Stg fimbria impairs the monocyte--bacteria association, as deduced by the lower level of invasion to macrophage-like cells observed when the stg fimbrial cluster was overexpressed. Nevertheless, since other CU fimbrial structures increase the entry of S. Typhi into macrophages, and considering that transcriptomic analyses revealed that stg operon is indeed expressed in macrophages, we reassessed the role of the stg operon in the interaction between S. Typhi strain STH2370 and human cells, including macrophage-like cells and mononuclear cells directly taken from human peripheral blood. We compared S. Typhi STH2370 WT, a Chilean clinical strain, and the S. Typhi STH2370 Δstg mutant with respect to association and invasion using epithelial and macrophage-like cells. We observed that deletion of stg operon reduced the association and invasion of S. Typhi, in both cellular types. The presence of the cloned stg operon restored the WT phenotype in all the cases. Moreover, we compared Salmonella enterica sv. Typhimurium 14028s (S. Typhimurium, a serovar lacking stg operon) and S. Typhimurium heterologously expressing S. Typhi stg. We found that the latter presents an increased cell disruption of polarized epithelial cells and an increased association in both epithelial and macrophage-like cells. S. Typhi stg operon encodes a functional adhesin that participates in the interaction bacteria-eukaryotic cells, including epithelial cells and macrophages-like cells. The phenotypes associated to stg operon include increased association and consequent invasion in bacteria-eukaryotic cells, and cell disruption.

  8. The Bacterial Species Campylobacter jejuni Induce Diverse Innate Immune Responses in Human and Avian Intestinal Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Daniel A. John

    2017-09-01

    Full Text Available Campylobacter remain the major cause of human gastroenteritis in the Developed World causing a significant burden to health services. Campylobacter are pathogens in humans and chickens, although differences in mechanistic understanding are incomplete, in part because phenotypic strain diversity creates inconsistent findings. Here, we took Campylobacter jejuni isolates (n = 100 from multi-locus sequence typed collections to assess their pathogenic diversity, through their inflammatory, cytotoxicity, adhesion, invasion and signaling responses in a high-throughput model using avian and human intestinal epithelial cells. C. jejuni induced IL-8 and CXCLi1/2 in human and avian epithelial cells, respectively, in a MAP kinase-dependent manner. In contrast, IL-10 responses in both cell types were PI 3-kinase/Akt-dependent. C. jejuni strains showed diverse levels of invasion with high invasion dependent on MAP kinase signaling in both cell lines. C. jejuni induced diverse cytotoxic responses in both cell lines with cdt-positive isolates showing significantly higher toxicity. Blockade of endocytic pathways suggested that invasion by C. jejuni was clathrin- and dynamin-dependent but caveolae- independent in both cells. In contrast, IL-8 (and CXCLi1/2 production was dependent on clathrin, dynamin, and caveolae. This study is important because of its scale, and the data produced, suggesting that avian and human epithelial cells use similar innate immune pathways where the magnitude of the response is determined by the phenotypic diversity of the Campylobacter species.

  9. Retinoic Acid Signaling in Thymic Epithelial Cells Regulates Thymopoiesis

    DEFF Research Database (Denmark)

    Wendland, Kerstin; Niss, Kristoffer; Kotarsky, Knut

    2018-01-01

    Despite the essential role of thymic epithelial cells (TEC) in T cell development, the signals regulating TEC differentiation and homeostasis remain incompletely understood. In this study, we show a key in vivo role for the vitamin A metabolite, retinoic acid (RA), in TEC homeostasis. In the abse......Despite the essential role of thymic epithelial cells (TEC) in T cell development, the signals regulating TEC differentiation and homeostasis remain incompletely understood. In this study, we show a key in vivo role for the vitamin A metabolite, retinoic acid (RA), in TEC homeostasis...

  10. Regulation of defensive function on gingival epithelial cells can prevent periodontal disease.

    Science.gov (United States)

    Fujita, Tsuyoshi; Yoshimoto, Tetsuya; Kajiya, Mikihito; Ouhara, Kazuhisa; Matsuda, Shinji; Takemura, Tasuku; Akutagawa, Keiichi; Takeda, Katsuhiro; Mizuno, Noriyoshi; Kurihara, Hidemi

    2018-05-01

    Periodontal disease is a bacterial biofilm-associated inflammatory disease that has been implicated in many systemic diseases. A new preventive method for periodontal disease needs to be developed in order to promote the health of the elderly in a super-aged society. The gingival epithelium plays an important role as a mechanical barrier against bacterial invasion and a part of the innate immune response to infectious inflammation in periodontal tissue. The disorganization of cell-cell interactions and subsequent inflammation contribute to the initiation of periodontal disease. These make us consider that regulation of host defensive functions, epithelial barrier and neutrophil activity, may become novel preventive methods for periodontal inflammation. Based on this concept, we have found that several agents regulate the barrier function of gingival epithelial cells and suppress the accumulation of neutrophils in the gingival epithelium. We herein introduce the actions of irsogladine maleate, azithromycin, amphotericin B, and Houttuynia cordata (dokudami in Japanese), which is commonly used in traditional medicine, on the epithelial barrier and neutrophil migration in gingival epithelial cells in vivo and in vitro , in order to provide support for the clinical application of these agents to the prevention of periodontal inflammation.

  11. Change in cell shape is required for matrix metalloproteinase-induced epithelial-mesenchymal transition of mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Nelson, Celeste M.; Khauv, Davitte; Bissell, Mina J.; Radisky, Derek C.

    2008-06-26

    Cell morphology dictates response to a wide variety of stimuli, controlling cell metabolism, differentiation, proliferation, and death. Epithelial-mesenchymal transition (EMT) is a developmental process in which epithelial cells acquire migratory characteristics, and in the process convert from a 'cuboidal' epithelial structure into an elongated mesenchymal shape. We had shown previously that matrix metalloproteinase-3 (MMP3) can stimulate EMT of cultured mouse mammary epithelial cells through a process that involves increased expression of Rac1b, a protein that stimulates alterations in cytoskeletal structure. We show here that cells treated with MMP-3 or induced to express Rac1b spread to cover a larger surface, and that this induction of cell spreading is a requirement of MMP-3/Rac1b-induced EMT. We find that limiting cell spreading, either by increasing cell density or by culturing cells on precisely defined micropatterned substrata, blocks expression of characteristic markers of EMT in cells treated with MMP-3. These effects are not caused by general disruptions in cell signaling pathways, as TGF-{beta}-induced EMT is not affected by similar limitations on cell spreading. Our data reveal a previously unanticipated cell shape-dependent mechanism that controls this key phenotypic alteration and provide insight into the distinct mechanisms activated by different EMT-inducing agents.

  12. Long-Term Live Cell Imaging of Cell Migration: Effects of Pathogenic Fungi on Human Epithelial Cell Migration.

    Science.gov (United States)

    Wöllert, Torsten; Langford, George M

    2016-01-01

    Long-term live cell imaging was used in this study to determine the responses of human epithelial cells to pathogenic biofilms formed by Candida albicans. Epithelial cells of the skin represent the front line of defense against invasive pathogens such as C. albicans but under certain circumstances, especially when the host's immune system is compromised, the skin barrier is breached. The mechanisms by which the fungal pathogen penetrates the skin and invade the deeper layers are not fully understood. In this study we used keratinocytes grown in culture as an in vitro model system to determine changes in host cell migration and the actin cytoskeleton in response to virulence factors produced by biofilms of pathogenic C. albicans. It is clear that changes in epithelial cell migration are part of the response to virulence factors secreted by biofilms of C. albicans and the actin cytoskeleton is the downstream effector that mediates cell migration. Our goal is to understand the mechanism by which virulence factors hijack the signaling pathways of the actin cytoskeleton to alter cell migration and thereby invade host tissues. To understand the dynamic changes of the actin cytoskeleton during infection, we used long-term live cell imaging to obtain spatial and temporal information of actin filament dynamics and to identify signal transduction pathways that regulate the actin cytoskeleton and its associated proteins. Long-term live cell imaging was achieved using a high resolution, multi-mode epifluorescence microscope equipped with specialized light sources, high-speed cameras with high sensitivity detectors, and specific biocompatible fluorescent markers. In addition to the multi-mode epifluorescence microscope, a spinning disk confocal long-term live cell imaging system (Olympus CV1000) equipped with a stage incubator to create a stable in vitro environment for long-term real-time and time-lapse microscopy was used. Detailed descriptions of these two long-term live

  13. Recreational physical inactivity and mortality in women with invasive epithelial ovarian cancer

    DEFF Research Database (Denmark)

    Cannioto, Rikki A.; LaMonte, Michael J.; Kelemen, Linda E

    2016-01-01

    Background: Little is known about modifiable behaviours that may be associated with epithelial ovarian cancer (EOC) survival. We conducted a pooled analysis of 12 studies from the Ovarian Cancer Association Consortium to investigate the association between pre-diagnostic physical inactivity...... and mortality. Methods: Participants included 6806 women with a primary diagnosis of invasive EOC. In accordance with the Physical Activity Guidelines for Americans, women reporting no regular, weekly recreational physical activity were classified as inactive. We utilised Cox proportional hazard models...... to estimate hazard ratios (HRs) and 95% confidence intervals (CIs) representing the associations of inactivity with mortality censored at 5 years. Results: In multivariate analysis, inactive women had significantly higher mortality risks, with (HR=1.34, 95% CI: 1.18-1.52) and without (HR=1.22, 95% CI: 1...

  14. Human thymic epithelial cells express functional HLA-DP molecules

    DEFF Research Database (Denmark)

    Jørgensen, A; Röpke, C; Nielsen, M

    1996-01-01

    T lymphocytes, we examined whether human thymic epithelial cells (TEC) expressed HLA-DP molecules. We present evidence that TEC obtained from short time culture express low but significant levels of HLA-DP molecules. The expression of HLA-DP molecules was comparable to or higher than the expression...... of HLA-DP allospecific primed lymphocyte typing (PLT) CD4 T cell lines. IFN-gamma treatment strongly upregulated the HLA-DP allospecific PLT responses whereas other PLT responses remained largely unchanged. In conclusion, these data indicate that human thymus epithelial cells express significant levels...

  15. Inflammatory Cytokine Tumor Necrosis Factor α Confers Precancerous Phenotype in an Organoid Model of Normal Human Ovarian Surface Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Joseph Kwong

    2009-06-01

    Full Text Available In this study, we established an in vitro organoid model of normal human ovarian surface epithelial (HOSE cells. The spheroids of these normal HOSE cells resembled epithelial inclusion cysts in human ovarian cortex, which are the cells of origin of ovarian epithelial tumor. Because there are strong correlations between chronic inflammation and the incidence of ovarian cancer, we used the organoid model to test whether protumor inflammatory cytokine tumor necrosis factor α would induce malignant phenotype in normal HOSE cells. Prolonged treatment of tumor necrosis factor α induced phenotypic changes of the HOSE spheroids, which exhibited the characteristics of precancerous lesions of ovarian epithelial tumors, including reinitiation of cell proliferation, structural disorganization, epithelial stratification, loss of epithelial polarity, degradation of basement membrane, cell invasion, and overexpression of ovarian cancer markers. The result of this study provides not only an evidence supporting the link between chronic inflammation and ovarian cancer formation but also a relevant and novel in vitro model for studying of early events of ovarian cancer.

  16. ELF5 in epithelial ovarian carcinoma tissues and biological behavior in ovarian carcinoma cells.

    Science.gov (United States)

    Yan, Hongchao; Qiu, Linglin; Xie, Xiaolei; Yang, He; Liu, Yongli; Lin, Xiaoman; Huang, Hongxiang

    2017-03-01

    The expression of E74-like factor 5 (ELF5) in epithelial ovarian carcinoma tissues and its effects on biological behavior in ovarian carcinoma cells were assessed in search for a new approach for gene treatment of epithelial ovarian carcinoma. RT-PCR technology was applied to detect the expression of ELF5 mRNA in epithelial ovarian carcinoma (n=49), borderline ovarian epithelial tumor (n=19), benign ovarian epithelial tumor (n=31) and normal ovarian tissues (n=40). Then, we transfected recombinant plasmid pcDNA3.1‑ELF5+EGFP into human ovarian carcinoma SKOV3 cells (recombinant plasmid group) in vitro and screened out stably transfected cells to conduct multiplication culture. Western blot analysis was performed to detect the expression of ELF5 protein in the different groups. Flow cytometry was employed to detect cell apoptosis and cycles. ELF5 mRNA in epithelial ovarian carcinoma and borderline ovarian epithelial tumor tissues were significantly lower (Pepithelial tumor and normal ovarian tissues. ELF5 protein expression in the cells of recombinant plasmid group was significantly higher compared with empty plasmid and blank control groups. The capacity of cell reproductive recombinant plasmid group at each time point decreased (P<0.05). Flow cytometry detection showed that 67.03% of cells in recombinant plasmid group was blocked in G0/G1 phase (P<0.05), compared with empty plasmid group (37.17%) and blank control group (38.24%). Apoptotic rate of recombinant plasmid group was significantly lower (31.4±1.9%; P<0.05), compared with that of empty plasmid group (9.1±2.2%) and blank control group (8.7±1.5%), and the differences were statistically significant. In conclusion, ELF5 interfered with cell cycle of human ovarian carcinoma SKOV3 cells and promoted apoptosis of human ovarian carcinoma SKOV3 cells inhibiting their growth and invasive capacity; and thus providing a new approach to gene treatment of ovarian carcinoma.

  17. Association between BRCA1 and BRCA2 mutations and survival in women with invasive epithelial ovarian cancer

    DEFF Research Database (Denmark)

    Bolton, Kelly L; Chenevix-Trench, Georgia; Goh, Cindy

    2012-01-01

    Approximately 10% of women with invasive epithelial ovarian cancer (EOC) carry deleterious germline mutations in BRCA1 or BRCA2. A recent article suggested that BRCA2-related EOC was associated with an improved prognosis, but the effect of BRCA1 remains unclear....

  18. In vivo imaging of the retinal pigment epithelial cells

    Science.gov (United States)

    Morgan, Jessica Ijams Wolfing

    The retinal pigment epithelial (RPE) cells form an important layer of the retina because they are responsible for providing metabolic support to the photoreceptors. Techniques to image the RPE layer include autofluorescence imaging with a scanning laser ophthalmoscope (SLO). However, previous studies were unable to resolve single RPE cells in vivo. This thesis describes the technique of combining autofluorescence, SLO, adaptive optics (AO), and dual-wavelength simultaneous imaging and registration to visualize the individual cells in the RPE mosaic in human and primate retina for the first time in vivo. After imaging the RPE mosaic non-invasively, the cell layer's structure and regularity were characterized using quantitative metrics of cell density, spacing, and nearest neighbor distances. The RPE mosaic was compared to the cone mosaic, and RPE imaging methods were confirmed using histology. The ability to image the RPE mosaic led to the discovery of a novel retinal change following light exposure; 568 nm exposures caused an immediate reduction in autofluorescence followed by either full recovery or permanent damage in the RPE layer. A safety study was conducted to determine the range of exposure irradiances that caused permanent damage or transient autofluorescence reductions. Additionally, the threshold exposure causing autofluorescence reduction was determined and reciprocity of radiant exposure was confirmed. Light exposures delivered by the AOSLO were not significantly different than those delivered by a uniform source. As all exposures tested were near or below the permissible light levels of safety standards, this thesis provides evidence that the current light safety standards need to be revised. Finally, with the retinal damage and autofluorescence reduction thresholds identified, the methods of RPE imaging were modified to allow successful imaging of the individual cells in the RPE mosaic while still ensuring retinal safety. This thesis has provided a

  19. Formation of reactive oxygen species in rat epithelial cells upon ...

    Indian Academy of Sciences (India)

    In our study, we investigated the influence of fly ash on the promotion of early inflammatory reactions like the formation of reactive oxygen species (ROS) in rat lung epithelial cells (RLE-6TN). Furthermore, we determined the formation of nitric oxide (NO). The cells show a clear dose-response relationship concerning the ...

  20. Sphingolipid trafficking and protein sorting in epithelial cells

    NARCIS (Netherlands)

    Slimane, TA; Hoekstra, D

    2002-01-01

    Sphingolipids represent a minor, but highly dynamic subclass of lipids in all eukaryotic cells. They are involved in functions that range from structural protection to signal transduction and protein sorting, and participate in lipid raft assembly. In polarized epithelial cells, which display an

  1. Prostaglandins in Cancer Cell Adhesion, Migration, and Invasion

    Directory of Open Access Journals (Sweden)

    David G. Menter

    2012-01-01

    Full Text Available Prostaglandins exert a profound influence over the adhesive, migratory, and invasive behavior of cells during the development and progression of cancer. Cyclooxygenase-2 (COX-2 and microsomal prostaglandin E2 synthase-1 (mPGES-1 are upregulated in inflammation and cancer. This results in the production of prostaglandin E2 (PGE2, which binds to and activates G-protein-coupled prostaglandin E1-4 receptors (EP1-4. Selectively targeting the COX-2/mPGES-1/PGE2/EP1-4 axis of the prostaglandin pathway can reduce the adhesion, migration, invasion, and angiogenesis. Once stimulated by prostaglandins, cadherin adhesive connections between epithelial or endothelial cells are lost. This enables cells to invade through the underlying basement membrane and extracellular matrix (ECM. Interactions with the ECM are mediated by cell surface integrins by “outside-in signaling” through Src and focal adhesion kinase (FAK and/or “inside-out signaling” through talins and kindlins. Combining the use of COX-2/mPGES-1/PGE2/EP1-4 axis-targeted molecules with those targeting cell surface adhesion receptors or their downstream signaling molecules may enhance cancer therapy.

  2. Risk of cervical intra-epithelial neoplasia and invasive cancer of the cervix in DES daughters

    NARCIS (Netherlands)

    H. Verloop (Herman); F.E. van Leeuwen (F.); T.J.M. Helmerhorst (Theo); I.M.C.M. de Kok (Inge); van Erp, E.J.M.; H.H. van Boven (Hester); M.A. Rookus (Matti)

    2017-01-01

    textabstractObjective: Women exposed to diethylstilbestrol in utero (DES) have an increased risk of clear cell adenocarcinoma (CCA) of the vagina and cervix, while their risk of non-CCA invasive cervical cancer is still unclear. Methods: We studied the risk of pre-cancerous (CIN) lesions and non-CCA

  3. Exosomes Promote Ovarian Cancer Cell Invasion through Transfer of CD44 to Peritoneal Mesothelial Cells.

    Science.gov (United States)

    Nakamura, Koji; Sawada, Kenjiro; Kinose, Yasuto; Yoshimura, Akihiko; Toda, Aska; Nakatsuka, Erika; Hashimoto, Kae; Mabuchi, Seiji; Morishige, Ken-Ichirou; Kurachi, Hirohisa; Lengyel, Ernst; Kimura, Tadashi

    2017-01-01

    Epithelial ovarian cancer (EOC) cells metastasize within the peritoneal cavity and directly encounter human peritoneal mesothelial cells (HPMC) as the initial step of metastasis. The contact between ovarian cancer cells and the single layer of mesothelial cells involves direct communications that modulate cancer progression but the mechanisms are unclear. One candidate mediating cell-cell communications is exosomes, 30-100 nm membrane vesicles of endocytic origin, through the cell-cell transfer of proteins, mRNAs, or microRNAs. Therefore, the goal was to mechanistically characterize how EOC-derived exosomes modulate metastasis. Exosomes from ovarian cancer cells were fluorescently labeled and cocultured with HPMCs which internalized the exosomes. Upon exosome uptake, HPMCs underwent a change in cellular morphology to a mesenchymal, spindle phenotype. CD44, a cell surface glycoprotein, was found to be enriched in the cancer cell-derived exosomes, transferred, and internalized to HPMCs, leading to high levels of CD44 in HPMCs. This increased CD44 expression in HPMCs promoted cancer invasion by inducing the HPMCs to secrete MMP9 and by cleaning the mesothelial barrier for improved cancer cell invasion. When CD44 expression was knocked down in cancer cells, exosomes had fewer effects on HPMCs. The inhibition of exosome release from cancer cells blocked CD44 internalization in HPMCs and suppressed ovarian cancer invasion. In ovarian cancer omental metastasis, positive CD44 expression was observed in those mesothelial cells that directly interacted with cancer cells, whereas CD44 expression was negative in the mesothelial cells remote from the invading edge. This study indicates that ovarian cancer-derived exosomes transfer CD44 to HPMCs, facilitating cancer invasion. Mechanistic insight from the current study suggests that therapeutic targeting of exosomes may be beneficial in treating ovarian cancer. Mol Cancer Res; 15(1); 78-92. ©2016 AACR. ©2016 American

  4. Protein Profiling of Isolated Leukocytes, Myofibroblasts, Epithelial, Basal, and Endothelial Cells from Normal, Hyperplastic, Cancerous, and Inflammatory Human Prostate Tissues

    Directory of Open Access Journals (Sweden)

    Zahraa I. Khamis, Kenneth A. Iczkowski, Ziad J. Sahab, Qing-Xiang Amy Sang

    2010-01-01

    Full Text Available In situ neoplastic prostate cells are not lethal unless they become invasive and metastatic. For cells to become invasive, the prostate gland must undergo degradation of the basement membrane and disruption of the basal cell layer underneath the luminal epithelia. Although the roles of proteinases in breaking down the basement membrane have been well-studied, little is known about the factors that induce basal cell layer disruption, degeneration, and its eventual disappearance in invasive cancer. It is hypothesized that microenvironmental factors may affect the degradation of the basal cell layer, which if protected may prevent tumor progression and invasion. In this study, we have revealed differential protein expression patterns between epithelial and stromal cells isolated from different prostate pathologies and identified several important epithelial and stromal proteins that may contribute to inflammation and malignant transformation of human benign prostate tissues to cancerous tissues using matrix-assisted laser desorption ionization time-of-flight mass spectrometry and proteomics methods. Cellular retinoic acid-binding protein 2 was downregulated in basal cells of benign prsotate. Caspase-1 and interleukin-18 receptor 1 were highly expressed in leukocytes of prostate cancer. Proto-oncogene Wnt-3 was downregulated in endothelial cells of prostatitis tissue and tyrosine phosphatase non receptor type 1 was only found in normal and benign endothelial cells. Poly ADP-ribose polymerase 14 was downregulated in myofibroblasts of prostatitis tissue. Interestingly, integrin alpha-6 was upregulated in epithelial cells but not detected in myofibroblasts of prostate cancer. Further validation of these proteins may generate new strategies for the prevention of basal cell layer disruption and subsequent cancer invasion.

  5. Proliferating cells in HIV and pamidronate-associated collapsing focal segmental glomerulosclerosis are parietal epithelial cells.

    NARCIS (Netherlands)

    Dijkman, H.B.P.M.; Weening, J.J.; Smeets, B.; Verrijp, K.; Kuppevelt, A.H.M.S.M. van; Assmann, K.K.; Steenbergen, E.; Wetzels, J.F.M.

    2006-01-01

    Collapsing focal segmental glomerulosclerosis (cFSGS) is characterized by hyperplasia of glomerular epithelial cells. In a mouse model of FSGS and in a patient with recurrent idiopathic FSGS, we identified the proliferating cells as parietal epithelial cells (PECs). In the present study, we have

  6. Proliferating cells in HIV and pamidronate-associated collapsing focal segmental glomerulosclerosis are parietal epithelial cells

    NARCIS (Netherlands)

    Dijkman, H. B. P. M.; Weening, J. J.; Smeets, B.; Verrijp, K. C. N.; van Kuppevelt, T. H.; Assmann, K. K. J. M.; Steenbergen, E. J.; Wetzels, J. F. M.

    2006-01-01

    Collapsing focal segmental glomerulosclerosis (cFSGS) is characterized by hyperplasia of glomerular epithelial cells. In a mouse model of FSGS and in a patient with recurrent idiopathic FSGS, we identified the proliferating cells as parietal epithelial cells (PECs). In the present study, we have

  7. Nicotine transport in lung and non-lung epithelial cells.

    Science.gov (United States)

    Takano, Mikihisa; Kamei, Hidetaka; Nagahiro, Machi; Kawami, Masashi; Yumoto, Ryoko

    2017-11-01

    Nicotine is rapidly absorbed from the lung alveoli into systemic circulation during cigarette smoking. However, mechanism underlying nicotine transport in alveolar epithelial cells is not well understood to date. In the present study, we characterized nicotine uptake in lung epithelial cell lines A549 and NCI-H441 and in non-lung epithelial cell lines HepG2 and MCF-7. Characteristics of [ 3 H]nicotine uptake was studied using these cell lines. Nicotine uptake in A549 cells occurred in a time- and temperature-dependent manner and showed saturation kinetics, with a Km value of 0.31mM. Treatment with some organic cations such as diphenhydramine and pyrilamine inhibited nicotine uptake, whereas treatment with organic cations such as carnitine and tetraethylammonium did not affect nicotine uptake. Extracellular pH markedly affected nicotine uptake, with high nicotine uptake being observed at high pH up to 11.0. Modulation of intracellular pH with ammonium chloride also affected nicotine uptake. Treatment with valinomycin, a potassium ionophore, did not significantly affect nicotine uptake, indicating that nicotine uptake is an electroneutral process. For comparison, we assessed the characteristics of nicotine uptake in another lung epithelial cell line NCI-H441 and in non-lung epithelial cell lines HepG2 and MCF-7. Interestingly, these cell lines showed similar characteristics of nicotine uptake with respect to pH dependency and inhibition by various organic cations. The present findings suggest that a similar or the same pH-dependent transport system is involved in nicotine uptake in these cell lines. A novel molecular mechanism of nicotine transport is proposed. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Oral epithelial cell reaction after exposure to Invisalign plastic material.

    Science.gov (United States)

    Premaraj, Thyagaseely; Simet, Samantha; Beatty, Mark; Premaraj, Sundaralingam

    2014-01-01

    Invisalign plastic aligners (Align Technology, Santa Clara, Calif) are used to correct malocclusions. The aligners wrap around the teeth and are in contact with gingival epithelium during treatment. The purpose of this study was to evaluate the cellular responses of oral epithelium exposed to Invisalign plastic in vitro. Oral epithelial cells were exposed to eluate obtained by soaking Invisalign plastic in either saline solution or artificial saliva for 2, 4, and 8 weeks. Cells grown in media containing saline solution or saliva served as controls. Morphologic changes were assessed by light microscopy. The 3-[4, 5-dimethythiazol- 2-yl]-2, 5-diphenyl tetrazolium bromide assay and flow cytometry were used to determine cell viability and membrane integrity, respectively. Cellular adhesion and micromotion of epithelial cells were measured in real time by electrical cell-substrate impedance sensing. Cells exposed to saline-solution eluate appeared rounded, were lifted from the culture plates, and demonstrated significantly increased metabolic inactivity or cell death (P <0.05). Saliva eluates did not induce significant changes in cell viability compared with untreated cells. Flow cytometry and electric cell-substrate impedance sensing showed that cells treated with saline-solution eluate exhibited compromised membrane integrity, and reduced cell-to-cell contact and mobility when compared with saliva-eluate treatment. Exposure to Invisalign plastic caused changes in viability, membrane permeability, and adhesion of epithelial cells in a saline-solution environment. Microleakage and hapten formation secondary to compromised epithelial integrity might lead to isocyanate allergy, which could be systemic or localized to gingiva. However, these results suggest that saliva might offer protection. Copyright © 2014 American Association of Orthodontists. Published by Mosby, Inc. All rights reserved.

  9. Campylobacter jejuni cocultured with epithelial cells reduces surface capsular polysaccharide expression.

    LENUS (Irish Health Repository)

    Corcionivoschi, N

    2012-02-01

    The host cell environment can alter bacterial pathogenicity. We employed a combination of cellular and molecular techniques to study the expression of Campylobacter jejuni polysaccharides cocultured with HCT-8 epithelial cells. After two passages, the amount of membrane-bound high-molecular-weight polysaccharide was considerably reduced. Microarray profiling confirmed significant downregulation of capsular polysaccharide (CPS) locus genes. Experiments using conditioned media showed that sugar depletion occurred only when the bacterial and epithelial cells were cocultured. CPS depletion occurred when C. jejuni organisms were exposed to conditioned media from a different C. jejuni strain but not when exposed to conditioned media from other bacterial species. Proteinase K or heat treatment of conditioned media under coculture conditions abrogated the effect on the sugars, as did formaldehyde fixation and cycloheximide treatment of host cells or chloramphenicol treatment of the bacteria. However, sugar depletion was not affected in flagellar export (fliQ) and quorum-sensing (luxS) gene mutants. Passaged C. jejuni showed reduced invasiveness and increased serum sensitivity in vitro. C. jejuni alters its surface polysaccharides when cocultured with epithelial cells, suggesting the existence of a cross talk mechanism that modulates CPS expression during infection.

  10. Proteomic analysis of protein interactions between Eimeria maxima sporozoites and chicken jejunal epithelial cells by shotgun LC-MS/MS.

    Science.gov (United States)

    Huang, Jingwei; Liu, Tingqi; Li, Ke; Song, Xiaokai; Yan, Ruofeng; Xu, Lixin; Li, Xiangrui

    2018-04-04

    Eimeria maxima initiates infection by invading the jejunal epithelial cells of chicken. However, the proteins involved in invasion remain unknown. The research of the molecules that participate in the interactions between E. maxima sporozoites and host target cells will fill a gap in our understanding of the invasion system of this parasitic pathogen. In the present study, chicken jejunal epithelial cells were isolated and cultured in vitro. Western blot was employed to analyze the soluble proteins of E. maxima sporozoites that bound to chicken jejunal epithelial cells. Co-immunoprecipitation (co-IP) assay was used to separate the E. maxima proteins that bound to chicken jejunal epithelial cells. Shotgun LC-MS/MS technique was used for proteomics identification and Gene Ontology was employed for the bioinformatics analysis. The results of Western blot analysis showed that four proteins bands from jejunal epithelial cells co-cultured with soluble proteins of E. maxima sporozoites were recognized by the positive sera, with molecular weights of 70, 90, 95 and 130 kDa. The co-IP dilutions were analyzed by shotgun LC-MS/MS. A total of 204 proteins were identified in the E. maxima protein database using the MASCOT search engine. Thirty-five proteins including microneme protein 3 and 7 had more than two unique peptide counts and were annotated using Gene Ontology for molecular function, biological process and cellular localization. The results revealed that of the 35 annotated peptides, 22 (62.86%) were associated with binding activity and 15 (42.86%) were involved in catalytic activity. Our findings provide an insight into the interaction between E. maxima and the corresponding host cells and it is important for the understanding of molecular mechanisms underlying E. maxima invasion.

  11. Embryonic chicken transplantation is a promising model for studying the invasive behaviour of melanoma cells.

    Directory of Open Access Journals (Sweden)

    Aparna eJayachandran

    2015-02-01

    Full Text Available Epithelial-to-mesenchymal transition is a hallmark event in the metastatic cascade conferring invasive ability to tumor cells. There are ongoing efforts to replicate the physiological events occurring during mobilization of tumor cells in model systems. However, few systems are able to capture these complex in vivo events. The embryonic chicken transplantation model has emerged as a useful system to assess melanoma cells including functions that are relevant to the metastatic process, namely invasion and plasticity. The chicken embryo represents an accessible and economical 3-dimensional in vivo model for investigating melanoma cell invasion as it exploits the ancestral relationship between melanoma and its precursor neural crest cells. We describe a methodology which enables the interrogation of melanoma cell motility within the developing avian embryo. This model involves the injection of melanoma cells into the neural tube of chicken embryos. Melanoma cells are labelled using fluorescent tracker dye, Vybrant DiO, then cultured as hanging drops for 24 hours to aggregate the cells. Groups of approximately 700 cells are placed into the neural tube of chicken embryos prior to the onset of neural crest migration at the hindbrain level (embryonic day 1.5 or trunk level (embryonic day 2.5. Chick embryos are reincubated and analysed after 48 hours for the location of melanoma cells using fluorescent microscopy on whole mounts and cross-sections of the embryos. Using this system, we compared the in vivo invasive behavior of epithelial-like and mesenchymal-like melanoma cells. We report that the developing embryonic microenvironment confers motile abilities to both types of melanoma cells. Hence the embryonic chicken transplantation model has potential to become a valuable tool for in vivo melanoma invasion studies. Importantly, it may provide novel insights into and reveal previously unknown mediators of the metastatic steps of invasion and

  12. Phenotype-dependent effects of EpCAM expression on growth and invasion of human breast cancer cell lines

    International Nuclear Information System (INIS)

    Martowicz, Agnieszka; Spizzo, Gilbert; Gastl, Guenther; Untergasser, Gerold

    2012-01-01

    The epithelial cell adhesion molecule (EpCAM) has been shown to be overexpressed in breast cancer and stem cells and has emerged as an attractive target for immunotherapy of breast cancer patients. This study analyzes the effects of EpCAM on breast cancer cell lines with epithelial or mesenchymal phenotype. For this purpose, shRNA-mediated knockdown of EpCAM gene expression was performed in EpCAM high breast cancer cell lines with epithelial phenotype (MCF-7, T47D and SkBR3). Moreover, EpCAM low breast carcinoma cell lines with mesenchymal phenotype (MDA-MB-231, Hs578t) and inducible overexpression of EpCAM were used to study effects on proliferation, migration and in vivo growth. In comparison to non-specific silencing controls (n/s-crtl) knockdown of EpCAM (E#2) in EpCAM high cell lines resulted in reduced cell proliferation under serum-reduced culture conditions. Moreover, DNA synthesis under 3D culture conditions in collagen was significantly reduced. Xenografts of MCF-7 and T47D cells with knockdown of EpCAM formed smaller tumors that were less invasive. EpCAM low cell lines with tetracycline-inducible overexpression of EpCAM showed no increased cell proliferation or migration under serum-reduced growth conditions. MDA-MB-231 xenografts with EpCAM overexpression showed reduced invasion into host tissue and more infiltrates of chicken granulocytes. The role of EpCAM in breast cancer strongly depends on the epithelial or mesenchymal phenotype of tumor cells. Cancer cells with epithelial phenotype need EpCAM as a growth- and invasion-promoting factor, whereas tumor cells with a mesenchymal phenotype are independent of EpCAM in invasion processes and tumor progression. These findings might have clinical implications for EpCAM-based targeting strategies in patients with invasive breast cancer

  13. [Frequency of oral squamous cell carcinoma and oral epithelial dysplasia in oral and oropharyngeal mucosa in Chile].

    Science.gov (United States)

    Martínez, Carolina; Hernández, Marcela; Martínez, Benjamín; Adorno, Daniela

    2016-02-01

    Oral cancer in Chile corresponds approximately to 1.6% of all cancer cases. There are few studies about oral epithelial dysplasia and oral squamous cell carcinoma in the Chilean population. To determine the frequency of hyperkeratosis, mild, moderate and severe oral epithelial dysplasia, in situ carcinoma and squamous cell carcinoma of the oral and oropharyngeal mucosa in a registry of the Oral Pathology Reference Institute of the Faculty of Dentistry, Universidad de Chile, in a ten years period. Review of clinical records and pathological plates of 389 patients, obtained between 1990 and 2009. Cases were selected according to their pathological diagnosis, including hyperkeratosis, oral epithelial dysplasia, in situ carcinoma, squamous cell carcinoma and verrucous carcinoma. Forty four percent of cases were squamous cell carcinoma, followed by hyperkeratosis in 37% and mild epithelial dysplasia in 11%. Squamous cell carcinoma was more common in men aged over 50 years. Most of the potentially malignant disorders presented clinically as leukoplakia and squamous cell carcinoma were clinically recognized as cancer. In this study, men aged over 50 years are the highest risk group for oral cancer. Early diagnosis is deficient since most of these lesions were diagnosed when squamous cell carcinoma became invasive. Leukoplakia diagnosis is mostly associated with hyperkeratosis and epithelial dysplasia, therefore biopsy of these lesions is mandatory to improve early diagnosis.

  14. Slug/SNAI2 regulates cell proliferation and invasiveness of metastatic prostate cancer cell lines.

    Science.gov (United States)

    Emadi Baygi, Modjtaba; Soheili, Zahra-Soheila; Essmann, Frank; Deezagi, Abdolkhaleg; Engers, Rainer; Goering, Wolfgang; Schulz, Wolfgang A

    2010-08-01

    Many metastatic cancers recapitulate the epithelial-to-mesenchymal transition (EMT) resulting in enhanced cell motility and invasiveness. The EMT is regulated by several transcription factors, including the zinc finger protein SNAI2, also named Slug, which appears to exert additional functions during development and cancer progression. We have studied the function of SNAI2 in prostate cancer cells. Quantitative RT-PCR analysis showed strong SNAI2 expression particularly in the PC-3 and PC3-16 prostate carcinoma cell lines. Knockdown of SNAI2 by specific siRNA induced changes in EMT markers and inhibited invasion of both cell lines into a matrigel matrix. SNAI2 siRNA-treated cells did not tolerate detachment from the culture plates, likely at least in part due to downregulation of integrin alpha6beta4. SNAI2 knockdown disturbed the microtubular and actin cytoskeletons, especially severely in PC-3 cells, resulting in grossly enlarged, flattened, and sometimes multinuclear cells. Knockdown also decreased cell proliferation, with a prominent G0/G1 arrest in PC3-16. Together, our data imply that SNAI2 exerts strong effects on the cytoskeleton and adhesion of those prostate cancer cells that express it and is necessary for their proliferation and invasiveness.

  15. The Regenerative Potential of Parietal Epithelial Cells in Adult Mice

    OpenAIRE

    Berger, Katja; Schulte, Kevin; Boor, Peter; Kuppe, Christoph; van Kuppevelt, Toin H.; Floege, Jürgen; Smeets, Bart; Moeller, Marcus J.

    2014-01-01

    Previously, we showed that some podocytes in juvenile mice are recruited from cells lining Bowman’s capsule, suggesting that parietal epithelial cells (PECs) are a progenitor cell population for podocytes. To investigate whether PECs also replenish podocytes in adult mice, PECs were genetically labeled in an irreversible fashion in 5-week-old mice. No significant increase in labeled podocytes was observed, even after 18 months. To accelerate a potential regenerative mechanism, progressive glo...

  16. Cancer Stem Cells and Epithelial-to-Mesenchymal Transition (EMT)-Phenotypic Cells: Are They Cousins or Twins?

    International Nuclear Information System (INIS)

    Kong, Dejuan; Li, Yiwei; Wang, Zhiwei; Sarkar, Fazlul H.

    2011-01-01

    Cancer stem cells (CSCs) are cells within a tumor that possess the capacity to self-renew and maintain tumor-initiating capacity through differentiation into the heterogeneous lineages of cancer cells that comprise the whole tumor. These tumor-initiating cells could provide a resource for cells that cause tumor recurrence after therapy. Although the cell origin of CSCs remains to be fully elucidated, mounting evidence has demonstrated that Epithelial-to-Mesenchymal Transition (EMT), induced by different factors, is associated with tumor aggressiveness and metastasis and these cells share molecular characteristics with CSCs, and thus are often called cancer stem-like cells or tumor-initiating cells. The acquisition of an EMT phenotype is a critical process for switching early stage carcinomas into invasive malignancies, which is often associated with the loss of epithelial differentiation and gain of mesenchymal phenotype. Recent studies have demonstrated that EMT plays a critical role not only in tumor metastasis but also in tumor recurrence and that it is tightly linked with the biology of cancer stem-like cells or cancer-initiating cells. Here we will succinctly summarize the state-of-our-knowledge regarding the molecular similarities between cancer stem-like cells or CSCs and EMT-phenotypic cells that are associated with tumor aggressiveness focusing on solid tumors

  17. Cancer Stem Cells and Epithelial-to-Mesenchymal Transition (EMT-Phenotypic Cells: Are They Cousins or Twins?

    Directory of Open Access Journals (Sweden)

    Fazlul H. Sarkar

    2011-02-01

    Full Text Available Cancer stem cells (CSCs are cells within a tumor that possess the capacity to self-renew and maintain tumor-initiating capacity through differentiation into the heterogeneous lineages of cancer cells that comprise the whole tumor. These tumor-initiating cells could provide a resource for cells that cause tumor recurrence after therapy. Although the cell origin of CSCs remains to be fully elucidated, mounting evidence has demonstrated that Epithelial-to-Mesenchymal Transition (EMT, induced by different factors, is associated with tumor aggressiveness and metastasis and these cells share molecular characteristics with CSCs, and thus are often called cancer stem-like cells or tumor-initiating cells. The acquisition of an EMT phenotype is a critical process for switching early stage carcinomas into invasive malignancies, which is often associated with the loss of epithelial differentiation and gain of mesenchymal phenotype. Recent studies have demonstrated that EMT plays a critical role not only in tumor metastasis but also in tumor recurrence and that it is tightly linked with the biology of cancer stem-like cells or cancer-initiating cells. Here we will succinctly summarize the state-of-our-knowledge regarding the molecular similarities between cancer stem-like cells or CSCs and EMT-phenotypic cells that are associated with tumor aggressiveness focusing on solid tumors.

  18. Suppressor of fused (Sufu) promotes epithelial-mesenchymal transition (EMT) in cervical squamous cell carcinoma

    Science.gov (United States)

    Zhang, Ziyu; Zou, Yang; Liang, Meirong; Chen, Yuanting; Luo, Yong; Yang, Bicheng; Liu, Faying; Qin, Yunna; He, Deming; Wang, Feng; Huang, Ouping

    2017-01-01

    Suppressor of fused is essential for the maximal activation of Sonic Hedgehog signaling in development and tumorigenesis. However, the role of Sufu in cervical carcinoma remains unknown. Here, we report new findings of Sufu in regulating the epithelial-to-mesenchymal transition through the FoxM1 transcriptional modulation by 14-3-3ζ protein in cervical carcinoma. Sufu is overexpressed in cervical squamous cell carcinoma and its level in clinical tumor tissues is positively correlated with 14-3-3ζ. Functionanlly, siSufu remarkably prevents the cancer cell migration and invasion. We further demonstrate that the transcriptional activity of Sufu is increased by FoxM1, of which stability is promoted by 14-3-3ζ. Knockdown FoxM1 decreases the invasion of SiHa cells and reconstitution of Sufu rescues the invasion of these cells.Finally, overexpression of Sufu is significantly associated with differentiation grade, FIGO stage, Depth of stromal invasion and vascular cancer embolus. Our findings highlight a novel role for Sufu in cervical carcinogenesis. PMID:29371981

  19. Cytotoxic effects of air freshener biocides in lung epithelial cells.

    Science.gov (United States)

    Kwon, Jung-Taek; Lee, Mimi; Seo, Gun-Baek; Kim, Hyun-Mi; Shim, Ilseob; Lee, Doo-Hee; Kim, Taksoo; Seo, Jung Kwan; Kim, Pilje; Choi, Kyunghee

    2013-09-01

    This study evaluated the cytotoxicity of mixtures of citral (CTR) and either benzisothiazolinone (BIT, Mix-CTR-BIT) or triclosan (TCS, Mix-CTR-TCS) in human A549 lung epithelial cells. We investigated the effects of various mix ratios of these common air freshener ingredients on cell viability, cell proliferation, reactive oxygen species (ROS) generation, and DNA damage. Mix-CTR-BIT and Mix-CTR-TCS significantly decreased the viability of lung epithelial cells and inhibited cell growth in a dose-dependent manner. In addition, both mixtures increased ROS generation, compared to that observed in control cells. In particular, cell viability, growth, and morphology were affected upon increase in the proportion of BIT or TCS in the mixture. However, comet analysis showed that treatment of cells with Mix-CTR-BIT or Mix-CTR-TCS did not increase DNA damage. Taken together, these data suggested that increasing the content of biocides in air fresheners might induce cytotoxicity, and that screening these compounds using lung epithelial cells may contribute to hazard assessment.

  20. Renal epithelial cells can release ATP by vesicular fusion

    Directory of Open Access Journals (Sweden)

    Randi G Bjaelde

    2013-09-01

    Full Text Available Renal epithelial cells have the ability to release nucleotides as paracrine factors. In the intercalated cells of the collecting duct, ATP is released by connexin30 (cx30, which is selectively expressed in this cell type. However, ATP is released by virtually all renal epithelia and the aim of the present study was to identify possible alternative nucleotide release pathways in a renal epithelial cell model. We used MDCK (type1 cells to screen for various potential ATP release pathways. In these cells, inhibition of the vesicular H+-ATPases (bafilomycin reduced both the spontaneous and hypotonically (80%-induced nucleotide release. Interference with vesicular fusion using N-ethylamide markedly reduced the spontaneous nucleotide release, as did interference with trafficking from the endoplasmic reticulum to the Golgi apparatus (brefeldin A1 and vesicular transport (nocodazole. These findings were substantiated using a siRNA directed against SNAP-23, which significantly reduced spontaneous ATP release. Inhibition of pannexin and connexins did not affect the spontaneous ATP release in this cell type, which consists of ∼90% principal cells. TIRF-microscopy of either fluorescently-labeled ATP (MANT-ATP or quinacrine-loaded vesicles, revealed that spontaneous release of single vesicles could be promoted by either hypoosmolality (50% or ionomycin. This vesicular release decreased the overall cellular fluorescence by 5.8% and 7.6% respectively. In summary, this study supports the notion that spontaneous and induced ATP release can occur via exocytosis in renal epithelial cells.

  1. Membrane dynamics and the regulation of epithelial cell polarity

    NARCIS (Netherlands)

    van der Wouden, JM; Maier, O; van IJzendoorn, SCD; Hoekstra, D

    2003-01-01

    Plasma membranes of epithelial cells consist of two domains, an apical and a basolateral domain, the surfaces of which differ in composition. The separation of these domains by a tight junction and the fact that specific transport pathways exist for intracellular communication between these domains

  2. Epithelial cell detachment by Porphyromonas gingivalis biofilm and planktonic cultures

    NARCIS (Netherlands)

    Huang, L.; van Loveren, C.; Ling, J.; Wei, X.; Crielaard, W.; Deng, D.M.

    2016-01-01

    Porphyromonas gingivalis is present as a biofilm at the sites of periodontal infections. The detachment of gingival epithelial cells induced by P. gingivalis biofilms was examined using planktonic cultures as a comparison. Exponentially grown planktonic cultures or 40-h biofilms were co-incubated

  3. Subtotal ablation of parietal epithelial cells induces crescent formation.

    NARCIS (Netherlands)

    Sicking, E.M.; Fuss, A.; Uhlig, S.; Jirak, P.; Dijkman, H.; Wetzels, J.; Engel, D.R.; Urzynicok, T.; Heidenreich, S.; Kriz, W.; Kurts, C.; Ostendorf, T.; Floege, J.; Smeets, B.; Moeller, M.J.

    2012-01-01

    Parietal epithelial cells (PECs) of the renal glomerulus contribute to the formation of both cellular crescents in rapidly progressive GN and sclerotic lesions in FSGS. Subtotal transgenic ablation of podocytes induces FSGS but the effect of specific ablation of PECs is unknown. Here, we established

  4. Parietal epithelial cells and podocytes in glomerular diseases

    NARCIS (Netherlands)

    Smeets, B.; Moeller, M.J.

    2012-01-01

    In recent years, it has become apparent that parietal epithelial cells (PECs) play an important role within the renal glomerulus, in particular in diseased conditions. In this review, we examine current knowledge about the role of PECs and their interactions with podocytes in development and under

  5. Metabolic cooperativity between epithelial cells and adipocytes of mice

    International Nuclear Information System (INIS)

    Bartley, J.C.; Emerman, J.T.; Bissell, M.J.

    1981-01-01

    We have demonstrated that glycogen and lipid synthesis in adipocytes is modulated by the lactational state and that this modulation in mammary adipocytes requires the presence of the adjacent epithelial cells. Glycogen and lipid synthesis from [ 14 C]glucose was measured in mammary fat pads cleared of epithelium, in abdominal fat pads, and in adipocytes from both sources and from intact mammary gland of mature virgin, pregnant, and lactating mice. Accumulation of glycogen, the activity of glycogen synthase, and the lipogenic rate in abdominal and mammary adipocytes remained high during pregnancy but decreased to insignificant levels by early lactation. The depressant effects of lactation were observed solely in those mammary adipocytes isolated from intact glands. The presence of mammary epithelial cells was also required to effect the stimulated lipogenesis in mammary adipocytes during pregnancy. We conclude that the metabolic activity of adipocytes is modulated both during pregnancy and lactation to channel nutrients to the mammary epithelial cell. The fact that the changes occur in mammary adipocytes only when epithelial cells are present indicates that local as well as systemic factors are operating in these modulations

  6. Parietal epithelial cells: their role in health and disease.

    Science.gov (United States)

    Romagnani, Paola

    2011-01-01

    Parietal epithelial cells of Bowman's capsules were first described by Sir William Bowman in 1842 in his paper On the Structure and Use of the Malpighian Bodies of the Kidney [London, Taylor, 1842], but since then their functions have remained poorly understood. A large body of evidence has recently suggested that parietal epithelial cells represent a reservoir of renal progenitors in adult human kidney which generate novel podocytes during childhood and adolescence, and can regenerate injured podocytes. The discovery that parietal epithelial cells represent a potential source for podocyte regeneration suggests that podocyte injury can be repaired. However, recent results also suggest that an abnormal proliferative response of renal progenitors to podocyte injury can generate hyperplastic glomerular lesions that are observed in crescentic glomerulonephritis and other types of glomerular disorders. Taken together, these results establish an entirely novel view that changes the way of thinking about renal physiology and pathophysiology, and suggest that understanding how self-renewal and fate decision of parietal epithelial cells in response to podocyte injury may be perturbed or modulated will be crucial for obtaining novel tools for prevention and treatment of glomerulosclerosis. Copyright © 2011 S. Karger AG, Basel.

  7. Host-pathogen interactions in bovine mammary epithelial cells and HeLa cells by Staphylococcus aureus isolated from subclinical bovine mastitis.

    Science.gov (United States)

    Castilho, Ivana G; Dantas, Stéfani Thais Alves; Langoni, Hélio; Araújo, João P; Fernandes, Ary; Alvarenga, Fernanda C L; Maia, Leandro; Cagnini, Didier Q; Rall, Vera L M

    2017-08-01

    Staphylococcus aureus is a common pathogen that causes subclinical bovine mastitis due to several virulence factors. In this study, we analyzed S. aureus isolates collected from the milk of cows with subclinical mastitis that had 8 possible combinations of bap, icaA, and icaD genes, to determine their capacity to produce biofilm on biotic (bovine primary mammary epithelial cells and HeLa cells) and abiotic (polystyrene microplates) surfaces, and their ability to adhere to and invade these cells. We also characterized isolates for microbial surface components recognizing adhesive matrix molecules (MSCRAMM) and agr genes, and for their susceptibility to cefquinome sulfate in the presence of biofilm. All isolates adhered to and invaded both cell types, but invasion indexes were higher in bovine primary mammary epithelial cells. Using tryptic soy broth + 1% glucose on abiotic surfaces, 5 out of 8 isolates were biofilm producers, but only the bap + icaA + icaD + isolate was positive in Dulbecco's Modified Eagle's medium. The production of biofilm on biotic surfaces occurred only with this isolate and only on HeLa cells, because the invasion index for bovine primary mammary epithelial cells was too high, making it impossible to use these cells in this assay. Of the 5 biofilm producers in tryptic soy broth + 1% glucose, 4 presented with the bap/fnbA/clfA/clfB/eno/fib/ebpS combination, and all were protected from cefquinome sulfate. We found no predominance of any agr group. The high invasive potential of S. aureus made it impossible to observe biofilm in bovine primary mammary epithelial cells, and we concluded that cells with lower invasion rates, such as HeLa cells, were more appropriate for this assay. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  8. Modeling Alveolar Epithelial Cell Behavior In Spatially Designed Hydrogel Microenvironments

    Science.gov (United States)

    Lewis, Katherine Jean Reeder

    The alveolar epithelium consists of two cell phenotypes, elongated alveolar type I cells (AT1) and rounded alveolar type II cells (ATII), and exists in a complex three-dimensional environment as a polarized cell layer attached to a thin basement membrane and enclosing a roughly spherical lumen. Closely surrounding the alveolar cysts are capillary endothelial cells as well as interstitial pulmonary fibroblasts. Many factors are thought to influence alveolar epithelial cell differentiation during lung development and wound repair, including physical and biochemical signals from the extracellular matrix (ECM), and paracrine signals from the surrounding mesenchyme. In particular, disrupted signaling between the alveolar epithelium and local fibroblasts has been implicated in the progression of several pulmonary diseases. However, given the complexity of alveolar tissue architecture and the multitude of signaling pathways involved, designing appropriate experimental platforms for this biological system has been difficult. In order to isolate key factors regulating cellular behavior, the researcher ideally should have control over biophysical properties of the ECM, as well as the ability to organize multiple cell types within the scaffold. This thesis aimed to develop a 3D synthetic hydrogel platform to control alveolar epithelial cyst formation, which could then be used to explore how extracellular cues influence cell behavior in a tissue-relevant cellular arrangement. To accomplish this, a poly(ethylene glycol) (PEG) hydrogel network containing enzymatically-degradable crosslinks and bioadhesive pendant peptides was employed as a base material for encapsulating primary alveolar epithelial cells. First, an array of microwells of various cross-sectional shapes was photopatterned into a PEG gel containing photo-labile crosslinks, and primary ATII cells were seeded into the wells to examine the role of geometric confinement on differentiation and multicellular arrangement

  9. Tumor budding cells, cancer stem cells and epithelial-mesenchymal transition-type cells in pancreatic cancer.

    Science.gov (United States)

    Karamitopoulou, Eva

    2012-01-01

    Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers with a 5-year survival rate of less than 5%. Moreover, PDAC escapes early detection and resists treatment. Multiple combinations of genetic alterations are known to occur in PDAC including mutational activation of KRAS, inactivation of p16/CDKN2A and SMAD4 (DPC4) and dysregulation of PTEN/PI3K/AKT signaling. Through their interaction with Wingless-INT pathway, the downstream molecules of these pathways have been implicated in the promotion of epithelial-mesenchymal transition (EMT). Emerging evidence has demonstrated that cancer stem cells (CSCs), small populations of which have been identified in PDAC, and EMT-type cells play critical roles in drug resistance, invasion, and metastasis in pancreatic cancer. EMT may be histologically represented by the presence of tumor budding which is described as the occurrence of single tumor cells or small clusters (<5) of dedifferentiated cells at the invasive front of gastrointestinal (including colorectal, oesophageal, gastric, and ampullary) carcinomas and is linked to poor prognosis. Tumor budding has recently been shown to occur frequently in PDAC and to be associated with adverse clinicopathological features and decreased disease-free and overall survival. The aim of this review is to present a short overview on the morphological and molecular aspects that underline the relationship between tumor budding cells, CSCs, and EMT-type cells in PDAC.

  10. Tumor Budding Cells, Cancer Stem Cells and Epithelial-Mesenchymal Transition-type Cells in Pancreatic Cancer

    Directory of Open Access Journals (Sweden)

    Eva eKaramitopoulou

    2013-01-01

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC is one of the most lethal cancers with a 5-year survival rate of less than 5%. Moreover, PDAC escapes early detection and resists treatment. Multiple combinations of genetic alterations are known to occur in PDAC including mutational activation of KRAS, inactivation of p16/CDKN2A and SMAD4 (DPC4 and dysregulation of PTEN/PI3K/AKT signaling. Through their interaction with WNT pathway, the downstream molecules of these pathways have been implicated in the promotion of epithelial-mesenchymal transition (EMT. Emerging evidence has demonstrated that cancer stem cells (CSCs, small populations of which have been identified in PDAC, and EMT-type cells play critical roles in drug resistance, invasion and metastasis in pancreatic cancer. EMT may be histologically represented by the presence of tumor budding which is described as the occurrence of single tumor cells or small clusters (<5 of dedifferentiated cells at the invasive front of gastrointestinal (including colorectal, oesophageal, gastric and ampullary carcinomas and is linked to poor prognosis. Tumor budding has recently been shown to occur frequently in PDAC and to be associated with adverse clinicopathological features and decreased disease-free and overall survival. The aim of this review is to present a short overview on the morphological and molecular aspects that underline the relationship between tumor budding cells, CSCs and EMT-type cells in PDAC.

  11. Amaranthus caudatus extract inhibits the invasion of E. coli into uroepithelial cells.

    Science.gov (United States)

    Mohanty, Soumitra; Zambrana, Silvia; Dieulouard, Soizic; Kamolvit, Witchuda; Nilsén, Vera; Gonzales, Eduardo; Östenson, Claes-Göran; Brauner, Annelie

    2018-06-28

    Amaranthus caudatus is traditionally used to treat infections. Based on its traditional usage, we investigated the effect of A. caudatus on the bladder epithelial cells in the protection of E. coli infection. The direct antimicrobial effects of A. caudatus on uropathogenic bacteria were investigated using minimum inhibitory concentration (MIC) assay. Bladder epithelial cell lines T24 and 5637 and uropathogenic E. coli strain #12 were used to investigate the effect of A. caudatus. Bacterial adhesion and invasion into bladder cells treated with A. caudatus was analyzed. Expression of uroplakin-1a (UPK1A), β1 integrin (ITGB1), caveolin-1 (CAV1) and the antimicrobial peptides human β defensin-2 (DEFB4A) and LL-37 (CAMP) was evaluated using RT-PCR. No direct antibacterial effect on E. coli or any of the tested uropathogenic strains was observed by A. caudatus. However, we demonstrated reduced mRNA expression of uroplakin-1a and caveolin-1, but not β1 integrin after treatment of uroepithelial cells, mirrored by the decreased adhesion and invasion of E. coli. A. caudatus treatment did not induce increased gene expression of the antimicrobial peptides, LL-37 and human β-defensin-2. Our results showed that A. caudatus has a protective role on bladder epithelial cells against uropathogenic E. coli infection by decreasing the bacterial adhesion and invasion, thereby preventing infection. Copyright © 2018 Elsevier B.V. All rights reserved.

  12. Nesfatin-1 inhibits ovarian epithelial carcinoma cell proliferation in vitro

    International Nuclear Information System (INIS)

    Xu, Yang; Pang, Xiaoyan; Dong, Mei; Wen, Fang; Zhang, Yi

    2013-01-01

    Highlights: •Nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest. •Nesfatin-1 enhances HO-8910 cell apoptosis. •Nesfatin-1 inhibits HO-8910 cell proliferation via mTOR and RhoA/ROCK signaling pathway. •The first report of nesfatin-1-mediated proliferation in ovarian epithelial carcinoma. -- Abstract: Nesfatin-1, an 82-amino-acid peptide derived from a 396-amino-acid precursor protein nucleobindin 2 (NUCB2), was originally identified in hypothalamic nuclei involved in the regulation of food intake. It was recently reported that nesfatin-1 is a novel depot specific adipokine preferentially produced by subcutaneous tissue, with obesity- and food deprivation-regulated expression. Although a relation between ovarian cancer mortality and obesity has been previously established, a role of nesfatin-1 in ovarian epithelial carcinoma remains unknown. The aim of the present study is to examine the effect of nesfatin-1 on ovary carcinoma cells proliferation. We found that nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest, this inhibition could be abolished by nesfatin-1 neutralizing antibody. Nesfatin-1 enhances HO-8910 cell apoptosis, activation of mammalian target of rapamycin (mTOR) and RhoA/ROCK signaling pathway block the effects of nesfatin-1-induced apoptosis, therefore reverses the inhibition of HO-8910 cell proliferation by nesfatin-1. In conclusion, the present study demonstrated that nesfatin-1 can inhibit the proliferation in human ovarian epithelial carcinoma cell line HO-8910 cells through inducing apoptosis via mTOR and RhoA/ROCK signaling pathway. This study provides a novel regulatory signaling pathway of nesfatin-1-regulated ovarian epithelial carcinoma growth and may contribute to ovarian cancer prevention and therapy, especially in obese patients

  13. Nesfatin-1 inhibits ovarian epithelial carcinoma cell proliferation in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Yang; Pang, Xiaoyan; Dong, Mei; Wen, Fang, E-mail: wenfang64@hotmail.com; Zhang, Yi, E-mail: syzi960@yahoo.com

    2013-11-01

    Highlights: •Nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest. •Nesfatin-1 enhances HO-8910 cell apoptosis. •Nesfatin-1 inhibits HO-8910 cell proliferation via mTOR and RhoA/ROCK signaling pathway. •The first report of nesfatin-1-mediated proliferation in ovarian epithelial carcinoma. -- Abstract: Nesfatin-1, an 82-amino-acid peptide derived from a 396-amino-acid precursor protein nucleobindin 2 (NUCB2), was originally identified in hypothalamic nuclei involved in the regulation of food intake. It was recently reported that nesfatin-1 is a novel depot specific adipokine preferentially produced by subcutaneous tissue, with obesity- and food deprivation-regulated expression. Although a relation between ovarian cancer mortality and obesity has been previously established, a role of nesfatin-1 in ovarian epithelial carcinoma remains unknown. The aim of the present study is to examine the effect of nesfatin-1 on ovary carcinoma cells proliferation. We found that nesfatin-1 inhibits the proliferation and growth of HO-8910 cells by G1 phase arrest, this inhibition could be abolished by nesfatin-1 neutralizing antibody. Nesfatin-1 enhances HO-8910 cell apoptosis, activation of mammalian target of rapamycin (mTOR) and RhoA/ROCK signaling pathway block the effects of nesfatin-1-induced apoptosis, therefore reverses the inhibition of HO-8910 cell proliferation by nesfatin-1. In conclusion, the present study demonstrated that nesfatin-1 can inhibit the proliferation in human ovarian epithelial carcinoma cell line HO-8910 cells through inducing apoptosis via mTOR and RhoA/ROCK signaling pathway. This study provides a novel regulatory signaling pathway of nesfatin-1-regulated ovarian epithelial carcinoma growth and may contribute to ovarian cancer prevention and therapy, especially in obese patients.

  14. Uranium induces oxidative stress in lung epithelial cells

    International Nuclear Information System (INIS)

    Periyakaruppan, Adaikkappan; Kumar, Felix; Sarkar, Shubhashish; Sharma, Chidananda S.; Ramesh, Govindarajan T.

    2007-01-01

    Uranium compounds are widely used in the nuclear fuel cycle, antitank weapons, tank armor, and also as a pigment to color ceramics and glass. Effective management of waste uranium compounds is necessary to prevent exposure to avoid adverse health effects on the population. Health risks associated with uranium exposure includes kidney disease and respiratory disorders. In addition, several published results have shown uranium or depleted uranium causes DNA damage, mutagenicity, cancer and neurological defects. In the current study, uranium toxicity was evaluated in rat lung epithelial cells. The study shows uranium induces significant oxidative stress in rat lung epithelial cells followed by concomitant decrease in the antioxidant potential of the cells. Treatment with uranium to rat lung epithelial cells also decreased cell proliferation after 72 h in culture. The decrease in cell proliferation was attributed to loss of total glutathione and superoxide dismutase in the presence of uranium. Thus the results indicate the ineffectiveness of antioxidant system's response to the oxidative stress induced by uranium in the cells. (orig.)

  15. The fate of epithelial cells in the human large intestine.

    Science.gov (United States)

    Barkla, D H; Gibson, P R

    1999-08-01

    One hundred and forty biopsies of the colon and rectum, collected during routine colonoscopies of 51 patients aged 19 to 74 years, were examined using light microscopy and transmission and scanning electron microscopy. The results indicated that surface epithelial cells undergo apoptosis, passing through fenestrations in the basement membrane to where they enter the lamina propria and are taken up by macrophages; and it is hypothesized that apoptotic cells are carried through the fenestrations on a current of fluid. The study also found that epithelial cells positioned over the crypts are better attached and more robust than those more distant from the crypt opening; and it is further hypothesized that, after reaching the top of the crypts, some goblet cells cease secreting mucus and pass onto the surface compartment of absorptive cells. An unexpected finding was that the lower regions of the crypts commonly contain isolated necrotic colonocytes. Apoptotic cells were rarely observed in the crypt epithelium. The findings of this study support the "recycling" model of epithelial cell death in the surface compartment of the human colon.

  16. Mesenchymal Stem Cells Induce Epithelial to Mesenchymal Transition in Colon Cancer Cells through Direct Cell-to-Cell Contact

    Directory of Open Access Journals (Sweden)

    Hidehiko Takigawa

    2017-05-01

    Full Text Available We previously reported that in an orthotopic nude mouse model of human colon cancer, bone marrow–derived mesenchymal stem cells (MSCs migrated to the tumor stroma and promoted tumor growth and metastasis. Here, we evaluated the proliferation and migration ability of cancer cells cocultured with MSCs to elucidate the mechanism of interaction between cancer cells and MSCs. Proliferation and migration of cancer cells increased following direct coculture with MSCs but not following indirect coculture. Thus, we hypothesized that direct contact between cancer cells and MSCs was important. We performed a microarray analysis of gene expression in KM12SM colon cancer cells directly cocultured with MSCs. Expression of epithelial-mesenchymal transition (EMT–related genes such as fibronectin (FN, SPARC, and galectin 1 was increased by direct coculture with MSCs. We also confirmed the upregulation of these genes with real-time polymerase chain reaction. Gene expression was not elevated in cancer cells indirectly cocultured with MSCs. Among the EMT-related genes upregulated by direct coculture with MSCs, we examined the immune localization of FN, a well-known EMT marker. In coculture assay in chamber slides, expression of FN was seen only at the edges of cancer clusters where cancer cells directly contacted MSCs. FN expression in cancer cells increased at the tumor periphery and invasive edge in orthotopic nude mouse tumors and human colon cancer tissues. These results suggest that MSCs induce EMT in colon cancer cells via direct cell-to-cell contact and may play an important role in colon cancer metastasis.

  17. HAb18G/CD147 is involved in TGF-β-induced epithelial-mesenchymal transition and hepatocellular carcinoma invasion.

    Science.gov (United States)

    Ru, Ning-Yu; Wu, Jiao; Chen, Zhi-Nan; Bian, Huijie

    2015-01-01

    Epithelial-mesenchymal transition (EMT) induced by the transforming growth factor beta (TGF-β) is involved in hepatocarcinogenesis and hepatocellular carcinoma (HCC) metastasis. HAb18G/CD147, a member of the immunoglobulin family, plays an important role in tumor invasion and metastasis. HAb18G/CD147 promotes EMT of hepatocytes through TGF-β signaling and is transcriptionally regulated by Slug. We investigated the role of HAb18G/CD147 in TGF-β-induced EMT in HCC invasion. Two human HCC cell lines, SMMC-7721 and HepG2, were used to determine the role of HAb18G/CD147 in EMT. Upregulation of HAb18G/CD147 induced by the high doses of TGF-β1 in SMMC-7721 (5 ng/mL) and HepG2 cells (10 ng/mL) (P CD147 upregulation was coupled with upregulation of Snail1 and Slug. CD147 knockout significantly decreased the expression of N-cadherin and vimentin, and colony formation ability of SMMC-7721 cells. TGF-β1 enhanced the migration capacity of SMMC-7721 cells, which was markedly attenuated by CD147 knockdown. Thus, HAb18G/CD147 is involved in TGF-β-induced EMT and HCC invasion. © 2014 International Federation for Cell Biology.

  18. Del-1 overexpression potentiates lung cancer cell proliferation and invasion

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Seung-Hwan; Kim, Dong-Young; Jing, Feifeng; Kim, Hyesoon [Department of Biomedical Sciences, University of Ulsan College of Medicine, Seoul (Korea, Republic of); Yun, Chae-Ok [Department of Bioengineering, College of Engineering, Hanyang University, Seoul (Korea, Republic of); Han, Deok-Jong [Department of Surgery, Asan Medical Center, University of Ulsan College of Medicine, Seoul (Korea, Republic of); Choi, Eun Young, E-mail: choieun@ulsan.ac.kr [Department of Biomedical Sciences, University of Ulsan College of Medicine, Seoul (Korea, Republic of)

    2015-12-04

    Developmental endothelial locus-1 (Del-1) is an endogenous anti-inflammatory molecule that is highly expressed in the lung and the brain and limits leukocyte migration to these tissues. We previously reported that the expression of Del-1 is positively regulated by p53 in lung endothelial cells. Although several reports have implicated the altered expression of Del-1 gene in cancer patients, little is known about its role in tumor cells. We here investigated the effect of Del-1 on the features of human lung carcinoma cells. Del-1 mRNA was found to be significantly decreased in the human lung adenocarcinoma cell lines A549 (containing wild type of p53), H1299 (null for p53) and EKVX (mutant p53), compared to in human normal lung epithelial BEAS-2B cells and MRC-5 fibroblasts. The decrease of Del-1 expression was dependent on the p53 activity in the cell lines, but not on the expression of p53. Neither treatment with recombinant human Del-1 protein nor the introduction of adenovirus expressing Del-1 altered the expression of the apoptosis regulators BAX, PUMA and Bcl-2. Unexpectedly, the adenovirus-mediated overexpression of Del-1 gene into the lung carcinoma cell lines promoted proliferation and invasion of the lung carcinoma cells, as revealed by BrdU incorporation and transwell invasion assays, respectively. In addition, overexpression of the Del-1 gene enhanced features of epithelial–mesenchymal transition (EMT), such as increasing vimentin while decreasing E-cadherin in A549 cells, and increases in the level of Slug, an EMT-associated transcription regulator. Our findings demonstrated for the first time that there are deleterious effects of high levels of Del-1 in lung carcinoma cells, and suggest that Del-1 may be used as a diagnostic or prognostic marker for cancer progression, and as a novel therapeutic target for lung carcinoma. - Highlights: • Developmental Endothelial Locus-1 (Del-1) expression is downregulated in human lung cancer cells.

  19. Del-1 overexpression potentiates lung cancer cell proliferation and invasion

    International Nuclear Information System (INIS)

    Lee, Seung-Hwan; Kim, Dong-Young; Jing, Feifeng; Kim, Hyesoon; Yun, Chae-Ok; Han, Deok-Jong; Choi, Eun Young

    2015-01-01

    Developmental endothelial locus-1 (Del-1) is an endogenous anti-inflammatory molecule that is highly expressed in the lung and the brain and limits leukocyte migration to these tissues. We previously reported that the expression of Del-1 is positively regulated by p53 in lung endothelial cells. Although several reports have implicated the altered expression of Del-1 gene in cancer patients, little is known about its role in tumor cells. We here investigated the effect of Del-1 on the features of human lung carcinoma cells. Del-1 mRNA was found to be significantly decreased in the human lung adenocarcinoma cell lines A549 (containing wild type of p53), H1299 (null for p53) and EKVX (mutant p53), compared to in human normal lung epithelial BEAS-2B cells and MRC-5 fibroblasts. The decrease of Del-1 expression was dependent on the p53 activity in the cell lines, but not on the expression of p53. Neither treatment with recombinant human Del-1 protein nor the introduction of adenovirus expressing Del-1 altered the expression of the apoptosis regulators BAX, PUMA and Bcl-2. Unexpectedly, the adenovirus-mediated overexpression of Del-1 gene into the lung carcinoma cell lines promoted proliferation and invasion of the lung carcinoma cells, as revealed by BrdU incorporation and transwell invasion assays, respectively. In addition, overexpression of the Del-1 gene enhanced features of epithelial–mesenchymal transition (EMT), such as increasing vimentin while decreasing E-cadherin in A549 cells, and increases in the level of Slug, an EMT-associated transcription regulator. Our findings demonstrated for the first time that there are deleterious effects of high levels of Del-1 in lung carcinoma cells, and suggest that Del-1 may be used as a diagnostic or prognostic marker for cancer progression, and as a novel therapeutic target for lung carcinoma. - Highlights: • Developmental Endothelial Locus-1 (Del-1) expression is downregulated in human lung cancer cells.

  20. Glomerular parietal epithelial cells in kidney physiology, pathology, and repair

    OpenAIRE

    Shankland, Stuart J.; Anders, Hans-Joachim; Romagnani, Paola

    2013-01-01

    Purpose of review We have summarized recently published glomerular parietal epithelial cell (PEC) research, focusing on their roles in glomerular development and physiology, and in certain glomerular diseases. The rationale is that PECs have been largely ignored until the recent availability of cell lineage tracing studies, human and murine PEC culture systems, and potential therapeutic interventions of PECs. Recent findings Several new paradigms involving PECs have emerged demonstrating thei...

  1. Intestinal epithelial cell-specific RARα depletion results in aberrant epithelial cell homeostasis and underdeveloped immune system.

    Science.gov (United States)

    Jijon, H B; Suarez-Lopez, L; Diaz, O E; Das, S; De Calisto, J; Yaffe, M B; Pittet, M J; Mora, J R; Belkaid, Y; Xavier, R J; Villablanca, E J

    2018-05-01

    Retinoic acid (RA), a dietary vitamin A metabolite, is crucial in maintaining intestinal homeostasis. RA acts on intestinal leukocytes to modulate their lineage commitment and function. Although the role of RA has been characterized in immune cells, whether intestinal epithelial cells (IECs) rely on RA signaling to exert their immune-regulatory function has not been examined. Here we demonstrate that lack of RA receptor α (RARα) signaling in IECs results in deregulated epithelial lineage specification, leading to increased numbers of goblet cells and Paneth cells. Mechanistically, lack of RARα resulted in increased KLF4 + goblet cell precursors in the distal bowel, whereas RA treatment inhibited klf4 expression and goblet cell differentiation in zebrafish. These changes in secretory cells are associated with increased Reg3g, reduced luminal bacterial detection, and an underdeveloped intestinal immune system, as evidenced by an almost complete absence of lymphoid follicles and gut resident mononuclear phagocytes. This underdeveloped intestinal immune system shows a decreased ability to clear infection with Citrobacter rodentium. Collectively, our findings indicate that epithelial cell-intrinsic RARα signaling is critical to the global development of the intestinal immune system.

  2. Acrolein stimulates eicosanoid release from bovine airway epithelial cells

    International Nuclear Information System (INIS)

    Doupnik, C.A.; Leikauf, G.D.

    1990-01-01

    Injury to the airway mucosa after exposure to environmental irritants is associated with pulmonary inflammation and bronchial hyperresponsiveness. To better understand the relationships between mediator release and airway epithelial cell injury during irritant exposures, we studied the effects of acrolein, a low-molecular-weight aldehyde found in cigarette smoke, on arachidonic acid metabolism in cultured bovine tracheal epithelial cells. Confluent airway epithelial cell monolayers, prelabeled with [3H]arachidonic acid, released significant levels of 3H activity when exposed (20 min) to 100 microM acrolein. [3H]arachidonic acid products were resolved using reverse-phase high-performance liquid chromatography. Under control conditions the released 3H activity coeluted predominantly with the cyclooxygenase product, prostaglandin (PG) E2. After exposure to acrolein, significant peaks in 3H activity coeluted with the lipoxygenase products 12-hydroxyeicosatetraenoic acid (HETE) and 15-HETE, as well as with PGE2, PGF2 alpha, and 6-keto-PGF1 alpha. Dose-response relationships for acrolein-induced release of immunoreactive PGF2 alpha and PGE2 from unlabeled epithelial monolayers demonstrated 30 microM acrolein as the threshold dose, with 100 microM acrolein inducing nearly a fivefold increase in both PGF2 alpha and PGE2. Cellular viability after exposure to 100 microM acrolein, determined by released lactate dehydrogenase activity, was not affected until exposure periods were greater than or equal to 2 h. These results implicate the airway epithelial cell as a possible source of eicosanoids after exposure to acrolein

  3. Distinct effects of EGFR inhibitors on epithelial- and mesenchymal-like esophageal squamous cell carcinoma cells.

    Science.gov (United States)

    Yoshioka, Masahiro; Ohashi, Shinya; Ida, Tomomi; Nakai, Yukie; Kikuchi, Osamu; Amanuma, Yusuke; Matsubara, Junichi; Yamada, Atsushi; Miyamoto, Shin'ichi; Natsuizaka, Mitsuteru; Nakagawa, Hiroshi; Chiba, Tsutomu; Seno, Hiroshi; Muto, Manabu

    2017-08-01

    Epidermal growth factor receptor (EGFR) plays a pivotal role in the pathophysiology of esophageal squamous cell carcinoma (ESCC). However, the clinical effects of EGFR inhibitors on ESCC are controversial. This study sought to identify the factors determining the therapeutic efficacy of EGFR inhibitors in ESCC cells. Immortalized-human esophageal epithelial cells (EPC2-hTERT), transformed-human esophageal epithelial cells (T-Epi and T-Mes), and ESCC cells (TE-1, TE-5, TE-8, TE-11, TE-11R, and HCE4) were treated with the EGFR inhibitors erlotinib or cetuximab. Inhibitory effects on cell growth were assessed by cell counting or cell-cycle analysis. The expression levels of genes and proteins such as involucrin and cytokeratin13 (a squamous differentiation marker), E-cadherin, and vimentin were evaluated by real-time polymerase chain reaction or western blotting. To examine whether mesenchymal phenotype influenced the effects of EGFR inhibitors, we treated T-Epi cells with TGF-β1 to establish a mesenchymal phenotype (mesenchymal T-Epi cells). We then compared the effects of EGFR inhibitors on parental T-Epi cells and mesenchymal T-Epi cells. TE-8 (mesenchymal-like ESCC cells)- or TE-11R (epithelial-like ESCC cells)-derived xenograft tumors in mice were treated with cetuximab, and the antitumor effects of EGFR inhibitors were evaluated. Cells were classified as epithelial-like or mesenchymal-like phenotypes, determined by the expression levels of E-cadherin and vimentin. Both erlotinib and cetuximab reduced cell growth and the ratio of cells in cell-cycle S phase in epithelial-like but not mesenchymal-like cells. Additionally, EGFR inhibitors induced squamous cell differentiation (defined as increased expression of involucrin and cytokeratin13) in epithelial-like but not mesenchymal-like cells. We found that EGFR inhibitors did not suppress the phosphorylation of EGFR in mesenchymal-like cells, while EGFR dephosphorylation was observed after treatment with EGFR

  4. The Crosstalk between Nrf2 and TGF-β1 in the Epithelial-Mesenchymal Transition of Pancreatic Duct Epithelial Cells.

    Directory of Open Access Journals (Sweden)

    Sarah Arfmann-Knübel

    Full Text Available Nrf2 and TGF-β1 both affect tumorigenesis in a dual fashion, either by preventing carcinogen induced carcinogenesis and suppressing tumor growth, respectively, or by conferring cytoprotection and invasiveness to tumor cells during malignant transformation. Given the involvement of Nrf2 and TGF-β1 in the adaptation of epithelial cells to persistent inflammatory stress, e.g. of the pancreatic duct epithelium during chronic pancreatitis, a crosstalk between Nrf2 and TGF-β1 can be envisaged. By using premalignant human pancreatic duct cells (HPDE and the pancreatic ductal adenocarcinoma cell line Colo357, we could show that Nrf2 and TGF-β1 independently but additively conferred an invasive phenotype to HPDE cells, whereas acting synergistically in Colo357 cells. This was accompanied by differential regulation of EMT markers like vimentin, Slug, L1CAM and E-cadherin. Nrf2 activation suppressed E-cadherin expression through an as yet unidentified ARE related site in the E-cadherin promoter, attenuated TGF-β1 induced Smad2/3-activity and enhanced JNK-signaling. In Colo357 cells, TGF-β1 itself was capable of inducing Nrf2 whereas in HPDE cells TGF-β1 per-se did not affect Nrf2 activity, but enhanced Nrf2 induction by tBHQ. In Colo357, but not in HPDE cells, the effects of TGF-β1 on invasion were sensitive to Nrf2 knock-down. In both cell lines, E-cadherin re-expression inhibited the proinvasive effect of Nrf2. Thus, the increased invasion of both cell lines relates to the Nrf2-dependent downregulation of E-cadherin expression. In line, immunohistochemistry analysis of human pancreatic intraepithelial neoplasias in pancreatic tissues from chronic pancreatitis patients revealed strong Nrf2 activity already in premalignant epithelial duct cells, accompanied by partial loss of E-cadherin expression. Our findings indicate that Nrf2 and TGF-β1 both contribute to malignant transformation through distinct EMT related mechanisms accounting for an

  5. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M

    1998-01-01

    human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...... induced apoptosis in several activated T-cell populations and T-cell lines, including T-cell antigen receptor (TCR)-CD3-negative T-cell lines. In contrast, RPE cells induced little or no apoptosis in resting peripheral T cells. Major histocompatibility complex (MHC) class II monoclonal antibodies, which...

  6. Analysis of interactions of Salmonella type three secretion mutants with 3-D intestinal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Andrea L Radtke

    Full Text Available The prevailing paradigm of Salmonella enteropathogenesis based on monolayers asserts that Salmonella pathogenicity island-1 Type Three Secretion System (SPI-1 T3SS is required for bacterial invasion into intestinal epithelium. However, little is known about the role of SPI-1 in mediating gastrointestinal disease in humans. Recently, SPI-1 deficient nontyphoidal Salmonella strains were isolated from infected humans and animals, indicating that SPI-1 is not required to cause enteropathogenesis and demonstrating the need for more in vivo-like models. Here, we utilized a previously characterized 3-D organotypic model of human intestinal epithelium to elucidate the role of all characterized Salmonella enterica T3SSs. Similar to in vivo reports, the Salmonella SPI-1 T3SS was not required to invade 3-D intestinal cells. Additionally, Salmonella strains carrying single (SPI-1 or SPI-2, double (SPI-1/2 and complete T3SS knockout (SPI-1/SPI-2: flhDC also invaded 3-D intestinal cells to wildtype levels. Invasion of wildtype and TTSS mutants was a Salmonella active process, whereas non-invasive bacterial strains, bacterial size beads, and heat-killed Salmonella did not invade 3-D cells. Wildtype and T3SS mutants did not preferentially target different cell types identified within the 3-D intestinal aggregates, including M-cells/M-like cells, enterocytes, or Paneth cells. Moreover, each T3SS was necessary for substantial intracellular bacterial replication within 3-D cells. Collectively, these results indicate that T3SSs are dispensable for Salmonella invasion into highly differentiated 3-D models of human intestinal epithelial cells, but are required for intracellular bacterial growth, paralleling in vivo infection observations and demonstrating the utility of these models in predicting in vivo-like pathogenic mechanisms.

  7. Generation of stratified squamous epithelial progenitor cells from mouse induced pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Satoru Yoshida

    Full Text Available BACKGROUND: Application of induced pluripotent stem (iPS cells in regenerative medicine will bypass ethical issues associated with use of embryonic stem cells. In addition, patient-specific IPS cells can be useful to elucidate the pathophysiology of genetic disorders, drug screening, and tailor-made medicine. However, in order to apply iPS cells to mitotic tissue, induction of tissue stem cells that give rise to progeny of the target organ is required. METHODOLOGY/PRINCIPAL FINDINGS: We induced stratified epithelial cells from mouse iPS cells by co-culture with PA6 feeder cells (SDIA-method with use of BMP4. Clusters of cells positive for the differentiation markers KRT1 or KRT12 were observed in KRT14-positive colonies. We successfully cloned KRT14 and p63 double-positive stratified epithelial progenitor cells from iPS-derived epithelial cells, which formed stratified epithelial sheets consisting of five- to six-polarized epithelial cells in vitro. When these clonal cells were cultured on denuded mouse corneas, a robust stratified epithelial layer was observed with physiological cell polarity including high levels of E-cadherin, p63 and K15 expression in the basal layer and ZO-1 in the superficial layer, recapitulating the apico-basal polarity of the epithelium in vivo. CONCLUSIONS/SIGNIFICANCE: These results suggest that KRT14 and p63 double-positive epithelial progenitor cells can be cloned from iPS cells in order to produce polarized multilayer epithelial cell sheets.

  8. Fra-1 induces morphological transformation and increases in vitro invasiveness and motility of epithelioid adenocarcinoma cells

    DEFF Research Database (Denmark)

    Kustikova, O.; Kramerov, D.; Grigorian, M.

    1998-01-01

    in vitro nor in vivo. CSML100 possesses all characteristics of a highly progressive carcinoma. These cells do not form tight contacts, are highly invasive in vitro, and are metastatic in vivo. AP-1 activity was considerably higher in CSML100 cells than in CSML0 cells. There was a common predominant Jun...... from tumors of epithelial origin revealed a correlation of fra-1 expression with mesenchymal characteristics of carcinoma cells. Moreover, we show here for the first time that the expression of exogenous Fra-1 in epithelioid cells results in morphological changes that resemble fibroblastoid conversion...

  9. Trichomonas vaginalis perturbs the junctional complex in epithelial cells

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    Trichomonas vaginalis, a protist parasite of the urogenital tract in humans, is the causative agent of trichomonosis,which in recent years have been associated with the cervical cancer development. In the present study we analyzed the modifications at the junctional complex level of Caco-2 cells after interaction with two isolates of T. vaginalis and the influence of the iron concentration present in the parasite's culture medium on the interaction effects. Our results show that T. vaginalis adheres to the epithelial cell causing alterations in the junctional complex, such as: (a) a decrease in transepithelial electrical resistance; (b) alteration in the pattern of junctional complex proteins distribution as obseryed for E-cadherin, occludin and ZO-1; and (c) enlargement of the spaces between epithelial cells. These effects were dependent on (a) the degree of the parasite virulence isolate, (b) the iron concentration in the culture medium, and (c) the expression of adhesin proteins on the parasite surface.

  10. Human airway xenograft models of epithelial cell regeneration

    Directory of Open Access Journals (Sweden)

    Puchelle Edith

    2000-10-01

    Full Text Available Abstract Regeneration and restoration of the airway epithelium after mechanical, viral or bacterial injury have a determinant role in the evolution of numerous respiratory diseases such as chronic bronchitis, asthma and cystic fibrosis. The study in vivo of epithelial regeneration in animal models has shown that airway epithelial cells are able to dedifferentiate, spread, migrate over the denuded basement membrane and progressively redifferentiate to restore a functional respiratory epithelium after several weeks. Recently, human tracheal xenografts have been developed in immunodeficient severe combined immunodeficiency (SCID and nude mice. In this review we recall that human airway cells implanted in such conditioned host grafts can regenerate a well-differentiated and functional human epithelium; we stress the interest in these humanized mice in assaying candidate progenitor and stem cells of the human airway mucosa.

  11. Cigarette smoke alters the secretome of lung epithelial cells.

    Science.gov (United States)

    Mossina, Alessandra; Lukas, Christina; Merl-Pham, Juliane; Uhl, Franziska E; Mutze, Kathrin; Schamberger, Andrea; Staab-Weijnitz, Claudia; Jia, Jie; Yildirim, Ali Ö; Königshoff, Melanie; Hauck, Stefanie M; Eickelberg, Oliver; Meiners, Silke

    2017-01-01

    Cigarette smoke is the most relevant risk factor for the development of lung cancer and chronic obstructive pulmonary disease. Many of its more than 4500 chemicals are highly reactive, thereby altering protein structure and function. Here, we used subcellular fractionation coupled to label-free quantitative MS to globally assess alterations in the proteome of different compartments of lung epithelial cells upon exposure to cigarette smoke extract. Proteomic profiling of the human alveolar derived cell line A549 revealed the most pronounced changes within the cellular secretome with preferential downregulation of proteins involved in wound healing and extracellular matrix organization. In particular, secretion of secreted protein acidic and rich in cysteine, a matricellular protein that functions in tissue response to injury, was consistently diminished by cigarette smoke extract in various pulmonary epithelial cell lines and primary cells of human and mouse origin as well as in mouse ex vivo lung tissue cultures. Our study reveals a previously unrecognized acute response of lung epithelial cells to cigarette smoke that includes altered secretion of proteins involved in extracellular matrix organization and wound healing. This may contribute to sustained alterations in tissue remodeling as observed in lung cancer and chronic obstructive pulmonary disease. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Wnt Signalling in Gastrointestinal Epithelial Stem Cells

    Directory of Open Access Journals (Sweden)

    Dustin J. Flanagan

    2018-03-01

    Full Text Available Wnt signalling regulates several cellular functions including proliferation, differentiation, apoptosis and migration, and is critical for embryonic development. Stem cells are defined by their ability for self-renewal and the ability to be able to give rise to differentiated progeny. Consequently, they are essential for the homeostasis of many organs including the gastrointestinal tract. This review will describe the huge advances in our understanding of how stem cell functions in the gastrointestinal tract are regulated by Wnt signalling, including how deregulated Wnt signalling can hijack these functions to transform cells and lead to cancer.

  13. Epithelial rotation is preceded by planar symmetry breaking of actomyosin and protects epithelial tissue from cell deformations.

    Science.gov (United States)

    Viktorinová, Ivana; Henry, Ian; Tomancak, Pavel

    2017-11-01

    Symmetry breaking is involved in many developmental processes that form bodies and organs. One of them is the epithelial rotation of developing tubular and acinar organs. However, how epithelial cells move, how they break symmetry to define their common direction, and what function rotational epithelial motions have remains elusive. Here, we identify a dynamic actomyosin network that breaks symmetry at the basal surface of the Drosophila follicle epithelium of acinar-like primitive organs, called egg chambers, and may represent a candidate force-generation mechanism that underlies the unidirectional motion of this epithelial tissue. We provide evidence that the atypical cadherin Fat2, a key planar cell polarity regulator in Drosophila oogenesis, directs and orchestrates transmission of the intracellular actomyosin asymmetry cue onto a tissue plane in order to break planar actomyosin symmetry, facilitate epithelial rotation in the opposite direction, and direct the elongation of follicle cells. In contrast, loss of this rotational motion results in anisotropic non-muscle Myosin II pulses that are disorganized in plane and causes cell deformations in the epithelial tissue of Drosophila eggs. Our work demonstrates that atypical cadherins play an important role in the control of symmetry breaking of cellular mechanics in order to facilitate tissue motion and model epithelial tissue. We propose that their functions may be evolutionarily conserved in tubular/acinar vertebrate organs.

  14. Destructive impact of t-lymphocytes, NK and mast cells on basal cell layers: implications for tumor invasion

    International Nuclear Information System (INIS)

    Yuan, Hongyan; Hsiao, Yi-Hsuan; Zhang, Yiyu; Wang, Jinlian; Yin, Chao; Shen, Rong; Su, Yiping

    2013-01-01

    Our previous studies have suggested that the primary impact of immune cell infiltration into the normal or pre-invasive tissue component is associated with the physical destruction of epithelial capsules, which may promote tumor progression and invasion. Our current study attempted to further verify our previous observations and determine the primary type(s) of infiltrating immune cells and the possible mechanism associated with physical destructions of the epithelial capsules. In total, the study was conducted with 250 primary breast and prostate tumors, the primary immune cell of cytotoxic T-lymphocytes (CTL), Natural killer cells (NK) and Mast cells were analyzed by immunohistochemistry, fluorescent labeling and apoptosis assay. qRT-PCR was used for gene expression analysis. Our current study assessed the physical disruption of these immune cells and potential impact on the epithelial capsule of human breast and prostate tumors. Our study yield several clinically-relevant findings that have not been studied before. (1) A vast majority of these infiltrating immune cells are distributed in the normal-appearing or pre-invasive tissue components rather than in invasive cancer tissues. (2) These cells often form rings or semilunar structures that either surround focally-disrupted basal cell layers or physically attach to the basal cells. (3) Basal cells physically associated with these immune cells generally displayed distinct signs of degeneration, including substantially elevated apoptosis, necrosis, and reduced tumor suppressor p63 expression. In contrast, luminal cells overlying focally disrupted basal cell layers had a substantially increased proliferation rate and elevated expression of stem cell markers compared to their adjacent morphologically similar counterparts that overlie a non-disrupted capsule. Our findings suggest that at the early stage of tumor invasion, CTL, NK and Mast cells are the main types of tumor infiltrating immune cells involved in focal

  15. Osmoregulation of chloride channels in epithelial cells

    NARCIS (Netherlands)

    C.H. Lim (Christina)

    2008-01-01

    markdownabstract__Abstract__ The plasma membrane of mammalian cells is formed by two layers of lipids (lipid bilayer), primarily phospholipids, glycolipids and cholesterol, in which many different proteins are embedded. Phospholipid consists of a glycerol backbone esterified to fatty acids

  16. microRNA-494 is a potential prognostic marker and inhibits cellular proliferation, migration and invasion by targeting SIRT1 in epithelial ovarian cancer.

    Science.gov (United States)

    Yang, Aijun; Wang, Xuenan; Yu, Chunna; Jin, Zhenzhen; Wei, Lingxia; Cao, Jinghe; Wang, Qin; Zhang, Min; Zhang, Lin; Zhang, Lei; Hao, Cuifang

    2017-09-01

    Ovarian cancer is one of the most common types of gynecological malignancy worldwide, and is the fourth leading cause of cancer-associated mortality among women. Despite improvements in therapeutic treatments, the prognosis for epithelial ovarian cancer (EOC) remains poor, mainly due to the rapid growth and metastasis of ovarian cancer tumors. An increasing number of studies have indicated that microRNAs (miRNAs) are involved in the carcinogenesis and progression of human cancer, suggesting that miRNAs may be used in clinical prognosis and as a therapeutic target in EOC. The aim of the present study was to investigate the expression levels of miRNA-494 in EOC tissues and cell lines. The clinical significance of miRNA-494 in patients with EOC was also evaluated. The results demonstrated that miRNA-494 was significantly downregulated in EOC tissues and cell lines. Low expression levels of miRNA-494 were associated with poor prognostic features, including International Federation of Gynecology and Obstetrics stage, tumor size and lymph node metastasis. In vitro functional studies demonstrated that overexpression of miRNA-494 inhibited proliferation, migration and invasion in EOC cells. By contrast, knockdown of miRNA-494 enhanced cell growth, migration and invasion in EOC cells. Notably, sirtuin 1 (SIRT1) was identified as a direct target of miRNA-494 in EOC. Furthermore, MTT, cell migration and invasion assays verified that EOC cell proliferation, migration and invasion were completely restored with forced miRNA-494 expression and SIRT1 restoration. Together, these findings suggest that miRNA-494 is a potential prognostic marker, and may provide novel therapeutic regimens of targeted therapy for EOC.

  17. Epithelial-Mesenchymal Transition in Kidney Tubular Epithelial Cells Induced by Globotriaosylsphingosine and Globotriaosylceramide.

    Directory of Open Access Journals (Sweden)

    Yeo Jin Jeon

    Full Text Available Fabry disease is a lysosomal storage disorder caused by deficiency of alpha-galactosidase A (α-gal A, which results in the deposition of globotriaosylceramide (Gb3 in the vascular endothelium. Globotriaosylsphingosine (lyso-Gb3, a deacylated Gb3, is also increased in the plasma of patients with Fabry disease. Renal fibrosis is a key feature of advanced Fabry disease patients. Therefore, we evaluated the association of Gb3 and lyso-Gb3 accumulation and the epithelial-mesenchymal transition (EMT on tubular epithelial cells of the kidney. In HK2 cells, exogenous treatments of Gb3 and lyso-Gb3 increased the expression of TGF-β, EMT markers (N-cadherin and α-SMA, and phosphorylation of PI3K/AKT, and decreased the expression of E-cadherin. Lyso-Gb3, rather than Gb3, strongly induced EMT in HK2 cells. In the mouse renal mesangial cell line, SV40 MES 13 cells, Gb3 strongly induced phenotype changes. The EMT induced by Gb3 was inhibited by enzyme α-gal A treatment, but EMT induced by lyso-Gb3 was not abrogated by enzyme treatment. However, TGF-β receptor inhibitor (TRI, SB525334 inhibited the activation of TGF-β and EMT markers in HK2 cells with Gb3 and lyso-Gb3 treatments. This study suggested that increased plasma lyso-Gb3 has a crucial role in the development of renal fibrosis through the cell-specific induction of the EMT in Fabry disease, and that TRI treatment, alongside enzyme replacement therapy, could be a potential therapeutic option for patients with Fabry disease.

  18. Antiadhesive and cytotoxic effect of Iranian Vipera lebetina snake venom on lung epithelial cancer cells.

    Science.gov (United States)

    Oghalaie, Akbar; Kazemi-Lomedasht, Fatemeh; Zareinejad, Mohammad Reza; Shahbazzadeh, Delavar

    2017-01-01

    Cancer is one of the major health problems worldwide. Hence, finding potent therapeutics from natural sources seems necessary. Snake venom of Vipera lebetina contains potential component with anticancer activities such as antiproliferation, migration, invasion, adhesion, and angiogenesis effect. Evaluation of cytotoxic and antiadhesive effect of V. lebetina venom on lung epithelial cancer tumor cell (TC-1) was the main aim of this study. Here, we purified snake venom of V. lebetina by fast protein liquid chromatography (FPLC) using Sephacryl S-200 hr column. The fractions collected and evaluated by SDS-PAGE analysis. The cytotoxicity and antiadhesive effect of crude venom and fractions on TC-1 cells were demonstrated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and adhesion assay, respectively. Our results showed six fractions in FPLC diagram. V. lebetina crude venom and fractions showed dose-dependent cytotoxic effect on TC-1 cells. Fractions 2 and 5 showed high cytotoxic effect with high IC50 value (IC50 = 6 μg/ml for fraction 2 and IC50 = 7.3 μg/ml for fraction 5). Fractions 2 and 5 selected for analysis antiadhesive effect on TC-1 cells. Furthermore, our results showed that both fractions 2 and 5 had antiadhesive effect on TC-1 cells. Because of potent cytotoxic and antiadhesive effect of V. lebetina fractions on lung epithelial cancer cell line, it could be promising tools for further analysis as anticancer therapeutic development.

  19. Antiadhesive and cytotoxic effect of Iranian Vipera lebetina snake venom on lung epithelial cancer cells

    Directory of Open Access Journals (Sweden)

    Akbar Oghalaie

    2017-01-01

    Full Text Available Background: Cancer is one of the major health problems worldwide. Hence, finding potent therapeutics from natural sources seems necessary. Snake venom of Vipera lebetina contains potential component with anticancer activities such as antiproliferation, migration, invasion, adhesion, and angiogenesis effect. Evaluation of cytotoxic and antiadhesive effect of V. lebetina venom on lung epithelial cancer tumor cell (TC-1 was the main aim of this study. Materials and Methods: Here, we purified snake venom of V. lebetina by fast protein liquid chromatography (FPLC using Sephacryl S-200 hr column. The fractions collected and evaluated by SDS-PAGE analysis. The cytotoxicity and antiadhesive effect of crude venom and fractions on TC-1 cells were demonstrated using 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide and adhesion assay, respectively. Results: Our results showed six fractions in FPLC diagram. V. lebetina crude venom and fractions showed dose-dependent cytotoxic effect on TC-1 cells. Fractions 2 and 5 showed high cytotoxic effect with high IC50 value (IC50 = 6 μg/ml for fraction 2 and IC50 = 7.3 μg/ml for fraction 5. Fractions 2 and 5 selected for analysis antiadhesive effect on TC-1 cells. Furthermore, our results showed that both fractions 2 and 5 had antiadhesive effect on TC-1 cells. Conclusion: Because of potent cytotoxic and antiadhesive effect of V. lebetina fractions on lung epithelial cancer cell line, it could be promising tools for further analysis as anticancer therapeutic development.

  20. Lentiviral Vector Mediated Claudin1 Silencing Inhibits Epithelial to Mesenchymal Transition in Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Xianqi Zhao

    2015-06-01

    Full Text Available Breast cancer has a high incidence and mortality rate worldwide. Several viral vectors including lentiviral, adenoviral and adeno-associated viral vectors have been used in gene therapy for various forms of human cancer, and have shown promising effects in controlling tumor development. Claudin1 (CLDN1 is a member of the tetraspan transmembrane protein family that plays a major role in tight junctions and is associated with tumor metastasis. However, the role of CLDN1 in breast cancer is largely unexplored. In this study, we tested the therapeutic potential of silencing CLDN1 expression in two breast cancer (MDA-MB-231 and MCF7 cell lines using lentiviral vector mediated RNA interference. We found that a CLDN1 short hairpin (shRNA construct efficiently silenced CLDN1 expression in both breast cancer cell lines, and CLDN1 knockdown resulted in reduced cell proliferation, survival, migration and invasion. Furthermore, silencing CLDN1 inhibited epithelial to mesenchymal transition (EMT by upregulating the epithelial cell marker, E-cadherin, and downregulating mesenchymal markers, smooth muscle cell alpha-actin (SMA and Snai2. Our data demonstrated that lentiviral vector mediated CLDN1 RNA interference has great potential in breast cancer gene therapy by inhibiting EMT and controlling tumor cell growth.

  1. Mesenchymal precursor cells maintain the differentiation and proliferation potentials of breast epithelial cells

    Science.gov (United States)

    2014-01-01

    Introduction Stromal-epithelial interactions play a fundamental role in tissue homeostasis, controlling cell proliferation and differentiation. Not surprisingly, aberrant stromal-epithelial interactions contribute to malignancies. Studies of the cellular and molecular mechanisms underlying these interactions require ex vivo experimental model systems that recapitulate the complexity of human tissue without compromising the differentiation and proliferation potentials of human primary cells. Methods We isolated and characterized human breast epithelial and mesenchymal precursors from reduction mammoplasty tissue and tagged them with lentiviral vectors. We assembled heterotypic co-cultures and compared mesenchymal and epithelial cells to cells in corresponding monocultures by analyzing growth, differentiation potentials, and gene expression profiles. Results We show that heterotypic culture of non-immortalized human primary breast epithelial and mesenchymal precursors maintains their proliferation and differentiation potentials and constrains their growth. We further describe the gene expression profiles of stromal and epithelial cells in co-cultures and monocultures and show increased expression of the tumor growth factor beta (TGFβ) family member inhibin beta A (INHBA) in mesenchymal cells grown as co-cultures compared with monocultures. Notably, overexpression of INHBA in mesenchymal cells increases colony formation potential of epithelial cells, suggesting that it contributes to the dynamic reciprocity between breast mesenchymal and epithelial cells. Conclusions The described heterotypic co-culture system will prove useful for further characterization of the molecular mechanisms mediating interactions between human normal or neoplastic breast epithelial cells and the stroma, and will provide a framework to test the relevance of the ever-increasing number of oncogenomic alterations identified in human breast cancer. PMID:24916766

  2. Cytotoxicity of ophthalmic solutions with and without preservatives to human corneal endothelial cells, epithelial cells and conjunctival epithelial cells.

    Science.gov (United States)

    Ayaki, Masahiko; Yaguchi, Shigeo; Iwasawa, Atsuo; Koide, Ryohei

    2008-08-01

    The cytotoxicity of a range of commercial ophthalmic solutions in the presence and absence of preservatives was assessed in human corneal endothelial cells (HCECs), corneal epithelia and conjunctival epithelia using in vitro techniques. Cell survival was measured using the WST-1 assay for endothelial cells and the MTT assay for epithelial cells. Commercially available timolol, carteolol, cromoglicate, diclofenac, bromfenac and hyaluronic acid ophthalmic solutions were assessed for cytotoxicity in the presence and absence of preservatives. The preservatives benzalkonium, chlorobutanol and polysorbate were also tested. The survival of cells exposed to test ophthalmic solutions was expressed as a percentage of cell survival in the control solution (distilled water added to media) after 48 h exposure. HCEC survival was 20-30% in ophthalmic solutions diluted 10-fold. The survival of HCEC was significantly greater in all solutions in the absence of preservative than in the presence of preservative. The survival of corneal and conjunctival epithelia was consistent with that of HCECs for all test ophthalmic solutions. The preservatives polysorbate and benzalkonium were highly cytotoxic with cell survival decreasing to 20% at the concentration estimated in commercial ophthalmic solutions. By comparison, the survival of cells exposed to chlorobutanol was 80% or greater. The cytotoxicity of ophthalmic solutions to HCEC, corneal epithelia and conjunctival epithelia decreased in the absence of preservative.

  3. Syndecan-1 suppresses epithelial-mesenchymal transition and migration in human oral cancer cells.

    Science.gov (United States)

    Wang, Xiaofeng; He, Jinting; Zhao, Xiaoming; Qi, Tianyang; Zhang, Tianfu; Kong, Chenfei

    2018-04-01

    Epithelial-mesenchymal transition (EMT) is one of the major processes that contribute to the occurrence of cancer metastasis. EMT has been associated with the development of oral cancer. Syndecan‑1 (SDC1) is a key cell‑surface adhesion molecule and its expression level inversely correlates with tumor differentiation and prognosis. In the present study, we aimed to determine the role of SDC1 in oral cancer progression and investigate the molecular mechanisms through which SDC1 regulates the EMT and invasiveness of oral cancer cells. We demonstrated that basal SDC1 expression levels were lower in four oral cancer cell lines (KB, Tca8113, ACC2 and CAL‑27), than in normal human periodontal ligament fibroblasts. Ectopic overexpression of SDC1 resulted in morphological transformation, decreased expression of EMT‑associated markers, as well as decreased migration, invasiveness and proliferation of oral cancer cells. In contrast, downregulation of the expression of SDC1 caused the opposite results. Furthermore, the knockdown of endogenous SDC1 activated the extracellular signal‑regulated kinase (ERK) cascade, upregulated the expression of Snail and inhibited the expression of E‑cadherin. In conclusion, our findings revealed that SDC1 suppressed EMT via the modulation of the ERK signaling pathway that, in turn, negatively affected the invasiveness of human oral cancer cells. Our results provided useful evidence about the potential use of SDC1 as a molecular target for therapeutic interventions in human oral cancer.

  4. Ethane dimethanesulfonate (EDS) perturbs epididymal epithelial cell function in vitro

    International Nuclear Information System (INIS)

    Klinefelter, G.

    1990-01-01

    The formation of sperm granulomas in the epididymis following exposure to EDS, a Leydig cell toxicant, was reported by Cooper and Jackson in 1970. Recent work suggests that EDS may effect the epididymis directly. An in vitro system was developed to determine the nature of any direct effect. The caput epididymis from adult rats was dissected free of connective tissue and small pieces of the tissue were enzymatically digested until plaques of epididymal epithelial cells were obtained. Plaques were cultured on an extracellular matrix gelled on top of a semipermeable filter creating dual-compartment environments. The epithelial cells maintained typical morphology and protein secretion in this culture system for several days. Beginning on day 3, EDS (1 mM) was added to the basal compartment, with or without 35 S-methionine. After 24 hours, 35 S-labelled culture medium was taken from the apical compartment and analyzed by SDS-PAGE and fluorography. EDS caused decreased secretion of several proteins, including a 39 Kd molecule. Interestingly, a 39 Kd protein was also shown to disappear from sperm taken from the caput epididymidis following in vivo exposure to EDS. Unlabelled cultures were fixed and processed for light microscopy. No alterations in morphological integrity were observed. Thus, epididymal epithelial cell function is directly altered by EDS exposure

  5. CCL20, (gamma)(delta) T cells, and IL-22 in corneal epithelial healing

    Science.gov (United States)

    After corneal epithelial abrasion, leukocytes and platelets rapidly enter the corneal stroma, and CCR6 (+) IL-17(+) gamma delta T cells migrate into the epithelium. Gamma delta T-cell-deficient (TCRd(-/-)) mice have significantly reduced inflammation and epithelial wound healing. Epithelial CCL20 mR...

  6. Protective effects of trehalose on the corneal epithelial cells.

    Science.gov (United States)

    Aragona, Pasquale; Colosi, Pietro; Rania, Laura; Colosi, Francesca; Pisani, Antonina; Puzzolo, Domenico; Micali, Antonio

    2014-01-01

    Aim of the present work was to evaluate the effects of the trehalose on the corneal epithelium undergoing alcohol delamination. Twelve patients undergoing laser subepithelial keratomileusis (LASEK) were consecutively included in the study. The right eyes were pretreated with 3% trehalose eye drops, whilst left eyes were used as control. Epithelial specimens were processed for cells vitality assessment, apoptosis, and light and transmission electron microscopy; a morphometric analysis was performed in both groups. In both trehalose-untreated eyes (TUE) and trehalose-treated eyes (TTE), the percentage of vital cells was similar and no apoptotic cells were observed. In TUE, the corneal epithelium showed superficial cells with reduced microfolds, wing cells with vesicles and dilated intercellular spaces, and dark basal cells with vesicles and wide clefts. In TTE, superficial and wing cells were better preserved, and basal cells were generally clear with intracytoplasmatic vesicles. The morphometric analysis showed statistically significant differences between the two groups: the TTE epithelial height was higher, the basal cells showed larger area and clearer cytoplasm. The distribution of desmosomes and hemidesmosomes was significantly different between the groups. Trehalose administration better preserved morphological and morphometric features of alcohol-treated corneal epithelium, when compared to controls.

  7. Protective Effects of Trehalose on the Corneal Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Pasquale Aragona

    2014-01-01

    Full Text Available Purpose. Aim of the present work was to evaluate the effects of the trehalose on the corneal epithelium undergoing alcohol delamination. Methods. Twelve patients undergoing laser subepithelial keratomileusis (LASEK were consecutively included in the study. The right eyes were pretreated with 3% trehalose eye drops, whilst left eyes were used as control. Epithelial specimens were processed for cells vitality assessment, apoptosis, and light and transmission electron microscopy; a morphometric analysis was performed in both groups. Results. In both trehalose-untreated eyes (TUE and trehalose-treated eyes (TTE, the percentage of vital cells was similar and no apoptotic cells were observed. In TUE, the corneal epithelium showed superficial cells with reduced microfolds, wing cells with vesicles and dilated intercellular spaces, and dark basal cells with vesicles and wide clefts. In TTE, superficial and wing cells were better preserved, and basal cells were generally clear with intracytoplasmatic vesicles. The morphometric analysis showed statistically significant differences between the two groups: the TTE epithelial height was higher, the basal cells showed larger area and clearer cytoplasm. The distribution of desmosomes and hemidesmosomes was significantly different between the groups. Conclusions. Trehalose administration better preserved morphological and morphometric features of alcohol-treated corneal epithelium, when compared to controls.

  8. Snail1 induces epithelial-to-mesenchymal transition and tumor initiating stem cell characteristics

    International Nuclear Information System (INIS)

    Dang, Hien; Ding, Wei; Emerson, Dow; Rountree, C Bart

    2011-01-01

    Tumor initiating stem-like cells (TISCs) are a subset of neoplastic cells that possess distinct survival mechanisms and self-renewal characteristics crucial for tumor maintenance and propagation. The induction of epithelial-mesenchymal-transition (EMT) by TGFβ has been recently linked to the acquisition of TISC characteristics in breast cancer. In HCC, a TISC and EMT phenotype correlates with a worse prognosis. In this work, our aim is to elucidate the underlying mechanism by which cells acquire tumor initiating characteristics after EMT. Gene and protein expression assays and Nanog-promoter luciferase reporter were utilized in epithelial and mesenchymal phenotype liver cancer cell lines. EMT was analyzed with migration/invasion assays. TISC characteristics were analyzed with tumor-sphere self-renewal and chemotherapy resistance assays. In vivo tumor assay was performed to investigate the role of Snail1 in tumor initiation. TGFβ induced EMT in epithelial cells through the up-regulation of Snail1 in Smad-dependent signaling. Mesenchymal liver cancer post-EMT demonstrates TISC characteristics such as tumor-sphere formation but are not resistant to cytotoxic therapy. The inhibition of Snail1 in mesenchymal cells results in decreased Nanog promoter luciferase activity and loss of self-renewal characteristics in vitro. These changes confirm the direct role of Snail1 in some TISC traits. In vivo, the down-regulation of Snail1 reduced tumor growth but was not sufficient to eliminate tumor initiation. In summary, TGFβ induces EMT and TISC characteristics through Snail1 and Nanog up-regulation. In mesenchymal cells post-EMT, Snail1 directly regulates Nanog expression, and loss of Snail1 regulates tumor growth without affecting tumor initiation

  9. Mechanisms of Cell Polarity-Controlled Epithelial Homeostasis and Immunity in the Intestine

    NARCIS (Netherlands)

    Klunder, Leon J.; Faber, Klaas Nico; Dijkstra, Gerard; van IJzendoorn, Sven C. D.

    Intestinal epithelial cell polarity is instrumental to maintain epithelial homeostasis and balance communications between the gut lumen and bodily tissue, thereby controlling the defense against gastrointestinal pathogens and maintenance of immune tolerance to commensal bacteria. In this review, we

  10. Inositol induces mesenchymal-epithelial reversion in breast cancer cells through cytoskeleton rearrangement.

    Science.gov (United States)

    Dinicola, Simona; Fabrizi, Gianmarco; Masiello, Maria Grazia; Proietti, Sara; Palombo, Alessandro; Minini, Mirko; Harrath, Abdel Halim; Alwasel, Saleh H; Ricci, Giulia; Catizone, Angela; Cucina, Alessandra; Bizzarri, Mariano

    2016-07-01

    Inositol displays multi-targeted effects on many biochemical pathways involved in epithelial-mesenchymal transition (EMT). As Akt activation is inhibited by inositol, we investigated if such effect could hamper EMT in MDA-MB-231 breast cancer cells. In cancer cells treated with pharmacological doses of inositol E-cadherin was increased, β-catenin was redistributed behind cell membrane, and metalloproteinase-9 was significantly reduced, while motility and invading capacity were severely inhibited. Those changes were associated with a significant down-regulation of PI3K/Akt activity, leading to a decrease in downstream signaling effectors: NF-kB, COX-2, and SNAI1. Inositol-mediated inhibition of PS1 leads to lowered Notch 1 release, thus contributing in decreasing SNAI1 levels. Overall, these data indicated that inositol inhibits the principal molecular pathway supporting EMT. Similar results were obtained in ZR-75, a highly metastatic breast cancer line. These findings are coupled with significant changes on cytoskeleton. Inositol slowed-down vimentin expression in cells placed behind the wound-healing edge and stabilized cortical F-actin. Moreover, lamellipodia and filopodia, two specific membrane extensions enabling cell migration and invasiveness, were no longer detectable after inositol addiction. Additionally, fascin and cofilin, two mandatory required components for F-actin assembling within cell protrusions, were highly reduced. These data suggest that inositol may induce an EMT reversion in breast cancer cells, suppressing motility and invasiveness through cytoskeleton modifications. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. A second wave of Salmonella T3SS1 activity prolongs the lifespan of infected epithelial cells.

    Directory of Open Access Journals (Sweden)

    Ciaran E Finn

    2017-04-01

    Full Text Available Type III secretion system 1 (T3SS1 is used by the enteropathogen Salmonella enterica serovar Typhimurium to establish infection in the gut. Effector proteins translocated by this system across the plasma membrane facilitate invasion of intestinal epithelial cells. One such effector, the inositol phosphatase SopB, contributes to invasion and mediates activation of the pro-survival kinase Akt. Following internalization, some bacteria escape from the Salmonella-containing vacuole into the cytosol and there is evidence suggesting that T3SS1 is expressed in this subpopulation. Here, we investigated the post-invasion role of T3SS1, using SopB as a model effector. In cultured epithelial cells, SopB-dependent Akt phosphorylation was observed at two distinct stages of infection: during and immediately after invasion, and later during peak cytosolic replication. Single cell analysis revealed that cytosolic Salmonella deliver SopB via T3SS1. Although intracellular replication was unaffected in a SopB deletion mutant, cells infected with ΔsopB demonstrated a lack of Akt phosphorylation, earlier time to death, and increased lysis. When SopB expression was induced specifically in cytosolic Salmonella, these effects were restored to levels observed in WT infected cells, indicating that the second wave of SopB protects this infected population against cell death via Akt activation. Thus, T3SS1 has two, temporally distinct roles during epithelial cell colonization. Additionally, we found that delivery of SopB by cytosolic bacteria was translocon-independent, in contrast to canonical effector translocation across eukaryotic membranes, which requires formation of a translocon pore. This mechanism was also observed for another T3SS1 effector, SipA. These findings reveal the functional and mechanistic adaptability of a T3SS that can be harnessed in different microenvironments.

  12. BMP2 induces PANC-1 cell invasion by MMP-2 overexpression through ROS and ERK.

    Science.gov (United States)

    Liu, Jun; Ben, Qi-Wen; Yao, Wei-Yan; Zhang, Jian-Jun; Chen, Da-Fan; He, Xiang-Yi; Li, Lei; Yuan, Yao-Zong

    2012-06-01

    The emerging roles of bone morphogenetic proteins (BMPs) in the initiation and progression of multiple cancers have drawn great attention in cancer research. We hypothesized that BMP2 promotes cancer metastasis by modulating MMP-2 secretion and activity through intracellular ROS regulation and ERK activation in human pancreatic cancer. Our data show that stimulation of PANC-1 cells with BMP2 induced MMP-2 secretion and activation, associated with decreased E-cadherin expression, resulting in epithelial-to-mesenchymal transformation (EMT) and cell invasion. Blockade of ROS by the ROS scavenger, 2-MPG, abolished cell invasion, inhibited the EMT process and decreased MMP-2 expression, suggesting ROS accumulation caused an increase in MMP-2 expression in BMP2-stimulated PANC-1 cell invasion. Furthermore, treatment of PANC-1 cells with 2-MPG or ERK inhibitor PD98059 reduced the phosphorylation of ERK, resulting in attenuation of BMP2-induced cell invasion and MMP-2 activation. Taken together, these results suggest that BMP2 induces the cell invasion of PANC-1 cells by enhancing MMP-2 secretion and acting through ROS accumulation and ERK activation.

  13. Transformation of Epithelial Ovarian Cancer Stemlike Cells into Mesenchymal Lineage via EMT Results in Cellular Heterogeneity and Supports Tumor Engraftment

    Science.gov (United States)

    Jiang, Hua; Lin, Xiaolong; Liu, Yingtao; Gong, Wenjia; Ma, Xiaoling; Yu, Yinhua; Xie, Yi; Sun, Xiaoxi; Feng, Youji; Janzen, Viktor; Chen, Tong

    2012-01-01

    Ovarian cancers are heterogeneous and contain stemlike cells that are able to self-renew and are responsible for sustained tumor growth. Metastasis in the peritoneal cavity occurs more frequently in ovarian cancer than in other malignancies, but the underlying mechanism remains largely unknown. We have identified that ovarian cancer stemlike cells (CSCs), which were defined as side population (SP) cells, were present in patients’ ascitic fluid and mesenchymally transformed cell lines, ES-2 and HO-8910PM. SP cells, which were sorted from both cell lines and implanted into immunocompromised mice, were localized to the xenografted tumor boundary. In addition, SP cells exhibited an epithelial phenotype and showed a distinct gene expression profile with reduced expression of cell adhesion molecules (CAMs), indicating that SP cells exert an important role in ovarian cancer progression on the basis of their delicate interaction with the surrounding microenvironment and anatomical localization in tumors. In contrast, non-SP cells exhibited a more mesenchymal phenotype and showed more increased invasive potential than SP cells. This heterogeneity was observed as an endogenous transformation via the epithelial–mesenchymal transition (EMT) process. Inhibition of the EMT process by Snail1 silencing reduced the SP cell frequency, and affected their invasive capacity and engraftment. These findings illustrate the interplay between epithelial ovarian CSCs and the EMT, and exert a link to explain tumor heterogeneity and its necessity for ovarian cancer maintenance, metastasis and progression. PMID:22801793

  14. Epithelial cells derived from human embryonic stem cells display p16INK4A senescence, hypermotility, and differentiation properties shared by many P63+ somatic cell types

    DEFF Research Database (Denmark)

    Dabelsteen, Sally; Hercule, Paula; Barron, Patricia

    2009-01-01

    hESderK cells and keratinocytes a substantially extended lifespan. When exposed to transforming growth factor beta or to an incompletely processed form of Laminin-332, three lifespan-extended or immortalized hESderK lines that we studied became directionally hypermotile, a wound healing and invasion......(+)/K14(+) urothelial and tracheobronchial epithelial cells. Primary and immortalized lines of these cell types had growth requirements and hypermotility responses similar to keratinocytes and bmi1 expression facilitated their immortalization by engineering to express the catalytic subunit of telomerase...

  15. TCDD alters medial epithelial cell differentiation during palatogenesis

    International Nuclear Information System (INIS)

    Abbott, B.D.; Birnbaum, L.S.

    1989-01-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a widely distributed, persistent environmental contaminant that is teratogenic in mice, where it induces hydronephrosis and cleft palate. The incidence of clefting has been shown to be dose dependent after exposure on either gestation Day (GD) 10 or 12, although the embryo is more susceptible on GD 12. TCDD-exposed palatal shelves meet but do not fuse, and programmed cell death of the medial epithelial cells is inhibited. The mechanism of action through which TCDD alters the program of medial cell development has not been examined in earlier studies, and it is not known whether the mechanism is the same regardless of the dose or developmental stage of exposure. In this study, C57BL/6N mice, a strain sensitive to TCDD, were dosed orally on GD 10 or 12 with 0, 6, 12, 24, or 30 micrograms/kg body wt, in 10 ml corn oil/kg. Embryonic palatal shelves were examined on GD 14, 15, or 16. The degree of palatal closure, epithelial surface morphology, and cellular ultrastructure, the incorporation of [3H]TdR, the expression of EGF receptors, and the binding of 125I-EGF were assessed. After exposure on GD 10 or 12, TCDD altered the differentiation pathway of the medial epithelial cells. The palatal shelves were of normal size and overall morphology, but fusion of the medial epithelia of the opposing shelves did not occur. TCDD prevented programmed cell death of the medial peridermal cells. The expression of EGF receptors by medial cells continued through Day 16 and the receptors were able to bind ligand. The medial cells differentiated into a stratified, squamous, keratinizing epithelium. The shift in phenotype to an oral-like epithelium occurred after exposure on either GD 10 or 12. At the lower dose (6 micrograms/kg), fewer cleft palates were produced, but those shelves which did respond had a fully expressed shift in differentiation

  16. Isolation, separation, and characterization of epithelial and connective cells from rat palate

    Energy Technology Data Exchange (ETDEWEB)

    Terranova, Victor Paul [Univ. of Rochester, NY (United States)

    1979-01-01

    Epithelial and connective tissue cells were isolated from rat palate by sequential collagenase, hyaluronidase and trypsin digestion of the extracellular matrix. Differences between the two populations were noted with respect to total cell protein, total cell water, proline uptake and incorporation, percent collagen synthesized, effects of parathyroid hormone, metabolism of D-valine and cell density. Basal epithelial cells were subsequently separated from the heterogeneous epithelial cell population on shallow linear density gradients by velocity centrifugation. The type of collagen synthesized by the basal epithelial cells was compared to the type of collagen synthesized by the connective tissue cells by means of labeled amino acid incorporation ratios. Cells isolated from the epithelial and connective tissue were compared. From these studies it can be concluded that epithelial and connective tissue cells can be isolated from rat palate as viable and distinct populations with respect to the biochemical parameters examined. Furthermore, subpopulations can be separated and biochemically characterized.

  17. Stonin 2 Overexpression is Correlated with Unfavorable Prognosis and Tumor Invasion in Epithelial Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Xiaoying Sun

    2017-07-01

    Full Text Available Stonin 2 (STON2, which functions in adjusting endocytotic complexes, is probably involved in the monitoring of the internalization of dopamine D2 receptors which have an inhibitory action of dopamine on tumor progression. However, its clinical significance in tumor progression and prognosis remains unclear. We explored the association between STON2 and the clinicopathological characteristics of epithelial ovarian cancer (EOC. The STON2 levels in ovarian cancer and normal cell lines and tissues were detected by real-time PCR and Western blot analyses. STON2 protein expression was also detected by an immunohistochemical analysis. The clinical significance of STON2 expression in ovarian cancer was statistically analyzed. STON2 significantly increased in the ovarian cancer cell lines and tissues compared to the normal ones. In the 89 EOC samples tested, STON2 expression was significantly correlated with intraperitoneal metastasis, intestinal metastasis, intraperitoneal recurrence, ascites containing tumor cells, and CA153 level. Moreover, patients with STON2 protein overexpression were more likely to exhibit platinum resistance and to have undergone neoadjuvant chemotherapy. Patients with high STON2 protein expression had a tendency to have a shorter overall survival and a poor prognosis. A multivariate analysis showed that STON2 was an independent prognostic predictor for EOC patients. In conclusion, STON2 plays an important role in the progression and prognosis of ovarian carcinoma, especially in platinum resistance, intraperitoneal metastasis, and recurrence. STON2 can be a novel antitumor drug target and biomarker which predicts an unfavorable prognosis for EOC patients.

  18. Oral epithelial cells are susceptible to cell-free and cell-associated HIV-1 infection in vitro

    International Nuclear Information System (INIS)

    Moore, Jennifer S.; Rahemtulla, Firoz; Kent, Leigh W.; Hall, Stacy D.; Ikizler, Mine R.; Wright, Peter F.; Nguyen, Huan H.; Jackson, Susan

    2003-01-01

    Epithelial cells lining the oral cavity are exposed to HIV-1 through breast-feeding and oral-genital contact. Genital secretions and breast milk of HIV-1-infected subjects contain both cell-free and cell-associated virus. To determine if oral epithelial cells can be infected with HIV-1 we exposed gingival keratinocytes and adenoid epithelial cells to cell-free virus and HIV-1-infected peripheral blood mononuclear cells and monocytes. Using primary isolates we determined that gingival keratinocytes are susceptible to HIV-1 infection via cell-free CD4-independent infection only. R5 but not X4 viral strains were capable of infecting the keratinocytes. Further, infected cells were able to release infectious virus. In addition, primary epithelial cells isolated from adenoids were also susceptible to infection; both cell-free and cell-associated virus infected these cells. These data have potential implications in the transmission of HIV-1 in the oral cavity

  19. Entrance and Survival of Brucella pinnipedialis Hooded Seal Strain in Human Macrophages and Epithelial Cells

    Science.gov (United States)

    Briquemont, Benjamin; Sørensen, Karen K.; Godfroid, Jacques

    2013-01-01

    Marine mammal Brucella spp. have been isolated from pinnipeds (B. pinnipedialis) and cetaceans (B. ceti) from around the world. Although the zoonotic potential of marine mammal brucellae is largely unknown, reports of human disease exist. There are few studies of the mechanisms of bacterial intracellular invasion and multiplication involving the marine mammal Brucella spp. We examined the infective capacity of two genetically different B. pinnipedialis strains (reference strain; NTCT 12890 and a hooded seal isolate; B17) by measuring the ability of the bacteria to enter and replicate in cultured phagocytes and epithelial cells. Human macrophage-like cells (THP-1), two murine macrophage cell lines (RAW264.7 and J774A.1), and a human malignant epithelial cell line (HeLa S3) were challenged with bacteria in a gentamicin protection assay. Our results show that B. pinnipedialis is internalized, but is then gradually eliminated during the next 72 – 96 hours. Confocal microscopy revealed that intracellular B. pinnipedialis hooded seal strain colocalized with lysosomal compartments at 1.5 and 24 hours after infection. Intracellular presence of B. pinnipedialis hooded seal strain was verified by transmission electron microscopy. By using a cholesterol-scavenging lipid inhibitor, entrance of B. pinnipedialis hooded seal strain in human macrophages was significantly reduced by 65.8 % (± 17.3), suggesting involvement of lipid-rafts in intracellular entry. Murine macrophages invaded by B. pinnipedialis do not release nitric oxide (NO) and intracellular bacterial presence does not induce cell death. In summary, B. pinnipedialis hooded seal strain can enter human and murine macrophages, as well as human epithelial cells. Intracellular entry of B. pinnipedialis hooded seal strain involves, but seems not to be limited to, lipid-rafts in human macrophages. Brucella pinnipedialis does not multiply or survive for prolonged periods intracellulary. PMID:24376851

  20. Quantitative imaging of epithelial cell scattering identifies specific inhibitors of cell motility and cell-cell dissociation

    NARCIS (Netherlands)

    Loerke, D.; le Duc, Q.; Blonk, I.; Kerstens, A.; Spanjaard, E.; Machacek, M.; Danuser, G.; de Rooij, J.

    2012-01-01

    The scattering of cultured epithelial cells in response to hepatocyte growth factor (HGF) is a model system that recapitulates key features of metastatic cell behavior in vitro, including disruption of cell-cell adhesions and induction of cell migration. We have developed image analysis tools that

  1. Transcriptional profiling of putative human epithelial stem cells

    Directory of Open Access Journals (Sweden)

    Koçer Salih S

    2008-07-01

    may be enriched for stem cells. This study is the first comprehensive gene expression profile of putative human epithelial stem cells and their progeny that were isolated directly from neonatal foreskin tissue. Our study is important for understanding self renewal and differentiation of epidermal stem cells, and for elucidating signaling pathways involved in those processes. The generated data base may serve those working with other human epithelial tissue progenitors.

  2. Cooperation between epithelial cells demonstrated by potassium transfer

    International Nuclear Information System (INIS)

    Ledbetter, M.L.; Young, G.J.; Wright, E.R.

    1986-01-01

    Junction-mediated communication can be measured in fibroblast cultures by determining the ability of mixed cultures of cells sensitive and resistant to ouabain to concentrate K+ in the presence of ouabain. We now report the extension of this assay procedure to cultured epithelial cells. Hamster kidney (HaK) cells maintain their ability to concentrate K+ in ouabain at levels inhibitory to dog kidney (MDCK) cells. When HaK and MDCK cells were cultured together in ouabain-containing medium, the K+ (measured as 86Rb+) in the mixed population was greater than expected if the cells were not interacting. The degree of enhancement, expressed as index of cooperation, depended on the numbers of cells in the cultures, their opportunity for cell-to-cell contact, and (above a certain permissive level) the concentration of ouabain. As with other cell types, protein synthesis in MDCK cells depends on maintenance of cell K+. Autoradiography of cells incubated with [3H]leucine demonstrated that MDCK cells in ouabain-treated mixed cultures were able to synthesize proteins only when physically adjacent to HaK cells. The transmission of labeled nucleosides among the cells provides independent evidence of the phenomenon of cooperation, probably mediated by gap junctions. This system offers promise for investigation of stimuli modulating junctional communication

  3. Persistent Transmissible Gastroenteritis Virus Infection Enhances Enterotoxigenic Escherichia coli K88 Adhesion by Promoting Epithelial-Mesenchymal Transition in Intestinal Epithelial Cells.

    Science.gov (United States)

    Xia, Lu; Dai, Lei; Yu, Qinghua; Yang, Qian

    2017-11-01

    Transmissible gastroenteritis virus (TGEV) is a coronavirus characterized by diarrhea and high morbidity rates, and the mortality rate is 100% in piglets less than 2 weeks old. Pigs infected with TGEV often suffer secondary infection by other pathogens, which aggravates the severity of diarrhea, but the mechanisms remain unknown. Here, we hypothesized that persistent TGEV infection stimulates the epithelial-mesenchymal transition (EMT), and thus enterotoxigenic Escherichia coli (ETEC) can more easily adhere to generating cells. Intestinal epithelial cells are the primary targets of TGEV and ETEC infections. We found that TGEV can persistently infect porcine intestinal columnar epithelial cells (IPEC-J2) and cause EMT, consistent with multiple changes in key cell characteristics. Infected cells display fibroblast-like shapes; exhibit increases in levels of mesenchymal markers with a corresponding loss of epithelial markers; have enhanced expression levels of interleukin-1β (IL-1β), IL-6, IL-8, transforming growth factor β (TGF-β), and tumor necrosis factor alpha (TNF-α) mRNAs; and demonstrate increases in migratory and invasive behaviors. Additional experiments showed that the activation of the phosphatidylinositol 3-kinase (PI3K)/Akt and extracellular signal-regulated kinase (ERK) signaling pathways via TGF-β is critical for the TGEV-mediated EMT process. Cellular uptake is also modified in cells that have undergone EMT. TGEV-infected cells have higher levels of integrin α5 and fibronectin and exhibit enhanced ETEC K88 adhesion. Reversal of EMT reduces ETEC K88 adhesion and inhibits the expression of integrin α5 and fibronectin. Overall, these results suggest that TGEV infection induces EMT in IPEC-J2 cells, increasing the adhesion of ETEC K88 in the intestine and facilitating dual infection. IMPORTANCE Transmissible gastroenteritis virus (TGEV) causes pig diarrhea and is often followed by secondary infection by other pathogens. In this study, we showed

  4. Studies Using an in Vitro Model Show Evidence of Involvement of Epithelial-Mesenchymal Transition of Human Endometrial Epithelial Cells in Human Embryo Implantation*

    Science.gov (United States)

    Uchida, Hiroshi; Maruyama, Tetsuo; Nishikawa-Uchida, Sayaka; Oda, Hideyuki; Miyazaki, Kaoru; Yamasaki, Akiko; Yoshimura, Yasunori

    2012-01-01

    Human embryo implantation is a critical multistep process consisting of embryo apposition/adhesion, followed by penetration and invasion. Through embryo penetration, the endometrial epithelial cell barrier is disrupted and remodeled by an unknown mechanism. We have previously developed an in vitro model for human embryo implantation employing the human choriocarcinoma cell line JAR and the human endometrial adenocarcinoma cell line Ishikawa. Using this model we have shown that stimulation with ovarian steroid hormones (17β-estradiol and progesterone, E2P4) and suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, enhances the attachment and adhesion of JAR spheroids to Ishikawa. In the present study we showed that the attachment and adhesion of JAR spheroids and treatment with E2P4 or SAHA individually induce the epithelial-mesenchymal transition (EMT) in Ishikawa cells. This was evident by up-regulation of N-cadherin and vimentin, a mesenchymal cell marker, and concomitant down-regulation of E-cadherin in Ishikawa cells. Stimulation with E2P4 or SAHA accelerated Ishikawa cell motility, increased JAR spheroid outgrowth, and enhanced the unique redistribution of N-cadherin, which was most prominent in proximity to the adhered spheroids. Moreover, an N-cadherin functional blocking antibody attenuated all events but not JAR spheroid adhesion. These results collectively provide evidence suggesting that E2P4- and implanting embryo-induced EMT of endometrial epithelial cells may play a pivotal role in the subsequent processes of human embryo implantation with functional control of N-cadherin. PMID:22174415

  5. Candida albicans: The Ability to Invade Epithelial Cells and Survive under Oxidative Stress Is Unlinked to Hyphal Length

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    Paloma K. Maza

    2017-07-01

    Full Text Available In its hyphal form, Candida albicans invades epithelial and endothelial cells by two distinct mechanisms: active penetration and induced endocytosis. The latter is dependent on a reorganization of the host cytoskeleton (actin/cortactin recruitment, whilst active penetration does not rely on the host's cellular machinery. The first obstacle for the fungus to reach deep tissues is the epithelial barrier and this interaction is crucial for commensal growth, fungal pathogenicity and host defense. This study aimed to characterize in vitro epithelial HeLa cell invasion by four different isolates of C. albicans with distinct clinical backgrounds, including a C. albicans SC5314 reference strain. All isolates invaded HeLa cells, recruited actin and cortactin, and induced the phosphorylation of both Src-family kinases (SFK and cortactin. Curiously, L3881 isolated from blood culture of a patient exhibited the highest resistance to oxidative stress, although this isolate showed reduced hyphal length and displayed the lowest cell damage and invasion rates. Collectively, these data suggest that the ability of C. albicans to invade HeLa cells, and to reach and adapt to the host's blood, including resistance to oxidative stress, may be independent of hyphal length.

  6. Ectopic expression of aquaporin-5 in noncancerous epithelial MDCK cells changes cellular morphology and actin fiber formation without inducing epithelial-to-mesenchymal transition.

    Science.gov (United States)

    Jensen, Helene H; Holst, Mikkel R; Login, Frédéric H; Morgen, Jeanette J; Nejsum, Lene N

    2018-06-01

    Aquaporin-5 (AQP5) is a plasma membrane water channel mainly expressed in secretory glands. Increased expression of AQP5 is observed in multiple cancers, including breast cancer, where high expression correlates with the degree of metastasis and poor prognosis. Moreover, studies in cancer cells have suggested that AQP5 activates Ras signaling, drives morphological changes, and in particular increased invasiveness. To design intervention strategies, it is of utmost importance to characterize and dissect the cell biological changes induced by altered AQP5 expression. To isolate the effect of AQP5 overexpression from the cancer background, AQP5 was overexpressed in normal epithelial MDCK cells which have no endogenous AQP5 expression. AQP5 overexpression promoted actin stress fiber formation and lamellipodia dynamics. Moreover, AQP5 decreased cell circularity. Phosphorylation of AQP5 on serine 156 in the second intracellular loop has been shown to activate the Ras pathway. When serine 156 was mutated to alanine to mimic the nonphosphorylated state, the decrease in cell circularity was reversed, indicating that the AQP5-Ras axis is involved in the effect on cell shape. Interestingly, the cellular changes mediated by AQP5 were not associated with induction of epithelial-to-mesenchymal transition. Thus, AQP5 may contribute to cancer by altering cellular morphology and actin organization, which increase the metastatic potential.

  7. Matrix Metalloproteinase Stromelysin-1 Triggers a Cascade of Molecular Alterations that leads to stable epithelial-to-Mesenchymal Conversion and a Premalignant Phenotype in Mammary Epithelial Cells

    Energy Technology Data Exchange (ETDEWEB)

    Lochter, A.; Galosy, S.; Muschler, J.; Freedman, N.; Werb, Z.; Bissell, M.J.

    1997-08-11

    Matrix metalloproteinases (MMPs) regulate ductal morphogenesis, apoptosis, and neoplastic progression in mammary epithelial cells. To elucidate the direct effects of MMPs on mammary epithelium, we generated functionally normal cells expressing an inducible autoactivating stromelysin-1 (SL-1) transgene. Induction of SL-1 expression resulted in cleavage of E-cadherin, and triggered progressive phenotypic conversion characterized by disappearance of E-cadherin and catenins from cell-cell contacts, downregulation of cytokeratins, upregulation of vimentin, induction of keratinocyte growth factor expression and activation, and upregulation of endogenous MMPs. Cells expressing SL-1 were unable to undergo lactogenic differentiation and became invasive. Once initiated, this phenotypic conversion was essentially stable, and progressed even in the absence of continued SL-1 expression. These observations demonstrate that inappropriate expression of SL-1 initiates a cascade of events that may represent a coordinated program leading to loss of the differentiated epithelial phenotype and gain of some characteristics of tumor cells. Our data provide novel insights into how MMPs function in development and neoplastic conversion.

  8. Proteomic profiling of acrolein adducts in human lung epithelial cells

    Science.gov (United States)

    Spiess, Page C.; Deng, Bin; Hondal, Robert J.; Matthews, Dwight E.; van der Vliet, Albert

    2011-01-01

    Acrolein (2,3-propenal) is a major indoor and outdoor air pollutant originating largely from tobacco smoke or organic combustion. Given its high reactivity, the adverse effects of inhaled acrolein are likely due to direct interactions with the airway epithelium, resulting in altered epithelial function, but only limited information exists to date regarding the primary direct cellular targets for acrolein. Here, we describe a global proteomics approach to characterize the spectrum of airway epithelial protein targets for Michael adduction in acrolein-exposed bronchial epithelial (HBE1) cells, based on biotin hydrazide labeling and avidin purification of biotinylated proteins or peptides for analysis by LC-MS/MS. Identified protein targets included a number of stress proteins, cytoskeletal proteins, and several key proteins involved in redox signaling, including thioredoxin reductase, thioredoxin, peroxiredoxins, and glutathione S-transferase π. Because of the central role of thioredoxin reductase in cellular redox regulation, additional LC-MS/MS characterization was performed on purified mitochondrial thioredoxin reductase to identify the specific site of acrolein adduction, revealing the catalytic selenocysteine residue as the target responsible for enzyme inactivation. Our findings indicate that these approaches are useful in characterizing major protein targets for acrolein, and will enhance mechanistic understanding of the impact of acrolein on cell biology. PMID:21704744

  9. Low Temperature Plasma for the Treatment of Epithelial Cancer Cells

    Science.gov (United States)

    Mohades, Soheila

    Biomedical applications of low temperature plasmas (LTP) may lead to a paradigm shift in treating various diseases by conducting fundamental research on the effects of LTP on cells, tissues, organisms (plants, insects, and microorganisms). This is a rapidly growing interdisciplinary research field that involves engineering, physics, life sciences, and chemistry to find novel solutions for urgent medical needs. Effects of different LTP sources have shown the anti-tumor properties of plasma exposure; however, there are still many unknowns about the interaction of plasma with eukaryotic cells which must be elucidated in order to evaluate the practical potential of plasma in cancer treatment. Plasma, the fourth state of matter, is composed of electrons, ions, reactive molecules (radicals and non-radicals), excited species, radiation, and heat. A sufficient dose (time) of plasma exposure can induce death in cancer cells. The plasma pencil is employed to study the anti-tumor properties of this treatment on epithelial cells. The plasma pencil has been previously used for the inactivation of bacteria, destroying amyloid fibrils, and the killing of various cancer cells. Bladder cancer is the 9th leading cause of cancer. In this dissertation, human urinary bladder tissue with the squamous cell carcinoma disease (SCaBER cells) is treated with LTP utilizing two different approaches: direct plasma exposure and Plasma Activated Media (PAM) as an advancement to the treatment. PAM is produced by exposing a liquid cell culture medium to the plasma pencil. Direct LTP treatment of cancer cells indicates a dose-dependent killing effect at post-treatment times. Similarly, PAM treatment shows an anti-cancer effect by inducing substantial cell death. Reactive oxygen species (ROS) and reactive nitrogen species (RNS) have an important role in the biomedical effects of LTP treatment. This study demonstrates the capability of the plasma pencil to transport ROS/RNS into cell culture media

  10. ATP7B detoxifies silver in ciliated airway epithelial cells

    International Nuclear Information System (INIS)

    Ibricevic, Aida; Brody, Steven L.; Youngs, Wiley J.; Cannon, Carolyn L.

    2010-01-01

    Silver is a centuries-old antibiotic agent currently used to treat infected burns. The sensitivity of a wide range of drug-resistant microorganisms to silver killing suggests that it may be useful for treating refractory lung infections. Toward this goal, we previously developed a methylated caffeine silver acetate compound, SCC1, that exhibits broad-spectrum antimicrobial activity against clinical strains of bacteria in vitro and when nebulized to lungs in mouse infection models. Preclinical testing of high concentrations of SCC1 in primary culture mouse tracheal epithelial cells (mTEC) showed selective ciliated cell death. Ciliated cell death was induced by both silver- and copper-containing compounds but not by the methylated caffeine portion of SCC1. We hypothesized that copper transporting P-type ATPases, ATP7A and ATP7B, play a role in silver detoxification in the airway. In mTEC, ATP7A was expressed in non-ciliated cells, whereas ATP7B was expressed only in ciliated cells. The exposure of mTEC to SCC1 induced the trafficking of ATP7B, but not ATP7A, suggesting the presence of a cell-specific silver uptake and detoxification mechanisms. Indeed, the expression of the copper uptake protein CTR1 was also restricted to ciliated cells. A role of ATP7B in silver detoxification was further substantiated when treatment of SCC1 significantly increased cell death in ATP7B shRNA-treated HepG2 cells. In addition, mTEC from ATP7B -/- mice showed enhanced loss of ciliated cells compared to wild type. These studies are the first to demonstrate a cell type-specific expression of the Ag + /Cu + transporters ATP7A, ATP7B, and CTR1 in airway epithelial cells and a role for ATP7B in detoxification of these metals in the lung.

  11. Studies in human skin epithelial cell carcinogenesis

    International Nuclear Information System (INIS)

    Lehman, T.A.

    1987-01-01

    Metabolism and DNA adduct formation of benzo[a]pyrene (BP) by human epidermal keratinocytes pretreated with inhibitors or inducer of cytochrame P450 was studied. To study DNA adduct analysis, cultures were pretreated as described above, and then treated with non-radiolabeled BP. DNA was prepared from these cultures, digested to the nucleotide level, and 32 P-postlabeled for adduct analysis. Cultures pretreated with BHA, 7,8-BF or disulfiralm formed significantly fewer BPDE I-dB adducts than non-pretreated cultures, while cultures pretreated with MeBHA formed more BPDE-I-dG adducts. MeBHA increased BP activation and adduct formation inhuman keratinocyte in cultures by inducing a specific isoenzyme of cytochrome P450 which preferentially increases the oxidative metabolism of BP to 7,8 diol BP and 7,8 diol BP to BPDE I. To approximate an in vivo human system, metabolism of BPDE I by human skin xenografts treated with cell cycles modulators was studied. When treated with BPDE I, specific carcinogen-DNA adducts were formed. Separation and identification of these adducts by the 32 P-postlabeling technique indicated that the 7R- and 7S-BPDE I-dG adducts were the major adducts

  12. Epithelial cells from smokers modify dendritic cell responses in the context of influenza infection

    Science.gov (United States)

    Epidemiologic evidence suggests that cigarette smoking is a risk factor for infection with influenza, but the mechanisms underlying this susceptibility remain unknown. To ascertain if airway epithelial cells from smokers demonstrate a decreased ability to orchestrate an influenza...

  13. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M

    1998-01-01

    PURPOSE: The immune privilege of the eye has been thought to be dependent on physical barriers and absence of lymphatic vessels. However, the immune privilege may also involve active immunologic processes, as recent studies have indicated. The purpose of the present study was to investigate whether...... human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...

  14. Salinomycin repressed the epithelial–mesenchymal transition of epithelial ovarian cancer cells via downregulating Wnt/β-catenin pathway

    Directory of Open Access Journals (Sweden)

    Li R

    2017-02-01

    Full Text Available Rui Li,* Taotao Dong,* Chen Hu, Jingjing Lu, Jun Dai, Peishu Liu Department of Obstetrics and Gynecology, Qilu Hospital of Shandong University, Jinan, Shandong, People’s Republic of China *These authors contributed equally to this work Abstract: Epithelial ovarian cancer (EOC is the leading cause of death among all gynecological malignancies. Most patients are diagnosed in the advanced stage and have distant metastasis ultimately. Salinomycin has been demonstrated to reduce invasive capacity of multiple tumor cells. The objective of this study was to investigate the effects of salinomycin on EOC cells. The cell counting kit 8 (CCK-8 and Boyden chamber assays showed that salinomycin could effectively reduce the abilities of proliferation, migration and invasion in EOC cells. The western blot assay showed that salinomycin could increase the expression of epithelial markers (E-cadherin and Keratin while decrease the expression of mesenchymal markers (N-cadherin and vimentin in a dose-dependent manner. These results were ascertained by reverse transcription polymerase chain reaction (RT-PCR. Besides, salinomycin could downregulate the expression of proteins associated with the Wnt/β-catenin pathway and repress the nuclear translocation of β-catenin. It was also shown that salinomycin could reverse the aberrant activation of the canonical Wnt pathway induced by GSK-3β inhibitor (SB216763. Our results revealed that salinomycin could inhibit the proliferation, migration and invasion in EOC cells. In addition, the inhibitive effect of salinomycin on the invasive ability was mediated by repressing the epithelial–mesenchymal transition (EMT program, which may be achieved through its inhibition of the Wnt/β-catenin pathway. Keywords: salinomycin, epithelial–mesenchymal transition, epithelial ovarian cancer, Wnt/β-catenin pathway

  15. UV-induced changes in cell cycle and gene expression within rabbit lens epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Sidjanin, D. [Northern Illinois Univ., De Kalb, IL (United States). Dept. of Biological Sciences; Grdina, D. [Argonne National Lab., IL (United States); Woloschak, G.E. [Northern Illinois Univ., De Kalb, IL (United States). Dept. of Biological Sciences

    1994-11-01

    Damage to lens epithelial cells is a probable initiation process in cataract formation induced by ultraviolet radiation. These experiments investigated the ability of 254 nm radiation on cell cycle progression and gene expression in rabbit lens epithelial cell line N/N1003A. No changes in expression of c-fos, c-jun, alpha- tubulin, or vimentin was observed following UV exposure. Using flow cytometry, an accumulation of cells in G1/S phase of the cell cycle 1 hr following exposure. The observed changes in gene expression, especially the decreased histone transcripts reported here may play a role in UV induced inhibition of cell cycle progression.

  16. Inhibition of EV71 by curcumin in intestinal epithelial cells.

    Science.gov (United States)

    Huang, Hsing-I; Chio, Chi-Chong; Lin, Jhao-Yin

    2018-01-01

    EV71 is a positive-sense single-stranded RNA virus that belongs to the Picornaviridae family. EV71 infection may cause various symptoms ranging from hand-foot-and-mouth disease to neurological pathological conditions such as aseptic meningitis, ataxia, and acute transverse myelitis. There is currently no effective treatment or vaccine available. Various compounds have been examined for their ability to restrict EV71 replication. However, most experiments have been performed in rhabdomyosarcoma or Vero cells. Since the gastrointestinal tract is the entry site for this pathogen, we anticipated that orally ingested agents may exert beneficial effects by decreasing virus replication in intestinal epithelial cells. In this study, curcumin (diferuloylmethane, C21H20O6), an active ingredient of turmeric (Curcuma longa Linn) with anti-cancer properties, was investigated for its anti-enterovirus activity. We demonstrate that curcumin treatment inhibits viral translation and increases host cell viability. Curcumin does not exert its anti-EV71 effects by modulating virus attachment or virus internal ribosome entry site (IRES) activity. Furthermore, curcumin-mediated regulation of mitogen-activated protein kinase (MAPK) signaling pathways is not involved. We found that protein kinase C delta (PKCδ) plays a role in virus translation in EV71-infected intestinal epithelial cells and that curcumin treatment decreases the phosphorylation of this enzyme. In addition, we show evidence that curcumin also limits viral translation in differentiated human intestinal epithelial cells. In summary, our data demonstrate the anti-EV71 properties of curcumin, suggesting that ingestion of this phytochemical may protect against enteroviral infections.

  17. Inhibition of EV71 by curcumin in intestinal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Hsing-I Huang

    Full Text Available EV71 is a positive-sense single-stranded RNA virus that belongs to the Picornaviridae family. EV71 infection may cause various symptoms ranging from hand-foot-and-mouth disease to neurological pathological conditions such as aseptic meningitis, ataxia, and acute transverse myelitis. There is currently no effective treatment or vaccine available. Various compounds have been examined for their ability to restrict EV71 replication. However, most experiments have been performed in rhabdomyosarcoma or Vero cells. Since the gastrointestinal tract is the entry site for this pathogen, we anticipated that orally ingested agents may exert beneficial effects by decreasing virus replication in intestinal epithelial cells. In this study, curcumin (diferuloylmethane, C21H20O6, an active ingredient of turmeric (Curcuma longa Linn with anti-cancer properties, was investigated for its anti-enterovirus activity. We demonstrate that curcumin treatment inhibits viral translation and increases host cell viability. Curcumin does not exert its anti-EV71 effects by modulating virus attachment or virus internal ribosome entry site (IRES activity. Furthermore, curcumin-mediated regulation of mitogen-activated protein kinase (MAPK signaling pathways is not involved. We found that protein kinase C delta (PKCδ plays a role in virus translation in EV71-infected intestinal epithelial cells and that curcumin treatment decreases the phosphorylation of this enzyme. In addition, we show evidence that curcumin also limits viral translation in differentiated human intestinal epithelial cells. In summary, our data demonstrate the anti-EV71 properties of curcumin, suggesting that ingestion of this phytochemical may protect against enteroviral infections.

  18. Inhibition of EV71 by curcumin in intestinal epithelial cells

    Science.gov (United States)

    Chio, Chi-Chong; Lin, Jhao-Yin

    2018-01-01

    EV71 is a positive-sense single-stranded RNA virus that belongs to the Picornaviridae family. EV71 infection may cause various symptoms ranging from hand-foot-and-mouth disease to neurological pathological conditions such as aseptic meningitis, ataxia, and acute transverse myelitis. There is currently no effective treatment or vaccine available. Various compounds have been examined for their ability to restrict EV71 replication. However, most experiments have been performed in rhabdomyosarcoma or Vero cells. Since the gastrointestinal tract is the entry site for this pathogen, we anticipated that orally ingested agents may exert beneficial effects by decreasing virus replication in intestinal epithelial cells. In this study, curcumin (diferuloylmethane, C21H20O6), an active ingredient of turmeric (Curcuma longa Linn) with anti-cancer properties, was investigated for its anti-enterovirus activity. We demonstrate that curcumin treatment inhibits viral translation and increases host cell viability. Curcumin does not exert its anti-EV71 effects by modulating virus attachment or virus internal ribosome entry site (IRES) activity. Furthermore, curcumin-mediated regulation of mitogen-activated protein kinase (MAPK) signaling pathways is not involved. We found that protein kinase C delta (PKCδ) plays a role in virus translation in EV71-infected intestinal epithelial cells and that curcumin treatment decreases the phosphorylation of this enzyme. In addition, we show evidence that curcumin also limits viral translation in differentiated human intestinal epithelial cells. In summary, our data demonstrate the anti-EV71 properties of curcumin, suggesting that ingestion of this phytochemical may protect against enteroviral infections. PMID:29370243

  19. Loss of prostasin (PRSS8) in human bladder transitional cell carcinoma cell lines is associated with epithelial-mesenchymal transition (EMT)

    International Nuclear Information System (INIS)

    Chen, Li-Mei; Verity, Nicole J; Chai, Karl X

    2009-01-01

    The glycosylphosphatidylinositol (GPI)-anchored epithelial extracellular membrane serine protease prostasin (PRSS8) is expressed abundantly in normal epithelia and essential for terminal epithelial differentiation, but down-regulated in human prostate, breast, and gastric cancers and invasive cancer cell lines. Prostasin is involved in the extracellular proteolytic modulation of the epidermal growth factor receptor (EGFR) and is an invasion suppressor. The aim of this study was to evaluate prostasin expression states in the transitional cell carcinomas (TCC) of the human bladder and in human TCC cell lines. Normal human bladder tissues and TCC on a bladder cancer tissue microarray (TMA) were evaluated for prostasin expression by means of immunohistochemistry. A panel of 16 urothelial and TCC cell lines were evaluated for prostasin and E-cadherin expression by western blot and quantitative PCR, and for prostasin gene promoter region CpG methylation by methylation-specific PCR (MSP). Prostasin is expressed in the normal human urothelium and in a normal human urothelial cell line, but is significantly down-regulated in high-grade TCC and lost in 9 (of 15) TCC cell lines. Loss of prostasin expression in the TCC cell lines correlated with loss of or reduced E-cadherin expression, loss of epithelial morphology, and promoter DNA hypermethylation. Prostasin expression could be reactivated by demethylation or inhibition of histone deacetylase. Re-expression of prostasin or a serine protease-inactive variant resulted in transcriptional up-regulation of E-cadherin. Loss of prostasin expression in bladder transitional cell carcinomas is associated with epithelial-mesenchymal transition (EMT), and may have functional implications in tumor invasion and resistance to chemotherapy

  20. Loss of prostasin (PRSS8 in human bladder transitional cell carcinoma cell lines is associated with epithelial-mesenchymal transition (EMT

    Directory of Open Access Journals (Sweden)

    Chai Karl X

    2009-10-01

    Full Text Available Abstract Background The glycosylphosphatidylinositol (GPI-anchored epithelial extracellular membrane serine protease prostasin (PRSS8 is expressed abundantly in normal epithelia and essential for terminal epithelial differentiation, but down-regulated in human prostate, breast, and gastric cancers and invasive cancer cell lines. Prostasin is involved in the extracellular proteolytic modulation of the epidermal growth factor receptor (EGFR and is an invasion suppressor. The aim of this study was to evaluate prostasin expression states in the transitional cell carcinomas (TCC of the human bladder and in human TCC cell lines. Methods Normal human bladder tissues and TCC on a bladder cancer tissue microarray (TMA were evaluated for prostasin expression by means of immunohistochemistry. A panel of 16 urothelial and TCC cell lines were evaluated for prostasin and E-cadherin expression by western blot and quantitative PCR, and for prostasin gene promoter region CpG methylation by methylation-specific PCR (MSP. Results Prostasin is expressed in the normal human urothelium and in a normal human urothelial cell line, but is significantly down-regulated in high-grade TCC and lost in 9 (of 15 TCC cell lines. Loss of prostasin expression in the TCC cell lines correlated with loss of or reduced E-cadherin expression, loss of epithelial morphology, and promoter DNA hypermethylation. Prostasin expression could be reactivated by demethylation or inhibition of histone deacetylase. Re-expression of prostasin or a serine protease-inactive variant resulted in transcriptional up-regulation of E-cadherin. Conclusion Loss of prostasin expression in bladder transitional cell carcinomas is associated with epithelial-mesenchymal transition (EMT, and may have functional implications in tumor invasion and resistance to chemotherapy.

  1. Epithelial Cell Inflammasomes in Intestinal Immunity and Inflammation

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    Andrea C. Lei-Leston

    2017-09-01

    Full Text Available Pattern recognition receptors (PRR, such as NOD-like receptors (NLRs, sense conserved microbial signatures, and host danger signals leading to the coordination of appropriate immune responses. Upon activation, a subset of NLR initiate the assembly of a multimeric protein complex known as the inflammasome, which processes pro-inflammatory cytokines and mediates a specialized form of cell death known as pyroptosis. The identification of inflammasome-associated genes as inflammatory bowel disease susceptibility genes implicates a role for the inflammasome in intestinal inflammation. Despite the fact that the functional importance of inflammasomes within immune cells has been well established, the contribution of inflammasome expression in non-hematopoietic cells remains comparatively understudied. Given that intestinal epithelial cells (IEC act as a barrier between the host and the intestinal microbiota, inflammasome expression by these cells is likely important for intestinal immune homeostasis. Accumulating evidence suggests that the inflammasome plays a key role in shaping epithelial responses at the host–lumen interface with many inflammasome components highly expressed by IEC. Recent studies have exposed functional roles of IEC inflammasomes in mucosal immune defense, inflammation, and tumorigenesis. In this review, we present the main features of the predominant inflammasomes and their effector mechanisms contributing to intestinal homeostasis and inflammation. We also discuss existing controversies in the field and open questions related to their implications in disease. A comprehensive understanding of the molecular basis of intestinal inflammasome signaling could hold therapeutic potential for clinical translation.

  2. Reconstitution of mammary epithelial morphogenesis by murine embryonic stem cells undergoing hematopoietic stem cell differentiation.

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    Shuxian Jiang

    2010-03-01

    Full Text Available Mammary stem cells are maintained within specific microenvironments and recruited throughout lifetime to reconstitute de novo the mammary gland. Mammary stem cells have been isolated through the identification of specific cell surface markers and in vivo transplantation into cleared mammary fat pads. Accumulating evidence showed that during the reformation of mammary stem cell niches by dispersed epithelial cells in the context of the intact epithelium-free mammary stroma, non-mammary epithelial cells may be sequestered and reprogrammed to perform mammary epithelial cell functions and to adopt mammary epithelial characteristics during reconstruction of mammary epithelium in regenerating mammary tissue in vivo.To examine whether other types of progenitor cells are able to contribute to mammary branching morphogenesis, we examined the potential of murine embryonic stem (mES cells, undergoing hematopoietic differentiation, to support mammary reconstitution in vivo. We observed that cells from day 14 embryoid bodies (EBs under hematopoietic differentiation condition, but not supernatants derived from these cells, when transplanted into denuded mammary fat pads, were able to contribute to both the luminal and myoepithelial lineages in branching ductal structures resembling the ductal-alveolar architecture of the mammary tree. No teratomas were observed when these cells were transplanted in vivo.Our data provide evidence for the dominance of the tissue-specific mammary stem cell niche and its role in directing mES cells, undergoing hematopoietic differentiation, to reprogram into mammary epithelial cells and to promote mammary epithelial morphogenesis. These studies should also provide insights into regeneration of damaged mammary gland and the role of the mammary microenvironment in reprogramming cell fate.

  3. Downregulation of Connective Tissue Growth Factor by Three-Dimensional Matrix Enhances Ovarian Carcinoma Cell Invasion

    Science.gov (United States)

    Barbolina, Maria V.; Adley, Brian P.; Kelly, David L.; Shepard, Jaclyn; Fought, Angela J.; Scholtens, Denise; Penzes, Peter; Shea, Lonnie D.; Sharon Stack, M

    2010-01-01

    Epithelial ovarian carcinoma (EOC) is a leading cause of death from gynecologic malignancy, due mainly to the prevalence of undetected metastatic disease. The process of cell invasion during intra-peritoneal anchoring of metastatic lesions requires concerted regulation of many processes, including modulation of adhesion to the extracellular matrix and localized invasion. Exploratory cDNA microarray analysis of early response genes (altered after 4 hours of 3-dimensional collagen culture) coupled with confirmatory real-time RT-PCR, multiple three-dimensional cell culture matrices, Western blot, immunostaining, adhesion, migration, and invasion assays were used to identify modulators of adhesion pertinent to EOC progression and metastasis. cDNA microarray analysis indicated a dramatic downregulation of connective tissue growth factor (CTGF) in EOC cells placed in invasion-mimicking conditions (3-dimensional type I collagen). Examination of human EOC specimens revealed that CTGF expression was absent in 46% of the tested samples (n=41), but was present in 100% of normal ovarian epithelium samples (n=7). Reduced CTGF expression occurs in many types of cells and may be a general phenomenon displayed by cells encountering a 3D environment. CTGF levels were inversely correlated with invasion such that downregulation of CTGF increased, while its upregulation reduced, collagen invasion. Cells adhered preferentially to a surface comprised of both collagen I and CTGF relative to either component alone using α6β1 and α3β1 integrins. Together these data suggest that downregulation of CTGF in EOC cells may be important for cell invasion through modulation of cell-matrix adhesion. PMID:19382180

  4. Potential of confocal laser scanning microscopy for non-invasive diagnostics of malignant epithelial skin tumors in the course of dermatoheliosis progression

    Directory of Open Access Journals (Sweden)

    E. S. Snarskaya

    2016-01-01

    Full Text Available Most cases of malignant epithelial skin neoplasms including actinic keratosis and basal cell carcinoma, which are characterized by the most complicated course and numerous clinical and morphological options, involve dermatoheliosis progression. The risk of actinic keratosis transformation into basal cell carcinoma varies from 0.1% to 20% and up to 80% in cases of multiple AK lesion foci. A non-invasive method known as reflectance confocal laser scanning microscopy is the most promising one for the purposes of early diagnostics of signs pointing at epithelial skin neoplasm development and makes it possible to monitor the tumor in progress in vivo to diagnose the presence of a pool of squamous cells on a timely basis. The confocal laser scanning microscopy method provides high-contrast images of for any horizontal-oriented morphologic structures in the epidermis and upper dermis with a resolution comparable to those characteristic of traditional optical microscopy of skin tissue samples. According to our data obtained as a result of studying dynamic changes and morphologic structures in actinic keratosis foci (50 cases using the confocal laser scanning microscopy method, we discovered a number of morphologic features, and their further analysis will distinguish the signs of progressing carcinogenesis in case of dermatoheliosis.

  5. Sublethal irradiation promotes invasiveness of neuroblastoma cells

    International Nuclear Information System (INIS)

    Schweigerer, Lothar; Rave-Fraenk, Margret; Schmidberger, Heinz; Hecht, Monica

    2005-01-01

    Neuroblastoma is the most frequent extracranial solid tumour of childhood. Despite multiple clinical efforts, clinical outcome has remained poor. Neuroblastoma is considered to be radiosensitive, but some clinical studies including the German trial NB90 failed to show a clinical benefit of radiation therapy. The mechanisms underlying this apparent discrepancy are still unclear. We have therefore investigated the effects of radiation on neuroblastoma cell behaviour in vitro. We show that sublethal doses of irradiation up-regulated the expression of the hepatocyte growth factor (HGF) and its receptor c-Met in some neuroblastoma cell lines. The increase in HGF/c-Met expression was correlated with enhanced invasiveness and activation of proteases degrading the extracellular matrix. Thus, irradiation at sublethal doses may promote the metastatic dissemination of neuroblastoma cells through activating the HGF/c-Met pathway and triggering matrix degradation

  6. Colonic epithelial cell activation and the paradoxical effects of butyrate.

    Science.gov (United States)

    Gibson, P R; Rosella, O; Wilson, A J; Mariadason, J M; Rickard, K; Byron, K; Barkla, D H

    1999-04-01

    Butyrate may have paradoxical effects on epithelial cells of similar origin. This study aimed to examine the hypothesis that one mechanism that dictates a cell's response to butyrate is its state of activation. First, the responses to 24 h exposure to butyrate (1-2 mM) of normal and neoplastic human colonic epithelial cells activated by their isolation and primary culture, and of colon cancer cell lines, LIM1215 and Caco-2, were examined. In primary cultures of normal and cancer cells, butyrate had no effect on alkaline phosphatase activities but significantly suppressed urokinase receptor expression by a mean +/- SEM of 30 +/- 12% and 36 +/- 9%, respectively. Interleukin-8 secretion was suppressed by 44 +/- 7% in normal cells (P 50%, urokinase receptor expression >2-fold and interleukin-8 secretion >3-fold in response to butyrate. Secondly, the effect of butyrate on Caco-2 cells was examined with or without prior exposure to a specific activating stimulus [tumour necrosis factor alpha (TNF alpha)]. Interleukin-8 secretion increased by 145 +/- 23% and 132 +/- 17% on 24 h exposure to 2 mM butyrate or 0.1 microM TNF alpha alone, respectively. However, in cells pre-treated with TNF alpha, butyrate significantly inhibited secretion by 34 +/- 7% below unstimulated levels. The response to butyrate of urokinase receptor, whose expression was not stimulated by TNF alpha, was unchanged. These effects were mimicked by trichostatin A, an inhibitor of histone deacetylase, suggesting that butyrate's paradoxical effects may have been operating by the same mechanism. In conclusion, some of the paradoxical effects of butyrate do not appear to represent inherent differences between normal and transformed cells. Rather, the response may be determined by the state of activation of the cells.

  7. Knockdown of BAG3 induces epithelial-mesenchymal transition in thyroid cancer cells through ZEB1 activation.

    Science.gov (United States)

    Meng, X; Kong, D-H; Li, N; Zong, Z-H; Liu, B-Q; Du, Z-X; Guan, Y; Cao, L; Wang, H-Q

    2014-02-27

    The process by which epithelial features are lost in favor of a mesenchymal phenotype is referred to as epithelial-mesenchymal transition (EMT). Most carcinomas use this mechanism to evade into neighboring tissues. Reduction or a loss of E-cadherin expression is a well-established hallmark of EMT. As a potent suppressor of E-cadherin, transcription factor ZEB1 is one of the key inducers of EMT, whose expression promotes tumorigenesis and metastasis of carcinomas. Bcl-2-associated athanogene 3 (BAG3) affects multifaceted cellular functions, including proliferation, apoptosis, cell adhesion and invasion, viral infection, and autophagy. Recently, we have reported a novel role of BAG3 implicated in EMT, while the mechanisms are poorly elucidated. The current study demonstrated that knockdown of BAG3 induced EMT, and increased cell migratory and invasiveness in thyroid cancer cells via transcriptional activation of ZEB1. We also found that BAG3 knockdown led to nuclear accumulation of β-catenin, which was responsible for the transcriptional activation of ZEB1. These results indicate BAG3 as a regulator of ZEB1 expression in EMT and as a regulator of metastasis in thyroid cancer cells, providing potential targets to prevent and/or treat thyroid cancer cell invasion and metastasis.

  8. Milk modulates campylobacter invasion into caco-2 intestinal epithelial cells

    NARCIS (Netherlands)

    Louwen, R.; Neerven, van R.J.J.

    2015-01-01

    Raw milk is a recognized source of Campylobacter outbreaks, but pasteurization is an effective way to eliminate the causative agent of Campylobacteriosis. Whereas breastfeeding is protective against infectious diseases, consumption of formula milk is thought to be not. However, in relation to

  9. MicroRNA and protein profiles in invasive versus non-invasive oral tongue squamous cell carcinoma cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Korvala, Johanna, E-mail: johanna.korvala@oulu.fi [Cancer and Translational Medicine Research Unit, University of Oulu, The Medical Research Center Oulu, Oulu University Hospital, Aapistie 5A, 90014 Oulu (Finland); Jee, Kowan [Department of Pathology, University of Turku, Turku University Hospital, Turku (Finland); Department of Pathology, Haartman Institute, University of Helsinki, Helsinki (Finland); Porkola, Emmi [Cancer and Translational Medicine Research Unit, University of Oulu, The Medical Research Center Oulu, Oulu University Hospital, Aapistie 5A, 90014 Oulu (Finland); Almangush, Alhadi [Department of Pathology, Haartman Institute, University of Helsinki, Helsinki (Finland); Mosakhani, Neda [Department of Pathology, HUSLAB, Helsinki (Finland); Bitu, Carolina [Cancer and Translational Medicine Research Unit, University of Oulu, The Medical Research Center Oulu, Oulu University Hospital, Aapistie 5A, 90014 Oulu (Finland); Cervigne, Nilva K. [Department of Oral Diagnosis, School of Dentistry, University of Campinas (UNICAMP), Av. Limeira, 901 – Bairro Areião, CEP: 13414-903 Piracicaba, São Paulo (Brazil); Department of Clinical and Pathology, Faculty of Medicine of Jundiai - FMJ, Jundiai, SP (Brazil); Zandonadi, Flávia S.; Meirelles, Gabriela V.; Leme, Adriana Franco Paes [Laboratório Nacional de Biociências, LNBio, CNPEM, Rua Giuseppe Máximo Scolfaro, 10.000, Polo II de Alta Tecnologia de Campinas, Campinas/SP, P.O.Box 6192, CEP 13083-970 Campinas, São Paulo (Brazil); Coletta, Ricardo D. [Department of Oral Diagnosis, School of Dentistry, University of Campinas (UNICAMP), Av. Limeira, 901 – Bairro Areião, CEP: 13414-903 Piracicaba, São Paulo (Brazil); and others

    2017-01-01

    Complex molecular pathways regulate cancer invasion. This study overviewed proteins and microRNAs (miRNAs) involved in oral tongue squamous cell carcinoma (OTSCC) invasion. The human highly aggressive OTSCC cell line HSC-3 was examined in a 3D organotypic human leiomyoma model. Non-invasive and invasive cells were laser-captured and protein expression was analyzed using mass spectrometry-based proteomics and miRNA expression by microarray. In functional studies the 3D invasion assay was replicated after silencing candidate miRNAs, miR-498 and miR-940, in invasive OTSCC cell lines (HSC-3 and SCC-15). Cell migration, proliferation and viability were also studied in the silenced cells. In HSC-3 cells, 67 proteins and 53 miRNAs showed significant fold-changes between non-invasive vs. invasive cells. Pathway enrichment analyses allocated “Focal adhesion” and “ECM-receptor interaction” as most important for invasion. Significantly, in HSC-3 cells, miR-498 silencing decreased the invasion area and miR-940 silencing reduced invasion area and depth. Viability, proliferation and migration weren’t significantly affected. In SCC-15 cells, down-regulation of miR-498 significantly reduced invasion and migration. This study shows HSC-3 specific miRNA and protein expression in invasion, and suggests that miR-498 and miR-940 affect invasion in vitro, the process being more influenced by mir-940 silencing in aggressive HSC-3 cells than in the less invasive SCC-15.

  10. Epithelial-to-mesenchymal plasticity of cancer stem cells: therapeutic targets in hepatocellular carcinoma

    Directory of Open Access Journals (Sweden)

    Aparna Jayachandran

    2016-08-01

    Full Text Available Abstract Hepatocellular carcinoma (HCC remains one of the most common and lethal malignancies worldwide despite the development of various therapeutic strategies. A better understanding of the mechanisms responsible for HCC initiation and progression is essential for the development of more effective therapies. The cancer stem cell (CSC model has provided new insights into the development and progression of HCC. CSCs are specialized tumor cells that are capable of self-renewal and have long-term repopulation potential. As they are important mediators of tumor proliferation, invasion, metastasis, therapy resistance, and cancer relapse, the selective targeting of this crucial population of cells has the potential to improve HCC patient outcomes and survival. In recent years, the role of epithelial-to-mesenchymal transition (EMT in the advancement of HCC has gained increasing attention. This multi-step reprograming process resulting in a phenotype switch from an epithelial to a mesenchymal cellular state has been closely associated with the acquisition of stem cell-like attributes in tumors. Moreover, CSC mediates tumor metastasis by maintaining plasticity to transition between epithelial or mesenchymal states. Therefore, understanding the molecular mechanisms of the reprograming switches that determine the progression through EMT and generation of CSC is essential for developing clinically relevant drug targets. This review provides an overview of the proposed roles of CSC in HCC and discusses recent results supporting the emerging role of EMT in facilitating hepatic CSC plasticity. In particular, we discuss how these important new insights may facilitate rational development of combining CSC- and EMT-targeted therapies in the future.

  11. Immunoregulation by airway epithelial cells (AECs against respiratory virus infection

    Directory of Open Access Journals (Sweden)

    Yan YAN

    2017-11-01

    Full Text Available The respiratory tract is primary contact site of the body and environment, and it is ventilated by 10-20 thousand liters of air per day. Inevitably, the respiratory system comes into contact with airborne microbes, which contain the disease-causing pathogens. Airway epithelial cells (AECs are known to have innate sensor functions, which are similar to the "professional" immune cells, such as alveolar macrophage and sub- or intra-epithelial dendritic cells (DCs. Thus AECs are able to detect invading microbial danger including different types of respiratory viruses, and mount a potent host response, for example, activating type Ⅰ interferon signaling pathway genes. To avoid chronic inflammation and maintain the immunological homeostasis, the pulmonary system has developed intrinsic mechanisms to control local immune responses. Most recently, the role of AECs in control of local immunity has gained much attention, as 1 AECs express the pattern recognition receptors (PRRs, such as Toll-like receptors, retinoic acid inducible gene Ⅰ (RIG-I-like receptor, and so on, thus AECs are equipped to participate in innate detection of microbial encounter; 2 To keep immunological homeostasis in the respiratory tract, AECs behave not only as innate immune sensors but also as immune modulators in parallel, through modulating the sensitivity of innate immune sensing of both AECs per se and sub- or intra-epithelial immune cells; 3 Loss of modularity capacity of AECs might be involved in the development of chronic airway diseases. In present review, how the AECs act will be intensively discussed in response to respiratory viruses and modulate the local immunity through cis- and trans-factors (direct and indirect factors, as well as the consequence of impairment of this control of local immunity, in the development and exacerbation of airway diseases, such as acute and chronic rhinosinusitis. DOI: 10.11855/j.issn.0577-7402.2017.10.02

  12. Emodin suppresses TGF-β1-induced epithelial-mesenchymal transition in alveolar epithelial cells through Notch signaling pathway

    International Nuclear Information System (INIS)

    Gao, Rundi; Chen, Ruilin; Cao, Yu; Wang, Yuan; Song, Kang; Zhang, Ya; Yang, Junchao

    2017-01-01

    Pulmonary fibrosis is characterized by the destruction of lung tissue architecture and the formation of fibrous foci, currently has no satisfactory treatment. Emodin is a component of Chinese herb that has been reported to be medicament on pancreatic fibrosis and liver fibrosis. However, its role in pulmonary fibrosis has not been established yet. In the present study, we investigated the hypothesis that Emodin plays an inhibitory role in TGF-β1 induced epithelial-mesenchymal transition (EMT) of alveolar epithelial cell, and Emodin exerts its effect through the Notch signaling pathway. Emodin inhibits the proliferation of Rat alveolar type II epithelial cells RLE-6TN in a concentration-dependent manner; reduces the expression of Collagen I, α-SMA and Vimentin, promotes the expression of E-cadherin. Moreover, Emodin could regulate the expression patterns of the Notch signaling pathway-related factors and reduce the Notch-1 nucleus translocation. Knockdown of Notch-1 enhances the inhibitory effect of Emodin on TGF-β1-induced EMT in RLE-6TN cells. In conclusion, the data of the present study suggests that Emodin suppresses TGF-β1-induced EMT in alveolar epithelial cells through Notch signaling pathway and shows the potential to be effective in the treatment of pulmonary fibrosis. - Highlights: • Emodin inhibits TGF-β1-induced EMT in alveolar epithelial cells. • Emodin regulates the expression patterns of the Notch signaling pathway-related factors. • Emodin inhibits TGF-β1-induced Notch-1 nucleus translocation and activation.

  13. Emodin suppresses TGF-β1-induced epithelial-mesenchymal transition in alveolar epithelial cells through Notch signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Rundi; Chen, Ruilin; Cao, Yu [Department of Respiration, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China); Wang, Yuan [Department of Pulmonary Function, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China); Song, Kang [Department of Respiration, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China); Zhang, Ya [Zhejiang Chinese Medicine University, No. 548, Binwen Road, Binjiang District, Hangzhou, Zhejiang Province 310006 (China); Yang, Junchao, E-mail: yangjunchaozj@zcmu.edu.cn [Department of Respiration, The First Affiliated Hospital of Zhejiang Chinese Medicine University, NO. 56, Youdian Road, Shangcheng District, Hangzhou, Zhejiang Province 310006 (China)

    2017-03-01

    Pulmonary fibrosis is characterized by the destruction of lung tissue architecture and the formation of fibrous foci, currently has no satisfactory treatment. Emodin is a component of Chinese herb that has been reported to be medicament on pancreatic fibrosis and liver fibrosis. However, its role in pulmonary fibrosis has not been established yet. In the present study, we investigated the hypothesis that Emodin plays an inhibitory role in TGF-β1 induced epithelial-mesenchymal transition (EMT) of alveolar epithelial cell, and Emodin exerts its effect through the Notch signaling pathway. Emodin inhibits the proliferation of Rat alveolar type II epithelial cells RLE-6TN in a concentration-dependent manner; reduces the expression of Collagen I, α-SMA and Vimentin, promotes the expression of E-cadherin. Moreover, Emodin could regulate the expression patterns of the Notch signaling pathway-related factors and reduce the Notch-1 nucleus translocation. Knockdown of Notch-1 enhances the inhibitory effect of Emodin on TGF-β1-induced EMT in RLE-6TN cells. In conclusion, the data of the present study suggests that Emodin suppresses TGF-β1-induced EMT in alveolar epithelial cells through Notch signaling pathway and shows the potential to be effective in the treatment of pulmonary fibrosis. - Highlights: • Emodin inhibits TGF-β1-induced EMT in alveolar epithelial cells. • Emodin regulates the expression patterns of the Notch signaling pathway-related factors. • Emodin inhibits TGF-β1-induced Notch-1 nucleus translocation and activation.

  14. Proliferation of Prostate Stromal Cell Induced by Benign Prostatic Hyperplasia Epithelial Cell Stimulated With Trichomonas vaginalis via Crosstalk With Mast Cell.

    Science.gov (United States)

    Kim, Jung-Hyun; Kim, Sang-Su; Han, Ik-Hwan; Sim, Seobo; Ahn, Myoung-Hee; Ryu, Jae-Sook

    2016-11-01

    Chronic inflammation has a role in the pathogenesis of benign prostatic hyperplasia (BPH) and prostate cancer. Mast cells have been detected in chronic inflammatory infiltrate of the prostate, and it is possible that the interaction between prostate epithelial cells and Trichomonas vaginalis influences the activity of mast cells in the prostate stroma. Activated mast cells might influence the biological functions of nearby tissues and cells. In this study, we investigated whether mast cells reacted with the culture supernatant of BPH epithelial cells infected with T. vaginalis may induce the proliferation of prostate stromal cells. To measure the proliferation of prostate stromal cells in response to chronic inflammation caused by the infection of BPH-1 cells with T. vaginalis, the CCK-8 assay and wound healing assay were used. ELISAs, quantitative real-time PCR, western blotting and immunofluorescence were used to measure the production and expression of inflammatory cytokine and cytokine receptor. BPH-1 cells incubated with live trichomonads produced increased levels of CCL2, IL-1β, IL-6, and CXCL8, and induced the migration of mast cells and monocytes. When the culture supernatant of BPH-1 cells stimulated with trichomonads (TCM) was added to mast cells, they became activated, as confirmed by release of β-hexosaminidase and CXCL8. Prostate stromal cells incubated with the culture supernatant of mast cells activated with TCM (M-TCM) proliferated and expressed increased levels of CXCL8, CCL2, and the cytokine receptors CXCR1 and CCR2. Blocking the chemokine receptors reduced the proliferation of stromal cells and also decreased the production of CXCL8 and CCL2. Moreover, the expression of FGF2, cyclin D1, and Bcl-2 was increased in the proliferated stromal cells stimulated with M-TCM. Additionally, the M-TCM-treated stromal cells were more invasive than control cells. The inflammatory mediators released by BPH epithelial cells in response to infection by

  15. An In-Vitro Model of Ciprofloxacin and Minocycline Transport by Oral Epithelial Cells

    Science.gov (United States)

    Brayton, James J.; Yang, Qing; Nakkula, Robin J.; Walters, John D.

    2008-01-01

    Background Fluoroquinolones and tetracyclines can penetrate epithelial cells, but the mechanism by which they cross the plasma membrane is unclear. In this study, a cell line derived from oral epithelium was used as a model to demonstrate a role for active transport. Methods Transport of ciprofloxacin and minocycline by confluent cell monolayers was assayed by measuring the increase in cell-associated fluorescence. Results Uptake of both agents was saturable and was inhibited at low temperatures. At 37° C, the cells transported ciprofloxacin and minocycline with Km values of 351 and 133 μg/ml, respectively, and maximum velocities of 5.11 and 13.4 ng/min/μg cell protein, respectively. When ciprofloxacin and minocycline were removed from the extracellular medium, the intracellular levels of both agents decreased. Ciprofloxacin efflux from loaded cells occurred more rapidly than with minocycline. Cells accumulated intracellular drug levels that were at least 8-fold higher than extracellular levels for ciprofloxacin and at least 40-fold higher for minocycline. Transport of ciprofloxacin and minocycline was significantly influenced by pH and was most favorable at pH 7.7 and 7.2, respectively. While ciprofloxacin transport was Na+-independent, minocycline transport was strongly inhibited when sodium in the medium was replaced with choline. Transport of both agents was inhibited by a variety of organic cations, but the pattern of inhibition was different. Papaverine, phenylephrine and doxycycline competitively inhibited minocycline transport, but inhibited ciprofloxacin transport by a noncompetitive mechanism. Conclusions Epithelial cells take up ciprofloxacin and minocycline via different active transport systems. These transporters may play an important role in enhancing the effectiveness of these agents against invasive pathogens. PMID:12479629

  16. Inflammasome-independent NLRP3 is required for epithelial-mesenchymal transition in colon cancer cells.

    Science.gov (United States)

    Wang, Hong; Wang, Yajing; Du, Qianming; Lu, Ping; Fan, Huimin; Lu, Jinrong; Hu, Rong

    2016-03-15

    Inflammasome NLRP3 plays a crucial role in the process of colitis and colitis--associated colon cancer. Even though much is known regarding the NLRP3 inflammasome that regulates pro-inflammatory cytokine release in innate immune cells, the role of NLRP3 in non-immune cells is still unclear. In this study, we showed that NLRP3 was highly expressed in mesenchymal-like colon cancer cells (SW620), and was upregulated by tumor necrosis factors-α (TNF-α) and transforming growth factor-β1 (TGF-β1) respectively, during EMT in colon cancer epithelial cells HCT116 and HT29. Knockdown of NLRP3 retained epithelial spindle-like morphology of HCT116 and HT29 cells and reversed the mesenchymal characteristic of SW620 cells, indicated by the decreased expression of vimentin and MMP9 and increased expression of E-cadherin. In addition, knockdown of NLRP3 in colorectal carcinoma cells displayed diminished cell migration and invasion. Interestingly, during the EMT process induced by TNF-α or TGF-β1, the cleaved caspase-1 and ASC speck were not detected, indicating that NLRP3 functions in an inflammasome-independent way. Further studies demonstrated that NLRP3 protein expression was regulated by NF-κB signaling in TNF-α or TGF-β1-induced EMT, as verified by the NF-κB inhibitor Bay 11-7082. Moreover, NLRP3 knockdown reduced the expression of Snail1, indicating that NLRP3 may promote EMT through regulating Snail1. In summary, our results showed that the NLRP3 expression, not the inflammasome activation, was required for EMT in colorectal cancer cells. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. Dielectric spectroscopy platform to measure MCF10A epithelial cell aggregation as a model for spheroidal cell cluster analysis.

    Science.gov (United States)

    Heileman, K L; Tabrizian, M

    2017-05-02

    3-Dimensional cell cultures are more representative of the native environment than traditional cell cultures on flat substrates. As a result, 3-dimensional cell cultures have emerged as a very valuable model environment to study tumorigenesis, organogenesis and tissue regeneration. Many of these models encompass the formation of cell aggregates, which mimic the architecture of tumor and organ tissue. Dielectric impedance spectroscopy is a non-invasive, label free and real time technique, overcoming the drawbacks of established techniques to monitor cell aggregates. Here we introduce a platform to monitor cell aggregation in a 3-dimensional extracellular matrix using dielectric spectroscopy. The MCF10A breast epithelial cell line serves as a model for cell aggregation. The platform maintains sterile conditions during the multi-day assay while allowing continuous dielectric spectroscopy measurements. The platform geometry optimizes dielectric measurements by concentrating cells within the electrode sensing region. The cells show a characteristic dielectric response to aggregation which corroborates with finite element analysis computer simulations. By fitting the experimental dielectric spectra to the Cole-Cole equation, we demonstrated that the dispersion intensity Δε and the characteristic frequency f c are related to cell aggregate growth. In addition, microscopy can be performed directly on the platform providing information about cell position, density and morphology. This platform could yield many applications for studying the electrophysiological activity of cell aggregates.

  18. Osteopontin mediates Citrobacter rodentium-induced colonic epithelial cell hyperplasia and attaching-effacing lesions.

    Science.gov (United States)

    Wine, Eytan; Shen-Tu, Grace; Gareau, Mélanie G; Goldberg, Harvey A; Licht, Christoph; Ngan, Bo-Yee; Sorensen, Esben S; Greenaway, James; Sodek, Jaro; Zohar, Ron; Sherman, Philip M

    2010-09-01

    Although osteopontin (OPN) is up-regulated in inflammatory bowel diseases, its role in disease pathogenesis remains controversial. The objective of this study was to determine the role of OPN in host responses to a non-invasive bacterial pathogen, Citrobacter rodentium, which serves as a murine infectious model of colitis. OPN gene knockout and wild-type mice were infected orogastrically with either C. rodentium or Luria-Bertani (LB) broth. Mouse-derived OPN(+/+) and OPN(-/-) fibroblasts were incubated with C. rodentium and attaching-effacing lesions were demonstrated using transmission electron microscopy and immunofluorescence. Colonic expression of OPN was increased by C. rodentium infection of wild-type mice. Furthermore, colonic epithelial cell hyperplasia, the hallmark of C. rodentium infection, was reduced in OPN(-/-) mice, and spleen enlargement by infection was absent in OPN(-/-) mice. Rectal administration of OPN to OPN(-/-) mice restored these effects. There was an 8- to 17-fold reduction in bacterial colonization in OPN(-/-) mice, compared with wild-type mice, which was accompanied by reduced attaching-effacing lesions, both in infected OPN(-/-) mice and OPN(-/-) mouse fibroblasts. Moreover, adhesion pedestals were restored in OPN(-/-) cells complemented with human OPN. Therefore, lack of OPN results in decreased pedestal formation, colonization, and colonic epithelial cell hyperplasia responses to C. rodentium infection, indicating that OPN impacts disease pathogenesis through bacterial attachment and altered host immune responses.

  19. Cytotoxicity of Different Excipients on RPMI 2650 Human Nasal Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Tamás Horváth

    2016-05-01

    Full Text Available The nasal route receives a great deal of attention as a non-invasive method for the systemic administration of drugs. For nasal delivery, specific formulations containing excipients are used. Because of the sensitive respiratory mucosa, not only the active ingredients, but also additives need to be tested in appropriate models for toxicity. The aim of the study was to measure the cytotoxicity of six pharmaceutical excipients, which could help to reach larger residence time, better permeability, and increased solubility dissolution rate. The following excipients were investigated on RPMI 2650 human nasal septum tumor epithelial cells: β-d-mannitol, sodium hyaluronate, α and β-cyclodextrin, polyvinyl alcohol and methylcellulose. 3-(4,5-dimethyltiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT dye conversion assay and real-time impedance analysis were used to investigate cytotoxicity. No excipient showed toxicity at 0.3% (w/v concentration or below while 1% concentration a significantly reduced metabolic activity was measured by MTT assay for methylcellulose and cyclodextrins. Using impedance measurements, only β-cyclodextrin (1% was toxic to cells. Mannitol at 1% concentration had a barrier opening effect on epithelial cells, but caused no cellular damage. Based on the results, all additives at 0.3%, sodium hyaluronate and polyvinyl alcohol at 1% concentrations can be safely used for nasal formulations.

  20. Aquaporin 2 promotes cell migration and epithelial morphogenesis.

    Science.gov (United States)

    Chen, Ying; Rice, William; Gu, Zhizhan; Li, Jian; Huang, Jianmin; Brenner, Michael B; Van Hoek, Alfred; Xiong, Jianping; Gundersen, Gregg G; Norman, Jim C; Hsu, Victor W; Fenton, Robert A; Brown, Dennis; Lu, Hua A Jenny

    2012-09-01

    The aquaporin 2 (AQP2) water channel, expressed in kidney collecting ducts, contributes critically to water homeostasis in mammals. Animals lacking or having significantly reduced levels of AQP2, however, have not only urinary concentrating abnormalities but also renal tubular defects that lead to neonatal mortality from renal failure. Here, we show that AQP2 is not only a water channel but also an integrin-binding membrane protein that promotes cell migration and epithelial morphogenesis. AQP2 expression modulates the trafficking and internalization of integrin β1, facilitating its turnover at focal adhesions. In vitro, disturbing the interaction between AQP2 and integrin β1 by mutating the RGD motif led to reduced endocytosis, retention of integrin β1 at the cell surface, and defective cell migration and tubulogenesis. Similarly, in vivo, AQP2-null mice exhibited significant retention of integrin β1 at the basolateral membrane and had tubular abnormalities. In summary, these data suggest that the water channel AQP2 interacts with integrins to promote renal epithelial cell migration, contributing to the structural and functional integrity of the mammalian kidney.

  1. Optimization of Invasion-Specific Effects of Betulin Derivatives on Prostate Cancer Cells through Lead Development.

    Directory of Open Access Journals (Sweden)

    Ville Härmä

    Full Text Available The anti-invasive and anti-proliferative effects of betulins and abietane derivatives was systematically tested using an organotypic model system of advanced, castration-resistant prostate cancers. A preliminary screen of the initial set of 93 compounds was performed in two-dimensional (2D growth conditions using non-transformed prostate epithelial cells (EP156T, an androgen-sensitive prostate cancer cell line (LNCaP, and the castration-resistant, highly invasive cell line PC-3. The 25 most promising compounds were all betulin derivatives. These were selected for a focused secondary screen in three-dimensional (3D growth conditions, with the goal to identify the most effective and specific anti-invasive compounds. Additional sensitivity and cytotoxicity tests were then performed using an extended cell line panel. The effects of these compounds on cell cycle progression, mitosis, proliferation and unspecific cytotoxicity, versus their ability to specifically interfere with cell motility and tumor cell invasion was addressed. To identify potential mechanisms of action and likely compound targets, multiplex profiling of compound effects on a panel of 43 human protein kinases was performed. These target de-convolution studies, combined with the phenotypic analyses of multicellular organoids in 3D models, revealed specific inhibition of AKT signaling linked to effects on the organization of the actin cytoskeleton as the most likely driver of altered cell morphology and motility.

  2. Fabrication of corneal epithelial cell sheets maintaining colony-forming cells without feeder cells by oxygen-controlled method.

    Science.gov (United States)

    Nakajima, Ryota; Takeda, Shizu

    2014-01-01

    The use of murine 3T3 feeder cells needs to be avoided when fabricating corneal epithelial cell sheets for use in treating ocular surface diseases. However, the expression level of the epithelial stem/progenitor cell marker, p63, is down-regulated in feeder-free culture systems. In this study, in order to fabricate corneal epithelial cell sheets that maintain colony-forming cells without using any feeder cells, we investigated the use of an oxygen-controlled method that was developed previously to fabricate cell sheets efficiently. Rabbit limbal epithelial cells were cultured under hypoxia (1-10% O2) and under normoxia during stratification after reaching confluence. Multilayered corneal epithelial cell sheets were fabricated using an oxygen-controlled method, and immunofluorescence analysis showed that cytokeratin 3 and p63 was expressed in appropriate localization in the cell sheets. The colony-forming efficiency of the cell sheets fabricated by the oxygen-controlled method without feeder cells was significantly higher than that of cell sheets fabricated under 20% O2 without feeder cells. These results indicate that the oxygen-controlled method has the potential to achieve a feeder-free culture system for fabricating corneal epithelial cell sheets for corneal regeneration. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Effects of mechanical stress and vitreous samples in retinal pigment epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, Eri, E-mail: eritakahashi@fc.kuh.kumamoto-u.ac.jp; Fukushima, Ayako; Haga, Akira; Inomata, Yasuya; Ito, Yasuhiro; Fukushima, Mikiko; Tanihara, Hidenobu

    2016-02-12

    In rhegmatogenous retinal detachment (RRD), scattered RPE cells from the basement membrane into the vitreous cavity undergo an epithelial mesenchymal transition (EMT) and form the intraocular fibrous membrane in response to vitreous fluid. We investigated whether exposure to vitreous samples was associated with EMT-associated signals and mesenchymal characters. Human vitreous samples were collected from patients with RRD, epiretinal membrane (ERM), or macular hole (MH). We evaluated the effects of vitreous on ARPE-19 cells in suspension cultures using poly 2-hydroxyethyl methacrylate-coated dishes and three-dimensional (3D) Matrigel cultures. We found that exposure to vitreous samples did not induce morphological changes or accelerate wound closure in monolayers. Several samples showed increased phosphorylation of Smad2 and nuclear translocation of nuclear factor-κB. Mechanical stress triggered an elevation of phosphorylation levels in Smad2. In addition, exposure to vitreous fluid increased the phosphorylation of p38 mitogen-activated protein kinase in cell suspension cultures after mechanical stress. Moreover, ARPE-19 cells showed a stellate invasive phenotype in 3D Matrigel cultures with vitreous samples. In this study, we demonstrated that mechanical stress and vitreous were associated with EMT-associated signals and invasive phenotypes in 3D cultures but not in monolayers. These results have important implications for the role of vitreous humor in the induction of EMT and intraocular fibrosis.

  4. Hypoxia inducible factor-1α-dependent epithelial to mesenchymal transition under hypoxic conditions in prostate cancer cells.

    Science.gov (United States)

    Li, Mingchuan; Wang, Yong Xing; Luo, Yong; Zhao, Jiahui; Li, Qing; Zhang, Jiao; Jiang, Yongguang

    2016-07-01

    Prostate cancer is the most commonly diagnosed cancer in men and the second leading cause of cancer death. Hypoxia is an environmental stimulus that plays an important role in the development and cancer progression especially for solid tumors. The key regulator under hypoxic conditions is stabilized hypoxia-inducible factor (HIF)-1α. In the present study, immune-fluorescent staining, siRNAs, qRT-PC, immunoblotting, cell migration and invasion assays were carried out to test typical epithelial to mesenchymal transition under hypoxia and the key regulators of this process in PC3, a human prostate cancer cell line. Our data demonstrated that hypoxia induces diverse molecular, phenotypic and functional changes in prostate cancer cells that are consistent with EMT. We also showed that a cell signal factor such as HIF-1α, which might be stabilized under hypoxic environment, is involved in EMT and cancer cell invasive potency. The induced hypoxia could be blocked by HIF-1α gene silencing and reoxygenation of EMT in prostate cancer cells, hypoxia partially reversed accompanied by a process of mesenchymal-epithelial reverting transition (MErT). EMT might be induced by activation of HIF-1α-dependent cell signaling in hypoxic prostate cancer cells.

  5. Phototoxicity and cytotoxicity of fullerol in human lens epithelial cells

    International Nuclear Information System (INIS)

    Roberts, Joan E.; Wielgus, Albert R.; Boyes, William K.; Andley, Usha; Chignell, Colin F.

    2008-01-01

    The water-soluble, hydroxylated fullerene [fullerol, nano-C 60 (OH) 22-26 ] has several clinical applications including use as a drug carrier to bypass the blood ocular barriers. We have assessed fullerol's potential ocular toxicity by measuring its cytotoxicity and phototoxicity induced by UVA and visible light in vitro with human lens epithelial cells (HLE B-3). Accumulation of nano-C 60 (OH) 22-26 in the cells was confirmed spectrophotometrically at 405 nm and cell viability estimated using MTS and LDH assays. Fullerol was cytotoxic to HLE B-3 cells maintained in the dark at concentrations higher than 20 μM. Exposure to either UVA or visible light in the presence of > 5 μM fullerol-induced phototoxic damage. When cells were pretreated with non-toxic antioxidants: 20 μM lutein, 1 mM N-acetyl cysteine, or 1 mM L-ascorbic acid prior to irradiation, only the singlet oxygen quencher-lutein significantly protected against fullerol photodamage. Apoptosis was observed in lens cells treated with fullerol whether or not the cells were irradiated, in the order UVA > visible light > dark. Dynamic light scattering (DLS) showed that in the presence of the endogenous lens protein α-crystallin, large aggregates of fullerol were reduced. In conclusion, fullerol is both cytotoxic and phototoxic to human lens epithelial cells. Although the acute toxicity of water-soluble nano-C 60 (OH) 22-26 is low, these compounds are retained in the body for long periods, raising concern for their chronic toxic effect. Before fullerols are used to deliver drugs to the eye, they should be tested for photo- and cytotoxicity in vivo

  6. Rap1 integrates tissue polarity, lumen formation, and tumorigenicpotential in human breast epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Itoh, Masahiko; Nelson, Celeste M.; Myers, Connie A.; Bissell,Mina J.

    2006-09-29

    Maintenance of apico-basal polarity in normal breast epithelial acini requires a balance between cell proliferation, cell death, and proper cell-cell and cell-extracellular matrix signaling. Aberrations in any of these processes can disrupt tissue architecture and initiate tumor formation. Here we show that the small GTPase Rap1 is a crucial element in organizing acinar structure and inducing lumen formation. Rap1 activity in malignant HMT-3522 T4-2 cells is appreciably higher than in S1 cells, their non-malignant counterparts. Expression of dominant-negative Rap1 resulted in phenotypic reversion of T4-2 cells, led to formation of acinar structures with correct apico-basal polarity, and dramatically reduced tumor incidence despite the persistence of genomic abnormalities. The resulting acini contained prominent central lumina not observed when other reverting agents were used. Conversely, expression of dominant-active Rap1 in T4-2 cells inhibited phenotypic reversion and led to increased invasiveness and tumorigenicity. Thus, Rap1 acts as a central regulator of breast architecture, with normal levels of activation instructing apical polarity during acinar morphogenesis, and increased activation inducing tumor formation and progression to malignancy.

  7. In vitro culture and characterization of a mammary epithelial cell line from Chinese Holstein dairy cow.

    Directory of Open Access Journals (Sweden)

    Han Hu

    Full Text Available BACKGROUND: The objective of this study was to establish a culture system and elucidate the unique characteristics of a bovine mammary epithelial cell line in vitro. METHODOLOGY: Mammary tissue from a three year old lactating dairy cow (ca. 100 d relative to parturition was used as a source of the epithelial cell line, which was cultured in collagen-coated tissue culture dishes. Fibroblasts and epithelial cells successively grew and extended from the culturing mammary tissue at the third day. Pure epithelial cells were obtained by passages culture. PRINCIPAL FINDINGS: The strong positive immunostaining to cytokeratin 18 suggested that the resulting cell line exhibited the specific character of epithelial cells. Epithelial cells cultured in the presence of 10% FBS, supraphysiologic concentrations of insulin, and hydrocortisone maintained a normal diploid chromosome modal number of 2n=60. Furthermore, they were capable of synthesizing beta-casein (CSN2, acetyl-CoA carboxylase-alpha (ACACA and butyrophilin (BTN1A1. An important finding was that frozen preservation in a mixture of 90% FBS and 10% DMSO did not influence the growth characteristics, chromosome number, or protein secretion of the isolated epithelial cell line. CONCLUSIONS: The obtained mammary epithelial cell line had normal morphology, growth characteristics, cytogenetic and secretory characteristics, thus, it might represent an useful tool for studying the function of Chinese Holstein dairy cows mammary epithelial cell (CMECs.

  8. Cellular and Nuclear Alignment Analysis for Determining Epithelial Cell Chirality

    Science.gov (United States)

    Raymond, Michael J.; Ray, Poulomi; Kaur, Gurleen; Singh, Ajay V.; Wan, Leo Q.

    2015-01-01

    Left-right (LR) asymmetry is a biologically conserved property in living organisms that can be observed in the asymmetrical arrangement of organs and tissues and in tissue morphogenesis, such as the directional looping of the gastrointestinal tract and heart. The expression of LR asymmetry in embryonic tissues can be appreciated in biased cell alignment. Previously an in vitro chirality assay was reported by patterning multiple cells on microscale defined geometries and quantified the cell phenotype–dependent LR asymmetry, or cell chirality. However, morphology and chirality of individual cells on micropatterned surfaces has not been well characterized. Here, a Python-based algorithm was developed to identify and quantify immunofluorescence stained individual epithelial cells on multicellular patterns. This approach not only produces results similar to the image intensity gradient-based method reported previously, but also can capture properties of single cells such as area and aspect ratio. We also found that cell nuclei exhibited biased alignment. Around 35% cells were misaligned and were typically smaller and less elongated. This new imaging analysis approach is an effective tool for measuring single cell chirality inside multicellular structures and can potentially help unveil biophysical mechanisms underlying cellular chiral bias both in vitro and in vivo. PMID:26294010

  9. Ouabain Increases Gap Junctional Communication in Epithelial Cells

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    Arturo Ponce

    2014-11-01

    Full Text Available Background/Aims: The finding that endogenous ouabain acts as a hormone prompted efforts to elucidate its physiological function. In previous studies, we have shown that 10 nM ouabain (i.e., a concentration within the physiological range modulates cell-cell contacts such as tight junctions and apical/basolateral polarity. In this study, we examined whether 10 nM ouabain affects another important cell-cell feature: gap junction communication (GJC. Methods: We employed two different approaches: 1 analysis of the cell-to-cell diffusion of neurobiotin injected into a particular MDCK cell (epithelial cells from dog kidneys in a confluent monolayer by counting the number of neighboring cells reached by the probe and 2 measurement of the electrical capacitance. Results: We found that 10 nM ouabain increase GJC by 475% within 1 hour. The Na+-K+-ATPase acts as a receptor of ouabain. In previous works we have shown that ouabain activates c-Src and ERK1/2 in 1 hour; in the present study we show that the inhibition of these proteins block the effect of ouabain on GJC. This increase in GJC does not require synthesis of new protein components, because the inhibitors cycloheximide and actinomycin D did not affect this phenomenon. Using silencing assays we also demonstrate that this ouabain-induced enhancement of GJC involves connexins 32 and 43. Conclusion: Ouabain 10 nM increases GJC in MDCK cells.

  10. The role of epithelial-mesenchymal transition in squamous cell carcinoma of the oral cavity.

    Science.gov (United States)

    Zidar, Nina; Boštjančič, Emanuela; Malgaj, Marija; Gale, Nina; Dovšak, Tadej; Didanovič, Vojko

    2018-02-01

    Epithelial-mesenchymal transition (EMT) has emerged as a possible mechanism of cancer metastasizing, but strong evidence for EMT involvement in human cancer is lacking. Our aim was to compare oral spindle cell carcinoma (SpCC) as an example of EMT with oral conventional squamous cell carcinoma (SCC) with and without nodal metastases to test the hypothesis that EMT contributes to metastasizing in oral SCC. Thirty cases of oral SCC with and without nodal metastasis and 15 cases of SpCC were included. Epithelial (cytokeratin, E-cadherin), mesenchymal (vimentin, N-cadherin), and stem cell markers (ALDH-1, CD44, Nanog, Sox-2) and transcription repressors (Snail, Slug, Twist) were analyzed immunohistochemically. We also analyzed the expression of microRNAs miR-141, miR-200 family, miR-205, and miR-429. SpCC exhibited loss of epithelial markers and expression of mesenchymal markers or coexpression of both up-regulation of transcription repressors and down-regulation of the investigated microRNAs. SCC showed only occasional focal expression of mesenchymal markers at the invasive front. No other differences were observed between SCC with and without nodal metastases except for a higher expression of ALDH-1 in SCC with metastases. Our results suggest that SpCC is an example of true EMT but do not support the hypothesis that EMT is involved in metastasizing of conventional SCC. Regarding oral SCC progression and metastasizing, we have been facing a shift from the initial enthusiasm for the EMT concept towards a more critical approach with "EMT-like" and "partial EMT" concepts. The real question, though, is, is there no EMT at all?

  11. Klebsiella pneumoniae triggers a cytotoxic effect on airway epithelial cells

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    Llobet-Brossa Enrique

    2009-08-01

    Full Text Available Abstract Background Klebsiella pneumoniae is a capsulated Gram negative bacterial pathogen and a frequent cause of nosocomial infections. Despite its clinical relevance, little is known about the features of the interaction between K. pneumoniae and lung epithelial cells on a cellular level, neither about the role of capsule polysaccharide, one of its best characterised virulence factors, in this interaction. Results The interaction between Klebsiella pneumoniae and cultured airway epithelial cells was analysed. K. pneumoniae infection triggered cytotoxicity, evident by cell rounding and detachment from the substrate. This effect required the presence of live bacteria and of capsule polysaccharide, since it was observed with isolates expressing different amounts of capsule and/or different serotypes but not with non-capsulated bacteria. Cytotoxicity was analysed by lactate dehydrogenase and formazan measurements, ethidium bromide uptake and analysis of DNA integrity, obtaining consistent and complementary results. Moreover, cytotoxicity of non-capsulated strains was restored by addition of purified capsule during infection. While a non-capsulated strain was avirulent in a mouse infection model, capsulated K. pneumoniae isolates displayed different degrees of virulence. Conclusion Our observations allocate a novel role to K. pneumoniae capsule in promotion of cytotoxicity. Although this effect is likely to be associated with virulence, strains expressing different capsule levels were not equally virulent. This fact suggests the existence of other bacterial requirements for virulence, together with capsule polysaccharide.

  12. Hyaluronan in aged collagen matrix increases prostate epithelial cell proliferation

    Science.gov (United States)

    Damodarasamy, Mamatha; Vernon, Robert B.; Chan, Christina K.; Plymate, Stephen R.; Wight, Thomas N.

    2015-01-01

    The extracellular matrix (ECM) of the prostate, which is comprised primarily of collagen, becomes increasingly disorganized with age, a property that may influence the development of hyperplasia and cancer. Collageous ECM extracted from the tails of aged mice exhibits many characteristics of collagen in aged tissues, including the prostate. When polymerized into a 3-dimensional (3D) gel, these collagen extracts can serve as models for the study of specific cell-ECM interactions. In the present study, we examined the behaviors of human prostatic epithelial cell lines representing normal prostate epithelial cells (PEC), benign prostatic hyperplasia (BPH-1), and adenocarcinoma (LNCaP) cultured in contact with 3D gels made from collagen extracts of young and aged mice. We found that proliferation of PEC, BPH-1, and LNCaP cells were all increased by culture on aged collagen gels relative to young collagen gels. In examining age-associated differences in the composition of the collagen extracts, we found that aged and young collagen had a similar amount of several collagen-associated ECM components, but aged collagen had a much greater content of the glycosaminoglycan hyaluronan (HA) than young collagen. The addition of HA (of similar size and concentration to that found in aged collagen extracts) to cells placed in young collagen elicited significantly increased proliferation in BPH-1 cells, but not in PEC or LNCaP cells, relative to controls not exposed to HA. Of note, histochemical analyses of human prostatic tissues showed significantly higher expression of HA in BPH and prostate cancer stroma relative to stroma of normal prostate. Collectively, these results suggest that changes in ECM involving increased levels of HA contribute to the growth of prostatic epithelium with aging. PMID:25124870

  13. Human cancer cells express Slug-based epithelial-mesenchymal transition gene expression signature obtained in vivo

    International Nuclear Information System (INIS)

    Anastassiou, Dimitris; Rumjantseva, Viktoria; Cheng, Weiyi; Huang, Jianzhong; Canoll, Peter D; Yamashiro, Darrell J; Kandel, Jessica J

    2011-01-01

    The biological mechanisms underlying cancer cell motility and invasiveness remain unclear, although it has been hypothesized that they involve some type of epithelial-mesenchymal transition (EMT). We used xenograft models of human cancer cells in immunocompromised mice, profiling the harvested tumors separately with species-specific probes and computationally analyzing the results. Here we show that human cancer cells express in vivo a precise multi-cancer invasion-associated gene expression signature that prominently includes many EMT markers, among them the transcription factor Slug, fibronectin, and α-SMA. We found that human, but not mouse, cells express the signature and Slug is the only upregulated EMT-inducing transcription factor. The signature is also present in samples from many publicly available cancer gene expression datasets, suggesting that it is produced by the cancer cells themselves in multiple cancer types, including nonepithelial cancers such as neuroblastoma. Furthermore, we found that the presence of the signature in human xenografted cells was associated with a downregulation of adipocyte markers in the mouse tissue adjacent to the invasive tumor, suggesting that the signature is triggered by contextual microenvironmental interactions when the cancer cells encounter adipocytes, as previously reported. The known, precise and consistent gene composition of this cancer mesenchymal transition signature, particularly when combined with simultaneous analysis of the adjacent microenvironment, provides unique opportunities for shedding light on the underlying mechanisms of cancer invasiveness as well as identifying potential diagnostic markers and targets for metastasis-inhibiting therapeutics

  14. Epithelial Cell-Derived Secreted and Transmembrane 1a Signals to Activated Neutrophils during Pneumococcal Pneumonia.

    Science.gov (United States)

    Kamata, Hirofumi; Yamamoto, Kazuko; Wasserman, Gregory A; Zabinski, Mary C; Yuen, Constance K; Lung, Wing Yi; Gower, Adam C; Belkina, Anna C; Ramirez, Maria I; Deng, Jane C; Quinton, Lee J; Jones, Matthew R; Mizgerd, Joseph P

    2016-09-01

    Airway epithelial cell responses are critical to the outcome of lung infection. In this study, we aimed to identify unique contributions of epithelial cells during lung infection. To differentiate genes induced selectively in epithelial cells during pneumonia, we compared genome-wide expression profiles from three sorted cell populations: epithelial cells from uninfected mouse lungs, epithelial cells from mouse lungs with pneumococcal pneumonia, and nonepithelial cells from those same infected lungs. Of 1,166 transcripts that were more abundant in epithelial cells from infected lungs compared with nonepithelial cells from the same lungs or from epithelial cells of uninfected lungs, 32 genes were identified as highly expressed secreted products. Especially strong signals included two related secreted and transmembrane (Sectm) 1 genes, Sectm1a and Sectm1b. Refinement of sorting strategies suggested that both Sectm1 products were induced predominantly in conducting airway epithelial cells. Sectm1 was induced during the early stages of pneumococcal pneumonia, and mutation of NF-κB RelA in epithelial cells did not diminish its expression. Instead, type I IFN signaling was necessary and sufficient for Sectm1 induction in lung epithelial cells, mediated by signal transducer and activator of transcription 1. For target cells, Sectm1a bound to myeloid cells preferentially, in particular Ly6G(bright)CD11b(bright) neutrophils in the infected lung. In contrast, Sectm1a did not bind to neutrophils from uninfected lungs. Sectm1a increased expression of the neutrophil-attracting chemokine CXCL2 by neutrophils from the infected lung. We propose that Sectm1a is an epithelial product that sustains a positive feedback loop amplifying neutrophilic inflammation during pneumococcal pneumonia.

  15. Y-27632, a ROCK Inhibitor, Promoted Limbal Epithelial Cell Proliferation and Corneal Wound Healing.

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    Chi-Chin Sun

    Full Text Available Transplantation of ex vivo cultured limbal epithelial cells is proven effective in restoring limbal stem cell deficiency. The present study aimed to investigate the promoting effect of Y-27632 on limbal epithelial cell proliferation. Limbal explants isolated from human donor eyes were expanded three weeks on culture dishes and outgrowth of epithelial cells was subsequently subcultured for in vitro experiments. In the presence of Y-27632, the ex vivo limbal outgrowth was accelerated, particularly the cells with epithelial cell-like morphology. Y-27632 dose-dependently promoted the proliferation of in vitro cultured human limbal epithelial cells as examined by phase contrast microscopy and luminescent cell-viability assay 30 hours after the treatment. The colony forming efficacy determined 7 days after the treatment was enhanced by Y-27632 also in a dose-dependent manner. The number of p63- or Ki67-positive cells was dose-dependently increased in Y-27632-treated cultures as detected by immunofluorescent staining and western blotanalysis. Cell cycle analysis by flow cytometric method revealed an increase in S-phase proliferating cells. The epithelial woundclosure rate was shown to be faster in experimental group received topical treatment withY-27632 than the sham control using a rat corneal wounding model. These resultsdemonstrate that Y-27632 can promote both the ex vivo and in vitro proliferation oflimbal epithelial cell proliferation. The in vivo enhanced epithelial wound healingfurther implies that the Y-27632 may act as a new strategy for treating limbal stem cell deficiency.

  16. CXCL9 Regulates TGF-β1-Induced Epithelial to Mesenchymal Transition in Human Alveolar Epithelial Cells.

    Science.gov (United States)

    O'Beirne, Sarah L; Walsh, Sinead M; Fabre, Aurélie; Reviriego, Carlota; Worrell, Julie C; Counihan, Ian P; Lumsden, Robert V; Cramton-Barnes, Jennifer; Belperio, John A; Donnelly, Seamas C; Boylan, Denise; Marchal-Sommé, Joëlle; Kane, Rosemary; Keane, Michael P

    2015-09-15

    Epithelial to mesenchymal cell transition (EMT), whereby fully differentiated epithelial cells transition to a mesenchymal phenotype, has been implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). CXCR3 and its ligands are recognized to play a protective role in pulmonary fibrosis. In this study, we investigated the presence and extent of EMT and CXCR3 expression in human IPF surgical lung biopsies and assessed whether CXCR3 and its ligand CXCL9 modulate EMT in alveolar epithelial cells. Coexpression of the epithelial marker thyroid transcription factor-1 and the mesenchymal marker α-smooth muscle actin and CXCR3 expression was examined by immunohistochemical staining of IPF surgical lung biopsies. Epithelial and mesenchymal marker expression was examined by quantitative real-time PCR, Western blotting, and immunofluorescence in human alveolar epithelial (A549) cells treated with TGF-β1 and CXCL9, with Smad2, Smad3, and Smad7 expression and cellular localization examined by Western blotting. We found that significantly more cells were undergoing EMT in fibrotic versus normal areas of lung in IPF surgical lung biopsy samples. CXCR3 was expressed by type II pneumocytes and fibroblasts in fibrotic areas in close proximity to cells undergoing EMT. In vitro, CXCL9 abrogated TGF-β1-induced EMT. A decrease in TGF-β1-induced phosphorylation of Smad2 and Smad3 occurred with CXCL9 treatment. This was associated with increased shuttling of Smad7 from the nucleus to the cytoplasm where it inhibits Smad phosphorylation. This suggests a role for EMT in the pathogenesis of IPF and provides a novel mechanism for the inhibitory effects of CXCL9 on TGF-β1-induced EMT. Copyright © 2015 by The American Association of Immunologists, Inc.

  17. γδ T cells in homeostasis and host defence of epithelial barrier tissues

    DEFF Research Database (Denmark)

    Nielsen, Morten M.; Witherden, Deborah A.; Havran, Wendy L.

    2017-01-01

    Epithelial surfaces line the body and provide a crucial interface between the body and the external environment. Tissue-resident epithelial γδ T cells represent a major T cell population in the epithelial tissues and are ideally positioned to carry out barrier surveillance and aid in tissue...... homeostasis and repair. In this Review, we focus on the intraepithelial γδ T cell compartment of the two largest epithelial tissues in the body — namely, the epidermis and the intestine — and provide a comprehensive overview of the crucial contributions of intraepithelial γδ T cells to tissue integrity...

  18. Production of arachidonic and linoleic acid metabolites by guinea pig tracheal epithelial cells

    International Nuclear Information System (INIS)

    Oosthuizen, M.J.; Engels, F.; Van Esch, B.; Henricks, P.A.; Nijkamp, F.P.

    1990-01-01

    Pulmonary epithelial cells may be responsible for regulating airway smooth muscle function, in part by release of fatty acid-derived mediators. Incubation of isolated guinea pig tracheal epithelial cells with radiolabeled arachidonic acid (AA) leads to the production of 5- and 15-hydroxyeicosatetraenoic acid (5- and 15-HETE) and smaller amounts of leukotriene (LT) B4 and C4 and 12-hydroxyheptadecatrienoic acid (HHT). Epithelial cells also are able to release linoleic acid (LA) metabolites. Incubation with radiolabeled linoleic acid leads to the formation of 9- and 13-hydroxyoctadecadienoic acid (9- and 13-HODE). The biological significance of these mediators produced by epithelial cells is discussed

  19. Cell-ECM Interactions During Cancer Invasion

    Science.gov (United States)

    Jiang, Yi

    The extracellular matrix (ECM), a fibrous material that forms a network in a tissue, significantly affects many aspects of cellular behavior, including cell movement and proliferation. Transgenic mouse tumor studies indicate that excess collagen, a major component of ECM, enhances tumor formation and invasiveness. Clinically, tumor associated collagen signatures are strong markers for breast cancer survival. However, the underlying mechanisms are unclear since the properties of ECM are complex, with diverse structural and mechanical properties depending on various biophysical parameters. We have developed a three-dimensional elastic fiber network model, and parameterized it with in vitro collagen mechanics. Using this model, we study ECM remodeling as a result of local deformation and cell migration through the ECM as a network percolation problem. We have also developed a three-dimensional, multiscale model of cell migration and interaction with ECM. Our model reproduces quantitative single cell migration experiments. This model is a first step toward a fully biomechanical cell-matrix interaction model and may shed light on tumor associated collagen signatures in breast cancer. This work was partially supported by NIH-U01CA143069.

  20. Cigarette smoke suppresses Bik to cause epithelial cell hyperplasia and mucous cell metaplasia.

    Science.gov (United States)

    Mebratu, Yohannes A; Schwalm, Kurt; Smith, Kevin R; Schuyler, Mark; Tesfaigzi, Yohannes

    2011-06-01

    Aberrant regulation of airway epithelial cell numbers in airways leads to increased mucous secretions in chronic lung diseases such as chronic bronchitis. Because the Bcl-2 family of proteins is crucial for airway epithelial homeostasis, identifying the players that reduce cigarette smoke (CS)-induced mucous cell metaplasia can help to develop effective therapies. To identify the Bcl-2 family of proteins that play a role in reducing CS-induced mucous cell metaplasia. We screened for dysregulated expression of the Bcl-2 family members. We identified Bik to be significantly reduced in bronchial brushings of patients with chronic epithelial cell hyperplasia compared with nondiseased control subjects. Reduced Bik but increased MUC5AC mRNA levels were also detected when normal human airway epithelial cells (HAECs) were exposed to CS or when autopsy tissues from former smokers with and without chronic bronchitis were compared. Similarly, exposure of C57Bl/6 mice to CS resulted in increased numbers of epithelial and mucous cells per millimeter of basal lamina, along with reduced Bik but increased Muc5ac expression, and this change was sustained even when mice were allowed to recover in filtered air for 8 weeks. Restoring Bik expression significantly suppressed CS-induced mucous cell metaplasia in differentiated primary HAEC cultures and in airways of mice in vivo. Bik blocked nuclear translocation of phospho-ERK1/2 to induce apoptosis of HAECs. The conserved Leu61 within Bik and ERK1/2 activation were essential to induce cell death in hyperplastic mucous cells. These studies show that CS suppresses Bik expression to block airway epithelia cell death and thereby increases epithelial cell hyperplasia in chronic bronchitis.

  1. Use of scanning electron microscopy to monitor nanofibre/cell interaction in digestive epithelial cells.

    Science.gov (United States)

    Millaku, Agron; Drobne, Damjana; Torkar, Matjaz; Novak, Sara; Remškar, Maja; Pipan-Tkalec, Živa

    2013-09-15

    We provide data obtained by scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) on the interaction of ingested tungsten nanofibers with epithelial cells of the digestive tubes of a test organism Porcellio scaber. Conventional toxicity endpoints including feeding behaviour, weight loss and mortality were also measured in each investigated animal. No toxicity was detected in any of exposed animals after 14 days of feeding on tungsten nanofiber dosed food, but when nanofibers enter the digestive system they can react with epithelial cells of the digestive tubes, becoming physically inserted into the cells. In this way, nanofibers can injure the epithelial cells of digestive gland tubes when they are ingested with food. Our SEM data suggest that peristaltic forces may have an important role, not predicted by in vitro experiments, in the interactions of nanomaterials with digestive intestinal cells. Copyright © 2013 Elsevier B.V. All rights reserved.

  2. Differentiation and molecular profiling of human embryonic stem cell-derived corneal epithelial cells.

    Science.gov (United States)

    Brzeszczynska, J; Samuel, K; Greenhough, S; Ramaesh, K; Dhillon, B; Hay, D C; Ross, J A

    2014-06-01

    It has been suggested that the isolation of scalable populations of limbal stem cells may lead to radical changes in ocular therapy. In particular, the derivation and transplantation of corneal stem cells from these populations may result in therapies providing clinical normality of the diseased or damaged cornea. Although feasible in theory, the lack of donor material in sufficient quantity and quality currently limits such a strategy. A potential scalable source of corneal cells could be derived from pluripotent stem cells (PSCs). We developed an in vitro and serum-free corneal differentiation model which displays significant promise. Our stepwise differentiation model was designed with reference to development and gave rise to cells which displayed similarities to epithelial progenitor cells which can be specified to cells displaying a corneal epithelial phenotype. We believe our approach is novel, provides a robust model of human development and in the future, may facilitate the generation of corneal epithelial cells that are suitable for clinical use. Additionally, we demonstrate that following continued cell culture, stem cell-derived corneal epithelial cells undergo transdifferentiation and exhibit squamous metaplasia and therefore, also offer an in vitro model of disease.

  3. Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

    Directory of Open Access Journals (Sweden)

    Magnusson Magnus K

    2010-07-01

    Full Text Available Abstract Background Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D co-culture assay. Methods Breast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy. Results In co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors. Conclusion Breast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

  4. Equine tracheal epithelial membrane strips - An alternate method for examining epithelial cell arachidonic acid metabolism

    International Nuclear Information System (INIS)

    Gray, P.R.; Derksen, F.J.; Robinson, N.E.; Peter-Golden, M.L.

    1990-01-01

    Arachidonic acid metabolism by tracheal epithelium can be studied using enzymatically dispersed cell suspensions or cell cultures. Both techniques require considerable tissue disruption and manipulation and may not accurately represent in vivo activity. The authors have developed an alternate method for obtaining strips of equine tracheal epithelium without enzymatic digestion. In the horse, a prominent elastic lamina supports the tracheal epithelium. By physical splitting this lamina, they obtained strips (≤12 x 1.5 cm) of pseudostratified columnar epithelium attached to a layer of elastic tissue 30-100 μm thick. Epithelial strips (1.2 x 0.5 cm) were attached to plexiglass rods and incubated with [ 3 H]arachidonic acid in M199 medium (0.5 μCi/ml) for 24 hours at 37C. The strips incorporated 36±4% (mean ± SEM) of the total radioactivity and released 8.0±1.2% of incorporated radioactivity when stimulated by 5.0 μM calcium ionophore A23187. The extracted supernatant was processed using HPLC, resulting in peaks of radioactivity that co-eluted with authentic PGE 2 , PGF 2 α, and 12-HETE standards. The greatest activity corresponded to the PGE 2 and PGF 2 α standards, which is a similar pattern to that reported for cultured human tracheal epithelium

  5. miR-146a Suppresses Invasion of Pancreatic Cancer Cells

    Science.gov (United States)

    Li, Yiwei; VandenBoom, Timothy G.; Wang, Zhiwei; Kong, Dejuan; Ali, Shadan; Philip, Philip A.; Sarkar, Fazlul H.

    2010-01-01

    The aggressive course of pancreatic cancer is believed to reflect its unusually invasive and metastatic nature, which is associated with epidermal growth factor receptor (EGFR) overexpression and NF-κB activation. MicroRNAs (miRNA) have been implicated in the regulation of various pathobiological processes in cancer, including metastasis in pancreatic cancer and in other human malignancies. In this study, we report lower expression of miR-146a in pancreatic cancer cells compared with normal human pancreatic duct epithelial cells. Reexpression of miR-146a inhibited the invasive capacity of pancreatic cancer cells with concomitant downregulation of EGFR and the NF-κB regulatory kinase interleukin 1 receptor–associated kinase 1 (IRAK-1). Cellular mechanism studies revealed crosstalk between EGFR, IRAK-1, IκBα, NF-κB, and MTA-2, a transcription factor that regulates metastasis. Treatment of pancreatic cancer cells with the natural products 3,3′-diinodolylmethane (DIM) or isoflavone, which increased miR-146a expression, caused a downregulation of EGFR, MTA-2, IRAK-1, and NF-κB, resulting in an inhibition of pancreatic cancer cell invasion. Our findings reveal DIM and isoflavone as nontoxic activators of a miRNA that can block pancreatic cancer cell invasion and metastasis, offering starting points to design novel anticancer agents. PMID:20124483

  6. NoxO1 Controls Proliferation of Colon Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Franziska Moll

    2018-05-01

    Full Text Available AimReactive oxygen species (ROS produced by enzymes of the NADPH oxidase family serve as second messengers for cellular signaling. Processes such as differentiation and proliferation are regulated by NADPH oxidases. In the intestine, due to the exceedingly fast and constant renewal of the epithelium both processes have to be highly controlled and balanced. Nox1 is the major NADPH oxidase expressed in the gut, and its function is regulated by cytosolic subunits such as NoxO1. We hypothesize that the NoxO1-controlled activity of Nox1 contributes to a proper epithelial homeostasis and renewal in the gut.ResultsNoxO1 is highly expressed in the colon. Knockout of NoxO1 reduces the production of superoxide in colon crypts and is not subsidized by an elevated expression of its homolog p47phox. Knockout of NoxO1 increases the proliferative capacity and prevents apoptosis of colon epithelial cells. In mouse models of dextran sulfate sodium (DSS-induced colitis and azoxymethane/DSS induced colon cancer, NoxO1 has a protective role and may influence the population of natural killer cells.ConclusionNoxO1 affects colon epithelium homeostasis and prevents inflammation.

  7. Adrenomedullin stimulates cyclic AMP production in the airway epithelial cells of guinea-pigs and in the human epithelial cell line

    Directory of Open Access Journals (Sweden)

    Takashi Kawaguchi

    1999-01-01

    Full Text Available This study was designed to examine the effects of adrenomedullin (AM on airway epithelial cells. Primary cultures of guinea-pig tracheal epithelial cells and the human bronchiolar epithelial cell line NCI-H441 were used. Intracellular cyclic adenosine monophosphate (cAMP, cyclic guanosine monophosphate (cGMP, prostaglandin E2 (PGE2, and stable end-products of nitric oxide were assayed. Adrenomedullin (10−6 mol/L stimulated cAMP production in guinea-pig epithelial cells. Indomethacin (10−5 mol/L significantly decreased the basal level of intracellular cAMP in guinea-pig epithelial cells, but not in NCI-H441 cells. However, AM did not stimulate production of PGE2, a major product that can increase cAMP formation. In the case of NCI-H441 cells, AM (10−8 – 10−6 mol/L did not significantly affect intracellular cGMP levels or nitrite content in conditioned medium. Adrenomedullin and calcitonin gene-related peptide (CGRP each stimulated cAMP production in NCI-H441 cells, but AM-stimulated cAMP production was antagonized by the CGRP fragment CGRP8–37. These findings suggest that AM stimulates cAMP production and functionally competes with CGRP for binding sites in airway epithelial cells, at least in human epithelial cells, but that it does not stimulate the release of PGE2 and nitric oxide. Though cyclooxygenase products contribute to some extent to cAMP formation in guinea-pigs, AM independently stimulates intracellular cAMP formation in airway epithelial cells.

  8. Monocarboxylate transporters MCT1 and MCT4 regulate migration and invasion of pancreatic ductal adenocarcinoma cells

    DEFF Research Database (Denmark)

    Kong, Su Chii; Nøhr-Nielsen, Asbjørn; Zeeberg, Katrine

    2016-01-01

    , localization, activity, and function were explored in human PDAC cells (MIAPaCa-2, Panc-1, BxPC-3, AsPC-1) and normal human pancreatic ductal epithelial (HPDE) cells, by quantitative polymerase chain reaction, immunoblotting, immunocytochemistry, lactate flux, migration, and invasion assays. RESULTS: MCT1......, or knockdown of MCT1 or MCT4. PDAC cell migration was largely unaffected by MCT1/MCT2 inhibition or MCT1 knockdown but was reduced by 4-CIN and by MCT4 knockdown (BxPC-3). Invasion measured in Boyden chamber (BxPC-3, Panc-1) and spheroid outgrowth (BxPC-3) assays was attenuated by 4-CIN and AR-C155858...

  9. CENPI is overexpressed in colorectal cancer and regulates cell migration and invasion.

    Science.gov (United States)

    Ding, Na; Li, Rongxin; Shi, Wenhao; He, Cui

    2018-06-21

    Centromere protein I (CENPI),an important member of centromere protein family, has been suggest to serve as a oncogene in breast cancer, but the clinical significance and biological function of CENPI in colorectal cancer (CRC) is still unclear. In our results, we found CENPI was overexpressed in CRC tissues and cells, and associated with clinical stage, tumor depth, lymph node metastasis, distant metastasis and differentiation in CRC patients. However, there was no significant association between CENPI protein expression and overall survival time in colon cancer patients and rectal cancer patients through analyzing TCGA survival data. Moreover, CENPI mRNA and protein were increased in metastatic lymph nodes compared with primary CRC tissues. Down-regulation of CENPI expression suppresses CRC cell migration, invasion and epithelial mesenchymal transition process. In conclusion, CENPI is overexpressed in CRC and functions as oncogene in modulating CRC cell migration, invasion and EMT process. Copyright © 2018. Published by Elsevier B.V.

  10. Cyclin-dependent kinases regulate apoptosis of intestinal epithelial cells

    Science.gov (United States)

    Bhattacharya, Sujoy; Ray, Ramesh M.; Johnson, Leonard R.

    2014-01-01

    Homeostasis of the gastrointestinal epithelium is dependent upon a balance between cell proliferation and apoptosis. Cyclin-dependent kinases (Cdks) are well known for their role in cell proliferation. Previous studies from our group have shown that polyamine-depletion of intestinal epithelial cells (IEC-6) decreases cyclin-dependent kinase 2 (Cdk2) activity, increases p53 and p21Cip1 protein levels, induces G1 arrest, and protects cells from camptothecin (CPT)-induced apoptosis. Although emerging evidence suggests that members of the Cdk family are involved in the regulation of apoptosis, their roles directing apoptosis of IEC-6 cells are not known. In this study, we report that inhibition of Cdk1, 2, and 9 (with the broad range Cdk inhibitor, AZD5438) in proliferating IEC-6 cells triggered DNA damage, activated p53 signaling, inhibited proliferation, and induced apoptosis. By contrast, inhibition of Cdk2 (with NU6140) increased p53 protein and activity, inhibited proliferation, but had no effect on apoptosis. Notably, AZD5438 sensitized, whereas, NU6140 rescued proliferating IEC-6 cells from CPT-induced apoptosis. However, in colon carcinoma (Caco2) cells with mutant p53, treatment with either AZD5438 or NU6140 blocked proliferation, albeit more robustly with AZD5438. Both Cdk inhibitors induced apoptosis in Caco2 cells in a p53-independent manner. In serum starved quiescent IEC-6 cells, both AZD5438 and NU6140 decreased TNF- /CPT-induced activation of p53 and, consequently, rescued cells from apoptosis, indicating that sustained Cdk activity is required for apoptosis of quiescent cells. Furthermore, AZD5438 partially reversed the protective effect of polyamine depletion whereas NU6140 had no effect. Together, these results demonstrate that Cdks possess opposing roles in the control of apoptosis in quiescent and proliferating cells. In addition, Cdk inhibitors uncouple proliferation from apoptosis in a p53-dependent manner. PMID:24242917

  11. CD8+ T cells induce thyroid epithelial cell hyperplasia and fibrosis.

    Science.gov (United States)

    Yu, Shiguang; Fang, Yujiang; Sharav, Tumenjargal; Sharp, Gordon C; Braley-Mullen, Helen

    2011-02-15

    CD8(+) T cells can be important effector cells in autoimmune inflammation, generally because they can damage target cells by cytotoxicity. This study shows that activated CD8(+) T cells induce thyroid epithelial cell hyperplasia and proliferation and fibrosis in IFN-γ(-/-) NOD.H-2h4 SCID mice in the absence of CD4(+) T cells. Because CD8(+) T cells induce proliferation rather than cytotoxicity of target cells, these results describe a novel function for CD8(+) T cells in autoimmune disease. In contrast to the ability of purified CD8(+) T cells to induce thyrocyte proliferation, CD4(+) T cells or CD8 T cell-depleted splenocytes induced only mild thyroid lesions in SCID recipients. T cells in both spleens and thyroids highly produce TNF-α. TNF-α promotes proliferation of thyrocytes in vitro, and anti-TNF-α inhibits development of thyroid epithelial cell hyperplasia and proliferation in SCID recipients of IFN-γ(-/-) splenocytes. This suggests that targeting CD8(+) T cells and/or TNF-α may be effective for treating epithelial cell hyperplasia and fibrosis.

  12. Comparison of para-aminophenol cytotoxicity in rat renal epithelial cells and hepatocytes.

    Science.gov (United States)

    Li, Ying; Bentzley, Catherine M; Tarloff, Joan B

    2005-04-01

    Several chemicals, including para-aminophenol (PAP), produce kidney damage in the absence of hepatic damage. Selective nephrotoxicity may be related to the ability of the kidney to reabsorb filtered water, thereby raising the intraluminal concentration of toxicants and exposing tubular epithelial cells to higher concentrations than would be present in other tissues. The present experiments tested the hypothesis that hepatocytes and renal epithelial cells exposed to equivalent concentrations of PAP would be equally susceptible to toxicity. Hepatocytes and renal epithelial cells were prepared by collagenase digestion of tissues obtained from female Sprague-Dawley rats. Toxicity was monitored using trypan blue exclusion, oxygen consumption and ATP content. We measured the rate of PAP clearance and formation of PAP-glutathione conjugate by HPLC. We found that renal epithelial cells accumulated trypan blue and showed declines in oxygen consumption and ATP content at significantly lower concentrations of PAP and at earlier time points than hepatocytes. The half-life of PAP in hepatocyte incubations was significantly shorter (0.71+/-0.07 h) than in renal epithelial cell incubations (1.33+/-0.23 h), suggesting that renal epithelial cells were exposed to PAP for longer time periods than hepatocytes. Renal epithelial cells formed significantly less glutathione conjugates of PAP (PAP-SG) than did hepatocytes, consistent with less efficient detoxification of reactive PAP intermediates by renal epithelial cells. Finally, hepatocytes contained significant more reduced glutathione (NPSH) than did renal epithelial cells, possibly explaining the enhanced formation of PAP-SG by this cell population. In conclusion, our data indicates that renal epithelial cells are intrinsically more susceptible to PAP cytotoxicity than are hepatocytes. This enhanced cytotoxicity may be due to longer exposure to PAP and/or reduced detoxification of reactive intermediates due to lower concentrations

  13. A method for isolating identifying and culturing of rat trachea-bronchia epithelial cells

    International Nuclear Information System (INIS)

    Cui Fengmei; Su Shibiao; Nie Jihua; Li Bingyan; Tong Jian

    2005-01-01

    Objective: To explore a method for isolating identifying and culturing the rat trachea-bronchia epithelial cells. Methods: The rat trachea-bronchia epithelial cells were isolated by digestion with pronase and brushing with cell brush, identified using confocul and cultured in entire F12 media with no serum. Results: With this method, cells in high purity and high viability could be obtained, and about 10 6 cells per rat. The cells grow well in entire F12 media with no serum. Conclusion: The method is useful for isolating rate trachea-bronchia epithelial cells and the entire F12 media with no serum is effective for culturing. (authors)

  14. Altered CXCR3 isoform expression regulates prostate cancer cell migration and invasion

    Directory of Open Access Journals (Sweden)

    Wu Qian

    2012-01-01

    Full Text Available Abstract Background Carcinoma cells must circumvent the normally suppressive signals to disseminate. While often considered 'stop' signals for adherent cells, CXCR3-binding chemokines have recently been correlated positively with cancer progression though the molecular basis remains unclear. Results Here, we examined the expression and function of two CXCR3 variants in human prostate cancer biopsies and cell lines. Globally, both CXCR3 mRNA and protein were elevated in localized and metastatic human cancer biopsies compared to normal. Additionally, CXCR3A mRNA level was upregulated while CXCR3B mRNA was downregulated in these prostate cancer specimens. In contrast to normal prostate epithelial cells (RWPE-1, CXCR3A was up to half the receptor in the invasive and metastatic DU-145 and PC-3 prostate cancer cells, but not in the localized LNCaP cells. Instead of inhibiting cell migration as in RWPE-1 cells, the CXCR3 ligands CXCL4/PF4 and CXCL10/IP10 promoted cell motility and invasiveness in both DU-145 and PC-3 cells via PLCβ3 and μ-calpain activation. CXCR3-mediated diminution of cell motility in RWPE-1 cells is likely a result of cAMP upregulation and m-calpain inhibition via CXCR3B signal transduction. Interestingly, overexpression of CXCR3B in DU-145 cells decreased cell movement and invasion. Conclusion These data suggest that the aberrant expression of CXCR3A and down-regulation of CXCR3B may switch a progression "stop" to a "go" signal to promote prostate tumor metastasis via stimulating cell migration and invasion.

  15. Centriole movements in mammalian epithelial cells during cytokinesis

    Directory of Open Access Journals (Sweden)

    Tanke Hans J

    2010-05-01

    Full Text Available Abstract Background In cytokinesis, when the cleavage furrow has been formed, the two centrioles in each daughter cell separate. It has been suggested that the centrioles facilitate and regulate cytokinesis to some extent. It has been postulated that termination of cytokinesis (abscission depends on the migration of a centriole to the intercellular bridge and then back to the cell center. To investigate the involvement of centrioles in cytokinesis, we monitored the movements of centrioles in three mammalian epithelial cell lines, HeLa, MCF 10A, and the p53-deficient mouse mammary tumor cell line KP-7.7, by time-lapse imaging. Centrin1-EGFP and α-Tubulin-mCherry were co-expressed in the cells to visualize respectively the centrioles and microtubules. Results Here we report that separated centrioles that migrate from the cell pole are very mobile during cytokinesis and their movements can be characterized as 1 along the nuclear envelope, 2 irregular, and 3 along microtubules forming the spindle axis. Centriole movement towards the intercellular bridge was only seen occasionally and was highly cell-line dependent. Conclusions These findings show that centrioles are highly mobile during cytokinesis and suggest that the repositioning of a centriole to the intercellular bridge is not essential for controlling abscission. We suggest that centriole movements are microtubule dependent and that abscission is more dependent on other mechanisms than positioning of centrioles.

  16. Effect of Formaldehyde on Human Middle Ear Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Shin Hye Kim

    2018-01-01

    Full Text Available Formaldehyde (FA is a familiar indoor air pollutant found in everything from cosmetics to clothing, but its impact on the middle ear is unknown. This study investigated whether FA causes cytotoxicity, inflammation, or induction of apoptosis in human middle ear epithelial cells (HMEECs. Cell viability was investigated using the trypan blue assay and a cell counting kit (CCK-8 in HMEECs treated with FA for 4 or 24 h. The expression of genes encoding the inflammatory cytokine tumor necrosis factor alpha (TNF-α and mucin (MUC5AC was analyzed using RT-PCR. Activation of the apoptosis pathway was determined by measuring mitochondrial membrane potential (MMP, cytochrome oxidase, caspase-9/Mch6/Apaf 3, and Caspase-Glo® 3/7 activities. The CCK-8 assay and trypan blue assay results showed a reduction in cell viability in FA-treated HMEECs. FA also increased the cellular expression of TNF-α and MUC5AC and reduced the activities of MMP and cytochrome oxidase. Caspase-9 activity increased in cells stimulated for 4 h, as well as caspase-3/7 activity in cells stimulated for 24 h. The decreased cell viability, the induction of inflammation and mucin gene expression, and the activation of the apoptosis pathway together indicate a link between environmental FA exposure and the development of otitis media.

  17. The regenerative potential of parietal epithelial cells in adult mice.

    Science.gov (United States)

    Berger, Katja; Schulte, Kevin; Boor, Peter; Kuppe, Christoph; van Kuppevelt, Toin H; Floege, Jürgen; Smeets, Bart; Moeller, Marcus J

    2014-04-01

    Previously, we showed that some podocytes in juvenile mice are recruited from cells lining Bowman's capsule, suggesting that parietal epithelial cells (PECs) are a progenitor cell population for podocytes. To investigate whether PECs also replenish podocytes in adult mice, PECs were genetically labeled in an irreversible fashion in 5-week-old mice. No significant increase in labeled podocytes was observed, even after 18 months. To accelerate a potential regenerative mechanism, progressive glomerular hypertrophy was induced by progressive partial nephrectomies. Again, no significant podocyte replenishment was observed. Rather, labeled PECs exclusively invaded segments of the tuft affected by glomerulosclerosis, consistent with our previous findings. We next reassessed PEC recruitment in juvenile mice using a different reporter mouse and confirmed significant recruitment of labeled PECs onto the glomerular tuft. Moreover, some labeled cells on Bowman's capsule expressed podocyte markers, and cells on Bowman's capsule were also directly labeled in juvenile podocyte-specific Pod-rtTA transgenic mice. In 6-week-old mice, however, cells on Bowman's capsule no longer expressed podocyte-specific markers. Similarly, in human kidneys, some cells on Bowman's capsule expressed the podocyte marker synaptopodin from 2 weeks to 2 years of age but not at 7 years of age. In summary, podocyte regeneration from PECs could not be detected in aging mice or models of glomerular hypertrophy. We propose that a small fraction of committed podocytes reside on Bowman's capsule close to the vascular stalk and are recruited onto the glomerular tuft during infancy to adolescence in mice and humans.

  18. Depleted uranium induces neoplastic transformation in human lung epithelial cells.

    Science.gov (United States)

    Xie, Hong; LaCerte, Carolyne; Thompson, W Douglas; Wise, John Pierce

    2010-02-15

    Depleted uranium (DU) is commonly used in military armor and munitions, and thus, exposure of soldiers and noncombatants is frequent and widespread. Previous studies have shown that DU has both chemical and radiological toxicity and that the primary route of exposure of DU to humans is through inhalation and ingestion. However, there is limited research information on the potential carcinogenicity of DU in human bronchial cells. Accordingly, we determined the neoplastic transforming ability of particulate DU to human bronchial epithelial cells (BEP2D). We observed the loss of contact inhibition and anchorage independent growth in cells exposed to DU after 24 h. We also characterized these DU-induced transformed cell lines and found that 40% of the cell lines exhibit alterations in plating efficiency and no significant changes in the cytotoxic response to DU. Cytogenetic analyses showed that 53% of the DU-transformed cell lines possess a hypodiploid phenotype. These data indicate that human bronchial cells are transformed by DU and exhibit significant chromosome instability consistent with a neoplastic phenotype.

  19. Regulation of potassium transport in human lens epithelial cells.

    Science.gov (United States)

    Lauf, Peter K; Warwar, Ronald; Brown, Thomas L; Adragna, Norma C

    2006-01-01

    The major K influx pathways and their response to thiol modification by N-ethylmaleimide (NEM) and protein kinase and phosphatase inhibitors were characterized in human lens epithelial B3 (HLE-B3) cells with Rb as K congener. Ouabain (0.1 mM) and bumetanide (5 microM) discriminated between the Na/K pump ( approximately 35% of total Rb influx) and Na-K-2Cl cotransport (NKCC) ( approximately 50%). Cl-replacement with nitrate or sulfamate revealed 100 microM, activated the Na/K pump and abolished NKCC but did not affect KCC. The data suggest at least partial inverse regulation of KCC and NKCC in HLE-B3 cells by signaling cascades involving serine, threonine and tyrosine phosphorylation/dephosphorylation equilibria.

  20. Mechanisms of Cell Polarity-Controlled Epithelial Homeostasis and Immunity in the Intestine.

    Science.gov (United States)

    Klunder, Leon J; Faber, Klaas Nico; Dijkstra, Gerard; van IJzendoorn, Sven C D

    2017-07-05

    Intestinal epithelial cell polarity is instrumental to maintain epithelial homeostasis and balance communications between the gut lumen and bodily tissue, thereby controlling the defense against gastrointestinal pathogens and maintenance of immune tolerance to commensal bacteria. In this review, we highlight recent advances with regard to the molecular mechanisms of cell polarity-controlled epithelial homeostasis and immunity in the human intestine. Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.

  1. Urokinase and the intestinal mucosa: evidence for a role in epithelial cell turnover

    OpenAIRE

    Gibson, P; Birchall, I; Rosella, O; Albert, V; Finch, C; Barkla, D; Young, G

    1998-01-01

    Background—The functions of urokinase in intestinal epithelia are unknown. 
Aims—To determine the relation of urokinase expressed by intestinal epithelial cells to their position in the crypt-villus/surface axis and of mucosal urokinase activity to epithelial proliferative kinetics in the distal colon. 
Methods—Urokinase expression was examined immunohistochemically in human intestinal mucosa. Urokinase activity was measured colorimetrically in epithelial cells isolated sequ...

  2. Canine Mammary Cancer Stem Cells are Radio- and Chemo-Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype

    International Nuclear Information System (INIS)

    Pang, Lisa Y.; Cervantes-Arias, Alejandro; Else, Rod W.; Argyle, David J.

    2011-01-01

    Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs) can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT) has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology

  3. Canine Mammary Cancer Stem Cells are Radio- and Chemo-Resistant and Exhibit an Epithelial-Mesenchymal Transition Phenotype

    Energy Technology Data Exchange (ETDEWEB)

    Pang, Lisa Y., E-mail: lisa.pang@ed.ac.uk; Cervantes-Arias, Alejandro; Else, Rod W.; Argyle, David J. [Royal (Dick) School of Veterinary Studies and Roslin Institute, The University of Edinburgh, Easter Bush, Midlothian, EH25 9RG (United Kingdom)

    2011-03-30

    Canine mammary carcinoma is the most common cancer among female dogs and is often fatal due to the development of distant metastases. In humans, solid tumors are made up of heterogeneous cell populations, which perform different roles in the tumor economy. A small subset of tumor cells can hold or acquire stem cell characteristics, enabling them to drive tumor growth, recurrence and metastasis. In veterinary medicine, the molecular drivers of canine mammary carcinoma are as yet undefined. Here we report that putative cancer stem cells (CSCs) can be isolated form a canine mammary carcinoma cell line, REM134. We show that these cells have an increased ability to form tumorspheres, a characteristic of stem cells, and that they express embryonic stem cell markers associated with pluripotency. Moreover, canine CSCs are relatively resistant to the cytotoxic effects of common chemotherapeutic drugs and ionizing radiation, indicating that failure of clinical therapy to eradicate canine mammary cancer may be due to the survival of CSCs. The epithelial to mesenchymal transition (EMT) has been associated with cancer invasion, metastasis, and the acquisition of stem cell characteristics. Our results show that canine CSCs predominantly express mesenchymal markers and are more invasive than parental cells, indicating that these cells have a mesenchymal phenotype. Furthermore, we show that canine mammary cancer cells can be induced to undergo EMT by TGFβ and that these cells have an increased ability to form tumorspheres. Our findings indicate that EMT induction can enrich for cells with CSC properties, and provide further insight into canine CSC biology.

  4. Muscarinic receptor agonists stimulate matrix metalloproteinase 1-dependent invasion of human colon cancer cells

    International Nuclear Information System (INIS)

    Raufman, Jean-Pierre; Cheng, Kunrong; Saxena, Neeraj; Chahdi, Ahmed; Belo, Angelica; Khurana, Sandeep; Xie, Guofeng

    2011-01-01

    Highlights: ► Muscarinic receptor agonists stimulated robust human colon cancer cell invasion. ► Anti-matrix metalloproteinase1 antibody pre-treatment blocks cell invasion. ► Bile acids stimulate MMP1 expression, cell migration and MMP1-dependent invasion. -- Abstract: Mammalian matrix metalloproteinases (MMPs) which degrade extracellular matrix facilitate colon cancer cell invasion into the bloodstream and extra-colonic tissues; in particular, MMP1 expression correlates strongly with advanced colon cancer stage, hematogenous metastasis and poor prognosis. Likewise, muscarinic receptor signaling plays an important role in colon cancer; muscarinic receptors are over-expressed in colon cancer compared to normal colon epithelial cells. Muscarinic receptor activation stimulates proliferation, migration and invasion of human colon cancer cells. In mouse intestinal neoplasia models genetic ablation of muscarinic receptors attenuates carcinogenesis. In the present work, we sought to link these observations by showing that MMP1 expression and activation plays a mechanistic role in muscarinic receptor agonist-induced colon cancer cell invasion. We show that acetylcholine, which robustly increases MMP1 expression, stimulates invasion of HT29 and H508 human colon cancer cells into human umbilical vein endothelial cell monolayers – this was abolished by pre-incubation with atropine, a non-selective muscarinic receptor inhibitor, and by pre-incubation with anti-MMP1 neutralizing antibody. Similar results were obtained using a Matrigel chamber assay and deoxycholyltaurine (DCT), an amidated dihydroxy bile acid associated with colon neoplasia in animal models and humans, and previously shown to interact functionally with muscarinic receptors. DCT treatment of human colon cancer cells resulted in time-dependent, 10-fold increased MMP1 expression, and DCT-induced cell invasion was also blocked by pre-treatment with anti-MMP1 antibody. This study contributes to understanding

  5. Muscarinic receptor agonists stimulate matrix metalloproteinase 1-dependent invasion of human colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Raufman, Jean-Pierre, E-mail: jraufman@medicine.umaryland.edu [Division of Gastroenterology and Hepatology, University of Maryland School of Medicine, Baltimore, MD (United States); Cheng, Kunrong; Saxena, Neeraj; Chahdi, Ahmed; Belo, Angelica; Khurana, Sandeep; Xie, Guofeng [Division of Gastroenterology and Hepatology, University of Maryland School of Medicine, Baltimore, MD (United States)

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Muscarinic receptor agonists stimulated robust human colon cancer cell invasion. Black-Right-Pointing-Pointer Anti-matrix metalloproteinase1 antibody pre-treatment blocks cell invasion. Black-Right-Pointing-Pointer Bile acids stimulate MMP1 expression, cell migration and MMP1-dependent invasion. -- Abstract: Mammalian matrix metalloproteinases (MMPs) which degrade extracellular matrix facilitate colon cancer cell invasion into the bloodstream and extra-colonic tissues; in particular, MMP1 expression correlates strongly with advanced colon cancer stage, hematogenous metastasis and poor prognosis. Likewise, muscarinic receptor signaling plays an important role in colon cancer; muscarinic receptors are over-expressed in colon cancer compared to normal colon epithelial cells. Muscarinic receptor activation stimulates proliferation, migration and invasion of human colon cancer cells. In mouse intestinal neoplasia models genetic ablation of muscarinic receptors attenuates carcinogenesis. In the present work, we sought to link these observations by showing that MMP1 expression and activation plays a mechanistic role in muscarinic receptor agonist-induced colon cancer cell invasion. We show that acetylcholine, which robustly increases MMP1 expression, stimulates invasion of HT29 and H508 human colon cancer cells into human umbilical vein endothelial cell monolayers - this was abolished by pre-incubation with atropine, a non-selective muscarinic receptor inhibitor, and by pre-incubation with anti-MMP1 neutralizing antibody. Similar results were obtained using a Matrigel chamber assay and deoxycholyltaurine (DCT), an amidated dihydroxy bile acid associated with colon neoplasia in animal models and humans, and previously shown to interact functionally with muscarinic receptors. DCT treatment of human colon cancer cells resulted in time-dependent, 10-fold increased MMP1 expression, and DCT-induced cell invasion was also blocked by pre

  6. Loss of RUNX3 expression inhibits bone invasion of oral squamous cell carcinoma.

    Science.gov (United States)

    Park, Junhee; Kim, Hyun-Jeong; Kim, Ki Rim; Lee, Sun Kyoung; Kim, Hyungkeun; Park, Kwang-Kyun; Chung, Won-Yoon

    2017-02-07

    High recurrence and lower survival rates in patients with oral squamous cell carcinoma (OSCC) are associated with its bone invasion. We identified the oncogenic role of RUNX3 during bone invasion by OSCC. Tumor growth and the generation of osteolytic lesions were significantly inhibited in mice that were subcutaneously inoculated with RUNX3-knockdown human OSCC cells. RUNX3 knockdown enhanced TGF-β-induced growth arrest and inhibited OSCC cell migration and invasion in the absence or presence of transforming growth factor-β (TGF-β), a major growth factor abundant in the bone microenvironment. RUNX3 knockdown induced cell cycle arrest at the G1 and G2 phases and promoted G2 arrest by TGF-β in Ca9.22 OSCC cells. RUNX3 knockdown also inhibited both the basal and TGF-β-induced epithelial-to-mesenchymal transition by increasing E-cadherin expression and suppressing the nuclear translocation of β-catenin. In addition, the expression and TGF-β-mediated induction of parathyroid hormone-related protein (PTHrP), one of key osteolytic factors, was blocked in RUNX3-knockdown OSCC cells. Furthermore, treating human osteoblastic cells with conditioned medium derived from RUNX3-knockdown OSCC cells reduced the receptor activator of nuclear factor-kappaB ligand (RANKL)/osteoprotegerin ratio compared with treatment with conditioned medium from RUNX3-expressing cells. These findings indicate that RUNX3 expression in OSCC cells contributes to their bone invasion and the resulting osteolysis by inducing their malignant behaviors and production of osteolytic factors. RUNX3 alone or in combination with TGF-β and PTHrP may be a useful predictive biomarker and therapeutic target for bone invasion by oral cancer.

  7. Effect of Lead on Human Middle Ear Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Shin Hye Kim

    2018-01-01

    Full Text Available Lead is a ubiquitous metal in the environment, but no studies have examined lead toxicity on the middle ear. Here, we investigated lead toxicity and its mechanism in human middle ear epithelial cells (HMEECs. Moreover, we investigated the protective effects of amniotic membrane extract (AME and chorionic membrane extract (CME against lead toxicity in HMEECs. Cell viability was analyzed using the cell counting kit, and reactive oxygen species (ROS activity was measured using a cellular ROS detection kit. After lead(II acetate trihydrate treatment, mRNA levels of various genes were assessed by semiquantitative real-time polymerase chain reaction. Following treatment with AME or CME after lead exposure, the changes in cell viability, ROS activity, and gene expression were analyzed. Exposure to >100 μg/mL of lead(II acetate trihydrate caused a significant decrease in cell viability and increased ROS production in HMEECs. Lead exposure significantly increased the mRNA expression of genes encoding inflammatory cytokines and mucins. Administration of AME or CME restored cell viability, reduced ROS activity, and ameliorated mRNA levels. Our findings suggest that environmental lead exposure is related to the development of otitis media, and AME and CME may have antioxidative and anti-inflammatory effects against lead toxicity.

  8. Elevated oxidized glutathione in cystinotic proximal tubular epithelial cells.

    Science.gov (United States)

    Wilmer, Martijn J G; de Graaf-Hess, Adriana; Blom, Henk J; Dijkman, Henry B P M; Monnens, Leo A; van den Heuvel, Lambertus P; Levtchenko, Elena N

    2005-11-18

    Cystinosis, the most frequent cause of inborn Fanconi syndrome, is characterized by the lysosomal cystine accumulation, caused by mutations in the CTNS gene. To elucidate the pathogenesis of cystinosis, we cultured proximal tubular cells from urine of cystinotic patients (n = 9) and healthy controls (n = 9), followed by immortalization with human papilloma virus (HPV E6/E7). Obtained cell lines displayed basolateral polarization, alkaline phosphatase activity, and presence of aminopeptidase N (CD-13) and megalin, confirming their proximal tubular origin. Cystinotic cell lines exhibited elevated cystine levels (0.86 +/- 0.95 nmol/mg versus 0.09 +/- 0.01 nmol/mg protein in controls, p = 0.03). Oxidized glutathione was elevated in cystinotic cells (1.16 +/- 0.83 nmol/mg versus 0.29 +/- 0.18 nmol/mg protein, p = 0.04), while total glutathione, free cysteine, and ATP contents were normal in these cells. In conclusion, elevated oxidized glutathione in cystinotic proximal tubular epithelial cell lines suggests increased oxidative stress, which may contribute to tubular dysfunction in cystinosis.

  9. Ionizing radiation induces heritable disruption of epithelial cell interactions

    Science.gov (United States)

    Park, Catherine C.; Henshall-Powell, Rhonda L.; Erickson, Anna C.; Talhouk, Rabih; Parvin, Bahram; Bissell, Mina J.; Barcellos-Hoff, Mary Helen; Chatterjee, A. (Principal Investigator)

    2003-01-01

    Ionizing radiation (IR) is a known human breast carcinogen. Although the mutagenic capacity of IR is widely acknowledged as the basis for its action as a carcinogen, we and others have shown that IR can also induce growth factors and extracellular matrix remodeling. As a consequence, we have proposed that an additional factor contributing to IR carcinogenesis is the potential disruption of critical constraints that are imposed by normal cell interactions. To test this hypothesis, we asked whether IR affected the ability of nonmalignant human mammary epithelial cells (HMEC) to undergo tissue-specific morphogenesis in culture by using confocal microscopy and imaging bioinformatics. We found that irradiated single HMEC gave rise to colonies exhibiting decreased localization of E-cadherin, beta-catenin, and connexin-43, proteins necessary for the establishment of polarity and communication. Severely compromised acinar organization was manifested by the majority of irradiated HMEC progeny as quantified by image analysis. Disrupted cell-cell communication, aberrant cell-extracellular matrix interactions, and loss of tissue-specific architecture observed in the daughters of irradiated HMEC are characteristic of neoplastic progression. These data point to a heritable, nonmutational mechanism whereby IR compromises cell polarity and multicellular organization.

  10. Lens epithelial cell apoptosis is an early event in the development of UVB-induced cataract.

    Science.gov (United States)

    Li, W C; Spector, A

    1996-01-01

    Epidemiological and experimental studies have revealed that exposure to UV can induce cataractogenesis. To investigate the mechanism of this induction, viability of the lens epithelial cells from UVB-treated rat lenses were examined. Irradiation of the cultured rat lenses with 8 J/s/m2 UVB for 60 min triggers lens epithelial cell apoptosis as determined by terminal deoxyribonucleotide transferase (TdT) labeling and DNA fragmentation assays. The apoptotic lens epithelial cells were initially found in the equatorial region and then quickly appeared in both equatorial and central regions. The percentage of apoptotic cells continuously increased during the postirradiation incubation. After a 5-h post-UVB incubation, more than 50% of the lens epithelial cells were apoptotic. By 24 h, all of the lens epithelial cells in the irradiated lenses were dead through apoptosis. Associated with this apoptotic process is a large upregulation of the proto-oncogene, c-fos. Opacification appears to follow the death of lens epithelial cells occurring first in the equatorial region and then in the central area. This is also true of classical cataract parameters such as non-protein thiol and wet weight, which are significantly modified only after appreciable epithelial cell apoptosis. Together, these results suggest that the rapid apoptotic death of the lens epithelial cells induced by UVB initiates cataract development.

  11. Fusion of intestinal epithelial cells with bone marrow derived cells is dispensable for tissue homeostasis

    NARCIS (Netherlands)

    de Jong, Joan H.; Rodermond, Hans M.; Zimberlin, Cheryl D.; Lascano, Valeria; de Sousa E Melo, Felipe; Richel, Dick J.; Medema, Jan Paul; Vermeulen, Louis

    2012-01-01

    The epithelial lining of the intestine is characterized by an immense cellular turn-over ascertaining an extensive regenerative capacity. Multiple reports suggest that besides the local intestinal stem cell pool, circulating cells of bone marrow origin (BMDCs) contribute to this process by fusing

  12. Suppression of actopaxin impairs hepatocellular carcinoma metastasis through modulation of cell migration and invasion.

    Science.gov (United States)

    Ng, Lui; Tung-Ping Poon, Ronnie; Yau, Simon; Chow, Ariel; Lam, Colin; Li, Hung-Sing; Chung-Cheung Yau, Thomas; Law, Wai-Lun; Pang, Roberta

    2013-08-01

    Early reports suggested that actopaxin, a member of the focal adhesion proteins, regulates cell migration. Here we investigated whether actopaxin is involved in hepatocellular carcinoma (HCC) progression and metastasis. We examined actopaxin expression in human HCC samples using immunohistochemistry and western blotting. The functional and molecular effect of actopaxin was studied in vitro by overexpression in a nonmetastatic HCC cell line, as well as repression in a metastatic cell line. The in vivo effect of actopaxin repression was studied in nonobese diabetic and severe combined immunodeficient mice. We found that actopaxin was frequently overexpressed in human HCC patients and its overexpression positively correlated with tumor size, stage, and metastasis. Actopaxin expression also correlated with the metastatic potential of HCC cell lines. Actopaxin overexpression induced the invasion and migration ability of nonmetastatic HCC cells, whereas down-regulation of actopaxin reverted the invasive phenotypes and metastatic potential of metastatic HCC cells through regulating the protein expression of certain focal adhesion proteins including ILK, PINCH, paxillin, and cdc42, as well as regulating the epithelial-mesenchymal transition pathway. Furthermore, there was a close association between actopaxin and CD29. HCC cells with stronger CD29 expression showed a higher actopaxin level, whereas actopaxin repression attenuated CD29 activity. Finally, actopaxin down-regulation enhanced the chemosensitivity of HCC cells towards oxaliplatin treatment by way of a collective result of suppression of survivin protein, β-catenin, and mammalian target of rapamycin pathways and up-regulation of p53. This study provides concrete evidence of a significant role of actopaxin in HCC progression and metastasis, by way of regulation of cell invasiveness and motility, an epithelial-mesenchymal transition process, and chemosensitivity to cytotoxic drugs. Copyright © 2013 by the

  13. Identification of six new susceptibility loci for invasive epithelial ovarian cancer

    NARCIS (Netherlands)

    Kuchenbaecker, K.B.; Ramus, S.J.; Tyrer, J.; Lee, A.; Shen, H.C.; Beesley, J.; Lawrenson, K.; McGuffog, L.; Healey, S.; Lee, J.M.; Spindler, T.J.; Lin, Y.G.; Pejovic, T.; Bean, Y.; Li, Q.; Coetzee, S.; Hazelett, D.; Miron, A.; Southey, M.; Terry, M.B.; Goldgar, D.E.; Buys, S.S.; Janavicius, R.; Dorfling, C.M.; Rensburg, E.J. van; Neuhausen, S.L.; Ding, Y.C.; Hansen, T.V.; Jonson, L.; Gerdes, A.M.; Ejlertsen, B.; Barrowdale, D.; Dennis, J.; Benitez, J.; Osorio, A.; Garcia, M.J.; Komenaka, I.; Weitzel, J.N.; Ganschow, P.; Peterlongo, P.; Bernard, L.; Viel, A.; Bonanni, B.; Peissel, B.; Manoukian, S.; Radice, P.; Papi, L.; Ottini, L.; Fostira, F.; Konstantopoulou, I.; Garber, J.; Frost, D.; Perkins, J.; Platte, R.; Ellis, S.; Embrace, .; Godwin, A.K.; Schmutzler, R.K.; Meindl, A.; Engel, C.; Sutter, C.; Sinilnikova, O.M.; Damiola, F.; Mazoyer, S.; Stoppa-Lyonnet, D.; Claes, K.; Leeneer, K. De; Kirk, J.; Rodriguez, G.C.; Piedmonte, M.; O'Malley, D.M.; Hoya, M. de la; Caldes, T.; Aittomaki, K.; Nevanlinna, H.; Collee, J.M.; Rookus, M.A.; Oosterwijk, J.C; Tihomirova, L.; Tung, N.; Hamann, U.; Isaccs, C.; Tischkowitz, M.; Imyanitov, E.N.; Caligo, M.A.; Campbell, I.G.; Hogervorst, F.B.; Olah, E.; Diez, O.; Blanco, I.; Brunet, J.; Lazaro, C.; Pujana, M.A.; Jakubowska, A.; Gronwald, J.; Lubinski, J.; Sukiennicki, G.; Massuger, L.F.A.G.; Altena, A.M. van; Aben, K.K.H.; Kiemeney, B.; Mensenkamp, A.R.; Kets, M.; Hoogerbrugge, N.; Ligtenberg, M.J.L.; et al.,

    2015-01-01

    Genome-wide association studies (GWAS) have identified 12 epithelial ovarian cancer (EOC) susceptibility alleles. The pattern of association at these loci is consistent in BRCA1 and BRCA2 mutation carriers who are at high risk of EOC. After imputation to 1000 Genomes Project data, we assessed

  14. Identification of six new susceptibility loci for invasive epithelial ovarian cancer

    NARCIS (Netherlands)

    K.B. Kuchenbaecker (Karoline); S.J. Ramus (Susan); J.P. Tyrer (Jonathan); A. Lee (Andrew); H.C. Shen (Howard C.); J. Beesley (Jonathan); K. Lawrenson (Kate); L. McGuffog (Lesley); S. Healey (Sue); J.M. Lee (Janet M.); T.J. Spindler (Tassja J.); Y.G. Lin (Yvonne G.); T. Pejovic (Tanja); Y. Bean (Yukie); Q. Li (Qiyuan); S. Coetzee (Simon); D. Hazelett (Dennis); A. Miron (Alexander); M.C. Southey (Melissa); M.B. Terry (Mary Beth); D. Goldgar (David); S.S. Buys (Saundra); R. Janavicius (Ramunas); C.M. Dorfling (Cecilia); E.J. van Rensburg (Elizabeth); S.L. Neuhausen (Susan); Y.C. Ding (Yuan); T.V.O. Hansen (Thomas); L. Jønson (Lars); A.-M. Gerdes (Anne-Marie); B. Ejlertsen (Bent); D. Barrowdale (Daniel); J. Dennis (Joe); J. Benítez (Javier); A. Osorio (Ana); M.J. Garcia (Maria Jose); I. Komenaka (Ian); J.N. Weitzel (Jeffrey); P. Ganschow (Pamela); P. Peterlongo (Paolo); L. Bernard (Loris); A. Viel (Alessandra); B. Bonnani (Bernardo); B. Peissel (Bernard); S. Manoukian (Siranoush); P. Radice (Paolo); L. Papi (Laura); L. Ottini (Laura); F. Fostira (Florentia); I. Konstantopoulou (I.); J. Garber (Judy); D. Frost (Debra); J. Perkins (Jo); R. Platte (Radka); S.D. Ellis (Steve); A.K. Godwin (Andrew K.); R.K. Schmutzler (Rita); A. Meindl (Alfons); C. Engel (Christoph); C. Sutter (Christian); O. Sinilnikova (Olga); F. Damiola (Francesca); S. Mazoyer (Sylvie); D. Stoppa-Lyonnet (Dominique); K.B.M. Claes (Kathleen B.M.); K. De Leeneer (Kim); J. Kirk (Judy); G. Rodriguez (Gustavo); M. Piedmonte (Marion); D.M. O'Malley (David M.); M. de La Hoya (Miguel); T. Caldes (Trinidad); K. Aittomäki (Kristiina); H. Nevanlinna (Heli); J.M. Collée (Margriet); M.A. Rookus (Matti); J.C. Oosterwijk (Jan); L. Tihomirova (Laima); N. Tung (Nadine); U. Hamann (Ute); C. Isaccs (Claudine); M. Tischkowitz (Marc); E.N. Imyanitov (Evgeny); M.A. Caligo (Maria); I. Campbell (Ian); F.B.L. Hogervorst (Frans); E. Olah; O. Díez (Orland); I. Blanco (Ignacio); J. Brunet (Joan); C. Lazaro (Conxi); M.A. Pujana (Miguel); A. Jakubowska (Anna); J. Gronwald (Jacek); J. Lubinski (Jan); G. Sukiennicki (Grzegorz); R.B. Barkardottir (Rosa); M. Plante (Marie); J. Simard (Jacques); P. Soucy (Penny); M. Montagna (Marco); S. Tognazzo (Silvia); P.J. Teixeira; V.S. Pankratz (Shane); X. Wang (Xianshu); N.M. Lindor (Noralane); C. Szabo (Csilla); N. Kauff (Noah); J. Vijai (Joseph); C.A. Aghajanian (Carol A.); G. Pfeiler (Georg); A. Berger (Andreas); C.F. Singer (Christian); M.-K. Tea; C. Phelan (Catherine); M.H. Greene (Mark H.); P.L. Mai (Phuong); G. Rennert (Gad); A.-M. Mulligan (Anna-Marie); S. Tchatchou (Sandrine); I.L. Andrulis (Irene); G. Glendon (Gord); A.E. Toland (Amanda); U.B. Jensen (Uffe Birk); T.A. Kruse (Torben); M. Thomassen (Mads); A. Bojesen (Anders); J. Zidan (Jamal); E. Friedman (Eitan); Y. Laitman (Yael); M. Soller (Maria); A. Liljegren (Annelie); B. Arver (Brita Wasteson); Z. Einbeigi (Zakaria); M. Stenmark-Askmalm (Marie); O.I. Olopade (Olufunmilayo I.); R.L. Nussbaum (Robert L.); T.R. Rebbeck (Timothy R.); K.L. Nathanson (Katherine); S.M. Domchek (Susan); K.H. Lu (Karen); B.Y. Karlan (Beth Y.); C. Walsh (Christine); K.J. Lester (Kathryn); R. Hein (Rebecca); A.B. Ekici (Arif); M.W. Beckmann (Matthias); P.A. Fasching (Peter); D. Lambrechts (Diether); E. Van Nieuwenhuysen (Els); I. Vergote (Ignace); S. Lambrechts (Sandrina); E. Dicks (Ed); J.A. Doherty (Jennifer A.); K.G. Wicklund (Kristine G.); M.A. Rossing (Mary Anne); A. Rudolph (Anja); J. Chang-Claude (Jenny); S. Wang-Gohrke (Shan); U. Eilber (Ursula); K.B. Moysich (Kirsten B.); K. Odunsi (Kunle); L. Sucheston (Lara); S. Lele (Shashi); L. Wilkens (Lynne); M.T. Goodman (Marc); P.J. Thompson (Pamela J.); Y.B. Shvetsov (Yurii B.); I.B. Runnebaum (Ingo); M. Dürst (Matthias); P. Hillemanns (Peter); T. Dörk (Thilo); N.N. Antonenkova (Natalia); N.V. Bogdanova (Natalia); A. Leminen (Arto); L.M. Pelttari (Liisa); R. Butzow (Ralf); F. Modugno (Francesmary); J.L. Kelley (Joseph L.); R. Edwards (Robert); R.B. Ness (Roberta); A. Du Bois (Andreas); P.U. Heitz; I. Schwaab (Ira); P. Harter (Philipp); K. Matsuo (Keitaro); N. Hosono (Naoya); S. Orsulic (Sandra); A. Jensen (Allan); M. Kjaer (Michael); E. Høgdall (Estrid); H.N. Hasmad (Hanis Nazihah); M.A. Noor Azmi (Mat Adenan); S.-H. Teo (Soo-Hwang); Y.L. Woo (Yin Ling); B.L. Fridley (Brooke); E.L. Goode (Ellen); J.M. Cunningham (Julie); R.A. Vierkant (Robert); F. Bruinsma (Fiona); G.G. Giles (Graham G.); D. Liang (Dong); M.A.T. Hildebrandt (Michelle A.T.); X. Wu (Xifeng); D.A. Levine (Douglas); M. Bisogna (Maria); A. Berchuck (Andrew); E. Iversen (Erik); J.M. Schildkraut (Joellen); P. Concannon (Patrick); R.P. Weber (Rachel Palmieri); D.W. Cramer (Daniel); K.L. Terry (Kathryn); E.M. Poole (Elizabeth); S. Tworoger (Shelley); E.V. Bandera (Elisa); I. Orlow (Irene); S.H. Olson (Sara); C. Krakstad (Camilla); H.B. Salvesen (Helga); I.L. Tangen (Ingvild L.); L. Bjorge (Line); A.M. van Altena (Anne); K.K.H. Aben (Katja); L.A.L.M. Kiemeney (Bart); L.F. Massuger (Leon); M. Kellar (Melissa); A. Brooks-Wilson (Angela); L.E. Kelemen (Linda); L.S. Cook (Linda S.); N.D. Le (Nhu D.); C. Cybulski (Cezary); H. Yang (Hannah); J. Lissowska (Jolanta); L.A. Brinton (Louise); N. Wentzensen (N.); C.K. Høgdall (Claus); L. Lundvall (Lene); L. Nedergaard (Lotte); H. Baker (Helen); H. Song (Honglin); D. Eccles (Diana); I. McNeish (Ian); J. Paul (James); K. Carty (Karen); N. Siddiqui (Nadeem); R. Glasspool (Rosalind); A.S. Whittemore (Alice S.); J.H. Rothstein (Joseph H.); W.P. McGuire; W. Sieh (Weiva); B.-T. Ji (Bu-Tian); W. Zheng (Wei); X.-O. Shu (Xiao-Ou); Y. Gao; B. Rosen (Barry); H. Risch (Harvey); J. McLaughlin (John); S.A. Narod (Steven A.); A.N.A. Monteiro (Alvaro N.); A. Chen (Ann); H.-Y. Lin (Hui-Yi); J. Permuth-Wey (Jenny); T.F. Sellers; Y.-Y. Tsai (Ya-Yu); Z. Chen (Zhihua); A. Ziogas (Argyrios); H. Anton-Culver (Hoda); A. Gentry-Maharaj (Aleksandra); U. Menon (Usha); P. harrington (Patricia); A.W. Lee (Alice W.); A.H. Wu (Anna H.); C.L. Pearce (Celeste); G. Coetzee (Gerry); M.C. Pike (Malcolm C.); A. Dansonka-Mieszkowska (Agnieszka); A. Timorek (Agnieszka); I.K. Rzepecka (Iwona); J. Kupryjanczyk (Jolanta); M. Freedman (Matthew); H. Noushmehr (Houtan); D.F. Easton (Douglas F.); K. Offit (Kenneth); F.J. Couch (Fergus); S.A. Gayther (Simon); P.P.D.P. Pharoah (Paul P.D.P.); A.C. Antoniou (Antonis C.); G. Chenevix-Trench (Georgia)

    2015-01-01

    textabstractGenome-wide association studies (GWAS) have identified 12 epithelial ovarian cancer (EOC) susceptibility alleles. The pattern of association at these loci is consistent in BRCA1 and BRCA2 mutation carriers who are at high risk of EOC. After imputation to 1000 Genomes Project data, we

  15. Synergy between type 1 fimbriae expression and C3 opsonisation increases internalisation of E. coli by human tubular epithelial cells.

    Science.gov (United States)

    Li, Ke; Zhou, Wuding; Hong, Yuzhi; Sacks, Steven H; Sheerin, Neil S

    2009-03-31

    Bacterial infection of the urinary tract is a common clinical problem with E. coli being the most common urinary pathogen. Bacterial uptake into epithelial cells is increasingly recognised as an important feature of infection. Bacterial virulence factors, especially fimbrial adhesins, have been conclusively shown to promote host cell invasion. Our recent study reported that C3 opsonisation markedly increases the ability of E. coli strain J96 to internalise into human proximal tubular epithelial cells via CD46, a complement regulatory protein expressed on host cell membrane. In this study, we further assessed whether C3-dependent internalisation by human tubular epithelial cells is a general feature of uropathogenic E. coli and investigated features of the bacterial phenotype that may account for any heterogeneity. In 31 clinical isolates of E. coli tested, C3-dependent internalisation was evident in 10 isolates. Type 1 fimbriae mediated-binding is essential for C3-dependent internalisation as shown by phenotypic association, type 1 fimbrial blockade with soluble ligand (mannose) and by assessment of a type 1 fimbrial mutant. we propose that efficient internalisation of uropathogenic E. coli by the human urinary tract depends on co-operation between type 1 fimbriae-mediated adhesion and C3 receptor -ligand interaction.

  16. Connective tissue growth factor activates pluripotency genes and mesenchymal-epithelial transition in head and neck cancer cells.

    Science.gov (United States)

    Chang, Cheng-Chi; Hsu, Wen-Hao; Wang, Chen-Chien; Chou, Chun-Hung; Kuo, Mark Yen-Ping; Lin, Been-Ren; Chen, Szu-Ta; Tai, Shyh-Kuan; Kuo, Min-Liang; Yang, Muh-Hwa

    2013-07-01

    The epithelial-mesenchymal transition (EMT) is a key mechanism in both embryonic development and cancer metastasis. The EMT introduces stem-like properties to cancer cells. However, during somatic cell reprogramming, mesenchymal-epithelial transition (MET), the reverse process of EMT, is a crucial step toward pluripotency. Connective tissue growth factor (CTGF) is a multifunctional secreted protein that acts as either an oncoprotein or a tumor suppressor among different cancers. Here, we show that in head and neck squamous cell carcinoma (HNSCC), CTGF promotes the MET and reduces invasiveness. Moreover, we found that CTGF enhances the stem-like properties of HNSCC cells and increases the expression of multiple pluripotency genes. Mechanistic studies showed that CTGF induces c-Jun expression through αvβ3 integrin and that c-Jun directly activates the transcription of the pluripotency genes NANOG, SOX2, and POU5F1. Knockdown of CTGF in TW2.6 cells was shown to reduce tumor formation and attenuate E-cadherin expression in xenotransplanted tumors. In HNSCC patient samples, CTGF expression was positively correlated with the levels of CDH1, NANOG, SOX2, and POU5F1. Coexpression of CTGF and the pluripotency genes was found to be associated with a worse prognosis. These findings are valuable in elucidating the interplay between epithelial plasticity and stem-like properties during cancer progression and provide useful information for developing a novel classification system and therapeutic strategies for HNSCC. ©2013 AACR.

  17. Establishment and characterization of novel epithelial-like cell lines derived from human periodontal ligament tissue in vitro.

    Science.gov (United States)

    Tansriratanawong, Kallapat; Ishikawa, Hiroshi; Toyomura, Junko; Sato, Soh

    2017-10-01

    In this study, novel human-derived epithelial-like cells (hEPLCs) lines were established from periodontal ligament (PDL) tissues, which were composed of a variety of cell types and exhibited complex cellular activities. To elucidate the putative features distinguishing these from epithelial rest of Malassez (ERM), we characterized hEPLCs based on cell lineage markers and tight junction protein expression. The aim of this study was, therefore, to establish and characterize hEPLCs lines from PDL tissues. The hEPLCs were isolated from PDL of third molar teeth. Cellular morphology and cell organelles were observed thoroughly. The characteristics of epithelial-endothelial-mesenchymal-like cells were compared in several markers by gene expression and immunofluorescence, to ERM and human umbilical-vein endothelial cells (HUVECs). The resistance between cellular junctions was assessed by transepithelial electron resistance, and inflammatory cytokines were detected by ELISA after infecting hEPLCs with periodontopathic bacteria. The hEPLCs developed into small epithelial-like cells in pavement appearance similar to ERM. However, gene expression patterns and immunofluorescence results were different from ERM and HUVECs, especially in tight junction markers (Claudin, ZO-1, and Occludins), and endothelial markers (vWF, CD34). The transepithelial electron resistance indicated higher resistance in hEPLCs, as compared to ERM. Periodontopathic bacteria were phagocytosed with upregulation of inflammatory cytokine secretion within 24 h. In conclusion, hEPLCs that were derived using the single cell isolation method formed tight multilayers colonies, as well as strongly expressed tight junction markers in gene expression and immunofluorescence. Novel hEPLCs lines exhibited differently from ERM, which might provide some specific functions such as metabolic exchange and defense mechanism against bacterial invasion in periodontal tissue.

  18. Physiology and pathophysiology of apoptosis in epithelial cells of the liver, pancreas, and intestine.

    Science.gov (United States)

    Jones, B A; Gores, G J

    1997-12-01

    Cell death of gastrointestinal epithelial cells occurs by a process referred to as apoptosis. In this review, we succinctly define apoptosis and summarize the role of apoptosis in the physiology and pathophysiology of epithelial cells in the liver, pancreas, and small and large intestine. The physiological mediators regulating apoptosis in gastrointestinal epithelial cells, when known, are discussed. Selected pathophysiological consequences of excessive apoptosis and inhibition of apoptosis are used to illustrate the significance of apoptosis in disease processes. These examples demonstrate that excessive apoptosis may result in epithelial cell atrophy, injury, and dysfunction, whereas inhibition of apoptosis results in hyperplasia and promotes malignant transformation. The specific cellular mechanisms responsible for dysregulation of epithelial cell apoptosis during pathophysiological disturbances are emphasized. Potential future areas of physiological research regarding apoptosis in gastrointestinal epithelia are highlighted when appropriate.

  19. Characterization of a continuous feline mammary epithelial cell line susceptible to feline epitheliotropic viruses.

    Science.gov (United States)

    Pesavento, Patricia; Liu, Hongwei; Ossiboff, Robert J; Stucker, Karla M; Heymer, Anna; Millon, Lee; Wood, Jason; van der List, Deborah; Parker, John S L

    2009-04-01

    Mucosal epithelial cells are the primary targets for many common viral pathogens of cats. Viral infection of epithelia can damage or disrupt the epithelial barrier that protects underlying tissues. In vitro cell culture systems are an effective means to study how viruses infect and disrupt epithelial barriers, however no true continuous or immortalized feline epithelial cell culture lines are available. A continuous cell culture of feline mammary epithelial cells (FMEC UCD-04-2) that forms tight junctions with high transepithelial electrical resistance (>2000Omegacm(-1)) 3-4 days after reaching confluence was characterized. In addition, it was shown that FMECs are susceptible to infection with feline calicivirus (FCV), feline herpesvirus (FHV-1), feline coronavirus (FeCoV), and feline panleukopenia virus (FPV). These cells will be useful for studies of feline viral disease and for in vitro studies of feline epithelia.

  20. Donor-derived stem-cells and epithelial mesenchymal transition in squamous cell carcinoma in transplant recipients.

    Science.gov (United States)

    Verneuil, Laurence; Leboeuf, Christophe; Bousquet, Guilhem; Brugiere, Charlotte; Elbouchtaoui, Morad; Plassa, Louis-François; Peraldi, Marie-Noelle; Lebbé, Celeste; Ratajczak, Philippe; Janin, Anne

    2015-12-08

    Skin squamous-cell-carcinoma (SCC), is the main complication in long-term kidney-transplant recipients, and it can include donor-derived cells. Preclinical models demonstrated the involvement of epithelial mesenchymal transition (EMT) in the progression of skin SCC, and the role of Snail, an EMT transcription factor, in cancer stem-cell survival and expansion.Here, we studied stem-cells and EMT expression in SCCs and concomitant actinic keratoses (AK) in kidney-transplant recipients. In SCC and AK in 3 female recipients of male kidney-transplants, donor-derived Y chromosome in epidermal stem cells was assessed using combined XY-FISH/CD133 immunostaining, and digital-droplet-PCR on laser-microdissected CD133 expressing epidermal cells.For EMT study, double immunostainings of CD133 with vimentin or snail and slug, electron microscopy and immunostainings of keratinocytes junctions were performed. Digital droplet PCR was used to check CDH1 (E-cadherin) expression level in laser-microdissected cells co-expressing CD133 and vimentin or snail and slug.The numbers of Y-chromosome were assessed using digital droplet PCR in laser-microdissected cells co-expressing CD133 and vimentin, or snail and slug, and in CD133 positive cells not expressing any EMT maker. We identified donor-derived stem-cells in basal layers and invasive areas in all skin SCCs and in concomitant AKs, but not in surrounding normal skin.The donor-derived stem-cells expressed the EMT markers, vimentin, snail and slug in SCCs but not in AKs. The expression of the EMT transcription factor, SNAI1, was higher in stem-cells when they expressed vimentin. They were located in invasive areas of SCCs. In these areas, the expressions of claudin-1 and desmoglein 1 were reduced or absent, and within the basal layer there were features of basal membrane disappearance.Donor-derived stem cells were in larger numbers in stem cells co-expressing vimentin or snail and slug than in stem cells not expressing any EMT marker

  1. Polystyrene nanoparticles activate ion transport in human airway epithelial cells

    Directory of Open Access Journals (Sweden)

    McCarthy J

    2011-06-01

    Full Text Available J McCarthy1, X Gong2, D Nahirney2, M Duszyk2, MW Radomski11School of Pharmacy and Pharmaceutical Sciences, Panoz Institute, Trinity College Dublin, Dublin, Ireland; 2Department of Physiology, University of Alberta, Edmonton, Alberta, CanadaBackground: Over the last decade, nanotechnology has provided researchers with new nanometer materials, such as nanoparticles, which have the potential to provide new therapies for many lung diseases. In this study, we investigated the acute effects of polystyrene nanoparticles on epithelial ion channel function.Methods: Human submucosal Calu-3 cells that express cystic fibrosis transmembrane conductance regulator (CFTR and baby hamster kidney cells engineered to express the wild-type CFTR gene were used to investigate the actions of negatively charged 20 nm polystyrene nanoparticles on short-circuit current in Calu-3 cells by Ussing chamber and single CFTR Cl- channels alone and in the presence of known CFTR channel activators by using baby hamster kidney cell patches.Results: Polystyrene nanoparticles caused sustained, repeatable, and concentration-dependent increases in short-circuit current. In turn, these short-circuit current responses were found to be biphasic in nature, ie, an initial peak followed by a plateau. EC50 values for peak and plateau short-circuit current responses were 1457 and 315.5 ng/mL, respectively. Short-circuit current was inhibited by diphenylamine-2-carboxylate, a CFTR Cl- channel blocker. Polystyrene nanoparticles activated basolateral K+ channels and affected Cl- and HCO3- secretion. The mechanism of short-circuit current activation by polystyrene nanoparticles was found to be largely dependent on calcium-dependent and cyclic nucleotide-dependent phosphorylation of CFTR Cl- channels. Recordings from isolated inside-out patches using baby hamster kidney cells confirmed the direct activation of CFTR Cl- channels by the nanoparticles.Conclusion: This is the first study to identify

  2. Invasive Glioblastoma Cells Acquire Stemness and Increased Akt Activation

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    Jennifer R. Molina

    2010-06-01

    Full Text Available Glioblastoma multiforme (GBM is the most frequent and most aggressive brain tumor in adults. The dismal prognosis is due to postsurgery recurrences arising from escaped invasive tumor cells. The signaling pathways activated in invasive cells are under investigation, and models are currently designed in search for therapeutic targets. We developed here an in vivo model of human invasive GBM in mouse brain from a GBM cell line with moderate tumorigenicity that allowed simultaneous primary tumor growth and dispersal of tumor cells in the brain parenchyma. This strategy allowed for the first time the isolation and characterization of matched sets of tumor mass (Core and invasive (Inv cells. Both cell populations, but more markedly Inv cells, acquired stem cell markers, neurosphere renewal ability, and resistance to rapamycin-induced apoptosis relative to parental cells. The comparative phenotypic analysis between Inv and Core cells showed significantly increased tumorigenicity in vivo and increased invasion with decreased proliferation in vitro for Inv cells. Examination of a large array of signaling pathways revealed extracellular signal-regulated kinase (Erk down-modulation and Akt activation in Inv cells and an opposite profile in Core cells. Akt activation correlated with the increased tumorigenicity, stemness, and invasiveness, whereas Erk activation correlated with the proliferation of the cells. These results underscore complementary roles of the Erk and Akt pathways for GBM proliferation and dispersal and raise important implications for a concurrent inhibitory therapy.

  3. Quantification of epithelial cells in coculture with fibroblasts by fluorescence image analysis.

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    Krtolica, Ana; Ortiz de Solorzano, Carlos; Lockett, Stephen; Campisi, Judith

    2002-10-01

    To demonstrate that senescent fibroblasts stimulate the proliferation and neoplastic transformation of premalignant epithelial cells (Krtolica et al.: Proc Natl Acad Sci USA 98:12072-12077, 2001), we developed methods to quantify the proliferation of epithelial cells cocultured with fibroblasts. We stained epithelial-fibroblast cocultures with the fluorescent DNA-intercalating dye 4,6-diamidino-2-phenylindole (DAPI), or expressed green fluorescent protein (GFP) in the epithelial cells, and then cultured them with fibroblasts. The cocultures were photographed under an inverted microscope with appropriate filters, and the fluorescent images were captured with a digital camera. We modified an image analysis program to selectively recognize the smaller, more intensely fluorescent epithelial cell nuclei in DAPI-stained cultures and used the program to quantify areas with DAPI fluorescence generated by epithelial nuclei or GFP fluorescence generated by epithelial cells in each field. Analysis of the image areas with DAPI and GFP fluorescences produced nearly identical quantification of epithelial cells in coculture with fibroblasts. We confirmed these results by manual counting. In addition, GFP labeling permitted kinetic studies of the same coculture over multiple time points. The image analysis-based quantification method we describe here is an easy and reliable way to monitor cells in coculture and should be useful for a variety of cell biological studies. Copyright 2002 Wiley-Liss, Inc.

  4. Urokinase and the intestinal mucosa: evidence for a role in epithelial cell turnover

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    Gibson, P; Birchall, I; Rosella, O; Albert, V; Finch, C; Barkla, D; Young, G

    1998-01-01

    Background—The functions of urokinase in intestinal epithelia are unknown. 
Aims—To determine the relation of urokinase expressed by intestinal epithelial cells to their position in the crypt-villus/surface axis and of mucosal urokinase activity to epithelial proliferative kinetics in the distal colon. 
Methods—Urokinase expression was examined immunohistochemically in human intestinal mucosa. Urokinase activity was measured colorimetrically in epithelial cells isolated sequentially from the crypt-villus axis of the rat small intestine. In separate experiments, urokinase activity and epithelial kinetics (measured stathmokinetically) were measured in homogenates of distal colonic mucosa of 14 groups of eight rats fed diets known to alter epithelial turnover. 
Results—From the crypt base, an ascending gradient of expression and activity of urokinase was associated with the epithelial cells. Median mucosal urokinase activities in each of the dietary groups of rats correlated positively with autologous median number of metaphase arrests per crypt (r=0.68; p<0.005) and per 100 crypt cells (r=0.75; p<0.001), but not with crypt column height. 
Conclusions—Localisation of an enzyme capable of leading to digestion of cell substratum in the region where cells are loosely attached to their basement membrane, and the association of its activity with indexes of cell turnover, suggest a role for urokinase in facilitating epithelial cell loss in the intestine. 

 Keywords: urokinase; intestinal epithelium; colon; epithelial proliferation PMID:9824347

  5. PKC activation induces inflammatory response and cell death in human bronchial epithelial cells.

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    Hyunhee Kim

    Full Text Available A variety of airborne pathogens can induce inflammatory responses in airway epithelial cells, which is a crucial component of host defence. However, excessive inflammatory responses and chronic inflammation also contribute to different diseases of the respiratory system. We hypothesized that the activation of protein kinase C (PKC is one of the essential mechanisms of inflammatory response in airway epithelial cells. In the present study, we stimulated human bronchial lung epithelial (BEAS-2B cells with the phorbol ester Phorbol 12, 13-dibutyrate (PDBu, and examined gene expression profile using microarrays. Microarray analysis suggests that PKC activation induced dramatic changes in gene expression related to multiple cellular functions. The top two interaction networks generated from these changes were centered on NFκB and TNF-α, which are two commonly known pathways for cell death and inflammation. Subsequent tests confirmed the decrease in cell viability and an increase in the production of various cytokines. Interestingly, each of the increased cytokines was differentially regulated at mRNA and/or protein levels by different sub-classes of PKC isozymes. We conclude that pathological cell death and cytokine production in airway epithelial cells in various situations may be mediated through PKC related signaling pathways. These findings suggest that PKCs can be new targets for treatment of lung diseases.

  6. Roles of Wnt/β-catenin signaling in epithelial differentiation of mesenchymal stem cells

    International Nuclear Information System (INIS)

    Wang, Yajing; Sun, Zhaorui; Qiu, Xuefeng; Li, Yan; Qin, Jizheng; Han, Xiaodong

    2009-01-01

    Bone marrow-derived mesenchymal stem cells (MSCs) have been demonstrated to be able to differentiate into epithelial lineage, but the precise mechanisms controlling this process are unclear. Our aim is to explore the roles of Wnt/β-catenin in the epithelial differentiation of MSCs. Using indirect co-culture of rat MSCs with rat airway epithelial cells (RTE), MSCs expressed several airway epithelial markers (cytokeratin 18, tight junction protein occudin, cystic fibrosis transmembrance regulator). The protein levels of some important members in Wnt/β-catenin signaling were determined, suggested down-regulation of Wnt/β-catenin with epithelial differentiation of MSCs. Furthermore, Wnt3α can inhibit the epithelial differentiation of MSCs. A loss of β-catenin induced by Dickkopf-1 can enhance MSCs differentiation into epithelial cells. Lithium chloride transiently activated β-catenin expression and subsequently decreased β-catenin level and at last inhibited MSCs to differentiate into airway epithelium. Taken together, our study indicated that RTE cells can trigger epithelial differentiation of MSCs. Blocking Wnt/β-catenin signaling may promote MSCs to differentiate towards airway epithelial cells.

  7. Hormonal regulation of Na -K -ATPase in cultured epithelial cells

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    Johnson, J.P.; Jones, D.; Wiesmann, W.P.

    1986-08-01

    Aldosterone and insulin stimulate Na transport through mechanisms involving protein synthesis. Na -K -ATPase has been implicated in the action of both hormones. The authors examined the effect of aldosterone and insulin on Na -K -ATPase in epithelial cells in culture derived from toad urinary bladder (TB6C) and toad kidney (A6). Aldosterone, but not insulin, increases short-circuit current (I/sub sc/) in TB6C cells. Aldosterone increases Na -K -(TSP)ATPase activity after 18 h of incubation, but no effect can be seen at 3 and 6 h. Amiloride, which inhibits aldosterone-induced increases in I/sub sc/, has no effect on either basal or aldosterone stimulated enzyme activity. Both aldosterone and insulin increase I/sub sc/ in A6 cells and when added together are synergistic. Aldosterone stimulates enzyme activity in A6 cells, but insulin alone has no effect. However, aldosterone and insulin together stimulate enzyme activity more than aldosterone alone. It appears that stimulation of Na -K -ATPase activity is involved in aldosterone action in both cell lines but does not appear to be due to increased Na entry, since enhanced enzyme activity is not inhibited by amiloride. In contrast, insulin alone has no direct effect on Na -K -ATPase, although the increased enzyme activity following both agents in combination may explain their synergism on I/sub sc/.

  8. Ibuprofen regulation of microtubule dynamics in cystic fibrosis epithelial cells.

    Science.gov (United States)

    Rymut, Sharon M; Kampman, Claire M; Corey, Deborah A; Endres, Tori; Cotton, Calvin U; Kelley, Thomas J

    2016-08-01

    High-dose ibuprofen, an effective anti-inflammatory therapy for the treatment of cystic fibrosis (CF), has been shown to preserve lung function in a pediatric population. Despite its efficacy, few patients receive ibuprofen treatment due to potential renal and gastrointestinal toxicity. The mechanism of ibuprofen efficacy is also unclear. We have previously demonstrated that CF microtubules are slower to reform after depolymerization compared with respective wild-type controls. Slower microtubule dynamics in CF cells are responsible for impaired intracellular transport and are related to inflammatory signaling. Here, it is identified that high-dose ibuprofen treatment in both CF cell models and primary CF nasal epithelial cells restores microtubule reformation rates to wild-type levels, as well as induce extension of microtubules to the cell periphery. Ibuprofen treatment also restores microtubule-dependent intracellular transport monitored by measuring intracellular cholesterol transport. These effects are specific to ibuprofen as other cyclooxygenase inhibitors have no effect on these measures. Effects of ibuprofen are mimicked by stimulation of AMPK and blocked by the AMPK inhibitor compound C. We conclude that high-dose ibuprofen treatment enhances microtubule formation in CF cells likely through an AMPK-related pathway. These findings define a potential mechanism to explain the efficacy of ibuprofen therapy in CF. Copyright © 2016 the American Physiological Society.

  9. Epigenetic regulation of CpG promoter methylation in invasive prostate cancer cells

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    Farrar William L

    2010-10-01

    Full Text Available Abstract Background Recently, much attention has been focused on gaining a better understanding of the different populations of cells within a tumor and their contribution to cancer progression. One of the most commonly used methods to isolate a more aggressive sub-population of cells utilizes cell sorting based on expression of certain cell adhesion molecules. A recently established method we developed is to isolate these more aggressive cells based on their properties of increased invasive ability. These more invasive cells have been previously characterized as tumor initiating cells (TICs that have a stem-like genomic signature and express a number of stem cell genes including Oct3/4 and Nanog and are more tumorigenic compared to their 'non-invasive' counterpart. They also have a profile reminiscent of cells undergoing a classic pattern of epithelial to mesenchymal transition or EMT. Using this model of invasion, we sought to investigate which genes are under epigenetic control in this rare population of cells. Epigenetic modifications, specifically DNA methylation, are key events regulating the process of normal human development. To determine the specific methylation pattern in these invasive prostate cells, and if any developmental genes were being differentially regulated, we analyzed differences in global CpG promoter methylation. Results Differentially methylated genes were determined and select genes were chosen for additional analyses. The non-receptor tyrosine kinase BMX and transcription factor SOX1 were found to play a significant role in invasion. Ingenuity pathway analysis revealed the methylated gene list frequently displayed genes from the IL-6/STAT3 pathway. Cells which have decreased levels of the targets BMX and SOX1 also display loss of STAT3 activity. Finally, using Oncomine, it was determined that more aggressive metastatic prostate cancers in humans also have higher levels of both Stat3 and Sox1. Conclusions Using this

  10. Lack of relationship between TIMP-1 tumour cell immunoreactivity, treatment efficacy and prognosis in patients with advanced epithelial ovarian cancer

    DEFF Research Database (Denmark)

    Steffensen, Karina Dahl; Waldstrøm, Marianne; Christensen, Rikke Kølby

    2010-01-01

    BACKGROUND: Tissue inhibitor of metalloproteinase 1 (TIMP-1) is a natural inhibitor of the matrix metalloproteinases (MMPs) which are proteolytic enzymes involved in degradation of extracellular matrix thereby favoring tumour cell invasion and metastasis. TIMP-1 activity in tumour tissue may ther...... immunoreactivity in tumour tissue from patients with primary epithelial ovarian cancer did not correlate with patient survival or response to combination platinum/cyclophosphamide therapy.......BACKGROUND: Tissue inhibitor of metalloproteinase 1 (TIMP-1) is a natural inhibitor of the matrix metalloproteinases (MMPs) which are proteolytic enzymes involved in degradation of extracellular matrix thereby favoring tumour cell invasion and metastasis. TIMP-1 activity in tumour tissue may...... therefore play an essential role in the progression of a malignant tumour.The primary aim of the present study was to evaluate TIMP-1 protein immunoreactivity in tissue from primary ovarian cancer patients and associate these findings with the course of the disease including response to treatment...

  11. Role of Corneal Stromal Cells on Epithelial Cell Function during Wound Healing

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    Bhavani S. Kowtharapu

    2018-02-01

    Full Text Available Following injury, corneal stromal keratocytes transform into repair-phenotype of activated stromal fibroblasts (SFs and participate in wound repair. Simultaneously, ongoing bi-directional communications between corneal stromal-epithelial cells also play a vital role in mediating the process of wound healing. Factors produced by stromal cells are known to induce proliferation, differentiation, and motility of corneal epithelial cells, which are also subsequently the main processes that occur during wound healing. In this context, the present study aims to investigate the effect of SFs conditioned medium (SFCM on corneal epithelial cell function along with substance P (SP. Antibody microarrays were employed to profile differentially expressed cell surface markers and cytokines in the presence of SFCM and SP. Antibody microarray data revealed enhanced expression of the ITGB1 in corneal epithelial cells following stimulation with SP whereas SFCM induced abundant expression of IL-8, ITGB1, PD1L1, PECA1, IL-15, BDNF, ICAM1, CD8A, CD44 and NTF4. All these proteins have either direct or indirect roles in epithelial cell growth, movement and adhesion related signaling cascades during tissue regeneration. We also observed activation of MAPK signaling pathway along with increased expression of focal adhesion kinase (FAK, paxillin, vimentin, β-catenin and vasodilator-stimulated phosphoprotein (VASP phosphorylation. Additionally, epithelial-to-mesenchymal transition (EMT regulating transcription factors Slug and ZEB1 expression were enhanced in the presence of SFCM. SP enriched the expression of integrin subunits α4, α5, αV, β1 and β3 whereas SFCM increased α4, α5, αV, β1 and β5 integrin subunits. We also observed increased expression of Serpin E1 following SP and SFCM treatment. Wound healing scratch assay revealed enhanced migration of epithelial cells following the addition of SFCM. Taken together, we conclude that SFCM-mediated sustained

  12. Human Bronchial Epithelial Cell Response to Heavy Particle Exposure

    Science.gov (United States)

    Story, Michael; Ding, Liang-Hao; Minna, John; Park, Seong-mi; Peyton, Michael; Larsen, Jill

    2012-07-01

    A battery of non-oncogenically immortalized human bronchial epithelial cells (HBECs) are being used to examine the molecular changes that lead to lung carcinogenesis after exposure to heavy particles found in the free space environment. The goal is to ultimately identify biomarkers of radioresponse that can be used for prediction of carcinogenic risk for fatal lung cancer. Our initial studies have focused on the cell line HBEC3 KT and the isogenic variant HBEC3 KTR53, which overexpresses the RASv12 mutant and where p53 has been knocked down by shRNA, and is considered to be a more oncogenically progressed variant. We have previously described the response of HBEC3 KT at the cellular and molecular level, however, the focus here is on the rate of cellular transformation after HZE radiation exposure and the molecular changes in transformed cells. When comparing the two cell lines we find that there is a maximum rate of cellular transformation at 0.25 Gy when cells are exposed to 1 GeV Fe particles, and, for the HBEC3 KTR53 there are multiple pathways upregulated that promote anchorage independent growth including the mTOR pathway, the TGF-1 pathway, RhoA signaling and the ERK/MAPK pathway as early as 2 weeks after radiation. This does not occur in the HBEC3 KT cell line. Transformed HBEC3 KT cells do not show any morphologic or phenotypic changes when grown as cell cultures. HBEC3 KTR53 cells on the other hand show substantial changes in morphology from a cobblestone epithelial appearance to a mesenchymal appearance with a lack of contact inhibition. This epithelial to mesenchymal change in morphology is accompanied by the expression of vimentin and a reduction in the expression of E-cadherin, which are hallmarks of epithelial to mesenchymal transition. Interestingly, for HBEC3 KT transformed cells there are no mutations in the p53 gene, 2 of 15 clones were found to be heterozygous for the RASV12 mutation, and 3 of 15 clones expressed high levels of BigH3, a TGFB

  13. Collective cell streams in epithelial monolayers depend on cell adhesion

    International Nuclear Information System (INIS)

    Czirók, András; Varga, Katalin; Méhes, Előd; Szabó, András

    2013-01-01

    We report spontaneously emerging, randomly oriented, collective streaming behavior within a monolayer culture of a human keratinocyte cell line, and explore the effect of modulating cell adhesions by perturbing the function of calcium-dependent cell adhesion molecules. We demonstrate that decreasing cell adhesion induces narrower and more anisotropic cell streams, reminiscent of decreasing the Taylor scale of turbulent liquids. To explain our empirical findings, we propose a cell-based model that represents the dual nature of cell–cell adhesions. Spring-like connections provide mechanical stability, while a cellular Potts model formalism represents surface-tension driven attachment. By changing the relevance and persistence of mechanical links between cells, we are able to explain the experimentally observed changes in emergent flow patterns. (paper)

  14. Gene expression in epithelial cells in response to pneumovirus infection

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    Rosenberg Helene F

    2001-05-01

    Full Text Available Abstract Respiratory syncytial virus (RSV and pneumonia virus of mice (PVM are viruses of the family Paramyxoviridae, subfamily pneumovirus, which cause clinically important respiratory infections in humans and rodents, respectively. The respiratory epithelial target cells respond to viral infection with specific alterations in gene expression, including production of chemoattractant cytokines, adhesion molecules, elements that are related to the apoptosis response, and others that remain incompletely understood. Here we review our current understanding of these mucosal responses and discuss several genomic approaches, including differential display reverse transcription-polymerase chain reaction (PCR and gene array strategies, that will permit us to unravel the nature of these responses in a more complete and systematic manner.

  15. Expression of ICAM-1 in colon epithelial cells

    DEFF Research Database (Denmark)

    Vainer, Ben; Sørensen, Susanne; Seidelin, Jakob

    2003-01-01

    Studies have suggested that in ulcerative colitis (UC), intercellular adhesion molecule-1 (IC