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Sample records for surrounding epithelial cells

  1. Isolation of a sesquiterpene synthase expressing in specialized epithelial cells surrounding the secretory cavities in rough lemon (Citrus jambhiri).

    Science.gov (United States)

    Uji, Yuya; Ozawa, Rika; Shishido, Hodaka; Taniguchi, Shiduku; Takabayashi, Junji; Akimitsu, Kazuya; Gomi, Kenji

    2015-05-15

    Volatile terpenoids such as monoterpenes and sesquiterpenes play multiple roles in plant responses and are synthesized by terpene synthases (TPSs). We have previously isolated a partial TPS gene, RlemTPS4, that responds to microbial attack in rough lemon. In this study, we isolated a full length RlemTPS4 cDNA from rough lemon. RlemTPS4 localized in the cytosol. The recombinant RlemTPS4 protein was obtained using a prokaryotic expression system and GC-MS analysis of the terpenes produced by the RlemTPS4 enzymatic reaction determined that RlemTPS4 produces some sesquiterpenes such as δ-elemene. The RlemTPS4 gene was specifically expressed in specialized epithelial cells surrounding the oil secretory cavities in rough lemon leaf tissue.

  2. 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,

  3. Epithelial cell-cell junctions and plasma membrane domains

    NARCIS (Netherlands)

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

    2009-01-01

    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, comp

  4. Mammary epithelial cell

    DEFF Research Database (Denmark)

    Kass, Laura; Erler, Janine Terra; Dembo, Micah

    2007-01-01

    a repertoire of transmembrane receptors, of which integrins are the best characterized. Integrins modulate cell fate by reciprocally transducing biochemical and biophysical cues between the cell and the extracellular matrix, facilitating processes such as embryonic branching morphogenesis and lactation...... in the mammary gland. During breast development and cancer progression, the extracellular matrix is dynamically altered such that its composition, turnover, processing and orientation change dramatically. These modifications influence mammary epithelial cell shape, and modulate growth factor and hormonal...

  5. Coronaviruses in polarized epithelial cells

    NARCIS (Netherlands)

    Rossen, J W; Bekker, C P; Voorhout, W F; Horzinek, M C; Van der Ende, A; Strous, G J; Rottier, P J

    1995-01-01

    Coronaviruses have a marked tropism for epithelial cells. In this paper the interactions of the porcine transmissible gastroenteritis virus (TGEV) and mouse hepatitis virus (MHV-A59) with epithelial cells are compared. Porcine (LLC-PK1) and murine (mTAL) epithelial cells were grown on permeable supp

  6. Integrins and epithelial cell polarity.

    Science.gov (United States)

    Lee, Jessica L; Streuli, Charles H

    2014-08-01

    Cell polarity is characterised by differences in structure, composition and function between at least two poles of a cell. In epithelial cells, these spatial differences allow for the formation of defined apical and basal membranes. It has been increasingly recognised that cell-matrix interactions and integrins play an essential role in creating epithelial cell polarity, although key gaps in our knowledge remain. This Commentary will discuss the mounting evidence for the role of integrins in polarising epithelial cells. We build a model in which both inside-out signals to polarise basement membrane assembly at the basal surface, and outside-in signals to control microtubule apical-basal orientation and vesicular trafficking are required for establishing and maintaining the orientation of epithelial cell polarity. Finally, we discuss the relevance of the basal integrin polarity axis to cancer. This article is part of a Minifocus on Establishing polarity.

  7. 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.

  8. Airway epithelial cell responses to ozone injury

    Energy Technology Data Exchange (ETDEWEB)

    Leikauf, G.D.; Simpson, L.G.; Zhao, Qiyu [Univ. of Cincinnati Medical Center, OH (United States)] [and others

    1995-03-01

    The airway epithelial cell is an important target in ozone injury. Once activated, the airway epithelium responds in three phases. The initial, or immediate phase, involves activation of constitutive cells, often through direct covalent interactions including the formation of secondary ozonolysis products-hydroxyhydroperoxides, aldehydes, and hydrogen peroxide. Recently, we found hydroxyhydroperoxides to be potent agonists; of bioactive eicosanoid formation by human airway epithelial cells in culture. Other probable immediate events include activation and inactivation of enzymes present on the epithelial surface (e.g., neutral endopeptidase). During the next 2 to 24 hr, or early phase, epithelial cells respond by synthesis and release of chemotactic factors, including chemokines-macrophage inflammatory protein-2, RANTES, and interleukin-8. Infiltrating leukocytes during this period also release elastase, an important agonist of epithelial cell mucus secretion and additional chemokine formation. The third (late) phase of ozone injury is characterized by eosinophil or monocyte infiltration. Cytokine expression leads to alteration of structural protein synthesis, with increases in fibronectin evident by in situ hybridization. Synthesis of epithelial antiproteases, e.g., secretary leukocyte protease inhibitor, may also increase locally 24 to 48 hr after elastase concentrations become excessive. Thus, the epithelium is not merely a passive barrier to ozone injury but has a dynamic role in directing the migration, activating, and then counteracting inflammatory cells. Through these complex interactions, epithelial cells can be viewed as the initiators (alpha) and the receptors (omega) of ozone-induced airway disease. 51 refs., 2 figs., 3 tabs.

  9. Polarized sorting and trafficking in epithelial cells

    Institute of Scientific and Technical Information of China (English)

    Xinwang Cao; Michal A Surma; Kai Simons

    2012-01-01

    The polarized distribution of proteins and lipids at the surface membrane of epithelial cells results in the formation of an apical and a basolateral domain,which are separated by tight junctions.The generation and maintenance of epithelial polarity require elaborate mechanisms that guarantee correct sorting and vectorial delivery of cargo molecules.This dynamic process involves the interaction of sorting signals with sorting machineries and the formation of transport carriers.Here we review the recent advances in the field of polarized sorting in epithelial cells.We especially highlight the role of lipid rafts in apical sorting.

  10. 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

  11. 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

  12. 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 concluded that the epithelial stromal cells of the thymus, by acting as veto cells, may be responsible for the negative intrathymic selection of self-reactive thymocytes leading to elimination of the vast majority of immature thymic lymphocytes....

  13. 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 concluded that the epithelial stromal cells of the thymus, by acting as veto cells, may be responsible for the negative intrathymic selection of self-reactive thymocytes leading to elimination of the vast majority of immature thymic lymphocytes....

  14. Wound healing of intestinal epithelial cells

    Institute of Scientific and Technical Information of China (English)

    Masahiro Iizuka; Shiho Konno

    2011-01-01

    The intestinal epithelial cells (IECs) form a selective permeability barrier separating luminal content from underlying tissues. Upon injury, the intestinal epithelium undergoes a wound healing process. Intestinal wound healing is dependent on the balance of three cellular events;restitution, proliferation, and differentiation of epithelial cells adjacent to the wounded area. Previous studies have shown that various regulatory peptides, including growth factors and cytokines, modulate intestinal epithelial wound healing. Recent studies have revealed that novel factors, which include toll-like receptors (TLRs), regulatory peptides, particular dietary factors, and some gastroprotective agents, also modulate intestinal epithelial wound repair. Among these factors, the activation of TLRs by commensal bacteria is suggested to play an essential role in the maintenance of gut homeostasis. Recent studies suggest that mutations and dysregulation of TLRs could be major contributing factors in the predisposition and perpetuation of inflammatory bowel disease. Additionally, studies have shown that specific signaling pathways are involved in IEC wound repair. In this review, we summarize the function of IECs, the process of intestinal epithelial wound healing, and the functions and mechanisms of the various factors that contribute to gut homeostasis and intestinal epithelial wound healing.

  15. Epithelial cell-extracellular matrix interactions and stem cells in airway epithelial regeneration.

    Science.gov (United States)

    Coraux, Christelle; Roux, Jacqueline; Jolly, Thomas; Birembaut, Philippe

    2008-08-15

    In healthy subjects, the respiratory epithelium forms a continuous lining to the airways and to the environment, and plays a unique role as a barrier against external deleterious agents to protect the airways from the insults. In respiratory diseases such as cystic fibrosis (CF), chronic obstructive pulmonary disease (COPD), chronic bronchitis, or asthma, the airway epithelium is frequently remodeled and injured, leading to the impairment of its defense functions. The rapid restoration of the epithelial barrier is crucial for these patients. The complete regeneration of the airway epithelium is a complex phenomenon, including not only the epithelial wound repair but also the epithelial differentiation to reconstitute a fully well differentiated and functional epithelium. The regeneration implies two partners: the epithelial stem/progenitor cells and factors able to regulate this process. Among these factors, epithelial cells-extracellular matrix (ECM) interactions play a crucial role. The secretion of a provisional ECM, the cell-ECM relationships through epithelial receptors, and the remodeling of the ECM by proteases (mainly matrix metalloproteinases) contribute not only to airway epithelial repair by modulating epithelial cell migration and proliferation, but also to the differentiation of repairing cells leading to the complete restoration of the wounded epithelium. A better characterization of resident stem cells and of effectors of the regeneration process is an essential prerequisite to propose new regenerative therapeutics to patients suffering from infectious/inflammatory respiratory diseases.

  16. Host epithelial geometry regulates breast cancer cell invasiveness

    Science.gov (United States)

    Boghaert, Eline; Gleghorn, Jason P.; Lee, KangAe; Gjorevski, Nikolce; Radisky, Derek C.; Nelson, Celeste M.

    2012-01-01

    Breast tumor development is regulated in part by cues from the local microenvironment, including interactions with neighboring nontumor cells as well as the ECM. Studies using homogeneous populations of breast cancer cell lines cultured in 3D ECM have shown that increased ECM stiffness stimulates tumor cell invasion. However, at early stages of breast cancer development, malignant cells are surrounded by normal epithelial cells, which have been shown to exert a tumor-suppressive effect on cocultured cancer cells. Here we explored how the biophysical characteristics of the host microenvironment affect the proliferative and invasive tumor phenotype of the earliest stages of tumor development, by using a 3D microfabrication-based approach to engineer ducts composed of normal mammary epithelial cells that contained a single tumor cell. We found that the phenotype of the tumor cell was dictated by its position in the duct: proliferation and invasion were enhanced at the ends and blocked when the tumor cell was located elsewhere within the tissue. Regions of invasion correlated with high endogenous mechanical stress, as shown by finite element modeling and bead displacement experiments, and modulating the contractility of the host epithelium controlled the subsequent invasion of tumor cells. Combining microcomputed tomographic analysis with finite element modeling suggested that predicted regions of high mechanical stress correspond to regions of tumor formation in vivo. This work suggests that the mechanical tone of nontumorigenic host epithelium directs the phenotype of tumor cells and provides additional insight into the instructive role of the mechanical tumor microenvironment. PMID:23150585

  17. Epithelial Cell Apoptosis and Lung Remodeling

    Institute of Scientific and Technical Information of China (English)

    Kazuyoshi Kuwano

    2007-01-01

    Lung epithelium is the primary site of lung damage in various lung diseases. Epithelial cell apoptosis has been considered to be initial event in various lung diseases. Apoptosis signaling is classically composed of two principle pathways. One is a direct pathway from death receptor ligation to caspase cascade activation and cell death. The other pathway triggered by stresses such as drugs, radiation, infectious agents and reactive oxygen species is mediated by mitochondria. Endoplasmic reticulum has also been shown to be the organelle to mediate apoptosis.Epithelial cell death is followed by remodeling processes, which consist of epithelial and fibroblast activation,cytokine production, activation of coagulation pathway, neoangiogenesis, re-epithelialization and fibrosis.Epithelial and mesenchymal interaction plays important roles in these processes. Further understanding of apoptosis signaling and its regulation by novel strategies may lead to effective treatments against various lung diseases. We review the recent advances in the understanding of apoptosis signaling and discuss the involvement of apoptosis in lung remodeling.

  18. 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 glycosphingolipid, together with an increase in plasmalogen, phosphatidylethanolamine, and cholesterol content, whereas the opposite changes took place during an epithelial-to-mesenchymal transition. Moreover, during polarization, the sphingolipids became longer, more saturated, and more hydroxylated as required...

  19. Lipid polarity and sorting in epithelial cells

    NARCIS (Netherlands)

    van Meer, G.; Simons, K.

    1988-01-01

    Apical and basolateral membrane domains of epithelial cell plasma membranes possess unique lipid compositions. The tight junction, the structure separating the two domains, forms a diffusion barrier for membrane components and thereby prevents intermixing of the two sets of lipids. The barrier appar

  20. Protons Sensitize Epithelial Cells to Mesenchymal Transition

    Science.gov (United States)

    Wang, Minli; Hada, Megumi; Saha, Janapriya; Sridharan, Deepa M.; Pluth, Janice M.; Cucinotta, Francis A.

    2012-01-01

    Proton radiotherapy has gained more favor among oncologists as a treatment option for localized and deep-seated tumors. In addition, protons are a major constituent of the space radiation astronauts receive during space flights. The potential for these exposures to lead to, or enhance cancer risk has not been well studied. Our objective is to study the biological effects of low energy protons on epithelial cells and its propensity to enhance transforming growth factor beta 1 (TGFβ1)-mediated epithelial-mesenchymal transition (EMT), a process occurring during tumor progression and critical for invasion and metastasis. Non-transformed mink lung epithelial cells (Mv1Lu) and hTERT- immortalized human esophageal epithelial cells (EPC) were used in this study. EMT was identified by alterations in cell morphology, EMT-related gene expression changes determined using real-time PCR, and EMT changes in specific cellular markers detected by immunostaining and western blotting. Although TGFβ1 treatment alone is able to induce EMT in both Mv1Lu and EPC cells, low energy protons (5 MeV) at doses as low as 0.1 Gy can enhance TGFβ1 induced EMT. Protons alone can also induce a mild induction of EMT. SD208, a potent TGFβ Receptor 1 (TGFβR1) kinase inhibitor, can efficiently block TGFβ1/Smad signaling and attenuate EMT induction. We suggest a model for EMT after proton irradiation in normal and cancerous tissue based on our results that showed that low and high doses of protons can sensitize normal human epithelial cells to mesenchymal transition, more prominently in the presence of TGFβ1, but also in the absence of TGFβ1. PMID:22844446

  1. Protons sensitize epithelial cells to mesenchymal transition.

    Directory of Open Access Journals (Sweden)

    Minli Wang

    Full Text Available Proton radiotherapy has gained more favor among oncologists as a treatment option for localized and deep-seated tumors. In addition, protons are a major constituent of the space radiation astronauts receive during space flights. The potential for these exposures to lead to, or enhance cancer risk has not been well studied. Our objective is to study the biological effects of low energy protons on epithelial cells and its propensity to enhance transforming growth factor beta 1 (TGFβ1-mediated epithelial-mesenchymal transition (EMT, a process occurring during tumor progression and critical for invasion and metastasis. Non-transformed mink lung epithelial cells (Mv1Lu and hTERT- immortalized human esophageal epithelial cells (EPC were used in this study. EMT was identified by alterations in cell morphology, EMT-related gene expression changes determined using real-time PCR, and EMT changes in specific cellular markers detected by immunostaining and western blotting. Although TGFβ1 treatment alone is able to induce EMT in both Mv1Lu and EPC cells, low energy protons (5 MeV at doses as low as 0.1 Gy can enhance TGFβ1 induced EMT. Protons alone can also induce a mild induction of EMT. SD208, a potent TGFβ Receptor 1 (TGFβR1 kinase inhibitor, can efficiently block TGFβ1/Smad signaling and attenuate EMT induction. We suggest a model for EMT after proton irradiation in normal and cancerous tissue based on our results that showed that low and high doses of protons can sensitize normal human epithelial cells to mesenchymal transition, more prominently in the presence of TGFβ1, but also in the absence of TGFβ1.

  2. Airway epithelial cells: current concepts and challenges.

    Science.gov (United States)

    Crystal, Ronald G; Randell, Scott H; Engelhardt, John F; Voynow, Judith; Sunday, Mary E

    2008-09-15

    The adult human bronchial tree is covered with a continuous layer of epithelial cells that play a critical role in maintaining the conduit for air, and which are central to the defenses of the lung against inhaled environmental concomitants. The epithelial sheet functions as an interdependent unit with the other lung components. Importantly, the structure and/or function of airway epithelium is deranged in major lung disorders, including chronic obstructive pulmonary disease, asthma, and bronchogenic carcinoma. Investigations regarding the airway epithelium have led to many advances over the past few decades, but new developments in genetics and stem cell/progenitor cell biology have opened the door to understanding how the airway epithelium is developed and maintained, and how it responds to environmental stress. This article provides an overview of the current state of knowledge regarding airway epithelial stem/progenitor cells, gene expression, cell-cell interactions, and less frequent cell types, and discusses the challenges for future areas of investigation regarding the airway epithelium in health and disease.

  3. Isolation of epithelial cells with hepatobiliary phenotype.

    Science.gov (United States)

    Castorina, Sergio; Luca, Tonia; Torrisi, Antonella; Privitera, Giovanna; Panebianco, Mariangela

    2008-01-01

    The regenerative capacity of the liver after partial hepatectomy or chemical injury is well known. In human liver, the resident progenitor cells are called "hepatic progenitor cells" (HPCs) while the term "oval cells" should be discouraged in order to indicate the stem cell compartment. The aim of our study was first to analyse the cellular aspects of liver regeneration through differentiation in cholangiocytes and hepatocytes, and then to characterise resident progenitor cells, using "primary cultured hepatocytes" derived from healthy adult human livers. Human hepatocytes were isolated from fresh surgical specimens of patients who underwent hepatic resections in our Clinical Centre surgery operating room. Hepatic differentiation and function were analysed by immunocytochemistry techniques and the presence of liver epithelial cell populations within normal adult human liver, was demonstrated by immunohistochemistry analysis. These cells expanded in vitro and showed the capacity for self-renewal and multipotent differentiation. Human liver stem cells expressed several mesenchymal markers, such as CD44, but not haematopoietic stem cell markers. In addition, these cells expressed alpha-fetoprotein, albumin, CK7 and CK19, indicating a partial commitment to hepatic and biliary cells. Interestingly the expression of both hepatocytes and biliary markers in HPCs reflects the bipotential nature of the hepatic stem cells toward both the hepatic and biliary lineage. According to their immature and bipotential phenotype, hepatic epithelial cells might represent a pool of precursors in the healthy human adult liver.

  4. The human airway epithelial basal cell transcriptome.

    Directory of Open Access Journals (Sweden)

    Neil R Hackett

    Full Text Available BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSION/SIGNIFICANCE: The human airway epithelial basal cell signature identified in the present study provides novel insights into the molecular phenotype and biology of

  5. Reversible transdifferentiation of alveolar epithelial cells.

    Science.gov (United States)

    Danto, S I; Shannon, J M; Borok, Z; Zabski, S M; Crandall, E D

    1995-05-01

    Alveolar epithelial type II (AT2) cells have been thought to be the progenitors of terminally differentiated type I (AT1) cells in the adult animal in vivo. In this study, we used an AT1 cell-specific monoclonal antibody (mAb VIII B2) to investigate expression of the AT1 cell phenotype accompanying reversible changes in expression of the AT2 cell phenotype. AT2 cells were isolated and cultured either on attached collagen gels or on gels detached 1 or 4 days after plating and maintained thereafter as floating gels. Monolayers on both attached and floating gels were harvested on days 4 and 8 and analyzed by electron microscopy for changes in morphology and binding of mAb VIII B2. Results indicate that: (1) alveolar epithelial cells (AEC) on attached gels develop characteristics of the AT1 cell phenotype, (2) AEC on gels detached on day 1 maintain features of the AT2 cell phenotype (and do not react with mAb VIII B2), and (3) the expression of AT1 cell phenotypic traits seen by day 4 on attached gels is reversed after detachment. We conclude that commitment to the AT1 and AT2 cell lineages requires continuous regulatory input to maintain the differentiated states, and that transdifferentiation between AT2 and AT1 cells may be reversible.

  6. 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.

  7. A method for establishing human primary gastric epithelial cell culture from fresh surgical gastric tissues.

    Science.gov (United States)

    Aziz, Faisal; Yang, Xuesong; Wen, Qingping; Yan, Qiu

    2015-08-01

    At present, biopsy specimens, cancer cell lines and tissues obtained by gastric surgery are used in the study and analysis of gastric cancer, including the molecular mechanisms and proteomics. However, fibroblasts and other tissue components may interfere with these techniques. Therefore, the present study aimed to develop a procedure for the isolation of viable human gastric epithelial cells from gastric surgical tissues. A method was developed to culture human gastric epithelial cells using fresh, surgically excised tissues and was evaluated using immunocytochemistry, periodic acid-Schiff (PAS) staining and cell viability assays. Low cell growth was observed surrounding the gastric tissue on the seventh day of tissue explant culture. Cell growth subsequently increased, and at 12 days post-explant a high number of pure epithelial cells were detected. The gastric cancer cells exhibited rapid growth with a doubling time of 13-52 h, as compared to normal cells, which had a doubling time of 20-53 h. Immunocytochemical analyses of primary gastric cells revealed positive staining for cytokeratin 18 and 19, which indicated that the culture was comprised of pure epithelial cells and contained no fibroblasts. Furthermore, PAS staining demonstrated that the cultured gastric cells produced neutral mucin. Granulin and carbohydrate antigen 724 staining confirmed the purity of gastric cancer and normal cells in culture. This method of cell culture indicated that the gastric cells in primary culture consisted of mucin-secreting gastric epithelial cells, which may be useful for the study of gastric infection with Helicobacter pylori and gastric cancer.

  8. Neutrophil-induced injury of rat pulmonary alveolar epithelial cells.

    Science.gov (United States)

    Simon, R H; DeHart, P D; Todd, R F

    1986-11-01

    The damage to pulmonary alveolar epithelial cells that occurs in many inflammatory conditions is thought to be caused in part by phagocytic neutrophils. To investigate this process, we exposed monolayers of purified rat alveolar epithelial cells to stimulated human neutrophils and measured cytotoxicity using a 51Cr-release assay. We found that stimulated neutrophils killed epithelial cells by a process that did not require neutrophil-generated reactive oxygen metabolites. Pretreatment of neutrophils with an antibody (anti-Mo1) that reduced neutrophil adherence to epithelial cells limited killing. Although a variety of serine protease inhibitors partially inhibited cytotoxicity, we found that neutrophil cytoplasts, neutrophil lysates, neutrophil-conditioned medium, purified azurophilic or specific granule contents, and purified human neutrophil elastase did not duplicate the injury. We conclude that stimulated neutrophils can kill alveolar epithelial cells in an oxygen metabolite-independent manner. Tight adherence of stimulated neutrophils to epithelial cell monolayers appears to promote epithelial cell killing.

  9. Intestinal epithelial cells in inflammatory bowel diseases

    Institute of Scientific and Technical Information of China (English)

    Giulia; Roda; Alessandro; Sartini; Elisabetta; Zambon; Andrea; Calafiore; Margherita; Marocchi; Alessandra; Caponi; Andrea; Belluzzi; Enrico; Roda

    2010-01-01

    The pathogenesis of inflammatory bowel diseases (IBDs) seems to involve a primary defect in one or more of the elements responsible for the maintenance of intestinal homeostasis and oral tolerance. The most important element is represented by the intestinal barrier, a complex system formed mostly by intestinal epithelial cells (IECs). IECs have an active role in producing mucus and regulating its composition; they provide a physical barrier capable of controlling antigen traff ic through the intestinal muco...

  10. Limbal stem cells: Central concepts of corneal epithelial homeostasis

    Institute of Scientific and Technical Information of China (English)

    Jinny; J; Yoon; Salim; Ismail; Trevor; Sherwin

    2014-01-01

    A strong cohort of evidence exists that supports the localisation of corneal stem cells at the limbus. The distinguishing characteristics of limbal cells as stem cells include slow cycling properties, high proliferative potential when required, clonogenicity, absence of differentiation marker expression coupled with positive expression of progenitor markers, multipotency, centripetal migration, requirement for a distinct niche environment and the ability of transplanted limbal cells to regenerate the entire corneal epithelium. The existence of limbal stem cells supports the prevailing theory of corneal homeostasis, known as the XYZ hypothesis where X represents proliferation and stratification of limbal basal cells, Y centripetal migration of basal cells and Z desquamation of superficial cells. To maintain the mass of cornea, the sum of X and Y must equal Z and very elegant cell tracking experiments provide strong evidence in support of this theory. However, several recent stud-ies have suggested the existence of oligopotent stem cells capable of corneal maintenance outside of the limbus. This review presents a summary of data which led to the current concepts of corneal epithelial homeostasis and discusses areas of controversy surrounding the existence of a secondary stem cell reservoir on the corneal surface

  11. Characterization of Human Mammary Epithelial Stem Cells

    Science.gov (United States)

    2010-10-01

    breast is highly expressed by luminal epithelial cells and is less expressed by basal cells19,20. In contrast, CD49f (a6 integrin) has an inverse pattern...mouse stretched on its back. The hose and nose cone from the anesthetic vaporizer are securely attached to one side of the plate, and a heated pad is...the mouse by a nose cone. Check that the mouse has reached surgical anesthesia by loss of pedal withdrawal reflex . ! cautIon Institutional review

  12. Cell competition with normal epithelial cells promotes apical extrusion of transformed cells through metabolic changes.

    Science.gov (United States)

    Kon, Shunsuke; Ishibashi, Kojiro; Katoh, Hiroto; Kitamoto, Sho; Shirai, Takanobu; Tanaka, Shinya; Kajita, Mihoko; Ishikawa, Susumu; Yamauchi, Hajime; Yako, Yuta; Kamasaki, Tomoko; Matsumoto, Tomohiro; Watanabe, Hirotaka; Egami, Riku; Sasaki, Ayana; Nishikawa, Atsuko; Kameda, Ikumi; Maruyama, Takeshi; Narumi, Rika; Morita, Tomoko; Sasaki, Yoshiteru; Enoki, Ryosuke; Honma, Sato; Imamura, Hiromi; Oshima, Masanobu; Soga, Tomoyoshi; Miyazaki, Jun-Ichi; Duchen, Michael R; Nam, Jin-Min; Onodera, Yasuhito; Yoshioka, Shingo; Kikuta, Junichi; Ishii, Masaru; Imajo, Masamichi; Nishida, Eisuke; Fujioka, Yoichiro; Ohba, Yusuke; Sato, Toshiro; Fujita, Yasuyuki

    2017-05-01

    Recent studies have revealed that newly emerging transformed cells are often apically extruded from epithelial tissues. During this process, normal epithelial cells can recognize and actively eliminate transformed cells, a process called epithelial defence against cancer (EDAC). Here, we show that mitochondrial membrane potential is diminished in RasV12-transformed cells when they are surrounded by normal cells. In addition, glucose uptake is elevated, leading to higher lactate production. The mitochondrial dysfunction is driven by upregulation of pyruvate dehydrogenase kinase 4 (PDK4), which positively regulates elimination of RasV12-transformed cells. Furthermore, EDAC from the surrounding normal cells, involving filamin, drives the Warburg-effect-like metabolic alteration. Moreover, using a cell-competition mouse model, we demonstrate that PDK-mediated metabolic changes promote the elimination of RasV12-transformed cells from intestinal epithelia. These data indicate that non-cell-autonomous metabolic modulation is a crucial regulator for cell competition, shedding light on the unexplored events at the initial stage of carcinogenesis.

  13. 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.

  14. Single cell migration in oral squamous cell carcinoma - possible evidence of epithelial-mesenchymal transition in vivo

    DEFF Research Database (Denmark)

    Jensen, David H; Reibel, Jesper; Mackenzie, Ian C

    2015-01-01

    BACKGROUND: The invasion of cancer cells into the surrounding normal tissue is one of the defining features of cancer. While the phenomena of tumour budding, epithelial-mesenchymal transition and the presence of myofibroblasts have independently been shown to be related to a poor prognosis of ora...

  15. Growth of cultured porcine retinal pigment epithelial cells

    DEFF Research Database (Denmark)

    Wiencke, A.K.; Kiilgaard, Jens Folke; Nicolini, Jair;

    2003-01-01

    To establish and characterize cultures of porcine retinal pigment epithelial (pRPE) cells in order to produce confluent monolayers of cells for transplantation.......To establish and characterize cultures of porcine retinal pigment epithelial (pRPE) cells in order to produce confluent monolayers of cells for transplantation....

  16. Cell reintegration: Stray epithelial cells make their way home.

    Science.gov (United States)

    Wilson, Tyler J; Bergstralh, Dan T

    2017-06-01

    Ongoing work shows that misplaced epithelial cells have the capacity to reintegrate back into tissue layers. This movement appears to underlie tissue stability and may also control aspects of tissue structure. A recent study reveals that cell reintegration in at least one tissue, the Drosophila follicular epithelium, is based on adhesion molecules that line lateral cell surfaces. In this article we will review these observations, discuss their implications for epithelial tissue development and maintenance, and identify future directions for study. © 2017 WILEY Periodicals, Inc.

  17. Cells of Origin of Epithelial Ovarian Cancers

    Science.gov (United States)

    2015-09-01

    AWARD NUMBER: W81XWH-14-1-0280 TITLE: Cells of Origin of Epithelial Ovarian Cancers PRINCIPAL INVESTIGATOR: Zhe Li, PhD CONTRACTING...Xie, Zhe Li 5d. PROJECT NUMBER 5e. TASK NUMBER E-Mail: zli4@rics.bwh.harvard.edu 5f. WORK UNIT NUMBER 7. PERFORMING ORGANIZATION NAME(S) AND...Lined Inclusion Cysts or Teratomas. PLoS ONE 8, e65067. Sherman-Baust, C.A., Kuhn, E., Valle, B.L., Shih Ie, M., Kurman, R.J., Wang , T.L., Amano, T

  18. Neutrophil-induced injury of rat pulmonary alveolar epithelial cells.

    OpenAIRE

    Simon, R H; DeHart, P D; Todd, R F

    1986-01-01

    The damage to pulmonary alveolar epithelial cells that occurs in many inflammatory conditions is thought to be caused in part by phagocytic neutrophils. To investigate this process, we exposed monolayers of purified rat alveolar epithelial cells to stimulated human neutrophils and measured cytotoxicity using a 51Cr-release assay. We found that stimulated neutrophils killed epithelial cells by a process that did not require neutrophil-generated reactive oxygen metabolites. Pretreatment of neut...

  19. Characterization of protocadherin-1 expression in primary bronchial epithelial cells : association with epithelial cell differentiation

    NARCIS (Netherlands)

    Koning, Henk; Sayers, Ian; Stewart, Ceri E.; de Jong, Debora; ten Hacken, Nick H. T.; Postma, Dirkje S.; van Oosterhout, Antoon J. M.; Nawijn, Martijn C.; Koppelman, Gerard H.

    2012-01-01

    Protocadherin-1 (PCDH1) is a novel susceptibility gene for asthma that is expressed in airway epithelium. We aimed to characterize PCDH1 mRNA transcripts and protein expression in primary bronchial epithelial cells and to determine regulation of PCDH1 during mucociliary differentiation. Total RNA an

  20. The plasticity of human breast carcinoma cells is more than epithelial to mesenchymal conversion

    Energy Technology Data Exchange (ETDEWEB)

    Petersen, Ole William; Nielsen, Helga Lind; Gudjonsson, Thorarinn; Villadsen, Ren& #233; ; Ronnov-Jessen, Lone; Bissell, Mina J.

    2001-05-12

    The human breast comprises three lineages: the luminal epithelial lineage, the myoepithelial lineage, and the mesenchymal lineage. It has been widely accepted that human breast neoplasia pertains only to the luminal epithelial lineage. In recent years, however, evidence has accumulated that neoplastic breast epithelial cells may be substantially more plastic in their differentiation repertoire than previously anticipated. Thus, along with an increasing availability of markers for the myoepithelial lineage, at least a partial differentiation towards this lineage is being revealed frequently. It has also become clear that conversions towards the mesenchymal lineage actually occur, referred to as epithelial to mesenchymal transitions. Indeed, some of the so-called myofibroblasts surrounding the tumor may indeed have an epithelial origin rather than a mesenchymal origin. Because myoepithelial cells, epithelial to mesenchymal transition-derived cells, genuine stromal cells and myofibroblasts share common markers, we now need to define a more ambitious set of markers to distinguish these cell types in the microenvironment of the tumors. This is necessary because the different microenvironments may confer different clinical outcomes. The aim of this commentary is to describe some of the inherent complexities in defining cellular phenotypes in the microenvironment of breast cancer and to expand wherever possible on the implications for tumor suppression and progression.

  1. 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.

  2. 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.

  3. 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.

  4. The plasticity of human breast carcinoma cells is more than epithelial to mesenchymal conversion

    DEFF Research Database (Denmark)

    Petersen, Ole William; Nielsen, Helga Lind; Gudjonsson, Thorarinn;

    2001-01-01

    The human breast comprises three lineages: the luminal epithelial lineage, the myoepithelial lineage, and the mesenchymal lineage. It has been widely accepted that human breast neoplasia pertains only to the luminal epithelial lineage. In recent years, however, evidence has accumulated that neopl......The human breast comprises three lineages: the luminal epithelial lineage, the myoepithelial lineage, and the mesenchymal lineage. It has been widely accepted that human breast neoplasia pertains only to the luminal epithelial lineage. In recent years, however, evidence has accumulated...... that neoplastic breast epithelial cells may be substantially more plastic in their differentiation repertoire than previously anticipated. Thus, along with an increasing availability of markers for the myoepithelial lineage, at least a partial differentiation towards this lineage is being revealed frequently....... It has also become clear that conversions towards the mesenchymal lineage actually occur, referred to as epithelial to mesenchymal transitions. Indeed, some of the so-called myofibroblasts surrounding the tumor may have an epithelial origin rather than a mesenchymal origin. Because myoepithelial cells...

  5. Silk film topography directs collective epithelial cell migration.

    Directory of Open Access Journals (Sweden)

    Brian D Lawrence

    Full Text Available 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.

  6. Uranium induces apoptosis in lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Periyakaruppan, Adaikkappan; Sarkar, Shubhashish; Sadanandan, Bindu; Thomas, Renard; Wilson, Bobby L. [Texas Southern University, Environmental Toxicology Program, Department of Chemistry, Houston, TX (United States); Ravichandran, Prabakaran; Sharma, Chidananda S.; Ramesh, Vani; Hall, Joseph C.; Ramesh, Govindarajan T. [Norfolk State University, Molecular Toxicology Laboratory, Department of Biology, Center for Biotechnology and Biomedical Sciences, Norfolk, VA (United States)

    2009-06-15

    Uranium is a naturally occurring radioactive material present everywhere in the environment. It is toxic because of its chemical or radioactive properties. Uranium enters environment mainly from mines and industry and cause threat to human health by accumulating in lungs as a result of inhalation. In our previous study, we have shown the effectiveness of antioxidant system response to the oxidative stress induced by uranyl acetate (UA) in rat lung epithelial (LE) cells. As part of our continuing studies; here, we investigated the mechanism underlying when LE cells are exposed to different concentration of UA. Oxidative stress may lead to apoptotic signaling pathways. LE cells treated with 0.25, 0.5 and 1 mM of UA results in dose and time-dependent increase in activity of both caspases-3 and -8. Increase in the concentration of cytochrome-c oxidase in cytosol was seen in LE cells treated with 1 mM UA as a result of mitochondria membrane permeability. The cytochrome-c leakage may trigger the apoptotic pathway. TUNEL assay performed in LE cells treated with 1 mM of UA showed significant incorporation of dNTPs in the nucleus after 24 h. In the presence of the caspase inhibitors, we observed the significant decrease in the activity of caspases-8 and -3 in 0.5 and 1 mM UA-treated LE cells. (orig.)

  7. Mechanical stretch triggers rapid epithelial cell division through Piezo1.

    Science.gov (United States)

    Gudipaty, S A; Lindblom, J; Loftus, P D; Redd, M J; Edes, K; Davey, C F; Krishnegowda, V; Rosenblatt, J

    2017-03-02

    Despite acting as a barrier for the organs they encase, epithelial cells turn over at some of the fastest rates in the body. However, epithelial cell division must be tightly linked to cell death to preserve barrier function and prevent tumour formation. How does the number of dying cells match those dividing to maintain constant numbers? When epithelial cells become too crowded, they activate the stretch-activated channel Piezo1 to trigger extrusion of cells that later die. However, it is unclear how epithelial cell division is controlled to balance cell death at the steady state. Here we show that mammalian epithelial cell division occurs in regions of low cell density where cells are stretched. By experimentally stretching epithelia, we find that mechanical stretch itself rapidly stimulates cell division through activation of the Piezo1 channel. To stimulate cell division, stretch triggers cells that are paused in early G2 phase to activate calcium-dependent phosphorylation of ERK1/2, thereby activating the cyclin B transcription that is necessary to drive cells into mitosis. Although both epithelial cell division and cell extrusion require Piezo1 at the steady state, the type of mechanical force controls the outcome: stretch induces cell division, whereas crowding induces extrusion. How Piezo1-dependent calcium transients activate two opposing processes may depend on where and how Piezo1 is activated, as it accumulates in different subcellular sites with increasing cell density. In sparse epithelial regions in which cells divide, Piezo1 localizes to the plasma membrane and cytoplasm, whereas in dense regions in which cells extrude, it forms large cytoplasmic aggregates. Because Piezo1 senses both mechanical crowding and stretch, it may act as a homeostatic sensor to control epithelial cell numbers, triggering extrusion and apoptosis in crowded regions and cell division in sparse regions.

  8. Progressive transformation of immortalized esophageal epithelial cells

    Institute of Scientific and Technical Information of China (English)

    Zhong-Ying Shen; Li-Yan Xu; Min-Hua Chen; Jian Shen; Wei-Jia Cai; Yi Zeng

    2002-01-01

    .CONCLUSION: In continual cultivation of fetal esophageal epithelial cells with transduction of HPV18E6E7, cells from the 10th to the 85th passage were changed gradually from preimmortal, immortal, precancerous to malignantly transformed stages. All of these changes were in a dynamic progressive process. The establishment of a continuous line of esophageal epithelium may provide a in vitro model of carcinogenesis induced by HPV.

  9. Epithelial cell identity in hyperplastic precursors of breast cancer

    Institute of Scientific and Technical Information of China (English)

    Danila Coradini; Patrizia Boracchi; Saro Oriana; Elia Biganzoli; Federico Ambrogi

    2015-01-01

    Introduction:In the adult human breast, hyperplastic enlarged lobular unit (HELU) and atypical ductal hyperplasia (ADH) are two common abnormalities that frequently coexist with ductal carcinoma in situ (DCIS). For this reason, they have been proposed as the early steps in a biological continuum toward breast cancer. Methods:We investigated in silico the expression of 369 genes experimentally recognized as involved in establishing and maintaining epithelial cell identity and mammary gland remodeling, in HELUs or ADHs with respect to the corresponding patient-matched normal tissue. Results:Despite the common luminal origin, HELUs and ADHs proved to be characterized by distinct gene profiles that overlap for 5 genes only. While HELUs were associated with the overexpression of progesterone receptor (PGR), ADHs were characterized by the overexpression of estrogen receptor 1 (ESR1) coupled with the overexpression of some proliferation-associated genes. Conclusions:This unexpected finding contradicts the notion that in differentiated luminal cells the expression of estrogen receptor (ER) is dissociated from cell proliferation and suggests that the establishing of an ER-dependent signaling is able to sustain cell proliferation in an autocrine manner as an early event in tumor initiation. Although clinical evidence indicates that only a fraction of HELUs and ADHs evolve to invasive cancer, present findings warn that exposure to synthetic progestins, frequently administered as hormone-replacement therapy, and estrogens, when abnormally produced by adipose cells and persistently present in the stroma surrounding the mammary gland, may cause these hyperplastic lesions.

  10. 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.

  11. Sonic Hedgehog regulates thymic epithelial cell differentiation.

    Science.gov (United States)

    Saldaña, José Ignacio; Solanki, Anisha; Lau, Ching-In; Sahni, Hemant; Ross, Susan; Furmanski, Anna L; Ono, Masahiro; Holländer, Georg; Crompton, Tessa

    2016-04-01

    Sonic Hedgehog (Shh) is expressed in the thymus, where it regulates T cell development. Here we investigated the influence of Shh on thymic epithelial cell (TEC) development. Components of the Hedgehog (Hh) signalling pathway were expressed by TEC, and use of a Gli Binding Site-green fluorescence protein (GFP) transgenic reporter mouse demonstrated active Hh-dependent transcription in TEC in the foetal and adult thymus. Analysis of Shh-deficient foetal thymus organ cultures (FTOC) showed that Shh is required for normal TEC differentiation. Shh-deficient foetal thymus contained fewer TEC than wild type (WT), the proportion of medullary TEC was reduced relative to cortical TEC, and cell surface expression of MHC Class II molecules was increased on both cortical and medullary TEC populations. In contrast, the Gli3-deficient thymus, which shows increased Hh-dependent transcription in thymic stroma, had increased numbers of TEC, but decreased cell surface expression of MHC Class II molecules on both cortical and medullary TEC. Neutralisation of endogenous Hh proteins in WT FTOC led to a reduction in TEC numbers, and in the proportion of mature Aire-expressing medullary TEC, but an increase in cell surface expression of MHC Class II molecules on medullary TEC. Likewise, conditional deletion of Shh from TEC in the adult thymus resulted in alterations in TEC differentiation and consequent changes in T cell development. TEC numbers, and the proportion of mature Aire-expressing medullary TEC were reduced, and cell surface expression of MHC Class II molecules on medullary TEC was increased. Differentiation of mature CD4 and CD8 single positive thymocytes was increased, demonstrating the regulatory role of Shh production by TEC on T cell development. Treatment of human thymus explants with recombinant Shh or neutralising anti-Shh antibody indicated that the Hedgehog pathway is also involved in regulation of differentiation from DP to mature SP T cells in the human thymus.

  12. Potential uses of milk epithelial cells: a review

    OpenAIRE

    2002-01-01

    International audience; Secretions collected from the mammary gland of different species contain heterogeneous populations of cells including lymphocytes, neutrophils, macrophages and epithelial cells in different species. Several factors influence the somatic cell count in milk and the distribution of cell types, such as species, infection status, physiological status and management practices. The epithelial cells are shed into milk during the lactation process. Most of them are viable and e...

  13. Desmosome dynamics in migrating epithelial cells requires the actin cytoskeleton

    Science.gov (United States)

    Roberts, Brett J.; Pashaj, Anjeza; Johnson, Keith R.; Wahl, James K.

    2011-01-01

    Re-modeling of epithelial tissues requires that the cells in the tissue rearrange their adhesive contacts in order to allow cells to migrate relative to neighboring cells. Desmosomes are prominent adhesive structures found in a variety of epithelial tissues that are believed to inhibit cell migration and invasion. Mechanisms regulating desmosome assembly and stability in migrating cells are largely unknown. In this study we established a cell culture model to examine the fate of desmosomal components during scratch wound migration. Desmosomes are rapidly assembled between epithelial cells at the lateral edges of migrating cells and structures are transported in a retrograde fashion while the structures become larger and mature. Desmosome assembly and dynamics in this system are dependent on the actin cytoskeleton prior to being associated with the keratin intermediate filament cytoskeleton. These studies extend our understanding of desmosome assembly and provide a system to examine desmosome assembly and dynamics during epithelial cell migration. PMID:21945137

  14. Observing planar cell polarity in multiciliated mouse airway epithelial cells.

    Science.gov (United States)

    Vladar, Eszter K; Lee, Yin Loon; Stearns, Tim; Axelrod, Jeffrey D

    2015-01-01

    The concerted movement of cilia propels inhaled contaminants out of the lungs, safeguarding the respiratory system from toxins, pathogens, pollutants, and allergens. Motile cilia on the multiciliated cells (MCCs) of the airway epithelium are physically oriented along the tissue axis for directional motility, which depends on the planar cell polarity (PCP) signaling pathway. The MCCs of the mouse respiratory epithelium have emerged as an important model for the study of motile ciliogenesis and the PCP signaling mechanism. Unlike other motile ciliated or planar polarized tissues, airway epithelial cells are relatively easily accessible and primary cultures faithfully model many of the essential features of the in vivo tissue. There is growing interest in understanding how cells acquire and polarize motile cilia due to the impact of mucociliary clearance on respiratory health. Here, we present methods for observing and quantifying the planar polarized orientation of motile cilia both in vivo and in primary culture airway epithelial cells. We describe how to acquire and evaluate electron and light microscopy images of ciliary ultrastructural features that reveal planar polarized orientation. Furthermore, we describe the immunofluorescence localization of PCP pathway components as a simple readout for airway epithelial planar polarization and ciliary orientation. These methods can be adapted to observe ciliary orientation in other multi- and monociliated cells and to detect PCP pathway activity in any tissue or cell type.

  15. The Preliminary Experimental Study of Induced Differentiation of Embryonic Stem Cells into Corneal Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    Ling Yu; Jian Ge; Zhichong Wang; Bing Huang; Keming Yu; Chongde Long; Xigu Chen

    2001-01-01

    Purpose:To study preliminarily induced differentiation of embryonic stem cells intocorneal epithelial cells in vitro.Methods: Murine embryonic stem cells were co-cultured with Rabbit limbal cornealepithelial cells in Transwell system to induce differentiation. Mophological andimmunohistochemical examination were implemented.Results: The induced cells from embryonic stem cells have an epithelial appearance.The cells formed a network and were confluent into film gradually after beingco-cultured with rabbit limbal corneal epithelial cells for 24 ~ 96 hours. The cells rangedmosaic structure and localized together with clear rim. Most of the cells showedpolygonal appearance. Transmission electron microscope showed lots of microvilli on thesurface of induced cells and tight junctions between them. These epithelial-like cellsexpressed the corneal epithelial cell specific marker cytokeratin3/cytokeratinl2.Conclusion: The potential mechanism of the differentiation of murine embryonic stemcells into corneal epithelial cells induced by limbal corneal epithelial cell-derivedinducing activity is to be further verified.

  16. 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

  17. Establishment of Hertwig’s Epithelial Root Sheath/Epithelial Rests of Malassez Cell Line from Human Periodontium

    OpenAIRE

    Nam, Hyun; Kim, Ji-Hye; Kim, Jae-Won; Seo, Byoung-Moo; Park, Joo-Cheol; Kim, Jung-Wook; Lee, Gene

    2014-01-01

    Human Hertwig’s epithelial root sheath/epithelial rests of Malassez (HERS/ERM) cells are epithelial remnants of teeth residing in the periodontium. Although the functional roles of HERS/ERM cells have yet to be elucidated, they are a unique epithelial cell population in adult teeth and are reported to have stem cell characteristics. Therefore, HERS/ERM cells might play a role as an epithelial component for the repair or regeneration of dental hard tissues; however, they are very rare populati...

  18. 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...

  19. Quantitative assessment of cytosolic Salmonella in epithelial cells.

    Directory of Open Access Journals (Sweden)

    Leigh A Knodler

    Full Text Available Within mammalian cells, Salmonella enterica serovar Typhimurium (S. Typhimurium inhabits a membrane-bound vacuole known as the Salmonella-containing vacuole (SCV. We have recently shown that wild type S. Typhimurium also colonizes the cytosol of epithelial cells. Here we sought to quantify the contribution of cytosolic Salmonella to the total population over a time course of infection in different epithelial cell lines and under conditions of altered vacuolar escape. We found that the lysosomotropic agent, chloroquine, acts on vacuolar, but not cytosolic, Salmonella. After chloroquine treatment, vacuolar bacteria are not transcriptionally active or replicative and appear degraded. Using a chloroquine resistance assay, in addition to digitonin permeabilization, we found that S. Typhimurium lyses its nascent vacuole in numerous epithelial cell lines, albeit with different frequencies, and hyper-replication in the cytosol is also widespread. At later times post-infection, cytosolic bacteria account for half of the total population in some epithelial cell lines, namely HeLa and Caco-2 C2Bbe1. Both techniques accurately measured increased vacuole lysis in epithelial cells upon treatment with wortmannin. By chloroquine resistance assay, we also determined that Salmonella pathogenicity island-1 (SPI-1, but not SPI-2, the virulence plasmid nor the flagellar apparatus, was required for vacuolar escape and cytosolic replication in epithelial cells. Together, digitonin permeabilization and the chloroquine resistance assay will be useful, complementary tools for deciphering the mechanisms of SCV lysis and Salmonella replication in the epithelial cell cytosol.

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

    NARCIS (Netherlands)

    P. Aparicio-Domingo (Patricia); M. Romera-Hernandez (Monica); 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

  1. Epithelial cells as alternative human biomatrices for comet assay

    Directory of Open Access Journals (Sweden)

    Emilio eRojas

    2014-11-01

    Full Text Available 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 thirty year period, the comet assay in epithelial cells has been litlle 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.

  2. 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

  3. Ion transport in epithelial spheroids derived from human airway cells

    DEFF Research Database (Denmark)

    Pedersen, P S; Frederiksen, O; Holstein-Rathlou, N H

    1999-01-01

    In the present study, we describe a novel three-dimensional airway epithelial explant preparation and demonstrate its use for ion transport studies by electrophysiological technique. Suspension cultures of sheets of epithelial cells released by protease treatment from cystic fibrosis (CF) and non...

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

    Indian Academy of Sciences (India)

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

    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-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 -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.

  5. Endoplasmic reticulum protein 29 regulates epithelial cell integrity during the mesenchymal-epithelial transition in breast cancer cells.

    Science.gov (United States)

    Bambang, I F; Lee, Y K; Richardson, D R; Zhang, D

    2013-03-07

    The epithelial-mesenchymal transition (EMT) correlates with disruption of cell-cell adhesion, loss of cell polarity and development of epithelial cell malignancy. Identifying novel molecules that inhibit EMT has profound potential for developing mechanism-based therapeutics. We previously demonstrated that the endoplasmic reticulum protein 29 (ERp29) is a novel factor that can drive mesenchymal-epithelial transition (MET) and induce cell growth arrest in MDA-MB-231 cells. Here, we show that ERp29 is an important molecule in establishing epithelial cell integrity during the MET. We demonstrate that ERp29 regulates MET in a cell context-dependent manner. ERp29 overexpression induced a complete MET in mesenchymal MDA-MB-231 cells through downregulating the expression of transcriptional repressors (for example, Slug, Snai1, ZEB2 and Twist) of E-cadherin. In contrast, overexpression of ERp29 induces incomplete MET in basal-like BT549 cells in which the expression of EMT-related markers (for example, vimentin; cytokeratin 19 (CK19) and E-cadherin) and the transcriptional repressors of E-cadherin were not altered. However, ERp29 overexpression in both cell-types resulted in loss of filamentous stress fibers, formation of cortical actin and restoration of an epithelial phenotype. Mechanistic studies revealed that overexpression of ERp29 in both cell-types upregulated the expression of TJ proteins (zonula-occludens-1 (ZO-1) and occludin) and the core apical-basal polarity proteins (Par3 and Scribble) at the membrane to enhance cell-cell contact and cell polarization. Knockdown of ERp29 in the epithelial MCF-7 cells decreased the expression of these proteins, leading to the disruption of cell-cell adhesion. Taken together, ERp29 is a novel molecule that regulates MET and epithelial cell integrity in breast cancer cells.

  6. Mitosis orientation in prostate epithelial cells changed by endocrine effect

    Institute of Scientific and Technical Information of China (English)

    Xiang-yun LIU; Dong-mei Li; Xiao-fang ZHANG; Jian-hui WU; Zu-yue SUN

    2008-01-01

    Aim: The aim of the present study was to investigate the effect of androgen and estrogen on mitosis orientation in the prostate epithelial cells of male rats. Methods: Castrated rats were treated with a single injection of testosterone propionate (TP) or benzogynestry (E2). There were 8 rats in the control group and TP-treated or E2-treated group. Prostate, liver, a specimen of skin, and a segment of the jejunum and colon were removed after the corresponding treatment. The results were observed through immunohistochemistry and iron hematoxylin-eosin staining.Results: All mitoses found in the prostate epithelial cells of castrated rats with TP were oriented parallel to the basement membrane; however, mitoses found in the prostate epithelial cells of castrated rats in E2 and the control group were oriented perpendicular to the basement membrane. TP treatment resulted in marked changes in mitosis orientation in the prostate epithelial cells. Bromodeoxyuridine-labeled positive cells could be seen throughout the stroma and prostate epithelial cells with an injection of TP; however, the positive cells could only be seen in the stroma of prostate with an injection of E2, and the positive cells could hardly be seen in the control group. Conclusion: We found a novel effect of TP in the prostate as a marked change of mitosis orientation in prostate epithelial cells.

  7. Cell cycle regulation by glucosamine in human pulmonary epithelial cells.

    Science.gov (United States)

    Chuang, Kun-Han; Lu, Chih-Shen; Kou, Yu Ru; Wu, Yuh-Lin

    2013-04-01

    Airway epithelial cells play an important role against intruding pathogens. Glucosamine, a commonly used supplemental compound, has recently begun to be regarded as a potential anti-inflammatory molecule. This study aimed to uncover how glucosamine impacts on cellular proliferation in human alveolar epithelial cells (A549) and bronchial epithelial cells (HBECs). With trypan blue-exclusion assay, we observed that glucosamine (10, 20, 50 mM) caused a decrease in cell number at 24 and 48 h; with a flow cytometric analysis, we also noted an enhanced cell accumulation within the G(0)/G(1) phase at 24 h and induction of late apoptosis at 24 and 48 h by glucosamine (10, 20, 50 mM) in A549 cells and HBECs. Examination of phosphorylation in retinoblastoma (Rb) protein, we found an inhibitory effect by glucosamine at 20 and 50 mM. Glucosamine at 50 mM was demonstrated to elevate both the mRNA and protein expression of p53 and heme oxygenase-1 (HO-1), but also caused a reduction in p21 protein expression. In addition, glucosamine attenuated p21 protein stability via the proteasomal proteolytic pathway, as well as inducing p21 nuclear accumulation. Altogether, our results suggest that a high dose of glucosamine may inhibit cell proliferation through apoptosis and disturb cell cycle progression with a halt at G(0)/G(1) phase, and that this occurs, at least in part, by a reduction in Rb phosphorylation together with modulation of p21, p53 and HO-1 expression, and nuclear p21 accumulation.

  8. Simvastatin Attenuates TGF-β1-Induced Epithelial-Mesenchymal Transition in Human Alveolar Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Tuo Yang

    2013-06-01

    Full Text Available Background: Transforming growth factor-β1 (TGF-β1-induced epithelial-mesenchymal transition (EMT of alveolar epithelial cells (AEC may contribute to idiopathic pulmonary fibrosis (IPF. TGF-β1-induced EMT in A549 cells (a human AEC cell line resulted in the adoption of mesenchymal responses that were predominantly mediated via the TGF-β1-Smad2/3 signaling pathway. Simvastatin (Sim, a 3-hydroxy-3-methylglutaryl CoA (HMG-CoA reductase inhibitor, has been previously reported to inhibit EMT in human proximal tubular epithelial cells and porcine lens epithelial cells and to suppress Smad2/3 phosphorylation in animal models. However, whether Sim can attenuate TGF-β1-induced EMT in A549 cells and its underlying mechanisms remains unknown. Methods: Cells were incubated with TGF-β1 in the presence or absence of Sim. The epithelial marker E-cadherin (E-Cad and the mesenchymal markers, α-smooth muscle actin (α-SMA, vimentin (Vi and fibronectin (FN, were detected using western blotting analyses and immunofluorescence. Phosphorylated Smad2 and Smad3 levels and connective tissue growth factor (CTGF were analyzed using western blotting. In addition, a cell migration assay was performed. Moreover, the levels of matrix metalloproteinase (MMP-2 and -9 in the culture medium were examined using ELISA. Results: Sim significantly attenuated the TGF-β1-induced decrease in E-Cad levels and elevated the levels of α-SMA, Vi and FN via the suppression of Smad2 and Smad3 phosphorylation. Furthermore, Sim inhibited the mesenchymal-like responses in A549 cells, including cell migration, CTGF expression and secretion of MMP-2 and -9. However, Sim failed to reverse the cell morphologial changes induced by TGF-β1 in A549 cells. Conclusion: Sim attenuated TGF-β1-induced EMT in A549 cells and might be a promising therapeutic agent for treating IPF.

  9. Cell volume regulation in epithelial physiology and cancer

    DEFF Research Database (Denmark)

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

    2013-01-01

    The physiological function of epithelia is transport of ions, nutrients, and fluid either in secretory or absorptive direction. All of these processes are closely related to cell volume changes, which are thus an integrated part of epithelial function. Transepithelial transport and cell volume...... 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...... 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....

  10. Rac promotes epithelial cell rearrangement during tracheal tubulogenesis in Drosophila.

    Science.gov (United States)

    Chihara, Takahiro; Kato, Kagayaki; Taniguchi, Misako; Ng, Julian; Hayashi, Shigeo

    2003-04-01

    Cell rearrangement, accompanied by the rapid assembly and disassembly of cadherin-mediated cell adhesions, plays essential roles in epithelial morphogenesis. Various in vitro and cell culture studies on the small GTPase Rac have suggested it to be a key regulator of cell adhesion, but this notion needs to be verified in the context of embryonic development. We used the tracheal system of Drosophila to investigate the function of Rac in the epithelial cell rearrangement, with a special attention to its role in regulating epithelial cadherin activity. We found that a reduced Rac activity led to an expansion of cell junctions in the embryonic epidermis and tracheal epithelia, which was accompanied by an increase in the amount of Drosophila E-Cadherin-Catenin complexes by a post-transcriptional mechanism. Reduced Rac activity inhibited dynamic epithelial cell rearrangement. Hyperactivation of Rac, on the other hand, inhibited assembly of newly synthesized E-Cadherin into cell junctions and caused loss of tracheal cell adhesion, resulting in cell detachment from the epithelia. Thus, in the context of Drosophila tracheal development, Rac activity must be maintained at a level necessary to balance the assembly and disassembly of E-Cadherin at cell junctions. Together with its role in cell motility, Rac regulates plasticity of cell adhesion and thus ensures smooth remodeling of epithelial sheets into tubules.

  11. Potential uses of milk epithelial cells: a review.

    Science.gov (United States)

    Boutinaud, Marion; Jammes, Hélène

    2002-01-01

    Secretions collected from the mammary gland of different species contain heterogeneous populations of cells including lymphocytes, neutrophils, macrophages and epithelial cells in different species. Several factors influence the somatic cell count in milk and the distribution of cell types, such as species, infection status, physiological status and management practices. The epithelial cells are shed into milk during the lactation process. Most of them are viable and exhibit the characteristics of fully differentiated alveolar cells. Primary cultures of epithelial cells from colostrum and milk of humans, baboons, cows and goats together with established cell lines from human and goat milk, provide a good model for the study of lactogenesis, immunity transmission, cancer research and infection by viruses. The RNA extracted from milk cells have been shown to be representative of gene expression in the mammary gland and thus provide a source of material for molecular studies of gene expression and environmental interactions.

  12. Immunochemical Detection of Metallothionein in Specific Epithelial Cells of Rat Organs

    Science.gov (United States)

    Danielson, Keith G.; Ohi, Seigo; Huang, P. C.

    1982-04-01

    The distribution of a heavy metal binding protein, metallothionein, was studied immunocytochemically by using antimetallothionein antibody and the immunoperoxidase staining technique on histological sections of liver, kidney, intestine, lung, and testis from cadmium-treated rats. These tissues either accumulate heavy metals (e.g., liver, kidney, and testis) or are exposed to metal by ingestion or inhalation (intestine and lung). Staining for metallothionein was observed intracellularly in epithelial parenchymal cells of the liver and kidney; all hepatocytes and most renal tubular cells stained for the protein. Accumulation of metallothionein was not seen in connective tissue cells surrounding either blood vessels or renal tubules. Extracellular localization of metallothionein was also observed in the liver sinusoids and within the lumina of the renal tubules, suggesting a metal transport or excretory function for this protein. Surface columnar epithelial cells of the intestinal villi indicated the presence of metallothionein but connective tissue cells of the lamina propria were negative for the protein. The granular secretory Paneth cells of the small intestine also stained strongly for metallothionein as did respiratory epithelial cells of the lung. In the testis, metallothionein was detected in the Sertoli cells and interstitial cells but not within the spermatogonia. Sertoli cells are closely associated with the developing spermatogonia and appear to serve a nutritive role in spermatogenesis. Because of the secretory, absorptive, or nutritive function of the metallothionein-localizing cells in the organs studied, we suggest that metallothionein may be involved in metal storage or transport in addition to its commonly proposed detoxification role.

  13. In Vitro transformation of LW13 Rat liver epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    SHICAN; KARLFETNANSKY; 等

    1992-01-01

    A rat liver epithelial cell line designated LW 13 was established using a sequential sedimentation method.The cell line retained many normal proerties of liver epithelial cells and showed some structural and functional features resembling those of liver parenchymal cells,LW13 cells became malignant after the intrduction of exogenous transforming EJ Ha ras gene,Tumors produced by inoculation of the transformed cells into baby rats contained areas of poorly differentialted hepatocellular carcinoma,In situ hybridization analysis confirmed the random rather than specific integration of exogenous ras gene into host chromosomes.Furthermore,an at least tenfold increase in the expression of the endogenous c mys gene was detected among transformed cell lines,suggesting the involvement of the c myc proto oncogene in the in vitro transformation of rat liver epithelial cells by EJ Ha ras oncogene.

  14. 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.

  15. Connective Tissue Growth Factor Expression in Human Bronchial Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    Amrita DOSANJH

    2006-01-01

    Connective tissue growth factor (CTGF) is a cysteine-rich protein that promotes extracellular matrix deposition. CTGF is selectively induced by transforming growth factor β and des-Arg kallidin in lung fibroblasts and increases steady-state mRNA levels of α type I collagen, 5α-integrin and fibronectin in fibroblasts. Bronchial epithelial cells have been proposed to functionally interact with lung fibroblasts. We therefore investigated if bronchial epithelial cells are able to synthesize CTGF. Human bronchial epithelial cells were grown to subconfluence in standard growth media. Proliferating cells grown in small airway growth media were harvested following starvation for up to 24 h. Expression of CTGF transcripts was measured by PCR. Immunocytochemistry was also completed using a commercially available antibody.The cells expressed readily detectable CTGF transcripts. Starvation of these cells resulted in a quantitative decline of CTGF transcripts. Direct sequencing of the PCR product identified human CTGF. Immunocytochemistry confirmed intracellular CTGF in the cells and none in negative control cells. We conclude that bronchial epithelial cells could be a novel source of CTGF. Bronchial epithelial cell-derived CTGF could thus directly influence the deposition of collagen in certain fibrotic lung diseases.

  16. Runx1 stabilizes the mammary epithelial cell phenotype and prevents epithelial to mesenchymal transition.

    Science.gov (United States)

    Hong, Deli; Messier, Terri L; Tye, Coralee E; Dobson, Jason R; Fritz, Andrew J; Sikora, Kenneth R; Browne, Gillian; Stein, Janet L; Lian, Jane B; Stein, Gary S

    2017-03-14

    Runx1 is a well characterized transcription factor essential for hematopoietic differentiation and Runx1 mutations are the cause of leukemias. Runx1 is highly expressed in normal epithelium of most glands and recently has been associated with solid tumors. Notably, the function of Runx1 in the mammary gland and how it is involved in initiation and progression of breast cancer is still unclear. Here we demonstrate the consequences of Runx1 loss in normal mammary epithelial and breast cancer cells. We first observed that Runx1 is decreased in tumorigenic and metastatic breast cancer cells. We also observed loss of Runx1 expression upon induction of epithelial-mesenchymal transition (EMT) in MCF10A (normal-like) cells. Furthermore depletion of Runx1 in MCF10A cells resulted in striking changes in cell shape, leading to mesenchymal cell morphology. The epithelial phenotype could be restored in breast cancer cells by re-expressing Runx1. Analyses of breast tumors and patient data revealed that low Runx1 expression is associated with poor prognosis and decreased survival. We addressed mechanisms for the function of Runx1 in maintaining the epithelial phenotype and find Runx1 directly regulates E-cadherin; and serves as a downstream transcription factor mediating TGFβ signaling. We also observed through global gene expression profiling of growth factor depleted cells that induction of EMT and loss of Runx1 is associated with activation of TGFβ and WNT pathways. Thus these findings have identified a novel function for Runx1 in sustaining normal epithelial morphology and preventing EMT and suggest Runx1 levels could be a prognostic indicator of tumor progression.

  17. 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.

  18. Attachment of epithelial cells and fibroblasts to ceramic materials.

    Science.gov (United States)

    Niederauer, G G; McGee, T D; Keller, J C; Zaharias, R S

    1994-04-01

    This study examined in vitro gingival epithelial and fibroblast cell attachment to ceramic materials made of tricalcium phosphate and/or magnesium aluminate spinel. The composite made of tricalcium phosphate and spinel is called 'osteoceramic'. These ceramics had various compositions and surface structures, which were initially characterized. Cell attachment assays were performed using both cell types to compare cellular response to the ceramic materials. Specimens were also prepared for scanning electron microscopy to investigate cellular morphology. The highest levels of cell attachment for gingival epithelial cells were observed on the rough osteoceramic surface, whereas gingival fibroblasts attached least to the rough osteoceramic surface.

  19. Trefoil peptides promote restitution of wounded corneal epithelial cells.

    Science.gov (United States)

    Göke, M N; Cook, J R; Kunert, K S; Fini, M E; Gipson, I K; Podolsky, D K

    2001-04-01

    The ocular surface shares many characteristics with mucosal surfaces. In both, healing is regulated by peptide growth factors, cytokines, and extracellular matrix proteins. However, these factors are not sufficient to ensure most rapid healing. Trefoil peptides are abundantly expressed epithelial cell products which exert protective effects and are key regulators of gastrointestinal epithelial restitution, the critical early phase of cell migration after mucosal injury. To assess the role of trefoil peptides in corneal epithelial wound healing, the effects of intestinal trefoil factor (ITF/TFF3) and spasmolytic polypeptide (SP/TFF2) on migration and proliferation of corneal epithelial cells were analyzed. Both ITF and SP enhanced restitution of primary rabbit corneal epithelial cells in vitro. While the restitution-enhancing effects of TGF-alpha and TGF-beta were both inhibited by neutralizing anti-TGF-beta-antibodies, trefoil peptide stimulation of restitution was not. Neither trefoil peptide significantly affected proliferation of primary corneal epithelial cells. ITF but not SP or pS2 mRNA was present in rabbit corneal and conjunctival tissues. In summary, the data indicate an unanticipated role of trefoil peptides in healing of ocular surface and demand rating their functional actions beyond the gastrointestinal tract.

  20. 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.

  1. Characterisation of cell adhesion in airway epithelial cell types using electric cell-substrate impedance sensing

    NARCIS (Netherlands)

    Heijink, I H; Brandenburg, S M; Noordhoek, J A; Postma, D S; Slebos, D-J; van Oosterhout, A J M

    Research on epithelial cell lines and primary epithelium is required to dissect the mechanisms underlying the structural abnormalities in airway epithelium observed for respiratory diseases, including asthma and chronic obstructive pulmonary disease. The novel electric cell-substrate impedance

  2. Probiotic Properties of Lactobacillus crispatus 2,029: Homeostatic Interaction with Cervicovaginal Epithelial Cells and Antagonistic Activity to Genitourinary Pathogens.

    Science.gov (United States)

    Abramov, Vyacheslav; Khlebnikov, Valentin; Kosarev, Igor; Bairamova, Guldana; Vasilenko, Raisa; Suzina, Natalia; Machulin, Andrey; Sakulin, Vadim; Kulikova, Natalia; Vasilenko, Nadezhda; Karlyshev, Andrey; Uversky, Vladimir; Chikindas, Michael L; Melnikov, Vyacheslav

    2014-12-01

    Lactobacillus crispatus 2029 isolated upon investigation of vaginal lactobacilli of healthy women of reproductive age was selected as a probiotic candidate. The aim of the present study was elucidation of the role of L. crispatus 2029 in resistance of the female reproductive tract to genitourinary pathogens using cervicovaginal epithelial model. Lactobacillus crispatus 2029 has surface layers (S-layers), which completely surround cells as the outermost component of their envelope. S-layers are responsible for the adhesion of lactobacilli on the surface of cervicovaginal epithelial cells. Study of interactions between L. crispatus 2029 and a type IV collagen, a major molecular component of epithelial cell extracellular matrix, showed that 125I-labeled type IV collagen binds to lactobacilli with high affinity (Kd = (8.0 ± 0.7) × 10(-10) M). Lactobacillus crispatus 2029 consistently colonized epithelial cells. There were no toxicity, epithelial damage and apoptosis after 24 h of colonization. Electronic microscope images demonstrated intimate association between L. crispatus 2029 and epithelial cells. Upon binding to epithelial cells, lactobacilli were recognized by toll-like 2/6 receptors. Lactobacillus crispatus induced NF-κB activation in epithelial cells and did not induce expression of innate immunity mediators IL-8, IL-1β, IL-1α and TNF-α. Lactobacillus crispatus 2029 inhibited IL-8 production in epithelial cells induced by MALP-2 and increased production of anti-inflammatory cytokine IL-6, maintaining the homeostasis of female reproductive tract. Lactobacillus crispatus 2029 produced H2O2 and provided wide spectrum of antagonistic activity increasing colonization resistance to urinary tract infections by bacterial vaginosis and vulvovaginal candidiasis associated agents.

  3. Lingual Epithelial Stem Cells and Organoid Culture of Them.

    Science.gov (United States)

    Hisha, Hiroko; Tanaka, Toshihiro; Ueno, Hiroo

    2016-01-28

    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.

  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. Starved epithelial cells uptake extracellular matrix for survival

    Science.gov (United States)

    Muranen, Taru; Iwanicki, Marcin P.; Curry, Natasha L.; Hwang, Julie; DuBois, Cory D.; Coloff, Jonathan L.; Hitchcock, Daniel S.; Clish, Clary B.; Brugge, Joan S.; Kalaany, Nada Y.

    2017-01-01

    Extracellular matrix adhesion is required for normal epithelial cell survival, nutrient uptake and metabolism. This requirement can be overcome by oncogene activation. Interestingly, inhibition of PI3K/mTOR leads to apoptosis of matrix-detached, but not matrix-attached cancer cells, suggesting that matrix-attached cells use alternate mechanisms to maintain nutrient supplies. Here we demonstrate that under conditions of dietary restriction or growth factor starvation, where PI3K/mTOR signalling is decreased, matrix-attached human mammary epithelial cells upregulate and internalize β4-integrin along with its matrix substrate, laminin. Endocytosed laminin localizes to lysosomes, results in increased intracellular levels of essential amino acids and enhanced mTORC1 signalling, preventing cell death. Moreover, we show that starved human fibroblasts secrete matrix proteins that maintain the growth of starved mammary epithelial cells contingent upon epithelial cell β4-integrin expression. Our study identifies a crosstalk between stromal fibroblasts and epithelial cells under starvation that could be exploited therapeutically to target tumours resistant to PI3K/mTOR inhibition. PMID:28071763

  6. Cell volume regulation in epithelial physiology and cancer

    DEFF Research Database (Denmark)

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

    2013-01-01

    The physiological function of epithelia is transport of ions, nutrients, and fluid either in secretory or absorptive direction. All of these processes are closely related to cell volume changes, which are thus an integrated part of epithelial function. Transepithelial transport and cell volume...... 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....... 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...

  7. Role of p53 in Mammary Epithelial Cell Senescence

    Science.gov (United States)

    2009-05-01

    culture of normal human breast epithelial cells. Methods Cell Biol 1980, 21B:107-135. 23. Easty GC, Easty DM, Monaghan P, Ormerod MG, Neville AM...27, 2006 Monitoring Editor: John Cleveland Polycomb group (PcG) protein Bmi-1 is an important regulator of cell proliferation. It regulates cellular

  8. A Three-Dimensional Cell Culture Model To Study Enterovirus Infection of Polarized Intestinal Epithelial Cells.

    Science.gov (United States)

    Drummond, Coyne G; Nickerson, Cheryl A; Coyne, Carolyn B

    2016-01-01

    epithelium lining the gastrointestinal tract early in infection. The lack of suitable in vivo and in vitro models to study CVB infection of the gastrointestinal epithelium has limited our understanding of the events that surround infection of these specialized cells. Here, we report on the development of a three-dimensional (3-D) organotypic cell culture model of human intestinal epithelial cells that better models the gastrointestinal epithelium in vivo. By applying this 3-D model, which recapitulates many aspects of the gastrointestinal epithelium in vivo, to the study of CVB infection, our work provides a new cell system to model the mechanisms by which CVB infects the intestinal epithelium, which may have a profound impact on CVB pathogenesis. Podcast: A podcast concerning this article is available.

  9. Epithelial cell division in the Xenopus laevis embryo during gastrulation.

    Science.gov (United States)

    Hatte, Guillaume; Tramier, Marc; Prigent, Claude; Tassan, Jean-Pierre

    2014-01-01

    How vertebrate epithelial cells divide in vivo and how the cellular environment influences cell division is currently poorly understood. A sine qua non condition to study cell division in situ is the ease of observation of cell division. This is fulfilled in the Xenopus embryo at the gastrula stage where polarized epithelial cells divide with a high frequency at the surface of the organism. Recently, using this model system, we have shown that epithelial cells divide by asymmetric furrowing and that the mode of cell division is regulated during development. Here, we further characterize epithelial cell division in situ. To this end, we used confocal microscopy to study epithelial cell division in the ectoderm of the Xenopus laevis gastrula. Cell division was followed either by indirect immunofluorescence in fixed embryos or by live imaging of embryos transiently expressing diverse fluorescent proteins. Here, we show that during cytokinesis, the plasma membranes of the two daughter cells are usually separated by a gap. For most divisions, daughter cells make contacts basally at a distance from the furrow tip which creates an inverted teardrop-like shaped volume tightly associated with the furrow. At the end of cytokinesis, the inverted teardrop is resorbed; thus it is a transient structure. Several proteins involved in cytokinesis are localized at the tip of the inverted teardrop suggesting that the formation of the gap could be an active process. We also show that intercalation of neighboring cells between daughter cells occasionally occurs during cytokinesis. Our results reveal an additional level of complexity in the relationship between dividing cells and also with their neighboring cells during cytokinesis in the Xenopus embryo epithelium.

  10. Change in Cell Shape Is Required for Matrix Metalloproteinase-Induced Epithelial-Mesenchymal Transition of Mammary Epithelial Cells

    Science.gov (United States)

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

    2010-01-01

    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-β-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. PMID:18506791

  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. Biomaterial surface proteomic signature determines interaction with epithelial cells.

    Science.gov (United States)

    Abdallah, Mohamed-Nur; Tran, Simon D; Abughanam, Ghada; Laurenti, Marco; Zuanazzi, David; Mezour, Mohamed A; Xiao, Yizhi; Cerruti, Marta; Siqueira, Walter L; Tamimi, Faleh

    2017-03-01

    Cells interact with biomaterials indirectly through extracellular matrix (ECM) proteins adsorbed onto their surface. Accordingly, it could be hypothesized that the surface proteomic signature of a biomaterial might determine its interaction with cells. Here, we present a surface proteomic approach to test this hypothesis in the specific case of biomaterial-epithelial cell interactions. In particular, we determined the surface proteomic signature of different biomaterials exposed to the ECM of epithelial cells (basal lamina). We revealed that the biomaterial surface chemistry determines the surface proteomic profile, and subsequently the interaction with epithelial cells. In addition, we found that biomaterials with surface chemistries closer to that of percutaneous tissues, such as aminated PMMA and aminated PDLLA, promoted higher selective adsorption of key basal lamina proteins (laminins, nidogen-1) and subsequently improved their interactions with epithelial cells. These findings suggest that mimicking the surface chemistry of natural percutaneous tissues can improve biomaterial-epithelial integration, and thus provide a rationale for the design of improved biomaterial surfaces for skin regeneration and percutaneous medical devices.

  13. 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...

  14. Immunolocalization of epithelial and mesenchymal matrix constituents in association with inner enamel epithelial cells.

    Science.gov (United States)

    Bosshardt, D D; Nanci, A

    1998-02-01

    After crown formation, the enamel organ reorganizes into Hertwig's epithelial root sheath (HERS). Although it is generally accepted that HERS plays an inductive role during root formation, it also has been suggested that it may contribute enamel-related proteins to cementum matrix. By analogy to the enamel-free area (EFA) in rat molars, in which epithelial cells express not only enamel proteins but also "typical" mesenchymal matrix constituents, it has been proposed that HERS cells may also have the potential to produce cementum proteins. To test this hypothesis, we examined the nature of the first matrix layer deposited along the cervical portion of root dentin and the characteristics of the associated cells. Rat molars were processed for postembedding colloidal gold immunolabeling with antibodies to amelogenin (AMEL), ameloblastin (AMBN), bone sialoprotein (BSP), and osteopontin (OPN). To minimize the possibility of false-negative results, several antibodies to AMEL were used. The labelings were compared with those obtained at the EFA. Initial cementum matrix was consistently observed at a time when epithelial cells from HERS covered most of the forming root surface. Cells with mesenchymal characteristics were rarely seen in proximity to the matrix. Both the EFA matrix and initial cementum exhibited collagen fibrils and were intensely immunoreactive for BSP and OPN. AMEL and AMBN were immunodetected at the EFA but not over the initial cementum proper. These two proteins were, however, present at the cervical-most portion of the root where enamel matrix extends for a short distance between dentin and cementum. These data suggest that epithelial cells along the root surface are likely responsible for the deposition of the initial cementum matrix and therefore, like the cells at the EFA, may be capable of producing mesenchymal proteins.

  15. Extracellular matrix proteins regulate epithelial-mesenchymal transition in mammary epithelial cells

    Science.gov (United States)

    Chen, Qike K.; Lee, KangAe; Radisky, Derek C.; Nelson, Celeste M.

    2013-01-01

    Mouse mammary epithelial cells undergo transdifferentiation via epithelial-mesenchymal transition (EMT) upon treatment with matrix metalloproteinase-3 (MMP3). In rigid microenvironments, MMP3 upregulates expression of Rac1b, which translocates to the cell membrane to promote induction of reactive oxygen species and EMT. Here we examine the role of the extracellular matrix (ECM) in this process. Our data show that the basement membrane protein laminin suppresses the EMT response in MMP3-treated cells, whereas fibronectin promotes EMT. These ECM proteins regulate EMT via interactions with their specific integrin receptors. α6-integrin sequesters Rac1b from the membrane and is required for inhibition of EMT by laminin. In contrast, α5-integrin maintains Rac1b at the membrane and is required for the promotion of EMT by fibronectin. Understanding the regulatory role of the ECM will provide insight into mechanisms underlying normal and pathological development of the mammary gland. PMID:23660532

  16. Effect of Helicobacter pylori on gastric epithelial cells

    Science.gov (United States)

    Alzahrani, Shatha; Lina, Taslima T; Gonzalez, Jazmin; Pinchuk, Irina V; Beswick, Ellen J; Reyes, Victor E

    2014-01-01

    The gastrointestinal epithelium has cells with features that make them a powerful line of defense in innate mucosal immunity. Features that allow gastrointestinal epithelial cells to contribute in innate defense include cell barrier integrity, cell turnover, autophagy, and innate immune responses. Helicobacter pylori (H. pylori) is a spiral shape gram negative bacterium that selectively colonizes the gastric epithelium of more than half of the world’s population. The infection invariably becomes persistent due to highly specialized mechanisms that facilitate H. pylori’s avoidance of this initial line of host defense as well as adaptive immune mechanisms. The host response is thus unsuccessful in clearing the infection and as a result becomes established as a persistent infection promoting chronic inflammation. In some individuals the associated inflammation contributes to ulcerogenesis or neoplasia. H. pylori has an array of different strategies to interact intimately with epithelial cells and manipulate their cellular processes and functions. Among the multiple aspects that H. pylori affects in gastric epithelial cells are their distribution of epithelial junctions, DNA damage, apoptosis, proliferation, stimulation of cytokine production, and cell transformation. Some of these processes are initiated as a result of the activation of signaling mechanisms activated on binding of H. pylori to cell surface receptors or via soluble virulence factors that gain access to the epithelium. The multiple responses by the epithelium to the infection contribute to pathogenesis associated with H. pylori. PMID:25278677

  17. AN IN VITRO MODEL FOR MURINE URETERIC EPITHELIAL CELLS

    Science.gov (United States)

    This report presents a model developed to study growth and differentiation of primary cultures of ureteric epithelial cells from embryonic C57BL/6N mouse urinary tracts. Single cells were resuspended in medium and plated onto transwells coated with collagen IV and laminin. Basa...

  18. Collective Movement of Epithelial Cells on a Collagen Gel Substrate

    OpenAIRE

    Haga, Hisashi; Irahara, Chikako; KOBAYASHI, Ryo; Nakagaki, Toshiyuki; Kawabata, Kazushige

    2004-01-01

    Collective cell movement acts as an efficient strategy in many physiological events, including wound healing, embryonic development, and morphogenesis. We found that epithelial cells (Madin-Darby canine kidney cell) migrated collectively along one direction on a collagen gel substrate. Time-lapse images of Madin-Darby canine kidney cells cultured on type-I collagen gels and glass substrates were captured by phase contrast microscopy equipped with an incubation system. On the gel substrate, th...

  19. Role of p53 Mammary Epithelial Cell Senescence

    Science.gov (United States)

    2005-05-01

    AD Award Number: DAMD17-02-1-0509 TITLE: Role of p53 Mammary Epithelial Cell Senescence PRINCIPAL INVESTIGATOR: Goberdhan P. Dimri, Ph.D. CONTRACTING ...type and However, Mucl , K-18, and ASMA were not expressed in luminal cell type groups [12,68]. Interestingly, a significant cells present in...13,17,27], the has also attracted a great interest in the field of breast cancer candidate mammary stem cells appear to be ESA+, Mucl -, research, and

  20. Oxidized alginate hydrogels as niche environments for corneal epithelial cells.

    Science.gov (United States)

    Wright, Bernice; De Bank, Paul A; Luetchford, Kim A; Acosta, Fernando R; Connon, Che J

    2014-10-01

    Chemical and biochemical modification of hydrogels is one strategy to create physiological constructs that maintain cell function. The aim of this study was to apply oxidised alginate hydrogels as a basis for development of a biomimetic niche for limbal epithelial stem cells that may be applied to treating corneal dysfunction. The stem phenotype of bovine limbal epithelial cells (LEC) and the viability of corneal epithelial cells (CEC) were examined in oxidised alginate gels containing collagen IV over a 3-day culture period. Oxidation increased cell viability (P ≤ 0.05) and this improved further with addition of collagen IV (P ≤ 0.01). Oxidised gels presented larger internal pores (diameter: 0.2-0.8 µm) than unmodified gels (pore diameter: 0.05-0.1 µm) and were significantly less stiff (P ≤ 0.001), indicating that an increase in pore size and a decrease in stiffness contributed to improved cell viability. The diffusion of collagen IV from oxidised alginate gels was similar to that of unmodified gels suggesting that oxidation may not affect the retention of extracellular matrix proteins in alginate gels. These data demonstrate that oxidised alginate gels containing corneal extracellular matrix proteins can influence corneal epithelial cell function in a manner that may impact beneficially on corneal wound healing therapy.

  1. Oxidized alginate hydrogels as niche environments for corneal epithelial cells

    Science.gov (United States)

    Wright, Bernice; De Bank, Paul A; Luetchford, Kim A; Acosta, Fernando R; Connon, Che J

    2014-01-01

    Chemical and biochemical modification of hydrogels is one strategy to create physiological constructs that maintain cell function. The aim of this study was to apply oxidised alginate hydrogels as a basis for development of a biomimetic niche for limbal epithelial stem cells that may be applied to treating corneal dysfunction. The stem phenotype of bovine limbal epithelial cells (LEC) and the viability of corneal epithelial cells (CEC) were examined in oxidised alginate gels containing collagen IV over a 3-day culture period. Oxidation increased cell viability (P ≤ 0.05) and this improved further with addition of collagen IV (P ≤ 0.01). Oxidised gels presented larger internal pores (diameter: 0.2–0.8 µm) than unmodified gels (pore diameter: 0.05–0.1 µm) and were significantly less stiff (P ≤ 0.001), indicating that an increase in pore size and a decrease in stiffness contributed to improved cell viability. The diffusion of collagen IV from oxidised alginate gels was similar to that of unmodified gels suggesting that oxidation may not affect the retention of extracellular matrix proteins in alginate gels. These data demonstrate that oxidised alginate gels containing corneal extracellular matrix proteins can influence corneal epithelial cell function in a manner that may impact beneficially on corneal wound healing therapy. © 2013 The Authors. Journal of Biomedical Materials Research Part A Published byWiley Periodicals, Inc. Part A: 102A: 3393–3400, 2014. PMID:24142706

  2. In vitro ultraviolet–induced damage in human corneal, lens, and retinal pigment epithelial cells

    OpenAIRE

    Youn, Hyun-Yi; McCanna, David J.; Sivak, Jacob G.; Jones, Lyndon W.

    2011-01-01

    Purpose The purpose was to develop suitable in vitro methods to detect ocular epithelial cell damage when exposed to UV radiation, in an effort to evaluate UV-absorbing ophthalmic biomaterials. Methods Human corneal epithelial cells (HCEC), lens epithelial cells (HLEC), and retinal pigment epithelial cells (ARPE-19) were cultured and Ultraviolet A/Ultraviolet B (UVA/UVB) blocking filters and UVB-only blocking filters were placed between the cells and a UV light source. Cells were irradiated w...

  3. Ultrastructural and immunohistochemical studies on Trichomonas vaginalis adhering to and phagocytizing genitourinary epithelial cells

    Institute of Scientific and Technical Information of China (English)

    陈文列; 陈金富; 钟秀容; 梁平; 林炜

    2004-01-01

    Background Trichomonas vaginalis (T. vaginalis) belongs to a common sexually transmitted disease pathogen causing genitourinary trichomoniasis in both sexes. We investigated the pathogenetic mechanism of genitourinary trichomoniasis.Methods Cultured T. vaginalis bodies were injected into the vaginas of rats, or incubated with genitourinary epithelial cells of female subjects, male subjects, and sperm. The ultrastructural and microscopic changes were observed via transmission and scanning electron microscopy and through microscopic histochemistry.Results Groups of T.vaginalis adhered to PAS positive columnar cells at the surface of stratified epithelium in the middle and upper portions of the vaginas. They also traversed under these cells. The parasites were shown to be PAS, cathepsin D, and actin positive, and they could release hydrolase into the cytoplasm of adhered epithelial cells. In the amebiform T.vaginalis, microfilaments were arranged into reticular formation. Similar phenomena were found during the interaction of T.vaginalis with host cells, both in vitro and in vivo. Usually several protozoa adhered to an epithelial cell and formed polymorphic pseudopodia or surface invaginations to surround and phagocytize the microvilli or other parts of the epithelial cytoplasm. Adhesion and phagocytosis of sperm by the protozoa occurred at 15-30 minutes of incubation. Digestion of sperm was found at 45-75 minutes and was complete at 90-105 minutes.Conclusions T.vaginalis tends to parasitize at the fornix of the vagina, because this is the site where columnar cells are rich in mucinogen granules and their microvilli are helpful for adhesion and nibbling. T.vaginalis possesses some invading and attacking abilities. Shape change, canalization, encystation, phagocytosis, digestion, the cell coat, cytoskeleton, and lysosome all play important roles in the process of adhesion. They have two methods of phagocytosis: nibbling and ingestion. Genitourinary epithelium may be

  4. Tiotropium attenuates IL-13-induced goblet cell metaplasia of human airway epithelial cells

    NARCIS (Netherlands)

    Kistemaker, Loes E. M.; Hiemstra, Pieter S.; Bos, I. Sophie T.; Bouwman, Susanne; van den Berge, Maarten; Hylkema, Machteld N.; Meurs, Herman; Kerstjens, Huib A. M.; Gosens, Reinoud

    2015-01-01

    BACKGROUND: It has been shown that acetylcholine is both a neurotransmitter and acts as a local mediator, produced by airway cells including epithelial cells. In vivo studies have demonstrated an indirect role for acetylcholine in epithelial cell differentiation. Here, we aimed to investigate direct

  5. Immortalized bovine mammary epithelial cells express stem cell markers and differentiate in vitro.

    Science.gov (United States)

    Hu, Han; Zheng, Nan; Gao, Haina; Dai, Wenting; Zhang, Yangdong; Li, Songli; Wang, Jiaqi

    2016-08-01

    The bovine mammary epithelial cell is a secretory cell, and its cell number and secretory activity determine milk production. In this study, we immortalized a bovine mammary epithelial cell line by SV40 large T antigen gene using a retrovirus based on Chinese Holstein primary mammary epithelial cells (CMEC) cultured in vitro. An immortalized bovine mammary epithelial cell line surpassed the 50-passage mark and was designated the CMEC-H. The immortalized mammary epithelial cells grew in close contact with each other and exhibited the typical cobblestone morphology characteristic with obvious boundaries. The telomerase expression of CMEC-H has consistently demonstrated the presence of telomerase activity as an immortalized cell line, but the cell line never induced tumor formation in nude mice. CMEC-H expressed epithelial (cytokeratins CK7, CK8, CK18, and CK19), mesenchymal (vimentin), and stem/progenitor (CD44 and p63) cell markers. The induced expression of milk proteins, αS1 -casein, β-casein, κ-casein, and butyrophilin, indicated that CMEC-H maintained the synthesis function of the mammary epithelial cells. The established immortalized bovine mammary epithelial cell line CMEC-H is capable of self-renewal and differentiation and can serve as a valuable reagent for studying the physiological mechanism of the mammary gland.

  6. Microarray analysis of human epithelial cell responses to bacterial interaction.

    Science.gov (United States)

    Mans, Jeffrey J; Lamont, Richard J; Handfield, Martin

    2006-09-01

    Host-pathogen interactions are inherently complex and dynamic. The recent use of human microarrays has been invaluable to monitor the effects of various bacterial and viral pathogens upon host cell gene expression programs. This methodology has allowed the host response transcriptome of several cell lines to be studied on a global scale. To this point, the great majority of reports have focused on the response of immune cells, including macrophages and dendritic cells. These studies revealed that the immune response to microbial pathogens is tailored to different microbial challenges. Conversely, the paradigm for epithelial cells has--until recently--held that the epithelium mostly served as a relatively passive physical barrier to infection. It is now generally accepted that the epithelial barrier contributes more actively to signaling events in the immune response. In light of this shift, this review will compare transcriptional profiling data from studies that involved host-pathogen interactions occurring with epithelial cells. Experiments that defined both a common core response, as well as pathogen-specific host responses will be discussed. This review will also summarize the contributions that transcriptional profiling analysis has made to our understanding of bacterial physio-pathogensis of infection. This will include a discussion of how host transcriptional responses can be used to infer the function of virulence determinants from bacterial pathogens interacting with epithelial mucosa. In particular, we will expand upon the lessons that have been learned from gastro-intestinal and oral pathogens, as well as from members of the commensal flora.

  7. Relationship between the number of cells surrounding oocytes and energy states of oocytes.

    Science.gov (United States)

    Munakata, Yasuhisa; Ichinose, Tomoya; Ogawa, Kaori; Itami, Nobuhiko; Tasaki, Hidetaka; Shirasuna, Koumei; Kuwayama, Takehito; Iwata, Hisataka

    2016-10-15

    Lipid content, ATP content, and histone acetylation are thought to reflect the energy state of cells. In addition, the energy state closely associates with growth and developmental ability of oocytes. Oocyte growth is accompanied by active proliferation of the surrounding granulosa cells (GCs), and GCs play a key role in the provision of energy substrates to the oocytes. In the present study, we first examined the relationship among the average number of GCs per follicle, the average number of cumulus cells (CCs) per oocyte, and the average lipid content in oocytes that developed in vivo within individual donor gilts. Second, we validated the relationship between the number of cells surrounding oocytes and the energy states of oocytes by using an IVC system of oocyte granulosa cell complexes (OGCs) derived from early antral follicles. We collected cumulus cells and oocyte complexes (COCs) from antral follicles (3-5 mm in diameter) and found that average lipid content in oocytes significantly correlated with the average number of both GCs/follicle and CCs/oocyte (P cell number of OGCs, as well as the lipid content, ATP content, and acetylation level of H4K12 in oocytes grown in vitro. In addition, glucose consumption by OGCs was calculated from the sample media collected at Days 13 and 14. The lipid content of oocytes grown in vitro, significantly correlated with the number of cells surrounding the oocytes (P number of cells surrounding the oocytes (P number of cells surrounding the oocytes, and glucose uptake by OGCs is crucial for lipid content and ATP content, and H4K12 acetylation in oocytes.

  8. Overexpression of c-myc induces epithelial mesenchymal transition in mammary epithelial cells.

    Science.gov (United States)

    Cho, Kyoung Bin; Cho, Min Kyong; Lee, Won Young; Kang, Keon Wook

    2010-07-28

    The c-myc gene is frequently overexpressed in human breast cancer and its target genes are involved in tumorigenesis. Epithelial mesenchymal transitions (EMT), where cells undergo a developmental switch from a polarized epithelial phenotype to a highly motile mesenchymal phenotype, are associated with invasion and motility of cancer cells. Basal E-cadherin expression was down-regulated in c-myc overexpressing MCF10A (c-myc-MCF10A) cells compared to GFP-overexpressing MCF10A (GFP-MCF10A) cells, while N-cadherin was distinctly increased in c-myc-MCF10A cells. Given that glycogen synthase kinase-3beta (GSK-3beta) and the snail axis have key roles in E-cadherin deregulation during EMT, we investigated the role of GSK-3beta/snail signaling pathways in the induction of EMT by c-myc overexpression. In contrast to GFP-MCF10A cells, both the transcriptional activity and the ubiquitination-dependent protein stability of snail were enhanced in c-myc-MCF10A cells, and this was reversed by GSK-3beta overexpression. We also found that c-myc overexpression inhibits GSK-3beta activity through activation of extracellular signal-regulated kinase (ERK). Inhibition of ERK by dominant negative mutant transfection or chemical inhibitor significantly suppressed snail gene transcription. These results suggest that c-myc overexpression during transformation of mammary epithelial cells (MEC) is involved in EMTs via ERK-dependent GSK-3beta inactivation and subsequent snail activation.

  9. Apoptotic cell clearance by bronchial epithelial cells critically influences airway inflammation

    Science.gov (United States)

    Juncadella, Ignacio J.; Kadl, Alexandra; Sharma, Ashish K.; Shim, Yun M.; Hochreiter-Hufford, Amelia; Borish, Larry; Ravichandran, Kodi S.

    2013-01-01

    Lung epithelial cells can influence immune responses to airway allergens1,2. Airway epithelial cells also undergo apoptosis after encountering environmental allergens3; yet, relatively little is known about how these are cleared, and their effect on airway inflammation. Here we show that airway epithelial cells efficiently engulf apoptotic epithelial cells and secrete anti-inflammatory cytokines, dependent upon intracellular signalling by the small GTPase Rac1. Inducible deletion of Rac1 expression specifically in airway epithelial cells in a mouse model resulted in defective engulfment by epithelial cells and aberrant anti-inflammatory cytokine production. Intranasal priming and challenge of these mice with house dust mite extract or ovalbumin as allergens led to exacerbated inflammation, augmented Th2 cytokines and airway hyper-responsiveness, with decreased interleukin (IL)-10 in bronchial lavages. Rac1-deficient epithelial cells produced much higher IL-33 upon allergen or apoptotic cell encounter, with increased numbers of nuocyte-like cells1,4,5. Administration of exogenous IL-10 ‘rescued’ the airway inflammation phenotype in Rac1-deficient mice, with decreased IL-33. Collectively, these genetic and functional studies suggest a new role for Rac1-dependent engulfment by airway epithelial cells and in establishing the anti-inflammatory environment, and that defects in cell clearance in the airways could contribute to inflammatory responses towards common allergens. PMID:23235830

  10. Oxidative Stress, Cell Death, and Other Damage to Alveolar Epithelial Cells Induced by Cigarette Smoke

    Directory of Open Access Journals (Sweden)

    Nagai A

    2003-09-01

    Full Text Available Abstract Cigarette smoking is a major risk factor in the development of various lung diseases, including pulmonary emphysema, pulmonary fibrosis, and lung cancer. The mechanisms of these diseases include alterations in alveolar epithelial cells, which are essential in the maintenance of normal alveolar architecture and function. Following cigarette smoking, alterations in alveolar epithelial cells induce an increase in epithelial permeability, a decrease in surfactant production, the inappropriate production of inflammatory cytokines and growth factors, and an increased risk of lung cancer. However, the most deleterious effect of cigarette smoke on alveolar epithelial cells is cell death, i.e., either apoptosis or necrosis depending on the magnitude of cigarette smoke exposure. Cell death induced by cigarette smoke exposure can largely be accounted for by an enhancement in oxidative stress. In fact, cigarette smoke contains and generates many reactive oxygen species that damage alveolar epithelial cells. Whether apoptosis and/or necrosis in alveolar epithelial cells is enhanced in healthy cigarette smokers is presently unclear. However, recent evidence indicates that the apoptosis of alveolar epithelial cells and alveolar endothelial cells is involved in the pathogenesis of pulmonary emphysema, an important cigarette smoke-induced lung disease characterized by the loss of alveolar structures. This review will discuss oxidative stress, cell death, and other damage to alveolar epithelial cells induced by cigarette smoke.

  11. Epithelial neoplasia in Drosophila entails switch to primitive cell states.

    Science.gov (United States)

    Khan, Sumbul J; Bajpai, Anjali; Alam, Mohammad Atif; Gupta, Ram P; Harsh, Sneh; Pandey, Ravi K; Goel-Bhattacharya, Surbhi; Nigam, Aditi; Mishra, Arati; Sinha, Pradip

    2013-06-11

    Only select cell types in an organ display neoplasia when targeted oncogenically. How developmental lineage hierarchies of these cells prefigure their neoplastic propensities is not yet well-understood. Here we show that neoplastic Drosophila epithelial cells reverse their developmental commitments and switch to primitive cell states. In a context of alleviated tissue surveillance, for example, loss of Lethal giant larvae (Lgl) tumor suppressor in the wing primordium induced epithelial neoplasia in its Homothorax (Hth)-expressing proximal domain. Transcriptional profile of proximally transformed mosaic wing epithelium and functional tests revealed tumor cooperation by multiple signaling pathways. In contrast, lgl(-) clones in the Vestigial (Vg)-expressing distal wing epithelium were eliminated by cell death. Distal lgl(-) clones, however, could transform when both tissue surveillance and cell death were compromised genetically and, alternatively, when the transcription cofactor of Hippo signaling pathway, Yorkie (Yki), was activated, or when Ras/EGFR signaling was up-regulated. Furthermore, transforming distal lgl(-) clones displayed loss of Vg, suggesting reversal of their terminal cell fate commitment. In contrast, reinforcing a distal (wing) cell fate commitment in lgl(-) clones by gaining Vg arrested their neoplasia and induced cell death. We also show that neoplasia in both distal and proximal lgl(-) clones could progress in the absence of Hth, revealing Hth-independent wing epithelial neoplasia. Likewise, neoplasia in the eye primordium resulted in loss of Elav, a retinal cell marker; these, however, switched to an Hth-dependent primitive cell state. These results suggest a general characteristic of "cells-of-origin" in epithelial cancers, namely their propensity for switch to primitive cell states.

  12. Surface position, not signaling from surrounding maternal tissues, specifies aleurone epidermal cell fate in maize.

    Science.gov (United States)

    Gruis, Darren Fred; Guo, Hena; Selinger, David; Tian, Qing; Olsen, Odd-Arne

    2006-07-01

    Maize (Zea mays) endosperm consists of an epidermal-like surface layer of aleurone cells, an underlying body of starchy endosperm cells, and a basal layer of transfer cells. To determine whether surrounding maternal tissues perform a role in specifying endosperm cell fates, a maize endosperm organ culture technique was established whereby the developing endosperm is completely removed from surrounding maternal tissues. Using cell type-specific fluorescence markers, we show that aleurone cell fate specification occurs exclusively in response to surface position and does not require specific, continued maternal signal input. The starchy endosperm and aleurone cell fates are freely interchangeable throughout the lifespan of the endosperm, with internalized aleurone cells converting to starchy endosperm cells and with starchy endosperm cells that become positioned at the surface converting to aleurone cells. In contrast to aleurone and starchy endosperm cells, transfer cells fail to develop in in vitro-grown endosperm, supporting earlier indications that maternal tissue interaction is required to fully differentiate this cell type. Several parameters confirm that the maize endosperm organ cultures described herein retain the main developmental features of in planta endosperm, including fidelity of aleurone mutant phenotypes, temporal and spatial control of cell type-specific fluorescent markers, specificity of cell type transcripts, and control of mitotic cell divisions.

  13. CXCL12 expression by healthy and malignant ovarian epithelial cells

    Directory of Open Access Journals (Sweden)

    Emilie Dominique

    2011-03-01

    Full Text Available Abstract Background CXCL12 has been widely reported to play a biologically relevant role in tumor growth and spread. In epithelial ovarian cancer (EOC, CXCL12 enhances tumor angiogenesis and contributes to the immunosuppressive network. However, its prognostic significance remains unclear. We thus compared CXCL12 status in healthy and malignant ovaries, to assess its prognostic value. Methods Immunohistochemistry was used to analyze CXCL12 expression in the reproductive tracts, including the ovaries and fallopian tubes, of healthy women, in benign and borderline epithelial tumors, and in a series of 183 tumor specimens from patients with advanced primary EOC enrolled in a multicenter prospective clinical trial of paclitaxel/carboplatin/gemcitabine-based chemotherapy (GINECO study. Univariate COX model analysis was performed to assess the prognostic value of clinical and biological variables. Kaplan-Meier methods were used to generate progression-free and overall survival curves. Results Epithelial cells from the surface of the ovary and the fallopian tubes stained positive for CXCL12, whereas the follicles within the ovary did not. Epithelial cells in benign, borderline and malignant tumors also expressed CXCL12. In EOC specimens, CXCL12 immunoreactivity was observed mostly in epithelial tumor cells. The intensity of the signal obtained ranged from strong in 86 cases (47% to absent in 18 cases ( Conclusion Our findings highlight the previously unappreciated constitutive expression of CXCL12 on healthy epithelia of the ovary surface and fallopian tubes, indicating that EOC may originate from either of these epithelia. We reveal that CXCL12 production by malignant epithelial cells precedes tumorigenesis and we confirm in a large cohort of patients with advanced EOC that CXCL12 expression level in EOC is not a valuable prognostic factor in itself. Trial Registration ClinicalTrials.gov: NCT00052468

  14. Mannheimia haemolytica biofilm formation on bovine respiratory epithelial cells.

    Science.gov (United States)

    Boukahil, Ismail; Czuprynski, Charles J

    2016-12-25

    Mannheimia haemolytica is the most important bacterial agent associated with the bovine respiratory disease complex (BRDC), which causes worldwide economic losses to the cattle industry. M. haemolytica cells initially colonize the tonsillar crypts in the upper respiratory tract of cattle, from where they can subsequently descend into the lungs to cause disease. Many bacteria exist as biofilms inside their hosts. We hypothesize that M. haemolytica colonization of cattle during its commensal state may include biofilm formation. To begin to assess this possibility, we developed an in vitro system to study biofilm formation directly on bovine respiratory epithelial cells. Using fixed primary bovine bronchial epithelial cells, we observed M. haemolytica biofilm formation after a 48h incubation period at 37°C. Addition of mucin, the main component of mucus present in the upper respiratory tract, decreased M. haemolytica biofilm formation on bovine epithelial cells. We investigated the effects of prior viral infection of the epithelial cells on subsequent biofilm formation by M. haemolytica and found negligible effects. Utilization of this model system will provide new insights into the potential role of biofilm formation by M. haemolytica in the pathogenesis of BRDC.

  15. Apicobasal Polarity Controls Lymphocyte Adhesion to Hepatic Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Natalia Reglero-Real

    2014-09-01

    Full Text Available Loss of apicobasal polarity is a hallmark of epithelial pathologies. Leukocyte infiltration and crosstalk with dysfunctional epithelial barriers are crucial for the inflammatory response. Here, we show that apicobasal architecture regulates the adhesion between hepatic epithelial cells and lymphocytes. Polarized hepatocytes and epithelium from bile ducts segregate the intercellular adhesion molecule 1 (ICAM-1 adhesion receptor onto their apical, microvilli-rich membranes, which are less accessible by circulating immune cells. Upon cell depolarization, hepatic ICAM-1 becomes exposed and increases lymphocyte binding. Polarized hepatic cells prevent ICAM-1 exposure to lymphocytes by redirecting basolateral ICAM-1 to apical domains. Loss of ICAM-1 polarity occurs in human inflammatory liver diseases and can be induced by the inflammatory cytokine tumor necrosis factor alpha (TNF-α. We propose that adhesion receptor polarization is a parenchymal immune checkpoint that allows functional epithelium to hamper leukocyte binding. This contributes to the haptotactic guidance of leukocytes toward neighboring damaged or chronically inflamed epithelial cells that expose their adhesion machinery.

  16. Integrin Signaling in Mammary Epithelial Cells and Breast Cancer

    OpenAIRE

    Lambert, Arthur W.; Sait Ozturk; Sam Thiagalingam

    2012-01-01

    Cells sense and respond to the extracellular matrix (ECM) by way of integrin receptors, which facilitate cell adhesion and intracellular signaling. Advances in understanding the mammary epithelial cell hierarchy are converging with new developments that reveal how integrins regulate the normal mammary gland. But in breast cancer, integrin signaling contributes to the development and progression of tumors. This paper highlights recent studies which examine the role of integrin signaling in mam...

  17. Intestinal epithelial cells and their role in innate mucosal immunity

    OpenAIRE

    Maldonado-Contreras, A. L.; McCormick, Beth A

    2010-01-01

    The mucosal surfaces of the respiratory, gastrointestinal and urogenital tracts are covered by a layer of epithelial cells that are responsible for sensing and promoting a host immune response in order to establish the limits not only for commensal microorganisms but also for foreign organisms or particles. This is a remarkable task as the human body represents a composite of about 10 trillion human-self cells plus non-self cells from autochthonous or indigenous microbes that outnumber human ...

  18. Epithelial cell adhesion and gastrointestinal colonization of Lactobacillus in poultry.

    Science.gov (United States)

    Spivey, Megan A; Dunn-Horrocks, Sadie L; Duong, Tri

    2014-11-01

    Administration of probiotic Lactobacillus cultures is an important alternative to the use of antibiotic growth promoters and has been demonstrated to improve animal health, growth performance, and preharvest food safety in poultry production. Whereas gastrointestinal colonization is thought to be critical to their probiotic functionality, factors important to Lactobacillus colonization in chickens are not well understood. In this study we investigate epithelial cell adhesion in vitro and colonization of Lactobacillusin vivo in broiler chickens. Adhesion of Lactobacillus cultures to epithelial cells was evaluated using the chicken LMH cell line. Lactobacillus cultures were able adhere effectively to LMH cells relative to Bacillus subtilis and Salmonella Typhimurium. Epithelial cell adhesion was similar for Lactobacillus crispatus TDCC 75, L. cristpatus TDCC 76, and Lactobacillus gallinarum TDCC 77, and all 3 were more adherent than L. gallinarum TDCC 78. However, when colonization was evaluated in the ileum and cecum of broiler chicks, L. crispatus TDCC 75 and L. gallinarum TDCC 77 were more persistent than L. crispatus TDCC 76 and L. gallinarum TDCC 78. The reduction of growth in medium supplemented with oxgal was greater for L. gallinarum TDCC 78 than L. gallinarum TDCC 77, suggesting that whereas adhesion was similar for the 2 strains, the difference in colonization between L. gallinarum strains may be due in part to their bile sensitivity. This study demonstrates that whereas adhesion to epithelial cells may be important in predicting gastrointestinal colonization, other factors including bile tolerance may also contribute to the colonization of Lactobacillus in poultry. Additionally, the chicken LMH cell line is expected to provide a platform for investigating mechanisms of Lactobacillus adhesion to epithelial tissue and evaluating the probiotic potential Lactobacillus in poultry.

  19. AM251 Suppresses Epithelial-Mesenchymal Transition of Renal Tubular Epithelial Cells

    Science.gov (United States)

    Yoshinaga, Tomoyo; Uwabe, Kenichiro; Naito, Shoichi; Higashino, Kenichi; Nakano, Toru; Numata, Yoshito

    2016-01-01

    Epithelial-mesenchymal transition (EMT) of renal tubular epithelial cells is one of the causative mechanisms of kidney fibrosis. In our study, we screened lipophilic compounds using a lipid library including approximately 200 lipids to identify those that suppressed EMT induced by a transforming growth factor (TGF)-β1 stimulus. Initial screening was performed with the immortalized HK-2 renal tubule epithelial cell line. The most promising compounds were further tested in RPTEC primary renal tubule epithelial cells. We found that the synthetic lipid AM251 suppressed two hallmark events associated with EMT, the upregulation of collagen 1A1 (COL1A1) and downregulation of E-cadherin. Though AM251 is known to act as an antagonist for the cannabinoid receptor type 1 (CB1) and an agonist for the G protein-coupled receptor 55 (GRP55), the suppression of EMT by AM251 was not mediated through either receptor. Microarray analyses revealed that AM251 inhibited induction of several EMT transcription factors such as SNAIL1, which is the key inducer of EMT, and the AP-1 transcription factors FOSB and JUNB. Activation of SMAD2/3 and p38 mitogen-activated protein kinase (MAPK) was inhibited by AM251, with greater inhibition of the latter, indicating that AM251 acted upstream of SMAD/p38 MAPK in the TGF-β signaling pathway. Our findings regarding the effects of AM251 on the TGF-β signaling pathway may inform development of a novel therapeutic agent suppressing EMT, thus preventing kidney fibrosis. PMID:27936102

  20. AM251 Suppresses Epithelial-Mesenchymal Transition of Renal Tubular Epithelial Cells.

    Science.gov (United States)

    Yoshinaga, Tomoyo; Uwabe, Kenichiro; Naito, Shoichi; Higashino, Kenichi; Nakano, Toru; Numata, Yoshito; Kihara, Akio

    2016-01-01

    Epithelial-mesenchymal transition (EMT) of renal tubular epithelial cells is one of the causative mechanisms of kidney fibrosis. In our study, we screened lipophilic compounds using a lipid library including approximately 200 lipids to identify those that suppressed EMT induced by a transforming growth factor (TGF)-β1 stimulus. Initial screening was performed with the immortalized HK-2 renal tubule epithelial cell line. The most promising compounds were further tested in RPTEC primary renal tubule epithelial cells. We found that the synthetic lipid AM251 suppressed two hallmark events associated with EMT, the upregulation of collagen 1A1 (COL1A1) and downregulation of E-cadherin. Though AM251 is known to act as an antagonist for the cannabinoid receptor type 1 (CB1) and an agonist for the G protein-coupled receptor 55 (GRP55), the suppression of EMT by AM251 was not mediated through either receptor. Microarray analyses revealed that AM251 inhibited induction of several EMT transcription factors such as SNAIL1, which is the key inducer of EMT, and the AP-1 transcription factors FOSB and JUNB. Activation of SMAD2/3 and p38 mitogen-activated protein kinase (MAPK) was inhibited by AM251, with greater inhibition of the latter, indicating that AM251 acted upstream of SMAD/p38 MAPK in the TGF-β signaling pathway. Our findings regarding the effects of AM251 on the TGF-β signaling pathway may inform development of a novel therapeutic agent suppressing EMT, thus preventing kidney fibrosis.

  1. Immunohistochemical demonstration of airway epithelial cell markers of guinea pig.

    Science.gov (United States)

    Li, Yong; Wang, Jing; He, Hai Yan; Ma, Ling Jie; Zeng, Jin; Deng, Guang Cun; Liu, Xiaoming; Engelhardt, John F; Wang, Yujiong

    2011-10-01

    The guinea pig (Cavea porcellus) is a mammalian non-rodent species in the Caviidae family. The sensitivity of the respiratory system and the susceptibility to infectious diseases allows the guinea pig to be a useful model for both infectious and non-infectious lung diseases such as asthma and tuberculosis. In this report, we demonstrated for the first time, the major cell types and composition in the guinea pig airway epithelium, using cell type-specific markers by immunohistochemical staining using the commercial available immunological reagents that cross-react with guinea pig. Our results revealed the availability of antibodies cross-reacting with airway epithelial cell types of basal, non-ciliated columnar, ciliated, Clara, goblet and alveolar type II cells, as well as those cells expressing Mucin 5AC, Mucin 2, Aquaporin 4 and Calcitonin Gene Related Peptide. The distribution of these various cell types were quantified in the guinea pig airway by immunohistochemical staining and were comparable with morphometric studies using an electron microscopy assay. Moreover, this study also demonstrated that goblet cells are the main secretory cell type in the guinea pig's airway, distinguishing this species from rats and mice. These results provide useful information for the understanding of airway epithelial cell biology and mechanisms of epithelial-immune integration in guinea pig models.

  2. The similarity between human embryonic stem cell-derived epithelial cells and ameloblast-lineage cells

    Institute of Scientific and Technical Information of China (English)

    Li-Wei Zheng; Logan Linthicum; Pamela K DenBesten; Yan Zhang

    2013-01-01

    This study aimed to compare epithelial cells derived from human embryonic stem cells (hESCs) to human ameloblast-lineage cells (ALCs), as a way to determine their potential use as a cell source for ameloblast regeneration. Induced by various concentrations of bone morphogenetic protein 4 (BMP4), retinoic acid (RA) and lithium chloride (LiCI) for 7 days, hESCs adopted cobble-stone epithelial phenotype (hESC-derived epithelial cells (ES-ECs)) and expressed cytokeratin 14. Compared with ALCs and oral epithelial cells (OE), ES-ECs expressed amelogenesis-associated genes similar to ALCs. ES-ECs were compared with human fetal skin epithelium, human fetal oral buccal mucosal epithelial cells and human ALCs for their expression pattern of cytokeratins as well. ALCs had relatively high expression levels of cytokeratin 76, which ,vas also found to be upregulated in ES-ECs. Based on the present study, with the similarity of gene expression with ALCs, ES-ECs are a promising potential cell source for regeneration, which are not available in erupted human teeth for regeneration of enamel.

  3. Sodium selectivity of semicircular canal duct epithelial cells

    Directory of Open Access Journals (Sweden)

    Harbidge Donald G

    2011-09-01

    Full Text Available Abstract Background Sodium absorption by semicircular canal duct (SCCD epithelial cells is thought to contribute to the homeostasis of the volume of vestibular endolymph. It was previously shown that the epithelial cells could absorb Na+ under control of a glucocorticoid hormone (dexamethasone and the absorptive transepithelial current was blocked by amiloride. The most commonly-observed target of amiloride is the epithelial sodium channel (ENaC, comprised of the three subunits α-, β- and γ-ENaC. However, other cation channels have also been observed to be sensitive in a similar concentration range. The aim of this study was to determine whether SCCD epithelial cells absorb only Na+ or also K+ through an amiloride-sensitive pathway. Parasensory K+ absorption could contribute to regulation of the transduction current through hair cells, as found to occur via vestibular transitional cells [S. H. Kim and D. C. Marcus. Regulation of sodium transport in the inner ear. Hear.Res. doi:10.1016/j.heares.2011.05.003, 2011]. Results We determined the molecular and functional expression of candidate cation channels with gene array (GEO GSE6197, whole-cell patch clamp and transepithelial recordings in primary cultures of rat SCCD. α-, β- and γ-ENaC were all previously reported as present. The selectivity of the amiloride-sensitive transepithelial and cell membrane currents was observed in Ussing chamber and whole-cell patch clamp recordings. The cell membrane currents were carried by Na+ but not K+, but the Na+ selectivity disappeared when the cells were cultured on impermeable supports. Transepithelial currents across SCCD were also carried exclusively by Na+. Conclusions These results are consistent with the amiloride-sensitive absorptive flux of SCCD mediated by a highly Na+-selective channel, likely αβγ-ENaC. These epithelial cells therefore absorb only Na+ via the amiloride-sensitive pathway and do not provide a parasensory K+ efflux from the

  4. 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

  5. Computational investigation of epithelial cell dynamic phenotype in vitro

    Directory of Open Access Journals (Sweden)

    Debnath Jayanta

    2009-05-01

    Full Text Available Abstract Background When grown in three-dimensional (3D cultures, epithelial cells typically form cystic organoids that recapitulate cardinal features of in vivo epithelial structures. Characterizing essential cell actions and their roles, which constitute the system's dynamic phenotype, is critical to gaining deeper insight into the cystogenesis phenomena. Methods Starting with an earlier in silico epithelial analogue (ISEA1 that validated for several Madin-Darby canine kidney (MDCK epithelial cell culture attributes, we built a revised analogue (ISEA2 to increase overlap between analogue and cell culture traits. Both analogues used agent-based, discrete event methods. A set of axioms determined ISEA behaviors; together, they specified the analogue's operating principles. A new experimentation framework enabled tracking relative axiom use and roles during simulated cystogenesis along with establishment of the consequences of their disruption. Results ISEA2 consistently produced convex cystic structures in a simulated embedded culture. Axiom use measures provided detailed descriptions of the analogue's dynamic phenotype. Dysregulating key cell death and division axioms led to disorganized structures. Adhering to either axiom less than 80% of the time caused ISEA1 to form easily identified morphological changes. ISEA2 was more robust to identical dysregulation. Both dysregulated analogues exhibited characteristics that resembled those associated with an in vitro model of early glandular epithelial cancer. Conclusion We documented the causal chains of events, and their relative roles, responsible for simulated cystogenesis. The results stand as an early hypothesis–a theory–of how individual MDCK cell actions give rise to consistently roundish, cystic organoids.

  6. Cell-extrinsic consequences of epithelial stress: activation of protumorigenic tissue phenotypes.

    Science.gov (United States)

    Fordyce, Colleen A; Patten, Kelley T; Fessenden, Tim B; DeFilippis, RosaAnna; Hwang, E Shelley; Zhao, Jianxin; Tlsty, Thea D

    2012-12-07

    damage. The response to cell-extrinsic DNA damage characterized in vitro is recapitulated in vivo in DCIS lesions, which exhibit telomere loss, heightened DNA damage response, and increased activin A and cyclooxygenase-2 expression. These lesions are surrounded by a stroma characterized by increased expression of α smooth muscle actin and endothelial and immune cell infiltration. Thus, synergy between stromal and epithelial interactions, even at the initiating stages of carcinogenesis, appears necessary for the acquisition of malignancy and provides novel insights into where, when, and how the tumor stroma develops, allowing new therapeutic strategies.

  7. A novel closed cell culture device for fabrication of corneal epithelial cell sheets.

    Science.gov (United States)

    Nakajima, Ryota; Kobayashi, Toyoshige; Moriya, Noboru; Mizutani, Manabu; Kan, Kazutoshi; Nozaki, Takayuki; Saitoh, Kazuo; Yamato, Masayuki; Okano, Teruo; Takeda, Shizu

    2015-11-01

    Automation technology for cell sheet-based tissue engineering would need to optimize the cell sheet fabrication process, stabilize cell sheet quality and reduce biological contamination risks. Biological contamination must be avoided in clinical settings. A closed culture system provides a solution for this. In the present study, we developed a closed culture device called a cell cartridge, to be used in a closed cell culture system for fabricating corneal epithelial cell sheets. Rabbit limbal epithelial cells were cultured on the surface of a porous membrane with 3T3 feeder cells, which are separate from the epithelial cells in the cell cartridges and in the cell-culture inserts as a control. To fabricate the stratified cell sheets, five different thicknesses of the membranes which were welded to the cell cartridge, were examined. Multilayered corneal epithelial cell sheets were fabricated in cell cartridges that were welded to a 25 µm-thick gas-permeable membrane, which was similar to the results with the cell-culture inserts. However, stratification of corneal epithelial cell sheets did not occur with cell cartridges that were welded to 100-300 µm-thick gas-permeable membranes. The fabricated cell sheets were evaluated by histological analyses to examine the expression of corneal epithelial-specific markers. Immunohistochemical analyses showed that a putative stem cell marker, p63, a corneal epithelial differentiation maker, CK3, and a barrier function marker, Claudin-1, were expressed in the appropriate position in the cell sheets. These results suggest that the cell cartridge is effective for fabricating corneal epithelial cell sheets.

  8. the genetic and molecular basis of bacterial invasion of epithelial cells

    African Journals Online (AJOL)

    DR. AMINU

    In this review, two of the most important virulence factors of bacterial invasion of epithelial cells. (adhesion and ... interaction such as surface change and hydrophobicity and that fimbrae .... attachment and invasion of cultured epithelial cells;.

  9. File list: ALL.Lng.20.AllAg.Tracheal_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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  12. File list: ALL.Lng.10.AllAg.Tracheal_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Lng.10.AllAg.Tracheal_epithelial_cells hg19 All antigens Lung Tracheal epitheli...barchive.biosciencedbc.jp/kyushu-u/hg19/assembled/ALL.Lng.10.AllAg.Tracheal_epithelial_cells.bed ...

  13. Etk/Bmx activation modulates barrier function in epithelial cells.

    Science.gov (United States)

    Hamm-Alvarez, S F; Chang, A; Wang, Y; Jerdeva, G; Lin, H H; Kim, K J; Ann, D K

    2001-06-01

    Etk/Bmx is a member of the Tec family of cytoplasmic non-receptor tyrosine kinases known to express in epithelial cells. We demonstrate herein that Etk activation in stably Etk-transfected epithelial Pa-4 cells resulted in a consistently increased transepithelial resistance (TER). After 24 h of hypoxic (1% O(2)) exposure, the TER and equivalent active ion transport rate (I(eq)) were reduced to <5% of the normoxia control in Pa-4 cells, whereas both TER and I(eq) were maintained at comparable and 60% levels, respectively, relative to their normoxic controls in cells with Etk activation. Moreover, Pa-4 cells exhibited an abundant actin stress fiber network with a diffuse distribution of beta-catenin at the cell periphery. By contrast, Etk-activated cells displayed a redistribution of actin to an exclusively peripheral network, with a discrete band of beta-catenin also concentrated at the cell periphery, and an altered occludin distribution profile. On the basis of these findings, we propose that Etk may be a novel regulator of epithelial junctions during physiological and pathophysiological conditions.

  14. Biomechanics of epithelial cell islands analyzed by modeling and experimentation

    CERN Document Server

    Coburn, Luke; Noppe, Adrian; Caldwell, Benjamin J; Moussa, Elliott; Yap, Chloe; Priya, Rashmi; Lobaskin, Vladimir; Roberts, Anthony P; Yap, Alpha S; Neufeld, Zoltan; Gomez, Guillermo A

    2016-01-01

    We generated a new computational approach to analyze the biomechanics of epithelial cell islands that combines both vertex and contact-inhibition-of-locomotion models to include both cell-cell and cell-substrate adhesion. Examination of the distribution of cell protrusions (adhesion to the substrate) in the model predicted high order profiles of cell organization that agree with those previously seen experimentally. Cells acquired an asymmetric distribution of protrusions (and traction forces) that decreased when moving from the edge to the island center. Our in silico analysis also showed that tension on cell-cell junctions (and monolayer stress) is not homogeneous across the island. Instead it is higher at the island center and scales up with island size, which we confirmed experimentally using laser ablation assays and immunofluorescence. Moreover, our approach has the minimal elements necessary to reproduce mechanical crosstalk between both cell-cell and cell substrate adhesion systems. We found that an i...

  15. Cytotoxic Effect of Lipophilic Bismuth Dimercaptopropanol Nanoparticles on Epithelial Cells.

    Science.gov (United States)

    Rene, Hernandez-Delgadillo; Badireddy, Appala Raju; José, Martínez-Sanmiguel Juan; Francisco, Contreras-Cordero Juan; Israel, Martinez-Gonzalez Gustavo; Isela, Sánchez-Nájera Rosa; Chellam, Shankararaman; Claudio, Cabral-Romero

    2016-01-01

    Bismuth nanoparticles have many interesting properties to be applied in biomedical and medicinal sectors, however their safety in humans have not been comprehensively investigated. The objective of this research was to determine the cytotoxic effect of bismuth dimercaptopropanol nanoparticles (BisBAL NPs) on epithelial cells. The nanoparticles are composed of 18.7 nm crystallites on average and have a rhombohedral structure, agglomerating into chains-like or clusters of small nanoparticles. Based on MTT viability assay and fluorescence microscopy, cytotoxicity was not observed on monkey kidney cells after growing with 5 µM of BisBAL NPs for 24 h. Employing same techniques, identical results were obtained with human epithelial cells (HeLa), showing a not strain-dependent phenomenon. The absence of toxic effects on epithelial cells growing with BisBAL NPs was corroborated with long-time experiments (24-72 hrs.), showing no difference in comparison with growing control (cells without nanoparticles). Further, genotoxicity assays, comet assay and fluorescent microscopy and electrophoresis in bromide-stained agarose gel revealed no damage to genomic DNA of MA104 cells after 24 h. of exposition to BisBAL NPs. Finally, the effect of bismuth nanoparticles on protein synthesis was studied in cells growing with BisBAL NPs for 24 h. SDS-PAGE assays showed no difference between treated and untreated cells, suggesting that BisBAL NPs did not interfere with protein synthesis. Hence BisBAL NPs do not appear to exert cytotoxic effects suggesting their biological compatibility with epithelial cells.

  16. 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.

  17. Uranium induces oxidative stress in lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Periyakaruppan, Adaikkappan; Kumar, Felix; Sarkar, Shubhashish; Sharma, Chidananda S.; Ramesh, Govindarajan T. [Texas Southern University, Molecular Neurotoxicology Laboratory/Proteomics Core, Department of Biology, Houston, TX (United States)

    2007-06-15

    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.)

  18. Expression and roles of CCN2 in dental epithelial cells.

    Science.gov (United States)

    Shimo, Tsuyoshi; Koyama, Eiki; Kurio, Naito; Matsumoto, Kenichi; Okui, Tatsuo; Ibaragi, Soichiro; Yoshioka, Norie; Sasaki, Akira

    2015-01-01

    Connective tissue growth factor (CCN2) regulates diverse cellular functions, including tooth development. In order to delineate the precise role of CCN2 in the epithelium during odontogenesis, we investigated how it is expressed and what roles it may have in primary cultures of epithelial cells derived from developing tooth germ of the bovine fetus. Ccn2 mRNA and protein were strongly expressed in the inner dental epithelium, which is consistent with the expression of transforming growth factor-β2 mRNA and proliferating cell nuclear antigen. Bone morphogenetic protein 4 (BMP4) and fibroblast growth factor 2 (FGF2) were also expressed in the inner dental epithelium, indicating that CCN2 functionally interacts with these factors in the epithelium. The stimulatory effects of FGF2 on cell proliferation and BMP4 on cell differentiation were additively up-regulated by CCN2 in a newly-established dental epithelium cell culture. Taken together, our data provide clear evidence that CCN2 is synthesized by inner dental epithelial cells, and appears to act as an autocrine factor, which regulates dental epithelial cell proliferation and differentiation in concert with growth factors. Copyright © 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  19. Proteomics demonstration that normal breast epithelial cells can induce apoptosis of breast cancer cells through insulin-like growth factor-binding protein-3 and maspin.

    Science.gov (United States)

    Toillon, Robert-Alain; Lagadec, Chann; Page, Adeline; Chopin, Valérie; Sautière, Pierre-Eric; Ricort, Jean-Marc; Lemoine, Jérôme; Zhang, Ming; Hondermarck, Hubert; Le Bourhis, Xuefen

    2007-07-01

    Normal breast epithelial cells are known to exert an apoptotic effect on breast cancer cells, resulting in a potential paracrine inhibition of breast tumor development. In this study we purified and characterized the apoptosis-inducing factors secreted by normal breast epithelial cells. Conditioned medium was concentrated by ultrafiltration and separated on reverse phase Sep-Pak C18 and HPLC. The proapoptotic activity of eluted fractions was tested on MCF-7 breast cancer cells, and nano-LC-nano-ESI-MS/MS allowed the identification of insulin-like growth factor-binding protein-3 (IGFBP-3) and maspin as the proapoptotic factors produced by normal breast epithelial cells. Western blot analysis of conditioned media confirmed the specific secretion of IGFBP-3 and maspin by normal cells but not by breast cancer cells. Immunodepletion of IGFBP-3 and maspin completely abolished the normal cell-induced apoptosis of cancer cells, and recombinant proteins reproduced the effect of normal cell-conditioned medium on apoptosis of breast cancer cells. Together our results indicated that normal breast epithelial cells can induce apoptosis of breast cancer cells through IGFBP-3 and maspin. These findings provide a molecular hypothesis for the long observed inhibitory effect of normal surrounding cells on breast cancer development.

  20. Directed differentiation of airway epithelial cells of human bone marrow mesenchymal stem cells.

    Science.gov (United States)

    Li, Jian-Dong

    2016-11-01

    The ability to generate lung and airway epithelial cells from human bone marrow mesenchymal stem cells (hBMSCs) would have applications in regenerative medicine, modeling of lung disease, drug screening, and studies of human lung development. In this research, hBMSCs were cultured in specialized airway epithelial cell growth media for differentiation of airway epithelial cells, including keratinocyte growth factor transferrin, bovine pituitary extract, epinephrine, triiodothyronine and retinoic acid. The surfactant protein C, a specific marker of type II pneumocytes, and its corresponding protein were demonstrated by immunofluorescence and western blotting after differentiation of airway epithelial cells, respectively. These cells were then transferred into an induced acute lung injury model. The results showed that the hBMSCs could induce differentiation in airway epithelial cells under the special conditions of the medium, the result for surfactant protein C was positive in differentiated airway epithelial cells using immunofluorescence and western blotting, and these cells were successfully colonized in the injured lung airway. In conclusion, our research shows that a population of airway epithelial cells can be specifically generated from hBMSCs and that induced cells may be allowed to participate in tissue repair.

  1. File list: DNS.Lng.20.AllAg.Tracheal_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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  7. File list: His.Brs.20.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  8. Epstein-Barr virus infection in ex vivo tonsil epithelial cell cultures of asymptomatic carriers.

    NARCIS (Netherlands)

    Pegtel, DM; Middeldorp, J.M.; Thorley-Lawson, DA

    2004-01-01

    Epstein-Barr virus (EBV) is found frequently in certain epithelial pathologies, such as nasopharyngeal carcinoma and oral hairy leukoplakia, indicating that the virus can infect epithelial cells in vivo. Recent studies of cell lines imply that epithelial cells may also play a role in persistent EBV

  9. File list: DNS.Brs.20.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  10. File list: Unc.Brs.05.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  11. File list: DNS.Brs.05.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  12. File list: Unc.Lng.20.AllAg.Tracheal_epithelial_cells [Chip-atlas[Archive

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  13. File list: Pol.Lng.05.AllAg.Tracheal_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  14. File list: Oth.Lng.10.AllAg.Tracheal_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

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    Lifescience Database Archive (English)

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  7. File list: His.Brs.05.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

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  12. File list: Oth.Brs.10.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

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  13. File list: Oth.Lng.20.AllAg.Tracheal_epithelial_cells [Chip-atlas[Archive

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  1. File list: Pol.Brs.20.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

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  2. Limbal Stromal Tissue Specific Stem Cells and Their Differentiation Potential to Corneal Epithelial Cells.

    Science.gov (United States)

    Katikireddy, Kishore Reddy; Jurkunas, Ula V

    2016-01-01

    From the derivation of the first human embryonic stem (hES) cell line to the development of induced pluripotent stem (iPS) cells; it has become evident that tissue specific stem cells are able to differentiate into a specific somatic cell types. The understanding of key processes such as the signaling pathways and the role of the microenvironment in epidermal/epithelial development has provided important clues for the derivation of specific epithelial cell types.Various differentiation protocols/methods were used to attain specific epithelial cell types. Here, we describe in detail the procedure to follow for isolation of tissue specific stem cells, mimicking their microenvironment to attain stem cell characteristics, and their potential differentiation to corneal epithelial cells.

  3. The PCP pathway regulates Baz planar distribution in epithelial cells

    OpenAIRE

    2016-01-01

    The localisation of apico-basal polarity proteins along the Z-axis of epithelial cells is well understood while their distribution in the plane of the epithelium is poorly characterised. Here we provide a systematic description of the planar localisation of apico-basal polarity proteins in the Drosophila ommatidial epithelium. We show that the adherens junction proteins Shotgun and Armadillo, as well as the baso-lateral complexes, are bilateral, i.e. present on both sides of cell interfaces. ...

  4. Established thymic epithelial progenitor/stem cell-like cell lines differentiate into mature thymic epithelial cells and support T cell development.

    Directory of Open Access Journals (Sweden)

    Pengfei Chen

    Full Text Available Common thymic epithelial progenitor/stem cells (TEPCs differentiate into cortical and medullary thymic epithelial cells (TECs, which are required for the development and selection of thymocytes. Mature TEC lines have been widely established. However, the establishment of TEPC lines is rarely reported. Here we describe the establishment of thymic epithelial stomal cell lines, named TSCs, from fetal thymus. TSCs express some of the markers present on tissue progenitor/stem cells such as Sca-1. Gene expression profiling verifies the thymic identity of TSCs. RANK stimulation of these cells induces expression of autoimmune regulator (Aire and Aire-dependent tissue-restricted antigens (TRAs in TSCs in vitro. TSCs could be differentiated into medullary thymic epithelial cell-like cells with exogenously expressed NF-κB subunits RelB and p52. Importantly, upon transplantation under the kidney capsules of nude mice, TSCs are able to differentiate into mature TEC-like cells that can support some limited development of T cells in vivo. These findings suggest that the TSC lines we established bear some characteristics of TEPC cells and are able to differentiate into functional TEC-like cells in vitro and in vivo. The cloned TEPC-like cell lines may provide useful tools to study the differentiation of mature TEC cells from precursors.

  5. Acrolein stimulates eicosanoid release from bovine airway epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Doupnik, C.A.; Leikauf, G.D. (Univ. of Cincinnati College of Medicine, OH (USA))

    1990-10-01

    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.

  6. In vitro methods to culture primary human breast epithelial cells.

    Science.gov (United States)

    Raouf, Afshin; Sun, Yu Jia

    2013-01-01

    Current evidence suggests that much like leukemia, breast tumors are maintained by a small subpopulation of tumor cells that have stem cell properties. These cancer stem cells are envisaged to be responsible for tumor formation and relapse. Therefore, knowledge about their nature will provide a platform to develop therapies to eliminate these breast cancer stem cells. This concept highlights the need to understand the mechanisms that regulate the normal functions of the breast stem cells and their immediate progeny as alterations to these same mechanisms can cause these primitive cells to act as cancer stem cells. The study of the primitive cell functions relies on the ability to isolate them from primary sources of breast tissue. This chapter describes processing of discarded tissue from reduction mammoplasty samples as sources of normal primary human breast epithelial cells and describes cell culture systems to grow single-cell suspensions prepared from these reduction samples in vitro.

  7. Estradiol increases mucus synthesis in bronchial epithelial cells.

    Directory of Open Access Journals (Sweden)

    Anthony Tam

    Full Text Available Airway epithelial mucus hypersecretion and mucus plugging are prominent pathologic features of chronic inflammatory conditions of the airway (e.g. asthma and cystic fibrosis and in most of these conditions, women have worse prognosis compared with male patients. We thus investigated the effects of estradiol on mucus expression in primary normal human bronchial epithelial cells from female donors grown at an air liquid interface (ALI. Treatment with estradiol in physiological ranges for 2 weeks caused a concentration-dependent increase in the number of PAS-positive cells (confirmed to be goblet cells by MUC5AC immunostaining in ALI cultures, and this action was attenuated by estrogen receptor beta (ER-β antagonist. Protein microarray data showed that nuclear factor of activated T-cell (NFAT in the nuclear fraction of NHBE cells was increased with estradiol treatment. Estradiol increased NFATc1 mRNA and protein in ALI cultures. In a human airway epithelial (1HAE0 cell line, NFATc1 was required for the regulation of MUC5AC mRNA and protein. Estradiol also induced post-translational modification of mucins by increasing total fucose residues and fucosyltransferase (FUT-4, -5, -6 mRNA expression. Together, these data indicate a novel mechanism by which estradiol increases mucus synthesis in the human bronchial epithelium.

  8. Alveolar epithelial type II cell: defender of the alveolus revisited

    Directory of Open Access Journals (Sweden)

    Fehrenbach Heinz

    2001-01-01

    Full Text Available Abstract In 1977, Mason and Williams developed the concept of the alveolar epithelial type II (AE2 cell as a defender of the alveolus. It is well known that AE2 cells synthesise, secrete, and recycle all components of the surfactant that regulates alveolar surface tension in mammalian lungs. AE2 cells influence extracellular surfactant transformation by regulating, for example, pH and [Ca2+] of the hypophase. AE2 cells play various roles in alveolar fluid balance, coagulation/fibrinolysis, and host defence. AE2 cells proliferate, differentiate into AE1 cells, and remove apoptotic AE2 cells by phagocytosis, thus contributing to epithelial repair. AE2 cells may act as immunoregulatory cells. AE2 cells interact with resident and mobile cells, either directly by membrane contact or indirectly via cytokines/growth factors and their receptors, thus representing an integrative unit within the alveolus. Although most data support the concept, the controversy about the character of hyperplastic AE2 cells, reported to synthesise profibrotic factors, proscribes drawing a definite conclusion today.

  9. Isolation and biological characterization of chicken amnion epithelial cells

    Directory of Open Access Journals (Sweden)

    Y. Gao

    2012-08-01

    Full Text Available Amniotic epithelial cells (AECs express Oct4, Nanog and Sox-2, which are necessary for maintaining the undifferentiated state of pluripotent stem cells. AECs additionally express CK19, which is a specific marker of epithelial cells, both in vivo and in vitro. In this research, we investigated the biological characteristics and potential for cell therapy of AECs from 6-day-old chicken embryos. We induced the AECs to differentiate into pancreatic islet-like cells (endoderm, adipocytes and osteoblasts (mesoderm and neural-like cells (ectoderm, and used immunofluorescence and RT-PCR to detect the expression of AECs specific markers. To assess the differentiation capacity of AECs, passage 3 cells were induced to differentiate into adipocytes, osteoblasts, pancreatic islet-like cells and neural-like cells. The AEC markers, Oct4, Nanog, Sox-2 and CK19, were all positively expressed. Cloning efficiency decreased with increasing passage number. Passage 3 AECs were successfully induced to differentiate into pancreatic islet-like cells, osteoblasts, adipocytes, and neural-like cells. These results suggested that AECs isolated from chicken embryos exhibited the characteristics of the multipotent stem cells. AECs may therefore be ideal candidates for cellular transplantation therapy and tissue engineering.

  10. Azithromycin Inhibits Mucus Hypersecretion from Airway Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Takeshi Shimizu

    2012-01-01

    Full Text Available To examine the in vivo effects of the 15-member macrolide, azithromycin (AZM, on mucus hypersecretion, we induced hypertrophic and metaplastic changes of goblet cells in rat nasal epithelium by intranasal instillation of ovalbumin (OVA in OVA-sensitized rats, or by intranasal lipopolysaccharides (LPS instillation. Oral administration of AZM (5–10 mg/kg or clarithromycin (CAM, 5–10 mg/kg significantly inhibited OVA- and LPS-induced mucus production, whereas josamycin (JM or ampicillin (ABPC showed no effect. In vitro effects of AZM on airway epithelial cells were examined using NCI-H292 cells and human nasal epithelial cells cultured in air-liquid interface. Mucus secretion was evaluated by enzyme-linked immunosorbent assay using an anti-MUC5AC monoclonal antibody. AZM or CAM significantly inhibited tumor necrosis factor-α (TNF-α (20 ng/mL-induced MUC5AC secretion from NCI-H292 cells at 10−6–10−7 M, whereas JM or ABPC showed no effect. AZM significantly inhibited TNF- (20 ng/mL-induced MUC5AC secretion from human nasal epithelial cells at 10−4 M. MUC5AC mRNA expression was also significantly inhibited. These results indicate that the 15-member macrolide, AZM, exerts direct inhibitory effects on mucus secretion from airway epithelial cells and that it may be useful for the treatment of mucus hypersecretion caused by allergic inflammation and LPS stimulation.

  11. 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...

  12. 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.

  13. 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.

  14. Epithelial cell extrusion leads to breaches in the intestinal epithelium.

    Science.gov (United States)

    Liu, Julia J; Davis, Elisabeth M; Wine, Eytan; Lou, Yuefei; Rudzinski, Jan K; Alipour, Misagh; Boulanger, Pierre; Thiesen, Aducio L; Sergi, Consolato; Fedorak, Richard N; Muruve, Daniel; Madsen, Karen L; Irvin, Randall T

    2013-04-01

    Two distinct forms of intestinal epithelial cell (IEC) extrusion are described: 1 with preserved epithelial integrity and 1 that introduced breaches in the epithelial lining. In this study, we sought to determine the mechanism underlying the IEC extrusion that alters the permeability of the gut epithelium. IEC extrusions in polarized T84 monolayer were induced with nigericin. Epithelial permeability was assessed with transepithelial electrical resistance and movements of latex microspheres and green fluorescent protein-transfected Escherichia coli across the monolayer. In vivo IEC extrusion was modulated in wild-type and a colitic (interleukin-10 knock-out) mouse model with caspase-1 activation and inhibition. Luminal aspirates and mucosal biopsies from control patients and patients with inflammatory bowel disease were analyzed for caspase-1 and caspase-3&7 activation. Caspase-1-induced IEC extrusion in T84 monolayers resulted in dose-dependent and time-dependent barrier dysfunction, reversible with caspase-1 inhibition. Moreover, the movements of microspheres and microbes across the treated epithelial monolayers were observed. Increased caspase-1-mediated IEC extrusion in interleukin-10 knock-out mice corresponded to enhanced permeation of dextran, microspheres, and translocation of E. coli compared with wild type. Caspase-1 inhibition in interleukin-10 knock-out mice resulted in a time-dependent reduction in cell extrusion and normalization of permeability to microspheres. Increased IEC extrusion in wild-type mice was induced with caspase-1 activation. In human luminal aspirates, the ratio of positively stained caspase-1 to caspase-3&7 cells were 1:1 and 2:1 in control patients and patients with inflammatory bowel disease, respectively; these observations were confirmed by cytochemical analysis of mucosal biopsies. IEC extrusion mediated by caspase-1 activation contributes to altered intestinal permeability in vitro and in vivo.

  15. Asbestos exposure increases human bronchial epithelial cell fibrinolytic activity.

    Science.gov (United States)

    Gross, T J; Cobb, S M; Gruenert, D C; Peterson, M W

    1993-03-01

    Chronic exposure to asbestos fibers results in fibrotic lung disease. The distal pulmonary epithelium is an early target of asbestos-mediated injury. Local plasmin activity may be important in modulating endoluminal inflammatory responses in the lung. We studied the effects of asbestos exposure on cell-mediated plasma clot lysis as a marker of pericellular plasminogen activation. Exposing human bronchial epithelial (HBE) cells to 100 micrograms/ml of asbestos fibers for 24 h resulted in increased plasma clot lysis. Fibrinolytic activity was augmented in a dose-dependent fashion, was not due to secreted protease, and occurred only when there was direct contact between the plasma clot and the epithelial monolayer. Further analysis showed that asbestos exposure increased HBE cell-associated urokinase-type plasminogen activator (uPA) activity in a time-dependent manner. The increased cell-associated PA activity could be removed by acid washing. The increase in PA activity following asbestos exposure required new protein synthesis because it was abrogated by treatment with either cycloheximide or actinomycin D. Therefore, asbestos exposure increases epithelial-mediated fibrinolysis by augmenting expression of uPA activity at the cell surface by mechanisms that require new RNA and protein synthesis. These observations suggest a novel mechanism whereby exposure of the distal epithelium to inhaled particulates may result in a chronic inflammatory response that culminates in the development of fibrotic lung disease.

  16. Expression of inducible nitric oxide in human lung epithelial cells.

    Science.gov (United States)

    Robbins, R A; Barnes, P J; Springall, D R; Warren, J B; Kwon, O J; Buttery, L D; Wilson, A J; Geller, D A; Polak, J M

    1994-08-30

    Nitric oxide (NO) is increased in the exhaled air of subjects with several airway disorders. To determine if cytokines could stimulate epithelial cells accounting for the increased NO, the capacity of the proinflammatory cytokines (cytomix: tumor necrosis factor-alpha, interleukin-1 beta, and interferon-gamma) to increase inducible nitric oxide synthase (iNOS) was investigated in A549 and primary cultures of human bronchial epithelial cells. Cytomix induced a time-dependent increase in nitrite levels in culture supernatant fluids (p < 0.05). Increased numbers of cells stained for iNOS and increased iNOS mRNA was detected in the cytokine-stimulated cells compared to control (p < 0.05). Dexamethasone diminished the cytokine-induced increase in nitrite, iNOS by immunocytochemistry, and iNOS mRNA. These data demonstrate that cytokines, such as those released by mononuclear cells, can induce lung epithelial iNOS expression and NO release, and that this is attenuated by dexamethasone.

  17. Expression of Connexin43 in Rat Epithelial Cells and Fibroblasts

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    To explore the role of connexin43 (Cx43) in gap junctional intercellular communication (GJIC) and propagated sensation along meridians, the expression of Cx43 in the rat epithelial cells and fibroblasts was studied both in vitro and in vivo. With the in vitro study, the rat epithelial cells and fibroblasts were cultured together, and the localization of Cx43 was detected by immunohistochemistry and indirect immunofluorescent cytochemistry and under confocal microscopy . And the expression of Cx43 on the surface of the cells was examined by flow cytometry. With the in vivo examination, 20 SD rats were randomized into control group (n = 10) and electrical acupuncture group (EAgroup, n=10). EA ( 0.5-1.5 V, 4-16 Hz , 30 min) was applied to"Zusanli"acupoint for 30 min at rat's hind paw, the localization of Cx43 was immunohistochemically detected.The immunohistochemical staining and indirect immunfluorescent cytochemistry showed that Cx43was localized on the surface of the cells and in the cytoplasm. The relative expression level of Cx43on the cellular membrane surfaces of the rat epithelial cells and fibroblasts, as determined by FACS, were 13.91 % and 29.53 % respectively. Our studied suggested that Cx43 might be involved in GJIC and propagated sensation along meridians.

  18. Radiogenic transformation of human mammary epithelial cells in vitro

    Science.gov (United States)

    Yang, T. C.; Georgy, K. A.; Tavakoli, A.; Craise, L. M.; Durante, M.

    1996-01-01

    Cancer induction by space radiations is a major concern for manned space exploration. Accurate assessment of radiation risk at low doses requires basic understanding of mechanism(s) of radiation carcinogenesis. For determining the oncogenic effects of ionizing radiation in human epithelial cells, we transformed a mammary epithelial cell line (185B5), which was immortalized by benzo(a)pyrene, with energetic heavy ions and obtained several transformed clones. These transformed cells showed growth properties on Matrigel similar to human mammary tumor cells. To better understand the mechanisms of radiogenic transformation of human cells, we systematically examined the alterations in chromosomes and cancer genes. Among 16 autosomes examined for translocations, by using fluorescence in situ hybridization (FISH) technique, chromosomes 3, 12, 13, 15, 16, and 18 appeared to be normal in transformed cells. Chromosomes 1, 4, 6, 8, and 17 in transformed cells, however, showed patterns different from those in nontransformed cells. Southern blot analyses indicated no detectable alterations in myc, ras, Rb, or p53 genes. Further studies of chromosome 17 by using in situ hybridization with unique sequence p53 gene probe and a centromere probe showed no loss of p53 gene in transformed cells. Experimental results from cell fusion studies indicated that the transforming gene(s) is recessive. The role of genomic instability and tumor suppressor gene(s) in radiogenic transformation of human breast cells remains to be identified.

  19. Generation of Spheres from Dental Epithelial Stem Cells

    Science.gov (United States)

    Natsiou, Despoina; Granchi, Zoraide; Mitsiadis, Thimios A.; Jimenez-Rojo, Lucia

    2017-01-01

    The in vitro three-dimensional sphere model has already been established as an important tool in fundamental sciences. This model facilitates the study of a variety of biological processes including stem cell/niche functions and tissue responses to injury and drugs. Here we describe the complete protocol for the in vitro formation of spheres originated from the epithelium of rodent incisors. In addition, we show that in these spheres cell proliferation is maintained, as well as the expression of several key molecules characterizing stem cells such as Sox2 and p63. These epithelial dentospheres could be used as an in vitro model system for stem cell research purposes. PMID:28154538

  20. Fusion of intestinal epithelial cells with bone marrow derived cells is dispensable for tissue homeostasis.

    Science.gov (United States)

    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 with the epithelial lineage. However, the functional relevance of these observations is unknown. In the present study we employ a model system in which we cannot only detect cell fusion but also examine the functional importance of this process in vivo. Our results indicate that fusion between BMDCs and intestinal epithelial cells is an extremely rare event under physiological conditions. More importantly, by employing a system in which fusion-derived cells can be specifically deleted after extensive tissue damage, we present evidence that cell fusion is not relevant for tissue regeneration. Our data decisively demonstrates that intestinal epithelial homeostasis and regeneration is not dependent on cell fusion involving BMDCs.

  1. Irsogladine maleate regulates gap junctional intercellular communication-dependent epithelial barrier in human nasal epithelial cells.

    Science.gov (United States)

    Miyata, Ryo; Nomura, Kazuaki; Kakuki, Takuya; Takano, Ken-Ichi; Kohno, Takayuki; Konno, Takumi; Sawada, Norimasa; Himi, Tetsuo; Kojima, Takashi

    2015-04-01

    The airway epithelium of the human nasal mucosa acts as the first physical barrier that protects against inhaled substances and pathogens. Irsogladine maleate (IM) is an enhancer of gastric mucosal protective factors via upregulation of gap junctional intercellular communication (GJIC). GJIC is thought to participate in the formation of functional tight junctions. However, the effects of IM on GJIC and the epithelial barrier in human nasal epithelial cells (HNECs) remain unknown. To investigate the effects of IM on GJIC and the tight junctional barrier in HNECs, primary cultures of HNECs transfected with human telomerase reverse transcriptase (hTERT-HNECs) were treated with IM and the GJIC inhibitors oleamide and 18β-GA. Some cells were pretreated with IM before treatment with TLR3 ligand poly(I:C) to examine whether IM prevented the changes via TLR3-mediated signal pathways. In hTERT-HNECs, GJIC blockers reduced the expression of tight junction molecules claudin-1, -4, -7, occludin, tricellulin, and JAM-A. IM induced GJIC activity and enhanced the expression of claudin-1, -4, and JAM-A at the protein and mRNA levels with an increase of barrier function. GJIC blockers prevented the increase of the tight junction proteins induced by IM. Furthermore, IM prevented the reduction of JAM-A but not induction of IL-8 and TNF-α induced by poly(I:C). In conclusion, IM can maintain the GJIC-dependent tight junctional barrier via regulation of GJIC in upper airway nasal epithelium. Therefore, it is possible that IM may be useful as a nasal spray to prevent the disruption of the epithelial barrier by viral infections and exposure to allergens in human nasal mucosa.

  2. Iodide handling by the thyroid epithelial cell.

    Science.gov (United States)

    Nilsson, M

    2001-01-01

    Iodination of thyroglobulin, the key event in the synthesis of thyroid hormone, is an extracellular process that takes place inside the thyroid follicles at the apical membrane surface that faces the follicular lumen. The supply of iodide involves two steps of TSH-regulated transport, basolateral uptake and apical efflux, that imprint the polarized phenotype of the thyroid cell. Iodide uptake is generated by the sodium/iodide symporter present in the basolateral plasma membrane. A candidate for the apical iodide-permeating mechanism is pendrin, a chloride/iodide transporting protein recently identified in the apical membrane. In physiological conditions, transepithelial iodide transport occurs without intracellular iodination, despite the presence of large amounts of thyroglobulin and thyroperoxidase inside the cells. The reason is that hydrogen peroxide, serving as electron acceptor in iodide-protein binding and normally produced at the apical cell surface, is rapidly degraded by cytosolic glutathione peroxidase once it enters the cells. Iodinated thyroglobulin in the lumen stores not only thyroid hormone but iodine incorporated in iodotyrosine residues as well. After endocytic uptake and degradation of thyroglobulin, intracellular deiodination provides a mechanism for recycling of iodide to participate in the synthesis of new thyroid hormone at the apical cell surface.

  3. 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.

  4. Molecular cloning, sequence analysis, and function of the intestinal epithelial stem cell marker Bmi1 in pig intestinal epithelial cells.

    Science.gov (United States)

    Li, C-M; Yan, H-C; Fu, H-L; Xu, G-F; Wang, X-Q

    2014-01-01

    In the present work, we cloned the full-length cDNA of the pig Bmi1 gene (BMI1 polycomb ring finger oncogene), which has been indicated as an intestinal epithelial stem cell (IESC) marker in other mammals. This paper provides the first report of the function of Bmi1 in pig intestinal epithelial cells and a brief description of its underlying mechanism. Rapid amplification of cDNA ends technology was used to clone the complete pig Bmi1 sequence, and a Bmi1-pcDNA3.1 vector was constructed for transfection into an intestinal porcine epithelial cell line (IPEC-1). The proliferation ability of the cells was estimated using the MTT assay and the EdU incorporation method at different time points after seeding. Cell cycle information was detected by flow cytometry. The mRNA abundances of cell cycle-related genes were also measured. The results indicated that the pig Bmi1 cDNA is 3,193 bp in length and consists of a 981 bp open reading frame, a 256 bp 5´ untranslated region (UTR), and a 1,956 bp 3' UTR. The transcript contains no signal peptides, and there are no transmembrane regions in the pig Bmi1 coded protein, which has a total of 326 AA. The overexpression of the pig Bmi1 in the IPEC-1 cells led to increased cell proliferation and a lower percentage of cells in the G1 and S phases (P cells in the G2 phase (P 0.05). Our data suggested that pig Bmi1 can increase the proliferation of IPEC-1 cells by promoting the G1/S transition and the overall cell cycle process.

  5. 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.

  6. Oral epithelial cell responses to multispecies microbial biofilms.

    Science.gov (United States)

    Peyyala, R; Kirakodu, S S; Novak, K F; Ebersole, J L

    2013-03-01

    This report describes the use of a novel model of multispecies biofilms to stimulate profiles of cytokines/chemokines from oral epithelial cells that contribute to local inflammation in the periodontium. Streptococcus gordonii (Sg)/S. oralis (So)/S. sanguinis (Ss) and Sg/Fusobacterium nucleatum (Fn)/Porphyromonas gingivalis (Pg) biofilms elicited significantly elevated levels of IL-1α and showed synergistic stimulatory activity compared with an additive effect of the 3 individual bacteria. Only the Sg/Actinomyces naeslundii (An)/Fn multispecies biofilms elicited IL-6 levels above those of control. IL-8 was a primary response to the Sg/An/Fn biofilms, albeit the level was not enhanced compared with a predicted composite level from the monospecies challenges. These results represent some of the first data documenting alterations in profiles of oral epithelial cell responses to multispecies biofilms.

  7. Evaluating alternative stem cell hypotheses for adultcorneal epithelial maintenance

    Institute of Scientific and Technical Information of China (English)

    John D West; Natalie J Dorà; Natalie J Dorà,

    2015-01-01

    In this review we evaluate evidence for three differenthypotheses that explain how the corneal epitheliumis maintained. The limbal epithelial stem cell (LESC)hypothesis is most widely accepted. This proposes thatstem cells in the basal layer of the limbal epithelium,at the periphery of the cornea, maintain themselvesand also produce transient (or transit) amplifying cells(TACs). TACs then move centripetally to the centre ofthe cornea in the basal layer of the corneal epitheliumand also replenish cells in the overlying suprabasallayers. The LESCs maintain the corneal epitheliumduring normal homeostasis and become more active torepair significant wounds. Second, the corneal epithelialstem cell (CESC) hypothesis postulates that, duringnormal homeostasis, stem cells distributed throughoutthe basal corneal epithelium, maintain the tissue.According to this hypothesis, LESCs are present in thelimbus but are only active during wound healing. We alsoconsider a third possibility, that the corneal epithelium ismaintained during normal homeostasis by proliferationof basal corneal epithelial cells without any input fromstem cells. After reviewing the published evidence,we conclude that the LESC and CESC hypotheses areconsistent with more of the evidence than the thirdhypothesis, so we do not consider this further. The LESCand CESC hypotheses each have difficulty accountingfor one main type of evidence so we evaluate the twokey lines of evidence that discriminate between them.Finally, we discuss how lineage-tracing experimentshave begun to resolve the debate in favour of theLESC hypothesis. Nevertheless, it also seems likely thatsome basal corneal epithelial cells can act as long-termprogenitors if limbal stem cell function is compromised.Thus, this aspect of the CESC hypothesis may have alasting impact on our understanding of corneal epithelialmaintenance, even if it is eventually shown that stemcells are restricted to the limbus as proposed by the

  8. 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

    2017-01-01

    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

  9. Advances in Classification and Research Methods of Lung Epithelial Stem 
and Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Minhua DENG

    2017-02-01

    Full Text Available Isolation and characterization of lung epithelial stem and progenitor cells and understanding of their specific role in lung physiopathology are critical for preventing and controlling lung diseases including lung cancer. In this review, we summarized recent advances in classification and research methods of lung epithelial stem and progenitor cells. Lung epithelial stem and progenitor cells were region-specific, which primarily included basal cells and duct cells in proximal airway, Clara cells, variant Clara cells, bronchioalveolar stem cells and induced krt5+ cells in bronchioles, type II alveolar cells and type II alveolar progenitor cells in alveoli. The research methods of lung epithelial stem and progenitor cells were mainly focused on lung injury models, lineage-tracing experiments, three dimensional culture, transplantation, chronic labeled cells and single-cell transcriptome analysis. Lastly, the potential relationship between lung epithelial stem and progenitor cells and lung cancer as well as lung cancer stem cell-targeted drug development were briefly reviewed.

  10. Continuous cytokine exposure of colonic epithelial cells induces DNA damage

    DEFF Research Database (Denmark)

    Seidelin, Jakob B; Nielsen, Ole Haagen

    2005-01-01

    Chronic inflammatory diseases of the intestinal tract are associated with an increased risk of colorectal cancer. As an example ulcerative colitis (UC) is associated with a production of reactive oxygen species (ROS), including nitrogen monoxide (NO), which is produced in high amounts by inducibl...... nitrogen oxide synthase (iNOS). NO as well as other ROS are potential DNA damaging agents. The aim was to determine the effect of long-term cytokine exposure on NO formation and DNA damage in epithelial cells....

  11. Structure and function of the interphotoreceptor matrix surrounding retinal photoreceptor cells.

    Science.gov (United States)

    Ishikawa, Makoto; Sawada, Yu; Yoshitomi, Takeshi

    2015-04-01

    The interphotoreceptor matrix (IPM) is a highly organized structure with interconnected domains surrounding cone and rod photoreceptor cells and extends throughout the subretinal space. Based on known roles of the extracellular matrix in other tissues, the IPM is thought to have several prominent functions including serving as a receptor for growth factors, regulating retinoid transport, participating in cytoskeletal organization in surrounding cells, and regulation of oxygen and nutrient transport. In addition, a number of studies suggest that the IPM also may play a significant role in the etiology of retinal degenerative disorders. In this review, we describe the present knowledge concerning the structure and function of the IPM under physiological and pathological conditions. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  12. Serum-Free Cryopreservation of Human Amniotic Epithelial Cells

    Directory of Open Access Journals (Sweden)

    H. Niknejad

    2013-04-01

    Full Text Available Introduction & Objective: One of the important issues in long term storage of cells is removal of animal serum from cell culture environments. The aim of this study was to evaluate amni-otic fluid (AF, which is full of growth factors, as substitute for fetal bovine serum (FBS in the cryopreservation protocol. Materials & Methods: In this experimental study human amniotic epithelial cells were isolated from placentas which were seronegative for microbial infections. The cells were preserved in 24 different patterns for 12 months in -196 ?C (liquid nitrogen and viability of cells were determined before and after cryopreservation by trypan blue and MTT assay. Moreover, Oct-4 expression was studied to determine pluripotency before and after cryopreservation with immunocytochemistry. Results were compared between groups with ANOVA (Tukey Post-Test. P.value under 0.01 and 0.05 was considered statistically significant. Results: The presence of DMEM, FBS or AF is necessary for amniotic cell cryopreservation. Trypan-blue, MTT and immunocytochemistry showed that there isn’t significant difference between using AF and FBS in viability and pluripotency of cells. Moreover, results showed that DMSO is a better cryoprotectant compared to glycerol. Conclusion : Results showed that amniotic fluid can be a proper substitute for FBS in amniotic epithelial cells cryopreservation. (Sci J Hamadan Univ Med Sci 2013; 20 (1:15-24

  13. Notch as a Driver of Gastric Epithelial Cell Proliferation.

    Science.gov (United States)

    Demitrack, Elise S; Samuelson, Linda C

    2017-05-01

    The gastric epithelium is sustained by a population of stem cells that replenish the various mature epithelial lineages throughout adulthood. Regulation of stem and progenitor cell proliferation occurs via basic developmental signaling pathways, including the Notch pathway, which recently was described to promote gastric stem cell proliferation in both mice and human beings. Current cancer theory proposes that adult stem cells that maintain gastrointestinal tissues accumulate mutations that promote cancerous growth, and that basic signaling pathways, such as Notch, which stimulate stem cell proliferation, can promote tumorigenesis. Accordingly, constitutive Notch activation leads to unchecked cellular proliferation and gastric tumors in genetic mouse models. Furthermore, there is emerging evidence suggesting that the Notch pathway may be activated in some human gastric cancers, supporting a potential role for Notch in gastric tumorigenesis. In this review, we first summarize the current understanding of gastric stem cells defined by genetic mouse studies, followed by discussion of the literature regarding Notch pathway regulation of gastric stem cell function in the mouse and human beings. Notch action to maintain gastric epithelial cell homeostasis and the cellular consequences of dysregulated signaling to promote tumorigenesis are discussed, including studies associating Notch activation with human gastric cancer. Finally, we compare and contrast Notch function in the stomach with other gastrointestinal tissues, including the intestine, to highlight the sensitivity of the stomach to Notch-induced tumors.

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

    DEFF Research Database (Denmark)

    Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla

    2011-01-01

    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...... 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...

  15. Vaginal epithelial dendritic cells renew from bone marrow precursors.

    Science.gov (United States)

    Iijima, Norifumi; Linehan, Melissa M; Saeland, Sem; Iwasaki, Akiko

    2007-11-27

    Dendritic cells (DCs) represent key professional antigen-presenting cells capable of initiating primary immune responses. A specialized subset of DCs, the Langerhans cells (LCs), are located in the stratified squamous epithelial layer of the skin and within the mucosal epithelial lining of the vaginal and oral cavities. The vaginal mucosa undergoes cyclic changes under the control of sex hormones, and the renewal characteristics of the vaginal epithelial DCs (VEDCs) remain unknown. Here, we examined the origin of VEDCs. In contrast to the skin epidermal LCs, the DCs in the epithelium of the vagina were found to be repopulated mainly by nonmonocyte bone-marrow-derived precursors, with a half-life of 13 days under steady-state conditions. Upon infection with HSV-2, the Gr-1(hi) monocytes were found to give rise to VEDCs. Furthermore, flow cytometric analysis of the VEDCs revealed the presence of at least three distinct populations, namely, CD11b(+)F4/80(hi), CD11b(+)F4/80(int), and CD11b(-)F4/80(-). Importantly, these VEDC populations expressed CD207 at low levels and had a constitutively more activated phenotype compared with the skin LCs. Collectively, our results revealed mucosa-specific features of the VEDCs with respect to their phenotype, activation status, and homeostatic renewal potential.

  16. Epithelial stem cell islands in the regenerated epidermis

    Institute of Scientific and Technical Information of China (English)

    Fu Xiaobing; Sun Xiaoqing; Li Xiaokun; Sheng Zhiyong

    2001-01-01

    Objective: The effects of growth factors on wound healing have been studied extensively, however,their molecular and genetic mechanisms that regulate epidermal regeneration are not fully understood. In this study,we explore the cell reversion characteristics and epithelial stem cell distribution in human regenerated epidermis treated with recombinant human epidermal growth factor (rhEGF). Methods:Tissue biospies from 8 regenerated skins treated with rhEGF were used to evaluate the cell reversion and stem cell distribution in epidermis . The expression of β1 integrin, keratin 19 (K19), keratin 14 (K14) and keratin 10 (K10) in skins was detected with SP immunohistochemical methods. Another 8 biopsies from the regenerated epidermis treated without rhEGF, fetus, children and adults were used as the controls. Results:Immunohistochemical stain for β1 integrin and keratin 19 showed that there were some new stem cell islands in the epidermis treated with rhEGF. These cells were small, containing low RNA content and exhibiting positive expression with β1 integrin and K19 stain. They were isolated, bearing no anatomic relation with the epithelial stem cells in the basal layer. The serial identification experiments indicated that there treated without rhEGF. All of these results supported that these β1 integrin and K19 positive stain cells were the stem cells. Conclusions: The results indicated that these stem cell islands were the specific and individual cell structures in rhEGF treated wounds and rhEGF is the main factor in inducing the stem cell island formation. These results offer a direct evidence for epidermal cell reversion from the differentiated cells to undifferentiated stem cells in vivo and may be useful in the rational use of this growth factor to promote wound healing in clinic.

  17. 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.

  18. Toxic mechanisms of copper oxide nanoparticles in epithelial kidney cells

    DEFF Research Database (Denmark)

    Thit, Amalie; Selck, Henriette; Bjerregaard, Henning F.

    2015-01-01

    CuO NPs have previously been reported as toxic to a range of cell cultures including kidney epithelial cells from the frog, Xenopus laevis (A6). Here we examine the molecular mechanisms affecting toxicity of Cu in different forms and particle sizes. A6 cells were exposed to ionic Cu (Cu2+) or Cu......O particles of three different sizes: CuO NPs of 6 nm (NP6), larger Poly-dispersed CuO NPs of toxic than NP6, Micro and Cu2+ to A6 cells, causing DNA damage, decreased cell viability...... and levels of reduced glutathione (GSH) and eventually cell death. We show that ROS (Reactive Oxygen Species) generation plays a key role and occurs early in Poly toxicity as Poly-induced DNA damage and cell death could be mitigated by the antioxidant NAC (N-acetyl-cysteine). Here we propose a model...

  19. Transplantation of Airway Epithelial Stem/Progenitor Cells: A Future for Cell-Based Therapy.

    Science.gov (United States)

    Ghosh, Moumita; Ahmad, Shama; White, Carl W; Reynolds, Susan D

    2017-01-01

    Cell therapy has the potential to cure disease through replacement of malfunctioning cells. Although the tissue stem cell (TSC) is thought to be the optimal therapeutic cell, transplantation of TSC/progenitor cell mixtures has saved lives. We previously purified the mouse tracheobronchial epithelial TSCs and reported that in vitro amplification generated numerous TSCs. However, these cultures also contained TSC-derived progenitor cells and TSC repurification by flow cytometry compromised TSC self-renewal. These limitations prompted us to determine if a TSC/progenitor cell mixture would repopulate the injured airway epithelium. We developed a cell transplantation protocol and demonstrate that transplanted mouse and human tracheobronchial epithelial TSC/progenitor cell mixtures are 20-25% of airway epithelial cells, actively contribute to epithelial repair, and persist for at least 43 days. At 2 weeks after transplantation, TSCs/progenitor cells differentiated into the three major epithelial cell types: basal, secretory, and ciliated. We conclude that cell therapy that uses adult tracheobronchial TSCs/progenitor cells is an effective therapeutic option.

  20. Rho GTPases and regulation of cell migration and polarization in human corneal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Aihua Hou

    Full Text Available PURPOSE: Epithelial cell migration is required for regeneration of tissues and can be defective in a number of ocular surface diseases. This study aimed to determine the expression pattern of Rho family small G-proteins in human corneal epithelial cells to test their requirement in directional cell migration. METHODS: Rho family small G-protein expression was assessed by reverse transcription-polymerase chain reaction. Dominant-inhibitory constructs encoding Rho proteins or Rho protein targeting small interfering RNA were transfected into human corneal epithelial large T antigen cells, and wound closure rate were evaluated by scratch wounding assay, and a complementary non-traumatic cell migration assay. Immunofluorescence staining was performed to study cell polarization and to assess Cdc42 downstream effector. RESULTS: Cdc42, Chp, Rac1, RhoA, TC10 and TCL were expressed in human corneal epithelial cells. Among them, Cdc42 and TCL were found to significantly affect cell migration in monolayer scratch assays. These results were confirmed through the use of validated siRNAs directed to Cdc42 and TCL. Scramble siRNA transfected cells had high percentage of polarized cells than Cdc42 or TCL siRNA transfected cells at the wound edge. We showed that the Cdc42-specific effector p21-activated kinase 4 localized predominantly to cell-cell junctions in cell monolayers, but failed to translocate to the leading edge in Cdc42 siRNA transfected cells after monolayer wounding. CONCLUSION: Rho proteins expressed in cultured human corneal epithelial cells, and Cdc42, TCL facilitate two-dimensional cell migration in-vitro. Although silencing of Cdc42 and TCL did not noticeably affect the appearance of cell adhesions at the leading edge, the slower migration of these cells indicates both GTP-binding proteins play important roles in promoting cell movement of human corneal epithelial cells.

  1. Prion infection of epithelial Rov cells is a polarized event.

    Science.gov (United States)

    Paquet, Sophie; Sabuncu, Elifsu; Delaunay, Jean-Louis; Laude, Hubert; Vilette, Didier

    2004-07-01

    During prion infections, the cellular glycosylphosphatidylinositol-anchored glycoprotein PrP is converted into a conformational isoform. This abnormal conformer is thought to recruit and convert the normal cellular PrP into a likeness of itself and is proposed to be the infectious agent. We investigated the distribution of the PrP protein on the surface of Rov cells, an epithelial cell line highly permissive to prion multiplication, and we found that PrP is primarily expressed on the apical side. We further show that prion transmission to Rov cells is much more efficient if infectivity contacts the apical side, indicating that the apical and basolateral sides of Rov cells are not equally competent for prion infection and adding prions to the list of the conventional infectious agents (viruses and bacteria) that infect epithelial cells in a polarized manner. These data raise the possibility that apically expressed PrP may be involved in this polarized process of infection. This would add further support for a crucial role of PrP at the cell surface in prion infection of target cells.

  2. Epithelial progenitor cell lines as models of normal breast morphogenesis and neoplasia

    DEFF Research Database (Denmark)

    Petersen, Ole William; Gudjonsson, Thorarinn; Villadsen, René;

    2003-01-01

    epithelial or the myoepithelial cell phenotype in primary cultures. Having succeeded in continuous propagation presumably without loss of markers, we could show that a subset of the luminal epithelial cells could convert to myoepithelial cells, signifying the possible existence of a progenitor cell...... cell lines. This suprabasal-derived epithelial cell line is able to generate both itself and differentiated luminal epithelial and myoepithelial cells, and in addition, is able to form elaborate terminal duct lobular unit (TDLU)-like structures within a reconstituted basement membrane. As more than 90...

  3. Slugging their way to immortality: driving mammary epithelial cells into a stem cell-like state.

    Science.gov (United States)

    Soady, Kelly; Smalley, Matthew J

    2012-09-10

    Delineating the molecular factors that define and maintain the mammary stem cell state is vital for understanding normal development and tumourigenesis. A recent study by Guo and colleagues identifies two master transcriptional regulators of mammary stem cells, Slug and Sox9, ectopic expression of which confers stem cell attributes on differentiated mammary epithelial cells. Slug and Sox9 expression was also shown to determine in vivo metastatic potential of human breast cancer cell lines. Understanding these factors in the context of normal lineage differentiation is an important step toward elucidating the mammary epithelial cell hierarchy and the origins of cancer stem cells.

  4. Effects of N-acetylcysteine on matrix metalloproteinase-9 secretion and cell migration of human corneal epithelial cells

    OpenAIRE

    Ramaesh, T; Ramaesh, K; Riley, S C; West, J.D.; Dhillon, B

    2012-01-01

    Matrix metalloproteinase-9 (MMP-9) secreted by corneal epithelial cells has a role in the remodelling of extracellular matrix and migration of epithelial cells. Elevated levels of MMP-9 activity in the ocular surface may be involved in the pathogenesis of corneal diseases. N-acetylcysteine (NAC) has been used to treat corneal diseases, including recurrent epithelial erosions. In this study, its effects on the MMP-9 secretion and human corneal epithelial (HCE) cell migration were evaluated in ...

  5. 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

  6. Stimulation of mucin secretion from human bronchial epithelial cells by mast cell chymase

    Institute of Scientific and Technical Information of China (English)

    Shao-heng HE; Jian ZHENG

    2004-01-01

    AIM: To investigate the effect ofchymase on the mucin secretion from human bronchial epithelial cells. METHODS:Primarily-cultured human bronchial epithelial (PCHBE) cells and normal human bronchial epithelial (NHBE) cells were cultured with chymase or other stimulus in a mixture of bronchial epithelial growth medium (BEGM) and Dulbecco's modified Eagle's medium (DMEM), and the quantities of stimulatory mucin release were recorded.MUC5AC mucin was measured with an ELISA and dolichos biflorus agglutinin (DBA) mucin was determined with an enzyme linked DBA assay. RESULTS: A dose-dependent secretion of DBA mucin from PCHBE cells was observed with chymase with a maximum secretion of 98 % above baseline being achieved following 3 h incubation.The action of chymase started from 1 h, peaked at 3 h and dramatically decreased at 20 h following incubation.Chymase was able to also stimulate approximately 38 % increase in MUC5AC mucin release from PCHBE cells, and about 121% increase in DBA mucin release from NHBE cells. A chymase inhibitor soybean trypsin inhibitor (SBTI)was able to inhibit up to 85 % chymase induced mucin release, indicating that the enzymatic activity was essential for the actions of chymase on bronchial epithelial cells. CONCLUSION: Chymase is a potent stimulus of mucin secretion from human bronchial epithelial cells. It can contribute to mucus hypersecretion process in the patients with chronic obstructive pulmonary disease or asthma.

  7. Impact of mechanical stretch on the cell behaviors of bone and surrounding tissues

    Directory of Open Access Journals (Sweden)

    Hye-Sun Yu

    2016-02-01

    Full Text Available Mechanical loading is recognized to play an important role in regulating the behaviors of cells in bone and surrounding tissues in vivo. Many in vitro studies have been conducted to determine the effects of mechanical loading on individual cell types of the tissues. In this review, we focus specifically on the use of the Flexercell system as a tool for studying cellular responses to mechanical stretch. We assess the literature describing the impact of mechanical stretch on different cell types from bone, muscle, tendon, ligament, and cartilage, describing individual cell phenotype responses. In addition, we review evidence regarding the mechanotransduction pathways that are activated to potentiate these phenotype responses in different cell populations.

  8. The search for truth and freedom: ethical issues surrounding human cloning and stem cell research.

    Science.gov (United States)

    Bruce, Alex

    2002-02-01

    The reality of cloning and stem cell research has provoked wonder, fear and anger. These developments have the potential fundamentally to alter humanity. But how well informed is the range of views being expressed? Is progress being threatened by understandable but uninformed fears? Or are scientists rushing toward an ethical abyss, so concerned with what they can do that they never stop to ask what they should do? This article identifies some of the fears and hopes surrounding cloning and stem cell research. It aims to provoke ethical debate in evaluating such research.

  9. EOTAXIN AND EOTAXIN-2 EXPRESSION IN HUMAN BRONCHIAL EPITHELIAL CELL

    Institute of Scientific and Technical Information of China (English)

    TANG Wei; DENG Wei-wu; Albert CHAN; Stanley CHIK; Adrain WU

    2005-01-01

    Objective To study the role of eotaxin and eotaxin-2 expression by Th2 cytokine and analyze their relationship in normal human bronchial epithelial cell line-BEAS-2B cell. Methods Levels of eotaxin mRNA and protein expression in the bronchial epithelial cell line BEAS-2B cell were determined with RT-PCR and ELISA. We also used RT-PCR to evaluate eotaxin-2 expression under the regulation of Th2 cytokine IL-4 and IL-13 as well as proinflammatory agent-TNFα. Results Eotaxin mRNA expression was the highest at the time point of 12h under the stimulation of TNF-α. While Th2 cytokine IL-4 and IL-13 had the amplification effect on the expression. Eotaxin protein was also elevated with the combination stimulation of proinflammatory agent TNF-α and IL-4 in dose and time dependent manner(P<0.01). These results were also seen when the cells were stimulated by TNF-α and IL-13. Eotaxin-2 mRNA expression was the highest at the time point of 8h. The expression evaluated by semi-quantitative RT-PCR also elevated under the co-stimulation of TNF-α and IL-4 or TNF-α and IL-13 and it should significantly correlate with Eotaxin(P<0.05). Conclusion This study demonstrated that Th2 cytokine like IL-4 and IL-13 enhances eotaxin and eotaxin-2 expression when co-stimulated with proinflammatory agent TNF-α. These results showed that Th2 cytokines existence is the strong evidence for bronchial epithelial cells taking part in the allergic inflammation especially in eosinophils recruitment.

  10. Cytotoxic effects of composite dust on human bronchial epithelial cells.

    Science.gov (United States)

    Cokic, Stevan M; Hoet, Peter; Godderis, Lode; Wiemann, Martin; Asbach, Christof; Reichl, Franz X; De Munck, Jan; Van Meerbeek, Bart; Van Landuyt, Kirsten L

    2016-12-01

    Previous research revealed that during routine abrasive procedures like polishing, shaping or removing of composites, high amounts of respirable dust particles (composite dust particles on bronchial epithelium cells. Composite dust of five commercial composites (one nano-composite, two nano-hybrid and two hybrid composites) was generated following a clinically relevant protocol. Polymerized composite samples were cut with a rough diamond bur (grain size 100μm, speed 200,000rpm) and all composite dust was collected in a sterile chamber. Human bronchial epithelial cells (16HBE14o-) were exposed to serially diluted suspensions of composite dust in cell culture medium at concentrations between 1.1 and 3.3mg/ml. After 24h-exposure, cell viability and membrane integrity were assessed by the WST-1 and the LDH leakage assay, respectively. The release of IL-1β and IL-6 was evaluated. The composite dust particles were characterized by transmission electron microscopy and by dynamic and electrophoretic light scattering. Neither membrane damage nor release of IL-1β was detected over the complete concentration range. However, metabolic activity gradually declined for concentrations higher than 660μg/ml and the release of IL-6 was reduced when cells were exposed to the highest concentrations of dust. Composite dust prepared by conventional dental abrasion methods only affected human bronchial epithelial cells in very high concentrations. Copyright © 2016 The Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

  11. Tissuelike 3D Assemblies of Human Broncho-Epithelial Cells

    Science.gov (United States)

    Goodwin, Thomas J.

    2010-01-01

    Three-dimensional (3D) tissuelike assemblies (TLAs) of human broncho-epithelial (HBE) cells have been developed for use in in vitro research on infection of humans by respiratory viruses. The 2D monolayer HBE cell cultures heretofore used in such research lack the complex cell structures and interactions characteristic of in vivo tissues and, consequently, do not adequately emulate the infection dynamics of in-vivo microbial adhesion and invasion. In contrast, the 3D HBE TLAs are characterized by more-realistic reproductions of the geometrical and functional complexity, differentiation of cells, cell-to-cell interactions, and cell-to-matrix interactions characteristic of human respiratory epithelia. Hence, the 3D HBE TLAs are expected to make it possible to perform at least some of the research in vitro under more-realistic conditions, without need to infect human subjects. The TLAs are grown on collagen-coated cyclodextran microbeads under controlled conditions in a nutrient liquid in the simulated microgravitational environment of a bioreactor of the rotating- wall-vessel type. Primary human mesenchymal bronchial-tracheal cells are used as a foundation matrix, while adult human bronchial epithelial immortalized cells are used as the overlying component. The beads become coated with cells, and cells on adjacent beads coalesce into 3D masses. The resulting TLAs have been found to share significant characteristics with in vivo human respiratory epithelia including polarization, tight junctions, desmosomes, and microvilli. The differentiation of the cells in these TLAs into tissues functionally similar to in vivo tissues is confirmed by the presence of compounds, including villin, keratins, and specific lung epithelium marker compounds, and by the production of tissue mucin. In a series of initial infection tests, TLA cultures were inoculated with human respiratory syncytial viruses and parainfluenza type 3 viruses. Infection was confirmed by photomicrographs that

  12. Inflammation and the Intestinal Barrier: Leukocyte–Epithelial Cell Interactions, Cell Junction Remodeling, and Mucosal Repair

    Science.gov (United States)

    Luissint, Anny-Claude; Parkos, Charles A.; Nusrat, Asma

    2017-01-01

    The intestinal tract is lined by a single layer of columnar epithelial cells that forms a dynamic, permeable barrier allowing for selective absorption of nutrients, while restricting access to pathogens and food-borne antigens. Precise regulation of epithelial barrier function is therefore required for maintaining mucosal homeostasis and depends, in part, on barrier-forming elements within the epithelium and a balance between pro- and anti-inflammatory factors in the mucosa. Pathologic states, such as inflammatory bowel disease, are associated with a leaky epithelial barrier, resulting in excessive exposure to microbial antigens, recruitment of leukocytes, release of soluble mediators, and ultimately mucosal damage. An inflammatory microenvironment affects epithelial barrier properties and mucosal homeostasis by altering the structure and function of epithelial intercellular junctions through direct and indirect mechanisms. We review our current understanding of complex interactions between the intestinal epithelium and immune cells, with a focus on pathologic mucosal inflammation and mechanisms of epithelial repair. We discuss leukocyte–epithelial interactions, as well as inflammatory mediators that affect the epithelial barrier and mucosal repair. Increased knowledge of communication networks between the epithelium and immune system will lead to tissue-specific strategies for treating pathologic intestinal inflammation. PMID:27436072

  13. 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...

  14. HAM56 and CD68 antigen presenting cells surrounding a sarcoidal granulomatous tattoo

    Directory of Open Access Journals (Sweden)

    Ana Maria Abreu Velez

    2011-01-01

    Full Text Available Context : Tattoos are produced by introducing colorants of various compositions into the skin, either accidentally or for cosmetic purposes. Case Report: A 62-year-old male presented with a cosmetic tattoo and requested a total excision of the lesion. Dermatopathologic analysis of the excised tissue with hematoxylin and eosin examination, as well as immunohistochemistry was performed. H&E staining demonstrated classic histologic features of a tattoo. Utilizing immunohistochemistry, dermal histiocytic antigen presenting cells stained with HAM56 and CD68 antibodies; the staining was present surrounding the tattoo pigment. Conclusions : We identified two macrophage markers (HAM56 and CD68 surrounding dermal tattoo pigment. A minimal dermal inflammatory immune was noted to the tattoo pigment. Moreover, the immune response and/or tolerance to tattoos is not well characterized. We suggest that tattoo materials and techniques could be utilized in therapeutic delivery for diseases such recessive dystrophic epidermolysis bullosa, potentially preventing immune rejection of gene therapy agents.

  15. Oral microbial biofilm stimulation of epithelial cell responses.

    Science.gov (United States)

    Peyyala, Rebecca; Kirakodu, Sreenatha S; Novak, Karen F; Ebersole, Jeffrey L

    2012-04-01

    Oral bacterial biofilms trigger chronic inflammatory responses in the host that can result in the tissue destructive events of periodontitis. However, the characteristics of the capacity of specific host cell types to respond to these biofilms remain ill-defined. This report describes the use of a novel model of bacterial biofilms to stimulate oral epithelial cells and profile select cytokines and chemokines that contribute to the local inflammatory environment in the periodontium. Monoinfection biofilms were developed with Streptococcus sanguinis, Streptococcus oralis, Streptococcus gordonii, Actinomyces naeslundii, Fusobacterium nucleatum, and Porphyromonas gingivalis on rigid gas-permeable contact lenses. Biofilms, as well as planktonic cultures of these same bacterial species, were incubated under anaerobic conditions with a human oral epithelial cell line, OKF4, for up to 24h. Gro-1α, IL1α, IL-6, IL-8, TGFα, Fractalkine, MIP-1α, and IP-10 were shown to be produced in response to a range of the planktonic or biofilm forms of these species. P. gingivalis biofilms significantly inhibited the production of all of these cytokines and chemokines, except MIP-1α. Generally, the biofilms of all species inhibited Gro-1α, TGFα, and Fractalkine production, while F. nucleatum biofilms stimulated significant increases in IL-1α, IL-6, IL-8, and IP-10. A. naeslundii biofilms induced elevated levels of IL-6, IL-8 and IP-10. The oral streptococcal species in biofilms or planktonic forms were poor stimulants for any of these mediators from the epithelial cells. The results of these studies demonstrate that oral bacteria in biofilms elicit a substantially different profile of responses compared to planktonic bacteria of the same species. Moreover, certain oral species are highly stimulatory when in biofilms and interact with host cell receptors to trigger pathways of responses that appear quite divergent from individual bacteria.

  16. 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...

  17. Epstein-Barr virus infection and persistence in nasopharyngeal epithelial cells

    Institute of Scientific and Technical Information of China (English)

    Chi Man Tsang; Wen Deng; Yim Ling Yip; Mu-Sheng Zeng; Kwok Wai Lo; Sai Wah Tsao

    2014-01-01

    Epstein-Barr virus (EBV) infection is closely associated with undifferentiated nasopharyngeal carcinoma (NPC), strongly implicating a role for EBV in NPC pathogenesis; conversely, EBV infection is rarely detected in normal nasopharyngeal epithelial tissues. In general, EBV does not show a strong tropism for infecting human epithelial cels, and EBV infection in oropharyngeal epithelial cels is believed to be lytic in nature. To establish life-long infection in humans, EBV has evolved efficient strategies to infect B cels and hijack their celular machinery for latent infection. Lytic EBV infection in oropharyngeal epithelial cels, though an infrequent event, is believed to be a major source of infectious EBV particles for salivary transmission. The biological events associated with nasopharyngeal epithelial cells are only beginning to be understood with the advancement of EBV infection methods and the availability of nasopharyngeal epithelial cel models for EBV infection studies. EBV infection in human epithelial cels is a highly inefficient process compared to that in B cels, which express the complement receptor type 2 (CR2) to mediate EBV infection. Although receptor(s) on the epithelial cell surface for EBV infection remain(s) to be identified, EBV infection in epithelial cels could be achieved via the interaction of glycoproteins on the viral envelope with surface integrins on epithelial cels, which might trigger membrane fusion to internalize EBV in cels. Normal nasopharyngeal epithelial cells are not permissive for latent EBV infection, and EBV infection in normal nasopharyngeal epithelial cells usually results in growth arrest. However, genetic alterations in premalignant nasopharyngeal epithelial cells, including p16 deletion and cyclin D1 overexpression, could override the growth inhibitory effect of EBV infection to support stable and latent EBV infection in nasopharyngeal epithelial cells. The EBV episome in NPC is clonal in nature, suggesting that NPC

  18. EBV infection is common in gingival epithelial cells of the periodontium and worsens during chronic periodontitis.

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    Séverine Vincent-Bugnas

    Full Text Available An amplifying role for oral epithelial cells (ECs in Epstein-Barr Virus (EBV infection has been postulated to explain oral viral shedding. However, while lytic or latent EBV infections of oro/nasopharyngeal ECs are commonly detected under pathological conditions, detection of EBV-infected ECs in healthy conditions is very rare. In this study, a simple non-surgical tissue sampling procedure was used to investigate EBV infection in the periodontal epithelium that surrounds and attaches teeth to the gingiva. Surprisingly, we observed that the gingival ECs of the periodontium (pECs are commonly infected with EBV and may serve as an important oral reservoir of latently EBV-infected cells. We also found that the basal level of epithelial EBV-infection is significantly increased in chronic periodontitis, a common inflammatory disease that undermines the integrity of tooth-supporting tissues. Moreover, the level of EBV infection was found to correlate with disease severity. In inflamed tissues, EBV-infected pECs appear to be prone to apoptosis and to produce larger amounts of CCL20, a pivotal inflammatory chemokine that controls tissue infiltration by immune cells. Our discovery that the periodontal epithelium is a major site of latent EBV infection sheds a new light on EBV persistence in healthy carriers and on the role of this ubiquitous virus in periodontitis. Moreover, the identification of this easily accessible site of latent infection may encourage new approaches to investigate and monitor other EBV-associated disorders.

  19. Milk Fat Globule Membrane Proteomics: A 'Snapshot' of Mammary Epithelial Cell Biology

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    Christelle Cebo

    2012-01-01

    Full Text Available Lipids are released in milk as fat globules, which are droplets of apolar lipids surrounded by a complex membrane deriving from the mammary epithelial cell (MEC and called the milk fat globule membrane (MFGM. The structure of the MFGM is highly complex and closely related to the mechanisms of milk fat globule secretion in the mammary epithelial cell. Indeed, MFGM is composed of two biological membranes, a phospholipid monolayer, deriving from the endoplasmic reticulum, and a phospholipid bilayer, which originates from the apical plasma membrane of the MEC, with variable amounts of cytoplasm trapped between. Biochemical techniques (i.e. sodium dodecyl sulphate-polyacrylamide gel electrophoresis followed by different staining procedures have been employed historically to characterize major MFGM proteins, namely MUC-1, fatty acid synthase, xanthine oxidase, butyrophilin, lactadherin, and adipophilin. However, recent advances in the field of proteomics (mostly development of one-dimensional gel electrophoresis approach coupled with tandem mass spectrometry have led to the identification of hundreds of proteins associated with the MFGM. Surprisingly, newly identified MFGM proteins were not only involved in lipid metabolic or exocytosis-related biological processes, but also in cell signalling, translation, or host defense-related mechanisms. Therefore, the milk fat globule should no longer be viewed as an inert structure only devoted to the delivery of lipids to the newborn, but rather as a dynamic and informative compartment which can contribute to the improvement of our comprehension of the mammary gland biology.

  20. High glucose stimulates the expression of erythropoietin in rat glomerular epithelial cells

    OpenAIRE

    Lim, Seul Ki; Park, Soo Hyun

    2011-01-01

    It has been reported that the levels of erythropoietin are associated with diabetes mellitus. Glomerular epithelial cells, located in the renal cortex, play an important role in the regulation of kidney function and hyperglycemia-induced cell loss of glomerular epithelial cells is implicated in the onset of diabetic nephropathy. This study investigated the effect of high glucose on erythropoietin and erythropoietin receptor expression in rat glomerular epithelial cells. We found that 25 mM D-...

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

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    Shuxian Jiang

    Full Text Available BACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: 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. CONCLUSIONS/SIGNIFICANCE: 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.

  2. Bordetella pertussis entry into respiratory epithelial cells and intracellular survival.

    Science.gov (United States)

    Lamberti, Yanina; Gorgojo, Juan; Massillo, Cintia; Rodriguez, Maria E

    2013-12-01

    Bordetella pertussis is the causative agent of pertussis, aka whooping cough. Although generally considered an extracellular pathogen, this bacterium has been found inside respiratory epithelial cells, which might represent a survival strategy inside the host. Relatively little is known, however, about the mechanism of internalization and the fate of B. pertussis inside the epithelia. We show here that B. pertussis is able to enter those cells by a mechanism dependent on microtubule assembly, lipid raft integrity, and the activation of a tyrosine-kinase-mediated signaling. Once inside the cell, a significant proportion of the intracellular bacteria evade phagolysosomal fusion and remain viable in nonacidic lysosome-associated membrane-protein-1-negative compartments. In addition, intracellular B. pertussis was found able to repopulate the extracellular environment after complete elimination of the extracellular bacteria with polymyxin B. Taken together, these data suggest that B. pertussis is able to survive within respiratory epithelial cells and by this means potentially contribute to host immune system evasion.

  3. Nerve growth factor modulate proliferation of cultured rabbit corneal endothelial cells and epithelial cells.

    Science.gov (United States)

    Li, Xinyu; Li, Zhongguo; Qiu, Liangxiu; Zhao, Changsong; Hu, Zhulin

    2005-01-01

    In order to investigate the effect of nerve growth factor (NGF) on the proliferation of rabbit corneal endothelial cells and epithelial cells, the in vitro cultured rabbit corneal endothelial cells and epithelial cells were treated with different concentrations of NGF. MTT assay was used to examine the clonal growth and proliferation of the cells by determining the absorbency values at 570 nm. The results showed that NGF with three concentrations ranging from 5 U/mL to 500 U/mL enhanced the proliferation of rabbit corneal endothelial cells in a concentration-dependent manner. 50 U/mL and 500 U/mL NGF got more increase of proliferation than that of 5 U/mL NGF did. Meanwhile, 50 U/mL and 500 U/mL NGF could promote the proliferation of the rabbit corneal epithelial cells significantly in a concentration-dependent manner. However, 5 U/mL NGF did not enhance the proliferation of epithelial cells. It was suggested that exogenous NGF can stimulate the proliferation of both rabbit corneal endothelial and epithelial cells, but the extent of modulation is different.

  4. Expression of ICAM-1 in colon epithelial cells

    DEFF Research Database (Denmark)

    Vainer, Ben; Sørensen, Susanne; Seidelin, Jakob;

    2003-01-01

    on monolayers of cancer cells. Conflicting results exist on epithelial ICAM-1 expression, and the aim of this study was to compare the expression in various models of colonic epithelium. MATERIALS AND METHODS: Colonic biopsies from four UC patients and four controls were examined by cryoimmuno......-electron microscopy using ICAM-1-antibodies. In four other controls, the epithelium was isolated from colonic biopsies, embedded in collagen, and evaluated similarly. Isolated crypts and cultured cancer cells were stimulated with interferon-gamma (IFN-gamma) or tumor necrosis factor-alpha (TNF-alpha). RESULTS: ICAM-1......, both colonocytes and HT29 cells were capable of expressing ICAM-1 on their apical membranes in response to supraphysiologic cytokine concentrations. These observations question the justification of extrapolating observations from colon cancer cell lines to in vivo inflammatory conditions....

  5. Human respiratory epithelial cells from nasal turbinate expressed stem cell genes even after serial passaging.

    Science.gov (United States)

    Ruszymah, B H I; Izham, B A Azrul; Heikal, M Y Mohd; Khor, S F; Fauzi, M B; Aminuddin, B S

    2011-12-01

    Current development in the field of tissue engineering led to the idea of repairing and regenerating the respiratory airway through in vitro reconstruction using autologous respiratory epithelial (RE). To ensure the capability of proliferation, the stem cell property of RE cells from the nasal turbinate should be evaluated. Respiratory epithelial cells from six human nasal turbinates were harvested and cultured in vitro. The gene expression of FZD-9 and BST-1 were expressed in passage 2 (P2) and passage 4 (P4). The levels of expression were not significant between both passages. The RE cells exhibit the stem cell properties, which remains even after serial passaging.

  6. Cell-mediated infection of cervix derived epithelial cells with primary isolates of human immunodeficiency virus.

    Science.gov (United States)

    Tan, X; Phillips, D M

    1996-01-01

    We have previously demonstrated that HIV-infected transformed T-cells or monocytes adhere to monolayers of CD4-negative epithelial cells. Adhesion is soon followed by budding of HIV from infected mononuclear cells onto the surface of epithelial cells. Epithelial cells subsequently take up virus and become productively infected. Based on these findings, we proposed that sexual transmission of HIV may involve cell-mediated infection of intact mucosal epithelia of the urogenital tract. However, it has become increasingly clear that primary cells and HIV strains isolated from patients are more appropriate models for HIV infection than established cell lines and lab strains of virus. In the studies described here, we infected cervix-derived epithelial monolayers with primary monocytes infected with patient isolates of non-syncytial inducing (NSI) macrophage-tropic strains of HIV. Under the culture conditions employed, HIV-infected primary monocytes do not remain adherent to the apical surface of the epithelium, as did HIV-infected transformed cells. Instead, following adherence, the primary cells migrate between epithelial cells. Virus is secreted from a pseudopod as HIV-infected primary monocytes pass between cells of the epithelium. Productive infection of the epithelium was detected by p24 ELISA and PCR Southern blot analysis. Infection can be blocked by sera from HIV-seropositive individuals or by certain sulfated polysaccharides. These findings support the supposition that transmission of HIV may occur via cell-mediated infection of intact epithelia. The observations also hint at the possibility that-HIV-infected monocyte/macrophages in semen or cervical-vaginal secretions could cross intact epithelia by passing between epithelial cells. Blocking studies suggest that it may be possible to inhibit sexual transmission of HIV either by antibodies in genital tract secretions or by a topical formulation containing certain sulfated polysaccharides.

  7. Interaction between submicron COD crystals and renal epithelial cells

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    Peng H

    2012-08-01

    Full Text Available Hua Peng1,2 Jian-Ming Ouyang1,2 Xiu-Qiong Yao1, Ru-E Yang11Department of Chemistry, Jinan University, 2Institute of Biomineralization and Lithiasis Research, Jinan University, Guangzhou, ChinaObjectives: This study aims to investigate the adhesion characteristics between submicron calcium oxalate dihydrate (COD with a size of 150 ± 50 nm and African green monkey kidney epithelial cells (Vero cells before and after damage, and to discuss the mechanism of kidney stone formation.Methods: Vero cells were oxidatively injured by hydrogen peroxide to establish a model of injured cells. Scanning electron microscopy was used to observe Vero–COD adhesion. Inductively coupled plasma emission spectrometry was used to quantitatively measure the amount of adhered COD microcrystals. Nanoparticle size analyzer and laser scanning confocal microscopy were performed to measure the change in the zeta potential on the Vero cell surface and the change in osteopontin expression during the adhesion process, respectively. The level of cell injury was evaluated by measuring the changes in malonaldehyde content, and cell viability during the adhesion process.Results: The adhesion capacity of Vero cells in the injury group to COD microcrystals was obviously stronger than that of Vero cells in the control group. After adhesion to COD, cell viability dropped, both malonaldehyde content and cell surface zeta potential increased, and the fluorescence intensity of osteopontin decreased because the osteopontin molecules were successfully covered by COD. Submicron COD further damaged the cells during the adhesion process, especially for Vero cells in the control group, leading to an elevated amount of attached microcrystals.Conclusion: Submicron COD can further damage injured Vero cells during the adhesion process. The amount of attached microcrystals is proportional to the degree of cell damage. The increased amount of microcrystals that adhered to the injured epithelial

  8. Effects of different Helicobacter pylori culture filtrates on growth of gastric epithelial cells

    Institute of Scientific and Technical Information of China (English)

    Yan-Guo Yan; Gang Zhao; Jin-Ping Ma; Shi-Rong Cai; Wen-Hua Zhan

    2008-01-01

    AIM: To study the effects of different Helicobacter pylori (H py/orl) culture filtrates on growth of gastric epithelial cells.METHODS: Broth culture filtrates of H pylori were prepared. Gastric epithelial cells were treated with the filtrates, and cell growth was determined by growth curve and flow cytometry. DNA damage of gastric epithelial cells was measured by single-cell microgel electrophoresis.RESULTS: Gastric epithelial cells proliferated actively when treated by CagA-gene-positive broth culture filtrates, and colony formation reached 40%. The number of cells in S phase increased compared to controls. Comet assay showed 41.2% comet cells in GES-1 cells treated with CagA-positive filtrates (P<0.05).CONCLUSION: CagA-positive filtrates enhance the changes in morphology and growth characteristics of human gastric epithelial tumor cells. DNA damage maybe one of the mechanisms involved in the growth changes.

  9. Apposition to endometrial epithelial cells activates mouse blastocysts for implantation.

    Science.gov (United States)

    Ruane, Peter T; Berneau, Stéphane C; Koeck, Rebekka; Watts, Jessica; Kimber, Susan J; Brison, Daniel R; Westwood, Melissa; Aplin, John D

    2017-09-01

    How do interactions between blastocyst-stage embryos and endometrial epithelial cells regulate the early stages of implantation in an in vitro model? Mouse blastocyst apposition with human endometrial epithelial cells initiates trophectoderm differentiation to trophoblast, which goes on to breach the endometrial epithelium. In vitro models using mouse blastocysts and human endometrial cell lines have proven invaluable in the molecular characterisation of embryo attachment to endometrial epithelium at the onset of implantation. Genes involved in embryonic breaching of the endometrial epithelium have not been investigated in such in vitro models. This study used an established in vitro model of implantation to examine cellular and molecular interactions during blastocyst attachment to endometrial epithelial cells. Mouse blastocysts developed from embryonic day (E) 1.5 in vitro were hatched and co-cultured with confluent human endometrial adenocarcinoma-derived Ishikawa cells in serum-free medium. A scale of attachment stability based on blastocyst oscillation upon agitation was devised. Blastocysts were monitored for 48 h to establish the kinetics of implantation, and optical sectioning using fluorescence microscopy revealed attachment and invasion interfaces. Quantitative PCR was used to determine blastocyst gene expression. Data from a total of 680 mouse blastocysts are reported, with 3-6 experimental replicates. T-test and ANOVA analyses established statistical significance at P cells over the first 24 h of co-culture, with intermediate and stable attachment occurring from 28 h (E5.5 + 4 h) in a hormone-independent manner. Attached embryos fixed after 48 h (E6.5) frequently exhibited outgrowths, characterised morphologically and with antibody markers as trophoblast giant cells (TGCs), which had breached the Ishikawa cell layer. Beginning co-culture at E5.5 also resulted in intermediate and stable attachment from E5.5 + 4 h; however, these embryos did not go on to

  10. Vulnerability of Normal Human Mammary Epithelial Cells to Oncogenic Transformation

    Science.gov (United States)

    2012-04-01

    algorithm for CpG-island detection. BMC Bioinformatics 7: 446. 17. Gardiner-Garden M, Frommer M (1987) CpG islands in vertebrate genomes. J Mol Biol...it does not have a CpG island according to the original criteria (Gardiner-Garden and Frommer 1987). H3K4me3 and H3Ac are present in miR-205...culture of normal human mammary epithelial cells. Cancer Res 69: 7557–7568. Gardiner-GardenM, Frommer M. 1987. CpG islands in vertebrate genomes. J Mol

  11. Matrix proteoglycans as effector molecules for epithelial cell function

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    C. W. Frevert

    2005-12-01

    Full Text Available Matrix proteoglycans are complex molecules composed of a core protein and glycosaminoglycan side chains. Once thought to be the molecular glue providing structural support and imparting biomechanical properties to lung tissue, it is now apparent that proteoglycans are important biological modifiers which regulate processes such as lung development, homeostasis, inflammation and wound healing. The diverse roles of proteoglycans in the extracellular matrix suggest that these molecules play a critical role in normal and diseased lungs. This short article will discuss the role extracellular matrix proteoglycans play in regulating epithelial cell function in the lungs.

  12. Efficient immortalization of primary nasopharyngeal epithelial cells for EBV infection study.

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    Yim Ling Yip

    Full Text Available Nasopharyngeal carcinoma (NPC is common among southern Chinese including the ethnic Cantonese population living in Hong Kong. Epstein-Barr virus (EBV infection is detected in all undifferentiated type of NPC in this endemic region. Establishment of stable and latent EBV infection in premalignant nasopharyngeal epithelial cells is an early event in NPC development and may contribute to its pathogenesis. Immortalized primary nasopharyngeal epithelial cells represent an important tool for investigation of EBV infection and its tumorigenic potential in this special type of epithelial cells. However, the limited availability and small sizes of nasopharyngeal biopsies have seriously restricted the establishment of primary nasopharyngeal epithelial cells for immortalization. A reliable and effective method to immortalize primary nasopharyngeal epithelial cells will provide unrestricted materials for EBV infection studies. An earlier study has reported that Bmi-1 expression could immortalize primary nasopharyngeal epithelial cells. However, its efficiency and actions in immortalization have not been fully characterized. Our studies showed that Bmi-1 expression alone has limited ability to immortalize primary nasopharyngeal epithelial cells and additional events are often required for its immortalization action. We have identified some of the key events associated with the immortalization of primary nasopharyngeal epithelial cells. Efficient immortalization of nasopharyngeal epithelial cells could be reproducibly and efficiently achieved by the combined actions of Bmi-1 expression, activation of telomerase and silencing of p16 gene. Activation of MAPK signaling and gene expression downstream of Bmi-1 were detected in the immortalized nasopharyngeal epithelial cells and may play a role in immortalization. Furthermore, these newly immortalized nasopharyngeal epithelial cells are susceptible to EBV infection and supported a type II latent EBV infection

  13. Chitin-Induced Airway Epithelial Cell Innate Immune Responses Are Inhibited by Carvacrol/Thymol

    OpenAIRE

    Ali Reza Khosravi; David J Erle

    2016-01-01

    Chitin is produced in large amounts by fungi, insects, and other organisms and has been implicated in the pathogenesis of asthma. Airway epithelial cells are in direct contact with environmental particles and serve as the first line of defense against inhaled allergens and pathogens. The potential contributions of airway epithelial cells to chitin-induced asthma remain poorly understood. We hypothesized that chitin directly stimulates airway epithelial cells to release cytokines that promote ...

  14. Barrier Epithelial Cells and the Control of Type 2 Immunity.

    Science.gov (United States)

    Hammad, Hamida; Lambrecht, Bart N

    2015-07-21

    Type-2-cell-mediated immunity, rich in eosinophils, basophils, mast cells, CD4(+) T helper 2 (Th2) cells, and type 2 innate lymphoid cells (ILC2s), protects the host from helminth infection but also drives chronic allergic diseases like asthma and atopic dermatitis. Barrier epithelial cells (ECs) represent the very first line of defense and express pattern recognition receptors to recognize type-2-cell-mediated immune insults like proteolytic allergens or helminths. These ECs mount a prototypical response made up of chemokines, innate cytokines such as interleukin-1 (IL-1), IL-25, IL-33, and thymic stromal lymphopoietin (TSLP), as well as the alarmins uric acid, ATP, HMGB1, and S100 proteins. These signals program dendritic cells (DCs) to mount Th2-cell-mediated immunity and in so doing boost ILC2, basophil, and mast cell function. Here we review the general mechanisms of how different stimuli trigger type-2-cell-mediated immunity at mucosal barriers and how this leads to protection or disease. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. 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.

  16. Sustained co-cultivation with human placenta-derived MSCs enhances ALK5/Smad3 signaling in human breast epithelial cells, leading to EMT and differentiation.

    Science.gov (United States)

    Yoo, Young A; Kang, Myoung Hee; Kim, Byung Soo; Kim, Jun Suk; Seo, Jae Hong

    2009-06-01

    The interaction between mammary epithelial cells and their surrounding microenvironment are important in the development of the mammary gland. Thus, mesenchymal stem cells (MSCs), which retain pluripotency for various mesenchymal lineages, may provide a permissive environment for the morphologic alteration and differentiation of mammary epithelial cells. To this end, we investigated whether the interactions between mammary epithelial cells and human placenta-derived MSCs (hPMSC) affect the morphology, proliferation, and differentiation of epithelial cells in a co-culture system. We show that after co-culture with hPMSCs, human mammary epithelial cell lines (MCF-10F and HEMC) underwent significant morphologic alterations and a dramatic increase in ductal-alveolar branching, which was accompanied by a decrease or loss of the epithelial marker E-cadherin and a gain of the mesenchymal markers, alpha-SMA and vimentin. MCF-10F and HEMC proliferation was also inhibited in the presence of hPMSCs, and this retardation in growth was due to cell cycle arrest. Furthermore, in MCF-10F and HMEC cells, hPMSCs induced the production of lipid droplets, milk fat globule protein, and milk protein lactoferrin, which are markers of functional mammary differentiation. We also noticed an elevation in ALK5 and phosphorylated Smad3 protein levels upon hPMSC co-culture. Strikingly, the changes in morphology, proliferation, and differentiation were reversed by treatment with ALK5 or Smad3 knockdown in MCF-10F/hPMSC co-cultures. Collectively, our findings suggest that co-cultivation with hPMSCs leads to epithelial to mesenchymal transition (EMT) and differentiation of human breast epithelial cells through the ALK5/Smad3 signaling pathway.

  17. AGE-RAGE interaction in the TGFβ2-mediated epithelial to mesenchymal transition of human lens epithelial cells.

    Science.gov (United States)

    Raghavan, Cibin T; Nagaraj, Ram H

    2016-08-01

    Basement membrane (BM) proteins accumulate chemical modifications with age. One such modification is glycation, which results in the formation of advanced glycation endproducts (AGEs). In a previous study, we reported that AGEs in the human lens capsule (BM) promote the TGFβ2-mediated epithelial-to-mesenchymal transition (EMT) of lens epithelial cells, which we proposed as a mechanism for posterior capsule opacification (PCO) or secondary cataract formation. In this study, we investigated the role of a receptor for AGEs (RAGE) in the TGFβ2-mediated EMT in a human lens epithelial cell line (FHL124). RAGE was present in FHL124 cells, and its levels were unaltered in cells cultured on either native or AGE-modified BM or upon treatment with TGFβ2. RAGE overexpression significantly enhanced the TGFβ2-mediated EMT responses in cells cultured on AGE-modified BM compared with the unmodified matrix. In contrast, treatment of cells with a RAGE antibody or EN-RAGE (an endogenous ligand for RAGE) resulted in a significant reduction in the TGFβ2-mediated EMT response. This was accompanied by a reduction in TGFβ2-mediated Smad signaling and ROS generation. These results imply that the interaction of matrix AGEs with RAGE plays a role in the TGFβ2-mediated EMT of lens epithelial cells and suggest that the blockade of RAGE could be a strategy to prevent PCO and other age-associated fibrosis.

  18. Cholesteatoma fibroblasts promote epithelial cell proliferation through overexpression of epiregulin.

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    Mamoru Yoshikawa

    Full Text Available To investigate whether keratinocytes proliferate in response to epiregulin produced by subepithelial fibroblasts derived from middle ear cholesteatoma. Tissue samples were obtained from patients undergoing tympanoplasty. The quantitative polymerase chain reaction and immunohistochemistry were performed to examine epiregulin expression and localization in cholesteatoma tissues and retroauricular skin tissues. Fibroblasts were cultured from cholesteatoma tissues and from normal retroauricular skin. These fibroblasts were used as feeder cells for culture with a human keratinocyte cell line (PHK16-0b. To investigate the role of epiregulin in colony formation by PHK16-0b cells, epiregulin mRNA expression was knocked down in fibroblasts by using short interfering RNA and epiregulin protein was blocked with a neutralizing antibody. Epiregulin mRNA expression was significantly elevated in cholesteatoma tissues compared with that in normal retroauricular skin. Staining for epiregulin was more intense in the epithelial cells and subepithelial fibroblasts of cholesteatoma tissues than in retroauricular skin. When PHK16-0b cells were cultured with cholesteatoma fibroblasts, their colony-forming efficiency was 50% higher than when these cells were cultured with normal skin fibroblasts. Also, knockdown of epiregulin mRNA in cholesteatoma fibroblasts led to greater suppression of colony formation than knockdown in skin fibroblasts. Furthermore, the colony-forming efficiency of PHK16-0b cells was significantly reduced after treatment with an epiregulin neutralizing antibody in co-culture with cholesteatoma fibroblasts, but not in co-culture with skin fibroblasts. These results suggest that keratinocyte hyperproliferation in cholesteatoma is promoted through overexpression of epiregulin by subepithelial fibroblasts via epithelial-mesenchymal interactions, which may play a crucial role in the pathogenesis of middle ear cholesteatoma.

  19. Nonhematopoietic cells are the primary source of bone marrow-derived lung epithelial cells.

    Science.gov (United States)

    Kassmer, Susannah H; Bruscia, Emanuela M; Zhang, Ping-Xia; Krause, Diane S

    2012-03-01

    Previous studies have demonstrated that bone marrow (BM)-derived cells differentiate into nonhematopoietic cells of multiple tissues. To date, it remains unknown which population(s) of BM cells are primarily responsible for this engraftment. To test the hypothesis that nonhematopoietic stem cells in the BM are the primary source of marrow-derived lung epithelial cells, either wild-type hematopoietic or nonhematopoietic BM cells were transplanted into irradiated surfactant-protein-C (SPC)-null mice. Donor-derived, SPC-positive type 2 pneumocytes were predominantly detected in the lungs of mice receiving purified nonhematopoietic cells and were absent from mice receiving purified hematopoietic stem and progenitor cells. We conclude that cells contained in the nonhematopoietic fraction of the BM are the primary source of marrow-derived lung epithelial cells. These nonhematopoietic cells may represent a primitive stem cell population residing in adult BM.

  20. 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.

  1. Effect of Lithium on Cell Cycle Progression of Pig Airway Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    陈文书; 吴人亮; 王曦; 李媛; 郝天玲

    2004-01-01

    To investigate the effect of lithium on cell cycle progression of airway epithelial cells,primary pig tracheobronchial epithelial cells were incubated with lithium chloride (LiCl) at different concentrations (0, 5 mmol/L, and 10 mmol/L) and time (12 h, 16 h and 24 h). After the treatment, cells were counted, cell cycle profile was measured by BrdU labeling and flow cytometry, and expression of cyclin D1 and cyclin B1 were detected by Western blotting. The results showed that after 24h of 10mmol/L but not 5mmol/L LiCl treatment, proliferation of cells was slowed down as manifested by delayed confluence and cell number accumulation (P<0.05). Lithium did not change the percentage of cells in S phase (P>0.05), but 24 h incubation with 10 mmol/L LiCl induced a G2/M cell cycle arrest. Furthermore, 10mmol/L LiCl elevated cyclin D1 expression after 12h treatment, while expression of cyclin B1 increased more significantly after 24h incubation. These data demonstrate that lithium inhibits proliferation of pig airway epithelial cells by inhibiting cell cycle progression, and suggest that lithium-sensitive molecule(s) such as glycogen synthase kinase 3 may have a role in the regulation of growth of airway epithelial cells.

  2. Lens Epithelial Cell Proliferation and Cell Density in Human Age-related Cataract

    Institute of Scientific and Technical Information of China (English)

    Xialin Liu; Yizhi Liu; Jianliang Zheng; Qiang Huang; Huling Zheng

    2000-01-01

    Purpose: To discuss the potential effect of the lens epithelial cell proliferation in age-related cataract.Methods: In vitro cell proliferation was assayed by MTT method to evaluate the lens epithelial cell density, index, and proliferation capacity in normal lens and all kinds of age-related cataract. Capsulotomy specimens from all kinds of patients who underwent cataract phacoemulsification extraction surgery were compared with the lens epithelial specimens from non-cataract lenses of Eye Bank eyes.Results: Lens epithelial cell density of central anterior capsule (LECD) in female normal lens was higher than that in male, LECD in nuclear cataract( > NⅢ ) was higher than that in normal lens, but in the mature cortical cataract, LF CD was lower. Mitotic index of three kinds of age-related cataracts in vivo had no statistical difference, neither did cell proliferation capacity of cultivated cells in vitro.Conclusion: The individual difference of lens epithelial cell density and proliferation capacity in vivo may be an important underlying cause for senile cataract in the cellular level, especially for nuclear cataract.

  3. Invasion of epithelial cells by Trichinella spiralis: in vitro observations

    Directory of Open Access Journals (Sweden)

    Romarís F.

    2001-06-01

    Full Text Available It has been known for many years that Trichinella spiralis initiates infection by penetrating the columnar epithelium of the small intestine, however, the mechanisms used by the parasite in the establishment of its intramulticellular niche in the intestine are unknown. The recent demonstration that invasion also occurs in vitro when infective larvae of T. spiralis are inoculated onto cultures of epithelial cells provides a model that allows the direct observation of the process by which the parasite recognizes, invades and migrates within the epithelium. The finding that penetration of the cell membrane or Induction of plasma membrane wounds by larvae do not always result in invasion argue in favor of some kind of host-parasite communication in successful invasion. In this sense, the in vitro model of invasion provides a readily manipulated and controlled system to investigate both parasite, and host cell requirements for invasion.

  4. Involvement of the autophagy pathway in trafficking of Mycobacterium tuberculosis bacilli through cultured human type II epithelial cells.

    Science.gov (United States)

    Fine, Kari L; Metcalfe, Maureen G; White, Elizabeth; Virji, Mumtaz; Karls, Russell K; Quinn, Frederick D

    2012-09-01

    Interactions between Mycobacterium tuberculosis bacilli and alveolar macrophages have been extensively characterized, while similar analyses in epithelial cells have not been performed. In this study, we microscopically examined endosomal trafficking of M. tuberculosis strain Erdman in A549 cells, a human type II pneumocyte cell line. Immuno-electron microscopic (IEM) analyses indicate that M. tuberculosis bacilli are internalized to a compartment labelled first with Rab5 and then with Rab7 small GTPase proteins. This suggests that, unlike macrophages, M. tuberculosis bacilli traffic to late endosomes in epithelial cells. However, fusion of lysosomes with the bacteria-containing compartment appears to be inhibited, as illustrated by IEM studies employing LAMP-2 and cathepsin-L antibodies. Examination by transmission electron microscopy and IEM revealed M. tuberculosis-containing compartments surrounded by double membranes and labelled with antibodies against the autophagy marker Lc3, providing evidence for involvement and intersection of the autophagy and endosomal pathways. Interestingly, inhibition of the autophagy pathway using 3-methyladenine improved host cell viability and decreased numbers of viable intracellular bacteria recovered after 72 h post infection. Collectively, these data suggest that trafficking patterns for M. tuberculosis bacilli in alveolar epithelial cells differ from macrophages, and that autophagy is involved this process.

  5. 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.

  6. Equine tracheal epithelial membrane strips - An alternate method for examining epithelial cell arachidonic acid metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Gray, P.R.; Derksen, F.J.; Robinson, N.E.; Peter-Golden, M.L. (Michigan State Univ., East Lansing (United States) Univ. of Michigan, Ann Arbor (United States))

    1990-02-26

    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 ({le}12 x 1.5 cm) of pseudostratified columnar epithelium attached to a layer of elastic tissue 30-100 {mu}m thick. Epithelial strips (1.2 x 0.5 cm) were attached to plexiglass rods and incubated with ({sup 3}H)arachidonic acid in M199 medium (0.5 {mu}Ci/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 {mu}M calcium ionophore A23187. The extracted supernatant was processed using HPLC, resulting in peaks of radioactivity that co-eluted with authentic PGE{sub 2}, PGF{sub 2}{alpha}, and 12-HETE standards. The greatest activity corresponded to the PGE{sub 2} and PGF{sub 2}{alpha} standards, which is a similar pattern to that reported for cultured human tracheal epithelium.

  7. Parietal Epithelial Cell Activation Marker in Early Recurrence of FSGS in the Transplant

    NARCIS (Netherlands)

    Fatima, H.; Moeller, M.J.; Smeets, B.; Yang, H.C.; D'Agati, V.D.; Alpers, C.E.; Fogo, A.B.

    2012-01-01

    BACKGROUND AND OBJECTIVES: Podocyte loss is key in glomerulosclerosis. Activated parietal epithelial cells are proposed to contribute to pathogenesis of glomerulosclerosis and may serve as stem cells that can transition to podocytes. CD44 is a marker for activated parietal epithelial cells. This stu

  8. Hypoxia is a key regulator of limbal epithelial stem cell growth and differentiation

    DEFF Research Database (Denmark)

    Bath, Chris; Yang, Sufang; Muttuvelu, Danson

    2013-01-01

    The aim of this study was to determine whether the growth and differentiation of limbal epithelial stem cell cultures could be controlled through manipulation of the oxygen tension. Limbal epithelial cells were isolated from corneoscleral disks, and cultured using either feeder cells in a growth ...

  9. WU Polyomavirus in Respiratory Epithelial Cells from Lung Transplant Patient with Job Syndrome

    Science.gov (United States)

    Siebrasse, Erica A.; Pastrana, Diana V.; Nguyen, Nang L.; Wang, Annie; Roth, Mark J.; Holland, Steven M.; Freeman, Alexandra F.; McDyer, John; Buck, Christopher B.

    2015-01-01

    We detected WU polyomavirus (WUPyV) in a bronchoalveolar lavage sample from lungs transplanted into a recipient with Job syndrome by using immunoassays specific for the WUPyV viral protein 1. Co-staining for an epithelial cell marker identified most WUPyV viral protein 1–positive cells as respiratory epithelial cells. PMID:25531075

  10. Epithelial Cell Coculture Models for Studying Infectious Diseases: Benefits and Limitations

    Directory of Open Access Journals (Sweden)

    Benjamin L. Duell

    2011-01-01

    Full Text Available Countless in vitro cell culture models based on the use of epithelial cell types of single lineages have been characterized and have provided insight into the mechanisms of infection for various microbial pathogens. Diverse culture models based on disease-relevant mucosal epithelial cell types derived from gastrointestinal, genitourinary, and pulmonary organ systems have delineated many key host-pathogen interactions that underlie viral, parasitic, and bacterial disease pathogenesis. An alternative to single lineage epithelial cell monoculture, which offers more flexibility and can overcome some of the limitations of epithelial cell culture models based on only single cell types, is coculture of epithelial cells with other host cell types. Various coculture models have been described, which incorporate epithelial cell types in culture combination with a wide range of other cell types including neutrophils, eosinophils, monocytes, and lymphocytes. This paper will summarize current models of epithelial cell coculture and will discuss the benefits and limitations of epithelial cell coculture for studying host-pathogen dynamics in infectious diseases.

  11. 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.

  12. Temporal Evolution of the Plasma Sheath Surrounding Solar Cells in Low Earth Orbit

    Science.gov (United States)

    Willis, Emily M.; Pour, Maria Z. A.

    2017-01-01

    High voltage solar array interactions with the space environment can have a significant impact on array performance and spacecraft charging. Over the past 10 years, data from the International Space Station has allowed for detailed observations of these interactions over long periods of time. Some of the surprising observations have been floating potential transients, which were not expected and are not reproduced by existing models. In order to understand the underlying processes producing these transients, the temporal evolution of the plasma sheath surrounding the solar cells in low Earth orbit is being investigated. This study includes lumped element modeling and particle-in-cell simulation methods. This presentation will focus on recent results from the on-going investigations.

  13. The PCP pathway regulates Baz planar distribution in epithelial cells

    Science.gov (United States)

    Aigouy, Benoit; Le Bivic, André

    2016-01-01

    The localisation of apico-basal polarity proteins along the Z-axis of epithelial cells is well understood while their distribution in the plane of the epithelium is poorly characterised. Here we provide a systematic description of the planar localisation of apico-basal polarity proteins in the Drosophila ommatidial epithelium. We show that the adherens junction proteins Shotgun and Armadillo, as well as the baso-lateral complexes, are bilateral, i.e. present on both sides of cell interfaces. In contrast, we report that other key adherens junction proteins, Bazooka and the myosin regulatory light chain (Spaghetti squash) are unilateral, i.e. present on one side of cell interfaces. Furthermore, we demonstrate that planar cell polarity (PCP) and not the apical determinants Crumbs and Par-6 control Bazooka unilaterality in cone cells. Altogether, our work unravels an unexpected organisation and combination of apico-basal, cytoskeletal and planar polarity proteins that is different on either side of cell-cell interfaces and unique for the different contacts of the same cell. PMID:27624969

  14. Infectious salmon anaemia virus infection of Atlantic salmon gill epithelial cells

    Directory of Open Access Journals (Sweden)

    Weli Simon

    2013-01-01

    Full Text Available Abstract Infectious salmon anaemia virus (ISAV, a member of the Orthomyxoviridae family, infects and causes disease in farmed Atlantic salmon (Salmo salar L.. Previous studies have shown Atlantic salmon endothelial cells to be the primary targets of ISAV infection. However, it is not known if cells other than endothelial cells play a role in ISAV tropism. To further assess cell tropism, we examined ISAV infection of Atlantic salmon gill epithelial cells in vivo and in vitro. We demonstrated the susceptibility of epithelial cells to ISAV infection. On comparison of primary gill epithelial cell cultures with ISAV permissive fish cell cultures, we found the virus yield in primary gill epithelial cells to be comparable with that of salmon head kidney (SHK-1 cells, but lower than TO or Atlantic salmon kidney (ASK-II cells. Light and transmission electron microscopy (TEM revealed that the primary gill cells possessed characteristics consistent with epithelial cells. Virus histochemistry showed that gill epithelial cells expressed 4-O-acetylated sialic acid which is recognized as the ISAV receptor. To the best of our knowledge, this is the first demonstration of ISAV infection in Atlantic salmon primary gill epithelial cells. This study thus broadens our understanding of cell tropism and transmission of ISAV in Atlantic salmon.

  15. 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.

  16. NK cells are necessary for recovery of corneal CD11c+ dendritic cells after epithelial abrasion injury

    Science.gov (United States)

    Mechanisms controlling CD11c(+) MHCII(+) DCs during corneal epithelial wound healing were investigated in a murine model of corneal abrasion. Selective depletion of NKp46(+) CD3- NK cells that normally migrate into the cornea after epithelial abrasion resulted in >85% reduction of the epithelial CD1...

  17. Cyclin-dependent kinases regulate apoptosis of intestinal epithelial cells.

    Science.gov (United States)

    Bhattacharya, Sujoy; Ray, Ramesh M; Johnson, Leonard R

    2014-03-01

    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 (Caco-2) cells with mutant p53, treatment with either AZD5438 or NU6140 blocked proliferation, albeit more robustly with AZD5438. Both Cdk inhibitors induced apoptosis in Caco-2 cells in a p53-independent manner. In serum starved quiescent IEC-6 cells, both AZD5438 and NU6140 decreased TNF-α/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.

  18. Cytokeratin 18 is necessary for initiation of TGF-β1-induced epithelial-mesenchymal transition in breast epithelial cells.

    Science.gov (United States)

    Jung, Hyejung; Kim, Bomin; Moon, Byung In; Oh, Eok-Soo

    2016-12-01

    During epithelial-mesenchymal transition (EMT), epithelial cells lose key phenotypic markers (e.g., E-cadherin and cytokeratin 18) and acquire mesenchymal markers (e.g., N-cadherin and vimentin). Although the loss of cytokeratin 18 is a hallmark of EMT, the regulatory role of cytokeratin 18 in EMT is not yet fully understood. Here, we report that cytokeratin 18 is involved in the regulation of transforming growth factor-beta1 (TGF-β1)-induced EMT in breast epithelial cells. When MCF10A cells were treated with TGF-β1 for 24 h, considerable morphological changes, indicative of the early stages of EMT (e.g., loss of cell-cell contact), were observed and cytokeratin 18 was downregulated. However, E-cadherin levels were not altered until a later time point. This suggests that cytokeratin 18 may play an active role during the earlier stages of EMT. Consistent with this notion, siRNA-mediated knockdown of cytokeratin 18 delayed TGF-β1-mediated EMT, and the associated downregulation of E-cadherin reduced the phosphorylation/nuclear localization of smad 2/3 and decreased the expression levels of snail and slug (which inhibit E-cadherin expression in epithelial cells as an early response to TGF-β1). Taken together, these results suggest that cytokeratin 18 critically contributes to initiating TGF-β1-induced EMT via the smad 2/3-mediated regulation of snail and slug expression in breast epithelial cells.

  19. Ferritinophagy drives uropathogenic Escherichia coli persistence in bladder epithelial cells.

    Science.gov (United States)

    Bauckman, Kyle A; Mysorekar, Indira U

    2016-05-01

    Autophagy is a cellular recycling pathway, which in many cases, protects host cells from infections by degrading pathogens. However, uropathogenic Escherichia coli (UPEC), the predominant cause of urinary tract infections (UTIs), persist within the urinary tract epithelium (urothelium) by forming reservoirs within autophagosomes. Iron is a critical nutrient for both host and pathogen, and regulation of iron availability is a key host defense against pathogens. Iron homeostasis depends on the shuttling of iron-bound ferritin to the lysosome for recycling, a process termed ferritinophagy (a form of selective autophagy). Here, we demonstrate for the first time that UPEC shuttles with ferritin-bound iron into the autophagosomal and lysosomal compartments within the urothelium. Iron overload in urothelial cells induces ferritinophagy in an NCOA4-dependent manner causing increased iron availability for UPEC, triggering bacterial overproliferation and host cell death. Addition of even moderate levels of iron is sufficient to increase and prolong bacterial burden. Furthermore, we show that lysosomal damage due to iron overload is the specific mechanism causing host cell death. Significantly, we demonstrate that host cell death and bacterial burden can be reversed by inhibition of autophagy or inhibition of iron-regulatory proteins, or chelation of iron. Together, our findings suggest that UPEC persist in host cells by taking advantage of ferritinophagy. Thus, modulation of iron levels in the bladder may provide a therapeutic avenue to controlling UPEC persistence, epithelial cell death, and recurrent UTIs.

  20. 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

  1. High-grade epithelial-myoepithelial carcinoma of the parotid gland with mucous cell differentiation.

    Science.gov (United States)

    Sentani, Kazuhiro; Ogawa, Ikuko; Uraoka, Naohiro; Ikeda, Masayuki; Hayashi, Naoki; Hattori, Takuya; Hattori, Yui; Oue, Naohide; Takata, Takashi; Yasui, Wataru

    2015-09-01

    Epithelial-myoepithelial carcinoma (EMC) is a rare salivary gland tumor with a low-grade malignancy, and EMC with high-grade histopathological features is exceedingly rare. Furthermore, EMC with intracellular mucin is also extremely rare. We report an uncommon case of a high-grade EMC of the parotid gland with mucous cell differentiation in a 66-year old Japanese woman who noticed a right palpable parotid mass increasing in size within a one-year period. The cytological specimen showed a focally biphasic structure and included isolated or discohesive piled-up clusters with hyaline globules surrounded by neoplastic cells with nuclear atypia. The gross examination revealed a relatively well-demarcated, multinodular gray-whitish and solid mass. Histologically, the tumor consisted of variably sized solid nests or trabeculae with central necrosis and increased mitotic activity, and invaded into adjacent skeletal muscles. Immunohistochemically, the biphasic ductal and myoepithelial differentiation of this tumor confirmed the diagnosis of high-grade EMC. Furthermore, numerous small nests with d-PAS and alcian blue-positive mucous cells predominated in about 5% of the whole tumor, and these mucous cells were encompassed by neoplastic myoepithelial cells. We should recognize this variant of EMC because we can't rule out the possibility of EMC even in the presence of mucous cells.

  2. Effect of bacteria on the wound healing behavior of oral epithelial cells.

    Science.gov (United States)

    Bhattacharya, Rupa; Xu, Fanxing; Dong, Guangyu; Li, Shuai; Tian, Chen; Ponugoti, Bhaskar; Graves, Dana T

    2014-01-01

    Wounded tissue offers opportunity to microflora to adhere, colonize, invade and infect surrounding healthy tissue. The bacteria of the oral cavity have the potential to alter the wound healing process by interacting with keratinocytes. The aim of this study was to investigate mechanisms through which oral bacteria may influence re-epithelialization by interacting with gingival keratinocytes. By an in vitro scratch assay we demonstrate that primary gingival keratinocytes have impaired closure when exposed to two well characterized oral bacteria, P. gingivalis, and to a lesser extent, F. nucleatum. P. gingivalis reduced wound closure by ∼ 40%, which was partially dependent on proteolytic activity, and bacteria was still present within infected cells 9 days later despite exposure to bacteria for only 24 h. Both oral bacteria caused keratinocyte apoptosis at the wound site with cell death being greatest at the wound edge. P. gingivalis and F. nucleatum adversely affected cell proliferation and the effect also had a spatial component being most striking at the edge. The impact of the bacteria was long lasting even when exposure was brief. Cell migration was compromised in bacteria challenged keratinocytes with P. gingivalis having more severe effect (pbacteria challenged cells showed that P. gingivalis and to a lesser extent F. nucleatum significantly downregulated cell cycle genes cyclin1, CDK1, and CDK4 (pcell migration such as integrin beta-3 and -6 were significantly downregulated by P. gingivalis (p<0.05).

  3. Cigarette smoke impairs airway epithelial barrier function and cell-cell contact recovery.

    Science.gov (United States)

    Heijink, I H; Brandenburg, S M; Postma, D S; van Oosterhout, A J M

    2012-02-01

    Cigarette smoking, the major cause of chronic obstructive pulmonary disease (COPD), induces aberrant airway epithelial structure and function. The underlying mechanisms are unresolved so far. We studied effects of cigarette smoke extract (CSE) on epithelial barrier function and wound regeneration in human bronchial epithelial 16HBE cells and primary bronchial epithelial cells (PBECs) from COPD patients, nonsmokers and healthy smokers. We demonstrate that CSE rapidly and transiently impairs 16HBE barrier function, largely due to disruption of cell-cell contacts. CSE induced a similar, but stronger and more sustained, defect in PBECs. Application of the specific epidermal growth factor receptor (EGFR) inhibitor AG1478 showed that EGFR activation contributes to the CSE-induced defects in both 16HBE cells and PBECs. Furthermore, our data indicate that the endogenous protease calpain mediates these defects through tight junction protein degradation. CSE also delayed the reconstitution of 16HBE intercellular contacts during wound healing and attenuated PBEC barrier function upon wound regeneration. These findings were comparable between PBECs from smokers, healthy smokers and COPD patients. In conclusion, we demonstrate for the first time that CSE reduces epithelial integrity, probably by EGFR and calpain-dependent disruption of intercellular contacts. This may increase susceptibility to environmental insults, e.g. inhaled pathogens. Thus, EGFR may be a promising target for therapeutic strategies to improve mucosal barrier function in cigarette smoking-related disease.

  4. Oxidant-induced corticosteroid unresponsiveness in human bronchial epithelial cells

    NARCIS (Netherlands)

    Heijink, Irene; van Oosterhout, Antoon; Kliphuis, Nathalie; Jonker, Marnix; Hoffmann, Roland; Telenga, Eef; Klooster, Karin; Slebos, Dirk-Jan; ten Hacken, Nick; Postma, Dirkje; van den Berge, Maarten

    2014-01-01

    Background We hypothesised that increased oxidative stress, as present in the airways of asthma and chronic obstructive pulmonary disease (COPD) patients, induces epithelial damage and reduces epithelial responsiveness to suppressive effects of corticosteroids on proinflammatory cytokine production

  5. Oxidant-induced corticosteroid unresponsiveness in human bronchial epithelial cells

    NARCIS (Netherlands)

    Heijink, Irene; van Oosterhout, Antoon; Kliphuis, Nathalie; Jonker, Marnix; Hoffmann, Roland; Telenga, Eef; Klooster, Karin; Slebos, Dirk-Jan; ten Hacken, Nick; Postma, Dirkje; van den Berge, Maarten

    2014-01-01

    Background We hypothesised that increased oxidative stress, as present in the airways of asthma and chronic obstructive pulmonary disease (COPD) patients, induces epithelial damage and reduces epithelial responsiveness to suppressive effects of corticosteroids on proinflammatory cytokine production

  6. Water fluxes through aquaporin-9 prime epithelial cells for rapid wound healing

    DEFF Research Database (Denmark)

    Karlsson, T.; Lagerholm, B. C.; Vikstrom, E.

    2013-01-01

    about the impact on migrating epithelial sheets during wound healing and epithelial renewal. Here, we investigate and compare the effects of AQP9 on single cell and epithelial sheet migration. To achieve this, MDCK-1 cells stably expressing AQP9 were subjected to migration assessment. We found that AQP9...... wound healing based on AQP-induced swelling and expansion of the monolayer. (C) 2012 Elsevier Inc. All rights reserved....

  7. Human Epithelial Cells Discriminate between Commensal and Pathogenic Interactions with Candida albicans.

    Science.gov (United States)

    Rast, Timothy J; Kullas, Amy L; Southern, Peter J; Davis, Dana A

    2016-01-01

    The commensal fungus, Candida albicans, can cause life-threatening infections in at risk individuals. C. albicans colonizes mucosal surfaces of most people, adhering to and interacting with epithelial cells. At low concentrations, C. albicans is not pathogenic nor does it cause epithelial cell damage in vitro; at high concentrations, C. albicans causes mucosal infections and kills epithelial cells in vitro. Here we show that while there are quantitative dose-dependent differences in exposed epithelial cell populations, these reflect a fundamental qualitative difference in host cell response to C. albicans. Using transcriptional profiling experiments and real time PCR, we found that wild-type C. albicans induce dose-dependent responses from a FaDu epithelial cell line. However, real time PCR and Western blot analysis using a high dose of various C. albicans strains demonstrated that these dose-dependent responses are associated with ability to promote host cell damage. Our studies support the idea that epithelial cells play a key role in the immune system by monitoring the microbial community at mucosal surfaces and initiating defensive responses when this community is dysfunctional. This places epithelial cells at a pivotal position in the interaction with C. albicans as epithelial cells themselves promote C. albicans stimulated damage.

  8. [In vitro development of rifampicin resistance in the epithelial cells].

    Science.gov (United States)

    Erokhina, M V; Aleksandrova, E A

    2006-01-01

    It has been first in vitro demonstrated on a model of epithelial cells that rifampicin may develop not only at the level of Mycobacterium tuberculosis, but also at the level of somatic cells. The mechanism of this phenomenon, its specificity (whether cross resistance to other antituberculous agents will occur), the way it puts into effect under the conditions of a microorganism, and how promptly it may be gone after discontinuation of the drug remain unknown. The effect of rifampicin on the functional activity of Pgp is an important factor that influences as a result not only the absorbability of drugs, but also normal transport processes in the body. These aspects seem to be topical and are the subject for further studies. The authors have obtained an epithelial cell line that resides in the presence of 100 microg/ml of rifampicin and that is 2-2.5 times more resistant to the drug as compared with the parental line. The cells of this line are 2-2.5 times more active in discharging the substrate rhodamine-123 for P-glycoprotein than those of the parental line, which suggests the enhanced functional activity of P-glycoprotein. The presence of P-glycoprotein in this line is confirmed by the action of this protein-specific blocker verapamil. At the same time rifampicin is not a substract for P-glycoprotein. Therefore, the mechanism of rifampicin resistance is unassociated with the functional activity of P-glycoprotein. The mechanism of the resistance remains open. At the same concentration (100 microg/ml), rifampicin can block the functional activity of P-glycoprotein. These results suggest the double mechanism of rifampicin in its long presence in the culture medium: as an inductor and a blocker of P-glycoprotein functional activity. The findings point to the fact that the pharmacokinetics of rifampicin and co-administered dtugs may change during their long use.

  9. Substrate stiffness regulates extracellular matrix deposition by alveolar epithelial cells

    Directory of Open Access Journals (Sweden)

    Jessica L Eisenberg

    2011-01-01

    Full Text Available Jessica L Eisenberg1,2, Asmahan Safi3, Xiaoding Wei3, Horacio D Espinosa3, GR Scott Budinger2, Desire Takawira1, Susan B Hopkinson1, Jonathan CR Jones1,21Department of Cell and Molecular Biology, 2Division of Pulmonary Medicine, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA; 3Department of Mechanical Engineering, Northwestern University, Evanston, IL, USAAim: The aim of the study was to address whether a stiff substrate, a model for pulmonary fibrosis, is responsible for inducing changes in the phenotype of alveolar epithelial cells (AEC in the lung, including their deposition and organization of extracellular matrix (ECM proteins.Methods: Freshly isolated lung AEC from male Sprague Dawley rats were seeded onto polyacrylamide gel substrates of varying stiffness and analyzed for expression and organization of adhesion, cytoskeletal, differentiation, and ECM components by Western immunoblotting and confocal immunofluorescence microscopy.Results: We observed that substrate stiffness influences cell morphology and the organization of focal adhesions and the actin cytoskeleton. Surprisingly, however, we found that substrate stiffness has no influence on the differentiation of type II into type I AEC, nor does increased substrate stiffness lead to an epithelial–mesenchymal transition. In contrast, our data indicate that substrate stiffness regulates the expression of the α3 laminin subunit by AEC and the organization of both fibronectin and laminin in their ECM.Conclusions: An increase in substrate stiffness leads to enhanced laminin and fibronectin assembly into fibrils, which likely contributes to the disease phenotype in the fibrotic lung.Keywords: alveolar epithelial cells, fibrosis, extracellular matrix, substrate stiffness

  10. Interactions between airway epithelial cells and dendritic cells during viral infections using an in vitro co-culture model

    Science.gov (United States)

    Rationale: Historically, single cell culture models have been limited in pathological and physiological relevance. A co-culture model of dendritic cells (DCs) and differentiated human airway epithelial cells was developed to examine potential interactions between these two cell t...

  11. Different Sensitivities to Apoptotic Induction by Camptothecin between Normal and Senescent Lens Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    Haike Guo; Haiying Jin; Liya Wang; Hongyang Zhang; Xin Yang

    2002-01-01

    Purpose: To investigate whether normal and senescent lens epithelial cells have different defense abilities to apoptotic induction factor in vitro.Methods: Rabbit lens epithelial cells were cultured, passed. When reaching confluence, cells from the first and seventh passage were stained by x-gal staining to detect cell senescence. Cell apoptosis was detected by TUNEL(Roche).10μmol/L camptothecin was used to induce cell apoptosis from the lens epithelial cells of the first and seventh passage to distinguish different sensitivities to apoptotic induction factor between normal and senescent cells.Results: The senescent cells (41.17% ± 5.24% ) were detected in the lens epithelial cell culture of the seventh passage, which are higher than those of the first passage (0.98% ±0. 39% ). There was no apoptotic cell detected in the cell cultures undisturbed. Exposure of the first passage cells to camptothecin resulted in death of approximately 23.87% ± 3.45% of the cells during a 36 hour exposure period. In contrast, significantly more lens epithelial cells died through the apoptosis (38.29% ±4. 01% ) from the seventh passage.Conclusion: Senescent cells increased with cell passage. Senescence lens epithelial cells do not undergo apoptosis if they were not disturbed. But the vulnerabilities to apoptotic induction between health and senescence cells were different.

  12. Rhinovirus infection induces cytotoxicity and delays wound healing in bronchial epithelial cells

    Directory of Open Access Journals (Sweden)

    Constantopoulos Andreas G

    2005-10-01

    Full Text Available Abstract Background Human rhinoviruses (RV, the most common triggers of acute asthma exacerbations, are considered not cytotoxic to the bronchial epithelium. Recent observations, however, have questioned this knowledge. The aim of this study was to evaluate the ability of RV to induce epithelial cytotoxicity and affect epithelial repair in-vitro. Methods Monolayers of BEAS-2B bronchial epithelial cells, seeded at different densities were exposed to RV serotypes 1b, 5, 7, 9, 14, 16. Cytotoxicity was assessed chromatometrically. Epithelial monolayers were mechanically wounded, exposed or not to RV and the repopulation of the damaged area was assessed by image analysis. Finally epithelial cell proliferation was assessed by quantitation of proliferating cell nuclear antigen (PCNA by flow cytometry. Results RV1b, RV5, RV7, RV14 and RV16 were able to induce considerable epithelial cytotoxicity, more pronounced in less dense cultures, in a cell-density and dose-dependent manner. RV9 was not cytotoxic. Furthermore, RV infection diminished the self-repair capacity of bronchial epithelial cells and reduced cell proliferation. Conclusion RV-induced epithelial cytotoxicity may become considerable in already compromised epithelium, such as in the case of asthma. The RV-induced impairment on epithelial proliferation and self-repair capacity may contribute to the development of airway remodeling.

  13. Long-term homeostasis and wound healing in an in vitro epithelial stem cell niche model

    Science.gov (United States)

    Miyashita, Hideyuki; Niwano, Hiroko; Yoshida, Satoru; Hatou, Shin; Inagaki, Emi; Tsubota, Kazuo; Shimmura, Shigeto

    2017-01-01

    Cultures of epithelial cells are limited by the proliferative capacity of primary cells and cell senescence. Herein we show that primary human epithelial cell sheets cultured without dermal equivalents maintained homeostasis in vitro for at least 1 year. Transparency of these sheets enabled live observation of pigmented melanocytes and Fluorescent Ubiquitination-based Cell Cycle Indicator (FUCCI) labeled epithelial cells during wound healing. Cell turn over and KRT15 expression pattern stabilized within 3 months, when KRT15 bright clusters often associated with niche-like melanocytes became apparent. EdU labels were retained in a subset of epithelial cells and melanocytes after 6 months chasing, suggesting their slow cell cycling property. FUCCI-labeling demonstrated robust cell migration and proliferation following wounding. Transparency and long-term (1 year) homeostasis of this model will be a powerful tool for the study of wound healing and cell linage tracing. PMID:28233843

  14. 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.

  15. Uterine epithelial cell proliferation and endometrial hyperplasia: evidence from a mouse model.

    Science.gov (United States)

    Gao, Yang; Li, Shu; Li, Qinglei

    2014-08-01

    In the uterus, epithelial cell proliferation changes during the estrous cycle and pregnancy. Uncontrolled epithelial cell proliferation results in implantation failure and/or cancer development. Transforming growth factor-β (TGF-β) signaling is a fundamental regulator of diverse biological processes and is indispensable for multiple reproductive functions. However, the in vivo role of TGF-β signaling in uterine epithelial cells remains poorly defined. We have shown that in the uterus, conditional deletion of the Type 1 receptor for TGF-β (Tgfbr1) using anti-Müllerian hormone receptor type 2 (Amhr2) Cre leads to myometrial defects. Here, we describe enhanced epithelial cell proliferation by immunostaining of Ki67 in the uteri of these mice. The aberration culminated in endometrial hyperplasia in aged females. To exclude the potential influence of ovarian steroid hormones, the proliferative status of uterine epithelial cells was assessed following ovariectomy. Increased uterine epithelial cell proliferation was also revealed in ovariectomized Tgfbr1 Amhr2-Cre conditional knockout mice. We further demonstrated that transcript levels for fibroblast growth factor 10 (Fgf10) were markedly up-regulated in Tgfbr1 Amhr2-Cre conditional knockout uteri. Consistently, treatment of primary uterine stromal cells with TGF-β1 significantly reduced Fgf10 mRNA expression. Thus, our findings suggest a potential involvement of TGFBR1-mediated signaling in the regulation of uterine epithelial cell proliferation, and provide genetic evidence supporting the role of uterine epithelial cell proliferation in the pathogenesis of endometrial hyperplasia.

  16. Activated alveolar epithelial cells initiate fibrosis through autocrine and paracrine secretion of connective tissue growth factor.

    Science.gov (United States)

    Yang, Jibing; Velikoff, Miranda; Canalis, Ernesto; Horowitz, Jeffrey C; Kim, Kevin K

    2014-04-15

    Fibrogenesis involves a pathological accumulation of activated fibroblasts and extensive matrix remodeling. Profibrotic cytokines, such as TGF-β, stimulate fibroblasts to overexpress fibrotic matrix proteins and induce further expression of profibrotic cytokines, resulting in progressive fibrosis. Connective tissue growth factor (CTGF) is a profibrotic cytokine that is indicative of fibroblast activation. Epithelial cells are abundant in the normal lung, but their contribution to fibrogenesis remains poorly defined. Profibrotic cytokines may activate epithelial cells with protein expression and functions that overlap with the functions of active fibroblasts. We found that alveolar epithelial cells undergoing TGF-β-mediated mesenchymal transition in vitro were also capable of activating lung fibroblasts through production of CTGF. Alveolar epithelial cell expression of CTGF was dramatically reduced by inhibition of Rho signaling. CTGF reporter mice demonstrated increased CTGF promoter activity by lung epithelial cells acutely after bleomycin in vivo. Furthermore, mice with lung epithelial cell-specific deletion of CTGF had an attenuated fibrotic response to bleomycin. These studies provide direct evidence that epithelial cell activation initiates a cycle of fibrogenic effector cell activation during progressive fibrosis. Therapy targeted at epithelial cell production of CTGF offers a novel pathway for abrogating this progressive cycle and limiting tissue fibrosis.

  17. Photodynamic injury of isolated crayfish neuron and surrounding glial cells: the role of p53

    Science.gov (United States)

    Sharifulina, S. A.; Uzdensky, A. B.

    2015-03-01

    The pro-apoptotic transcription factor p53 is involved in cell responses to injurious impacts. Using its inhibitor pifithrin- α and activators tenovin-1, RITA and WR-1065, we studied its potential participation in inactivation and death of isolated crayfish mechanoreceptor neuron and satellite glial cells induced by photodynamic treatment, a strong inducer of oxidative stress. In dark, p53 activation by tenovin-1 or WR-1065 shortened activity of isolated neurons. Tenovin-1 and WR-1065 induced apoptosis of glial cells, whereas pifithrin-α was anti-apoptotic. Therefore, p53 mediated glial apoptosis and suppression of neuronal activity after axotomy. Tenovin-1 but not other p53 modulators induced necrosis of axotomized neurons and surrounding glia, possibly, through p53-independent pathway. Under photodynamic treatment, p53 activators tenovin-1 and RITA enhanced glial apoptosis indicating the pro-apoptotic activity of p53. Photoinduced necrosis of neurons and glia was suppressed by tenovin-1 and, paradoxically, by pifithrin-α. Modulation of photoinduced changes in the neuronal activity and necrosis of neurons and glia was possibly p53-independent. The different effects of p53 modulators on neuronal and glial responses to axotomy and photodynamic impact were apparently associated with different signaling pathways in neurons and glial cells.

  18. Myosin Id is required for planar cell polarity in ciliated tracheal and ependymal epithelial cells.

    Science.gov (United States)

    Hegan, Peter S; Ostertag, Eric; Geurts, Aron M; Mooseker, Mark S

    2015-10-01

    In wild type (WT) tracheal epithelial cells, ciliary basal bodies are oriented such that all cilia on the cell surface beat in the same upward direction. This precise alignment of basal bodies and, as a result, the ciliary axoneme, is termed rotational planar cell polarity (PCP). Rotational PCP in the multi-ciliated epithelial cells of the trachea is perturbed in rats lacking myosin Id (Myo1d). Myo1d is localized in the F-actin and basal body rich subapical cortex of the ciliated tracheal epithelial cell. Scanning and transmission electron microscopy of Myo1d knock out (KO) trachea revealed that the unidirectional bending pattern is disrupted. Instead, cilia splay out in a disordered, often radial pattern. Measurement of the alignment axis of the central pair axonemal microtubules was much more variable in the KO, another indicator that rotational PCP is perturbed. The asymmetric localization of the PCP core protein Vangl1 is lost. Both the velocity and linearity of cilia-driven movement of beads above the tracheal mucosal surface was impaired in the Myo1d KO. Multi-ciliated brain ependymal epithelial cells exhibit a second form of PCP termed translational PCP in which basal bodies and attached cilia are clustered at the anterior side of the cell. The precise asymmetric clustering of cilia is disrupted in the ependymal cells of the Myo1d KO rat. While basal body clustering is maintained, left-right positioning of the clusters is lost.

  19. Isolation, growth, and characterization of human renal epithelial cells using traditional and 3D methods.

    Science.gov (United States)

    Gildea, John J; McGrath, Helen E; Van Sciver, Robert E; Wang, Dora Bigler; Felder, Robin A

    2013-01-01

    The kidney is a highly heterogeneous organ that is responsible for fluid and electrolyte balance. Much interest is focused on determining the function of specific renal epithelial cells in humans, which can only be accomplished through the isolation and growth of nephron segment-specific epithelial cells. However, human renal epithelial cells are notoriously difficult to maintain in culture. This chapter describes the isolation, growth, immortalization, and characterization of the human renal proximal tubule cell. In addition, we describe new paradigms in 3D cell culture which allow the cells to maintain more in vivo-like morphology and function.

  20. Culture of Oral Mucosal Epithelial Cells for the Purpose of Treating Limbal Stem Cell Deficiency

    Directory of Open Access Journals (Sweden)

    Tor Paaske Utheim

    2016-03-01

    Full Text Available The cornea is critical for normal vision as it allows allowing light transmission to the retina. The corneal epithelium is renewed by limbal epithelial cells (LEC, which are located in the periphery of the cornea, the limbus. Damage or disease involving LEC may lead to various clinical presentations of limbal stem cell deficiency (LSCD. Both severe pain and blindness may result. Transplantation of cultured autologous oral mucosal epithelial cell sheet (CAOMECS represents the first use of a cultured non-limbal autologous cell type to treat this disease. Among non-limbal cell types, CAOMECS and conjunctival epithelial cells are the only laboratory cultured cell sources that have been explored in humans. Thus far, the expression of p63 is the only predictor of clinical outcome following transplantation to correct LSCD. The optimal culture method and substrate for CAOMECS is not established. The present review focuses on cell culture methods, with particular emphasis on substrates. Most culture protocols for CAOMECS used amniotic membrane as a substrate and included the xenogeneic components fetal bovine serum and murine 3T3 fibroblasts. However, it has been demonstrated that tissue-engineered epithelial cell sheet grafts can be successfully fabricated using temperature-responsive culture surfaces and autologous serum. In the studies using different substrates for culture of CAOMECS, the quantitative expression of p63 was generally poorly reported; thus, more research is warranted with quantification of phenotypic data. Further research is required to develop a culture system for CAOMECS that mimics the natural environment of oral/limbal/corneal epithelial cells without the need for undefined foreign materials such as serum and feeder cells.

  1. Culture of Oral Mucosal Epithelial Cells for the Purpose of Treating Limbal Stem Cell Deficiency.

    Science.gov (United States)

    Utheim, Tor Paaske; Utheim, Øygunn Aass; Khan, Qalb-E-Saleem; Sehic, Amer

    2016-03-01

    The cornea is critical for normal vision as it allows allowing light transmission to the retina. The corneal epithelium is renewed by limbal epithelial cells (LEC), which are located in the periphery of the cornea, the limbus. Damage or disease involving LEC may lead to various clinical presentations of limbal stem cell deficiency (LSCD). Both severe pain and blindness may result. Transplantation of cultured autologous oral mucosal epithelial cell sheet (CAOMECS) represents the first use of a cultured non-limbal autologous cell type to treat this disease. Among non-limbal cell types, CAOMECS and conjunctival epithelial cells are the only laboratory cultured cell sources that have been explored in humans. Thus far, the expression of p63 is the only predictor of clinical outcome following transplantation to correct LSCD. The optimal culture method and substrate for CAOMECS is not established. The present review focuses on cell culture methods, with particular emphasis on substrates. Most culture protocols for CAOMECS used amniotic membrane as a substrate and included the xenogeneic components fetal bovine serum and murine 3T3 fibroblasts. However, it has been demonstrated that tissue-engineered epithelial cell sheet grafts can be successfully fabricated using temperature-responsive culture surfaces and autologous serum. In the studies using different substrates for culture of CAOMECS, the quantitative expression of p63 was generally poorly reported; thus, more research is warranted with quantification of phenotypic data. Further research is required to develop a culture system for CAOMECS that mimics the natural environment of oral/limbal/corneal epithelial cells without the need for undefined foreign materials such as serum and feeder cells.

  2. 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

  3. Human immunodeficiency virus type 1 infection of human uterine epithelial cells: viral shedding and cell contact-mediated infectivity.

    Science.gov (United States)

    Asin, Susana N; Wildt-Perinic, Dunja; Mason, Sarah I; Howell, Alexandra L; Wira, Charles R; Fanger, Michael W

    2003-05-15

    We examined the mechanism of human immunodeficiency virus (HIV) type 1 infection of human uterine epithelial cells to gain a clearer understanding of the events by which HIV-1 infects cells within the female reproductive tract. We demonstrated that these cells can be productively infected by HIV-1 and that infection is associated with viral RNA reverse transcription, DNA transcription, and secretion of infectious virus. Levels of viral DNA and secreted virus decreased gradually after infection. Moreover, virus released by the uterine epithelial cells shortly after infection was able to infect human T cell lines, but virus released later did not. In contrast, human CD4(+) T cell lines were infected after cocultivation with epithelial cells at both early and late stages of infection. These data demonstrated that HIV-1 infects human epithelial cells of upper reproductive tract origin and that productive viral infection of epithelial cells may be an important mechanism of transmission of HIV-1 infection in women.

  4. The effect of calprotectin on TSLP and IL-25 production from airway epithelial cells

    Directory of Open Access Journals (Sweden)

    Tomohisa Kato

    2017-04-01

    Conclusions: These results indicate that calprotectin enhances the allergen-induced Th2-type inflammatory responses in airway epithelial cells via the secretion of TSLP and IL-25, and that calprotectin secreted by the epithelial cells may be involved in the pathogenesis of ECRS.

  5. Cytotoxicity and induction of inflammation by pepsin in Acid in bronchial epithelial cells

    NARCIS (Netherlands)

    Bathoorn, Erik; Daly, Paul; Gaiser, Birgit; Sternad, Karl; Poland, Craig; Macnee, William; Drost, Ellen M

    2011-01-01

    Introduction. Gastroesophageal reflux has been associated with chronic inflammatory diseases and may be a cause of airway remodelling. Aspiration of gastric fluids may cause damage to airway epithelial cells, not only because acidity is toxic to bronchial epithelial cells, but also since it contains

  6. File list: ALL.Brs.05.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Brs.05.AllAg.Mammary_epithelial_cells mm9 All antigens Breast Mammary epithelia...SRX031066,SRX031214,SRX396750 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Brs.05.AllAg.Mammary_epithelial_cells.bed ...

  7. Pim1 kinase protects airway epithelial cells from cigarette smoke-induced damage and airway inflammation

    NARCIS (Netherlands)

    de Vries, M.; Heijink, Hilde; Gras, R.; den Boef, L. E.; Reinders-Luinge, M.; Pouwels, S. D.; Hylkema, Machteld; van der Toorn, Marco; Brouwer, U.; van Oosterhout, A. J. M.; Nawijn, M. C.

    2014-01-01

    Exposure to cigarette smoke (CS) is the main risk factor for developing chronic obstructive pulmonary disease and can induce airway epithelial cell damage, innate immune responses, and airway inflammation. We hypothesized that cell survival factors might decrease the sensitivity of airway epithelial

  8. Effects of putrescine, cadaverine, spermine, spermidine and beta-phenylethylamine on cultured bovine mammary epithelial cells

    DEFF Research Database (Denmark)

    Fusi, Eleonora; Baldi, Antonella; Cheli, Federica

    2008-01-01

    A bovine mammary epithelial cell line (BME-UV1) and three-dimensional collagen primary bovine organoids were used to evaluate the effects of cadaverine, putrescine, spermine, spermicline and beta-phenylethylamine on mammary epithelial cells. Each biogenic amine was diluted in several concentratio...

  9. Effects of putrescine, cadaverine, spermine, spermidine and beta-phenylethylamine on cultured bovine mammary epithelial cells

    DEFF Research Database (Denmark)

    Fusi, Eleonora; Baldi, Antonella; Cheli, Federica

    2008-01-01

    A bovine mammary epithelial cell line (BME-UV1) and three-dimensional collagen primary bovine organoids were used to evaluate the effects of cadaverine, putrescine, spermine, spermicline and beta-phenylethylamine on mammary epithelial cells. Each biogenic amine was diluted in several concentratio...

  10. File list: ALL.Brs.50.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Brs.50.AllAg.Mammary_epithelial_cells mm9 All antigens Breast Mammary epithelia...SRX396750,SRX031066,SRX031214 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Brs.50.AllAg.Mammary_epithelial_cells.bed ...

  11. File list: ALL.Brs.20.AllAg.Mammary_epithelial_cells [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available ALL.Brs.20.AllAg.Mammary_epithelial_cells mm9 All antigens Breast Mammary epithelia...SRX396750,SRX031066,SRX031214 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/ALL.Brs.20.AllAg.Mammary_epithelial_cells.bed ...

  12. Roles of Wnt/{beta}-catenin signaling in epithelial differentiation of mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yajing; Sun, Zhaorui; Qiu, Xuefeng [Immunology and Reproductive Biology Laboratory, Medical College of Nanjing University, Nanjing 210093 (China); Jiangsu Key Laboratory of Molecular Medicine, Nanjing 210093 (China); Li, Yan [Jiangsu Centers for Diseases Prevention and Control, Nanjing 210009 (China); Qin, Jizheng [Immunology and Reproductive Biology Laboratory, Medical College of Nanjing University, Nanjing 210093 (China); Jiangsu Key Laboratory of Molecular Medicine, Nanjing 210093 (China); Han, Xiaodong, E-mail: hanxd@nju.edu.cn [Immunology and Reproductive Biology Laboratory, Medical College of Nanjing University, Nanjing 210093 (China); Jiangsu Key Laboratory of Molecular Medicine, Nanjing 210093 (China)

    2009-12-25

    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/{beta}-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/{beta}-catenin signaling were determined, suggested down-regulation of Wnt/{beta}-catenin with epithelial differentiation of MSCs. Furthermore, Wnt3{alpha} can inhibit the epithelial differentiation of MSCs. A loss of {beta}-catenin induced by Dickkopf-1 can enhance MSCs differentiation into epithelial cells. Lithium chloride transiently activated {beta}-catenin expression and subsequently decreased {beta}-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/{beta}-catenin signaling may promote MSCs to differentiate towards airway epithelial cells.

  13. PKC activation induces inflammatory response and cell death in human bronchial epithelial cells.

    Directory of Open Access Journals (Sweden)

    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.

  14. Effect of curcumin on aging retinal pigment epithelial cells

    Directory of Open Access Journals (Sweden)

    Zhu W

    2015-09-01

    Full Text Available Wei Zhu,1,* Yan Wu,2,* Yi-Fang Meng,1 Jin-Yu Wang,1 Ming Xu,1 Jian-Jun Tao,1 Jiong Lu1 1Department of Ophthalmology, Changshu No 2 People’s Hospital, Changshu, 2Department of Ophthalmology, The First People’s Hospital of Kunshan Affiliated with Jiangsu University, Suzhou, People’s Republic of China *These authors contributed equally to this work Abstract: Age-related macular degeneration (AMD is now one of the leading causes of blindness in the elderly population. The antioxidative effects of curcumin on aging retinal pigment epithelial (RPE cells are still unclear. We conducted an in vitro study to investigate the effects of curcumin on aging RPE cells. A pulsed H2O2 exposure aging model was adopted. Aging RPE cells were treated with curcumin 20 µM, 40 µM, and 80 µM. Apoptosis of RPE cells was analyzed by flow cytometry. The intracellular reactive oxygen species concentration was detected using a specific probe and apoptosis-associated proteins were detected by Western blot. Expression of oxidative biomarkers, including superoxide dismutase, maleic dialdehyde, and glutathione, was detected commercially available assay kits. Compared with normal cells, lower cell viability, higher apoptosis rates, and more severe oxidation status were identified in the aging RPE cell model. Curcumin improved cell viability and decreased apoptosis and oxidative stress. Further, curcumin had a significant influence on expression of apoptosis-associated proteins and oxidative stress biomarkers. In conclusion, treatment with curcumin was able to regulate proliferation, oxidative stress, and apoptosis in aging RPE cells. Accordingly, application of curcumin may be a novel strategy to protect against age-related change in AMD. Keywords: curcumin, retinal pigment epithelium, apoptosis, age-related macular degeneration

  15. The Neisseria meningitidis ADP-Ribosyltransferase NarE Enters Human Epithelial Cells and Disrupts Epithelial Monolayer Integrity.

    Directory of Open Access Journals (Sweden)

    Maria Valeri

    Full Text Available Many pathogenic bacteria utilize ADP-ribosylating toxins to modify and impair essential functions of eukaryotic cells. It has been previously reported that Neisseria meningitidis possesses an ADP-ribosyltransferase enzyme, NarE, retaining the capacity to hydrolyse NAD and to transfer ADP-ribose moiety to arginine residues in target acceptor proteins. Here we show that upon internalization into human epithelial cells, NarE gains access to the cytoplasm and, through its ADP-ribosylating activity, targets host cell proteins. Notably, we observed that these events trigger the disruption of the epithelial monolayer integrity and the activation of the apoptotic pathway. Overall, our findings provide, for the first time, evidence for a biological activity of NarE on host cells, suggesting its possible involvement in Neisseria pathogenesis.

  16. Maximum geometrical hindrance to diffusion in brain extracellular space surrounding uniformly spaced convex cells.

    Science.gov (United States)

    Tao, L; Nicholson, C

    2004-07-07

    Brain extracellular space (ECS) constitutes a porous medium in which diffusion is subject to hindrance, described by tortuosity, lambda = (D/D*)1/2, where D is the free diffusion coefficient and D* is the effective diffusion coefficient in brain. Experiments show that lambda is typically 1.6 in normal brain tissue although variations occur in specialized brain regions. In contrast, different theoretical models of cellular assemblies give ambiguous results: they either predict lambda-values similar to experimental data or indicate values of about 1.2. Here we constructed three different ECS geometries involving tens of thousands of cells and performed Monte Carlo simulation of 3-D diffusion. We conclude that the geometrical hindrance in the ECS surrounding uniformly spaced convex cells is independent of the cell shape and only depends on the volume fraction alpha (the ratio of the ECS volume to the whole tissue volume). This dependence can be described by the relation lambda = ((3-alpha)/2)1/2, indicating that the geometrical hindrance in such ECS cannot account for lambda > 1.225. Reasons for the discrepancy between the theoretical and experimental tortuosity values are discussed.

  17. 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

  18. 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 (ICAM-1) is involved in migration of leukocytes toward the colonic epithelium. A suitable in vitro model of chronic colonic inflammation does not exist, and the role of the epithelium is based on monolayers ...... of cancer cells. Conflicting results exist on epithelial ICAM-1 expression, and the aim of this study was to compare the expression in various models of colonic epithelium.......Studies have suggested that in ulcerative colitis (UC), intercellular adhesion molecule-1 (ICAM-1) is involved in migration of leukocytes toward the colonic epithelium. A suitable in vitro model of chronic colonic inflammation does not exist, and the role of the epithelium is based on monolayers...

  19. Gene expression in epithelial cells in response to pneumovirus infection

    Directory of Open Access Journals (Sweden)

    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.

  20. Human amniotic epithelial cells express specific markers of nerve cells and migrate along the nerve fibers in the corpus callosum

    Institute of Scientific and Technical Information of China (English)

    Zhiyuan Wu; Guozhen Hui; Yi Lu; Tianjin Liu; Qin Huang; Lihe Guo

    2012-01-01

    Human amniotic epithelial cells were isolated from a piece of fresh amnion. Using immunocytochemical methods, we investigated the expression of neuronal phenotypes (microtubule-associated protein-2, glial fibrillary acidic protein and nestin) in human amniotic epithelial cells. The conditioned medium of human amniotic epithelial cells promoted the growth and proliferation of rat glial cells cultured in vitro, and this effect was dose-dependent. Human amniotic epithelial cells were further transplanted into the corpus striatum of healthy adult rats and the grafted cells could integrate with the host and migrate 1-2 mm along the nerve fibers in corpus callosum. Our experimental findings indicate that human amniotic epithelial cells may be a new kind of seed cells for use in neurograft.

  1. Interactions of virulent and avirulent leptospires with primary cultures of renal epithelial cells

    DEFF Research Database (Denmark)

    Ballard, S A; Williamson, M; Adler, B

    1986-01-01

    A primary culture system for the cells of mouse renal-tubular epithelium was established and used to observe the adhesion of leptospires. Virulent strains of serovars copenhageni and ballum attached themselves to epithelial cells within 3 h of infection whereas an avirulent variant of serovar...... copenhageni did not adhere to epithelial cells at all within the experimental period of 24 h. The saprophytic Leptospira biflexa serovar patoc became attached non-specifically to inert glass surfaces as well as to the cells. The adhesion of leptospires to epithelial cells was not inhibited by homologous...

  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. Interaction of oral bacteria with gingival epithelial cell multilayers.

    Science.gov (United States)

    Dickinson, B C; Moffatt, C E; Hagerty, D; Whitmore, S E; Brown, T A; Graves, D T; Lamont, R J

    2011-06-01

    Primary gingival epithelial cells were cultured in multilayers as a model for the study of interactions with oral bacteria associated with health and periodontal disease. Multilayers maintained at an air-liquid interface in low-calcium medium displayed differentiation and cytokeratin properties characteristic of junctional epithelium. Multilayers were infected with fluorescently labeled Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum or Streptococcus gordonii, and bacterial association was determined by confocal microscopy and quantitative image analysis. Porphyromonas gingivalis invaded intracellularly and spread from cell to cell; A. actinomycetemcomitans and F. nucleatum remained extracellular and showed intercellular movement through the multilayer; whereas S. gordonii remained extracellular and predominantly associated with the superficial cell layer. None of the bacterial species disrupted barrier function as measured by transepithelial electrical resistance. P. gingivalis did not elicit secretion of proinflammatory cytokines. However, A. actinomycetemcomitans and S. gordonii induced interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), IL-6 and IL-8 secretion; and F. nucleatum stimulated production of IL-1β and TNF-α. Aggregatibacter actinomycetemcomitans, F. nucleatum and S. gordonii, but not P. gingivalis, increased levels of apoptosis after 24 h infection. The results indicate that the organisms with pathogenic potential were able to traverse the epithelium, whereas the commensal bacteria did not. In addition, distinct host responses characterized the interaction between the junctional epithelium and oral bacteria.

  4. The Effects of Vitamin A Compounds on Hyaluronic Acid Released from Cultured Rabbit Corneal Epithelial Cells and Keratocytes

    National Research Council Canada - National Science Library

    TOSHIDA, Hiroshi; TABUCHI, Nobuhito; KOIKE, Daisuke; KOIDE, Misao; SUGIYAMA, Keikichi; NAKAYASU, Kiyoo; KANAI, Atsushi; MURAKAMI, Akira

    2012-01-01

    .... Hyaluronic acid is produced by corneal epithelial cells and keratocytes in the eye. We investigated whether rabbit corneal epithelial cells and keratocytes release hyaluronic acid after exposure to vitamin A compounds...

  5. File list: ALL.Utr.05.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  6. File list: ALL.Utr.20.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  7. File list: ALL.Utr.50.AllAg.Fallopian_tube_secretory_epithelial_cell [Chip-atlas[Archive

    Lifescience Database Archive (English)

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  8. In vitro isolation and cultivation of rabbit tracheal epithelial cells using tissue explant technique.

    Science.gov (United States)

    Shi, Hong-Can; Lu, Dan; Li, Hai-Jia; Han, Shi; Zeng, Yan-Jun

    2013-04-01

    Epithelial cells from tracheal mucosa offer significant potential as a cell source in development of tissue-engineered trachea. The purpose of this study was to investigate and optimize a suitable culture system for tracheal epithelial cells, including the methods of primary culture, passage, identification, and cryopreservation. Epithelial cells were isolated from rabbit tracheal mucosa using tissue explant technique and were subjected to immunohistochemistry, immunofluorescence, and cryopreservation after purification. Epithelial cells reached confluency at 14-15 d. Immunohistochemical staining for cytokeratin showed brown yellow-positive cytoplasm and blue-counterstained nuclei, while immunofluorescence staining for cytokeratin showed green-positive cytoplasm and clear cell outline, indicating that the cultured cells had properties of epithelial cells. After recovery, epithelial cells exhibited high survival and viability. The results demonstrated that in vitro isolation and cultivation model was successfully established to provide high proliferative capacity, typical morphology and characteristics of tracheal epithelial cells from trachea mucosa by the use of the tissue explant technique.

  9. Epithelial cells as active player in fibrosis: findings from an in vitro model.

    Directory of Open Access Journals (Sweden)

    Solange Moll

    Full Text Available Kidney fibrosis, a scarring of the tubulo-interstitial space, is due to activation of interstitial myofibroblasts recruited locally or systemically with consecutive extracellular matrix deposition. Newly published clinical studies correlating acute kidney injury (AKI to chronic kidney disease (CKD challenge this pathological concept putting tubular epithelial cells into the spotlight. In this work we investigated the role of epithelial cells in fibrosis using a simple controlled in vitro system. An epithelial/mesenchymal 3D cell culture model composed of human proximal renal tubular cells and fibroblasts was challenged with toxic doses of Cisplatin, thus injuring epithelial cells. RT-PCR for classical fibrotic markers was performed on fibroblasts to assess their modulation toward an activated myofibroblast phenotype in presence or absence of that stimulus. Epithelial cell lesion triggered a phenotypical modulation of fibroblasts toward activated myofibroblasts as assessed by main fibrotic marker analysis. Uninjured 3D cell culture as well as fibroblasts alone treated with toxic stimulus in the absence of epithelial cells were used as control. Our results, with the caveats due to the limited, but highly controllable and reproducible in vitro approach, suggest that epithelial cells can control and regulate fibroblast phenotype. Therefore they emerge as relevant target cells for the development of new preventive anti-fibrotic therapeutic approaches.

  10. Allergen recognition by innate immune cells: critical role of dendritic and epithelial cells

    Directory of Open Access Journals (Sweden)

    Fabian eSalazar

    2013-11-01

    Full Text Available Allergy is an exacerbated response of the immune system against non-self-proteins called allergens and is typically characterized by biased type-2 T helper cell and deleterious IgE mediated immune responses. The allergic cascade starts with the recognition of allergens by antigen presenting cells, mainly dendritic cells, culminating in mast cell sensitization and triggering. Dendritic cells have been demonstrated to play a crucial role in orchestrating allergic diseases. Using different C-type lectin receptors dendritic cells are able to recognize and internalize a number of allergens from diverse sources leading to sensitization. Furthermore, there is increasing evidence highlighting the role of epithelial cells in triggering and modulating immune responses to allergens. As well as providing a physical barrier, epithelial cells can interact with allergens and influence dendritic cells behaviour through the release of a number of Th2 promoting cytokines. In this review we will summarise current understanding of how allergens are recognised by dendritic cells and epithelial cells and what are the consequences of such interaction in the context of allergic sensitisation and downstream events leading to allergic inflammation. Better understanding of the molecular mechanisms of allergen recognition and associated signalling pathways could enable developing more effective therapeutic strategies that target the initial steps of allergic sensitisation hence hindering development or progression of allergic diseases.

  11. Cytotoxic effects of octenidine mouth rinse on human fibroblasts and epithelial cells - an in vitro study.

    Science.gov (United States)

    Schmidt, J; Zyba, V; Jung, K; Rinke, S; Haak, R; Mausberg, R F; Ziebolz, D

    2016-01-01

    This study compared the cytotoxicity of a new octenidine mouth rinse (MR) against gingival fibroblasts and epithelial cells with different established MRs. The following MRs were used: Octenidol (OCT), Chlorhexidine 0.2% (CHX), Listerine (LIS), Meridol (MER), Betaisodona (BET); and control (medium only). Human primary gingiva fibroblasts and human primary nasal epithelial cells were cultivated in cell-specific media (2 × 10(5) cells/ml) and treated with MR for 1, 5, and 15 min. Each test was performed 12 times. Metabolism activity was measured using a cytotoxicity assay. A cellometer analyzed cell viability, cell number, and cell diameter. The data were analyzed by two-way analysis of variance with subsequent Dunnett's test and additional t-tests. The cytotoxic effects of all MRs on fibroblasts and epithelial cells compared to the control depended on the contact time (p 0.005). Cell numbers of both cell types at all contact times revealed that OCT showed a less negative effect (p > 0.005), especially for epithelial cells compared to CHX after 15 min (p 0.005), but MER showed less influence than OCT in epithelial cells (p < 0.005). OCT is a potential alternative to CHX regarding cytotoxicity because of its lower cell-toxic effect against fibroblasts and epithelial cells.

  12. Cytotoxic effects of curcumin in human retinal pigment epithelial cells.

    Directory of Open Access Journals (Sweden)

    Margrit Hollborn

    Full Text Available BACKGROUND: Curcumin from turmeric is an ingredient in curry powders. Due to its antiinflammatory, antioxidant and anticarcinogenic effects, curcumin is a promising drug for the treatment of cancer and retinal diseases. We investigated whether curcumin alters the viability and physiological properties of human retinal pigment epithelial (RPE cells in vitro. METHODOLOGY/PRINCIPAL FINDINGS: Cellular proliferation was investigated with a bromodeoxy-uridine immunoassay, and chemotaxis was investigated with a Boyden chamber assay. Cell viability was determined by trypan blue exclusion. Apoptosis and necrosis rates were determined with a DNA fragmentation ELISA. Gene expression was determined by real-time PCR, and secretion of VEGF and bFGF was examined with ELISA. The phosphorylation level of proteins was revealed by Western blotting. The proliferation of RPE cells was slightly increased by curcumin at 10 µM and strongly reduced by curcumin above 50 µM. Curcumin at 50 µM increased slightly the chemotaxis of the cells. Curcumin reduced the expression and secretion of VEGF under control conditions and abolished the VEGF secretion induced by PDGF and chemical hypoxia. Whereas low concentrations of curcumin stimulated the expression of bFGF and HGF, high concentrations caused downregulation of both factors. Curcumin decreased dose-dependently the viability of RPE cells via induction of early necrosis (above 10 µM and delayed apoptosis (above 1 µM. The cytotoxic effect of curcumin involved activation of caspase-3 and calpain, intracellular calcium signaling, mitochondrial permeability, oxidative stress, increased phosphorylation of p38 MAPK and decreased phosphorylation of Akt protein. CONCLUSION: It is concluded that curcumin at concentrations described to be effective in the treatment of tumor cells and in inhibiting death of retinal neurons (∼10 µM has adverse effects on RPE cells. It is suggested that, during the intake of curcumin as

  13. Cytotoxic Effects of Curcumin in Human Retinal Pigment Epithelial Cells

    Science.gov (United States)

    Hollborn, Margrit; Chen, Rui; Wiedemann, Peter; Reichenbach, Andreas; Bringmann, Andreas; Kohen, Leon

    2013-01-01

    Backround Curcumin from turmeric is an ingredient in curry powders. Due to its antiinflammatory, antioxidant and anticarcinogenic effects, curcumin is a promising drug for the treatment of cancer and retinal diseases. We investigated whether curcumin alters the viability and physiological properties of human retinal pigment epithelial (RPE) cells in vitro. Methodology/Principal Findings Cellular proliferation was investigated with a bromodeoxy-uridine immunoassay, and chemotaxis was investigated with a Boyden chamber assay. Cell viability was determined by trypan blue exclusion. Apoptosis and necrosis rates were determined with a DNA fragmentation ELISA. Gene expression was determined by real-time PCR, and secretion of VEGF and bFGF was examined with ELISA. The phosphorylation level of proteins was revealed by Western blotting. The proliferation of RPE cells was slightly increased by curcumin at 10 µM and strongly reduced by curcumin above 50 µM. Curcumin at 50 µM increased slightly the chemotaxis of the cells. Curcumin reduced the expression and secretion of VEGF under control conditions and abolished the VEGF secretion induced by PDGF and chemical hypoxia. Whereas low concentrations of curcumin stimulated the expression of bFGF and HGF, high concentrations caused downregulation of both factors. Curcumin decreased dose-dependently the viability of RPE cells via induction of early necrosis (above 10 µM) and delayed apoptosis (above 1 µM). The cytotoxic effect of curcumin involved activation of caspase-3 and calpain, intracellular calcium signaling, mitochondrial permeability, oxidative stress, increased phosphorylation of p38 MAPK and decreased phosphorylation of Akt protein. Conclusion It is concluded that curcumin at concentrations described to be effective in the treatment of tumor cells and in inhibiting death of retinal neurons (∼10 µM) has adverse effects on RPE cells. It is suggested that, during the intake of curcumin as concomitant therapy of

  14. Elastase induces lung epithelial cell autophagy through placental growth factor

    Science.gov (United States)

    Hou, Hsin-Han; Cheng, Shih-Lung; Chung, Kuei-Pin; Kuo, Mark Yen-Ping; Yeh, Cheng-Chang; Chang, Bei-En; Lu, Hsuan-Hsuan; Wang, Hao-Chien; Yu, Chong-Jen

    2014-01-01

    Chronic obstructive pulmonary disease (COPD) is a devastating disease, which is associated with increasing mortality and morbidity. Therefore, there is a need to clearly define the COPD pathogenic mechanism and to explore effective therapies. Previous studies indicated that cigarette smoke (CS) induces autophagy and apoptosis in lung epithelial (LE) cells. Excessive ELANE/HNE (elastase, neutrophil elastase), a factor involved in protease-antiprotease imbalance and the pathogenesis of COPD, causes LE cell apoptosis and upregulates the expression of several stimulus-responsive genes. However, whether or not elastase induces autophagy in LE cell remains unknown. The level of PGF (placental growth factor) is higher in COPD patients than non-COPD controls. We hypothesize that elastase induces PGF expression and causes autophagy in LE cells. In this study, we demonstrated that porcine pancreatic elastase (PPE) induced PGF expression and secretion in LE cells in vitro and in vivo. The activation of MAPK8/JNK1 (mitogen-activated protein kinase 8) and MAPK14/p38alpha MAPK signaling pathways was involved in the PGF mediated regulation of the TSC (tuberous sclerosis complex) pathway and autophagy in LE cells. Notably, PGF-induced MAPK8 and MAPK14 signaling pathways mediated the inactivation of MTOR (mechanistic target of rapamycin), the upregulation of MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3 β) and the increase of autophagosome formation in mice. Furthermore, the PPE-induced autophagy promotes further apoptosis in vitro and in vivo. In summary, elastase-induced autophagy promotes LE cell apoptosis and pulmonary emphysema through the upregulation of PGF. PGF and its downstream MAPK8 and MAPK14 signaling pathways are potential therapeutic targets for the treatment of emphysema and COPD. PMID:24988221

  15. Distribution of natural killer cells and T-lymphocyte subsets in peripheral blood, gallbladder cancer and surrounding tissue

    Institute of Scientific and Technical Information of China (English)

    Gang Liu; Hong Ren; Xue-Jun Sun; Jing-Sen Shi

    2007-01-01

    BACKGROUND:The patient with malignant tumor always show immunologic function drawback and ingravescent with tumor development, especially in the aspect of cell-mediated immunity. This study was undertaken to deifne the relationship between the immune function of local cells and cancer development by investigating the distribution of natural killer (NK) cells and T-lymphocyte subsets in peripheral blood, the cancer tissue and the tissue surrounding gallbladder carcinoma. METHODS:The numbers of CD4+and CD8+T-lymphocytes and NK cells were measured by lfow cytometry in samples taken from gallbladder cancer tissue, the surrounding tissues and peripheral blood of 38 patients, and compared with the numbers in the peripheral blood and gallbladder tissue of 30 patients with cholecystitis as controls. RESULTS:The numbers of CD4+and CD8+T-cells and NK cells in gallbladder cancer tissues were signiifcantly higher than those in the surrounding tissue and gallbladder with gallstone. However, the ratio of CD4+/CD8+was lower in the cancer tissue than that in the surrounding tissue and tissue from gallbladders with gallstones. The distribution of CD4+ and CD8+ T-cells and NK cells in mucous membrane of cholecystitis gallbladder and that in the tissue surrounding gallbladder cancer were signiifcantly different. CONCLUSIONS:Disproportionate and imbalanced distri-bution of NK cells and subsets of T-lymphocytes occurs in the mucous membrane proper of gallbladder cancer and surrounding tissue. Although gallbladder cancer tissue has higher expressions of CD4+, CD8+and NK cells, the immune function is low or in an inhibited state. In gallbladder cancer immunization therapy, local cellular immunological function should be enhanced and the protective barrier improved.

  16. The Rho Target PRK2 Regulates Apical Junction Formation in Human Bronchial Epithelial Cells

    OpenAIRE

    Wallace, Sean W.; Magalhaes, Ana; Hall, Alan

    2010-01-01

    Rho GTPases regulate multiple signaling pathways to control a number of cellular processes during epithelial morphogenesis. To investigate the downstream pathways through which Rho regulates epithelial apical junction formation, we screened a small interfering RNA (siRNA) library targeting 28 known Rho target proteins in 16HBE human bronchial epithelial cells. This led to the identification of the serine-threonine kinase PRK2 (protein kinase C-related kinase 2, also called PKN2). Depletion of...

  17. Specific N-glycan alterations are coupled in epithelial-mesenchymal transition induced by EGF in GE11 epithelial cells.

    Science.gov (United States)

    Xu, Qingsong; Qu, Chen; Wang, Wenjing; Gu, Jianguo; Du, Yuguang; Song, Linsheng

    2017-02-01

    Epithelial-mesenchymal transition (EMT) is a phenomenon in cancer progression during which cancer cells undergo remarkable alteration acquiring highly invasive property. The aim of this study was to evaluate specific N-glycan alterations during EMT induced by epidermal growth factor (EGF) in GE11 epithelial cells. Herein, we demonstrated that EGF activated epidermal growth factor receptor (EGFR)/Akt/extracellular signal-regulated kinase (ERK) phosphorylation and promoted GE11 cell proliferation. Meanwhile, EGF stimulated the epithelial cells to undergo morphological alteration, destroying cell-cell inter-contact and exhibiting mesenchymal cells higher metastatic potential. A wound-healing assay showed the migratory ability increased 1.5-fold after EGF treatment. Moreover, the relative intensity of N-cadherin versus E-cadherin increased 2.6-fold, and the E-cadherin distribution in cell-cell junctions became jagged and faint after EGF incubation for 72 h. Interestingly, the amounts of bisecting GlcNAc structure were dramatically declined, by contrast, the formation of β1,6 GlcNAc branches on cell surface was upregulated during EMT induced by EGF. To understand the roles of N-glycans in EGF-induced EMT, the cells were stably transfected with N-acetylglucosaminyltransferase III (GnT-III), which catalyzes the bisecting GlcNAc structure formation. As the markers for EMT, EGF-induced E-cadherin decrease and fibronectin increase were delayed in GnT-III-overexpressing cells. Taken together, these results demonstrated that specific N-glycan alterations were coupled in EMT induced by EGF, which might be contributed to diagnosis and therapy of tumor metastasis.

  18. Epithelial cell kinetics of the gastric mucosa during Helicobacter pylori infection

    DEFF Research Database (Denmark)

    Holck, Susanne; Holm, I.L.; Holck, P.P.

    2007-01-01

    Helicobacter pylori is an important pathogen in major gastroduodenal diseases, including inflammation with ulceration and gastric malignancies. Alterations in H. pylori associated cell turnover in gastric epithelial cells are examined in relation to inflammatory activity, bacteria load and cytoki...

  19. Generation of corneal epithelial cells from induced pluripotent stem cells derived from human dermal fibroblast and corneal limbal epithelium.

    Directory of Open Access Journals (Sweden)

    Ryuhei Hayashi

    Full Text Available Induced pluripotent stem (iPS cells can be established from somatic cells. However, there is currently no established strategy to generate corneal epithelial cells from iPS cells. In this study, we investigated whether corneal epithelial cells could be differentiated from iPS cells. We tested 2 distinct sources: human adult dermal fibroblast (HDF-derived iPS cells (253G1 and human adult corneal limbal epithelial cells (HLEC-derived iPS cells (L1B41. We first established iPS cells from HLEC by introducing the Yamanaka 4 factors. Corneal epithelial cells were successfully induced from the iPS cells by the stromal cell-derived inducing activity (SDIA differentiation method, as Pax6(+/K12(+ corneal epithelial colonies were observed after prolonged differentiation culture (12 weeks or later in both the L1B41 and 253G1 iPS cells following retinal pigment epithelial and lens cell induction. Interestingly, the corneal epithelial differentiation efficiency was higher in L1B41 than in 253G1. DNA methylation analysis revealed that a small proportion of differentially methylated regions still existed between L1B41 and 253G1 iPS cells even though no significant difference in methylation status was detected in the specific corneal epithelium-related genes such as K12, K3, and Pax6. The present study is the first to demonstrate a strategy for corneal epithelial cell differentiation from human iPS cells, and further suggests that the epigenomic status is associated with the propensity of iPS cells to differentiate into corneal epithelial cells.

  20. Hexagonal packing of Drosophila wing epithelial cells by the planar cell polarity pathway.

    Science.gov (United States)

    Classen, Anne-Kathrin; Anderson, Kurt I; Marois, Eric; Eaton, Suzanne

    2005-12-01

    The mechanisms that order cellular packing geometry are critical for the functioning of many tissues, but they are poorly understood. Here, we investigate this problem in the developing wing of Drosophila. The surface of the wing is decorated by hexagonally packed hairs that are uniformly oriented by the planar cell polarity pathway. They are constructed by a hexagonal array of wing epithelial cells. Wing epithelial cells are irregularly arranged throughout most of development, but they become hexagonally packed shortly before hair formation. During the process, individual cell boundaries grow and shrink, resulting in local neighbor exchanges, and Cadherin is actively endocytosed and recycled through Rab11 endosomes. Hexagonal packing depends on the activity of the planar cell polarity proteins. We propose that these proteins polarize trafficking of Cadherin-containing exocyst vesicles during junction remodeling. This may be a common mechanism for the action of planar cell polarity proteins in diverse systems.

  1. Culture of human intestinal epithelial cell using the dissociating enzyme thermolysin and endothelin-3

    Directory of Open Access Journals (Sweden)

    Z. Liu

    2010-05-01

    Full Text Available Epithelium, a highly dynamic system, plays a key role in the homeostasis of the intestine. However, thus far a human intestinal epithelial cell line has not been established in many countries. Fetal tissue was selected to generate viable cell cultures for its sterile condition, effective generation, and differentiated character. The purpose of the present study was to culture human intestinal epithelial cells by a relatively simple method. Thermolysin was added to improve the yield of epithelial cells, while endothelin-3 was added to stimulate their growth. By adding endothelin-3, the achievement ratio (viable cell cultures/total cultures was enhanced to 60% of a total of 10 cultures (initiated from 8 distinct fetal small intestines, allowing the generation of viable epithelial cell cultures. Western blot, real-time PCR and immunofluorescent staining showed that cytokeratins 8, 18 and mouse intestinal mucosa-1/39 had high expression levels in human intestinal epithelial cells. Differentiated markers such as sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV also showed high expression levels in human intestinal epithelial cells. Differentiated human intestinal epithelial cells, with the expression of surface markers (cytokeratins 8, 18 and mouse intestinal mucosa-1/39 and secretion of cytokines (sucrase-isomaltase, aminopeptidase N and dipeptidylpeptidase IV, may be cultured by the thermolysin and endothelin-3 method and maintained for at least 20 passages. This is relatively simple, requiring no sophisticated techniques or instruments, and may have a number of varied applications.

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

    Science.gov (United States)

    Aparicio-Domingo, Patricia; Romera-Hernandez, Monica; Karrich, Julien J; Cornelissen, Ferry; Papazian, Natalie; Lindenbergh-Kortleve, Dicky J; Butler, James A; Boon, Louis; Coles, Mark C; Samsom, Janneke N; Cupedo, Tom

    2015-10-19

    Disruption 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 of this high mitotic activity, mucosal surfaces are frequently targeted by anticancer therapies, leading to dose-limiting side effects. The cellular mechanisms that control tissue protection and mucosal healing in response to intestinal damage remain poorly understood. Type 3 innate lymphoid cells (ILC3s) are regulators of homeostasis and tissue responses to infection at mucosal surfaces. We now demonstrate that ILC3s are required for epithelial activation and proliferation in response to small intestinal tissue damage induced by the chemotherapeutic agent methotrexate. Multiple subsets of ILC3s are activated after intestinal tissue damage, and in the absence of ILC3s, epithelial activation is lost, correlating with increased pathology and severe damage to the intestinal crypts. Using ILC3-deficient Lgr5 reporter mice, we show that maintenance of intestinal stem cells after damage is severely impaired in the absence of ILC3s or the ILC3 signature cytokine IL-22. These data unveil a novel function of ILC3s in limiting tissue damage by preserving tissue-specific stem cells.

  3. Tacrolimus Modulates TGF-β Signaling to Induce Epithelial-Mesenchymal Transition in Human Renal Proximal Tubule Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Jason Bennett

    2016-04-01

    Full Text Available Epithelial-mesenchymal transition (EMT, a process which describes the trans-differentiation of epithelial cells into motile mesenchymal cells, is pivotal in stem cell behavior, development and wound healing, as well as contributing to disease processes including fibrosis and cancer progression. Maintenance immunosuppression with calcineurin inhibitors (CNIs has become routine management for renal transplant patient, but unfortunately the nephrotoxicity of these drugs has been well documented. HK-2 cells were exposed to Tacrolimus (FK506 and EMT markers were assessed by RT PCR and western blot. FK506 effects on TGF-β mRNA were assessed by RT PCR and TGF-β secretion was measured by ELISA. The impact of increased TGF-β secretion on Smad signaling pathways was investigated. The impact of inhibition of TGF-β signaling on EMT processes was assessed by scratch-wound assay. The results presented in this study suggest that FK506 initiates EMT processes in the HK-2 cell line, with altered expression of epithelial and myofibroblast markers evident. Additionally, the study demonstrates that FK506 activation of the TGF-β/ SMAD pathways is an essential step in the EMT process. Overall the results demonstrate that EMT is heavily involved in renal fibrosis associated with CNI nephrotoxicity.

  4. Autophagy protects type II alveolar epithelial cells from Mycobacterium tuberculosis infection

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Xu-Guang [Center for Clinical Laboratory Medicine of PLA, Xijing Hospital, Fourth Military Medical University, Xi' an (China); Department of Laboratory Medicine, The Third Affiliated Hospital of Guangzhou Medical University, Guangzhou (China); Ji, Tian-Xing [Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou Medical University, Guangzhou (China); Xia, Yong, E-mail: gysyxy@gmail.com [Center for Clinical Laboratory Medicine of PLA, Xijing Hospital, Fourth Military Medical University, Xi' an (China); Ma, Yue-Yun, E-mail: cmbmayy@fmmu.edu.cn [Center for Clinical Laboratory Medicine of PLA, Xijing Hospital, Fourth Military Medical University, Xi' an (China)

    2013-03-08

    Highlights: ► We investigated the protective effect of autophagy pathway against MTB infection. ► MTB-infected A549 cells had higher LDH release. ► Inhibition of autophagy signaling significantly enhanced the MTB-induced necrosis. ► Autophagy prevents apoptosis and promotes cell survival in infected cells. -- Abstract: This study was designed to investigate the protective effect of the autophagy signaling pathway against Mycobacterium tuberculosis infection in type II alveolar epithelial cells. An in vitro M. tuberculosis system was established using human A549 cells. Infection-induced changes in the expression of the autophagic marker LC3 were assessed by reverse transcription-PCR and Western blotting. Morphological changes in autophagosomes were detected by transmission electron microscopy (TEM). The function of the autophagy signaling pathway during infection was assessed by measuring the level of cell death and the amount of lactate dehydrogenase (LDH) released in the presence or absence of the inhibitor 3-methyladenine (3-MA). In addition, effects on LDH release were assessed after the siRNA-mediated knockdown of the essential autophagosomal structural membrane protein Atg5. LC3 mRNA expression was significantly reduced in M.tuberculosis-infected A549 cells (16888.76 ± 1576.34 vs. uninfected: 12744.29 ± 1089.37; P < 0.05). TEM revealed M.tuberculosis bacilli-containing compartments that were surrounded by double membranes characteristic of the autophagic process. M.tuberculosis-infected A549 cells released more LDH (1.45 ± 0.12 vs. uninfected: 0.45 ± 0.04; P < 0.05). The inhibition of autophagy signaling significantly enhanced M.tuberculosis-induced necrosis (3-MA: 75 ± 5% vs. untreated: 15 ± 1%; P < 0.05) and LDH release (3-MA: 2.50 ± 0.24 vs. untreated: 0.45 ± 0.04; Atg5 knockdown: 3.19 ± 0.29 vs. untreated: 1.28 ± 0.11; P < 0.05). Our results indicate that autophagy signaling pathway prevents apoptosis in type II alveolar epithelial cells

  5. Phthalates stimulate the epithelial to mesenchymal transition through an HDAC6-dependent mechanism in human breast epithelial stem cells.

    Science.gov (United States)

    Hsieh, Tsung-Hua; Tsai, Cheng-Fang; Hsu, Chia-Yi; Kuo, Po-Lin; Lee, Jau-Nan; Chai, Chee-Yin; Hou, Ming-Feng; Chang, Chia-Cheng; Long, Cheng-Yu; Ko, Ying-Chin; Tsai, Eing-Mei

    2012-08-01

    Phthalates are environmental hormone-like molecules that are associated with breast cancer risk and are involved in metastasis, a process that requires the epithelial-mesenchymal transition (EMT). However, few studies have addressed the potential effects of phthalates on stem cells. Here we tested the hypothesis that phthalates such as butyl benzyl phthalate and di-n-butyl phthalate induce EMT in R2d cells, a stem cell-derived human breast epithelial cell line that is responsive to estradiol for tumor development. We observed that phthalates induced EMT as evidenced by morphological changes concomitant with increased expression of mesenchymal markers and decreased expression of epithelial markers. Molecular mechanism studies revealed that histone deacetylase 6 (HDAC6) is required for phthalate-induced cell migration and invasion during EMT in vitro and metastasis into the lungs of nude mice. We also constructed a series of mutant HDAC6 promoter fragments and found that the transcription factor AP-2a plays a novel role in regulating the HDAC6 promoter. Furthermore, phthalates stimulated estrogen receptors and triggered the downstream EGFR-PKA signaling cascade, leading to increased expression of AP-2a in the nucleus. We also observed that phthalates increased expression of the PP1/HDAC6 complex and caused Akt activation and GSK3β inactivation, leading to transcriptional activation of vimentin through the β-catenin-TCF-4/LEF1 pathway. Understanding the signaling cascades of phthalates that activate EMT through HDAC6 in breast epithelial stem cells provides the identification of novel therapeutic target for human breast cancer.

  6. Use of scanning electron microscopy to monitor nanofibre/cell interaction in digestive epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Millaku, Agron, E-mail: agron.mi@hotmail.com [Limnos-Company for Applied Ecology Ltd, Podlimbarskega 31, 1000 Ljubljana (Slovenia); Drobne, Damjana [University of Ljubljana, Biotechnical Faculty, Department of Biology, Večna pot 111, 1000 Ljubljana (Slovenia); Centre of Excellence, Advanced Materials and Technologies for the Future (CO NAMASTE), Jamova cesta 39, 1000 Ljubljana (Slovenia); Centre of Excellence, Nanoscience and Nanotechnology (Nanocentre), Jamova cesta 39, 1000 Ljubljana (Slovenia); Torkar, Matjaz [Institute of Metals and Technology IMT, Lepi pot 11, 1000 Ljubljana (Slovenia); Jožef Stefan Institute, Condensed Matter Physics Department, Jamova cesta 39, 1000 Ljubljana (Slovenia); Novak, Sara [University of Ljubljana, Biotechnical Faculty, Department of Biology, Večna pot 111, 1000 Ljubljana (Slovenia); Remškar, Maja [Jožef Stefan Institute, Condensed Matter Physics Department, Jamova cesta 39, 1000 Ljubljana (Slovenia); Pipan-Tkalec, Živa [University of Ljubljana, Biotechnical Faculty, Department of Biology, Večna pot 111, 1000 Ljubljana (Slovenia)

    2013-09-15

    Graphical abstract: Scanning electron microscopy is particularly well suited to the observation of nanofibre/cell interaction in the endothelial cells lining the hepatopancreas. (a) Tungsten oxide nanofibres, (b) test organism Porcellio scaber and schematic appearance of digestive tubes, (c) digestive tube (hepatopancreas) prepared for SEM investigation, (d) digestive gland cells (C) with nanofibres (NF) embedded in the cell membrane and (e) nanofibres inserted deeply in the cells and damaged nanofibres due to peristalsis. -- Highlights: • Tungsten oxide nanofibres react physically with digestive gland epithelial cells in Porcellio scaber. • Physical peristaltic forces of lead to insertion of nanofibres into the cells. • No toxic responses as measured by conventional toxicity biomarkers were detected. • Physical interactions were observed in a majority of the investigated animals. -- Abstract: 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.

  7. An Optimised Human Cell Culture Model for Alveolar Epithelial Transport

    Science.gov (United States)

    Birch, Nigel P.; Suresh, Vinod

    2016-01-01

    Robust and reproducible in vitro models are required for investigating the pathways involved in fluid homeostasis in the human alveolar epithelium. We performed functional and phenotypic characterisation of ion transport in the human pulmonary epithelial cell lines NCI-H441 and A549 to determine their similarity to primary human alveolar type II cells. NCI-H441 cells exhibited high expression of junctional proteins ZO-1, and E-cadherin, seal-forming claudin-3, -4, -5 and Na+-K+-ATPase while A549 cells exhibited high expression of pore-forming claudin-2. Consistent with this phenotype NCI-H441, but not A549, cells formed a functional barrier with active ion transport characterised by higher electrical resistance (529 ± 178 Ω cm2 vs 28 ± 4 Ω cm2), lower paracellular permeability ((176 ± 42) ×10−8 cm/s vs (738 ± 190) ×10−8 cm/s) and higher transepithelial potential difference (11.9 ± 4 mV vs 0 mV). Phenotypic and functional properties of NCI-H441 cells were tuned by varying cell seeding density and supplement concentrations. The cells formed a polarised monolayer typical of in vivo epithelium at seeding densities of 100,000 cells per 12-well insert while higher densities resulted in multiple cell layers. Dexamethasone and insulin-transferrin-selenium supplements were required for the development of high levels of electrical resistance, potential difference and expression of claudin-3 and Na+-K+-ATPase. Treatment of NCI-H441 cells with inhibitors and agonists of sodium and chloride channels indicated sodium absorption through ENaC under baseline and forskolin-stimulated conditions. Chloride transport was not sensitive to inhibitors of the cystic fibrosis transmembrane conductance regulator (CFTR) under either condition. Channels inhibited by 5-nitro-1-(3-phenylpropylamino) benzoic acid (NPPB) contributed to chloride secretion following forskolin stimulation, but not at baseline. These data precisely define experimental conditions for the application of NCI

  8. Culture models of human mammary epithelial cell transformation

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, Martha R.; Yaswen, Paul

    2000-11-10

    Human pre-malignant breast diseases, particularly ductal carcinoma in situ (DCIS)3 already display several of the aberrant phenotypes found in primary breast cancers, including chromosomal abnormalities, telomerase activity, inactivation of the p53 gene and overexpression of some oncogenes. Efforts to model early breast carcinogenesis in human cell cultures have largely involved studies in vitro transformation of normal finite lifespan human mammary epithelial cells (HMEC) to immortality and malignancy. We present a model of HMEC immortal transformation consistent with the know in vivo data. This model includes a recently described, presumably epigenetic process, termed conversion, which occurs in cells that have overcome stringent replicative senescence and are thus able to maintain proliferation with critically short telomeres. The conversion process involves reactivation of telomerase activity, and acquisition of good uniform growth in the absence and presence of TFGB. We propose th at overcoming the proliferative constraints set by senescence, and undergoing conversion, represent key rate-limiting steps in human breast carcinogenesis, and occur during early stage breast cancer progression.

  9. Proliferation of cultured mouse choroid plexus epithelial cells.

    Directory of Open Access Journals (Sweden)

    Basam Z Barkho

    Full Text Available The choroid plexus (ChP epithelium is a multifunctional tissue found in the ventricles of the brain. The major function of the ChP epithelium is to produce cerebrospinal fluid (CSF that bathes and nourishes the central nervous system (CNS. In addition to the CSF, ChP epithelial cells (CPECs produce and secrete numerous neurotrophic factors that support brain homeostasis, such as adult hippocampal neurogenesis. Accordingly, damage and dysfunction to CPECs are thought to accelerate and intensify multiple disease phenotypes, and CPEC regeneration would represent a potential therapeutic approach for these diseases. However, previous reports suggest that CPECs rarely divide, although this has not been extensively studied in response to extrinsic factors. Utilizing a cell-cycle reporter mouse line and live cell imaging, we identified scratch injury and the growth factors insulin-like growth factor 1 (IGF-1 and epidermal growth factor (EGF as extrinsic cues that promote increased CPEC expansion in vitro. Furthermore, we found that IGF-1 and EGF treatment enhances scratch injury-induced proliferation. Finally, we established whole tissue explant cultures and observed that IGF-1 and EGF promote CPEC division within the intact ChP epithelium. We conclude that although CPECs normally have a slow turnover rate, they expand in response to external stimuli such as injury and/or growth factors, which provides a potential avenue for enhancing ChP function after brain injury or neurodegeneration.

  10. Tungsten-induced carcinogenesis in human bronchial epithelial cells

    Science.gov (United States)

    Laulicht, Freda; Brocato, Jason; Cartularo, Laura; Vaughan, Joshua; Wu, Feng; Kluz, Thomas; Sun, Hong; Oksuz, Betul Akgol; Shen, Steven; Paena, Massimilano; Medici, Serenella; Zoroddu, Maria Antonietta; Costa, Max

    2015-01-01

    Metals such as arsenic, cadmium, beryllium, and nickel are known human carcinogens; however, other transition metals, such as tungsten (W), remain relatively uninvestigated with regard to their potential carcinogenic activity. Tungsten production for industrial and military applications has almost doubled over the past decade and continues to increase. Here, for the first time, we demonstrate tungsten’s ability to induce carcinogenic related endpoints including cell transformation, increased migration, xenograft growth in nude mice, and the activation of multiple cancer related pathways in transformed clones as determined by RNA seq. Human bronchial epithelial cell line (Beas-2B) exposed to tungsten developed carcinogenic properties. In a soft agar assay, tungsten-treated cells formed more colonies than controls and the tungsten-transformed clones formed tumors in nude mice. RNA-sequencing data revealed that the tungsten-transformed clones altered the expression of many cancer-associated genes when compared to control clones. Genes involved in lung cancer, leukemia, and general cancer genes were deregulated by tungsten. Taken together, our data shows the carcinogenic potential of tungsten. Further tests are needed, including in vivo and human studies, in order to validate tungsten as a carcinogen to humans. PMID:26164860

  11. Simulating properties of in vitro epithelial cell morphogenesis.

    Directory of Open Access Journals (Sweden)

    Mark R Grant

    2006-10-01

    Full Text Available How do individual epithelial cells (ECs organize into multicellular structures? ECs are studied in vitro to help answer that question. Characteristic growth features include stable cyst formation in embedded culture, inverted cyst formation in suspension culture, and lumen formation in overlay culture. Formation of these characteristic structures is believed to be a consequence of an intrinsic program of differentiation and de-differentiation. To help discover how such a program may function, we developed an in silico analogue in which space, events, and time are discretized. Software agents and objects represent cells and components of the environment. "Cells" act independently. The "program" governing their behavior is embedded within each in the form of axioms and an inflexible decisional process. Relationships between the axioms and recognized cell functions are specified. Interactions between "cells" and environment components during simulation give rise to a complex in silico phenotype characterized by context-dependent structures that mimic counterparts observed in four different in vitro culture conditions: a targeted set of in vitro phenotypic attributes was matched by in silico attributes. However, for a particular growth condition, the analogue failed to exhibit behaviors characteristic of functionally polarized ECs. We solved this problem by following an iterative refinement method that improved the first analogue and led to a second: it exhibited characteristic differentiation and growth properties in all simulated growth conditions. It is the first model to simultaneously provide a representation of nonpolarized and structurally polarized cell types, and a mechanism for their interconversion. The second analogue also uses an inflexible axiomatic program. When specific axioms are relaxed, growths strikingly characteristic of cancerous and precancerous lesions are observed. In one case, the simulated cause is aberrant matrix

  12. Metabolic detoxication pathways for sterigmatocystin in primary tracheal epithelial cells.

    Science.gov (United States)

    Cabaret, Odile; Puel, Olivier; Botterel, Françoise; Pean, Michel; Khoufache, Khaled; Costa, Jean-Marc; Delaforge, Marcel; Bretagne, Stéphane

    2010-11-15

    Human health effects of inhaled mycotoxins remain poorly documented, despite the large amounts present in bioaerosols. Among these mycotoxins, sterigmatocystin is one of the most prevalent. Our aim was to study the metabolism and cellular consequences of sterigmatocystin once it is in contact with the airway epithelium. Metabolites were analyzed first in vitro, using recombinant P450 1A1, 1A2, 2A6, 2A13, and 3A4 enzymes, and subsequently in porcine tracheal epithelial cell (PTEC) primary cultures at an air-liquid interface. Expressed enzymes and PTECs were exposed to sterigmatocystin, uniformly enriched with (13)C to confirm the relationship between sterigmatocystin and metabolites. Induction of the expression of xenobiotic-metabolizing enzymes upon sterigmatocystin exposure was examined by real-time quantitative real-time polymerase chain reaction. Incubation of 50 μM sterigmatocystin with recombinant P450 1A1 led to the formation of three metabolites: monohydroxy-sterigmatocystin (M1), dihydroxy-sterigmatocystin (M2), and one glutathione adduct (M3), the latter after the formation of a transient epoxide. Recombinant P450 1A2 also led to M1 and M3. P450 3A4 led to only M3. In PTEC, 1 μM sterigmatocystin metabolism resulted in a glucuro conjugate (M4) mainly excreted at the basal side of cells. If PTEC were treated with β-naphthoflavone prior to sterigmatocystin incubation, two other products were detected, i.e., a sulfo conjugate (M5) and a glucoro conjugate (M6) of hydroxy-sterigmatocystin. Exposure of PTEC for 24 h to 1 μM sterigmatocystin induced an 18-fold increase in the mRNA levels of P450 1A1, without significantly induced 7-ethoxyresorufin O-deethylation activity. These data suggest that sterigmatocystin is mainly detoxified and is unable to produce significant amounts of reactive epoxide metabolites in respiratory cells. However, sterigmatocystin increases the P450 1A1 mRNA levels with unknown long-term consequences. These in vitro results obtained in

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    Lifescience Database Archive (English)

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  8. Regulatory T Cells in Skin Facilitate Epithelial Stem Cell Differentiation.

    Science.gov (United States)

    Ali, Niwa; Zirak, Bahar; Rodriguez, Robert Sanchez; Pauli, Mariela L; Truong, Hong-An; Lai, Kevin; Ahn, Richard; Corbin, Kaitlin; Lowe, Margaret M; Scharschmidt, Tiffany C; Taravati, Keyon; Tan, Madeleine R; Ricardo-Gonzalez, Roberto R; Nosbaum, Audrey; Bertolini, Marta; Liao, Wilson; Nestle, Frank O; Paus, Ralf; Cotsarelis, George; Abbas, Abul K; Rosenblum, Michael D

    2017-06-01

    The maintenance of tissue homeostasis is critically dependent on the function of tissue-resident immune cells and the differentiation capacity of tissue-resident stem cells (SCs). How immune cells influence the function of SCs is largely unknown. Regulatory T cells (Tregs) in skin preferentially localize to hair follicles (HFs), which house a major subset of skin SCs (HFSCs). Here, we mechanistically dissect the role of Tregs in HF and HFSC biology. Lineage-specific cell depletion revealed that Tregs promote HF regeneration by augmenting HFSC proliferation and differentiation. Transcriptional and phenotypic profiling of Tregs and HFSCs revealed that skin-resident Tregs preferentially express high levels of the Notch ligand family member, Jagged 1 (Jag1). Expression of Jag1 on Tregs facilitated HFSC function and efficient HF regeneration. Taken together, our work demonstrates that Tregs in skin play a major role in HF biology by promoting the function of HFSCs. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Sequestration of human cytomegalovirus by human renal and mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Twite, Nicolas [Institute for Medical Immunology, Université Libre de Bruxelles, Rue A. Bolland 8, B-6041 Charleroi (Belgium); Andrei, Graciela [Laboratory of Virology and Chemotherapy, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven (Belgium); Kummert, Caroline [ImmuneHealth, Rue A. Bolland 8, B-6041 Charleroi (Belgium); Donner, Catherine [Department of Obstetrics and Gynecology, Erasme Hospital, Route de Lennik 808, 1070 Brussels (Belgium); Perez-Morga, David [Laboratory of Molecular Parasitology, Institut de Biologie et Médecine Moléculaires, Université Libre de Bruxelles, Gosselies (Belgium); De Vos, Rita [Pathology Department, U.Z. Leuven, Minderbroedersstraat 12, Leuven (Belgium); Snoeck, Robert, E-mail: Robert.Snoeck@Rega.kuleuven.be [Laboratory of Virology and Chemotherapy, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven (Belgium); Marchant, Arnaud, E-mail: arnaud.marchant@ulb.ac.be [Institute for Medical Immunology, Université Libre de Bruxelles, Rue A. Bolland 8, B-6041 Charleroi (Belgium); ImmuneHealth, Rue A. Bolland 8, B-6041 Charleroi (Belgium)

    2014-07-15

    Urine and breast milk represent the main routes of human cytomegalovirus (HCMV) transmission but the contribution of renal and mammary epithelial cells to viral excretion remains unclear. We observed that kidney and mammary epithelial cells were permissive to HCMV infection and expressed immediate early, early and late antigens within 72 h of infection. During the first 24 h after infection, high titers of infectious virus were measured associated to the cells and in culture supernatants, independently of de novo synthesis of virus progeny. This phenomenon was not observed in HCMV-infected fibroblasts and suggested the sequestration and the release of HCMV by epithelial cells. This hypothesis was supported by confocal and electron microscopy analyses. The sequestration and progressive release of HCMV by kidney and mammary epithelial cells may play an important role in the excretion of the virus in urine and breast milk and may thereby contribute to HCMV transmission. - Highlights: • Primary renal and mammary epithelial cells are permissive to HCMV infection. • HCMV is sequestered by epithelial cells and this phenomenon does not require viral replication. • HCMV sequestration by epithelial cells is reduced by antibodies and IFN-γ.

  10. Romo1 expression contributes to oxidative stress-induced death of lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Jung Ar [Department of Internal Medicine, Yonsei University College of Medicine, Yonsei University Health System, Seoul 135-270 (Korea, Republic of); Chung, Jin Sil [Laboratory of Molecular Cell Biology, College of Life Sciences and Biotechnology, Korea University, Seoul 136-713 (Korea, Republic of); Cho, Sang-Ho [Department of Pathology, Pochon CHA University, College of Medicine, Gyeonggi-do (Korea, Republic of); Kim, Hyung Jung, E-mail: khj57@yuhs.ac.kr [Department of Internal Medicine, Yonsei University College of Medicine, Yonsei University Health System, Seoul 135-270 (Korea, Republic of); Yoo, Young Do, E-mail: ydy1130@korea.ac.kr [Laboratory of Molecular Cell Biology, College of Life Sciences and Biotechnology, Korea University, Seoul 136-713 (Korea, Republic of)

    2013-09-20

    Highlights: •Romo1 mediates oxidative stress-induced mitochondrial ROS production. •Romo1 induction by oxidative stress plays an important role in oxidative stress-induced apoptosis. •Romo1 overexpression correlates with epithelial cell death in patients with IPF. -- Abstract: Oxidant-mediated death of lung epithelial cells due to cigarette smoking plays an important role in pathogenesis in lung diseases such as idiopathic pulmonary fibrosis (IPF). However, the exact mechanism by which oxidants induce epithelial cell death is not fully understood. Reactive oxygen species (ROS) modulator 1 (Romo1) is localized in the mitochondria and mediates mitochondrial ROS production through complex III of the mitochondrial electron transport chain. Here, we show that Romo1 mediates mitochondrial ROS production and apoptosis induced by oxidative stress in lung epithelial cells. Hydrogen peroxide (H{sub 2}O{sub 2}) treatment increased Romo1 expression, and Romo1 knockdown suppressed the cellular ROS levels and cell death triggered by H{sub 2}O{sub 2} treatment. In immunohistochemical staining of lung tissues from patients with IPF, Romo1 was mainly localized in hyperplastic alveolar and bronchial epithelial cells. Romo1 overexpression was detected in 14 of 18 patients with IPF. TUNEL-positive alveolar epithelial cells were also detected in most patients with IPF but not in normal controls. These findings suggest that Romo1 mediates apoptosis induced by oxidative stress in lung epithelial cells.

  11. Effects of Weaning on Intestinal Upper Villus Epithelial Cells of Piglets.

    Directory of Open Access Journals (Sweden)

    Huansheng Yang

    Full Text Available The intestinal upper villus epithelial cells represent the differentiated epithelial cells and play key role in digesting and absorbing lumenal nutrients. Weaning stress commonly results in a decrease in villus height and intestinal dysfunction in piglets. However, no study have been conducted to test the effects of weaning on the physiology and functions of upper villus epithelial cells. A total of 40 piglets from 8 litters were weaned at 14 days of age and one piglet from each litter was killed at 0 d (w0d, 1 d (w1d, 3 d (w3d, 5 d (w5d, and 7 d (w7d after weaning, respectively. The upper villus epithelial cells in mid-jejunum were isolated using the distended intestinal sac method. The expression of proteins in upper villus epithelial cells was analyzed using the isobaric tags for relative and absolute quantification or Western blotting. The expression of proteins involved in energy metabolism, Golgi vesicle transport, protein amino acid glycosylation, secretion by cell, transmembrane transport, ion transport, nucleotide catabolic process, translational initiation, and epithelial cell differentiation and apoptosis, was mainly reduced during the post-weaning period, and these processes may be regulated by mTOR signaling pathway. These results indicated that weaning inhibited various cellular processes in jejunal upper villus epithelial cells, and provided potential new directions for exploring the effects of weaning on the functions of intestine and improving intestinal functions in weaning piglets.

  12. 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....

  13. A Human Corneal Epithelial Cell Line Model for Limbal Stem Cell Biology and Limbal Immunobiology.

    Science.gov (United States)

    Shaharuddin, Bakiah; Ahmad, Sajjad; Md Latar, Nani; Ali, Simi; Meeson, Annette

    2016-10-14

    : Limbal stem cell (LSC) deficiency is a visually debilitating condition caused by abnormal maintenance of LSCs. It is treated by transplantation of donor-derived limbal epithelial cells (LECs), the success of which depends on the presence and quality of LSCs within the transplant. Understanding the immunobiological responses of these cells within the transplants could improve cell engraftment and survival. However, human corneal rings used as a source of LSCs are not always readily available for research purposes. As an alternative, we hypothesized that a human telomerase-immortalized corneal epithelial cell (HTCEC) line could be used as a model for studying LSC immunobiology. HTCEC constitutively expressed human leukocyte antigen (HLA) class I but not class II molecules. However, when stimulated by interferon-γ, HTCECs then expressed HLA class II antigens. Some HTCECs were also migratory in response to CXCL12 and expressed stem cell markers, Nanog, Oct4, and Sox2. In addition because both HTCECs and LECs contain side population (SP) cells, which are an enriched LSC population, we used these SP cells to show that some HTCEC SP cells coexpressed ABCG2 and ABCB5. HTCEC SP and non-side population (NSP) cells also expressed CXCR4, but the SP cells expressed higher levels. Both were capable of colony formation, but the NSP colonies were smaller and contained fewer cells. In addition, HTCECs expressed ΔNp63α. These results suggest the HTCEC line is a useful model for further understanding LSC biology by using an in vitro approach without reliance on a supply of human tissue. Limbal stem cell deficiency is a painful eye condition caused by abnormal maintenance of limbal stem cells. It is treated by transplantation of limbal epithelial cells derived from human tissue. The success of this treatment depends of the quality of the cells transplanted; however, some transplants fail. Understanding more about the immunobiology of these cells within the transplants could

  14. Surface proteins in normal and transformed rat liver epithelial cells in culture.

    Science.gov (United States)

    Bannikov, G. A.; Saint Vincent, L.; Montesano, R.

    1980-01-01

    The pattern of surface proteins of different types of normal and transformed rat liver cells have been studied in culture by means of lactoperoxidase-catalysed iodination procedures, followed by SDS-gel electrophoresis. The cells examined were primary cultures of epithelial liver cells, long-term cultures of epithelial liver cells, in vitro transformed epithelial liver cell lines and liver tumour-cell lines; mesenchymal cells from liver and skin were also examined. The principal surface proteins of primary cultures of epithelial cells from adult or neonatal rats had components with mol. wts of 140,000-160,000, 100,000 and 40,000-70,000. A band that had the same position as fibronectin from mesenchymal cells was also present and this band, as well as other iodinated components, were less sensitive to trypsin than fibroblastic fibronectin. A similar pattern of iodinated proteins was seen in long-term cultures of epithelial liver cells, with a great reduction in the number and intensity of the bands in the mol. wt region below 100,000. Almost all the in vitro transformed and tumour epithelial cell lines contain a protein with a mol. wt 135,000 as one of the major iodinated bands, and in contrast to the observation in transformed fibroblasts, the fibronectin was retained by most of these transformed cell lines. Images Fig. 1 Fig. 2 Fig. 3 PMID:7053205

  15. Regulation of epithelial sodium channel a-subunit expression by adenosine receptor A2a in alveolar epithelial cells

    Institute of Scientific and Technical Information of China (English)

    DENG Wang; WANG Dao-xin; ZHANG Wei; LI Chang-yi

    2011-01-01

    Background The amiloride-sensitive epithelial sodium channel a-subunit (a-ENaC) is an important factor for alveolar fluid clearance during acute lung injury. The relationship between adenosine receptor A2a (A2aAR) expressed in alveolar epithelial cells and aα-ENaC is poorly understood. We targeted the A2aAR in this study to investigate its role in the expression of αa-ENaC and in acute lung injury.Methods A549 cells were incubated with different concentrations of A2aAR agonist CGS-21680 and with 100 μmol/L CGS-21680 for various times. Rats were treated with lipopolysaccharide (LPS) after CGS-21680 was injected. Animals were sacrificed and tissue was harvested for evaluation of lung injury by analysis of the lung wet-to-dry weight ratio, lung permeability and myeloperoxidase activity. RT-PCR and Western blotting were used to determine the mRNA and protein expression levels of α-ENaC in A549 cells and alveolar type II epithelial cells.Results Both mRNA and protein levels of α-ENaC were markedly higher from 4 hours to 24 hours after exposure to 100μmol/L CGS-21680. There were significant changes from 0.1 umol/L to 100 μmol/L CGS-21680, with a positive correlation between increased concentrations of CGS-21680 and expression of α-ENaC. Treatment with CGS-21680during LPS induced lung injury protected the lung and promoted α-ENaC expression in the alveolar epithelial cells.Conclusion Activation of A2aAR has a protective effect during the lung injury, which may be beneficial to the prognosis of acute lung injury.

  16. Alveolar epithelial type II cells induce T cell tolerance to specific antigen

    DEFF Research Database (Denmark)

    Lo, Bernice; Hansen, Søren; Evans, Kathy

    2008-01-01

    The lungs face the immunologic challenge of rapidly eliminating inhaled pathogens while maintaining tolerance to innocuous Ags. A break in this immune homeostasis may result in pulmonary inflammatory diseases, such as allergies or asthma. The observation that alveolar epithelial type II cells (Ty...

  17. Blocking TGF-β expression inhibits silica particle-induced epithelial-mesenchymal transition in human lung epithelial cells.

    Science.gov (United States)

    Rong, Yi; Shen, Yan; Zhang, Zhihong; Cui, Xiuqing; Xiao, Lili; Liu, Yuewei; Luo, Xin; Chen, Weihong

    2015-11-01

    The main characteristic of silicosis is irreversible fibrosis. Certain studies have shown that epithelial-mesenchymal transition (EMT) regulated by transforming growth factor-β (TGF-β) is involved in fibrosis. Thus, we suggest that TGF-β regulated EMT may play an important role in silicosis. In this study, we determined the expression of TGF-β-Smad2/3, EMT- and ECM-related markers in lung epithelial cells treated with silica particle by RT-PCR, western-blot and ELISA. In order to explore the role of TGF-β, we used TGF-β inhibitor in the cell model. We found that the cells lost the expression of epithelial phenotypic markers and acquired increased expression of mesenchymal cells markers with ECM deposition after treatment with silica particle. Moreover, the changes of EMT-related event was restricted in response to TGF-β inhibitor. These findings suggest that EMT is essentially involved in the pathogenesis of fibrosis induced by silica particles and down-regulating the TGF-β expression can inhibit the process of EMT.

  18. Permissiveness of bovine epithelial cells from lung, intestine, placenta and udder for infection with Coxiella burnetii.

    Science.gov (United States)

    Sobotta, Katharina; Bonkowski, Katharina; Liebler-Tenorio, Elisabeth; Germon, Pierre; Rainard, Pascal; Hambruch, Nina; Pfarrer, Christiane; Jacobsen, Ilse D; Menge, Christian

    2017-04-12

    Ruminants are the main source of human infections with the obligate intracellular bacterium Coxiella (C.) burnetii. Infected animals shed high numbers of C. burnetii by milk, feces, and birth products. In goats, shedding by the latter route coincides with C. burnetii replication in epithelial (trophoblast) cells of the placenta, which led us to hypothesize that epithelial cells are generally implicated in replication and shedding of C. burnetii. We therefore aimed at analyzing the interactions of C. burnetii with epithelial cells of the bovine host (1) at the entry site (lung epithelium) which govern host immune responses and (2) in epithelial cells of gut, udder and placenta decisive for the quantity of pathogen excretion. Epithelial cell lines [PS (udder), FKD-R 971 (small intestine), BCEC (maternal placenta), F3 (fetal placenta), BEL-26 (lung)] were inoculated with C. burnetii strains Nine Mile I (NMI) and NMII at different cultivation conditions. The cell lines exhibited different permissiveness for C. burnetii. While maintaining cell viability, udder cells allowed the highest replication rates with formation of large cell-filling Coxiella containing vacuoles. Intestinal cells showed an enhanced susceptibility to invasion but supported C. burnetii replication only at intermediate levels. Lung and placental cells also internalized the bacteria but in strikingly smaller numbers. In any of the epithelial cells, both Coxiella strains failed to trigger a substantial IL-1β, IL-6 and TNF-α response. Epithelial cells, with mammary epithelial cells in particular, may therefore serve as a niche for C. burnetii replication in vivo without alerting the host's immune response.

  19. Kruppel-like factor 4 regulates intestinal epithelial cell morphology and polarity.

    Directory of Open Access Journals (Sweden)

    Tianxin Yu

    Full Text Available Krüppel-like factor 4 (KLF4 is a zinc finger transcription factor that plays a vital role in regulating cell lineage differentiation during development and maintaining epithelial homeostasis in the intestine. In normal intestine, KLF4 is predominantly expressed in the differentiated epithelial cells. It has been identified as a tumor suppressor in colorectal cancer. KLF4 knockout mice demonstrated a decrease in number of goblet cells in the colon, and conditional ablation of KLF4 from the intestinal epithelium led to altered epithelial homeostasis. However, the role of KLF4 in differentiated intestinal cells and colon cancer cells, as well as the mechanism by which it regulates homeostasis and represses tumorigenesis in the intestine is not well understood. In our study, KLF4 was partially depleted in the differentiated intestinal epithelial cells by a tamoxifen-inducible Cre recombinase. We found a significant increase in the number of goblet cells in the KLF4-deleted small intestine, suggesting that KLF4 is not only required for goblet cell differentiation, but also required for maintaining goblet cell numbers through its function in inhibiting cell proliferation. The number and position of Paneth cells also changed. This is consistent with the KLF4 knockout study using villin-Cre [1]. Through immunohistochemistry (IHC staining and statistical analysis, we found that a stem cell and/or tuft cell marker, DCAMKL1, and a proliferation marker, Ki67, are affected by KLF4 depletion, while an enteroendocrine cell marker, neurotensin (NT, was not affected. In addition, we found KLF4 depletion altered the morphology and polarity of the intestinal epithelial cells. Using a three-dimensional (3D intestinal epithelial cyst formation assay, we found that KLF4 is essential for cell polarity and crypt-cyst formation in human colon cancer cells. These findings suggest that, as a tumor suppressor in colorectal cancer, KLF4 affects intestinal epithelial cell

  20. Programmed Cell-to-Cell Variability in Ras Activity Triggers Emergent Behaviors during Mammary Epithelial Morphogenesis

    Directory of Open Access Journals (Sweden)

    Jennifer S. Liu

    2012-11-01

    Full Text Available Variability in signaling pathway activation between neighboring epithelial cells can arise from local differences in the microenvironment, noisy gene expression, or acquired genetic changes. To investigate the consequences of this cell-to-cell variability in signaling pathway activation on coordinated multicellular processes such as morphogenesis, we use DNA-programmed assembly to construct three-dimensional MCF10A microtissues that are mosaic for low-level expression of activated H-Ras. We find two emergent behaviors in mosaic microtissues: cells with activated H-Ras are basally extruded or lead motile multicellular protrusions that direct the collective motility of their wild-type neighbors. Remarkably, these behaviors are not observed in homogeneous microtissues in which all cells express the activated Ras protein, indicating that heterogeneity in Ras activity, rather than the total amount of Ras activity, is critical for these processes. Our results directly demonstrate that cell-to-cell variability in pathway activation within local populations of epithelial cells can drive emergent behaviors during epithelial morphogenesis.

  1. Epithelial monolayer culture system for real-time single-cell analyses.

    Science.gov (United States)

    Seo, Jong Bae; Moody, Mark; Koh, Duk-Su

    2014-01-01

    Abstract Many epithelial cells form polarized monolayers under in vivo and in vitro conditions. Typically, epithelial cells are cultured for differentiation on insert systems where cells are plated on a porous filter membrane. Although the cultured monolayers have been a standard system to study epithelial physiology, there are some limits: The epithelial cells growing inside the commercial inserts are not optimal to visualize directly through lenses on inverted microscopes. The cell images are optically distorted and background fluorescence is bright due to the filter membrane positioned between the cells and the lens. In addition, the cells are not easily accessible by electrodes due to the presence of tall side walls. Here, we present the design, fabrication, and practical applications of an improved system for analysis of polarized epithelial monolayers. This new system allows (1) direct imaging of cells without an interfering filter membrane, (2) electrophysiological measurements, and (3) detection of apical secretion with minimal dilution. Therefore, our culture method is optimized to study differentiated epithelial cells at the single-cell and subcellular levels, and can be extended to other cell types with minor modifications.

  2. Plasticity between Epithelial and Mesenchymal States Unlinks EMT from Metastasis-Enhancing Stem Cell Capacity

    Directory of Open Access Journals (Sweden)

    Evelyne Beerling

    2016-03-01

    Full Text Available Forced overexpression and/or downregulation of proteins regulating epithelial-to-mesenchymal transition (EMT has been reported to alter metastasis by changing migration and stem cell capacity of tumor cells. However, these manipulations artificially keep cells in fixed states, while in vivo cells may adapt transient and reversible states. Here, we have tested the existence and role of epithelial-mesenchymal plasticity in metastasis of mammary tumors without artificially modifying EMT regulators. In these tumors, we found by intravital microscopy that the motile tumor cells have undergone EMT, while their epithelial counterparts were not migratory. Moreover, we found that epithelial-mesenchymal plasticity renders any EMT-induced stemness differences, as reported previously, irrelevant for metastatic outgrowth, because mesenchymal cells that arrive at secondary sites convert to the epithelial state within one or two divisions, thereby obtaining the same stem cell potential as their arrived epithelial counterparts. We conclude that epithelial-mesenchymal plasticity supports migration but additionally eliminates stemness-enhanced metastatic outgrowth differences.

  3. Energy metabolism in intestinal epithelial cells during maturation along the crypt-villus axis

    Science.gov (United States)

    Yang, Huansheng; Wang, Xiaocheng; Xiong, Xia; Yin, Yulong

    2016-01-01

    Intestinal epithelial cells continuously migrate and mature along crypt-villus axis (CVA), while the changes in energy metabolism during maturation are unclear in neonates. The present study was conducted to test the hypothesis that the energy metabolism in intestinal epithelial cells would be changed during maturation along CVA in neonates. Eight 21-day-old suckling piglets were used. Intestinal epithelial cells were isolated sequentially along CVA, and proteomics was used to analyze the changes in proteins expression in epithelial cells along CVA. The identified differentially expressed proteins were mainly involved in cellular process, metabolic process, biological regulation, pigmentation, multicellular organizational process and so on. The energy metabolism in intestinal epithelial cells of piglets was increased from the bottom of crypt to the top of villi. Moreover, the expression of proteins related to the metabolism of glucose, most of amino acids, and fatty acids was increased in intestinal epithelial cells during maturation along CVA, while the expression of proteins related to glutamine metabolism was decreased from crypt to villus tip. The expression of proteins involved in citrate cycle was also increased intestinal epithelial cells during maturation along CVA. Moreover, dietary supplementation with different energy sources had different effects on intestinal structure of weaned piglets. PMID:27558220

  4. Entamoeba histolytica induces cell death of HT29 colonic epithelial cells via NOX1-derived ROS.

    Science.gov (United States)

    Kim, Kyeong Ah; Kim, Ju Young; Lee, Young Ah; Min, Arim; Bahk, Young Yil; Shin, Myeong Heon

    2013-02-01

    Entamoeba histolytica, which causes amoebic colitis and occasionally liver abscess in humans, is able to induce host cell death. However, signaling mechanisms of colon cell death induced by E. histolytica are not fully elucidated. In this study, we investigated the signaling role of NOX in cell death of HT29 colonic epithelial cells induced by E. histolytica. Incubation of HT29 cells with amoebic trophozoites resulted in DNA fragmentation that is a hallmark of apoptotic cell death. In addition, E. histolytica generate intracellular reactive oxygen species (ROS) in a contact-dependent manner. Inhibition of intracellular ROS level with treatment with DPI, an inhibitor of NADPH oxidases (NOXs), decreased Entamoeba-induced ROS generation and cell death in HT29 cells. However, pan-caspase inhibitor did not affect E. histolytica-induced HT29 cell death. In HT29 cells, catalytic subunit NOX1 and regulatory subunit Rac1 for NOX1 activation were highly expressed. We next investigated whether NADPH oxidase 1 (NOX1)-derived ROS is closely associated with HT29 cell death induced by E. histolytica. Suppression of Rac1 by siRNA significantly inhibited Entamoeba-induced cell death. Moreover, knockdown of NOX1 by siRNA, effectively inhibited E. histolytica-triggered DNA fragmentation in HT29 cells. These results suggest that NOX1-derived ROS is required for apoptotic cell death in HT29 colon epithelial cells induced by E. histolytica.

  5. Distribution and number of epidermal growth factor receptors in skin is related to epithelial cell growth

    DEFF Research Database (Denmark)

    Green, M R; Basketter, D A; Couchman, J R

    1983-01-01

    receptors are detected on the epithelial cells overlying the basement membranes of the epidermis, sebaceous gland, and regions of the hair follicle all of which have proliferative capacity. In marked contrast, tissues which have started to differentiate and lost their growth potential, carry either...... and temporal control of epithelial proliferation....

  6. Helicobacter pylori damages human gallbladder epithelial cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Dong-Feng Chen; Lu Hu; Ping Yi; Wei-Wen Liu; Dian-Chun Fang; Hong Cao

    2008-01-01

    AIM: To study the mechanism by which Helicobacter pylori (Hpy/orO damages human gallbladder epithelial cells (HGBEC).METHODS: H pylori isolated from gallbladder were cultured in a liquid medium. Different concentration supernatants and sonicated extracts of H pylori cells were then added to HGBEC in a primary culture. The morphological changes in HGBEC as well as changes in the levels of alkaline phosphatase (ALP), lactate dehydrogenase (LDH) and glutamyltransferase (GGT)were measured.RESULTS: According to the culture curve of HGBEC,it was convenient to study the changes in HGBEC by adding H pylori sonicated extracts and H pylori culture supernatants. Both H pylori sonicated extracts and H pylori culture supernatants had a significant influence on HGBEC morphology, i.e. HGBEC grew more slowly, their viability decreased and their detachment increased. Furthermore, HGBEC ruptured and died. The levels of ALP (33.84 ± 6.00 vs 27.01± 4.67, P < 0.05), LDH (168.37 ± 20.84 vs 55.51 ±17.17, P < 0.01) and GGT (42.01 ± 6.18 vs 25.34 ±4.33, P < 0.01) significantly increased in the HGBEC culture supernatant in a time- and concentrationdependent. The damage to HGBEC in Hpylori culture liquid was more significant than that in H pylori sonicated extracts.CONCLUSION: H pylori induces no obvious damage to HGBEC.

  7. Substrate stiffness regulates extracellular matrix deposition by alveolar epithelial cells

    Science.gov (United States)

    Eisenberg, Jessica L; Safi, Asmahan; Wei, Xiaoding; Espinosa, Horacio D; Budinger, GR Scott; Takawira, Desire; Hopkinson, Susan B; Jones, Jonathan CR

    2012-01-01

    Aim The aim of the study was to address whether a stiff substrate, a model for pulmonary fibrosis, is responsible for inducing changes in the phenotype of alveolar epithelial cells (AEC) in the lung, including their deposition and organization of extracellular matrix (ECM) proteins. Methods Freshly isolated lung AEC from male Sprague Dawley rats were seeded onto polyacrylamide gel substrates of varying stiffness and analyzed for expression and organization of adhesion, cytoskeletal, differentiation, and ECM components by Western immunoblotting and confocal immunofluorescence microscopy. Results We observed that substrate stiffness influences cell morphology and the organization of focal adhesions and the actin cytoskeleton. Surprisingly, however, we found that substrate stiffness has no influence on the differentiation of type II into type I AEC, nor does increased substrate stiffness lead to an epithelial–mesenchymal transition. In contrast, our data indicate that substrate stiffness regulates the expression of the α3 laminin subunit by AEC and the organization of both fibronectin and laminin in their ECM. Conclusions An increase in substrate stiffness leads to enhanced laminin and fibronectin assembly into fibrils, which likely contributes to the disease phenotype in the fibrotic lung. PMID:23204878

  8. Nanoceria have no genotoxic effect on human lens epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Pierscionek, Barbara K; Yasseen, Akeel A [School of Biomedical Sciences, University of Ulster, Coleraine, BT52 1SA (United Kingdom); Li, Yuebin; Schachar, Ronald A; Chen, Wei [Department of Physics, University of Texas at Arlington, Arlington, TX 76019 (United States); Colhoun, Liza M, E-mail: b.pierscionek@ulster.ac.uk, E-mail: weichen@uta.edu [Centre for Vision and Vascular Sciences, School of Medicine, Dentistry and Biomedical Sciences, Queen' s University Belfast, Grosvenor Road, Belfast, BT12 6BA (United Kingdom)

    2010-01-22

    There are no treatments for reversing or halting cataract, a disease of the structural proteins in the eye lens, that has associations with other age-related degenerative conditions such as Alzheimer's disease. The incidence of cataract and associated conditions is increasing as the average age of the population rises. Protein folding diseases are difficult to assess in vivo as proteins and their age-related changes are assessed after extraction. Nanotechnology can be used to investigate protein changes in the intact lens as well as for a potential means of drug delivery. Nanoparticles, such as cerium oxide (CeO{sub 2}) which have antioxidant properties, may even be used as a means of treating cataract directly. Prior to use in treatments, nanoparticle genotoxicity must be tested to assess the extent of any DNA or chromosomal damage. Sister chromatid exchanges were measured and DNA damage investigated using the alkaline COMET assay on cultured human lens epithelial cells, exposed to 5 and 10 {mu}g ml{sup -1} of CeO{sub 2} nanoparticles (nanoceria). Nanoceria at these dosages did not cause any DNA damage or significant increases in the number of sister chromatid exchanges. The absence of genotoxic effects on lens cells suggests that nanoceria, in the doses and exposures tested in this study, are not deleterious to the eye lens and have the potential for use in studying structural alterations, in developing non-surgical cataract treatments and in investigating other protein folding diseases.

  9. Thymic Epithelial Cell Development and Its Dysfunction in Human Diseases

    Directory of Open Access Journals (Sweden)

    Lina Sun

    2014-01-01

    Full Text Available Thymic epithelial cells (TECs are the key components in thymic microenvironment for T cells development. TECs, composed of cortical and medullary TECs, are derived from a common bipotent progenitor and undergo a stepwise development controlled by multiple levels of signals to be functionally mature for supporting thymocyte development. Tumor necrosis factor receptor (TNFR family members including the receptor activator for NFκB (RANK, CD40, and lymphotoxin β receptor (LTβR cooperatively control the thymic medullary microenvironment and self-tolerance establishment. In addition, fibroblast growth factors (FGFs, Wnt, and Notch signals are essential for establishment of functional thymic microenvironment. Transcription factors Foxn1 and autoimmune regulator (Aire are powerful modulators of TEC development, differentiation, and self-tolerance. Dysfunction in thymic microenvironment including defects of TEC and thymocyte development would cause physiological disorders such as tumor, infectious diseases, and autoimmune diseases. In the present review, we will summarize our current understanding on TEC development and the underlying molecular signals pathways and the involvement of thymus dysfunction in human diseases.

  10. Compound exocytosis of casein micelles in mammary epithelial cells.

    Science.gov (United States)

    Dylewski, D P; Keenan, T W

    1983-07-01

    Ultrastructure of lactating bovine and rat mammary epithelial cells was studied with emphasis on secretory vesicle interactions. In the apical zone of the cell, adjacent secretory vesicles formed ball and socket configurations at their points of apposition. Similar configurations were formed between plasma membrane and secretory vesicle membrane. These structures may be formed by the diffusion of water between vesicles with different osmotic potentials. Frequently, vesicular chains consisting of 10 or more linked secretory vesicles were observed. Prior to the exocytotic release of casein micelles, adjacent vesicles fused through fragmentation of the ball and socket membrane. These membrane fragments and the casein micelles appeared to be secreted into the alveolar lumen after passing from one vesicle into another and finally through a pore in the apical plasma membrane. Emptied vesicular chains appeared to collapse and fragmentation of their membrane was observed. Based on these observations, we suggest that most vesicular membrane does not directly contact or become incorporated into the plasma membrane during secretion of the nonfat phase of milk.

  11. Graphene-induced apoptosis in lung epithelial cells through EGFR

    Science.gov (United States)

    Tsai, Shih-Ming; Bangalore, Preeti; Chen, Eric Y.; Lu, David; Chiu, Meng-Hsuen; Suh, Andrew; Gehring, Matthew; Cangco, John P.; Garcia, Santiago G.; Chin, Wei-Chun

    2017-07-01

    Expanding interest in nanotechnology applied to electronic and biomedical fields has led to fast-growing development of various nanomaterials. Graphene is a single-atom thick, two-dimensional sheet of hexagonally arranged carbon atoms with unique physical and chemical properties. Recently, graphene has been used in many studies on electronics, photonics, composite materials, energy generation and storage, sensors, and biomedicine. However, the current health risk assessment for graphene has been relatively limited and inconclusive. This study evaluated the toxicity effects of graphene on the airway epithelial cell line BEAS-2B, which represents the first barrier of the human body to interact with airborne graphene particles. Our result showed that graphene can induce the cellular Ca2+ by phospholipase C (PLC) associated pathway by activating epidermal growth factor receptor (EGFR). Subsequently, inositol 1,4,5-triphosphate (IP3) receptors activate the release of Ca2+ from the endoplasmic reticulum (ER) Ca2+ stores. Those Ca2+ signals further trigger the calcium-regulated apoptosis in the cell. Furthermore, the stimulation can cause EGFR upregulation, which have been demonstrated to associate with diseases such as lung cancer, chronic obstructive pulmonary disease (COPD), and cardiovascular diseases. This study highlights the additional health risk considering that it can function as a contributing factor for other respiratory diseases.

  12. Pseudomonas aeruginosa Transmigrates at Epithelial Cell-Cell Junctions, Exploiting Sites of Cell Division and Senescent Cell Extrusion.

    Directory of Open Access Journals (Sweden)

    Guillaume Golovkine

    2016-01-01

    Full Text Available To achieve systemic infection, bacterial pathogens must overcome the critical and challenging step of transmigration across epithelial barriers. This is particularly true for opportunistic pathogens such as Pseudomonas aeruginosa, an agent which causes nosocomial infections. Despite extensive study, details on the mechanisms used by this bacterium to transmigrate across epithelial tissues, as well as the entry sites it uses, remain speculative. Here, using real-time microscopy and a model epithelial barrier, we show that P. aeruginosa employs a paracellular transmigration route, taking advantage of altered cell-cell junctions at sites of cell division or when senescent cells are expelled from the cell layer. Once a bacterium transmigrates, it is followed by a cohort of bacteria using the same entry point. The basal compartment is then invaded radially from the initial penetration site. Effective transmigration and propagation require type 4 pili, the type 3 secretion system (T3SS and a flagellum, although flagellum-deficient bacteria can occasionally invade the basal compartment from wounded areas. In the basal compartment, the bacteria inject the T3SS toxins into host cells, disrupting the cytoskeleton and focal contacts to allow their progression under the cells. Thus, P. aeruginosa exploits intrinsic host cell processes to breach the epithelium and invade the subcellular compartment.

  13. Identification of stem cells that maintain and regenerate lingual keratinized epithelial cells.

    Science.gov (United States)

    Tanaka, Toshihiro; Komai, Yoshihiro; Tokuyama, Yoko; Yanai, Hirotsugu; Ohe, Shuichi; Okazaki, Kazuichi; Ueno, Hiroo

    2013-05-01

    Lingual keratinized epithelial cells, which constitute the filiform papillae of the tongue, have one of the most rapid tissue turnover rates in the mammalian body and are thought to be the source of squamous cell carcinoma of the tongue. However, the mechanism of tissue maintenance and regeneration is largely unknown for these cells. Here, we show that stem cells positive for Bmi1, keratin 14 and keratin 5 are present in the base but not at the very bottom of the interpapillary pit (observed most frequently in the second or third layer (position +2 or +3) from the basal cells). Using a multicolour lineage tracing method, we demonstrated that one stem cell per interpapillary pit survives long-term. The cells were shown to be unipotent stem cells for keratinized epithelial cells but not for taste bud cells, and were found to usually be in a slow-growing or resting state; however, on irradiation-induced injury, the cells rapidly entered the cell cycle and regenerated tongue epithelium. The elimination of Bmi1-positive stem cells significantly suppressed the regeneration. Taken together, these results suggest that the stem cells identified in this study are important for tissue maintenance and regeneration of the lingual epithelium.

  14. The effect of N-acetylcysteine on chloride efflux from airway epithelial cells.

    Science.gov (United States)

    Varelogianni, Georgia; Oliynyk, Igor; Roomans, Godfried M; Johannesson, Marie

    2010-01-27

    Defective chloride transport in epithelial cells increases mucus viscosity and leads to recurrent infections with high oxidative stress in patients with CF (cystic fibrosis). NAC (N-acetylcysteine) is a well known mucolytic and antioxidant drug, and an indirect precursor of glutathione. Since GSNO (S-nitrosoglutathione) previously has been shown to be able to promote Cl- efflux from CF airway epithelial cells, it was investigated whether NAC also could stimulate Cl- efflux from CF and non-CF epithelial cells and through which mechanisms. CFBE (CF bronchial epithelial cells) and normal bronchial epithelial cells (16HBE) were treated with 1 mM, 5 mM, 10 mM or 15 mM NAC for 4 h at 37 degrees C. The effect of NAC on Cl- transport was measured by Cl- efflux measurements and by X-ray microanalysis. Cl- efflux from CFBE cells was stimulated by NAC in a dose-dependent manner, with 10 mM NAC causing a significant increase in Cl- efflux with nearly 80% in CFBE cells. The intracellular Cl- concentration in CFBE cells was significantly decreased up to 60% after 4 h treatment with 10 mM NAC. Moreover immunocytochemistry and Western blot experiments revealed expression of CFTR channel on CFBE cells after treatment with 10 mM NAC. The stimulation of Cl- efflux by NAC in CF airway epithelial cells may improve hydration of the mucus and thereby be beneficial for CF patients.

  15. Preliminary study on Herpes simplex virus type 1 infection of human oral epithelial cell in vitro

    Institute of Scientific and Technical Information of China (English)

    Jie Zhao; Weibin Sun; Juan Wang

    2008-01-01

    Objective: To explore the functions and mechanisms of herpes simplex virus type 1(HSV-1) while infecting human oral epithelial cells in vitro(being similar to the infection in vivo). Methods:An abundance of HSV-1 strains amplified in Vero cells were used to infect human oral epithelial cells. The culture supernatant was collected to infect Veto cells again. Morphology of HSV-1 was identified by inverted microscope and transmission electron microscope. Nucleic acid of the virus was detected by PCR. Results:The infected human oral epithelial cells didn't display an obvious cytopathic effect(CPE) under inverted microscope(while Veto cells which were infected by the culture supematant showed typical(CPE). The virus particles were not observed in the cytoplasm nor in nucleus of human oral epithelial cells, however under transmission electron microscope in the cytoplasm of Vero cells, the nucleic acid of HSV-1 could be detected in infected human oral epithelial cells, by PCR. Conclusion:HSV-1 can successfully infect human oral epithelial cells. This model may provide a useful approach for studying the pathogenesis of herpes virus-associated periodontal disease.

  16. Human amniotic epithelial cells combined with silk ifbroin scaffold in the repair of spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    Ting-gang Wang; Jie Xu; Ai-hua Zhu; Hua Lu; Zong-ning Miao; Peng Zhao; Guo-zhen Hui; Wei-jiangWu

    2016-01-01

    Treatment and functional reconstruction atfer central nervous system injury is a major medical and social challenge. An increasing number of researchers are attempting to use neural stem cells combined with artiifcial scaffold materials, such as ifbroin, for nerve repair. However, such approaches are challenged by ethical and practical issues. Amniotic tissue, a clinical waste product, is abundant, and amniotic epithe-lial cells are pluripotent, have low immunogenicity, and are not the subject of ethical debate. We hypothesized that amniotic epithelial cells combined with silk ifbroin scaffolds would be conducive to the repair of spinal cord injury. To test this, we isolated and cultured amniotic epithelial cells, and constructed complexes of these cells and silk ifbroin scaffolds. Implantation of the cell-scaffold complex into a rat model of spinal cord injury resulted in a smaller glial scar in the damaged cord tissue than in model rats that received a blank scaffold, or amniotic epithelial cells alone. In addition to a milder local immunological reaction, the rats showed less inlfammatory cell inifltration at the trans-plant site, milder host-versus-gratf reaction, and a marked improvement in motor function. hTese ifndings conifrm that the transplantation of amniotic epithelial cells combined with silk ifbroin scaffold can promote the repair of spinal cord injury. Silk ifbroin scaffold can provide a good nerve regeneration microenvironment for amniotic epithelial cells.

  17. Generation of Alveolar Epithelial Spheroids via Isolated Progenitor Cells from Human Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Shimpei Gotoh

    2014-09-01

    Full Text Available No methods for isolating induced alveolar epithelial progenitor cells (AEPCs from human embryonic stem cells (hESCs and induced pluripotent stem cells (hiPSCs have been reported. Based on a study of the stepwise induction of alveolar epithelial cells (AECs, we identified carboxypeptidase M (CPM as a surface marker of NKX2-1+ “ventralized” anterior foregut endoderm cells (VAFECs in vitro and in fetal human and murine lungs. Using SFTPC-GFP reporter hPSCs and a 3D coculture system with fetal human lung fibroblasts, we showed that CPM+ cells isolated from VAFECs differentiate into AECs, demonstrating that CPM is a marker of AEPCs. Moreover, 3D coculture differentiation of CPM+ cells formed spheroids with lamellar-body-like structures and an increased expression of surfactant proteins compared with 2D differentiation. Methods to induce and isolate AEPCs using CPM and consequently generate alveolar epithelial spheroids would aid human pulmonary disease modeling and regenerative medicine.

  18. Snail involves in the transforming growth factor β1-mediated epithelial-mesenchymal transition of retinal pigment epithelial cells.

    Directory of Open Access Journals (Sweden)

    Hui Li

    Full Text Available BACKGROUND: The proliferation of retinal pigment epithelium (RPE cells resulting from an epithelial-mesenchymal transition (EMT plays a key role in proliferative vitreoretinopathy (PVR, which leads to complex retinal detachment and the loss of vision. Genes of Snail family encode the zinc finger transcription factors that have been reported to be essential in EMT during embryonic development and cancer metastasis. However, the function of Snail in RPE cells undergoing EMT is largely unknown. PRINCIPAL FINDINGS: Transforming growth factor beta(TGF-β-1 resulted in EMT in human RPE cells (ARPE-19, which was characterized by the expected decrease in E-cadherin and Zona occludin-1(ZO-1 expression, and the increase in fibronectin and α-smooth muscle actin (α-SMA expression, as well as the associated increase of Snail expression at both mRNA and protein levels. Furthermore, TGF-β1 treatment caused a significant change in ARPE-19 cells morphology, with transition from a typical epithelial morphology to mesenchymal spindle-shaped. More interestingly, Snail silencing significantly attenuated TGF-β1-induced EMT in ARPE-19 cells by decreasing the mesenchymal markers fibronectin and a-SMA and increasing the epithelial marker E-cadherin and ZO-1. Snail knockdown could effectively suppress ARPE-19 cell migration. Finally, Snail was activated in epiretinal membranes from PVR patients. Taken together, Snail plays very important roles in TGF-β-1-induced EMT in human RPE cells and may contribute to the development of PVR. SIGNIFICANCE: Snail transcription factor plays a critical role in TGF-β1-induced EMT in human RPE cells, which provides deep insight into the pathogenesis of human PVR disease. The specific inhibition of Snail may provide a new approach to treat and prevent PVR.

  19. Influencing factors of rat small intestinal epithelial cell cultivation and effects of radiation on cell proliferation

    Institute of Scientific and Technical Information of China (English)

    Xin Ze Ran; Yong Ping Su; Yong Jiang Wei; Guo Ping Ai; Tian Min Cheng; Yuan Lin

    2001-01-01

    @@ INTRODUCTIONCrypt epithelial cells in normal small intestineproliferate at a high speed. But they are verydifficult to culture in vitro and passage stably. A lotof studies have been done[1-16]. Some domestic labsisolated and cultured crypt cells from embryonalintestines and aseptic animal intestine, but failed.We introduced normal rat epithelial cell line-IEC-6from the USA and its living condition for stablepassage was successfully established after trials. Thecell line was testified to be the small intestinalepithelial cell by electron microscopy,immunihistochemistry and enzymatic histoch-emistry. It has been applied to some relatedresearch work[17-21]. It was found that manyfactors were involved in the culture system. Ourpresent study focuses on the culture method and theinfluencing factors on IEC-6.

  20. Transcriptomic response of goat mammary epithelial cells to Mycoplasma agalactiae challenge – a preliminary study

    DEFF Research Database (Denmark)

    Ogorevc, Jernej; Mihevc, Sonja Prpar; Hedegaard, Jakob

    2015-01-01

    Mycoplasma agalactiae (Ma) is one of the main aetiological agents of intramammary infections in small ruminants, causing contagious agalactia. To better understand the underlying disease patterns a primary goat mammary epithelial cell (pgMEC) culture was established from the mammary tissue......, steroid metabolism, fatty acid metabolism, apoptosis signalling, transcription regulation, and cell cycle regulation. Based on the results we suggest that mammary epithelial cells in vivo contribute to the immune system by the induced expression of cytokines and other chemotactic agents, activation...

  1. GlyRα2, not GlyRα3, modulates the receptive field surround of OFF retinal ganglion cells.

    Science.gov (United States)

    Zhang, Chi; Nobles, Regina D; McCall, Maureen A

    2015-01-01

    Receptive fields (RFs) of most retinal ganglion cells (RGCs) consist of an excitatory center and suppressive surround. The RF center arises from the summation of excitatory bipolar cell glutamatergic inputs, whereas the surround arises from lateral inhibitory inputs. In the retina, both gamma amino butyric acid (GABA) and glycine are inhibitory neurotransmitters. A clear role for GABAergic inhibition modulating the RGC RF surround has been demonstrated across species. Glycinergic inhibition is more commonly associated with RF center modulation, although there is some evidence that it may contribute to the RF surround. The synaptic glycinergic chloride channels are formed by three homomeric β and two homomeric α subunits that can be glycine receptor (GlyR) α1, α2, α3, or α4. GlyRα composition is responsible for currents with distinct decay kinetics. Their expression within the inner plexiform laminae and neuronal subtypes also differ. We studied the role of GlyR subunit selective modulation of RGC RF surrounds, using mice lacking GlyRα2 (Glra2 -/-), GlyRα3 (Glra3 -/-), or both (Glra2/3 -/-). We chose this molecular genetic approach instead of pharmacological manipulation because there are no subunit selective antagonists and strychnine blocks all GlyRs. Comparisons of annulus-evoked responses among wild type (WT) and GlyRα knockouts (Glra2 -/-, Glra3 -/- and Glra2/3 -/-) show that GlyRα2 inhibition enhances RF surround suppression and post-stimulus excitation in only WT OFF RGCs. Similarities in the responses in Glra2 -/- and Glra2/3 -/- RGCs verify these conclusions. Based on previous and current data, we propose that GlyRα2-mediated input uses a crossover inhibitory circuit. Further, we suggest that GlyRα2 modulates the OFF RGC RF center and surround independently. In summary, our results define a selective GlyR subunit-specific control of RF surround suppression in OFF RGCs.

  2. Reactivity of alveolar epithelial cells in primary culture with type I cell monoclonal antibodies.

    Science.gov (United States)

    Danto, S I; Zabski, S M; Crandall, E D

    1992-03-01

    An understanding of the process of alveolar epithelial cell growth and differentiation requires the ability to trace and analyze the phenotypic transitions that the cells undergo. This analysis demands specific phenotypic probes to type II and, especially, type I pneumocytes. To this end, monoclonal antibodies have been generated to type I alveolar epithelial cells using an approach designed to enhance production of lung-specific clones from a crude lung membrane preparation. The monoclonal antibodies were screened by a combination of enzyme-linked immunosorbent assay and immunohistochemical techniques, with the determination of type I cell specificity resting primarily on immunoelectron microscopic localization. Two of these new markers of the type I pneumocyte phenotype (II F1 and VIII B2) were used to analyze primary cultures of type II cells growing on standard tissue culture plastic and on a variety of substrata reported to affect the morphology of these cells in culture. On tissue culture plastic, the antibodies fail to react with early (days 1 to 3) type II cell cultures. The cells become progressively more reactive with time in culture to a plateau of approximately 6 times background by day 8, with a maximum rate of increase between days 3 and 5. This finding is consistent with the hypothesis that type II cells in primary culture undergo at least partial differentiation into type I cells. Type II cells grown on laminin, which reportedly delays the loss of type II cell appearance, and on fibronectin, which has been reported to facilitate cell spreading and loss of type II cell features, develop the type I cell markers during cultivation in vitro with kinetics similar to those on uncoated tissue culture plastic. Cells on type I collagen and on tissue culture-treated Nuclepore filters, which have been reported to support monolayers with type I cell-like morphology, also increase their expression of the II F1 and VIII B2 epitopes around days 3 to 5. Taken

  3. In Vivo Tagging of Lung Epithelial Cells to Define the Early Steps of Tumor Cell Dissemination

    Science.gov (United States)

    2014-10-01

    derived lung cells, only the epithelium will be tagged by this method. Prior to using these animals for our studies, we will ensure that lineage labeled...1 AWARD NUMBER: W81XWH-13-1-0184 TITLE: In Vivo Tagging of Lung Epithelial Cells To Define...REPORT TYPE Annual 3. DATES COVERED 15Sep2013-14Sep2014 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER In Vivo Tagging of Lung

  4. 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.

  5. Isolation of mammary epithelial cells from three-dimensional mixed-cell spheroid co-culture.

    Science.gov (United States)

    Xu, Kun; Buchsbaum, Rachel J

    2012-04-30

    While enormous efforts have gone into identifying signaling pathways and molecules involved in normal and malignant cell behaviors(1-2), much of this work has been done using classical two-dimensional cell culture models, which allow for easy cell manipulation. It has become clear that intracellular signaling pathways are affected by extracellular forces, including dimensionality and cell surface tension(3-4). Multiple approaches have been taken to develop three-dimensional models that more accurately represent biologic tissue architecture(3). While these models incorporate multi-dimensionality and architectural stresses, study of the consequent effects on cells is less facile than in two-dimensional tissue culture due to the limitations of the models and the difficulty in extracting cells for subsequent analysis. The important role of the microenvironment around tumors in tumorigenesis and tumor behavior is becoming increasingly recognized(4). Tumor stroma is composed of multiple cell types and extracellular molecules. During tumor development there are bidirectional signals between tumor cells and stromal cells(5). Although some factors participating in tumor-stroma co-evolution have been identified, there is still a need to develop simple techniques to systematically identify and study the full array of these signals(6). Fibroblasts are the most abundant cell type in normal or tumor-associated stromal tissues, and contribute to deposition and maintenance of basement membrane and paracrine growth factors(7). Many groups have used three dimensional culture systems to study the role of fibroblasts on various cellular functions, including tumor response to therapies, recruitment of immune cells, signaling molecules, proliferation, apoptosis, angiogenesis, and invasion(8-15). We have optimized a simple method for assessing the effects of mammary fibroblasts on mammary epithelial cells using a commercially available extracellular matrix model to create three

  6. Ablation of lung epithelial cells deregulates FGF-10 expression and impairs lung branching morphogenesis.

    Science.gov (United States)

    Kim, Namjin; Yamamoto, Hiroaki; Pauling, Michelle Haynes; Lorizio, Walter; Vu, Thiennu H

    2009-01-01

    Epithelial-mesenchymal interactions are essential for tissue patterning during organogenesis. Distal lung epithelium and its adjacent mesenchyme comprise the epithelial-mesenchymal signaling unit that regulates lung branching morphogenesis. Tissue recombination experiments have demonstrated the importance of mesenchymal signals in inducing lung epithelial differentiation and branching, but the role of the epithelium in regulating mesenchymal signals has not been well characterized. Using transgenic mice, we ablated distal lung epithelial cells during lung development by inducing the expression of a constitutively active proapoptotic Bax protein under the surfactant protein C (SP-C) promoter. We found that epithelial cell ablation results in impaired lung branching morphogenesis, which progresses to emphysematous airspaces in the adults. Mesenchymal expression of fibroblast growth factor 10 (Fgf-10), whose strict spatial and temporal expression is critical for proper lung branching morphogenesis, is disrupted and loses its localized pattern. Interestingly, the expression of sonic hedgehog (Shh), an epithelial gene known to modulate Fgf-10 expression, is unchanged, indicating the existence of other distal epithelial signals that regulate mesenchymal Fgf-10expression. We propose that distal SP-C expressing lung epithelial cells provide essential signals for the downregulation of Fgf-10 expression in the distal mesenchyme during lung development. 292:123-130, 2009. (c) 2008 Wiley-Liss, Inc.

  7. Canine distemper virus epithelial cell infection is required for clinical disease but not for immunosuppression.

    Science.gov (United States)

    Sawatsky, Bevan; Wong, Xiao-Xiang; Hinkelmann, Sarah; Cattaneo, Roberto; von Messling, Veronika

    2012-04-01

    To characterize the importance of infection of epithelial cells for morbillivirus pathogenesis, we took advantage of the severe disease caused by canine distemper virus (CDV) in ferrets. To obtain a CDV that was unable to enter epithelial cells but retained the ability to enter immune cells, we transferred to its attachment (H) protein two mutations shown to interfere with the interaction of measles virus H with its epithelial receptor, human nectin-4. As expected for an epithelial receptor (EpR)-blind CDV, this virus infected dog and ferret epithelial cells inefficiently and did not cause cell fusion or syncytium formation. On the other hand, the EpR-blind CDV replicated in cells expressing canine signaling lymphocyte activation molecule (SLAM), the morbillivirus immune cell receptor, with similar kinetics to those of wild-type CDV. While ferrets infected with wild-type CDV died within 12 days after infection, after developing severe rash and fever, animals infected with the EpR-blind virus showed no clinical signs of disease. Nevertheless, both viruses spread rapidly and efficiently in immune cells, causing similar levels of leukopenia and inhibition of lymphocyte proliferation activity, two indicators of morbillivirus immunosuppression. Infection was documented for airway epithelia of ferrets infected with wild-type CDV but not for those of animals infected with the EpR-blind virus, and only animals infected with wild-type CDV shed virus. Thus, epithelial cell infection is necessary for clinical disease and efficient virus shedding but not for immunosuppression.

  8. Sequential Salinomycin Treatment Results in Resistance Formation through Clonal Selection of Epithelial-Like Tumor Cells

    Directory of Open Access Journals (Sweden)

    Florian Kopp

    2014-12-01

    Full Text Available Acquiring therapy resistance is one of the major obstacles in the treatment of patients with cancer. The discovery of the cancer stem cell (CSC–specific drug salinomycin raised hope for improved treatment options by targeting therapy-refractory CSCs and mesenchymal cancer cells. However, the occurrence of an acquired salinomycin resistance in tumor cells remains elusive. To study the formation of salinomycin resistance, mesenchymal breast cancer cells were sequentially treated with salinomycin in an in vitro cell culture assay, and the resulting differences in gene expression and salinomycin susceptibility were analyzed. We demonstrated that long-term salinomycin treatment of mesenchymal cancer cells resulted in salinomycin-resistant cells with elevated levels of epithelial markers, such as E-cadherin and miR-200c, a decreased migratory capability, and a higher susceptibility to the classic chemotherapeutic drug doxorubicin. The formation of salinomycin resistance through the acquisition of epithelial traits was further validated by inducing mesenchymal-epithelial transition through an overexpression of miR-200c. The transition from a mesenchymal to a more epithelial-like phenotype of salinomycin-treated tumor cells was moreover confirmed in vivo, using syngeneic and, for the first time, transgenic mouse tumor models. These results suggest that the acquisition of salinomycin resistance through the clonal selection of epithelial-like cancer cells could become exploited for improved cancer therapies by antagonizing the tumor-progressive effects of epithelial-mesenchymal transition.

  9. Sequential Salinomycin Treatment Results in Resistance Formation through Clonal Selection of Epithelial-Like Tumor Cells.

    Science.gov (United States)

    Kopp, Florian; Hermawan, Adam; Oak, Prajakta Shirish; Ulaganathan, Vijay Kumar; Herrmann, Annika; Elnikhely, Nefertiti; Thakur, Chitra; Xiao, Zhiguang; Knyazev, Pjotr; Ataseven, Beyhan; Savai, Rajkumar; Wagner, Ernst; Roidl, Andreas

    2014-12-01

    Acquiring therapy resistance is one of the major obstacles in the treatment of patients with cancer. The discovery of the cancer stem cell (CSC)-specific drug salinomycin raised hope for improved treatment options by targeting therapy-refractory CSCs and mesenchymal cancer cells. However, the occurrence of an acquired salinomycin resistance in tumor cells remains elusive. To study the formation of salinomycin resistance, mesenchymal breast cancer cells were sequentially treated with salinomycin in an in vitro cell culture assay, and the resulting differences in gene expression and salinomycin susceptibility were analyzed. We demonstrated that long-term salinomycin treatment of mesenchymal cancer cells resulted in salinomycin-resistant cells with elevated levels of epithelial markers, such as E-cadherin and miR-200c, a decreased migratory capability, and a higher susceptibility to the classic chemotherapeutic drug doxorubicin. The formation of salinomycin resistance through the acquisition of epithelial traits was further validated by inducing mesenchymal-epithelial transition through an overexpression of miR-200c. The transition from a mesenchymal to a more epithelial-like phenotype of salinomycin-treated tumor cells was moreover confirmed in vivo, using syngeneic and, for the first time, transgenic mouse tumor models. These results suggest that the acquisition of salinomycin resistance through the clonal selection of epithelial-like cancer cells could become exploited for improved cancer therapies by antagonizing the tumor-progressive effects of epithelial-mesenchymal transition.

  10. Airway Epithelial Cell Integrity Protects from Cytotoxicity of Pseudomonas aeruginosa Quorum-Sensing Signals.

    Science.gov (United States)

    Losa, Davide; Köhler, Thilo; Bacchetta, Marc; Saab, Joanna Bou; Frieden, Maud; van Delden, Christian; Chanson, Marc

    2015-08-01

    Cell-to-cell communication via gap junctions regulates airway epithelial cell homeostasis and maintains the epithelium host defense. Quorum-sensing molecules produced by Pseudomonas aeruginosa coordinate the expression of virulence factors by this respiratory pathogen. These bacterial signals may also incidentally modulate mammalian airway epithelial cell responses to the pathogen, a process called interkingdom signaling. We investigated the interactions between the P. aeruginosa N-3-oxo-dodecanoyl-L-homoserine lactone (C12) quorum-sensing molecule and human airway epithelial cell gap junctional intercellular communication (GJIC). C12 degradation and its effects on cells were monitored in various airway epithelial cell models grown under nonpolarized and polarized conditions. Its concentration was further monitored in daily tracheal aspirates of colonized intubated patients. C12 rapidly altered epithelial integrity and decreased GJIC in nonpolarized airway epithelial cells, whereas other quorum-sensing molecules had no effect. The effects of C12 were dependent on [Ca(2+)]i and could be prevented by inhibitors of Src tyrosine family and Rho-associated protein kinases. In contrast, polarized airway cells grown on Transwell filters were protected from C12 except when undergoing repair after wounding. In vivo during colonization of intubated patients, C12 did not accumulate, but it paralleled bacterial densities. In vitro C12 degradation, a reaction catalyzed by intracellular paraoxonase 2 (PON2), was impaired in nonpolarized cells, whereas PON2 expression was increased during epithelial polarization. The cytotoxicity of C12 on nonpolarized epithelial cells, combined with its impaired degradation allowing its accumulation, provides an additional pathogenic mechanism for P. aeruginosa infections.

  11. Cellular heterogeneity in the mouse esophagus implicates the presence of a nonquiescent epithelial stem cell population.

    Science.gov (United States)

    DeWard, Aaron D; Cramer, Julie; Lagasse, Eric

    2014-10-23

    Because the esophageal epithelium lacks a defined stem cell niche, it is unclear whether all basal epithelial cells in the adult esophagus are functionally equivalent. In this study, we showed that basal cells in the mouse esophagus contained a heterogeneous population of epithelial cells, similar to other rapidly cycling tissues such as the intestine or skin. Using a combination of cell-surface markers, we separated primary esophageal tissue into distinct cell populations that harbored differences in stem cell potential. We also used an in vitro 3D organoid assay to demonstrate that Sox2, Wnt, and bone morphogenetic protein signaling regulate esophageal self-renewal. Finally, we labeled proliferating basal epithelial cells in vivo to show differing cell-cycle profiles and proliferation kinetics. Based on our results, we propose that a nonquiescent stem cell population resides in the basal epithelium of the mouse esophagus.

  12. Cellular Heterogeneity in the Mouse Esophagus Implicates the Presence of a Nonquiescent Epithelial Stem Cell Population

    Directory of Open Access Journals (Sweden)

    Aaron D. DeWard

    2014-10-01

    Full Text Available Because the esophageal epithelium lacks a defined stem cell niche, it is unclear whether all basal epithelial cells in the adult esophagus are functionally equivalent. In this study, we showed that basal cells in the mouse esophagus contained a heterogeneous population of epithelial cells, similar to other rapidly cycling tissues such as the intestine or skin. Using a combination of cell-surface markers, we separated primary esophageal tissue into distinct cell populations that harbored differences in stem cell potential. We also used an in vitro 3D organoid assay to demonstrate that Sox2, Wnt, and bone morphogenetic protein signaling regulate esophageal self-renewal. Finally, we labeled proliferating basal epithelial cells in vivo to show differing cell-cycle profiles and proliferation kinetics. Based on our results, we propose that a nonquiescent stem cell population resides in the basal epithelium of the mouse esophagus.

  13. Sds22, a PP1 phosphatase regulatory subunit, regulates epithelial cell polarity and shape [Sds22 in epithelial morphology

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    Sung Hsin-Ho

    2009-02-01

    Full Text Available Abstract Background How epithelial cells adopt their particular polarised forms is poorly understood. In a screen for genes regulating epithelial morphology in Drosophila, we identified sds22, a conserved gene previously characterised in yeast. Results In the columnar epithelia of imaginal discs or follicle cells, mutation of sds22 causes contraction of cells along their apical-basal axis, resulting in a more cuboidal morphology. In addition, the mutant cells can also display altered cell polarity, forming multiple layers in follicle cells and leaving the epithelium in imaginal discs. In yeast, sds22 encodes a PP1 phosphatase regulatory subunit. Consistent with this, we show that Drosophila Sds22 binds to all four Drosophila PP1s and shares an overlapping phenotype with PP1beta9c. We also show that two previously postulated PP1 targets, Spaghetti Squash and Moesin are hyper-phosphorylated in sds22 mutants. This function is shared by the human homologue of Sds22, PPP1R7. Conclusion Sds22 is a conserved PP1 phosphatase regulatory subunit that controls cell shape and polarity.

  14. Nanoparticle-mediated delivery of the antimicrobial peptide plectasin against Staphylococcus aureus in infected epithelial cells

    DEFF Research Database (Denmark)

    Water, Jorrit Jeroen; Smart, Simon; Franzyk, Henrik

    2015-01-01

    intracellularly in Calu-3 epithelial cells and in THP-1 cells, whereas A549 cells did not show significant uptake of nanoparticles. Overall, encapsulation of plectasin into PLGA-based nanoparticles appears to be a viable strategy to improve the efficacy of plectasin against infections in epithelial tissues....... epithelial cells might thus be a promising approach to combat such infections. In this work, plectasin, which is a cationic AMP of the defensin class, was encapsulated into poly(lactic-co-glycolic acid) (PLGA) nanoparticles using the double emulsion solvent evaporation method. The nanoparticles displayed...... high plectasin encapsulation efficiency (71-90%) and mediated release of the peptide over 24h. The antimicrobial efficacy of the peptide-loaded nanoparticles was investigated using bronchiolar epithelial Calu-3 cell monolayers infected with S. aureus. The plectasin-loaded nanoparticles displayed...

  15. Hormonal regulation of leucine catabolism in mammary epithelial cells.

    Science.gov (United States)

    Lei, Jian; Feng, Dingyuan; Zhang, Yongliang; Dahanayaka, Sudath; Li, Xilong; Yao, Kang; Wang, Junjun; Wu, Zhenlong; Dai, Zhaolai; Wu, Guoyao

    2013-09-01

    Branched-chain amino acids (BCAA) are actively taken up and catabolized by the mammary gland during lactation for syntheses of glutamate, glutamine and aspartate. Available evidence shows that the onset of lactation is associated with increases in circulating levels of cortisol, prolactin and glucagon, but decreases in insulin and growth hormone. This study determined the effects of physiological concentrations of these hormones on the catabolism of leucine (a representative BCAA) in bovine mammary epithelial cells. Cells were incubated at 37 °C for 2 h in Krebs buffer containing 3 mM D-glucose, 0.5 mM L-leucine, L-[1-14C]leucine or L-[U-14C]leucine, and 0-50 μU/mL insulin, 0-20 ng/mL growth hormone 0-200 ng/mL prolactin, 0-150 nM cortisol or 0-300 pg/mL glucagon. Increasing extracellular concentrations of insulin did not affect leucine transamination or oxidative decarboxylation, but decreased the rate of oxidation of leucine carbons 2-6. Elevated levels of growth hormone dose dependently inhibited leucine catabolism, α-ketoisocaproate (KIC) production and the syntheses of glutamate plus glutamine. In contrast, cortisol and glucagon increased leucine transamination, leucine oxidative decarboxylation, KIC production, the oxidation of leucine 2-6 carbons and the syntheses of glutamate plus glutamine. Prolactin did not affect leucine catabolism in the cells. The changes in leucine degradation were consistent with alterations in abundances of BCAA transaminase and phosphorylated levels of branched-chain α-ketoacid dehydrogenase. Reductions in insulin and growth hormone but increases in cortisol and glucagon with lactation act in concert to stimulate BCAA catabolism for glutamate and glutamine syntheses. These coordinated changes in hormones may facilitate milk production in lactating mammals.

  16. Differentiated transplant derived airway epithelial cell cytokine secretion is not regulated by cyclosporine

    Science.gov (United States)

    2011-01-01

    Background While lung transplantation is an increasingly utilized therapy for advanced lung diseases, chronic rejection in the form of Bronchiolitis Obliterans Syndrome (BOS) continues to result in significant allograft dysfunction and patient mortality. Despite correlation of clinical events with eventual development of BOS, the causative pathophysiology remains unknown. Airway epithelial cells within the region of inflammation and fibrosis associated with BOS may have a participatory role. Methods Transplant derived airway epithelial cells differentiated in air liquid interface culture were treated with IL-1β and/or cyclosporine, after which secretion of cytokines and growth factor and gene expression for markers of epithelial to mesenchymal transition were analyzed. Results Secretion of IL-6, IL-8, and TNF-α, but not TGF-β1, was increased by IL-1β stimulation. In contrast to previous studies using epithelial cells grown in submersion culture, treatment of differentiated cells in ALI culture with cyclosporine did not elicit cytokine or growth factor secretion, and did not alter IL-6, IL-8, or TNF-α production in response to IL-1β treatment. Neither IL-1β nor cyclosporine elicited expression of markers of the epithelial to mesenchymal transition E-cadherin, EDN-fibronectin, and α-smooth muscle actin. Conclusion Transplant derived differentiated airway epithelial cell IL-6, IL-8, and TNF-α secretion is not regulated by cyclosporine in vitro; these cells thus may participate in local inflammatory responses in the setting of immunosuppression. Further, treatment with IL-1β did not elicit gene expression of markers of epithelial to mesenchymal transition. These data present a model of differentiated airway epithelial cells that may be useful in understanding epithelial participation in airway inflammation and allograft rejection in lung transplantation. PMID:21477368

  17. Origins and consequences of hyperosmolar stress in retinal pigmented epithelial cells.

    Science.gov (United States)

    Willermain, François; Libert, Sarah; Motulsky, Elie; Salik, Dany; Caspers, Laure; Perret, Jason; Delporte, Christine

    2014-01-01

    The retinal pigmented epithelium (RPE) is composed of retinal pigmented epithelial cells joined by tight junctions and represents the outer blood-retinal barrier (BRB). The inner BRB is made of endothelial cells joined by tight junctions and glial extensions surrounding all the retinal blood vessels. One of the functions of the RPE is to maintain an osmotic transepithelial gradient created by ionic pumps and channels, avoiding paracellular flux. Under such physiological conditions, transcellular water movement follows the osmotic gradient and flows normally from the retina to the choroid through the RPE. Several diseases, such as diabetic retinopathy, are characterized by the BRB breakdown leading to leakage of solutes, proteins, and fluid from the retina and the choroid. The prevailing hypothesis explaining macular edema formation during diabetic retinopathy incriminates the inner BRB breakdown resulting in increased osmotic pressure leading in turn to massive water accumulation that can affect vision. Under these conditions, it has been hypothesized that RPE is likely to be exposed to hyperosmolar stress at its apical side. This review summarizes the origins and consequences of osmotic stress in the RPE. Ongoing and further research advances will clarify the mechanisms, at the molecular level, involved in the response of the RPE to osmotic stress and delineate potential novel therapeutic targets and tools.

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

    Energy Technology Data Exchange (ETDEWEB)

    Kikuta, Kazuhiro [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Masamune, Atsushi, E-mail: amasamune@med.tohoku.ac.jp [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Watanabe, Takashi; Ariga, Hiroyuki; Itoh, Hiromichi; Hamada, Shin; Satoh, Kennichi [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan); Egawa, Shinichi; Unno, Michiaki [Department of Hepatobiliary-Pancreatic Surgery, Tohoku University Graduate School of Medicine, Sendai (Japan); Shimosegawa, Tooru [Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai (Japan)

    2010-12-17

    Research highlights: {yields} Recent studies have shown that pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. {yields} Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and scattered, fibroblast-like appearance. {yields} PSCs decreased the expression of epithelial markers but increased that of mesenchymal markers, along with increased migration. {yields} 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 {beta}-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

  19. Identification of DNA aptamers toward epithelial cell adhesion molecule via cell-SELEX.

    Science.gov (United States)

    Kim, Ji Won; Kim, Eun Young; Kim, Sun Young; Byun, Sang Kyung; Lee, Dasom; Oh, Kyoung-Jin; Kim, Won Kon; Han, Baek Soo; Chi, Seung-Wook; Lee, Sang Chul; Bae, Kwang-Hee

    2014-10-31

    The epithelial cell adhesion molecule (EpCAM, also known as CD326) is a transmembrane glycoprotein that is specifically detected in most adenocarcinomas and cancer stem cells. In this study, we performed a Cell systematic evolution of ligands by exponential enrichment (SELEX) experiment to isolate the aptamers against EpCAM. After seven round of Cell SELEX, we identified several aptamer candidates. Among the selected aptamers, EP166 specifically binds to cells expressing EpCAM with an equilibrium dissociation constant (Kd) in a micromolar range. On the other hand, it did not bind to negative control cells. Moreover, EP166 binds to J1ES cells, a mouse embryonic stem cell line. Therefore, the isolated aptamers against EpCAM could be used as a stem cell marker or in other applications in both stem cell and cancer studies.

  20. Characteristic and functional analysis of a newly established porcine small intestinal epithelial cell line.

    Directory of Open Access Journals (Sweden)

    Jing Wang

    Full Text Available The mucosal surface of intestine is continuously exposed to both potential pathogens and beneficial commensal microorganisms. Recent findings suggest that intestinal epithelial cells, which once considered as a simple physical barrier, are a crucial cell lineage necessary for maintaining intestinal immune homeostasis. Therefore, establishing a stable and reliable intestinal epithelial cell line for future research on the mucosal immune system is necessary. In the present study, we established a porcine intestinal epithelial cell line (ZYM-SIEC02 by introducing the human telomerase reverse transcriptase (hTERT gene into small intestinal epithelial cells derived from a neonatal, unsuckled piglet. Morphological analysis revealed a homogeneous cobblestone-like morphology of the epithelial cell sheets. Ultrastructural indicated the presence of microvilli, tight junctions, and a glandular configuration typical of the small intestine. Furthermore, ZYM-SIEC02 cells expressed epithelial cell-specific markers including cytokeratin 18, pan-cytokeratin, sucrase-isomaltase, E-cadherin and ZO-1. Immortalized ZYM-SIEC02 cells remained diploid and were not transformed. In addition, we also examined the host cell response to Salmonella and LPS and verified the enhanced expression of mRNAs encoding IL-8 and TNF-α by infection with Salmonella enterica serovars Typhimurium (S. Typhimurium. Results showed that IL-8 protein expression were upregulated following Salmonella invasion. TLR4, TLR6 and IL-6 mRNA expression were upregulated following stimulation with LPS, ZYM-SIEC02 cells were hyporeponsive to LPS with respect to IL-8 mRNA expression and secretion. TNFα mRNA levels were significantly decreased after LPS stimulation and TNF-α secretion were not detected challenged with S. Typhimurium neither nor LPS. Taken together, these findings demonstrate that ZYM-SIEC02 cells retained the morphological and functional characteristics typical of primary swine

  1. Identification of human fibroblast cell lines as a feeder layer for human corneal epithelial regeneration.

    Directory of Open Access Journals (Sweden)

    Rong Lu

    Full Text Available There is a great interest in using epithelium generated in vitro for tissue bioengineering. Mouse 3T3 fibroblasts have been used as a feeder layer to cultivate human epithelia including corneal epithelial cells for more than 3 decades. To avoid the use of xeno-components, we evaluated human fibroblasts as an alternative feeder supporting human corneal epithelial regeneration. Five human fibroblast cell lines were used for evaluation with mouse 3T3 fibroblasts as a control. Human epithelial cells isolated from fresh corneal limbal tissue were seeded on these feeders. Colony forming efficiency (CFE and cell growth capacity were evaluated on days 5-14. The phenotype of the regenerated epithelia was evaluated by morphology and immunostaining with epithelial markers. cDNA microarray was used to analyze the gene expression profile of the supportive human fibroblasts. Among 5 strains of human fibroblasts evaluated, two newborn foreskin fibroblast cell lines, Hs68 and CCD1112Sk, were identified to strongly support human corneal epithelial growth. Tested for 10 passages, these fibroblasts continually showed a comparative efficiency to the 3T3 feeder layer for CFE and growth capacity of human corneal epithelial cells. Limbal epithelial cells seeded at 1 × 10(4 in a 35-mm dish (9.6 cm(2 grew to confluence (about 1.87-2.41 × 10(6 cells in 12-14 days, representing 187-241 fold expansion with over 7-8 doublings on these human feeders. The regenerated epithelia expressed K3, K12, connexin 43, p63, EGFR and integrin β1, resembling the phenotype of human corneal epithelium. DNA microarray revealed 3 up-regulated and 10 down-regulated genes, which may be involved in the functions of human fibroblast feeders. These findings demonstrate that commercial human fibroblast cell lines support human corneal epithelial regeneration, and have potential use in tissue bioengineering for corneal reconstruction.

  2. Molecular basis of potassium channels in pancreatic duct epithelial cells

    DEFF Research Database (Denmark)

    Hayashi, M.; Novak, Ivana

    2013-01-01

    Potassium channels regulate excitability, epithelial ion transport, proliferation, and apoptosis. In pancreatic ducts, K channels hyperpolarize the membrane potential and provide the driving force for anion secretion. This review focuses on the molecular candidates of functional K channels...

  3. Molecular basis of potassium channels in pancreatic duct epithelial cells

    DEFF Research Database (Denmark)

    Hayashi, M.; Novak, Ivana

    2013-01-01

    Potassium channels regulate excitability, epithelial ion transport, proliferation, and apoptosis. In pancreatic ducts, K channels hyperpolarize the membrane potential and provide the driving force for anion secretion. This review focuses on the molecular candidates of functional K channels in pan...

  4. Distinct mesenchymal alterations in N-cadherin and E-cadherin positive primary renal epithelial cells.

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    Christof Keller

    Full Text Available BACKGROUND: Renal tubular epithelial cells of proximal and distal origin differ markedly in their physiological functions. Therefore, we hypothesized that they also differ in their capacity to undergo epithelial to mesenchymal alterations. RESULTS: We used cultures of freshly isolated primary human tubular cells. To distinguish cells of different tubular origin we took advantage of the fact that human proximal epithelial cells uniquely express N-cadherin instead of E-cadherin as major cell-cell adhesion molecule. To provoke mesenchymal alteration we treated these cocultures with TGF-β for up to 6 days. Within this time period, the morphology of distal tubular cells was barely altered. In contrast to tubular cell lines, E-cadherin was not down-regulated by TGF-β, even though TGF-β signal transduction was initiated as demonstrated by nuclear localization of Smad2/3. Analysis of transcription factors and miRNAs possibly involved in E-cadherin regulation revealed high levels of miRNAs of the miR200-family, which may contribute to the stability of E-cadherin expression in human distal tubular epithelial cells. By contrast, proximal tubular epithelial cells altered their phenotype when treated with TGF-β. They became elongated and formed three-dimensional structures. Rho-kinases were identified as modulators of TGF-β-induced morphological alterations. Non-specific inhibition of Rho-kinases resulted in stabilization of the epithelial phenotype, while partial effects were observed upon downregulation of Rho-kinase isoforms ROCK1 and ROCK2. The distinct reactivity of proximal and distal cells was retained when the cells were cultured as polarized cells. CONCLUSIONS: Interference with Rho-kinase signaling provides a target to counteract TGF-β-mediated mesenchymal alterations of epithelial cells, particularly in proximal tubular epithelial cells. Furthermore, primary distal tubular cells differed from cell lines by their high phenotypic stability

  5. Detonation Nanodiamond Toxicity in Human Airway Epithelial Cells Is Modulated by Air Oxidation

    Science.gov (United States)

    Detonational nanodiamonds (DND), a nanomaterial with an increasing range of industrial and biomedical applications, have previously been shown to induce a pro-inflammatory response in cultured human airway epithelial cells (HAEC). We now show that surface modifications induced by...

  6. Regulation of antiapoptotic and cytoprotective pathways in colonic epithelial cells in ulcerative colitis

    DEFF Research Database (Denmark)

    Seidelin, Jakob B

    2015-01-01

    Ulcerative colitis is an inflammatory bowel disease involving the colon resulting in bloody diarrhea and increased risk of colorectal cancer in certain patient subgroups. Increased apoptosis in the epithelial cell layer causes increased permeability, especially during flares; this leads...

  7. Role of Allergen Source-Derived Proteases in Sensitization via Airway Epithelial Cells

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    Yasuhiro Matsumura

    2012-01-01

    Full Text Available Protease activity is a characteristic common to many allergens. Allergen source-derived proteases interact with lung epithelial cells, which are now thought to play vital roles in both innate and adaptive immune responses. Allergen source-derived proteases act on airway epithelial cells to induce disruption of the tight junctions between epithelial cells, activation of protease-activated receptor-2, and the production of thymic stromal lymphopoietin. These facilitate allergen delivery across epithelial layers and enhance allergenicity or directly activate the immune system through a nonallergic mechanism. Furthermore, they cleave regulatory cell surface molecules involved in allergic reactions. Thus, allergen source-derived proteases are a potentially critical factor in the development of allergic sensitization and appear to be strongly associated with heightened allergenicity.

  8. Puerarin antagonizes peroxyntrite-induced injury in retinal pigment epithelial cells

    Institute of Scientific and Technical Information of China (English)

    Lina Hao; Xudong Zhang; Tao Yang; Junling Ma

    2012-01-01

    A rat model of diabetes mellitus was established by intraperitoneal injection of streptozotocin. Three days later, the rats were intraperitoneally administered 140 mg puerarin/kg daily, for a total of 60 successive days. DNA ladder results showed increased apoptosis over time in retinal pigment epithelial cells from rats with streptozotocin-induced diabetes mellitus. Western blot analysis, Reverse transcription-PCR, immunohistochemistry, and flow cytometry results showed increased expression of 3-nitrotyrosine, a peroxyntrite marker, as well as inducible nitric synthase and Fas/FasL, in retinal pigment epithelial cells. Puerarin reversed these changes, and results demonstrated that puerarin inhibited Fas/FasL expression and alleviated peroxyntrite injury to retinal pigment epithelial cells. These results suggested that puerarin inhibited production of inducible nitric oxide synthase and directly antagonized peroxyntrite injury in retinal pigment epithelial cells.

  9. Cytological analysis of the epithelial cells in patients with oral candidiasis.

    Science.gov (United States)

    Loss, Rafael; Sandrin, Rodrigo; França, Beatriz Helena Sottile; de Azevedo-Alanis, Luciana Reis; Grégio, Ana Maria Trindade; Machado, Maria Ângela Naval; de Lima, Antonio Adilson Soares

    2011-07-01

    The aim of this study was to evaluate oral epithelial cells of the oral mucosa infected by Candida albicans using exfoliative cytology. Oral smears were collected from clinically normal-appearing mucosa by liquid-based exfoliative cytology of 60 individuals (30 patients with oral candidiasis and 30 healthy controls matched for age and gender) and analysed for morphologic and cytomorphometric technique. Morphologically, candida-infected epithelial cells exhibited nuclear enlargement, perinuclear rings, discrete orangeophilia, and cytoplasmic vacuoles. The cytomorphometric analysis demonstrated that the cytoplasmic area (CA) of the epithelial cells was diminished in patients undergoing candidiasis as compared to the non-infected controls. In addition, there was an augmentation in nuclear area (NA) and NA/CA area ratio. This study revealed that oral mucosa of patients undergoing candidal infection exhibited significant changes in the size and shape of the oral epithelial cells.

  10. Quantitative analysis of epithelial cells in urine from men with and without urethritis: implications for studying epithelial: pathogen interactions in vivo

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    Whittington Kate

    2009-07-01

    Full Text Available Abstract Background Epithelial cells in first catch urine (FCU specimens from 87 men with and without urethritis were quantified. Epithelial cells were broadly categorised into transitional and squamous populations using morphological characteristics and immunostaining with anti-pan leukocyte and anti-cytokeratin monoclonal antibodies. Findings The majority (77/87 = 89% of samples contained both transitional (76/87 = 87%; range 1 × 104 – 6 × 105, median 6 × 104 and squamous (57/87 = 66%; range 1 × 104 – 8 × 105, median 2 × 104 epithelial cells. The number of transitional cells correlated with the number of squamous cells (Spearman's rho = 0.697 p Conclusion Further studies are required to explore the complexity of epithelial cell populations in urine. These would provide novel opportunities for studying cellular interactions of C. trachomatis in male urethral infections, about which little is currently known.

  11. Genomic dissection of conserved transcriptional regulation in intestinal epithelial cells

    Science.gov (United States)

    Camp, J. Gray; Weiser, Matthew; Cocchiaro, Jordan L.; Kingsley, David M.; Furey, Terrence S.; Sheikh, Shehzad Z.; Rawls, John F.

    2017-01-01

    The intestinal epithelium serves critical physiologic functions that are shared among all vertebrates. However, it is unknown how the transcriptional regulatory mechanisms underlying these functions have changed over the course of vertebrate evolution. We generated genome-wide mRNA and accessible chromatin data from adult intestinal epithelial cells (IECs) in zebrafish, stickleback, mouse, and human species to determine if conserved IEC functions are achieved through common transcriptional regulation. We found evidence for substantial common regulation and conservation of gene expression regionally along the length of the intestine from fish to mammals and identified a core set of genes comprising a vertebrate IEC signature. We also identified transcriptional start sites and other putative regulatory regions that are differentially accessible in IECs in all 4 species. Although these sites rarely showed sequence conservation from fish to mammals, surprisingly, they drove highly conserved IEC expression in a zebrafish reporter assay. Common putative transcription factor binding sites (TFBS) found at these sites in multiple species indicate that sequence conservation alone is insufficient to identify much of the functionally conserved IEC regulatory information. Among the rare, highly sequence-conserved, IEC-specific regulatory regions, we discovered an ancient enhancer upstream from her6/HES1 that is active in a distinct population of Notch-positive cells in the intestinal epithelium. Together, these results show how combining accessible chromatin and mRNA datasets with TFBS prediction and in vivo reporter assays can reveal tissue-specific regulatory information conserved across 420 million years of vertebrate evolution. We define an IEC transcriptional regulatory network that is shared between fish and mammals and establish an experimental platform for studying how evolutionarily distilled regulatory information commonly controls IEC development and physiology. PMID

  12. Hydrogen peroxide contributes to the epithelial cell death induced by the oral mitis group of streptococci.

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    Nobuo Okahashi

    Full Text Available Members of the mitis group of streptococci are normal inhabitants of the commensal flora of the oral cavity and upper respiratory tract of humans. Some mitis group species, such as Streptococcus oralis and Streptococcus sanguinis, are primary colonizers of the human oral cavity. Recently, we found that hydrogen peroxide (H2O2 produced by S. oralis is cytotoxic to human macrophages, suggesting that streptococcus-derived H2O2 may act as a cytotoxin. Since epithelial cells provide a physical barrier against pathogenic microbes, we investigated their susceptibility to infection by H2O2-producing streptococci in this study. Infection by S. oralis and S. sanguinis was found to stimulate cell death of Detroit 562, Calu-3 and HeLa epithelial cell lines at a multiplicity of infection greater than 100. Catalase, an enzyme that catalyzes the decomposition of H2O2, inhibited S. oralis cytotoxicity, and H2O2 alone was capable of eliciting epithelial cell death. Moreover, S. oralis mutants lacking the spxB gene encoding pyruvate oxidase, which are deficient in H2O2 production, exhibited reduced cytotoxicity toward Detroit 562 epithelial cells. In addition, enzyme-linked immunosorbent assays revealed that both S. oralis and H2O2 induced interleukin-6 production in Detroit 562 epithelial cells. These results suggest that streptococcal H2O2 is cytotoxic to epithelial cells, and promotes bacterial evasion of the host defense systems in the oral cavity and upper respiratory tracts.

  13. Epithelial cell differentiation regulated by MicroRNA-200a in mammary glands.

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    Kentaro Nagaoka

    Full Text Available Mammary gland epithelial cells undergo periodic cycles of proliferation, differentiation, and involution. Many studies have reported that miRNAs, which are small, non-coding RNAs, influence a variety of biological processes during posttranscriptional regulation. Here, we found that one miRNA, miR-200a, was relatively highly expressed in epithelial cell-rich organs such as mammary glands, lung, and kidney in mice. In mammary glands, miR-200a expression increased during mid-pregnancy through lactation; its expression was stimulated by lactogenic hormone treatment of mammary epithelial cells. Lactogenic hormone also induced the expression of milk protein ß-casein mRNA (a marker of cell differentiation and E-cadherin mRNA (a marker of epithelial cells. However, knockdown of miR-200a prevented increases in ß-casein and E-cadherin mRNA expression. Protein analysis revealed that E-cadherin signal was decreased and ZEB1 (a marker of EMT was increased following miR-200a knockdown. Finally, in a three-dimensional culture system modeling lumen-containing mammary ducts, miR-200a knockdown decreased the cavity formation rate and suppressed claudin-3 and par-6b expression, indicating reduced epithelial cell polarity. These observations suggest that miR-200a is important for maintaining the epithelial cell phenotype, which contributes to lactogenic hormone induction of cellular differentiation in mammary glands.

  14. Role of Porphyromonas gingivalis SerB in Gingival Epithelial Cell Cytoskeletal Remodeling and Cytokine Production▿

    Science.gov (United States)

    Hasegawa, Yoshiaki; Tribble, Gena D.; Baker, Henry V.; Mans, Jeffrey J.; Handfield, Martin; Lamont, Richard J.

    2008-01-01

    The SerB protein of Porphyromonas gingivalis is a HAD family serine phosphatase that plays a critical role in entry and survival of the organism in gingival epithelial cells. SerB is secreted by P. gingivalis upon contact with epithelial cells. Here it is shown by microarray analysis that SerB impacts the transcriptional profile of gingival epithelial cells, with pathways involving the actin cytoskeleton and cytokine production among those significantly overpopulated with differentially regulated genes. Consistent with the transcriptional profile, a SerB mutant of P. gingivalis exhibited defective remodeling of actin in epithelial cells. Interaction between gingival epithelial cells and isolated SerB protein resulted in actin rearrangement and an increase in the F/G actin ratio. SerB protein was also required for P. gingivalis to antagonize interleukin-8 accumulation following stimulation of epithelial cells with Fusobacterium nucleatum. SerB is thus capable of modulating host cell signal transduction that impacts the actin cytoskeleton and cytokine production. PMID:18391005

  15. Role of Porphyromonas gingivalis SerB in gingival epithelial cell cytoskeletal remodeling and cytokine production.

    Science.gov (United States)

    Hasegawa, Yoshiaki; Tribble, Gena D; Baker, Henry V; Mans, Jeffrey J; Handfield, Martin; Lamont, Richard J

    2008-06-01

    The SerB protein of Porphyromonas gingivalis is a HAD family serine phosphatase that plays a critical role in entry and survival of the organism in gingival epithelial cells. SerB is secreted by P. gingivalis upon contact with epithelial cells. Here it is shown by microarray analysis that SerB impacts the transcriptional profile of gingival epithelial cells, with pathways involving the actin cytoskeleton and cytokine production among those significantly overpopulated with differentially regulated genes. Consistent with the transcriptional profile, a SerB mutant of P. gingivalis exhibited defective remodeling of actin in epithelial cells. Interaction between gingival epithelial cells and isolated SerB protein resulted in actin rearrangement and an increase in the F/G actin ratio. SerB protein was also required for P. gingivalis to antagonize interleukin-8 accumulation following stimulation of epithelial cells with Fusobacterium nucleatum. SerB is thus capable of modulating host cell signal transduction that impacts the actin cytoskeleton and cytokine production.

  16. Psychological Stress-Derived Prolactin Modulates Occludin Expression in Vaginal Epithelial Cells to Compromise Barrier Function

    Directory of Open Access Journals (Sweden)

    Xueyan Li

    2015-08-01

    Full Text Available Background/Aims: The causative factors of the vaginitis are not fully understood yet. Epithelial barrier dysfunction plays a critical role in the pathogenesis of vaginitis. This study aims to investigate the role of prolactin (PRL in the causing the vaginal epithelial barrier dysfunction. Methods: Adult rats were treated with water-avoid-stress. The serum levels of PRL were determined by ELISA. T84 cell (T84 cells; a vaginal epithelial cell line monolayers were prepared to be used assessing the epithelial barrier functions. The expression of occludin in T84 cells was assessed by Chromatin immunoprecipitation assay, methylation specifIc PCR, real time quantitative RT-PCR and Western blotting. Results: The results showed that psychological stress markedly increased the serum levels of PRL in the rat vaginal epithelia. Exposure of T84 cells to PRL in the culture markedly increased the phosphorylation of STAT3 and suppressed the expression of occludin in the cells; the transepithelial electric resistance was decreased and the permeability to a macromolecular tracer was increased in the T84 monolayers, which was mimicked by blocking STAT3, or abolished by over expression of occludin in the epithelial cells. Conclusions: Psychological stress-derived PRL induces vaginal epithelial barrier dysfunction by inhibiting the expression of occludin.

  17. Chlamydia trachomatis Infection of Endocervical Epithelial Cells Enhances Early HIV Transmission Events.

    Science.gov (United States)

    Buckner, Lyndsey R; Amedee, Angela M; Albritton, Hannah L; Kozlowski, Pamela A; Lacour, Nedra; McGowin, Chris L; Schust, Danny J; Quayle, Alison J

    2016-01-01

    Chlamydia trachomatis causes a predominantly asymptomatic, but generally inflammatory, genital infection that is associated with an increased risk for HIV acquisition. Endocervical epithelial cells provide the major niche for this obligate intracellular bacterium in women, and the endocervix is also a tissue in which HIV transmission can occur. The mechanism by which CT infection enhances HIV susceptibility at this site, however, is not well understood. Utilizing the A2EN immortalized endocervical epithelial cell line grown on cell culture inserts, we evaluated the direct role that CT-infected epithelial cells play in facilitating HIV transmission events. We determined that CT infection significantly enhanced the apical-to-basolateral migration of cell-associated, but not cell-free, HIVBaL, a CCR5-tropic strain of virus, across the endocervical epithelial barrier. We also established that basolateral supernatants from CT-infected A2EN cells significantly enhanced HIV replication in peripheral mononuclear cells and a CCR5+ T cell line. These results suggest that CT infection of endocervical epithelial cells could facilitate both HIV crossing the mucosal barrier and subsequent infection or replication in underlying target cells. Our studies provide a mechanism by which this common STI could potentially promote the establishment of founder virus populations and the maintenance of local HIV reservoirs in the endocervix. Development of an HIV/STI co-infection model also provides a tool to further explore the role of other sexually transmitted infections in enhancing HIV acquisition.

  18. Alterations in Helicobacter pylori triggered by contact with gastric epithelial cells

    Directory of Open Access Journals (Sweden)

    Elizabeth M. Johnson

    2012-02-01

    Full Text Available Helicobacter pylori lives within the mucus layer of the human stomach, in close proximity to gastric epithelial cells. While a great deal is known about the effects of H. pylori on human cells and the specific bacterial products that mediate these effects, relatively little work has been done to investigate alterations in H. pylori that may be triggered by bacterial contact with human cells. In this review, we discuss the spectrum of changes in bacterial physiology and morphology that occur when H. pylori is in contact with gastric epithelial cells. Several studies have reported that cell contact causes alterations in H. pylori gene transcription. In addition, H. pylori contact with gastric epithelial cells promotes the formation of pilus-like structures at the bacteria-host cell interface. The formation of these structures requires multiple genes in the cag pathogenicity island, and these structures are proposed to have an important role in the type IV secretion system-dependent process through which CagA enters host cells. Finally, H. pylori contact with epithelial cells can promote bacterial replication and the formation of microcolonies, phenomena that are facilitated by the acquisition of iron and other nutrients from infected cells. In summary, the gastric epithelial cell surface represents an important niche for H. pylori, and upon entry into this niche, the bacteria alter their behavior in a manner that optimizes bacterial proliferation and persistent colonization of the host.

  19. Differentiation-Dependent KLF4 Expression Promotes Lytic Epstein-Barr Virus Infection in Epithelial Cells.

    Directory of Open Access Journals (Sweden)

    Dhananjay M Nawandar

    2015-10-01

    Full Text Available Epstein-Barr virus (EBV is a human herpesvirus associated with B-cell and epithelial cell malignancies. EBV lytically infects normal differentiated oral epithelial cells, where it causes a tongue lesion known as oral hairy leukoplakia (OHL in immunosuppressed patients. However, the cellular mechanism(s that enable EBV to establish exclusively lytic infection in normal differentiated oral epithelial cells are not currently understood. Here we show that a cellular transcription factor known to promote epithelial cell differentiation, KLF4, induces differentiation-dependent lytic EBV infection by binding to and activating the two EBV immediate-early gene (BZLF1 and BRLF1 promoters. We demonstrate that latently EBV-infected, telomerase-immortalized normal oral keratinocyte (NOKs cells undergo lytic viral reactivation confined to the more differentiated cell layers in organotypic raft culture. Furthermore, we show that endogenous KLF4 expression is required for efficient lytic viral reactivation in response to phorbol ester and sodium butyrate treatment in several different EBV-infected epithelial cell lines, and that the combination of KLF4 and another differentiation-dependent cellular transcription factor, BLIMP1, is highly synergistic for inducing lytic EBV infection. We confirm that both KLF4 and BLIMP1 are expressed in differentiated, but not undifferentiated, epithelial cells in normal tongue tissue, and show that KLF4 and BLIMP1 are both expressed in a patient-derived OHL lesion. In contrast, KLF4 protein is not detectably expressed in B cells, where EBV normally enters latent infection, although KLF4 over-expression is sufficient to induce lytic EBV reactivation in Burkitt lymphoma cells. Thus, KLF4, together with BLIMP1, plays a critical role in mediating lytic EBV reactivation in epithelial cells.

  20. Lysosomal Changes in Renal Proximal Tubular Epithelial Cells of Male Sprague Dawley Rats Following Decalin Exposure

    Science.gov (United States)

    1990-01-01

    decalin-treated animal. Note large, pale, rcd-staining lysosome (-). An exfoliated epithelial cell can iu- seen in the tubular lumen containing large...photomicrograph contains an exfoliated epithelial cell (-) with enlarged, intact lysosomes. The tubule on the left half of the photomicrograph contains an...metabolism of proteins. In: Cytology , GH Bourne and JF Danielli (eds). Academ- The Kidney: Physiology and Pathophysiology, DW ic Press, NY, pp. 251-300. - ~- i :- d .L n .- 2

  1. Thymopoiesis in mice depends on a Foxn1-positive thymic epithelial cell lineage

    OpenAIRE

    Corbeaux, Tatiana; Hess, Isabell; Swann, Jeremy B.; Kanzler, Benoît; Haas-Assenbaum, Annette; Boehm, Thomas

    2010-01-01

    The thymus is essential for T-cell development. Here, we focus on the role of the transcription factor Foxn1 in the development and function of thymic epithelial cells (TECs) of the mouse. TECs are of endodermal origin; they initially express Foxn1 and give rise to orthotopic (thoracic) and additional (cervical) thymi. Using Foxn1-directed cytoablation, we show that during embryogenesis, cervical thymi develop a few days after the thoracic lobes, and that bipotent epithelial progenitors of co...

  2. Connective Tissue Growth Factor Promotes Pulmonary Epithelial Cell Senescence and Is Associated with COPD Severity.

    Science.gov (United States)

    Jang, Jun-Ho; Chand, Hitendra S; Bruse, Shannon; Doyle-Eisele, Melanie; Royer, Christopher; McDonald, Jacob; Qualls, Clifford; Klingelhutz, Aloysius J; Lin, Yong; Mallampalli, Rama; Tesfaigzi, Yohannes; Nyunoya, Toru

    2017-04-01

    The purpose of this study was to determine whether expression of connective tissue growth factor (CTGF) protein in chronic obstructive pulmonary disease (COPD) is consistent in humans and animal models of COPD and to investigate the role of this protein in lung epithelial cells. CTGF in lung epithelial cells of ex-smokers with COPD was compared with ex-smokers without COPD by immunofluorescence. A total of twenty C57Bl/6 mice and sixteen non-human primates (NHPs) were exposed to cigarette smoke (CS) for 4 weeks. Ten mice of these CS-exposed mice and eight of the CS-exposed NHPs were infected with H3N2 influenza A virus (IAV), while the remaining ten mice and eight NHPs were mock-infected with vehicle as control. Both mRNA and protein expression of CTGF in lung epithelial cells of mice and NHPs were determined. The effects of CTGF overexpression on cell proliferation, p16 protein, and senescence-associated β-galactosidase (SA-β-gal) activity were examined in cultured human bronchial epithelial cells (HBECs). In humans, CTGF expression increased with increasing COPD severity. We found that protein expression of CTGF was upregulated in lung epithelial cells in both mice and NHPs exposed to CS and infected with IAV compared to those exposed to CS only. When overexpressed in HBECs, CTGF accelerated cellular senescence accompanied by p16 accumulation. Both CTGF and p16 protein expression in lung epithelia are positively associated with the severity of COPD in ex-smokers. These findings show that CTGF is consistently expressed in epithelial cells of COPD lungs. By accelerating lung epithelial senescence, CTGF may block regeneration relative to epithelial cell loss and lead to emphysema.

  3. Btbd7 is essential for region-specific epithelial cell dynamics and branching morphogenesis in vivo

    DEFF Research Database (Denmark)

    Daley, William P; Matsumoto, Kazue; Doyle, Andrew D

    2017-01-01

    Branching morphogenesis of developing organs requires coordinated but poorly understood changes in epithelial cell-cell adhesion and cell motility. We report that Btbd7 is a crucial regulator of branching morphogenesis in vivo. Btbd7 levels are elevated in peripheral cells of branching epithelial...... end buds, where it enhances cell motility and cell-cell adhesion dynamics. Genetic ablation of Btbd7 in mice disrupts branching morphogenesis of salivary gland, lung, and kidney. Btbd7 knockout results in more tightly packed outer bud cells, which display stronger E-cadherin localization, reduced cell...... motility, and decreased dynamics of transient cell separations associated with cleft formation; inner bud cells remain unaffected. Mechanistic analyses using in vitro MDCK cells to mimic outer bud cell behavior establish that Btbd7 promotes loss of E-cadherin from cell-cell adhesions with enhanced...

  4. Subcultured odontogenic epithelial cells in combination with dental mesenchymal cells produce enamel-dentin-like complex structures.

    Science.gov (United States)

    Honda, M J; Shinohara, Y; Hata, K I; Ueda, M

    2007-01-01

    We showed in a previous study that odontogenic epithelial cells can be selectively cultured from the enamel organ in serum-free medium and expanded using feeder layers of 3T3-J2 cells. The subcultured odontogenic epithelial cells retain the capacity for ameloblast-related gene expression, as shown by semiquantitative RT-PCR. The purpose of the present study was to evaluate the potential of subcultured odontogenic epithelial cells to form tooth structures in cell-polymer constructs maintained in vivo. Enamel organs from 6-month-old porcine third molars were dissociated into single odontogenic epithelial cells and subcultured on feeder layers of 3T3-J2 cells. Amelogenin expression was detected in the subcultured odontogenic epithelial cells by immunostaining and Western blotting. The subcultured odontogenic epithelial cells were seeded onto collagen sponge scaffolds in combination with fresh dental mesenchymal cells, and transplanted into athymic rats. After 4 weeks, enamel-dentin-like complex structures were present in the implanted constructs. These results show that our culture system produced differentiating ameloblast-like cells that were able to secrete amelogenin proteins and form enamel-like tissues in vivo. This application of the subculturing technique provides a foundation for further tooth-tissue engineering and for improving our understanding of ameloblast biology.

  5. Cross-talk between intestinal epithelial cells and immune cells in inflammatory bowel disease.

    Science.gov (United States)

    Al-Ghadban, Sara; Kaissi, Samira; Homaidan, Fadia R; Naim, Hassan Y; El-Sabban, Marwan E

    2016-07-15

    Inflammatory bowel disease (IBD) involves functional impairment of intestinal epithelial cells (IECs), concomitant with the infiltration of the lamina propria by inflammatory cells. We explored the reciprocal paracrine and direct interaction between human IECs and macrophages (MΦ) in a co-culture system that mimics some aspects of IBD. We investigated the expression of intercellular junctional proteins in cultured IECs under inflammatory conditions and in tissues from IBD patients. IECs establish functional gap junctions with IECs and MΦ, respectively. Connexin (Cx26) and Cx43 expression in cultured IECs is augmented under inflammatory conditions; while, Cx43-associated junctional complexes partners, E-cadherin, ZO-1, and β-catenin expression is decreased. The expression of Cx26 and Cx43 in IBD tissues is redistributed to the basal membrane of IEC, which is associated with decrease in junctional complex proteins' expression, collagen type IV expression and infiltration of MΦ. These data support the notion that the combination of paracrine and hetero-cellular communication between IECs and MΦs may regulate epithelial cell function through the establishment of junctional complexes between inflammatory cells and IECs, which ultimately contribute to the dys-regulation of intestinal epithelial barrier.

  6. Cross-talk between intestinal epithelial cells and immune cells in inflammatory bowel disease

    Science.gov (United States)

    Al-Ghadban, Sara; Kaissi, Samira; Homaidan, Fadia R.; Naim, Hassan Y.; El-Sabban, Marwan E.

    2016-01-01

    Inflammatory bowel disease (IBD) involves functional impairment of intestinal epithelial cells (IECs), concomitant with the infiltration of the lamina propria by inflammatory cells. We explored the reciprocal paracrine and direct interaction between human IECs and macrophages (MΦ) in a co-culture system that mimics some aspects of IBD. We investigated the expression of intercellular junctional proteins in cultured IECs under inflammatory conditions and in tissues from IBD patients. IECs establish functional gap junctions with IECs and MΦ, respectively. Connexin (Cx26) and Cx43 expression in cultured IECs is augmented under inflammatory conditions; while, Cx43-associated junctional complexes partners, E-cadherin, ZO-1, and β-catenin expression is decreased. The expression of Cx26 and Cx43 in IBD tissues is redistributed to the basal membrane of IEC, which is associated with decrease in junctional complex proteins’ expression, collagen type IV expression and infiltration of MΦ. These data support the notion that the combination of paracrine and hetero-cellular communication between IECs and MΦs may regulate epithelial cell function through the establishment of junctional complexes between inflammatory cells and IECs, which ultimately contribute to the dys-regulation of intestinal epithelial barrier. PMID:27417573

  7. Human gastric epithelial cells contribute to gastric immune regulation by providing retinoic acid to dendritic cells.

    Science.gov (United States)

    Bimczok, D; Kao, J Y; Zhang, M; Cochrun, S; Mannon, P; Peter, S; Wilcox, C M; Mönkemüller, K E; Harris, P R; Grams, J M; Stahl, R D; Smith, P D; Smythies, L E

    2015-05-01

    Despite the high prevalence of chronic gastritis caused by Helicobacter pylori, the gastric mucosa has received little investigative attention as a unique immune environment. Here, we analyzed whether retinoic acid (RA), an important homeostatic factor in the small intestinal mucosa, also contributes to gastric immune regulation. We report that human gastric tissue contains high levels of the RA precursor molecule retinol (ROL), and that gastric epithelial cells express both RA biosynthesis genes and RA response genes, indicative of active RA biosynthesis. Moreover, primary gastric epithelial cells cultured in the presence of ROL synthesized RA in vitro and induced RA biosynthesis in co-cultured monocytes through an RA-dependent mechanism, suggesting that gastric epithelial cells may also confer the ability to generate RA on gastric dendritic cells (DCs). Indeed, DCs purified from gastric mucosa had similar levels of aldehyde dehydrogenase activity and RA biosynthesis gene expression as small intestinal DCs, although gastric DCs lacked CD103. In H. pylori-infected gastric mucosa, gastric RA biosynthesis gene expression was severely disrupted, which may lead to reduced RA signaling and thus contribute to disease progression. Collectively, our results support a critical role for RA in human gastric immune regulation.

  8. Multipotent epithelial cells in the process of regeneration and asexual reproduction in colonial tunicates.

    Science.gov (United States)

    Kawamura, Kazuo; Sugino, Yasuo; Sunanaga, Takeshi; Fujiwara, Shigeki

    2008-01-01

    The cellular and molecular features of multipotent epithelial cells during regeneration and asexual reproduction in colonial tunicates are described in the present study. The epicardium has been regarded as the endodermal tissue-forming epithelium in the order Enterogona, because only body fragments having the epicardium exhibit the regenerative potential. Epicardial cells in Polycitor proliferus have two peculiar features; they always accompany coelomic undifferentiated cells, and they contain various kinds of organelles in the cytoplasm. During strobilation a large amount of organelles are discarded in the lumen, and then, each tissue-forming cell takes an undifferentiated configuration. Septum cells in the stolon are also multipotent in Enterogona. Free cells with a similar configuration to the septum inhabit the hemocoel. They may provide a pool for epithelial septum cells. At the distal tip of the stolon, septum cells are columnar in shape and apparently undifferentiated. They are the precursor of the stolonial bud. In Pleurogona, the atrial epithelium of endodermal origin is multipotent. In Polyandrocarpa misakiensis, it consists of pigmented squamous cells. The cells have ultrastructurally fine granules in the cytoplasm. During budding, coelomic cells with similar morphology become associated with the atrial epithelium. Then, cells of organ placodes undergo dedifferentiation, enter a cell division cycle, and commence morphogenesis. Retinoic acid-related molecules are involved in this dedifferentiation process of multipotent cells. We conclude that in colonial tunicates two systems support the flexibility of tissue remodeling during regeneration and asexual reproduction; dedifferentiation of epithelial cells and epithelial transformation of coelomic free cells.

  9. Plasticity between Epithelial and Mesenchymal States Unlinks EMT from Metastasis-Enhancing Stem Cell Capacity

    NARCIS (Netherlands)

    Beerling, Evelyne; Seinstra, Daniëlle; de Wit, Elzo; Kester, Lennart; van der Velden, Daphne; Maynard, Carrie; Schäfer, Ronny; van Diest, Paul; Voest, Emile; van Oudenaarden, Alexander; Vrisekoop, Nienke; van Rheenen, Jacco

    2016-01-01

    Forced overexpression and/or downregulation of proteins regulating epithelial-to-mesenchymal transition (EMT) has been reported to alter metastasis by changing migration and stem cell capacity of tumor cells. However, these manipulations artificially keep cells in fixed states, while in vivo cells m

  10. Polarity, cell division, and out-of-equilibrium dynamics control the growth of epithelial structures

    Science.gov (United States)

    Cerruti, Benedetta; Puliafito, Alberto; Shewan, Annette M.; Yu, Wei; Combes, Alexander N.; Little, Melissa H.; Chianale, Federica; Primo, Luca; Serini, Guido; Mostov, Keith E.; Celani, Antonio

    2013-01-01

    The growth of a well-formed epithelial structure is governed by mechanical constraints, cellular apico-basal polarity, and spatially controlled cell division. Here we compared the predictions of a mathematical model of epithelial growth with the morphological analysis of 3D epithelial structures. In both in vitro cyst models and in developing epithelial structures in vivo, epithelial growth could take place close to or far from mechanical equilibrium, and was determined by the hierarchy of time-scales of cell division, cell–cell rearrangements, and lumen dynamics. Equilibrium properties could be inferred by the analysis of cell–cell contact topologies, and the nonequilibrium phenotype was altered by inhibiting ROCK activity. The occurrence of an aberrant multilumen phenotype was linked to fast nonequilibrium growth, even when geometric control of cell division was correctly enforced. We predicted and verified experimentally that slowing down cell division partially rescued a multilumen phenotype induced by altered polarity. These results improve our understanding of the development of epithelial organs and, ultimately, of carcinogenesis. PMID:24145168

  11. Mammary epithelial cell: Influence of extracellular matrix composition and organization during development and tumorigenesis

    Science.gov (United States)

    Kass, Laura; Erler, Janine T.; Dembo, Micah; Weaver, Valerie M.

    2009-01-01

    Stromal–epithelial interactions regulate mammary gland development and are critical for the maintenance of tissue homeostasis. The extracellular matrix, which is a proteinaceous component of the stroma, regulates mammary epithelial growth, survival, migration and differentiation through a repertoire of transmembrane receptors, of which integrins are the best characterized. Integrins modulate cell fate by reciprocally transducing biochemical and biophysical cues between the cell and the extracellular matrix, facilitating processes such as embryonic branching morphogenesis and lactation in the mammary gland. During breast development and cancer progression, the extracellular matrix is dynamically altered such that its composition, turnover, processing and orientation change dramatically. These modifications influence mammary epithelial cell shape, and modulate growth factor and hormonal responses to regulate processes including branching morphogenesis and alveolar differentiation. Malignant transformation of the breast is also associated with significant matrix remodeling and a progressive stiffening of the stroma that can enhance mammary epithelial cell growth, perturb breast tissue organization, and promote cell invasion and survival. In this review, we discuss the role of stromal–epithelial interactions in normal and malignant mammary epithelial cell behavior. We specifically focus on how dynamic modulation of the biochemical and biophysical properties of the extracellular matrix elicit a dialogue with the mammary epithelium through transmembrane integrin receptors to influence tissue morphogenesis, homeostasis and malignant transformation. PMID:17719831

  12. SPLUNC1 regulation in airway epithelial cells: role of toll-like receptor 2 signaling

    Directory of Open Access Journals (Sweden)

    Smith Sean

    2010-11-01

    Full Text Available Abstract Background Respiratory infections including Mycoplasma pneumoniae (Mp contribute to various chronic lung diseases. We have shown that mouse short palate, lung, and nasal epithelium clone 1 (SPLUNC1 protein was able to inhibit Mp growth. Further, airway epithelial cells increased SPLUNC1 expression upon Mp infection. However, the mechanisms underlying SPLUNC1 regulation remain unknown. In the current study, we investigated if SPLUNC1 production following Mp infection is regulated through Toll-like receptor 2 (TLR2 signaling. Methods Airway epithelial cell cultures were utilized to reveal the contribution of TLR2 signaling including NF-κB to SPLUNC1 production upon bacterial infection and TLR2 agonist stimulation. Results Mp and TLR2 agonist Pam3CSK4 increased SPLUNC1 expression in tracheal epithelial cells from wild type, but not TLR2-/- BALB/c mice. RNA interference (short-hairpin RNA of TLR2 in normal human bronchial epithelial cells under air-liquid interface cultures significantly reduced SPLUNC1 levels in Mp-infected or Pam3CSK4-treated cells. Inhibition and activation of NF-κB pathway decreased and increased SPLUNC1 production in airway epithelial cells, respectively. Conclusions Our data for the first time suggest that airway epithelial TLR2 signaling is pivotal in mycoplasma-induced SPLUNC1 production, thus improving our understanding of the aberrant SPLUNC1 expression in airways of patients suffering from chronic lung diseases with bacterial infections.

  13. Dickkopf proteins influence lung epithelial cell proliferation in idiopathic pulmonary fibrosis.

    Science.gov (United States)

    Pfaff, E-M; Becker, S; Günther, A; Königshoff, M

    2011-01-01

    Idiopathic pulmonary fibrosis (IPF) is a fatal interstitial lung disease with unknown pathogenesis. The WNT/β-catenin pathway has recently been reported to be operative in epithelial cells in IPF. Dickkopf (DKK) proteins are known to regulate WNT signalling via interaction with Kremen (KRM) receptors, yet their expression and role in the adult lung and in IPF has not been addressed. We analysed the expression, localisation and function of DKK and KRM proteins in IPF lungs using Western blotting, quantitative RT-PCR, immunohistochemistry, ELISA and functional in vitro studies. Enhanced expression of DKK1 and DKK4 and KRM1 was detected in lung homogenates of IPF patients compared with transplant donors. Immunohistochemistry revealed that DKK1 was predominantly localised in basal bronchial epithelial cells. Furthermore, prominent expression of all proteins was observed in hyperplastic alveolar epithelial cells in IPF. Quantitative measurement of DKK1 revealed enhanced protein expression in the bronchoalveolar lumen of IPF patients. Finally, functional studies using human bronchial and alveolar epithelial cell lines demonstrated that WNT-induced epithelial cell proliferation is regulated by DKK1 in a dose-dependent fashion. In summary, DKK proteins are expressed in the lung epithelium in IPF. DKK proteins influence epithelial cell proliferation and may, therefore, be suitable therapeutic targets for IPF.

  14. Loss of γ-cytoplasmic actin triggers myofibroblast transition of human epithelial cells.

    Science.gov (United States)

    Lechuga, Susana; Baranwal, Somesh; Li, Chao; Naydenov, Nayden G; Kuemmerle, John F; Dugina, Vera; Chaponnier, Christine; Ivanov, Andrei I

    2014-10-15

    Transdifferentiation of epithelial cells into mesenchymal cells and myofibroblasts plays an important role in tumor progression and tissue fibrosis. Such epithelial plasticity is accompanied by dramatic reorganizations of the actin cytoskeleton, although mechanisms underlying cytoskeletal effects on epithelial transdifferentiation remain poorly understood. In the present study, we observed that selective siRNA-mediated knockdown of γ-cytoplasmic actin (γ-CYA), but not β-cytoplasmic actin, induced epithelial-to-myofibroblast transition (EMyT) of different epithelial cells. The EMyT manifested by increased expression of α-smooth muscle actin and other contractile proteins, along with inhibition of genes responsible for cell proliferation. Induction of EMyT in γ-CYA-depleted cells depended on activation of serum response factor and its cofactors, myocardial-related transcriptional factors A and B. Loss of γ-CYA stimulated formin-mediated actin polymerization and activation of Rho GTPase, which appear to be essential for EMyT induction. Our findings demonstrate a previously unanticipated, unique role of γ-CYA in regulating epithelial phenotype and suppression of EMyT that may be essential for cell differentiation and tissue fibrosis.

  15. A Comparative Study on Rat Intestinal Epithelial Cells and Resident Gut Bacteria (ii) Effect of Arsenite

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    In order to use facultative gut bacteria as an alternate to animals for the initial gastrointestinal toxicity screening of heavy metals, a comparative study on rat intestinal epithelial cells and resident gut bacteria was undertaken.Methods in vitro growth rate of four gut bacteria, dehydrogenase (DHA) and esterase (EA) activity test, intestinal epithelial and bacterial cell membrane enzymes and in situ effect of arsenite were analysed. Results Growth profile of mixed resident population of gut bacteria and pure isolates of Escherichia coli, Pseudomonas sp., Lactobacillus sp., and Staphylococcus sp.revealed an arsenite (2-20 ppm) concentration-dependent inhibition. The viability pattern of epithelial cells also showed similar changes. DHA and EA tests revealed significant inhibition (40%-72%) with arsenite exposure of 5 and 10 ppm in isolated gut bacteria and epithelial cells. Decrease in membrane alkaline phosphatase and Ca2+-Mg2+-ATPase activities was in the range of 33%-55% in four bacteria at the arsenite exposure of 10 ppm, whereas it was 60%-65% in intestinal epithelial villus cells. in situ incubation of arsenite using intestinal loops also showed more or less similar changes in membrane enzymes of resident gut bacterial population and epithelial cells. Conclusion The results indicate that facultative gut bacteria can be used as suitable in vitro model for the preliminary screening of arsenical gastrointestinal cytotoxic effects.

  16. Culturing of Human Nasal Epithelial Cells at the Air Liquid Interface

    Science.gov (United States)

    Müller, Loretta; Brighton, Luisa E.; Carson, Johnny L.; Fischer, William A.; Jaspers, Ilona

    2013-01-01

    In vitro models using human primary epithelial cells are essential in understanding key functions of the respiratory epithelium in the context of microbial infections or inhaled agents. Direct comparisons of cells obtained from diseased populations allow us to characterize different phenotypes and dissect the underlying mechanisms mediating changes in epithelial cell function. Culturing epithelial cells from the human tracheobronchial region has been well documented, but is limited by the availability of human lung tissue or invasiveness associated with obtaining the bronchial brushes biopsies. Nasal epithelial cells are obtained through much less invasive superficial nasal scrape biopsies and subjects can be biopsied multiple times with no significant side effects. Additionally, the nose is the entry point to the respiratory system and therefore one of the first sites to be exposed to any kind of air-borne stressor, such as microbial agents, pollutants, or allergens. Briefly, nasal epithelial cells obtained from human volunteers are expanded on coated tissue culture plates, and then transferred onto cell culture inserts. Upon reaching confluency, cells continue to be cultured at the air-liquid interface (ALI), for several weeks, which creates more physiologically relevant conditions. The ALI culture condition uses defined media leading to a differentiated epithelium that exhibits morphological and functional characteristics similar to the human nasal epithelium, with both ciliated and mucus producing cells. Tissue culture inserts with differentiated nasal epithelial cells can be manipulated in a variety of ways depending on the research questions (treatment with pharmacological agents, transduction with lentiviral vectors, exposure to gases, or infection with microbial agents) and analyzed for numerous different endpoints ranging from cellular and molecular pathways, functional changes, morphology, etc. In vitro models of differentiated human nasal epithelial

  17. Culturing of human nasal epithelial cells at the air liquid interface.

    Science.gov (United States)

    Müller, Loretta; Brighton, Luisa E; Carson, Johnny L; Fischer, William A; Jaspers, Ilona

    2013-10-08

    In vitro models using human primary epithelial cells are essential in understanding key functions of the respiratory epithelium in the context of microbial infections or inhaled agents. Direct comparisons of cells obtained from diseased populations allow us to characterize different phenotypes and dissect the underlying mechanisms mediating changes in epithelial cell function. Culturing epithelial cells from the human tracheobronchial region has been well documented, but is limited by the availability of human lung tissue or invasiveness associated with obtaining the bronchial brushes biopsies. Nasal epithelial cells are obtained through much less invasive superficial nasal scrape biopsies and subjects can be biopsied multiple times with no significant side effects. Additionally, the nose is the entry point to the respiratory system and therefore one of the first sites to be exposed to any kind of air-borne stressor, such as microbial agents, pollutants, or allergens. Briefly, nasal epithelial cells obtained from human volunteers are expanded on coated tissue culture plates, and then transferred onto cell culture inserts. Upon reaching confluency, cells continue to be cultured at the air-liquid interface (ALI), for several weeks, which creates more physiologically relevant conditions. The ALI culture condition uses defined media leading to a differentiated epithelium that exhibits morphological and functional characteristics similar to the human nasal epithelium, with both ciliated and mucus producing cells. Tissue culture inserts with differentiated nasal epithelial cells can be manipulated in a variety of ways depending on the research questions (treatment with pharmacological agents, transduction with lentiviral vectors, exposure to gases, or infection with microbial agents) and analyzed for numerous different endpoints ranging from cellular and molecular pathways, functional changes, morphology, etc. In vitro models of differentiated human nasal epithelial

  18. Wound repair and anti-oxidative capacity is regulated by ITGB4 in airway epithelial cells.

    Science.gov (United States)

    Liu, Chi; Liu, Hui-jun; Xiang, Yang; Tan, Yu-rong; Zhu, Xiao-lin; Qin, Xiao-qun

    2010-08-01

    Integrin beta 4 (ITGB4) is a structural adhesion molecule which engages in maintaining the integrity of airway epithelial cells. Its specific cytomembrane structural feature strongly indicates that ITGB4 may engage in many signaling pathways and physiologic processes. However, in addition to adhesion, the specific biologic significance of ITGB4 in airway epithelial cells is almost unknown. In this article, we investigated the expression and functional properties of ITGB4 in airway epithelial cells in vivo and in vitro. Human bronchial epithelial cell line (16HBE14O-cells) and primary rat tracheal epithelial cells (RTE cells) were used to determine ITGB4 expression under ozone tress or mechanical damage, respectively. An ovalbumin (OVA)-challenged asthma model was used to investigate ITGB4 expression after antigen exposure in vivo. In addition, an ITGB4 overexpression vector and ITGB4 silence virus vector were constructed and transfected into RTE cells. Then, wound repair ability and anti-oxidation capacity was evaluated. Our results demonstrated that, on the edge of mechanically wounded cell areas, ITGB4 expression was increased after mechanical injury. After ozone stress, upregulation expression of ITGB4 was also detected. In the OVA-challenged asthma model, ITGB4 expression was decreased on airway epithelial cells accompanying with structural disruption and damage of anti-oxidation capacity. Besides, our study revealed that upregulation of ITGB4 promotes wound repair ability and anti-oxidative ability, while such abilities were blocked when ITGB4 was silenced. Taken together, these results showed that ITGB4 was a new interesting molecule involved in the regulation of wound repair and anti-oxidation processes for airway epithelial cells.

  19. Epithelial tricellular junctions act as interphase cell shape sensors to orient mitosis.

    Science.gov (United States)

    Bosveld, Floris; Markova, Olga; Guirao, Boris; Martin, Charlotte; Wang, Zhimin; Pierre, Anaëlle; Balakireva, Maria; Gaugue, Isabelle; Ainslie, Anna; Christophorou, Nicolas; Lubensky, David K; Minc, Nicolas; Bellaïche, Yohanns

    2016-02-25

    The orientation of cell division along the long axis of the interphase cell--the century-old Hertwig's rule--has profound roles in tissue proliferation, morphogenesis, architecture and mechanics. In epithelial tissues, the shape of the interphase cell is influenced by cell adhesion, mechanical stress, neighbour topology, and planar polarity pathways. At mitosis, epithelial cells usually adopt a rounded shape to ensure faithful chromosome segregation and to promote morphogenesis. The mechanisms underlying interphase cell shape sensing in tissues are therefore unknown. Here we show that in Drosophila epithelia, tricellular junctions (TCJs) localize force generators, pulling on astral microtubules and orienting cell division via the Dynein-associated protein Mud independently of the classical Pins/Gαi pathway. Moreover, as cells round up during mitosis, TCJs serve as spatial landmarks, encoding information about interphase cell shape anisotropy to orient division in the rounded mitotic cell. Finally, experimental and simulation data show that shape and mechanical strain sensing by the TCJs emerge from a general geometric property of TCJ distributions in epithelial tissues. Thus, in addition to their function as epithelial barrier structures, TCJs serve as polarity cues promoting geometry and mechanical sensing in epithelial tissues.

  20. The keratin-binding protein Albatross regulates polarization of epithelial cells.

    Science.gov (United States)

    Sugimoto, Masahiko; Inoko, Akihito; Shiromizu, Takashi; Nakayama, Masanori; Zou, Peng; Yonemura, Shigenobu; Hayashi, Yuko; Izawa, Ichiro; Sasoh, Mikio; Uji, Yukitaka; Kaibuchi, Kozo; Kiyono, Tohru; Inagaki, Masaki

    2008-10-06

    The keratin intermediate filament network is abundant in epithelial cells, but its function in the establishment and maintenance of cell polarity is unclear. Here, we show that Albatross complexes with Par3 to regulate formation of the apical junctional complex (AJC) and maintain lateral membrane identity. In nonpolarized epithelial cells, Albatross localizes with keratin filaments, whereas in polarized epithelial cells, Albatross is primarily localized in the vicinity of the AJC. Knockdown of Albatross in polarized cells causes a disappearance of key components of the AJC at cell-cell borders and keratin filament reorganization. Lateral proteins E-cadherin and desmoglein 2 were mislocalized even on the apical side. Although Albatross promotes localization of Par3 to the AJC, Par3 and ezrin are still retained at the apical surface in Albatross knockdown cells, which retain intact microvilli. Analysis of keratin-deficient epithelial cells revealed that keratins are required to stabilize the Albatross protein, thus promoting the formation of AJC. We propose that keratins and the keratin-binding protein Albatross are important for epithelial cell polarization.

  1. Auxin induces cell proliferation in an experimental model of mammalian renal tubular epithelial cells.

    Science.gov (United States)

    Cernaro, Valeria; Medici, Maria Antonietta; Leonello, Giuseppa; Buemi, Antoine; Kohnke, Franz Heinrich; Villari, Antonino; Santoro, Domenico; Buemi, Michele

    2015-06-01

    Indole-3-acetic acid is the main auxin produced by plants and plays a key role in the plant growth and development. This hormone is also present in humans where it is considered as a uremic toxin deriving from tryptophan metabolism. However, beyond this peculiar aspect, the involvement of auxin in human pathophysiology has not been further investigated. Since it is a growth hormone, we evaluated its proliferative properties in an in vitro model of mammalian renal tubular epithelial cells. We employed an experimental model of renal tubular epithelial cells belonging to the LLC-PK1 cell line that is derived from the kidney of healthy male pig. Growth effects of auxin against LLC-PK1 cell lines were determined by a rapid colorimetric assay. Increasing concentrations of auxin (to give a final concentration from 1 to 1000 ng/mL) were added and microplates were incubated for 72 h. Each auxin concentration was assayed in four wells and repeated four times. Cell proliferation significantly increased, compared to control cells, 72 h after addition of auxin to cultured LLC-PK1 cells. Statistically significant values were observed when 100 ng/mL (p auxin influences cell growth not only in plants, where its role is well documented, but also in mammalian cell lines. This observation opens new scenarios in the field of tissue regeneration and may stimulate a novel line of research aiming at investigating whether this hormone really influences human physiology and pathophysiology and in particular, kidney regeneration.

  2. Diet-Derived Short Chain Fatty Acids Stimulate Intestinal Epithelial Cells To Induce Mucosal Tolerogenic Dendritic Cells.

    Science.gov (United States)

    Goverse, Gera; Molenaar, Rosalie; Macia, Laurence; Tan, Jian; Erkelens, Martje N; Konijn, Tanja; Knippenberg, Marlene; Cook, Emma C L; Hanekamp, Diana; Veldhoen, Marc; Hartog, Anita; Roeselers, Guus; Mackay, Charles R; Mebius, Reina E

    2017-03-01

    The gastrointestinal tract is continuously exposed to many environmental factors that influence intestinal epithelial cells and the underlying mucosal immune system. In this article, we demonstrate that dietary fiber and short chain fatty acids (SCFAs) induced the expression of the vitamin A-converting enzyme RALDH1 in intestinal epithelial cells in vivo and in vitro, respectively. Furthermore, our data showed that the expression levels of RALDH1 in small intestinal epithelial cells correlated with the activity of vitamin A-converting enzymes in mesenteric lymph node dendritic cells, along with increased numbers of intestinal regulatory T cells and a higher production of luminal IgA. Moreover, we show that the consumption of dietary fiber can alter the composition of SCFA-producing microbiota and SCFA production in the small intestines. In conclusion, our data illustrate that dietary adjustments affect small intestinal epithelial cells and can be used to modulate the mucosal immune system.

  3. Protective effects of Lactobacillus plantarum on epithelial barrier disruption caused by enterotoxigenic Escherichia coli in intestinal porcine epithelial cells.

    Science.gov (United States)

    Wu, Yunpeng; Zhu, Cui; Chen, Zhuang; Chen, Zhongjian; Zhang, Weina; Ma, Xianyong; Wang, Li; Yang, Xuefen; Jiang, Zongyong

    2016-04-01

    Tight junctions (TJs) play an important role in maintaining the mucosal barrier function and gastrointestinal health of animals. Lactobacillus plantarum (L. plantarum) was reported to protect the intestinal barrier function of early-weaned piglets against enterotoxigenic Escherichia coli (ETEC) K88 challenge; however, the underlying cellular mechanism of this protection was unclear. Here, an established intestinal porcine epithelia cell (IPEC-J2) model was used to investigate the protective effects and related mechanisms of L. plantarum on epithelial barrier damages induced by ETEC K88. Epithelial permeability, expression of inflammatory cytokines, and abundance of TJ proteins, were determined. Pre-treatment with L. plantarum for 6h prevented the reduction in transepithelial electrical resistance (TEER) (Pplantarum were higher (Pplantarum was shown to regulate proteins of nuclear factor κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways. These results indicated that L. plantarum may improve epithelial barrier function by maintenance of TEER, inhibiting the reduction of TJ proteins, and reducing the expression of proinflammatory cytokines induced by ETEC K88, possibly through modulation of TLRs, NF-κB and MAPK pathways.

  4. Enamel tissue engineering using subcultured enamel organ epithelial cells in combination with dental pulp cells.

    Science.gov (United States)

    Honda, Masaki J; Shinmura, Yuka; Shinohara, Yoshinori

    2009-01-01

    We describe a strategy for the in vitro engineering of enamel tissue using a novel technique for culturing enamel organ epithelial (EOE) cells isolated from the enamel organ using 3T3-J2 cells as a feeder layer. These subcultured EOE cells retain the capacity to produce enamel structures over a period of extended culture. In brief, enamel organs from 6-month-old porcine third molars were dissociated into single cells and subcultured on 3T3-J2 feeder cell layers. These subcultured EOE cells were then seeded onto a collagen sponge in combination with primary dental pulp cells isolated at an early stage of crown formation, and these constructs were transplanted into athymic rats. After 4 weeks, complex enamel-dentin structures were detected in the implants. These results show that our culture technique maintained ameloblast lineage cells that were able to produce enamel in vivo. This novel subculture technique provides an important tool for tooth tissue engineering.

  5. The value of the Lugol's iodine staining technique for the identification of vaginal epithelial cells.

    Science.gov (United States)

    Hausmann, R; Pregler, C; Schellmann, B

    1994-01-01

    This paper reports on the specificity of the Lugol's iodine staining technique for the detection of vaginal epithelial cells on penile swabs. Air-dried swabs taken from the glans of the penis of 153 hospital patients and from 50 healthy volunteers, whose last sexual intercourse had taken place at least 5 days previously, were stained with Lugol's solution. Glycogenated cells were found in more than 50% of the cases studied, even in healthy volunteers without urethritis. In almost all of these cases the smear contained at least a few polygonal nucleated epithelial cells showing an unequivocal positive Lugol reaction. These cells cannot be distinguished from superficial or intermediate vaginal cells, by cytomorphology or staining. Urinary tract infections had no influence on the glycogen content of male squamous epithelial cells. On the basis of these results the Lugol's method can no longer be assumed to prove the presence of vaginal cells in penile swabs.

  6. Taurine Protects Lens Epithelial Cells Against Ultraviolet B-Induced Apoptosis.

    Science.gov (United States)

    Dayang, Wu; Dongbo, Pang

    2017-10-01

    The massive uptake of compatible osmolytes is a self-protective response shared by lens exposed to hypertonic stress and ultraviolet stress. This study aimed to investigate the protective effects of taurine against ultraviolet B-induced cytotoxicity in the lens epithelial cells. Real-time PCR was used to measure osmolytes transport. Radioimmunoassay was used to measure osmolytes uptake. Cell counting kit-8 assays were used to measure cellular viability. Flow cytometry analysis was used to measure apoptosis level. Compared with normotonic stress, hypertonic stress-induced osmolytes uptake into the lens epithelial cells such as betaine, myoinositol and taurine. UVB exposure increased osmolytes transporter mRNA expression together with osmolytes uptake. Moreover, taurine suppressed UVB-induced cell apoptosis in the lens epithelial cells significantly. The effect of compatible osmolyte taurine on cell survival rate may play an important role in cell resistance and adaption to UVB exposure.

  7. ADAM17 deletion in thymic epithelial cells alters aire expression without affecting T cell developmental progression.

    Directory of Open Access Journals (Sweden)

    David M Gravano

    Full Text Available BACKGROUND: Cellular interactions between thymocytes and thymic stromal cells are critical for normal T cell development. Thymic epithelial cells (TECs are important stromal niche cells that provide essential growth factors, cytokines, and present self-antigens to developing thymocytes. The identification of genes that mediate cellular crosstalk in the thymus is ongoing. One candidate gene, Adam17, encodes a metalloprotease that functions by cleaving the ectodomain of several transmembrane proteins and regulates various developmental processes. In conventional Adam17 knockout mice, a non-cell autonomous role for ADAM17 in adult T cell development was reported, which strongly suggested that expression of ADAM17 in TECs was required for normal T cell development. However, knockdown of Adam17 results in multisystem developmental defects and perinatal lethality, which has made study of the role of Adam17 in specific cell types difficult. Here, we examined T cell and thymic epithelial cell development using a conditional knockout approach. METHODOLOGY/PRINCIPAL FINDINGS: We generated an Adam17 conditional knockout mouse in which floxed Adam17 is deleted specifically in TECs by Cre recombinase under the control of the Foxn1 promoter. Normal T cell lineage choice and development through the canonical αβ T cell stages was observed. Interestingly, Adam17 deficiency in TECs resulted in reduced expression of the transcription factor Aire. However, no alterations in the patterns of TEC phenotypic marker expression and thymus morphology were noted. CONCLUSIONS/SIGNIFICANCE: In contrast to expectation, our data clearly shows that absence of Adam17 in TECs is dispensable for normal T cell development. Differentiation of TECs is also unaffected by loss of Adam17 based on phenotypic markers. Surprisingly, we have uncovered a novel genetic link between Adam17and Aire expression in vivo. The cell type in which ADAM17 mediates its non-cell autonomous impact and

  8. Human Gastric Epithelial Cells Contribute to Gastric Immune Regulation by Providing Retinoic Acid to Dendritic Cells

    OpenAIRE

    Bimczok, Diane; John Y. Kao; Zhang, Min; Cochrun, Steven; Mannon, Peter; Peter, Shajan; Wilcox, Charles M.; Mönkemüller, Klaus E; Harris, Paul R.; Grams, Jayleen M.; Stahl, Richard D.; Smith, Phillip D.; Smythies, Lesley E.

    2014-01-01

    Despite the high prevalence of chronic gastritis caused by H. pylori, the gastric mucosa has received little investigative attention as a unique immune environment. Here, we analyzed whether retinoic acid (RA), an important homeostatic factor in the small intestinal mucosa, also contributes to gastric immune regulation. We report that human gastric tissue contains high levels of the RA precursor molecule, retinol, and that gastric epithelial cells express both RA biosynthesis genes and RA res...

  9. Interleukin-6 induces an epithelial-mesenchymal transition phenotype in human adamantinomatous craniopharyngioma cells and promotes tumor cell migration

    Science.gov (United States)

    Zhou, Jie; Zhang, Chao; Pan, Jun; Chen, Ligang; Qi, Song-Tao

    2017-01-01

    Total resection of adamantinomatous craniopharyngioma (ACP) is complex and often leads to postoperative recurrence. This is due to the tendency of the tumor to invade the surrounding brain tissue and the generation of a local inflammatory state between the tumor cells and parenchyma. While there is evidence to suggest that interleukin-6 (IL-6) induces craniopharyngioma (CP)-associated inflammation, particularly in ACP, the role of IL-6 in the progression of ACP remains unclear. The results of the present study demonstrated that CP inflammation was associated with pathological classification, extent of surgery, degree of calcification and postoperative hypothalamic status scale. Cytokine antibody arrays were conducted to measure the expression of IL-6 and other inflammatory factors in tumor tissues in response to various levels of inflammatory exposure. IL-6, IL-6 receptor (IL-6R) and glycoprotein 130 expression was detected by immunohistochemistry. In addition, an ELISA was performed to quantify the levels of soluble IL-6R (sIL-6R) in the cystic fluid and supernatants of ACP cells and tumor-associated fibroblasts. These measurements demonstrated that ACP cells produce IL-6 and its associated proteins. In addition, the results revealed that while the viability of ACP cells was not affected, the migration of ACP cells was promoted by IL-6 treatment in a concentration-dependent manner. Conversely, treatment with an IL-6-blocking monoclonal antibody significantly decreased the migration of ACP cells. In addition, IL-6 treatment increased the expression of vimentin and decreased the expression of E-cadherin in a dose-dependent manner. The findings of the present study demonstrate that IL-6 may promote migration in vitro via the classic- and trans-signaling pathways by inducing epithelial-mesenchymal transition in ACP cell cultures. PMID:28487953

  10. Melanin dependent survival of Apergillus fumigatus conidia in lung epithelial cells.

    Science.gov (United States)

    Amin, Shayista; Thywissen, Andreas; Heinekamp, Thorsten; Saluz, Hans Peter; Brakhage, Axel A

    2014-07-01

    Aspergillus fumigatus is the most important air-borne pathogenic fungus of humans. Upon inhalation of conidia, the fungus makes close contact with lung epithelial cells, which only possess low phagocytic activity. These cells are in particular interesting to address the question whether there is some form of persistence of conidia of A. fumigatus in the human host. Therefore, by also using uracil-auxotrophic mutant strains, we were able to investigate the interaction of A549 lung epithelial cells and A. fumigatus conidia in detail for long periods. Interestingly, unlike professional phagocytes, our study showed that the presence of conidial dihydroxynaphthalene (DHN) melanin enhanced the uptake of A. fumigatus conidia by epithelial cells when compared with non-pigmented pksP mutant conidia. Furthermore, conidia of A. fumigatus were able to survive within epithelial cells. This was due to the presence of DHN melanin in the cell wall of conidia, because melanised wild-type conidia showed a higher survival rate inside epithelial cells and led to inhibition of acidification of phagolysosomes. Both effects were not observed for white (non-melanised) conidia of the pksP mutant strain. Moreover, in contrast to pksP mutant conidia, melanised wild-type conidia were able to inhibit the extrinsic apoptotic pathway in A549 lung epithelial cells even for longer periods. The anti-apoptotic effect was not restricted to conidia, because both conidia-derived melanin ghosts (cell-free DHN melanin) and a different type of melanin, dihydroxyphenylalanine (DOPA) melanin, acted anti-apoptotically. Taken together, these data indicate the possibility of melanin-dependent persistence of conidia in lung epithelial cells.

  11. Helicobacter pylori infection inhibits phagocyte clearance of apoptotic gastric epithelial cells.

    Science.gov (United States)

    Bimczok, Diane; Smythies, Lesley E; Waites, Ken B; Grams, Jayleen M; Stahl, Richard D; Mannon, Peter J; Peter, Shajan; Wilcox, C Mel; Harris, Paul R; Das, Soumita; Ernst, Peter B; Smith, Phillip D

    2013-06-15

    Increased apoptotic death of gastric epithelial cells is a hallmark of Helicobacter pylori infection, and altered epithelial cell turnover is an important contributor to gastric carcinogenesis. To address the fate of apoptotic gastric epithelial cells and their role in H. pylori mucosal disease, we investigated phagocyte clearance of apoptotic gastric epithelial cells in H. pylori infection. Human gastric mononuclear phagocytes were analyzed for their ability to take up apoptotic epithelial cells (AECs) in vivo using immunofluorescence analysis. We then used primary human gastric epithelial cells induced to undergo apoptosis by exposure to live H. pylori to study apoptotic cell uptake by autologous monocyte-derived macrophages. We show that HLA-DR(+) mononuclear phagocytes in human gastric mucosa contain cytokeratin-positive and TUNEL-positive AEC material, indicating that gastric phagocytes are involved in AEC clearance. We further show that H. pylori both increased apoptosis in primary gastric epithelial cells and decreased phagocytosis of the AECs by autologous monocyte-derived macrophages. Reduced macrophage clearance of apoptotic cells was mediated in part by H. pylori-induced macrophage TNF-α, which was expressed at higher levels in H. pylori-infected, compared with uninfected, gastric mucosa. Importantly, we show that H. pylori-infected gastric mucosa contained significantly higher numbers of AECs and higher levels of nonphagocytosed TUNEL-positive apoptotic material, consistent with a defect in apoptotic cell clearance. Thus, as shown in other autoimmune and chronic inflammatory diseases, insufficient phagocyte clearance may contribute to the chronic and self-perpetuating inflammation in human H. pylori infection.

  12. ARP2, a novel pro-apoptotic protein expressed in epithelial prostate cancer LNCaP cells and epithelial ovary CHO transformed cells.

    Directory of Open Access Journals (Sweden)

    Jaime Mas-Oliva

    Full Text Available Neoplastic epithelial cells generate the most aggressive types of cancers such as those located in the lung, breast, colon, prostate and ovary. During advanced stages of prostate cancer, epithelial cells are associated to the appearance of androgen-independent tumors, an apoptotic-resistant phenotype that ultimately ov