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Sample records for mucosal epithelial cells

  1. Autologous Transplantation of Oral Mucosal Epithelial Cell Sheets Cultured on an Amniotic Membrane Substrate for Intraoral Mucosal Defects

    PubMed Central

    Amemiya, Takeshi; Nakamura, Takahiro; Yamamoto, Toshiro; Kinoshita, Shigeru; Kanamura, Narisato

    2015-01-01

    The human amniotic membrane (AM) is a thin intrauterine placental membrane that is highly biocompatible and possesses anti-inflammatory and anti-scarring properties. Using AM, we developed a novel method for cultivating oral mucosal epithelial cell sheets. We investigated the autologous transplantation of oral mucosal epithelial cells cultured on AM in patients undergoing oral surgeries. We obtained specimens of AM from women undergoing cesarean sections. This study included five patients without any history of a medical disorder who underwent autologous cultured oral epithelial transplantation following oral surgical procedures. Using oral mucosal biopsy specimens obtained from these patients, we cultured oral epithelial cells on an AM carrier. We transplanted the resultant cell sheets onto the oral mucosal defects. Patients were followed-up for at least 12 months after transplantation. After 2–3 weeks of being cultured on AM, epithelial cells were well differentiated and had stratified into five to seven layers. Immunohistochemistry revealed that the cultured cells expressed highly specific mucosal epithelial cell markers and basement membrane proteins. After the surgical procedures, no infection, bleeding, rejection, or sheet detachment occurred at the reconstructed sites, at which new oral mucous membranes were evident. No recurrence was observed in the long-term follow-up, and the postoperative course was excellent. Our results suggest that AM-cultured oral mucosal epithelial cell sheets represent a useful biomaterial and feasible method for oral mucosal reconstruction. However, our primary clinical study only evaluated their effects on a limited number of small oral mucosal defects. PMID:25915046

  2. Lymphotoxin beta receptor signaling limits mucosal damage through driving IL-23 production by epithelial cells.

    PubMed

    Macho-Fernandez, E; Koroleva, E P; Spencer, C M; Tighe, M; Torrado, E; Cooper, A M; Fu, Y-X; Tumanov, A V

    2015-03-01

    The immune mechanisms regulating epithelial cell repair after injury remain poorly defined. We demonstrate here that lymphotoxin beta receptor (LTβR) signaling in intestinal epithelial cells promotes self-repair after mucosal damage. Using a conditional gene-targeted approach, we demonstrate that LTβR signaling in intestinal epithelial cells is essential for epithelial interleukin-23 (IL-23) production and protection against epithelial injury. We further show that epithelial-derived IL-23 promotes mucosal wound healing by inducing the IL-22-mediated proliferation and survival of epithelial cells and mucus production. Additionally, we identified CD4(-)CCR6(+)T-bet(-) RAR-related orphan receptor gamma t (RORγt)(+) lymphoid tissue inducer cells as the main producers of protective IL-22 after epithelial damage. Thus, our results reveal a novel role for LTβR signaling in epithelial cells in the regulation of intestinal epithelial cell homeostasis to limit mucosal damage. PMID:25183367

  3. Autonomous immunity in mucosal epithelial cells: fortifying the barrier against infection.

    PubMed

    Ross, Karen F; Herzberg, Mark C

    2016-06-01

    Mucosal epithelial cells express an autonomous innate immune response that controls the overgrowth of invaded bacteria, mitigates the harmful effects of the bacteria carried within, and does not rely on other external arms of the immune response. Epithelial cell autonomous innate immunity "respects" the social biology of invading bacteria to achieve symbiosis, and is the primary protective mechanism against pathogens. PMID:27005450

  4. Fabrication of transplantable corneal epithelial and oral mucosal epithelial cell sheets using a novel temperature-responsive closed culture device.

    PubMed

    Nakajima, Ryota; Kobayashi, Toyoshige; Kikuchi, Tetsutaro; Kitano, Yuriko; Watanabe, Hiroya; Mizutani, Manabu; Nozaki, Takayuki; Senda, Naoko; Saitoh, Kazuo; Takagi, Ryo; Yamato, Masayuki; Okano, Teruo; Takeda, Shizu

    2015-05-01

    Temperature-responsive culture surfaces make it possible to harvest transplantable carrier-free cell sheets. Here, we applied temperature-responsive polymer for polycarbonate surfaces with previously developed closed culture devices for an automated culture system in order to fabricate transplantable stratified epithelial cell sheets. Histological and immunohistochemical analyses and colony-forming assays revealed that corneal epithelial and oral mucosal epithelial cell sheets could be harvested with the temperature-responsive closed culture devices. The results were similar to those obtained using temperature-responsive culture inserts. These results indicate that the novel temperature-responsive closed culture device is useful for fabricating transplantable stratified epithelial cell sheets. PMID:23475606

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

    PubMed Central

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

    2016-01-01

    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. PMID:26938569

  6. A Novel Peptide to Treat Oral Mucositis Blocks Endothelial and Epithelial Cell Apoptosis

    SciTech Connect

    Wu Xiaoyan; Chen Peili; Sonis, Stephen T.; Lingen, Mark W.; Berger, Ann; Toback, F. Gary

    2012-07-01

    Purpose: No effective agents currently exist to treat oral mucositis (OM) in patients receiving chemoradiation for the treatment of head-and-neck cancer. We identified a novel 21-amino acid peptide derived from antrum mucosal protein-18 that is cytoprotective, mitogenic, and motogenic in tissue culture and animal models of gastrointestinal epithelial cell injury. We examined whether administration of antrum mucosal protein peptide (AMP-p) could protect against and/or speed recovery from OM. Methods and Materials: OM was induced in established hamster models by a single dose of radiation, fractionated radiation, or fractionated radiation together with cisplatin to simulate conventional treatments of head-and-neck cancer. Results: Daily subcutaneous administration of AMP-p reduced the occurrence of ulceration and accelerated mucosal recovery in all three models. A delay in the onset of erythema after irradiation was observed, suggesting that a protective effect exists even before injury to mucosal epithelial cells occurs. To test this hypothesis, the effects of AMP-p on tumor necrosis factor-{alpha}-induced apoptosis were studied in an endothelial cell line (human dermal microvascular endothelial cells) as well as an epithelial cell line (human adult low-calcium, high-temperature keratinocytes; HaCaT) used to model the oral mucosa. AMP-p treatment, either before or after cell monolayers were exposed to tumor necrosis factor-{alpha}, protected against development of apoptosis in both cell types when assessed by annexin V and propidium iodide staining followed by flow cytometry or ligase-mediated polymerase chain reaction. Conclusions: These observations suggest that the ability of AMP-p to attenuate radiation-induced OM could be attributable, at least in part, to its antiapoptotic activity.

  7. Human oral mucosal epithelial cell sheets imaging with high-resolution phase-diversity homodyne OCT

    NASA Astrophysics Data System (ADS)

    Senda, Naoko; Osawa, Kentaro

    2015-03-01

    There is a need for development of non-invasive technique to evaluate regenerative tissues such as cell sheets for transplantation. We demonstrated non-invasive imaging inside living cell sheets of human oral mucosal epithelial cells by phase-diversity homodyne optical coherence tomography (OCT). The new method OCT developed in Hitachi enables cell imaging because of high resolution (axial resolution; ~2.6 μm, lateral resolution; ~1 μm, in the air). Nuclei inside cell sheets were imaged with sufficient spatial resolution to identify each cell. It suggested that the new method OCT could be useful for non-invasive cell sheet evaluation test.

  8. TLR-Dependent Human Mucosal Epithelial Cell Responses to Microbial Pathogens

    PubMed Central

    McClure, Ryan; Massari, Paola

    2014-01-01

    Toll-like receptor (TLR) signaling represents one of the best studied pathways to implement defense mechanisms against invading microbes in human being as well as in animals. TLRs respond to specific microbial ligands and to danger signals produced by the host during infection, and initiate downstream cascades that activate both innate and adaptive immunity. TLRs are expressed by professional immune cells and by the large majority of non-hematopoietic cells, including epithelial cells. In epithelial tissues, TLR functions are particularly important because these sites are constantly exposed to microorganisms, due to their location at the host interface with the environment. While at these sites specific defense mechanisms and inflammatory responses are initiated via TLR signaling against pathogens, suppression or lack of TLR activation is also observed in response to the commensal microbiota. The mechanisms by which TLR signaling is regulated in mucosal epithelial cells include differential expression and levels of TLRs (and their signaling partners), their cellular localization and positioning within the tissue in a fashion that favors responses to pathogens while dampening responses to commensals and maintaining tissue homeostasis in physiologic conditions. In this review, the expression and activation of TLRs in mucosal epithelial cells of several sites of the human body are examined. Specifically, the oral cavity, the ear canal and eye, the airways, the gut, and the reproductive tract are discussed, along with how site-specific host defense mechanisms are implemented via TLR signaling. PMID:25161655

  9. Intestinal epithelial tuft cells initiate type 2 mucosal immunity to helminth parasites.

    PubMed

    Gerbe, François; Sidot, Emmanuelle; Smyth, Danielle J; Ohmoto, Makoto; Matsumoto, Ichiro; Dardalhon, Valérie; Cesses, Pierre; Garnier, Laure; Pouzolles, Marie; Brulin, Bénédicte; Bruschi, Marco; Harcus, Yvonne; Zimmermann, Valérie S; Taylor, Naomi; Maizels, Rick M; Jay, Philippe

    2016-01-14

    Helminth parasitic infections are a major global health and social burden. The host defence against helminths such as Nippostrongylus brasiliensis is orchestrated by type 2 cell-mediated immunity. Induction of type 2 cytokines, including interleukins (IL) IL-4 and IL-13, induce goblet cell hyperplasia with mucus production, ultimately resulting in worm expulsion. However, the mechanisms underlying the initiation of type 2 responses remain incompletely understood. Here we show that tuft cells, a rare epithelial cell type in the steady-state intestinal epithelium, are responsible for initiating type 2 responses to parasites by a cytokine-mediated cellular relay. Tuft cells have a Th2-related gene expression signature and we demonstrate that they undergo a rapid and extensive IL-4Rα-dependent amplification following infection with helminth parasites, owing to direct differentiation of epithelial crypt progenitor cells. We find that the Pou2f3 gene is essential for tuft cell specification. Pou2f3(-/-) mice lack intestinal tuft cells and have defective mucosal type 2 responses to helminth infection; goblet cell hyperplasia is abrogated and worm expulsion is compromised. Notably, IL-4Rα signalling is sufficient to induce expansion of the tuft cell lineage, and ectopic stimulation of this signalling cascade obviates the need for tuft cells in the epithelial cell remodelling of the intestine. Moreover, tuft cells secrete IL-25, thereby regulating type 2 immune responses. Our data reveal a novel function of intestinal epithelial tuft cells and demonstrate a cellular relay required for initiating mucosal type 2 immunity to helminth infection. PMID:26762460

  10. Evidence of the Survival of Ectopically Transplanted Oral Mucosal Epithelial Stem Cells After Repeated Wounding of Cornea

    PubMed Central

    Sugiyama, Hiroaki; Yamato, Masayuki; Nishida, Kohji; Okano, Teruo

    2014-01-01

    Tissue engineering has become an essential tool in the development of regenerative medicine. We have developed cell sheet–based techniques for use in regenerative medicine that have already been successfully used in clinical applications. Native corneal epithelium is produced from limbal stem cells located in the transition zone between the cornea and the bulbar conjunctiva. Limbal stem cell deficiency (LSCD) is a severe defect of the limbal stem cells leading to vision loss due to conjunctival epithelial invasion and neovascularization. Rabbit LSCD models were treated with transplantable autologous oral mucosal epithelial cell (OEC) sheets fabricated on temperature-responsive cell culture surfaces, after which, the ocular surfaces were clear and smooth with no observable defects. The central part of the reconstructed ocular surface was scraped and wounded, after which proliferating epithelial cells covered the scraped area within a few days. The ocular surfaces were clear and smooth even after repeated scrapings and consisted of only OECs or heterogeneously mixed with corneal epithelial cells. This study demonstrates that transplanted cell sheets containing oral mucosal epithelial stem cells could reconstruct the ocular surface to maintain cornea homeostasis; moreover, they provide an ideal microenvironment to support the proliferation of remaining native limbal stem cells. PMID:24769908

  11. Mucosal production of uric acid by airway epithelial cells contributes to particulate matter-induced allergic sensitization.

    PubMed

    Gold, M J; Hiebert, P R; Park, H Y; Stefanowicz, D; Le, A; Starkey, M R; Deane, A; Brown, A C; Liu, G; Horvat, J C; Ibrahim, Z A; Sukkar, M B; Hansbro, P M; Carlsten, C; VanEeden, S; Sin, D D; McNagny, K M; Knight, D A; Hirota, J A

    2016-05-01

    Exposure to particulate matter (PM), a major component of air pollution, contributes to increased morbidity and mortality worldwide. PM induces innate immune responses and contributes to allergic sensitization, although the mechanisms governing this process remain unclear. Lung mucosal uric acid has also been linked to allergic sensitization. The links among PM exposure, uric acid, and allergic sensitization remain unexplored. We therefore investigated the mechanisms behind PM-induced allergic sensitization in the context of lung mucosal uric acid. PM10 and house dust mite exposure selectively induced lung mucosal uric acid production and secretion in vivo, which did not occur with other challenges (lipopolysaccharide, virus, bacteria, or inflammatory/fibrotic stimuli). PM10-induced uric acid mediates allergic sensitization and augments antigen-specific T-cell proliferation, which is inhibited by uricase. We then demonstrate that human airway epithelial cells secrete uric acid basally and after stimulation through a previously unidentified mucosal secretion system. Our work discovers a previously unknown mechanism of air pollution-induced, uric acid-mediated, allergic sensitization that may be important in the pathogenesis of asthma. PMID:26509876

  12. Early Mucosal Sensing of SIV Infection by Paneth Cells Induces IL-1β Production and Initiates Gut Epithelial Disruption

    PubMed Central

    Bourry, Olivier; Hu, William K.; Somrit, Monsicha; Sankaran-Walters, Sumathi; Gaulke, Chris A.; Fenton, Anne N.; Li, Jay A.; Crawford, Robert W.; Chuang, Frank; Tarara, Ross; Marco, Maria L.; Bäumler, Andreas J.; Cheng, Holland; Dandekar, Satya

    2014-01-01

    HIV causes rapid CD4+ T cell depletion in the gut mucosa, resulting in immune deficiency and defects in the intestinal epithelial barrier. Breakdown in gut barrier integrity is linked to chronic inflammation and disease progression. However, the early effects of HIV on the gut epithelium, prior to the CD4+ T cell depletion, are not known. Further, the impact of early viral infection on mucosal responses to pathogenic and commensal microbes has not been investigated. We utilized the SIV model of AIDS to assess the earliest host-virus interactions and mechanisms of inflammation and dysfunction in the gut, prior to CD4+ T cell depletion. An intestinal loop model was used to interrogate the effects of SIV infection on gut mucosal immune sensing and response to pathogens and commensal bacteria in vivo. At 2.5 days post-SIV infection, low viral loads were detected in peripheral blood and gut mucosa without CD4+ T cell loss. However, immunohistological analysis revealed the disruption of the gut epithelium manifested by decreased expression and mislocalization of tight junction proteins. Correlating with epithelial disruption was a significant induction of IL-1β expression by Paneth cells, which were in close proximity to SIV-infected cells in the intestinal crypts. The IL-1β response preceded the induction of the antiviral interferon response. Despite the disruption of the gut epithelium, no aberrant responses to pathogenic or commensal bacteria were observed. In fact, inoculation of commensal Lactobacillus plantarum in intestinal loops led to rapid anti-inflammatory response and epithelial tight junction repair in SIV infected macaques. Thus, intestinal Paneth cells are the earliest responders to viral infection and induce gut inflammation through IL-1β signaling. Reversal of the IL-1β induced gut epithelial damage by Lactobacillus plantarum suggests synergistic host-commensal interactions during early viral infection and identify these mechanisms as potential

  13. Neonatal Fc Receptor-Mediated IgG Transport Across Porcine Intestinal Epithelial Cells: Potentially Provide the Mucosal Protection.

    PubMed

    Guo, Jinyue; Li, Fei; He, Qigai; Jin, Hui; Liu, Mei; Li, Shaowen; Hu, Sishun; Xiao, Yuncai; Bi, Dingren; Li, Zili

    2016-06-01

    It has been well characterized that piglets can absorb colostrum IgG across the intestine to neonatal bloodstream and a certain level of IgG has been found in the mucosal secretions of the porcine intestinal tract. However, little is known about how the maternal IgG transport across the intestinal barrier and how IgG enter the lumen of intestinal tract. In this study, we demonstrated that the porcine neonatal Fc receptor (pFcRn) was expressed in a model of normal porcine intestinal epithelial cells (IPEC-J2) as well as in kidney cells (PK-15), and pFcRn was mainly distributed in the apical side of the polarized IPEC-J2 cells. Analyzing the phylogenetic relatedness of this gene we found that swine and human neonatal Fc receptor (FcRn) amino acid sequence are closer than rodents. We also showed that pFcRn mediated bidirectional IgG transport across polarized IPEC-J2 cells and bound to IgG in a pH-dependent manner. Furthermore, pFcRn-transcytosed viral-specific IgG reduced the transmissible gastroenteritis virus (TGEV) yield from the luminal direction by a 50% tissue culture infective dose (TCID50) assay. Our results indicate that pFcRn-dependent bidirectional IgG transport across the intestinal epithelium plays critical role in the acquisition of humoral immunity in early life and in host defense at mucosal surfaces. PMID:26982157

  14. Glutamate is the major anaplerotic substrate in the tricarboxylic acid cycle of isolated rumen epithelial and duodenal mucosal cells from beef cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study aimed to determine the contribution of substrates to tricarboxylic acid (TCA) cycle fluxes in rumen epithelial (REC) and duodenal mucosal (DMC) cells isolated from bulls (n = 6) fed either a 75% forage (HF) or 75% concentrate (HC) diet. In separate incubations, [13C6]glucose, [13C5]glutam...

  15. Muc1 Cell Surface Mucin Attenuates Epithelial Inflammation in Response to a Common Mucosal Pathogen*

    PubMed Central

    Guang, Wei; Ding, Hua; Czinn, Steven J.; Kim, K. Chul; Blanchard, Thomas G.; Lillehoj, Erik P.

    2010-01-01

    Helicobacter pylori infection of the gastric mucosa causes an active-chronic inflammation that is strongly linked to the development of duodenal and gastric ulcers and stomach cancer. However, greater than 80% of individuals infected with H. pylori are asymptomatic beyond histologic inflammation, and it is unknown what factors influence the incidence and character of bacterial-associated gastritis and related disorders. Because previous studies demonstrated that the Muc1 epithelial glycoprotein inhibited inflammation during acute lung infection by Pseudomonas aeruginosa, we asked whether Muc1 might also counter-regulate gastric inflammation in response to H. pylori infection. Muc1−/− mice displayed increased bacterial colonization of the stomach and greater TNF-α and keratinocyte chemoattractant transcript levels compared with Muc1+/+ mice after experimental H. pylori infection. Knockdown of Muc1 expression in AGS human gastric epithelial cells by RNA interference was associated with increased phosphorylation of IκBα, augmented activation and nuclear translocation of NF-κB, and enhanced production of interleulin-8 compared with Muc1-expressing cells. Conversely, Muc1 overexpression was correlated with decreased NF-κB activation, reduced interleulin-8 production, and diminished IκB kinase β (IKKβ)/IKKγ coimmunoprecipitation compared with cells expressing Muc1 endogenously. Cotransfection of AGS cells with Muc1 plus IKKβ, but not a catalytically inactive IKKβ mutant, reversed the Muc1 inhibitory effect. Finally, Muc1 formed a coimmunoprecipitation complex with IKKγ but not with IKKβ. These results are consistent with the hypothesis that Muc1 binds to IKKγ, thereby inhibiting formation of the catalytically active IKK complex and blocking the ability of H. pylori to stimulate IκBα phosphorylation, NF-κB activation, and downstream inflammatory responses. PMID:20430889

  16. Characterization of Binding of Candida albicans to Small Intestinal Mucin and Its Role in Adherence to Mucosal Epithelial Cells

    PubMed Central

    de Repentigny, Louis; Aumont, Francine; Bernard, Karine; Belhumeur, Pierre

    2000-01-01

    In order to approximate and adhere to mucosal epithelial cells, Candida must traverse the overlying mucus layer. Interactions of Candida species with mucin and human buccal epithelial cells (BECs) were thus investigated in vitro. Binding of the Candida species to purified small intestinal mucin showed a close correlation with their hierarchy of virulence. Significant differences (P < 0.05) were found among three categories of Candida species adhering highly (C. dubliniensis, C. tropicalis, and C. albicans), moderately (C. parapsilosis and C. lusitaniae) or weakly (C. krusei and C. glabrata) to mucin. Adherence of C. albicans to BECs was quantitatively inhibited by graded concentrations of mucin. However, inhibition of adherence was reversed by pretreatment of mucin with pronase or C. albicans secretory aspartyl proteinase Sap2p but not with sodium periodate. Saturable concentration- and time-dependent binding of mucin to C. albicans was abrogated by pronase or Sap2p treatment of mucin but was unaffected by β-mercaptoethanol, sodium periodate, neuraminidase, lectins, or potentially inhibitory sugars. Probing of membrane blots of the mucin with C. albicans revealed binding of the yeast to the 66-kDa cleavage product of the 118-kDa C-terminal glycopeptide of mucin. Although no evidence was found for the participation of C. albicans cell surface mannoproteins in specific receptor-ligand binding to mucin, inhibition of binding by p-nitrophenol (1 mM) and tetramethylurea (0.36 M) revealed that hydrophobic interactions are involved in adherence of C. albicans to mucin. These results suggest that C. albicans may both adhere to and enzymatically degrade mucins by the action of Saps, and that both properties may act to modulate Candida populations in the oral cavity and gastrointestinal tract. PMID:10816460

  17. NOD1 agonist iE-DAP reverses effects of cigarette smoke extract on NOD1 signal pathway in human oral mucosal epithelial cells

    PubMed Central

    Gao, Yafan; Jiang, Wenhui; Qian, Yajie; Zhou, Qian; Jiang, Hongliu; Wang, Xiang; Wang, Wenmei

    2015-01-01

    Smoking is a well-known risk factor for many systemic diseases and oral disorders. Smoking has been recognized to cause diminished defense, persistent inflammation and result in disease development. Nucleotide binding oligomerization domain 1 (NOD1) signal pathway plays a key role in innate immune and tissue homeostasis. Our recent studies confirmed that cigarette smoke extract (CSE) could inhibit NOD1 expression and affect expression levels of crucial molecules of NOD1 signaling in oral mucosal epithelial cells. In the present study, immortalized human oral mucosal epithelial (Leuk-1) cells were treated with CSE, iE-DAP (NOD1 agonist), CSE + iE-DAP, respectively. Western blotting analysis demonstrated that iE-DAP triggered NOD1 expression of leuk-1 cells in a dose-dependent manner. iE-DAP also reversed the suppressive effect of CSE on NOD1 expression and prevented the overactivation of RIP2 and P-NF-κB following CSE exposure. Real-time PCR and ELISA results confirmed that iE-DAP reversed CSE-mediated effects on the mRNA levels and releases of IL-6, IL-8, TNF-α and IFN-γ by Leuk-1 cells. Taken together, our results indicated that NOD1 activation with iE-DAP could reverse CSE-mediated effects on NOD1 signaling in human oral mucosal epithelial cells. PMID:26550162

  18. Epithelial MUC1 promotes cell migration, reduces apoptosis and affects levels of mucosal modulators during acetylsalicylic acid (aspirin)-induced gastropathy.

    PubMed

    Banerjee, Debashish; Fernandez, Harvey Robert; Patil, Pradeep Bhatu; Premaratne, Pushpa; Quiding-Järbrink, Marianne; Lindén, Sara Katarina

    2015-02-01

    MUC1 is a transmembrane mucin highly expressed in the stomach. Although extensive research has uncovered many of its roles in cancer, knowledge about the functions of MUC1 in normal tissues is limited. In the present study, we showed that acetylsalicylic acid (ASA; aspirin) up-regulated MUC1/Muc1 expression in the gastric mucosa of humans and wild-type (WT) mice. ASA induced mucosal injury in all mice to a similar extent; however, WT animals and those chimaeras with Muc1 on the epithelia recovered faster than Muc1-knockout (KO) mice and chimaeras carrying Muc1 on haemopoietic but not epithelial cells. MUC1 enhanced proliferation and migration of the human gastric cell line MKN-7 and increased resistance to apoptosis. The repeated treatment regime used caused a reduction in cyclo-oxygenase-1 (Cox-1) expression, though WT animals returned faster towards pre-treatment levels and had increased Cox-2 and vascular endothelial growth factor levels during recovery. Thus we found that epithelial Muc1 is more important for the healing process than haemopoietic Muc1 and Muc1/MUC1 facilitates wound healing by enhancing cell migration and proliferation, protecting against apoptosis and mediating expression of mucosal modulators. Thus MUC1 plays essential roles during wound healing and development of treatment modalities targeting enhanced expression of MUC1 may be beneficial to treat mucosal wounds. PMID:25387004

  19. Peptides from cytomegalovirus UL130 and UL131 proteins induce high titer antibodies that block viral entry into mucosal epithelial cells

    PubMed Central

    Saccoccio, Frances M.; Sauer, Anne L.; Cui, Xiaohong; Armstrong, Amy E.; Habib, EL-Sayed E.; Johnson, David C.; Ryckman, Brent J.; Klingelhutz, Aloysius J.; Adler, Stuart P.; McVoy, Michael A.

    2011-01-01

    Cytomegalovirus infections are an important cause of disease for which no licensed vaccine exists. Recent studies have focused on the gH/gL/UL128-131 complex as antibodies to gH/gL/UL128-131 neutralize viral entry into epithelial cells. Prior studies have used cells from the retinal pigment epithelium, while to prevent transmission, vaccine-induced antibodies may need to block viral infection of epithelial cells of the oral or genital mucosa. We found that gH/gL/UL128-131 is necessary for efficient viral entry into epithelial cells derived from oral and genital mucosa, that short peptides from UL130 and UL131 elicit high titer neutralizing antibodies in rabbits, and that such antibodies neutralize viral entry into epithelial cells derived from these relevant tissues. These results suggest that single subunits or peptides may be sufficient to elicit potent epithelial entry neutralizing responses and that secretory antibodies to such neutralizing epitopes have the potential to provide sterilizing immunity by blocking initial mucosal infection. PMID:21310190

  20. Mucosal dendritic cells shape mucosal immunity

    PubMed Central

    Chang, Sun-Young; Ko, Hyun-Jeong; Kweon, Mi-Na

    2014-01-01

    Dendritic cells (DCs) are key modulators that shape the immune system. In mucosal tissues, DCs act as surveillance systems to sense infection and also function as professional antigen-presenting cells that stimulate the differentiation of naive T and B cells. On the basis of their molecular expression, DCs can be divided into several subsets with unique functions. In this review, we focus on intestinal DC subsets and their function in bridging the innate signaling and adaptive immune systems to maintain the homeostasis of the intestinal immune environment. We also review the current strategies for manipulating mucosal DCs for the development of efficient mucosal vaccines to protect against infectious diseases. PMID:24626170

  1. Caspase-12 silencing attenuates inhibitory effects of cigarette smoke extract on NOD1 signaling and hBDs expression in human oral mucosal epithelial cells.

    PubMed

    Wang, Xiang; Qian, Ya-jie; Zhou, Qian; Ye, Pei; Duan, Ning; Huang, Xiao-feng; Zhu, Ya-nan; Li, Jing-jing; Hu, Li-ping; Zhang, Wei-yun; Han, Xiao-dong; Wang, Wen-mei

    2014-01-01

    Cigarette smoke exposure is associated with increased risk of various diseases. Epithelial cells-mediated innate immune responses to infectious pathogens are compromised by cigarette smoke. Although many studies have established that cigarette smoke exposure affects the expression of Toll-liked receptor (TLR), it remains unknown whether the nucleotide-binding oligomerization domain-containing protein 1 (NOD1) expression is affected by cigarette smoke exposure. In the study, we investigated effects of cigarette smoke extract (CSE) on NOD1 signaling in an immortalized human oral mucosal epithelial (Leuk-1) cell line. We first found that CSE inhibited NOD1 expression in a dose-dependent manner. Moreover, CSE modulated the expression of other crucial molecules in NOD1 signaling and human β defensin (hBD) 1, 2 and 3. We found that RNA interference-induced Caspase-12 silencing increased NOD1 and phospho-NF-κB (p-NF-κB) expression and down-regulated RIP2 expression. The inhibitory effects of CSE on NOD1 signaling can be attenuated partially through Caspase-12 silencing. Intriguingly, Caspase-12 silencing abrogated inhibitory effects of CSE on hBD1, 3 expression and augmented induced effect of CSE on hBD2 expression. Caspase-12 could play a vital role in the inhibitory effects of cigarette smoke on NOD1 signaling and hBDs expression in oral mucosal epithelial cells. PMID:25503380

  2. Caspase-12 Silencing Attenuates Inhibitory Effects of Cigarette Smoke Extract on NOD1 Signaling and hBDs Expression in Human Oral Mucosal Epithelial Cells

    PubMed Central

    Wang, Xiang; Qian, Ya-jie; Zhou, Qian; Ye, Pei; Duan, Ning; Huang, Xiao-feng; Zhu, Ya-nan; Li, Jing-jing; Hu, Li-ping; Zhang, Wei-yun; Han, Xiao-dong; Wang, Wen-mei

    2014-01-01

    Cigarette smoke exposure is associated with increased risk of various diseases. Epithelial cells-mediated innate immune responses to infectious pathogens are compromised by cigarette smoke. Although many studies have established that cigarette smoke exposure affects the expression of Toll-liked receptor (TLR), it remains unknown whether the nucleotide-binding oligomerization domain-containing protein 1 (NOD1) expression is affected by cigarette smoke exposure. In the study, we investigated effects of cigarette smoke extract (CSE) on NOD1 signaling in an immortalized human oral mucosal epithelial (Leuk-1) cell line. We first found that CSE inhibited NOD1 expression in a dose-dependent manner. Moreover, CSE modulated the expression of other crucial molecules in NOD1 signaling and human β defensin (hBD) 1, 2 and 3. We found that RNA interference-induced Caspase-12 silencing increased NOD1 and phospho-NF-κB (p-NF-κB) expression and down-regulated RIP2 expression. The inhibitory effects of CSE on NOD1 signaling can be attenuated partially through Caspase-12 silencing. Intriguingly, Caspase-12 silencing abrogated inhibitory effects of CSE on hBD1, 3 expression and augmented induced effect of CSE on hBD2 expression. Caspase-12 could play a vital role in the inhibitory effects of cigarette smoke on NOD1 signaling and hBDs expression in oral mucosal epithelial cells. PMID:25503380

  3. Human Immunodeficiency Virus Type 1 Stimulates the Expression and Production of Secretory Leukocyte Protease Inhibitor (SLPI) in Oral Epithelial Cells: a Role for SLPI in Innate Mucosal Immunity

    PubMed Central

    Jana, N. K.; Gray, L. R.; Shugars, D. C.

    2005-01-01

    The innate immune response is a key barrier against pathogenic microorganisms such as human immunodeficiency virus type 1 (HIV-1). Because HIV-1 is rarely transmitted orally, we hypothesized that oral epithelial cells participate in the innate immune defense against this virus. We further hypothesized that secretory leukocyte protease inhibitor (SLPI), a 12-kDa mucosal antiviral protein, is a component of the host immune response to this virus. Here we demonstrated constitutive expression and production of SLPI in immortalized human oral keratinocytes. Brief exposure of cells to HIV-1 BaL and HXB2 significantly increased SLPI mRNA and protein production compared to that in mock-exposed cells (P < 0.01), as evaluated by real-time quantitative reverse transcription-PCR and enzyme-linked immunosorbent assay. HIV-1-mediated stimulation of SLPI occurred at the transcriptional level, was dose and time dependent, was elicited by heat-inactivated and infectious viruses, and did not depend on cellular infection. Experiments with purified retroviral proteins showed that the stimulatory effect was induced specifically by external envelope glycoproteins from HIV-1 and simian immunodeficiency virus. SLPI responsiveness to HIV-1 was also observed in an unrelated oral epithelial cell line and in normal (nonimmortalized) human oral epithelial cells isolated from healthy uninfected gingival tissues. In this first report of SLPI regulation by HIV-1, we show that the expression and production of the antimicrobial and anti-inflammatory protein can be stimulated in oral epithelial cells by the virus through interactions with gp120 in the absence of direct infection. These findings indicate that SLPI is a component of the oral mucosal response to HIV-1. PMID:15858026

  4. Critical Roles of Intestinal Epithelial Vitamin D Receptor Signaling in Controlling Gut Mucosal Inflammation

    PubMed Central

    Li, Yan Chun; Chen, Yunzi; Du, Jie

    2015-01-01

    Although vitamin D receptor (VDR) is highly expressed in the intestine, the role of VDR signaling in the gut is not fully understood. Our recent studies unveil a regulatory circuit that centers gut epithelial VDR as a key molecule in the control of mucosal inflammation and colitis development. On the one hand, intestinal epithelial VDR signaling protects the integrity of the mucosal barrier by inhibiting inflammation-induced epithelial cell apoptosis. This barrier-protecting, anti-colitic activity is independent of the non-epithelial immune VDR actions. A healthy and intact mucosal barrier prevents bacterial invasion and thus reduces mucosal inflammation. On the other hand, inflammation in turn down-regulates epithelial VDR expression by inducing VDR-targeting microRNA-346, thus compromising mucosal barrier functions. Consistently, colonic epithelial VDR levels are markedly reduced in patients with inflammatory bowel diseases or in experimental colitis models, whereas vitamin D analog therapy that ameliorates colitis up-regulates epithelial VDR. Thus, gut epithelial VDR signaling appears to play an essential role in controlling mucosal inflammation and thus could be a useful therapeutic target in the management of inflammatory bowel diseases. PMID:25603468

  5. Neutrophil Interactions with Epithelial Expressed ICAM-1 Enhances Intestinal Mucosal Wound Healing

    PubMed Central

    Sumagin, R; Brazil, JC; Nava, P; Nishio, H; Alam, A; Luissint, AC; Weber, DA; Neish, AS; Nusrat, A; Parkos, CA

    2015-01-01

    A characteristic feature of gastrointestinal tract inflammatory disorders, such as inflammatory bowel disease, is polymorphonuclear neutrophil (PMN) transepithelial migration (TEM) and accumulation in the gut lumen. PMN accumulation within the intestinal mucosa contributes to tissue injury. While epithelial infiltration by large numbers of PMNs results in mucosal injury, we found that PMN interactions with luminal epithelial membrane receptors may also play a role in wound healing. Intercellular adhesion molecule-1 (ICAM-1) is a PMN ligand that is upregulated on apical surfaces of intestinal epithelial cells under inflammatory conditions. In our study, increased expression of ICAM-1 resulted in enhanced PMN binding to the apical epithelium, which was associated with reduced PMN apoptosis. Following TEM, PMN adhesion to ICAM-1 resulted in activation of Akt and β-catenin signaling, increased epithelial-cell proliferation, and wound healing. Such responses were ICAM-1 dependent as engagement of epithelial ICAM-1 by antibody-mediated cross-linking yielded similar results. Furthermore, using an in-vivo biopsy-based, colonic-mucosal-injury model, we demonstrated epithelial ICAM-1 plays an important role in activation of epithelial Akt and β-catenin signaling and wound healing. These findings suggest that post-migrated PMNs within the intestinal lumen can regulate epithelial homeostasis, thereby identifying ICAM-1 as a potential therapeutic target for promoting mucosal wound healing. PMID:26732677

  6. Neutrophil interactions with epithelial-expressed ICAM-1 enhances intestinal mucosal wound healing.

    PubMed

    Sumagin, R; Brazil, J C; Nava, P; Nishio, H; Alam, A; Luissint, A C; Weber, D A; Neish, A S; Nusrat, A; Parkos, C A

    2016-09-01

    A characteristic feature of gastrointestinal tract inflammatory disorders, such as inflammatory bowel disease, is polymorphonuclear neutrophil (PMN) transepithelial migration (TEM) and accumulation in the gut lumen. PMN accumulation within the intestinal mucosa contributes to tissue injury. Although epithelial infiltration by large numbers of PMNs results in mucosal injury, we found that PMN interactions with luminal epithelial membrane receptors may also play a role in wound healing. Intercellular adhesion molecule-1 (ICAM-1) is a PMN ligand that is upregulated on apical surfaces of intestinal epithelial cells under inflammatory conditions. In our study, increased expression of ICAM-1 resulted in enhanced PMN binding to the apical epithelium, which was associated with reduced PMN apoptosis. Following TEM, PMN adhesion to ICAM-1 resulted in activation of Akt and β-catenin signaling, increased epithelial-cell proliferation, and wound healing. Such responses were ICAM-1 dependent as engagement of epithelial ICAM-1 by antibody-mediated cross-linking yielded similar results. Furthermore, using an in-vivo biopsy-based, colonic-mucosal-injury model, we demonstrated epithelial ICAM-1 has an important role in activation of epithelial Akt and β-catenin signaling and wound healing. These findings suggest that post-migrated PMNs within the intestinal lumen can regulate epithelial homeostasis, thereby identifying ICAM-1 as a potential therapeutic target for promoting mucosal wound healing. PMID:26732677

  7. Epithelial Cell Regulation of Allergic Diseases.

    PubMed

    Gour, Naina; Lajoie, Stephane

    2016-09-01

    Allergic diseases, which have escalated in prevalence in recent years, arise as a result of maladaptive immune responses to ubiquitous environmental stimuli. Why only certain individuals mount inappropriate type 2 immune responses to these otherwise harmless allergens has remained an unanswered question. Mounting evidence suggests that the epithelium, by sensing its environment, is the central regulator of allergic diseases. Once considered to be a passive barrier to allergens, epithelial cells at mucosal surfaces are now considered to be the cornerstone of the allergic diathesis. Beyond their function as maintaining barrier at mucosal surfaces, mucosal epithelial cells through the secretion of mediators like IL-25, IL-33, and TSLP control the fate of downstream allergic immune responses. In this review, we will discuss the advances in recent years regarding the process of allergen recognition and secretion of soluble mediators by epithelial cells that shape the development of the allergic response. PMID:27534656

  8. No longer an innocent bystander: epithelial toll-like receptor signaling in the development of mucosal inflammation.

    PubMed

    Gribar, Steven C; Richardson, Ward M; Sodhi, Chhinder P; Hackam, David J

    2008-01-01

    Diseases of mucosal inflammation represent important causes of morbidity and mortality, and have led to intense research efforts to understand the factors that lead to their development. It is well accepted that a breakdown of the normally impermeant epithelial barrier of the intestine, the lung, and the kidney is associated with the development of inflammatory disease in these organs, yet significant controversy exists as to how this breakdown actually occurs, and how such a breakdown may lead to inflammation. In this regard, much work has focused upon the role of the epithelium as an "innocent bystander," a target of a leukocyte-mediated inflammatory cascade that leads to its destruction in the mucosal inflammatory process. However, recent evidence from a variety of laboratories indicates that the epithelium is not merely a passive component in the steps that lead to mucosal inflammation, but is a central participant in the process. In addressing this controversy, we and others have determined that epithelial cells express Toll-like receptors (TLRs) of the innate immune system, and that activation of TLRs by endogenous and exogenous ligands may play a central role in determining the balance between a state of "mucosal homeostasis," as is required for optimal organ function, and "mucosal injury," leading to mucosal inflammation and barrier breakdown. In particular, activation of TLRs within intestinal epithelial cells leads to the development of cellular injury and impairment in mucosal repair in the pathogenesis of intestinal inflammation, while activation of TLRs in the lung and kidney may participate in the development of pneumonitis and nephritis respectively. Recent work in support of these concepts is extensively reviewed, while essential areas of further study that are required to determine the significance of epithelial TLR signaling during states of health and disease are outlined. PMID:18584047

  9. PROXIMAL GUT MUCOSAL EPITHELIAL HOMEOSTASIS IN AGED IL-1 TYPE I RECEPTOR KNOCKOUT MICE AFTER STARVATION

    PubMed Central

    Song, Juquan; Wolf, Steven E.; Wu, Xiao-Wu; Finnerty, Celeste C.; Herndon, David N.; Jeschke, Marc G.

    2010-01-01

    Background Previous studies have shown that starvation induces small bowel atrophy, and that atrophy diminishes with aging. In this experiment, we assessed whether starvation-induced atrophy of proximal gut mucosa is associated with the Interleukin-1 receptor (IL-1R) signaling pathway in aged mice. Materials and Methods Thirty 26-month-old IL-1R knockout mice and age-matched wild-type C57BL/6 mice were randomly divided into two groups: ad libitum fed and fasted. Mice were euthanized 12 or 48 hours after starvation. The proximal small bowel was harvested for morphologic analysis. Gut epithelial cell proliferation was detected using immunohistochemical staining for proliferating cell nuclear antigen (PCNA), and apoptosis was identified using terminal deoxyuridine nick-end labeling (TUNEL) staining. Results Aged IL-1R knockout mice were larger than aged-matched wild-type mice (p<0.05). Proximal gut mucosal height and mucosal cell number were not different between aged IL-1R knockout and wild-type groups. The apoptosis index in gut epithelial cells was higher in fed IL-1R knockout versus wild-type mice (p<0.05), while no significant difference in cell proliferation between both groups. Mucosal atrophy was induced in both aged IL-1R knockout and wild-type groups by starvation (p<0.05), however, aged IL-1R knockout mice experienced greater losses in proximal gut weight, mucosal length, and corresponding cell number than did wild-type mice at the 12-hour time point (p<0.05). The apoptosis index in gut epithelial cells significantly increased in both groups after starvation (p<0.05). Starvation decreased cell proliferation in IL-1R knockout mice (p<0.05), but not in wild-type mice. Conclusions The response in aged IL-1R knockout mice differs from wild-type mice in that starvation increases atrophy and is associated with decreased cell proliferation rather than increased apoptosis. PMID:20605606

  10. Innate immunity in HIV-1 infection: epithelial and non-specific host factors of mucosal immunity- a workshop report.

    PubMed

    Nittayananta, W; Weinberg, A; Malamud, D; Moyes, D; Webster-Cyriaque, J; Ghosh, S

    2016-04-01

    The interplay between HIV-1 and epithelial cells represents a critical aspect in mucosal HIV-1 transmission. Epithelial cells lining the oral cavity cover subepithelial tissues, which contain virus-susceptible host cells including CD4(+) T lymphocytes, monocytes/macrophages, and dendritic cells. Oral epithelia are among the sites of first exposure to both cell-free and cell-associated virus HIV-1 through breast-feeding and oral-genital contact. However, oral mucosa is considered to be naturally resistant to HIV-1 transmission. Oral epithelial cells have been shown to play a crucial role in innate host defense. Nevertheless, it is not clear to what degree these local innate immune factors contribute to HIV-1 resistance of the oral mucosa. This review paper addressed the following issues that were discussed at the 7th World Workshop on Oral Health and Disease in AIDS held in Hyderabad, India, during November 6-9, 2014: (i) What is the fate of HIV-1 after interactions with oral epithelial cells?; (ii) What are the keratinocyte and other anti-HIV effector oral factors, and how do they contribute to mucosal protection?; (iii) How can HIV-1 interactions with oral epithelium affect activation and populations of local immune cells?; (iv) How can HIV-1 interactions alter functions of oral epithelial cells? PMID:27109285

  11. Protective mucosal immunity mediated by epithelial CD1d and IL-10.

    PubMed

    Olszak, Torsten; Neves, Joana F; Dowds, C Marie; Baker, Kristi; Glickman, Jonathan; Davidson, Nicholas O; Lin, Chyuan-Sheng; Jobin, Christian; Brand, Stephan; Sotlar, Karl; Wada, Koichiro; Katayama, Kazufumi; Nakajima, Atsushi; Mizuguchi, Hiroyuki; Kawasaki, Kunito; Nagata, Kazuhiro; Müller, Werner; Snapper, Scott B; Schreiber, Stefan; Kaser, Arthur; Zeissig, Sebastian; Blumberg, Richard S

    2014-05-22

    The mechanisms by which mucosal homeostasis is maintained are of central importance to inflammatory bowel disease. Critical to these processes is the intestinal epithelial cell (IEC), which regulates immune responses at the interface between the commensal microbiota and the host. CD1d presents self and microbial lipid antigens to natural killer T (NKT) cells, which are involved in the pathogenesis of colitis in animal models and human inflammatory bowel disease. As CD1d crosslinking on model IECs results in the production of the important regulatory cytokine interleukin (IL)-10 (ref. 9), decreased epithelial CD1d expression--as observed in inflammatory bowel disease--may contribute substantially to intestinal inflammation. Here we show in mice that whereas bone-marrow-derived CD1d signals contribute to NKT-cell-mediated intestinal inflammation, engagement of epithelial CD1d elicits protective effects through the activation of STAT3 and STAT3-dependent transcription of IL-10, heat shock protein 110 (HSP110; also known as HSP105), and CD1d itself. All of these epithelial elements are critically involved in controlling CD1d-mediated intestinal inflammation. This is demonstrated by severe NKT-cell-mediated colitis upon IEC-specific deletion of IL-10, CD1d, and its critical regulator microsomal triglyceride transfer protein (MTP), as well as deletion of HSP110 in the radioresistant compartment. Our studies thus uncover a novel pathway of IEC-dependent regulation of mucosal homeostasis and highlight a critical role of IL-10 in the intestinal epithelium, with broad implications for diseases such as inflammatory bowel disease. PMID:24717441

  12. A role for mucosal IL-22 production and Th22 cells in HIV-associated mucosal immunopathogenesis.

    PubMed

    Kim, C J; Nazli, A; Rojas, O L; Chege, D; Alidina, Z; Huibner, S; Mujib, S; Benko, E; Kovacs, C; Shin, L Y Y; Grin, A; Kandel, G; Loutfy, M; Ostrowski, M; Gommerman, J L; Kaushic, C; Kaul, R

    2012-11-01

    Interleukin-22 (IL-22) is a cytokine with epithelial reparative and regenerative properties that is produced by Th22 cells and by other immune cell subsets. Therefore, we explored the hypothesis that disruption of the gut barrier during HIV infection involves dysregulation of these cells in the gastrointestinal mucosa. Sigmoid IL-22-producing T cell and Th22 cells were dramatically depleted during chronic HIV infection, epithelial integrity was compromised, and microbial translocation was increased. These alterations were reversed after long-term antiretroviral therapy. While all mucosal IL-22-producing T-cell subsets were also depleted very early during HIV infection, at these early stages IL-22 production by non-T-cell populations (including NKp44+ cells) was increased and gut epithelial integrity was maintained. Circulating Th22 cells expressed a higher level of the HIV co-receptor/binding molecules CCR5 and α4β7 than CD4+ T-cell subsets in HIV-uninfected participants, but this was not the case after HIV infection. Finally, recombinant IL-22 was protective against HIV and tumor necrosis factor-α-induced gut epithelial damage in a validated in vitro gut epithelial system. We conclude that reduced IL-22 production and Th22 depletion in the gut mucosa are important factors in HIV mucosal immunopathogenesis. PMID:22854709

  13. Epithelial crosstalk at the microbiota-mucosal interface.

    PubMed

    Wells, Jerry M; Rossi, Oriana; Meijerink, Marjolein; van Baarlen, Peter

    2011-03-15

    This article provides an overview of how intestinal epithelial cells (IEC) recognize commensals and how they maintain host-bacterial symbiosis. Endocrine, goblet cells, and enterocytes of the intestinal epithelium express a range of pattern recognition receptors (PRR) to sense the presence of microbes. The best characterized are the Toll-like receptors (TLR) and nucleotide oligomerization domain-like receptors (NLR), which play a key role in pathogen recognition and the induction of innate effectors and inflammation. Several adaptations of PRR signaling have evolved in the gut to avoid uncontrolled and potentially destructive inflammatory responses toward the resident microbiota. PRR signaling in IEC serve to maintain the barrier functions of the epithelium, including the production of secretory IgA (sIgA). Additionally, IECs play a cardinal role in setting the immunosuppressive tone of the mucosa to inhibit overreaction against innocuous luminal antigens. This includes regulation of dendritic cells (DC), macrophage and lymphocyte functions by epithelial secreted cytokines. These immune mechanisms depend heavily on IEC recognition of microbes and are consistent with several studies in knockout mice that demonstrate TLR signaling in the epithelium has a profoundly beneficial role in maintaining homeostasis. PMID:20826446

  14. Dendritic cell-targeting DNA-based mucosal adjuvants for the development of mucosal vaccines

    PubMed Central

    Kataoka, Kosuke; Fujihashi, Kohtaro

    2009-01-01

    In order to establish effective mucosal immunity against various mucosal pathogens, vaccines must be delivered via the mucosal route and contain effective adjuvant(s). Since mucosal adjuvants can simply mix with the antigen, it is relatively easy to adapt them for different types of vaccine development. Even in simple admixture vaccines, the adjuvant itself must be prepared without any complications. Thus, CpG oligodeoxynucleotides or plasmids encoding certain cDNA(s) would be potent mucosal adjuvant candidates when compared with other substances that can be used as mucosal adjuvants. The strategy of a DNA-based mucosal adjuvant facilitates the targeting of mucosal dendritic cells, and thus is an effective and safe approach. It would also provide great flexibility for the development of effective vaccines for various mucosal pathogens. PMID:19722892

  15. Epithelial Cell Adhesion Molecule

    PubMed Central

    Trzpis, Monika; McLaughlin, Pamela M.J.; de Leij, Lou M.F.H.; Harmsen, Martin C.

    2007-01-01

    The epithelial cell adhesion molecule (EpCAM, CD326) is a glycoprotein of ∼40 kd that was originally identified as a marker for carcinoma, attributable to its high expression on rapidly proliferating tumors of epithelial origin. Normal epithelia express EpCAM at a variable but generally lower level than carcinoma cells. In early studies, EpCAM was proposed to be a cell-cell adhesion molecule. However, recent insights revealed a more versatile role for EpCAM that is not limited only to cell adhesion but includes diverse processes such as signaling, cell migration, proliferation, and differentiation. Cell surface expression of EpCAM may actually prevent cell-cell adhesion. Here, we provide a comprehensive review of the current knowledge on EpCAM biology in relation to other cell adhesion molecules. We discuss the implications of the newly identified functions of EpCAM in view of its prognostic relevance in carcinoma, inflammatory pathophysiology, and tissue development and regeneration as well as its role in normal epithelial homeostasis. PMID:17600130

  16. Candida albicans VPS4 contributes differentially to epithelial and mucosal pathogenesis

    PubMed Central

    Rane, Hallie S; Hardison, Sarah; Botelho, Claudia; Bernardo, Stella M; Wormley, Floyd; Lee, Samuel A

    2014-01-01

    We have previously demonstrated that the C. albicans pre-vacuolar protein sorting gene VPS4 is required for extracellular secretion of the secreted aspartyl proteases Sap2p and Saps4–6p. Furthermore, the vps4Δ null mutant has been shown to be markedly hypovirulent in a murine tail vein model of disseminated candidiasis. In these experiments, we sought to further define the role of the pre-vacuolar secretion pathway mediated by the pre-vacuolar sorting gene VPS4 in the pathogenesis of epithelial and mucosal infection using a broad range of virulence models. The C. albicans vps4Δ mutant demonstrates reduced tolerance of cell wall stresses compared to its isogenic, complemented control strain. In an in vitro oral epithelial model (OEM) of tissue invasion, the vps4Δ mutant caused reduced tissue damage compared to controls. Further, the vps4Δ mutant was defective in macrophage killing in vitro, and was attenuated in virulence in an in vivo Caenorhabditis elegans model representative of intestinal epithelial infection. In contrast, the vps4Δ mutant caused a similar degree of tissue damage in an in vitro uroepithelial model of Candida infection compared with controls. Furthermore, in an in vivo murine model of vaginal candidiasis there was no reduction in fungal colony burden and no differences in vaginal histopathology compared to wild-type and complemented controls. These results suggest that VPS4 contributes to several key aspects of oral epithelial but not uroepithelial infection, and in contrast to systemic infection, plays no major role in the pathogenesis of Candida vaginitis. By using a wide range of virulence models, we demonstrate that C. albicans VPS4 contributes to virulence according to the specific tissue that is infected. Thus, in order to gain a full understanding of C. albicans virulence in relation to a particular gene or pathway of interest, a selected range of infection models may need to be utilized. PMID:25483774

  17. Long-term result of autologous cultivated oral mucosal epithelial transplantation for severe ocular surface disease.

    PubMed

    Prabhasawat, Pinnita; Ekpo, Pattama; Uiprasertkul, Mongkol; Chotikavanich, Suksri; Tesavibul, Nattaporn; Pornpanich, Kanograt; Luemsamran, Panitee

    2016-09-01

    The present study aimed to investigate the clinical outcomes of autologous cultivated oral mucosal epithelial transplantation (COMET) on human amniotic membrane (AM) for corneal limbal stem cell deficiency (LSCD). In this prospective, noncomparative case series, 20 eyes (18 patients) with bilateral severe ocular surface disease were chosen to undergo COMET on human AM. The primary outcome was clinical success, and the secondary outcomes were the best-corrected visual acuity difference, corneal opacification, symblepharon formation, and complications. The mean patient age was 48.2 ± 15.5 years. The mean follow-up time was 31.9 ± 12.1 months (range 8-50 months). All except one eye exhibited complete epithelialization within the first postoperative week. A successful clinical outcome, defined as a stable ocular surface without epithelial defects, a clear cornea without fibrovascular tissue invasion at the pupillary area, and no or mild ocular surface inflammation, was obtained in 15 of 20 eyes (75 %). The clinical success rate at 1 year was 79.3 %, and that at 4 years (end of follow-up) was 70.5 %. Fourteen of 20 (70 %) eyes exhibited improvement in visual acuity after COMET, and some required subsequent cataract surgery (2 eyes), penetrating keratoplasty (3 eyes), or keratoprosthesis implantation (1 eye). Preoperative symblepharon was eliminated in most eyes (8 of 13, 61.5 %) after COMET combined with eyelid reconstruction when needed. The only complication was corneal perforation (1 eye) induced by a severe eyelid abnormality; treatment with a tectonic corneal graft was successful. COMET can successfully restore ocular surface damage in most eyes with corneal LSCD. PMID:27507558

  18. Epithelial stem cells.

    PubMed

    Draheim, Kyle M; Lyle, Stephen

    2011-01-01

    It is likely that adult epithelial stem cells will be useful in the treatment of diseases, such as ectodermal dysplasias, monilethrix, Netherton syndrome, Menkes disease, hereditary epidermolysis bullosa, and alopecias. Additionally, other skin problems such as burn wounds, chronic wounds, and ulcers will benefit from stem cell-related therapies. However, there are many questions that need to be answered before this goal can be realized. The most important of these questions is what regulates the adhesion of stem cells to the niche versus migration to the site of injury. We have started to identify the mechanisms involved in this decision-making process. PMID:21618097

  19. Chemokines and chemokine receptors in mucosal homeostasis at the intestinal epithelial barrier in inflammatory bowel disease.

    PubMed

    Zimmerman, Noah P; Vongsa, Rebecca A; Wendt, Michael K; Dwinell, Michael B

    2008-07-01

    Chemokines, a large family of small chemoattractive cytokines, and their receptors play an integral role in the regulation of the immune response and homeostasis. The ability of chemokines to attract specific populations of immune cells sets them apart from other chemoattractants. Chemokines produced within the gastrointestinal mucosa are critical players in directing the balance between physiological and pathophysiological inflammation in health, inflammatory bowel disease (IBD), and the progression to colon cancer. In addition to the well-characterized role of chemokines in directed trafficking of immune cells to the gut mucosa, the expression of chemokine receptors on the cells of the epithelium makes them active participants in the chemokine signaling network. Recent findings demonstrate an important role for chemokines and chemokine receptors in epithelial barrier repair and maintenance as well as an intricate involvement in limiting metastasis of colonic carcinoma. Increased recognition of the association between barrier defects and inflammation and the subsequent progression to cancer in IBD thus implicates chemokines as key regulators of mucosal homeostasis and disease pathogenesis. PMID:18452220

  20. Plasma Cell Mucositis of Oro- and Hypopharynx: A Case Report

    PubMed Central

    Puvanendran, Mark; Lieder, Anja; Issing, Wolfgang

    2012-01-01

    Objective. To raise awareness of plasma cell mucositis as a rare differential diagnosis for oral mucosal ulceration and its macroscopic similarity to malignancy. Method. We report a patient who presented with oral features suggestive of malignancy. A biopsy revealed plasma cell mucositis. Results. The patient successfully had a full excision of one lesion and a spontaneous resolution of the other. Conclusion. With the increasing incidence of oral mucosal pathology, physicians should be aware of this differential diagnosis. PMID:22953106

  1. Intestinal Epithelial Barrier Disruption through Altered Mucosal MicroRNA Expression in Human Immunodeficiency Virus and Simian Immunodeficiency Virus Infections

    PubMed Central

    Gaulke, Christopher A.; Porter, Matthew; Han, Yan-Hong; Sankaran-Walters, Sumathi; Grishina, Irina; George, Michael D.; Dang, Angeline T.; Ding, Shou-Wei; Jiang, Guochun; Korf, Ian

    2014-01-01

    ABSTRACT Epithelial barrier dysfunction during human immunodeficiency virus (HIV) infection has largely been attributed to the rapid and severe depletion of CD4+ T cells in the gastrointestinal (GI) tract. Although it is known that changes in mucosal gene expression contribute to intestinal enteropathy, the role of small noncoding RNAs, specifically microRNA (miRNA), has not been investigated. Using the simian immunodeficiency virus (SIV)-infected nonhuman primate model of HIV pathogenesis, we investigated the effect of viral infection on miRNA expression in intestinal mucosa. SIV infection led to a striking decrease in the expression of mucosal miRNA compared to that in uninfected controls. This decrease coincided with an increase in 5′-3′-exoribonuclease 2 protein and alterations in DICER1 and Argonaute 2 expression. Targets of depleted miRNA belonged to molecular pathways involved in epithelial proliferation, differentiation, and immune response. Decreased expression of several miRNA involved in maintaining epithelial homeostasis in the gut was localized to the proliferative crypt region of the intestinal epithelium. Our findings suggest that SIV-induced decreased expression of miRNA involved in epithelial homeostasis, disrupted expression of miRNA biogenesis machinery, and increased expression of XRN2 are involved in the development of epithelial barrier dysfunction and gastroenteropathy. IMPORTANCE MicroRNA (miRNA) regulate the development and function of intestinal epithelial cells, and many viruses disrupt normal host miRNA expression. In this study, we demonstrate that SIV and HIV disrupt expression of miRNA in the small intestine during infection. The depletion of several key miRNA is localized to the proliferative crypt region of the gut epithelium. These miRNA are known to control expression of genes involved in inflammation, cell death, and epithelial maturation. Our data indicate that this disruption might be caused by altered expression of mi

  2. Regulation of local immunity by airway epithelial cells.

    PubMed

    Mayer, Anja K; Dalpke, Alexander H

    2007-01-01

    Epithelial cells are the first line of defense against invading microbial pathogens. They are important contributors to innate mucosal immunity and generate various and sophisticated anti-microbial defense mechanisms, including the formation of a tight barrier and secretion of anti-microbial substances as well as inflammatory mediators. To provide these active defense mechanisms, epithelial cells functionally express various pattern-recognition receptors. Toll-like receptors have been shown to recognize conserved microbial patterns mediating inducible activation of innate immunity. Mucosal surfaces, however, are prone to contact with pathogenic as well as non-pathogenic microbes and, therefore, immune-recognition principles have to be strictly regulated to avoid uncontrolled permanent activation. This review will focus on mechanisms by which epithelial cells regulate mucosal immune responses, thus creating an organ-specific microenvironment. This includes local adaptations in microbial recognition, regulation of local immune homeostasis, and modulation of antigen-presenting cells and adaptive immune responses. These regulatory mechanisms serve the special needs of controlled microbial recognition in mucosal compartments. PMID:18060372

  3. Roles of M cells in infection and mucosal vaccines

    PubMed Central

    Wang, Miao; Gao, Zeqian; Zhang, Zhongwang; Pan, Li; Zhang, Yongguang

    2014-01-01

    The mucosal immune system plays a crucial part in the control of infection. Exposure of humans and animals to potential pathogens generally occurs through mucosal surfaces, thus, strategies that target the mucosa seem rational and efficient vaccination measures. Vaccination through the mucosal immune system can induce effective systemic immune responses simultaneously with mucosal immunity compared with parenteral vaccination. M cells are capable of transporting luminal antigens to the underlying lymphoid tissues and can be exploited by pathogens as an entry portal to invade the host. Therefore, targeting M-cell-specific molecules might enhance antigen entry, initiate the immune response, and induce protection against mucosal pathogens. Here, we outline our understanding of the distribution and function of M cells, and summarize the advances in mucosal vaccine strategies that target M cells. PMID:25483705

  4. Intestinal immune homeostasis is regulated by the crosstalk between epithelial cells and dendritic cells.

    PubMed

    Rimoldi, Monica; Chieppa, Marcello; Salucci, Valentina; Avogadri, Francesca; Sonzogni, Angelica; Sampietro, Gianluca M; Nespoli, Angelo; Viale, Giuseppe; Allavena, Paola; Rescigno, Maria

    2005-05-01

    The control of damaging inflammation by the mucosal immune system in response to commensal and harmful ingested bacteria is unknown. Here we show epithelial cells conditioned mucosal dendritic cells through the constitutive release of thymic stromal lymphopoietin and other mediators, resulting in the induction of 'noninflammatory' dendritic cells. Epithelial cell-conditioned dendritic cells released interleukins 10 and 6 but not interleukin 12, and they promoted the polarization of T cells toward a 'classical' noninflammatory T helper type 2 response, even after exposure to a T helper type 1-inducing pathogen. This control of immune responses seemed to be lost in patients with Crohn disease. Thus, the intimate interplay between intestinal epithelial cells and dendritic cells may help to maintain gut immune homeostasis. PMID:15821737

  5. Cytokine Production and Antigen Recognition by Human Mucosal Homing Conjunctival Effector Memory CD8+ T Cells

    PubMed Central

    Williams, Geraint P.; Pachnio, Annette; Long, Heather M.; Rauz, Saaeha; Curnow, S. John

    2014-01-01

    Purpose. Conjunctival epithelial T cells are dominated by CD3+CD56-TCRαβ+CD8αβ+ lymphocytes. In this study we explored the antigen experience status, mucosal homing phenotype, cytokine expression, and viral antigen recognition of conjunctival epithelial CD8+ T cells from healthy individuals. Methods. Following ocular surface impression cytology, conjunctival cells were recovered by gentle agitation and analyzed by flow cytometry for cell surface markers, cytokine production (stimulated by phorbol 12-myristate 13-acetate [PMA]/ionomycin), and Epstein-Barr virus (EBV)/cytomegalovirus (CMV) immunodominant epitope recognition using major histocompatibility complex (MHC) class I peptide tetramers. Results. In contrast to peripheral blood, conjunctival epithelial CD8+ T cells were dominantly CD45RA−CCR7− effector memory cells, and the vast majority expressed the mucosal homing integrin αEβ7. Conjunctival memory CD8+ T cells maintained effector functions with the ability to secrete IFN-γ and expression of Granzyme B, although they expressed significantly reduced amounts per cell compared to peripheral blood T cells. Interestingly, herpetic virus-specific CD8+ T cells recognizing epitopes derived from EBV and CMV could be detected in the conjunctival cells of healthy virus carriers, although they were generally at lower frequencies than in the peripheral blood of the same donor. Virus-specific conjunctival CD8+ T cells were dominated by CD45RA−CCR7− effector memory cells that expressed αEβ7. Conclusions. These data demonstrate that the majority of conjunctival epithelial CD8+ T cells are mucosal homing αEβ7+ effector memory T cells, which can recognize viral epitopes and are capable of secreting Granzyme B and IFN-γ. PMID:25395484

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

    PubMed Central

    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. PMID:27088599

  7. Canine oral mucosal mast cell tumours.

    PubMed

    Elliott, J W; Cripps, P; Blackwood, L; Berlato, D; Murphy, S; Grant, I A

    2016-03-01

    Mast cell tumours (MCTs) are the most common cutaneous tumours of dogs, however rarely they can arise from the oral mucosa. This subset of MCT is reported to demonstrate a more aggressive clinical course than those tumours on the haired skin and the authors hypothesised that dogs with oral, mucosal MCT would have a high incidence of local lymph node metastasis at presentation and that this would be a negative prognostic factor. An additional hypothesis was that mitotic index (MI) would be prognostic. This retrospective study examines 33 dogs with MCTs arising from the oral mucosa. The results suggest that oral mucosal MCTs in the dog have a high incidence of lymph node metastasis at diagnosis (55%) which results in a poor prognosis. MI and nodal metastasis is highly prognostic. Loco-regional progression is common in these patients and dogs with adequate local control of their tumour had an improved outcome. Despite a more aggressive clinical course, treatment can result in protracted survivals, even when metastasis is present. PMID:24215587

  8. Integrins and epithelial cell polarity

    PubMed Central

    Lee, Jessica L.; Streuli, Charles H.

    2014-01-01

    ABSTRACT 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. For further reading, please see related articles: ‘ERM proteins at a glance’ by Andrea McClatchey (J. Cell Sci. 127, 3199–3204). ‘Establishment of epithelial polarity – GEF who's minding the GAP?’ by Siu Ngok et al. (J. Cell Sci. 127, 3205–3215). PMID:24994933

  9. Potential effector and immunoregulatory functions of mast cells in mucosal immunity

    PubMed Central

    Reber, Laurent L; Sibilano, Riccardo; Mukai, Kaori; Galli, Stephen J

    2016-01-01

    Mast cells (MCs) are cells of hematopoietic origin that normally reside in mucosal tissues, often near epithelial cells, glands, smooth muscle cells, and nerves. Best known for their contributions to pathology during IgE-associated disorders such as food allergy, asthma, and anaphylaxis, MCs are also thought to mediate IgE-associated effector functions during certain parasite infections. However, various MC populations also can be activated to express functional programs – such as secreting pre-formed and/or newly synthesized biologically active products – in response to encounters with products derived from diverse pathogens, other host cells (including leukocytes and structural cells), damaged tissue, or the activation of the complement or coagulation systems, as well as by signals derived from the external environment (including animal toxins, plant products, and physical agents). In this review, we will discuss evidence suggesting that MCs can perform diverse effector and immunoregulatory roles that contribute to homeostasis or pathology in mucosal tissues. PMID:25669149

  10. Arhgap17, a RhoGTPase activating protein, regulates mucosal and epithelial barrier function in the mouse colon

    PubMed Central

    Lee, So-young; Kim, Hwain; Kim, Kyoungmi; Lee, Hyunji; Lee, Seungbok; Lee, Daekee

    2016-01-01

    Coordinated regulation of the actin cytoskeleton by the Rho GTPase family is required for the maintenance of polarity in epithelial cells as well as for their proliferation and migration. A RhoGTPase-activating protein 17 (Arhgap17) is known to be involved in multiple cellular processes in vitro, including the maintenance of tight junctions and vesicle trafficking. However, the function of Arhgap17 has not been studied in the physiological context. Here, we generated Arhgap17-deficient mice and examined the effect in the epithelial and mucosal barriers of the intestine. Reporter staining revealed that Arhgap17 expression is limited to the luminal epithelium of intestine. Arhgap17-deficient mice show an increased paracellular permeability and aberrant localization of the apical junction complex in the luminal epithelium, but do not develop spontaneous colitis. The inner mucus layer is impervious to the enteric bacteria irrespective of Tff3 downregulation in the Arhgap17-deficient mice. Interestingly however, treatment with dextran sulfate sodium (DSS) causes an increased accumulation of DSS and TNF production in intraluminal cells and rapid destruction of the inner mucus layer, resulting in increased severity of colitis in mutant mice. Overall, these data reveal that Arhgap17 has a novel function in regulating transcellular transport and maintaining integrity of intestinal barriers. PMID:27229483

  11. Human cytomegalovirus tropism for mucosal myeloid dendritic cells

    PubMed Central

    Hertel, Laura

    2014-01-01

    SUMMARY Human CMV infections are a serious source of morbidity and mortality for immunocompromised patients and for the developing fetus. Because of this, the development of new strategies to prevent CMV acquisition and transmission is a top priority. Myeloid dendritic cells (DC) residing in the oral and nasal mucosae are among the first immune cells to encounter CMV during entry, and greatly contribute to virus dissemination, reactivation from latency, and horizontal spread. Albeit affected by the immunoevasive tactics of CMV, mucosal DC remain potent inducers of cellular and humoral immune responses against this virus. Their natural functions could thus be exploited to generate long-lasting protective immunity against CMV by vaccination via the oro-nasal mucosae. Although related, epithelial Langerhans-type DC (LC) and dermal monocyte-derived DC (MDDC) interact with CMV in dramatically different ways. While immature MDDC are fully permissive to infection, for instance, immature LC are completely resistant. Understanding these differences is essential to design innovative vaccines and new antiviral compounds to protect these cells from CMV infection in vivo. PMID:24888709

  12. Interactions between bacteria and the intestinal mucosa: Do enteric neurotransmitters acting on epithelium cells influence mucosal colonization or infection?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mechanisms governing the ability of bacteria to adhere to and colonize human and animal hosts in health and disease are still incompletely understood. Throughout the extensive mucosal surfaces of the body that are in contact with the external environment, epithelial cells represent the first po...

  13. Response of corneal epithelial cells to Staphylococcus aureus

    PubMed Central

    2010-01-01

    Staphylococcus aureus is a leading cause of invasive infection. It also infects wet mucosal tissues including the cornea and conjunctiva. Conflicting evidence exists on the expression of Toll-like receptors by human corneal epithelial cells. It was therefore of interest to determine how epithelial cells from this immune privileged tissue respond to S. aureus. Further, it was of interest to determine whether cytolytic toxins, with the potential to cause ion flux or potentially permit effector molecule movement across the target cell membrane, alter the response. Microarrays were used to globally assess the response of human corneal epithelial cells to S. aureus. A large increase in abundance of transcripts encoding the antimicrobial dendritic cell chemokine, CCL20, was observed. CCL20 release into the medium was detected, and this response was found to be largely TLR2 and NOD2 independent. Corneal epithelial cells also respond to S. aureus by increasing the intracellular abundance of mRNA for inflammatory mediators, transcription factors, and genes related to MAP kinase pathways, in ways similar to other cell types. The corneal epithelial cell response was surprisingly unaffected by toxin exposure. Toxin exposure did, however, induce a stress response. Although model toxigenic and non-toxigenic strains of S. aureus were employed in the present study, the results obtained were strikingly similar to those reported for stimulation of vaginal epithelial cells by clinical toxic shock toxin expressing isolates, demonstrating that the initial epithelial cellular responses to S. aureus are largely independent of strain as well as epithelial cell tissue source. PMID:21178447

  14. A marvel of mucosal T cells and secretory antibodies for the creation of first lines of defense.

    PubMed

    Kunisawa, J; Kiyono, H

    2005-06-01

    The mucosal immune system acts as a first line of defense against bacterial and viral infections while also playing a crucial role in the establishment and maintenance of mucosal homeostasis between the host and the outside environment. In addition to epithelial cells and antigen-presenting cells (dendritic cells and macrophages), B and T lymphocytes form a dynamic mucosal network for the induction and regulation of secretory IgA (S-IgA) and cytotoxic T lymphocyte (CTL) responses. This review seeks to shed light on the pathways of induction and regulation of these responses and to elucidate the role they simultaneously play in fending off pathogen invasion and maintaining mucosal homeostasis. PMID:15971106

  15. Ion Channels in Epithelial Cells

    NASA Astrophysics Data System (ADS)

    Palmer, Lawrence G.

    Ion channels in epithelial cells serve to move ions, and in some cases fluid, between compartments of the body. This function of the transfer of material is fundamentally different from that of the transfer of information, which is the main job of most channels in excitable cells. Nevertheless the basic construction of the channels is similar in many respects in the two tissue types. This chapter reviews the nature of channels in epithelia and discusses how their functions have evolved to accomplish the basic tasks for which they are responsible. I will focus on three channel types: epithelial Na+ channels, inward-rectifier K+ channels, and CFTR Cl- channels.

  16. Gastric Mucosal Protection by Aegle Marmelos Against Gastric Mucosal Damage: Role of Enterochromaffin Cell and Serotonin

    PubMed Central

    Singh, Purnima; Dutta, Shubha R.; Guha, Debjani

    2015-01-01

    Background/Aims: Serotonin (5-hydroxytryptamine; 5-HT) released from enterochromaffin (EC) cells in gastric mucosa inhibits gastric acidity by increasing the gastric mucus secretion. In the present study, we evaluated the effect of aqueous extract of Aegle marmelos (AM) ripe fruit pulp (250 mg/kg body weight) on mean ulcer index (MUI), EC cells, 5-HT content, and adherent mucosal thickness of ulcerated gastric tissue in adult albino rats. Material and Methods: Ulceration was induced by using aspirin (500 mg/kg, p.o.), cerebellar nodular lesion and applying cold-restraint stress. Results: In all cases increased MUI in gastric tissue along with decreased EC cell count was observed with concomitant decrease of 5-HT content and adherent mucosal thickness (P < 0.05). Pretreatment with AM for 14 days decreased MUI, increased EC cell count, and 5-HT content as well as adherent mucosal thickness in all ulcerated group (P < 0.05). Conclusion: AM produces gastric mucosal protection mediated by increased EC cell count and 5-HT levels. PMID:25672237

  17. Galectin-4 Controls Intestinal Inflammation by Selective Regulation of Peripheral and Mucosal T Cell Apoptosis and Cell Cycle

    PubMed Central

    Paclik, Daniela; Danese, Silvio; Berndt, Uta; Wiedenmann, Bertram; Dignass, Axel; Sturm, Andreas

    2008-01-01

    Galectin-4 is a carbohydrate-binding protein belonging to the galectin family. Here we provide novel evidence that galectin-4 is selectively expressed and secreted by intestinal epithelial cells and binds potently to activated peripheral and mucosal lamina propria T-cells at the CD3 epitope. The carbohydrate-dependent binding of galectin-4 at the CD3 epitope is fully functional and inhibited T cell activation, cycling and expansion. Galectin-4 induced apoptosis of activated peripheral and mucosal lamina propria T cells via calpain-, but not caspase-dependent, pathways. Providing further evidence for its important role in regulating T cell function, galectin-4 blockade by antisense oligonucleotides reduced TNF-alpha inhibitor induced T cell death. Furthermore, in T cells, galectin-4 reduced pro-inflammatory cytokine secretion including IL-17. In a model of experimental colitis, galectin-4 ameliorated mucosal inflammation, induced apoptosis of mucosal T-cells and decreased the secretion of pro-inflammatory cytokines. Our results show that galectin-4 plays a unique role in the intestine and assign a novel role of this protein in controlling intestinal inflammation by a selective induction of T cell apoptosis and cell cycle restriction. Conclusively, after defining its biological role, we propose Galectin-4 is a novel anti-inflammatory agent that could be therapeutically effective in diseases with a disturbed T cell expansion and apoptosis such as inflammatory bowel disease. PMID:18612433

  18. Application of direct oral microscopy in evaluating mucosal margins around invasive oral squamous cell carcinoma

    PubMed Central

    Michcik, Adam; Michajłowski, Igor; Starzyńska, Anna

    2015-01-01

    Introduction Direct oral microscopy constitutes a novel, non-invasive diagnostic technique, which aids clinical examination of the oral cavity. The oral mucosa is examined at multiple magnifications and features such as sub-epithelial mucosal vessels, surface patterns, colour tone, transparency and the exact demarcation of mucosal lesions are estimated. The incidence of oral squamous cell carcinoma (OSCC) oscillates between 1.9% and 3.5%, which makes it the eighth most common carcinoma occurring around the world and in Poland. The 5-year survival rates oscillate between 20% and 30%. Aim The aim of the study was to evaluate clinically unchanged mucosal margins around OSCC by direct oral microscopy. The authors approached the question whether the borders of mucosal margins around OSCC established via direct oral microscopy differ from those established based on clinical examination. Material and methods Fifteen patients diagnosed with OSCC were enrolled. Patients were first clinically examined to evaluate the extent of the tumour and to plan resection margins. Eventually, direct oral microscopy was performed to establish the width of the subclinically unchanged mucosal margins based on a standard picture of healthy oral mucosae, followed by comparison with those established by clinical evaluation. Results Histopathologic results of biopsies from areas indicated by direct oral microscopy revealed dysplasia in 86.7% of patients, whereas biopsies from areas indicated by clinical examination revealed dysplasia only in 40% of individuals, resulting in the need for widening of mucosal margins. Conclusions Direct oral microscopy enables detection of dysplasia within clinically unaltered mucosal margins around OSCC, which results in more precise establishing of resection boundaries, contributing to improvement of resection totality. PMID:26759543

  19. Urokinase and the intestinal mucosa: evidence for a role in epithelial cell turnover

    PubMed Central

    Gibson, P; Birchall, I; Rosella, O; Albert, V; Finch, C; Barkla, D; Young, G

    1998-01-01

    Background—The functions of urokinase in intestinal epithelia are unknown. 
Aims—To determine the relation of urokinase expressed by intestinal epithelial cells to their position in the crypt-villus/surface axis and of mucosal urokinase activity to epithelial proliferative kinetics in the distal colon. 
Methods—Urokinase expression was examined immunohistochemically in human intestinal mucosa. Urokinase activity was measured colorimetrically in epithelial cells isolated sequentially from the crypt-villus axis of the rat small intestine. In separate experiments, urokinase activity and epithelial kinetics (measured stathmokinetically) were measured in homogenates of distal colonic mucosa of 14 groups of eight rats fed diets known to alter epithelial turnover. 
Results—From the crypt base, an ascending gradient of expression and activity of urokinase was associated with the epithelial cells. Median mucosal urokinase activities in each of the dietary groups of rats correlated positively with autologous median number of metaphase arrests per crypt (r=0.68; p<0.005) and per 100 crypt cells (r=0.75; p<0.001), but not with crypt column height. 
Conclusions—Localisation of an enzyme capable of leading to digestion of cell substratum in the region where cells are loosely attached to their basement membrane, and the association of its activity with indexes of cell turnover, suggest a role for urokinase in facilitating epithelial cell loss in the intestine. 

 Keywords: urokinase; intestinal epithelium; colon; epithelial proliferation PMID:9824347

  20. Mucosal Imprinting of Vaccine-Induced CD8+ T Cells Is Crucial to Inhibit the Growth of Mucosal Tumors

    PubMed Central

    Sandoval, Federico; Bureau, Michel-Francis; Freyburger, Ludovic; Clement, Olivier; Marcheteau, Elie; Gey, Alain; Fraisse, Guillaume; Bouguin, Cécilia; Merillon, Nathalie; Dransart, Estelle; Tran, Thi; Quintin-Colonna, Françoise; Autret, Gwennhael; Thiebaud, Marine; Suleman, Muhammad; Riffault, Sabine; Wu, Tzyy-Choou; Launay, Odile; Danel, Claire; Taieb, Julien; Richardson, Jennifer; Zitvogel, Laurence; Fridman, Wolf H.; Johannes, Ludger; Tartour, Eric

    2014-01-01

    Although many human cancers are located in mucosal sites, most cancer vaccines are tested against subcutaneous tumors in preclinical models. We therefore wondered whether mucosa-specific homing instructions to the immune system might influence mucosal tumor outgrowth. We showed that the growth of orthotopic head and neck or lung cancers was inhibited when a cancer vaccine was delivered by the intranasal mucosal route but not the intramuscular route. This antitumor effect was dependent on CD8+ T cells. Indeed, only intranasal vaccination elicited mucosal-specific CD8+ T cells expressing the mucosal integrin CD49a. Blockade of CD49a decreased intratumoral CD8+ T cell infiltration and the efficacy of cancer vaccine on mucosal tumor. We then showed that after intranasal vaccination, dendritic cells from lung parenchyma, but not those from spleen, induced the expression of CD49a on cocultured specific CD8+ T cells. Tumor-infiltrating lymphocytes from human mucosal lung cancer also expressed CD49a, which supports the relevance and possible extrapolation of these results in humans. We thus identified a link between the route of vaccination and the induction of a mucosal homing program on induced CD8+ T cells that controlled their trafficking. Immunization route directly affected the efficacy of the cancer vaccine to control mucosal tumors. PMID:23408053

  1. Induction of apoptosis in oral epithelial cells by Candida albicans.

    PubMed

    Villar, C Cunha; Chukwuedum Aniemeke, J; Zhao, X-R; Huynh-Ba, G

    2012-12-01

    During infection, interactions between Candida albicans and oral epithelial cells result in oral epithelial cell death. This is clinically manifested by the development of oral mucosal ulcerations generally associated with discomfort. In vitro studies have shown that C. albicans induces early apoptotic alterations in oral epithelial cells; however, these studies have also shown that treatment of infected cells with caspase inhibitors does not prevent their death. The reasons for these contradictory results are unknown and it is still not clear if C. albicans stimulates oral epithelial signaling pathways that promote apoptotic cell death. Activation of specific death pathways in response to microbial organisms plays an essential role in modulating the pathogenesis of a variety of infectious diseases. The aim of this study was to (i) characterize C. albicans-induced apoptotic morphological alterations in oral epithelial cells, and (ii) investigate the activation of apoptotic signaling pathways and expression of apoptotic genes during infection. Candida albicans induced early apoptotic changes in over 50% of oral epithelial cells. However, only 15% of those showed mid-late apoptotic alterations. At the molecular level, C. albicans caused a loss of the mitochondrial transmembrane potential and translocation of mitochondrial cytochrome c. Caspase-3/9 activities increased only during the first hours of infection. Moreover, poly[ADP ribose] polymerase 1 was cleaved into apoptotic and necrotic-like fragments. Finally, five anti-apoptotic genes were significantly upregulated and two pro-apoptotic genes were downregulated during infection. Altogether, these findings indicate that epithelial apoptotic pathways are activated in response to C. albicans, but fail to progress and promote apoptotic cell death. PMID:23134609

  2. Mast Cells Infiltrating Inflamed or Transformed Gut Alternatively Sustain Mucosal Healing or Tumor Growth.

    PubMed

    Rigoni, Alice; Bongiovanni, Lucia; Burocchi, Alessia; Sangaletti, Sabina; Danelli, Luca; Guarnotta, Carla; Lewis, Amy; Rizzo, Aroldo; Silver, Andrew R; Tripodo, Claudio; Colombo, Mario P

    2015-09-15

    Mast cells (MC) are immune cells located next to the intestinal epithelium with regulatory function in maintaining the homeostasis of the mucosal barrier. We have investigated MC activities in colon inflammation and cancer in mice either wild-type (WT) or MC-deficient (Kit(W-sh)) reconstituted or not with bone marrow-derived MCs. Colitis was chemically induced with dextran sodium sulfate (DSS). Tumors were induced by administering azoxymethane (AOM) intraperitoneally before DSS. Following DSS withdrawal, Kit(W-sh) mice showed reduced weight gain and impaired tissue repair compared with their WT littermates or Kit(W-sh) mice reconstituted with bone marrow-derived MCs. MCs were localized in areas of mucosal healing rather than damaged areas where they degraded IL33, an alarmin released by epithelial cells during tissue damage. Kit(W-sh) mice reconstituted with MC deficient for mouse mast cell protease 4 did not restore normal mucosal healing or reduce efficiently inflammation after DSS withdrawal. In contrast with MCs recruited during inflammation-associated wound healing, MCs adjacent to transformed epithelial cells acquired a protumorigenic profile. In AOM- and DSS-treated WT mice, high MC density correlated with high-grade carcinomas. In similarly treated Kit(W-sh) mice, tumors were less extended and displayed lower histologic grade. Our results indicate that the interaction of MCs with epithelial cells is dependent on the inflammatory stage, and on the activation of the tissue repair program. Selective targeting of MCs for prevention or treatment of inflammation-associated colon cancer should be timely pondered to allow tissue repair at premalignant stages or to reduce aggressiveness at the tumor stage. PMID:26206557

  3. High Concentrate Diet Induced Mucosal Injuries by Enhancing Epithelial Apoptosis and Inflammatory Response in the Hindgut of Goats

    PubMed Central

    Tao, Shiyu; Duanmu, Yongqian; Dong, Haibo; Ni, Yingdong; Chen, Jie; Shen, Xiangzhen; Zhao, Ruqian

    2014-01-01

    Purpose It is widely accepted that lipopolysaccharide and volatile fatty acids (VFA) accumulate in the digestive tract of ruminants fed diets containing high portions of grain. Compared to the ruminal epithelium, the hindgut epithelium is composed of a monolayer structure that is more “leaky” for lipopolysaccharide and susceptible to organic acid-induced damage. The aim of this study was to investigate changes in epithelial structure, apoptosis and inflammatory response in the hindgut of goats fed a high-concentrate diet for 6 weeks. Experimental Design Eight local Chinese goats with rumen cannulas were randomly assigned to two groups: one group was fed a high-concentrate diet (65% concentrate of dry matter, HC) and the other group was fed a low-concentrate diet (35% concentrate of dry matter, LC) for 6 wks. Ruminal fluid, plasma, and hindgut mucosa tissues were collected. Histological techniques, real-time PCR and western blotting were used to evaluate the tissues structure, cell apoptosis and local inflammation in the hindguts. Results Feeding HC diet for 6 wks resulted in a significant decrease of ruminal pH (p<0.01), and ruminal lipopolysaccharide concentrations were significantly increased in HC goats (p<0.05). Obvious damage was observed to mucosal epithelium of the hindgut and the intercellular tight junctions in HC, but not in LC, goats. The expression of MyD88 and caspase-8 mRNA was increased in colonic epithelium of HC goats compared to LC (p<0.05), and the expression of TLR-4 and caspase-3 showed a tendency to increase. In the cecum, interleukin-1β mRNA expression was decreased (p<0.05), and caspase-3 showed a potential increase (p = 0.07) in HC goats. The level of NF-κB protein was increased in colonic epithelium of HC goats. Caspase-3 activity was elevated in both colon and cecum, whereas caspase-8 activity was statistically increased only in colon. Conclusions Feeding a high-concentrate diet to goats for 6 wks led to hindgut mucosal injuries

  4. The yin and yang of intestinal epithelial cells in controlling dendritic cell function

    PubMed Central

    Iliev, Iliyan D.; Matteoli, Gianluca; Rescigno, Maria

    2007-01-01

    Recent work suggests that dendritic cells (DCs) in mucosal tissues are “educated” by intestinal epithelial cells (IECs) to suppress inflammation and promote immunological tolerance. After attack by pathogenic microorganisms, however, “non-educated” DCs are recruited from nearby areas, such as the dome of Peyer's patches (PPs) and the blood, to initiate inflammation and the ensuing immune response to the invader. Differential epithelial cell (EC) responses to commensals and pathogens may control these two tolorogenic and immunogenic functions of DCs. PMID:17893197

  5. Mucosal immunoglobulins and B cells of Teleost fish

    PubMed Central

    Salinas, Irene; Zhang, Yong-An; Sunyer, J. Oriol

    2012-01-01

    As physical barriers that separate teleost fish from the external environment, mucosae are also active immunological sites that protect them against exposure to microbes and stressors. In mammals, the sites where antigens are sampled from mucosal surfaces and where stimulation of naive T and B lymphocytes occurs are known as inductive sites and are constituted by mucosa-associated lymphoid tissue (MALT). According to anatomical location, the MALT in teleost fish is subdivided into gut-associated lymphoid tissue (GALT), skin-associated lymphoid tissue (SALT), and gill-associated lymphoid tissue (GIALT). All MALT contain a variety of leukocytes, including, but not limited to, T cells, B cells, plasma cells, macrophages and granulocytes. Secretory immunoglobulins are produced mainly by plasmablasts and plasma cells, and play key roles in the maintenance of mucosal homeostasis. Until recently, teleost fish B cells were thought to express only two classes of immunoglobulins, IgM and IgD, in which IgM was thought to be the only one responding to pathogens both in systemic and mucosal compartments. However, a third teleost immunoglobulin class, IgT/IgZ, was discovered in 2005, and it has recently been shown to behave as the prevalent immunoglobulin in gut mucosal immune responses. The purpose of this review is to summarise the current knowledge of mucosal immunoglobulins and B cells of fish MALT. Moreover, we attempt to integrate the existing knowledge on both basic and applied research findings on fish mucosal immune responses, with the goal to provide new directions that may facilitate the development of novel vaccination strategies that stimulate not only systemic, but also mucosal immunity. PMID:22133710

  6. Regeneration of Vocal Fold Mucosa Using Tissue-Engineered Structures with Oral Mucosal Cells

    PubMed Central

    Fukahori, Mioko; Chitose, Shun-ichi; Sato, Kiminori; Sueyoshi, Shintaro; Kurita, Takashi; Umeno, Hirohito; Monden, Yu; Yamakawa, Ryoji

    2016-01-01

    Objectives Scarred vocal folds result in irregular vibrations during phonation due to stiffness of the vocal fold mucosa. To date, a completely satisfactory corrective procedure has yet to be achieved. We hypothesize that a potential treatment option for this disease is to replace scarred vocal folds with organotypic mucosa. The purpose of this study is to regenerate vocal fold mucosa using a tissue-engineered structure with autologous oral mucosal cells. Study Design Animal experiment using eight beagles (including three controls). Methods A 3 mm by 3 mm specimen of canine oral mucosa was surgically excised and divided into epithelial and subepithelial tissues. Epithelial cells and fibroblasts were isolated and cultured separately. The proliferated epithelial cells were co-cultured on oriented collagen gels containing the proliferated fibroblasts for an additional two weeks. The organotypic cultured tissues were transplanted to the mucosa-deficient vocal folds. Two months after transplantation, vocal fold vibrations and morphological characteristics were observed. Results A tissue-engineered vocal fold mucosa, consisting of stratified epithelium and lamina propria, was successfully fabricated to closely resemble the normal layered vocal fold mucosa. Laryngeal stroboscopy revealed regular but slightly small mucosal waves at the transplanted site. Immunohistochemically, stratified epithelium expressed cytokeratin, and the distributed cells in the lamina propria expressed vimentin. Elastic Van Gieson staining revealed a decreased number of elastic fibers in the lamina propria of the transplanted site. Conclusion The fabricated mucosa with autologous oral mucosal cells successfully restored the vocal fold mucosa. This reconstruction technique could offer substantial clinical advantages for treating intractable diseases such as scarring of the vocal folds. PMID:26730600

  7. β-Arrestin1 inhibits chemotherapy-induced intestinal stem cell apoptosis and mucositis.

    PubMed

    Zhan, Y; Xu, C; Liu, Z; Yang, Y; Tan, S; Yang, Y; Jiang, J; Liu, H; Chen, J; Wu, B

    2016-01-01

    The mechanism of chemotherapy-induced gastrointestinal (GI) syndrome (CIGIS) is still controversial, and it is unclear whether chemotherapy induces intestinal stem cell (ISC) apoptosis. β-Arrestins are regulators and mediators of G protein-coupled receptor signaling in cell apoptosis, division and growth. In this study, we aimed to investigate whether chemotherapy induces ISC apoptosis to contribute to mucositis in CIGIS and whether β-arrestin1 (β-arr1) is involved in this apoptosis. Different chemotherapeutic agents were used to generate a CIGIS model. Lgr5-EGFP-IRES-creERT2(+/-) knock-in mice were used as a CIGIS model to investigate ISC apoptosis. β-arr1 knockout mice were used to determine whether β-arr1 is involved in the apoptosis in CIGIS. Intestinal histology was performed, the ISC apoptosis was analyzed and the mucosal barrier was examined. The effects of β-arr1 in apoptosis were investigated in the samples from humans and mice as well as in cell lines. Here, we demonstrate that chemotherapy induced intestinal mucositis by promoting crypt cell apoptosis, especially in Lgr5+ stem cells and Paneth cells but not in goblet cells, epithelial cells or vascular endothelial cells. Furthermore, β-arr1 deficiency exacerbated the Lgr5+ stem cell apoptosis, but not Paneth cell apoptosis, in CIGIS. In addition, the data showed that β-arr1 reduced the chemotherapy-induced Lgr5+ stem cell apoptosis by inhibiting endoplasmic reticulum stress-mediated mitochondrial apoptotic signaling. Our study indicates that β-arr1 inhibits chemotherapy-induced ISC apoptosis to alleviate intestinal mucositis in CIGIS. PMID:27195676

  8. β-Arrestin1 inhibits chemotherapy-induced intestinal stem cell apoptosis and mucositis

    PubMed Central

    Zhan, Y; Xu, C; Liu, Z; Yang, Y; Tan, S; Yang, Y; Jiang, J; Liu, H; Chen, J; Wu, B

    2016-01-01

    The mechanism of chemotherapy-induced gastrointestinal (GI) syndrome (CIGIS) is still controversial, and it is unclear whether chemotherapy induces intestinal stem cell (ISC) apoptosis. β-Arrestins are regulators and mediators of G protein-coupled receptor signaling in cell apoptosis, division and growth. In this study, we aimed to investigate whether chemotherapy induces ISC apoptosis to contribute to mucositis in CIGIS and whether β-arrestin1 (β-arr1) is involved in this apoptosis. Different chemotherapeutic agents were used to generate a CIGIS model. Lgr5-EGFP-IRES-creERT2+/− knock-in mice were used as a CIGIS model to investigate ISC apoptosis. β-arr1 knockout mice were used to determine whether β-arr1 is involved in the apoptosis in CIGIS. Intestinal histology was performed, the ISC apoptosis was analyzed and the mucosal barrier was examined. The effects of β-arr1 in apoptosis were investigated in the samples from humans and mice as well as in cell lines. Here, we demonstrate that chemotherapy induced intestinal mucositis by promoting crypt cell apoptosis, especially in Lgr5+ stem cells and Paneth cells but not in goblet cells, epithelial cells or vascular endothelial cells. Furthermore, β-arr1 deficiency exacerbated the Lgr5+ stem cell apoptosis, but not Paneth cell apoptosis, in CIGIS. In addition, the data showed that β-arr1 reduced the chemotherapy-induced Lgr5+ stem cell apoptosis by inhibiting endoplasmic reticulum stress-mediated mitochondrial apoptotic signaling. Our study indicates that β-arr1 inhibits chemotherapy-induced ISC apoptosis to alleviate intestinal mucositis in CIGIS. PMID:27195676

  9. Ethanol stimulation of HIV infection of oral epithelial cells.

    PubMed

    Zheng, Jun; Yang, Otto O; Xie, Yiming; Campbell, Richard; Chen, Irvin S Y; Pang, Shen

    2004-12-01

    Oral mucosal cells can be infected by exogenous HIV during receptive oral sex or breast-feeding. The risk of oral mucosal infection depends on the infection efficiency of the HIV strains present in the oral cavity, the viral titers, and the defense mechanisms in the oral cavity environment. It is expected that alcohol can weaken the host defense mechanism against HIV infection in the oral cavity. We modified an HIV strain, NL4-3, by inserting the enhanced green fluorescent protein gene and used this virus to infect oral epithelial cells obtained from patients. Various concentrations of ethanol (0%-4%) were added to the infected cells. HIV-infected cells were detected by fluorescent microscopy or fluorescence-activated cell sorting. We found that ethanol significantly increases HIV infection of primary oral epithelial cells (POEs). POEs pretreated with 4% ethanol for less than 10 minutes demonstrated 3- to 6-fold higher susceptibility to infection by the CXCR-4 HIV strain NL4-3. Our studies also demonstrated that HIV infects POEs through a gp120-independent mechanism. We tested an HIV CCR5 strain, JRCSF, and also found its infection efficiency to be stimulated by alcohol. Our results indicate that in cell culture conditions, the ranges of concentrations of alcohol that are commercially available are able to stimulate the infection efficiency of HIV in POEs. PMID:15602121

  10. Epithelial Sodium Channels in Pulmonary Epithelial Progenitor and Stem Cells

    PubMed Central

    Liu, Yang; Jiang, Bi-Jie; Zhao, Run-Zhen; Ji, Hong-Long

    2016-01-01

    Regeneration of the epithelium of mammalian lungs is essential for restoring normal function following injury, and various cells and mechanisms contribute to this regeneration and repair. Club cells, bronchioalveolar stem cells (BASCs), and alveolar type II epithelial cells (ATII) are dominant stem/progenitor cells for maintaining epithelial turnover and repair. Epithelial Na+ channels (ENaC), a critical pathway for transapical salt and fluid transport, are expressed in lung epithelial progenitors, including club and ATII cells. Since ENaC activity and expression are development- and differentiation-dependent, apically located ENaC activity has therefore been used as a functional biomarker of lung injury repair. ENaC activity may be involved in the migration and differentiation of local and circulating stem/progenitor cells with diverse functions, eventually benefiting stem cells spreading to re-epithelialize injured lungs. This review summarizes the potential roles of ENaC expressed in native progenitor and stem cells in the development and regeneration of the respiratory epithelium. PMID:27570489

  11. The Cytosolic Microbial Receptor Nod2 Regulates Small Intestinal Crypt Damage and Epithelial Regeneration following T Cell-Induced Enteropathy.

    PubMed

    Zanello, Galliano; Goethel, Ashleigh; Rouquier, Sandrine; Prescott, David; Robertson, Susan J; Maisonneuve, Charles; Streutker, Catherine; Philpott, Dana J; Croitoru, Kenneth

    2016-07-01

    Loss of function in the NOD2 gene is associated with a higher risk of developing Crohn's disease (CD). CD is characterized by activation of T cells and activated T cells are involved in mucosal inflammation and mucosal damage. We found that acute T cell activation with anti-CD3 mAb induced stronger small intestinal mucosal damage in NOD2(-/-) mice compared with wild-type mice. This enhanced mucosal damage was characterized by loss of crypt architecture, increased epithelial cell apoptosis, delayed epithelial regeneration and an accumulation of inflammatory cytokines and Th17 cells in the small intestine. Partial microbiota depletion with antibiotics did not decrease mucosal damage 1 d after anti-CD3 mAb injection, but it significantly reduced crypt damage and inflammatory cytokine secretion in NOD2(-/-) mice 3 d after anti-CD3 mAb injection, indicating that microbial sensing by Nod2 was important to control mucosal damage and epithelial regeneration after anti-CD3 mAb injection. To determine which cells play a key role in microbial sensing and regulation of mucosal damage, we engineered mice carrying a cell-specific deletion of Nod2 in villin and Lyz2-expressing cells. T cell activation did not worsen crypt damage in mice carrying either cell-specific deletion of Nod2 compared with wild-type mice. However, increased numbers of apoptotic epithelial cells and higher expression of TNF-α and IL-22 were observed in mice carrying a deletion of Nod2 in Lyz2-expressing cells. Taken together, our results demonstrate that microbial sensing by Nod2 is an important mechanism to regulate small intestinal mucosal damage following acute T cell activation. PMID:27206769

  12. Interplay between Helicobacter pylori and immune cells in immune pathogenesis of gastric inflammation and mucosal pathology

    PubMed Central

    Tsai, Hwei-Fang; Hsu, Ping-Ning

    2010-01-01

    Helicobacter pylori infection is associated with an inflammatory response in the gastric mucosa, leading to chronic gastritis, peptic ulcers, gastric carcinoma and gastric mucosa-associated lymphoid tissue (MALT) lymphomas. Recent studies have shown that apoptosis of gastric epithelial cells is increased during H. pylori infection. Apoptosis induced by microbial infections are factors implicated in the pathogenesis of H. pylori infection. The enhanced gastric epithelial cell apoptosis in H. pylori infection has been suggested to play an important role in the pathogenesis of chronic gastritis and gastric pathology. In addition to directly triggering apoptosis, H. pylori induces sensitivity to tumor-necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis in gastric epithelial cells via modulation of TRAIL apoptosis signaling. Moreover, H. pylori infection induces infiltration of T lymphocytes and triggers inflammation to augment apoptosis. In H. pylori infection, there was significantly increased CCR6+CD3+ T-cell infiltration in the gastric mucosa, and the CCR6 ligand, CCL20 chemokine, was selectively expressed in inflamed gastric tissues. These results implicate that the interaction between CCL20 and CCR6 may play a role in recruiting T cells to the sites of inflammation in the gastric mucosa during Helicobacter infection. Through these mechanisms, chemokine-mediated T lymphocyte trafficking into inflamed epithelium is initiated and the mucosal injury in Helicobacter infection is induced. This article will review the recent novel findings on the interactions of H. pylori with diverse host epithelial signaling pathways and events involved in the initiation of gastric pathology, including gastric inflammation, mucosal damage and development of MALT lymphomas. PMID:20190789

  13. Epithelial cells and Von Gierke's disease.

    PubMed

    Negishi, H; Benke, P J

    1977-08-01

    Epithelial cells and not fibroblasts from human liver and amniotic fluid contain inducible glucose-6-phosphatase (G-6-Pase) activity. The diagnosis of Von Gierke's disease has been made in a patient with hepatomegaly utilizing cultured epithelial cells grown from a liver biopsy. G-6-Pase activity in epithelial cells from this patient could not be induced by dibutyryl cyclic AMP and theophylline. This is the first use of epithelial cells for diagnosis of a metabolic disease. G-6-Pase activity in cloned epithelial cells from amniotic fluid increases 2- to 3-fold after 24-hr exposure to dibutyryl cyclic AMP and theophylline. The prenatal diagnosis of Von Gierke's disease may be possible in a laboratory experienced with these techniques if epithelial cell growth is obtained from amniotic fluid. PMID:196249

  14. Mycobacteria bypass mucosal NF-kB signalling to induce an epithelial anti-inflammatory IL-22 and IL-10 response.

    PubMed

    Lutay, Nataliya; Håkansson, Gisela; Alaridah, Nader; Hallgren, Oskar; Westergren-Thorsson, Gunilla; Godaly, Gabriela

    2014-01-01

    The mechanisms by which mycobacteria subvert the inflammatory defence to establish chronic infection remain an unresolved question in the pathogenesis of tuberculosis. Using primary epithelial cells, we have analysed mycobacteria induced epithelial signalling pathways from activation of TLRs to cytokine secretion. Mycobacterium bovis bacilli Calmette-Guerin induced phosphorylation of glycogen synthase kinase (GSK)3 by PI3K-Akt in the signalling pathway downstream of TLR2 and TLR4. Mycobacteria did not suppress NF-κB by activating the peroxisome proliferator-activated receptor γ. Instead the pro-inflammatory NF-κB was bypassed by mycobacteria induced GSK3 inhibition that promoted the anti-inflammatory transcription factor CREB. Mycobacterial infection did not thus induce mucosal pro-inflammatory response as measured by TNFα and IFNγ secretion, but led to an anti-inflammatory IL-10 and IL-22 production. Apart from CREB, MAP3Ks p38 and ERK1/2 activated the transcription factor AP-1 leading to IL-6 production. Interestingly, blocking of TLR4 before infection decreased epithelial IL-6 secretion, but increased the CREB-activated IL-10 production. Our data indicate that mycobacteria suppress epithelial pro-inflammatory production by suppressing NF-κB activation thereby shifting the infection towards an anti-inflammatory state. This balance between the host immune response and the pathogen could determine the outcome of infection. PMID:24489729

  15. Mycobacteria Bypass Mucosal NF-kB Signalling to Induce an Epithelial Anti-Inflammatory IL-22 and IL-10 Response

    PubMed Central

    Lutay, Nataliya; Håkansson, Gisela; Alaridah, Nader; Hallgren, Oskar; Westergren-Thorsson, Gunilla; Godaly, Gabriela

    2014-01-01

    The mechanisms by which mycobacteria subvert the inflammatory defence to establish chronic infection remain an unresolved question in the pathogenesis of tuberculosis. Using primary epithelial cells, we have analysed mycobacteria induced epithelial signalling pathways from activation of TLRs to cytokine secretion. Mycobacterium bovis bacilli Calmette-Guerin induced phosphorylation of glycogen synthase kinase (GSK)3 by PI3K–Akt in the signalling pathway downstream of TLR2 and TLR4. Mycobacteria did not supress NF-κB by activating the peroxisome proliferator-activated receptor γ. Instead the pro-inflammatory NF-κB was bypassed by mycobacteria induced GSK3 inhibition that promoted the anti-inflammatory transcription factor CREB. Mycobacterial infection did not thus induce mucosal pro-inflammatory response as measured by TNFα and IFNγ secretion, but led to an anti-inflammatory IL-10 and IL-22 production. Apart from CREB, MAP3Ks p38 and ERK1/2 activated the transcription factor AP-1 leading to IL-6 production. Interestingly, blocking of TLR4 before infection decreased epithelial IL-6 secretion, but increased the CREB-activated IL-10 production. Our data indicate that mycobacteria supress epithelial pro-inflammatory production by supressing NF-κB activation thereby shifting the infection towards an anti-inflammatory state. This balance between the host immune response and the pathogen could determine the outcome of infection. PMID:24489729

  16. Intestinal epithelial cells as mediators of the commensal–host immune crosstalk

    PubMed Central

    Goto, Yoshiyuki; Ivanov, Ivaylo I

    2014-01-01

    Commensal bacteria regulate the homeostasis of host effector immune cell subsets. The mechanisms involved in this commensal–host crosstalk are not well understood. Intestinal epithelial cells (IECs) not only create a physical barrier between the commensals and immune cells in host tissues, but also facilitate interactions between them. Perturbations of epithelial homeostasis or function lead to the development of intestinal disorders such as inflammatory bowel diseases (IBD) and intestinal cancer. IECs receive signals from commensals and produce effector immune molecules. IECs also affect the function of immune cells in the lamina propria. Here we discuss some of these properties of IECs that define them as innate immune cells. We focus on how IECs may integrate and transmit signals from individual commensal bacteria to mucosal innate and adaptive immune cells for the establishment of the unique mucosal immunological equilibrium. PMID:23318659

  17. Cell-associated HIV mucosal transmission: the neglected pathway.

    PubMed

    Anderson, Deborah J; Le Grand, Roger

    2014-12-15

    This supplement to The Journal of Infectious Diseases is devoted to the important and understudied topic of cell-associated human immunodeficiency virus Type 1 (HIV) mucosal transmission. It stems from a workshop held in Boston, Massachusetts, in October 2013, in which scientists discussed their research and insights regarding cell-associated HIV mucosal transmission. The 10 articles in this supplement present the case for cell-associated HIV transmission as an important element contributing to the HIV epidemic, review evidence for the efficacy of current HIV prevention strategies against cell-associated HIV transmission and opportunities for further development, and describe in vitro, ex vivo, and animal cell-associated transmission models that can be used to further elucidate the molecular mechanisms of cell-associated HIV mucosal transmission and test HIV prevention strategies. We hope that these articles will help to inform and invigorate the HIV prevention field and contribute to the development of more-effective vaccine, treatment, and microbicide strategies for HIV prevention. PMID:25414413

  18. Epithelial organization, cell polarity and tumorigenesis.

    PubMed

    McCaffrey, Luke Martin; Macara, Ian G

    2011-12-01

    Epithelial cells comprise the foundation for the majority of organs in the mammalian body, and are the source of approximately 90% of all human cancers. Characteristically, epithelial cells form intercellular adhesions, exhibit apical/basal polarity, and orient their mitotic spindles in the plane of the epithelial sheet. Defects in these attributes result in the tissue disorganization associated with cancer. Epithelia undergo self-renewal from stem cells, which might in some cases be the cell of origin for cancers. The PAR polarity proteins are master regulators of epithelial organization, and are closely linked to signaling pathways such as Hippo, which orchestrate proliferation and apoptosis to control organ size. 3D ex vivo culture systems can now faithfully recapitulate epithelial organ morphogenesis, providing a powerful approach to study both normal development and the initiating events in carcinogenesis. PMID:21782440

  19. Cultured mucosal cell sheet with a double layer of keratinocytes and fibroblasts on a collagen membrane.

    PubMed

    Imaizumi, Fumiko; Asahina, Izumi; Moriyama, Takashi; Ishii, Masatoshi; Omura, Ken

    2004-01-01

    The aim of this study was to develop a novel cultured mucosal membrane that was facile to prepare and easy to handle, and that could be applied to mucosal defects in the oral cavity. Human oral keratinocytes and fibroblasts were prepared from the oral mucosa. We made the following two types of cultured mucosal cell sheets: a monolayer sheet of keratinocytes cultured on a collagen membrane (K-S) and a double-layered sheet of keratinocytes and fibroblasts on a collagen membrane (KF-S). A collagen membrane was used as a control. Each type of sheet was transplanted onto dorsal skin defects of nude mice. The wound area was measured for the assessment of wound contraction and a specimen was harvested for histologic evaluation 1 week and 4 weeks after grafting. Wound contraction was minimal with KF-S grafts. Although histologic examination showed normal differentiation of the epithelium in all graft types, the involucrin expression pattern of KFS was most similar to that of normal epithelium. These results indicate that a double-layered sheet of keratinocytes and fibroblasts cultured on a collagen membrane may facilitate epithelial healing and prevent wound contraction. PMID:15265283

  20. Epithelial TRPV1 signaling accelerates gingival epithelial cell proliferation.

    PubMed

    Takahashi, N; Matsuda, Y; Yamada, H; Tabeta, K; Nakajima, T; Murakami, S; Yamazaki, K

    2014-11-01

    Transient receptor potential cation channel subfamily V member 1 (TRPV1), a member of the calcium-permeable thermosensitive transient receptor potential superfamily, is a sensor of thermal and chemical stimuli. TRPV1 is activated by noxious heat (> 43°C), acidic conditions (pH < 6.6), capsaicin, and endovanilloids. This pain receptor was discovered on nociceptive fibers in the peripheral nervous system. TRPV1 was recently found to be expressed by non-neuronal cells, such as epithelial cells. The oral gingival epithelium is exposed to multiple noxious stimuli, including heat and acids derived from endogenous and exogenous substances; however, whether gingival epithelial cells (GECs) express TRPV1 is unknown. We show that both TRPV1 mRNA and protein are expressed by GECs. Capsaicin, a TRPV1 agonist, elevated intracellular Ca(2+) levels in the gingival epithelial cell line, epi 4. Moreover, TRPV1 activation in epi 4 cells accelerated proliferation. These responses to capsaicin were inhibited by a specific TRPV1 antagonist, SB-366791. We also observed GEC proliferation in capsaicin-treated mice in vivo. No effects were observed on GEC apoptosis by epithelial TRPV1 signaling. To examine the molecular mechanisms underlying this proliferative effect, we performed complementary (c)DNA microarray analysis of capsaicin-stimulated epi 4 cells. Compared with control conditions, 227 genes were up-regulated and 232 genes were down-regulated following capsaicin stimulation. Several proliferation-related genes were validated by independent experiments. Among them, fibroblast growth factor-17 and neuregulin 2 were significantly up-regulated in capsaicin-treated epi 4 cells. Our results suggest that functional TRPV1 is expressed by GECs and contributes to the regulation of cell proliferation. PMID:25266715

  1. Interleukin-10 prevents epithelial cell apoptosis by regulating IFNγ and TNFα expression in rhesus macaque colon explants

    PubMed Central

    Pan, Diganta; Das, Arpita; Lala, Wendy; Kenway-Lynch, Carys S.; Liu, David X.; Veazey, Ronald S.; Pahar, Bapi

    2013-01-01

    Interleukin-10 (IL-10) is an important immunomodulatory cytokine that plays an obligate role in regulating inflammatory responses. Here we demonstrated the role of IL-10 in regulating crypts length and breadth as well as maintaining the survival of epithelial cells using rhesus colon explant cultures. Anti-IL-10 antibody treatment of colon explant cultures induced increased production of inflammatory cytokines/molecules like IFNγ, TNFα, CD107a and perforin as well as increased epithelial cell apoptosis compared to media controls tested. Our results suggest that IL-10 plays a crucial role in maintaining mucosal homeostasis by regulating mucosal IFNγ and TNFα cytokine production. PMID:23867612

  2. Interleukin-10 prevents epithelial cell apoptosis by regulating IFNγ and TNFα expression in rhesus macaque colon explants.

    PubMed

    Pan, Diganta; Das, Arpita; Lala, Wendy; Kenway-Lynch, Carys S; Liu, David X; Veazey, Ronald S; Pahar, Bapi

    2013-10-01

    Interleukin-10 (IL-10) is an important immunomodulatory cytokine that plays an obligate role in regulating inflammatory responses. Here we demonstrated the role of IL-10 in regulating crypts length and breadth as well as maintaining the survival of epithelial cells using rhesus colon explant cultures. Anti-IL-10 antibody treatment of colon explant cultures induced increased production of inflammatory cytokines/molecules like IFNγ, TNFα, CD107a and perforin as well as increased epithelial cell apoptosis compared to media controls tested. Our results suggest that IL-10 plays a crucial role in maintaining mucosal homeostasis by regulating mucosal IFNγ and TNFα cytokine production. PMID:23867612

  3. Characteristic and Functional Analysis of a Newly Established Porcine Small Intestinal Epithelial Cell Line

    PubMed Central

    Wang, Jing; Hu, Guangdong; Lin, Zhi; He, Lei; Xu, Lei; Zhang, Yanming

    2014-01-01

    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 intestinal epithelial

  4. Symmetry breaking mechanism for epithelial cell polarization

    NASA Astrophysics Data System (ADS)

    Veglio, A.; Gamba, A.; Nicodemi, M.; Bussolino, F.; Serini, G.

    2009-09-01

    In multicellular organisms, epithelial cells form layers separating compartments responsible for different physiological functions. At the early stage of epithelial layer formation, each cell of an aggregate defines an inner and an outer side by breaking the symmetry of its initial state, in a process known as epithelial polarization. By integrating recent biochemical and biophysical data with stochastic simulations of the relevant reaction-diffusion system, we provide evidence that epithelial cell polarization is a chemical phase-separation process induced by a local bistability in the signaling network at the level of the cell membrane. The early symmetry breaking event triggering phase separation is induced by adhesion-dependent mechanical forces localized in the point of convergence of cell surfaces when a threshold number of confluent cells is reached. The generality of the emerging phase-separation scenario is likely common to many processes of cell polarity formation.

  5. Genotoxicity of di-butyl-phthalate and di-iso-butyl-phthalate in human lymphocytes and mucosal cells.

    PubMed

    Kleinsasser, N H; Wallner, B C; Kastenbauer, E R; Weissacher, H; Harréus, U A

    2001-01-01

    The genotoxicity of phthalates, widely used plasticizers, has been shown previously for di-butyl-phthalate (DBP) and di-iso-butyl-phthalate (DBP) in human mucosal cells of the upper aerodigestive tract in a previous study using the Comet assay. Furthermore, higher genotoxic sensitivities of patients with squamous cell carcinomas of either the larynx or the oropharynx compared to non-tumor patients were described. Other authors have demonstrated DNA damage by a different phthalate in human lymphocytes. It was the aim of the present study to determine whether there is a correlation between the genotoxic sensitivities to DBP and its isomer DiBP in either mucosal cells or lymphocytes. The single-cell microgel electrophoresis assay (Comet assay) was applied to detect DNA strand breaks in human epithelial cells of the upper aerodigestive tract (n=132 specimens). Human mucosa was harvested from the oropharynx in non-tumor patients and patients with squamous cell carcinomas of the oropharynx. Laryngeal mucosa of patients with laryngeal squamous cell carcinomas was harvested as well. Peripheral lymphocytes (n=49 specimens) were separated from peripheral blood. Xenobiotics investigated were DBP, DiBP, and N'methyl-N'-nitro-N-nitrosoguanidine (MNNG) as positive control, respectively. For statistical analysis, the SPSS correlation analysis according to Pearson and the Wilcoxon test were performed. Genotoxicity was found for DBP and DiBP in epithelial cells and lymphocytes (P<0.001). MNNG caused severe DNA damage. In analyzing DBP and DiBP results, genotoxic impacts in mucosal cells showed an intermediate correlation (r=0.570). Correlation in lymphocytes was the same (r=0.570). Phthalates have been investigated as a potential health hazard for a variety of reasons, including possible xenoestrogenic impact, peroxisome proliferation, and membrane destabilization. The present investigation suggests a correlated DNA-damaging impact of DBP and DiBP in human mucosal cells and in

  6. Airway epithelial cell responses to ozone injury

    SciTech Connect

    Leikauf, G.D.; Simpson, L.G.; Zhao, Qiyu

    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.

  7. Mucosal Regulatory T Cells and T Helper 17 Cells in HIV-Associated Immune Activation

    PubMed Central

    Pandiyan, Pushpa; Younes, Souheil-Antoine; Ribeiro, Susan Pereira; Talla, Aarthi; McDonald, David; Bhaskaran, Natarajan; Levine, Alan D.; Weinberg, Aaron; Sekaly, Rafick P.

    2016-01-01

    Residual mucosal inflammation along with chronic systemic immune activation is an important feature in individuals infected with human immunodeficiency virus (HIV), and has been linked to a wide range of co-morbidities, including malignancy, opportunistic infections, immunopathology, and cardiovascular complications. Although combined antiretroviral therapy (cART) can reduce plasma viral loads to undetectable levels, reservoirs of virus persist, and increased mortality is associated with immune dysbiosis in mucosal lymphoid tissues. Immune-based therapies are pursued with the goal of improving CD4+ T-cell restoration, as well as reducing chronic immune activation in cART-treated patients. However, the majority of research on immune activation has been derived from analysis of circulating T cells. How immune cell alterations in mucosal tissues contribute to HIV immune dysregulation and the associated risk of non-infectious chronic complications is less studied. Given the significant differences between mucosal T cells and circulating T cells, and the immediate interactions of mucosal T cells with the microbiome, more attention should be devoted to mucosal immune cells and their contribution to systemic immune activation in HIV-infected individuals. Here, we will focus on mucosal immune cells with a specific emphasis on CD4+ T lymphocytes, such as T helper 17 cells and CD4+Foxp3+ regulatory T cells (Tregs), which play crucial roles in maintaining mucosal barrier integrity and preventing inflammation, respectively. We hypothesize that pro-inflammatory milieu in cART-treated patients with immune activation significantly contributes to enhanced loss of Th17 cells and increased frequency of dysregulated Tregs in the mucosa, which in turn may exacerbate immune dysfunction in HIV-infected patients. We also present initial evidence to support this hypothesis. A better comprehension of how pro-inflammatory milieu impacts these two types of cells in the mucosa will shed light

  8. Absence of inflammatory response from upper airway epithelial cells after X irradiation.

    PubMed

    Reiter, R; Deutschle, T; Wiegel, T; Riechelmann, H; Bartkowiak, D

    2009-03-01

    Radiotherapy of head and neck tumors causes adverse reactions in normal tissue, especially mucositis. The dose- and time-dependent response of upper airway cells to X radiation should be analyzed in terms of the pro-inflammatory potential. Immortalized BEAS-2B lung epithelial cells were treated with 2, 5 and 8 Gy. Out of 1232 genes, those that were transcribed differentially after 2, 6 and 24 h were assigned to biological themes according to the Gene Ontology Consortium. Enrichment of differentially regulated gene clusters was determined with GOTree ( http://bioinfo.vanderbilt.edu/gotm ). Eleven cytokines were measured in culture supernatants. The cell cycle response up to 24 h and induction of apoptosis up to 4 days after exposure were determined by flow cytometry. A significant dose- and time-dependent gene activation was observed for the categories response to DNA damage, oxidative stress, cell cycle arrest and cell death/apoptosis but not for immune/inflammatory response. This correlated with functional G(2) arrest and apoptosis. Pro-inflammatory cytokines accumulated in supernatants of control cells but not of X-irradiated cells. The complex gene expression pattern of X-irradiated airway epithelial cells is accompanied by cell cycle arrest and induction of apoptosis. In vivo, this may impair the epithelial barrier. mRNA and protein expression suggest at most an indirect contribution of epithelial cells to early radiogenic mucositis. PMID:19267554

  9. The anti-inflammatory activity of curcumin protects the genital mucosal epithelial barrier from disruption and blocks replication of HIV-1 and HSV-2.

    PubMed

    Ferreira, Victor H; Nazli, Aisha; Dizzell, Sara E; Mueller, Kristen; Kaushic, Charu

    2015-01-01

    Inflammation is a known mechanism that facilitates HIV acquisition and the spread of infection. In this study, we evaluated whether curcumin, a potent and safe anti-inflammatory compound, could be used to abrogate inflammatory processes that facilitate HIV-1 acquisition in the female genital tract (FGT) and contribute to HIV amplification. Primary, human genital epithelial cells (GECs) were pretreated with curcumin and exposed to HIV-1 or HIV glycoprotein 120 (gp120), both of which have been shown to disrupt epithelial tight junction proteins, including ZO-1 and occludin. Pre-treatment with curcumin prevented disruption of the mucosal barrier by maintaining ZO-1 and occludin expression and maintained trans-epithelial electric resistance across the genital epithelium. Curcumin pre-treatment also abrogated the gp120-mediated upregulation of the proinflammatory cytokines tumor necrosis factor-α and interleukin (IL)-6, which mediate barrier disruption, as well as the chemokines IL-8, RANTES and interferon gamma-induced protein-10 (IP-10), which are capable of recruiting HIV target cells to the FGT. GECs treated with curcumin and exposed to the sexually transmitted co-infecting microbes HSV-1, HSV-2 and Neisseria gonorrhoeae were unable to elicit innate inflammatory responses that indirectly induced activation of the HIV promoter and curcumin blocked Toll-like receptor (TLR)-mediated induction of HIV replication in chronically infected T-cells. Finally, curcumin treatment resulted in significantly decreased HIV-1 and HSV-2 replication in chronically infected T-cells and primary GECs, respectively. All together, our results suggest that the use of anti-inflammatory compounds such as curcumin may offer a viable alternative for the prevention and/or control of HIV replication in the FGT. PMID:25856395

  10. Cell Division Drives Epithelial Cell Rearrangements during Gastrulation in Chick.

    PubMed

    Firmino, Joao; Rocancourt, Didier; Saadaoui, Mehdi; Moreau, Chloe; Gros, Jerome

    2016-02-01

    During early embryonic development, cells are organized as cohesive epithelial sheets that are continuously growing and remodeled without losing their integrity, giving rise to a wide array of tissue shapes. Here, using live imaging in chick embryo, we investigate how epithelial cells rearrange during gastrulation. We find that cell division is a major rearrangement driver that powers dramatic epithelial cell intercalation events. We show that these cell division-mediated intercalations, which represent the majority of epithelial rearrangements within the early embryo, are absolutely necessary for the spatial patterning of gastrulation movements. Furthermore, we demonstrate that these intercalation events result from overall low cortical actomyosin accumulation within the epithelial cells of the embryo, which enables dividing cells to remodel junctions in their vicinity. These findings uncover a role for cell division as coordinator of epithelial growth and remodeling that might underlie various developmental, homeostatic, or pathological processes in amniotes. PMID:26859350

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

    PubMed

    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. PMID:26446290

  12. Effects of spaceflight on the proliferation of jejunal mucosal cells

    NASA Technical Reports Server (NTRS)

    Phillips, Robert W.; Moeller, C. L.; Sawyer, Heywood R.; Smirnov, K. L.

    1991-01-01

    The purpose of this project was to test the hypothesis that the generalized, whole body decrease in synthetic activity due to microgravity conditions encountered during spaceflight would be demonstrable in cells and tissues characterized by a rapid rate of turnover. Jejunal mucosal cells were chosen as a model since these cells are among the most rapidly proliferating in the body. Accordingly, the percentage of mitotic cells present in the crypts of Lieberkuhn in each of 5 rats flown on the COSMOS 2044 mission were compared to the percentage of mitotic cells present in the crypts in rats included in each of 3 ground control groups (i.e., vivarium, synchronous and caudal-elevated). No significant difference (p greater than .05) was detected in mitotic indices between the flight and vivarium group. Although the ability of jejunal mucosal cells to divide by mitosis was not impaired in flight group, there was, however, a reduction in the length of villi and depth of crypts. The concommitant reduction in villus length and crypth depth in the flight group probably reflects changes in connective tissue components within the core of villi.

  13. Odontogenic epithelial stem cells: hidden sources.

    PubMed

    Padma Priya, Sivan; Higuchi, Akon; Abu Fanas, Salem; Pooi Ling, Mok; Kumari Neela, Vasantha; Sunil, P M; Saraswathi, T R; Murugan, Kadarkarai; Alarfaj, Abdullah A; Munusamy, Murugan A; Kumar, Suresh

    2015-12-01

    The ultimate goal of dental stem cell research is to construct a bioengineered tooth. Tooth formation occurs based on the well-organized reciprocal interaction of epithelial and mesenchymal cells. The dental mesenchymal stem cells are the best explored, but because the human odontogenic epithelium is lost after the completion of enamel formation, studies on these cells are scarce. The successful creation of a bioengineered tooth is achievable only when the odontogenic epithelium is reconstructed to produce a replica of natural enamel. This article discusses the untapped sources of odontogenic epithelial stem cells in humans, such as those present in the active dental lamina in postnatal life, in remnants of dental lamina (the gubernaculum cord), in the epithelial cell rests of Malassez, and in reduced enamel epithelium. The possible uses of these stem cells in regenerative medicine, not just for enamel formation, are discussed. PMID:26367485

  14. Immunity and Tolerance Induced by Intestinal Mucosal Dendritic Cells.

    PubMed

    Aliberti, Julio

    2016-01-01

    Dendritic cells present in the digestive tract are constantly exposed to environmental antigens, commensal flora, and invading pathogens. Under steady-state conditions, these cells have high tolerogenic potential, triggering differentiation of regulatory T cells to protect the host from unwanted proinflammatory immune responses to innocuous antigens or commensals. On the other hand, these cells must discriminate between commensal flora and invading pathogens and mount powerful immune response against pathogens. A potential result of unbalanced tolerogenic versus proinflammatory responses mediated by dendritic cells is associated with chronic inflammatory conditions, such as Crohn's disease, ulcerative colitis, food allergies, and celiac disease. Herein, we review the dendritic cell population involved in mediating tolerance and immunity in mucosal surfaces, the progress in unveiling their development in vivo, and factors that can influence their functions. PMID:27034589

  15. Immunity and Tolerance Induced by Intestinal Mucosal Dendritic Cells

    PubMed Central

    Aliberti, Julio

    2016-01-01

    Dendritic cells present in the digestive tract are constantly exposed to environmental antigens, commensal flora, and invading pathogens. Under steady-state conditions, these cells have high tolerogenic potential, triggering differentiation of regulatory T cells to protect the host from unwanted proinflammatory immune responses to innocuous antigens or commensals. On the other hand, these cells must discriminate between commensal flora and invading pathogens and mount powerful immune response against pathogens. A potential result of unbalanced tolerogenic versus proinflammatory responses mediated by dendritic cells is associated with chronic inflammatory conditions, such as Crohn's disease, ulcerative colitis, food allergies, and celiac disease. Herein, we review the dendritic cell population involved in mediating tolerance and immunity in mucosal surfaces, the progress in unveiling their development in vivo, and factors that can influence their functions. PMID:27034589

  16. Human-restricted bacterial pathogens block shedding of epithelial cells by stimulating integrin activation.

    PubMed

    Muenzner, Petra; Bachmann, Verena; Zimmermann, Wolfgang; Hentschel, Jochen; Hauck, Christof R

    2010-09-01

    Colonization of mucosal surfaces is the key initial step in most bacterial infections. One mechanism protecting the mucosa is the rapid shedding of epithelial cells, also termed exfoliation, but it is unclear how pathogens counteract this process. We found that carcinoembryonic antigen (CEA)-binding bacteria colonized the urogenital tract of CEA transgenic mice, but not of wild-type mice, by suppressing exfoliation of mucosal cells. CEA binding triggered de novo expression of the transforming growth factor receptor CD105, changing focal adhesion composition and activating beta1 integrins. This manipulation of integrin inside-out signaling promotes efficient mucosal colonization and represents a potential target to prevent or cure bacterial infections. PMID:20813953

  17. In vitro Intestinal Mucosal Epithelial Responses to Wild-Type Salmonella Typhi and Attenuated Typhoid Vaccines.

    PubMed

    Fiorentino, Maria; Lammers, Karen M; Levine, Myron M; Sztein, Marcelo B; Fasano, Alessio

    2013-01-01

    Typhoid fever, caused by S. Typhi, is responsible for approximately 200,000 deaths per year worldwide. Little information is available regarding epithelium-bacterial interactions in S. Typhi infection. We have evaluated in vitro the effects of wild-type S. Typhi, the licensed Ty21a typhoid vaccine and the leading strains CVD 908-htrA and CVD 909 vaccine candidates on intestinal barrier function and immune response. Caco2 monolayers infected with wild-type S. Typhi exhibited alterations in the organization of tight junctions, increased paracellular permeability, and a rapid decrease in Trans-Epithelial Electrical Resistance as early as 4 h post-exposure. S. Typhi triggered the secretion of interleukin (IL)-8 and IL-6. Caco2 cells infected with the attenuated strains exhibited a milder pro-inflammatory response with minimal disruption of the barrier integrity. We conclude that wild-type S. Typhi causes marked transient alterations of the intestinal mucosa that are more pronounced than those observed with Ty21a or new generation attenuated typhoid vaccine candidates. PMID:23408152

  18. Reappraisal of the mucosal epithelial space associated with the surface of Hymenolepis diminuta and its effect on transport parameters.

    PubMed

    Murphy, W A; Lumsden, R D

    1984-08-01

    Using nonpermeating, radiolabeled solutes to estimate the magnitude of the "unstirred water layer" (="mucosal epithelial space") of the surface of Hymenolepis diminuta, a value approximating 1% of the worm's fluid volume (0.011-0.022 ml/g wet tissue) was obtained. This value was compared with those previously reported by other workers which were greater by an order of magnitude. The difference between these results appears to be related to the use in previous studies of a permeating marker (mannitol), and a failure to divest the surface of nonspecifically adherent bathing fluid in excess of the actual "unstirred layer". These parameters must be considered in future studies on this useful model for the study of transport. PMID:6438294

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

    PubMed Central

    Domachowske, Joseph B; Bonville, Cynthia A; Rosenberg, Helene F

    2001-01-01

    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. PMID:11686888

  20. Activation of rat intestinal mucosal mast cells by fat absorption.

    PubMed

    Ji, Yong; Sakata, Yasuhisa; Yang, Qing; Li, Xiaoming; Xu, Min; Yoder, Stephanie; Langhans, Wolfgang; Tso, Patrick

    2012-06-01

    Previous studies have linked certain types of gut mucosal immune cells with fat intake. We determined whether fat absorption activates intestinal mucosal mast cells (MMC), a key component of the gut mucosal immune system. Conscious intestinal lymph fistula rats were used. The mesenteric lymph ducts were cannulated, and the intraduodenal (i.d.) tubes were installed for the infusion of Liposyn II 20% (an intralipid emulsion). Lymphatic concentrations of histamine, rat MMC protease II (RMCPII), a specific marker of rat intestinal MMC degranulation, and prostaglandin D(2) (PGD(2)) were measured by ELISA. Intestinal MMC degranulation was visualized by immunofluorescent microscopy of jejunum sections taken at 1 h after Liposyn II gavage. Intraduodenal bolus infusion of Liposyn II 20% (4.4 kcal/3 ml) induced approximately a onefold increase in lymphatic histamine and PGD(2), ∼20-fold increase in lymphatic RMCPII, but only onefold increase in peripheral serum RMCPII concentrations. Release of RMCPII into lymph increased dose dependently with the amount of lipid fed. In addition, i.d. infusion of long-chain triacylglycerol trilinolein (C18:2 n-6, the major composite in Liposyn II) significantly increased the lymphatic RMCPII concentration, whereas medium-chain triacylglycerol tricaprylin (C8:0) did not alter lymph RMCPII secretion. Immunohistochemistry image revealed the degranulation of MMC into lamina propria after lipid feeding. These novel findings indicate that intestinal MMC are activated and degranulate to release MMC mediators to the circulation during fat absorption. This action of fatty acid is dose and chain length dependent. PMID:22461027

  1. Induced pluripotency of human prostatic epithelial cells.

    PubMed

    Zhao, Hongjuan; Sun, Ning; Young, Sarah R; Nolley, Rosalie; Santos, Jennifer; Wu, Joseph C; Peehl, Donna M

    2013-01-01

    Induced pluripotent stem (iPS) cells are a valuable resource for discovery of epigenetic changes critical to cell type-specific differentiation. Although iPS cells have been generated from other terminally differentiated cells, the reprogramming of normal adult human basal prostatic epithelial (E-PZ) cells to a pluripotent state has not been reported. Here, we attempted to reprogram E-PZ cells by forced expression of Oct4, Sox2, c-Myc, and Klf4 using lentiviral vectors and obtained embryonic stem cell (ESC)-like colonies at a frequency of 0.01%. These E-PZ-iPS-like cells with normal karyotype gained expression of pluripotent genes typical of iPS cells (Tra-1-81, SSEA-3, Nanog, Sox2, and Oct4) and lost gene expression characteristic of basal prostatic epithelial cells (CK5, CK14, and p63). E-PZ-iPS-like cells demonstrated pluripotency by differentiating into ectodermal, mesodermal, and endodermal cells in vitro, although lack of teratoma formation in vivo and incomplete demethylation of pluripotency genes suggested only partial reprogramming. Importantly, E-PZ-iPS-like cells re-expressed basal epithelial cell markers (CD44, p63, MAO-A) in response to prostate-specific medium in spheroid culture. Androgen induced expression of androgen receptor (AR), and co-culture with rat urogenital sinus further induced expression of prostate-specific antigen (PSA), a hallmark of secretory cells, suggesting that E-PZ-iPS-like cells have the capacity to differentiate into prostatic basal and secretory epithelial cells. Finally, when injected into mice, E-PZ-iPS-like cells expressed basal epithelial cell markers including CD44 and p63. When co-injected with rat urogenital mesenchyme, E-PZ-iPS-like cells expressed AR and expression of p63 and CD44 was repressed. DNA methylation profiling identified epigenetic changes in key pathways and genes involved in prostatic differentiation as E-PZ-iPS-like cells converted to differentiated AR- and PSA-expressing cells. Our results suggest that

  2. Roflumilast combined with adenosine increases mucosal hydration in human airway epithelial cultures after cigarette smoke exposure

    PubMed Central

    Tyrrell, Jean; Qian, Xiaozhong; Freire, Jose

    2015-01-01

    Chronic obstructive pulmonary disease (COPD) is a growing cause of morbidity and mortality worldwide. Recent studies have shown that cigarette smoke (CS) induces cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction, which leads to airway-surface liquid (ASL) dehydration. This in turn contributes to the mucus dehydration and impaired mucociliary clearance that are seen in the chronic bronchitis form of COPD. Roflumilast is a phosphodiesterase 4 inhibitor that may improve lung function and reduce the frequency of exacerbations in patients with COPD. Although roflumilast can affect cAMP metabolism, little is known about the downstream pharmacological effects in the airways. We hypothesized that roflumilast would increase ASL rehydration in human bronchial epithelial cultures (HBECs) after chronic CS exposure. cAMP production was measured by Förster resonance energy transfer in HEK293T cells and by ELISA in HBECs. ASL height was measured by xz-confocal microscopy after air exposure or following HBEC exposure to freshly produced CS. Roflumilast had little effect on cAMP or ASL height when applied on its own; however, roflumilast significantly potentiated adenosine-induced increases in cAMP and ASL height in CS-exposed HBECs. Roflumilast increased the rate of ASL height recovery in cultures after CS exposure compared with controls. In contrast, the β2-adrenergic receptor agonists isoproterenol and salmeterol failed to increase ASL height after CS exposure. Our data suggest that roflumilast can increase ASL hydration in CS-exposed HBECs, which is predicted to be beneficial for the treatment of mucus dehydration/mucus stasis in patients with COPD chronic bronchitis. PMID:25795727

  3. Roflumilast combined with adenosine increases mucosal hydration in human airway epithelial cultures after cigarette smoke exposure.

    PubMed

    Tyrrell, Jean; Qian, Xiaozhong; Freire, Jose; Tarran, Robert

    2015-05-15

    Chronic obstructive pulmonary disease (COPD) is a growing cause of morbidity and mortality worldwide. Recent studies have shown that cigarette smoke (CS) induces cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction, which leads to airway-surface liquid (ASL) dehydration. This in turn contributes to the mucus dehydration and impaired mucociliary clearance that are seen in the chronic bronchitis form of COPD. Roflumilast is a phosphodiesterase 4 inhibitor that may improve lung function and reduce the frequency of exacerbations in patients with COPD. Although roflumilast can affect cAMP metabolism, little is known about the downstream pharmacological effects in the airways. We hypothesized that roflumilast would increase ASL rehydration in human bronchial epithelial cultures (HBECs) after chronic CS exposure. cAMP production was measured by Förster resonance energy transfer in HEK293T cells and by ELISA in HBECs. ASL height was measured by xz-confocal microscopy after air exposure or following HBEC exposure to freshly produced CS. Roflumilast had little effect on cAMP or ASL height when applied on its own; however, roflumilast significantly potentiated adenosine-induced increases in cAMP and ASL height in CS-exposed HBECs. Roflumilast increased the rate of ASL height recovery in cultures after CS exposure compared with controls. In contrast, the β2-adrenergic receptor agonists isoproterenol and salmeterol failed to increase ASL height after CS exposure. Our data suggest that roflumilast can increase ASL hydration in CS-exposed HBECs, which is predicted to be beneficial for the treatment of mucus dehydration/mucus stasis in patients with COPD chronic bronchitis. PMID:25795727

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

    PubMed Central

    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. PMID:26730599

  5. Loss of ADAM17-Mediated Tumor Necrosis Factor Alpha Signaling in Intestinal Cells Attenuates Mucosal Atrophy in a Mouse Model of Parenteral Nutrition

    PubMed Central

    Feng, Yongjia; Tsai, Yu-Hwai; Xiao, Weidong; Ralls, Matthew W.; Stoeck, Alex; Wilson, Carole L.; Raines, Elaine W.

    2015-01-01

    Total parenteral nutrition (TPN) is commonly used clinically to sustain patients; however, TPN is associated with profound mucosal atrophy, which may adversely affect clinical outcomes. Using a mouse TPN model, removing enteral nutrition leads to decreased crypt proliferation, increased intestinal epithelial cell (IEC) apoptosis and increased mucosal tumor necrosis factor alpha (TNF-α) expression that ultimately produces mucosal atrophy. Upregulation of TNF-α signaling plays a central role in mediating TPN-induced mucosal atrophy without intact epidermal growth factor receptor (EGFR) signaling. Currently, the mechanism and the tissue-specific contributions of TNF-α signaling to TPN-induced mucosal atrophy remain unclear. ADAM17 is an ectodomain sheddase that can modulate the signaling activity of several cytokine/growth factor receptor families, including the TNF-α/TNF receptor and ErbB ligand/EGFR pathways. Using TPN-treated IEC-specific ADAM17-deficient mice, the present study demonstrates that a loss of soluble TNF-α signaling from IECs attenuates TPN-induced mucosal atrophy. Importantly, this response remains dependent on the maintenance of functional EGFR signaling in IECs. TNF-α blockade in wild-type mice receiving TPN confirmed that soluble TNF-α signaling is responsible for downregulation of EGFR signaling in IECs. These results demonstrate that ADAM17-mediated TNF-α signaling from IECs has a significant role in the development of the proinflammatory state and mucosal atrophy observed in TPN-treated mice. PMID:26283731

  6. Diesel Exhaust Particle-Exposed Human Bronchial Epithelial Cells Induce Dendritic Cell Maturation and Polarization via Thymic Stromal Lymphopoietin

    PubMed Central

    Bleck, Bertram; Tse, Doris B.; Curotto de Lafaille, Maria A.; Zhang, Feijie

    2009-01-01

    Human exposure to air pollutants, including ambient particulate matter, has been proposed as a mechanism for the rise in allergic disorders. Diesel exhaust particles, a major component of ambient particulate matter, induce sensitization to neoallergens, but the mechanisms by which sensitization occur remain unclear. We show that diesel exhaust particles upregulate thymic stromal lymphopoietin in human bronchial epithelial cells in an oxidant-dependent manner. Thymic stromal lymphopoietin induced by diesel exhaust particles was associated with maturation of myeloid dendritic cells, which was blocked by anti-thymic stromal lymphopoietin antibodies or silencing epithelial cell-derived thymic stromal lymphopoietin. Dendritic cells exposed to diesel exhaust particle-treated human bronchial epithelial cells induced Th2 polarization in a thymic stromal lymphopoietin-dependent manner. These findings provide new insight into the mechanisms by which diesel exhaust particles modify human lung mucosal immunity. PMID:18049884

  7. Role of the Amino-Terminal Region of Porphyromonas gingivalis Fimbriae in Adherence to Epithelial Cells

    PubMed Central

    Sojar, Hakimuddin T.; Han, Yiping; Hamada, Nobushiro; Sharma, Ashu; Genco, Robert J.

    1999-01-01

    Porphyromonas gingivalis fimbriae elicit many responses in eukaryotic cells, including mitogenicity, cytokine production, epithelial cell invasion, and cellular immune response. Specific domains of the major fimbrial protein (FimA) have been shown to be important in triggering some of these functions. The goal of the present study was to identify the domain(s) of P. gingivalis FimA responsible for specific interaction with human mucosal epithelial cells. Fimbriated P. gingivalis strains have been shown to bind to buccal epithelial cells, whereas nonfimbriated strains bind at low levels or not at all. This and other studies provide evidence that FimA mediates the adherence of P. gingivalis to oral epithelial cells. To determine the specific region(s) of P. gingivalis FimA involved in epithelial cell binding, specific antipeptide antibodies were used to inhibit the binding of iodinated purified fimbriae as well as the binding of P. gingivalis cells to epithelial cells. Antibodies directed against peptides 49 to 68 (VVMANTAGAMELVGKTLAEVK) and 69 to 90 (ALTTELTAENQEAAGLIMTAEP) were found to highly inhibit both the binding of fimbriae and the binding of P. gingivalis cells to epithelial cells. The antibody against FimA peptides 69 to 90 also reacted with P. gingivalis fimbriae in immunogold labeling and immunoblot analysis, thereby indicating that this peptide domain is exposed on the surface of fimbriae. Our results suggest that the amino-terminal domain corresponding to amino acid residues 49 to 90 of the fimbrillin protein is a major epithelial cell binding domain of P. gingivalis fimbriae. PMID:10531284

  8. Epithelial Cell Shedding and Barrier Function

    PubMed Central

    Williams, J. M.; Duckworth, C. A.; Burkitt, M. D.; Watson, A. J. M.; Campbell, B. J.

    2015-01-01

    The intestinal epithelium is a critical component of the gut barrier. Composed of a single layer of intestinal epithelial cells (IECs) held together by tight junctions, this delicate structure prevents the transfer of harmful microorganisms, antigens, and toxins from the gut lumen into the circulation. The equilibrium between the rate of apoptosis and shedding of senescent epithelial cells at the villus tip, and the generation of new cells in the crypt, is key to maintaining tissue homeostasis. However, in both localized and systemic inflammation, this balance may be disturbed as a result of pathological IEC shedding. Shedding of IECs from the epithelial monolayer may cause transient gaps or microerosions in the epithelial barrier, resulting in increased intestinal permeability. Although pathological IEC shedding has been observed in mouse models of inflammation and human intestinal conditions such as inflammatory bowel disease, understanding of the underlying mechanisms remains limited. This process may also be an important contributor to systemic and intestinal inflammatory diseases and gut barrier dysfunction in domestic animal species. This review aims to summarize current knowledge about intestinal epithelial cell shedding, its significance in gut barrier dysfunction and host-microbial interactions, and where research in this field is directed. PMID:25428410

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

    EPA Science Inventory

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

  10. Expression of the ATP-gated P2X7 Receptor on M Cells and Its Modulating Role in the Mucosal Immune Environment.

    PubMed

    Kim, Sae-Hae; Lee, Ha-Yan; Jang, Yong-Suk

    2015-02-01

    Interactions between microbes and epithelial cells in the gastrointestinal tract are closely associated with regulation of intestinal mucosal immune responses. Recent studies have highlighted the modulation of mucosal immunity by microbe-derived molecules such as ATP and short-chain fatty acids. In this study, we undertook to characterize the expression of the ATP-gated P2X7 receptor (P2X7R) on M cells and its role in gastrointestinal mucosal immune regulation because it was poorly characterized in Peyer's patches, although purinergic signaling via P2X7R and luminal ATP have been considered to play an important role in the gastrointestinal tract. Here, we present the first report on the expression of P2X7R on M cells and characterize the role of P2X7R in immune enhancement by ATP or LL-37. PMID:25713508

  11. Intranasal Immunization Strategy To Impede Pilin-Mediated Binding of Pseudomonas aeruginosa to Airway Epithelial Cells

    PubMed Central

    Hsieh, Jennifer C.; Tham, Doris M.; Feng, Weijun; Huang, Fan; Embaie, Selamawit; Liu, Keyi; Dean, Deborah; Hertle, Ralf; FitzGerald, David J.; Mrsny, Randall J.

    2005-01-01

    Prevention of pulmonary Pseudomonas aeruginosa infections represents a critical unmet medical need for cystic fibrosis (CF) patients. We have examined the tenet that a mucosal immunization approach can reduce interactions of a piliated form of this opportunistic pathogen with respiratory epithelial cells. Vaccinations were performed using ntPEpilinPAK, a protein chimera composed of a nontoxic form of P. aeruginosa exotoxin A (ntPE), where the C-terminal loop amino acid sequence of the PAK strain pilin protein was inserted in place of the ntPE Ib domain. Intranasal (i.n.) immunization of BALB/c mice with ntPEpilinPAK generated both serum and saliva immune responses. A series of in vitro studies showed that diluted samples of saliva obtained from immunized mice reduced pilin-dependent P. aeruginosa binding to polarized human tracheal epithelial cells, protected human pulmonary epithelial cells from cytotoxic actions associated with bacterial challenge, and reduced exotoxin A toxicity. Overall, i.n. administration of ntPEpilinPAK induced mucosal and systemic immune responses that may be beneficial for blocking early stage adhesion and/or infection events of epithelial cell-P. aeruginosa interactions at oropharyngeal surfaces. PMID:16239575

  12. Mucosal vaccines

    PubMed Central

    Nizard, Mevyn; Diniz, Mariana O; Roussel, Helene; Tran, Thi; Ferreira, Luis CS; Badoual, Cecile; Tartour, Eric

    2014-01-01

    The mucosal immune system displays several adaptations reflecting the exposure to the external environment. The efficient induction of mucosal immune responses also requires specific approaches, such as the use of appropriate administration routes and specific adjuvants and/or delivery systems. In contrast to vaccines delivered via parenteral routes, experimental, and clinical evidences demonstrated that mucosal vaccines can efficiently induce local immune responses to pathogens or tumors located at mucosal sites as well as systemic response. At least in part, such features can be explained by the compartmentalization of mucosal B and T cell populations that play important roles in the modulation of local immune responses. In the present review, we discuss molecular and cellular features of the mucosal immune system as well as novel immunization approaches that may lead to the development of innovative and efficient vaccines targeting pathogens and tumors at different mucosal sites. PMID:25424921

  13. Dendritic cell-derived tumor necrosis factor α modifies airway epithelial cell responses.

    PubMed

    Lutfi, R; Ledford, J R; Zhou, P; Lewkowich, I P; Page, K

    2012-01-01

    Mucosal dendritic cells (DC) are intimately associated with the airway epithelium and thus are ideally situated to be first responders to pathogens. We hypothesize that DC drive innate immune responses through early release of tumor necrosis factor (TNF) α, which drives airway epithelial cell responses. In a mouse model, TNFα release was significantly increased following a single exposure to German cockroach (GC) frass, an event independent of neutrophil recruitment into the airways. While lung epithelial cells and alveolar macrophages failed to release TNFα following GC frass exposure, bone marrow-derived DC (BMDC) produced substantial amounts of TNFα suggesting their importance as early responding cells. This was confirmed by flow cytometry of pulmonary myeloid DC. Addition of GC frass-pulsed BMDC or conditioned media from GC frass-pulsed BMDC to primary mouse tracheal epithelial cells (MTEC) or MLE-15 cells induced chemokine (C-C) motif ligand (CCL) 20 and granulocyte macrophage (GM) colony-stimulating factor (CSF), both of which are important for DC recruitment, survival and differentiation. Importantly, DC do not produce CCL20 or GM-CSF following allergen exposure. Blocking TNFα receptor 1 (TNFR1) completely abolished chemokine production, suggesting that BMDC-derived TNFα induced airway epithelial cell activation and enhancement of the innate immune response. Lastly, blocking TNFR1 in vivo resulted in significantly decreased CCL20 and GM-CSF production in the lungs of mice. Together, our data strongly suggest that DC-derived TNFα plays a crucial role in the initiation of innate immune responses through the modification of airway epithelial cell responses. PMID:22517116

  14. Innate lymphoid cells regulate intestinal epithelial cell glycosylation.

    PubMed

    Goto, Yoshiyuki; Obata, Takashi; Kunisawa, Jun; Sato, Shintaro; Ivanov, Ivaylo I; Lamichhane, Aayam; Takeyama, Natsumi; Kamioka, Mariko; Sakamoto, Mitsuo; Matsuki, Takahiro; Setoyama, Hiromi; Imaoka, Akemi; Uematsu, Satoshi; Akira, Shizuo; Domino, Steven E; Kulig, Paulina; Becher, Burkhard; Renauld, Jean-Christophe; Sasakawa, Chihiro; Umesaki, Yoshinori; Benno, Yoshimi; Kiyono, Hiroshi

    2014-09-12

    Fucosylation of intestinal epithelial cells, catalyzed by fucosyltransferase 2 (Fut2), is a major glycosylation mechanism of host-microbiota symbiosis. Commensal bacteria induce epithelial fucosylation, and epithelial fucose is used as a dietary carbohydrate by many of these bacteria. However, the molecular and cellular mechanisms that regulate the induction of epithelial fucosylation are unknown. Here, we show that type 3 innate lymphoid cells (ILC3) induced intestinal epithelial Fut2 expression and fucosylation in mice. This induction required the cytokines interleukin-22 and lymphotoxin in a commensal bacteria-dependent and -independent manner, respectively. Disruption of intestinal fucosylation led to increased susceptibility to infection by Salmonella typhimurium. Our data reveal a role for ILC3 in shaping the gut microenvironment through the regulation of epithelial glycosylation. PMID:25214634

  15. Role of mast cells in gastrointestinal mucosal defense.

    PubMed

    Penissi, Alicia B; Rudolph, María I; Piezzi, Ramón S

    2003-08-01

    The purpose of this review, based on studies from our laboratory as well as from others, is to summarize salient features of mast cell immunobiology and to describe their associations with gastrointestinal mucosal defense. Gastrointestinal mast cells are involved in many pathologic effects, such as food hypersensitivity. On the other hand, they also play a protective role in defense against parasitic and microbial infections. Thus, they have both positive and negative effects, but presently the mechanisms that control the balance of these various effects are poorly known. It has been suggested that stabilization of mast cells may be a key mechanism to protect the gastrointestinal tract from injury. Few molecules are known to possess both mast cell stabilizing and gastrointestinal cytoprotective activity. These include zinc compounds, sodium cromoglycate, FPL 52694, ketotifen, aloe vera, certain flavonoids such as quercetin, some sulfated proteoglycans such as chondroitin sulfate and dehydroleucodine. Dehydroleucodine, a sesquiterpene lactone isolated from Artemisia douglasiana Besser, exhibits anti-inflammatory and gastrointestinal cytoprotective action. The lactone stimulates mucus production, and inhibits histamine and serotonin release from intestinal mast cells. The lactone could act as a selective mast cell stabilizer by releasing cytoprotective factors and inhibiting pro-inflammatory mast cell mediators. PMID:14510234

  16. Postbiotic Modulation of Retinoic Acid Imprinted Mucosal-like Dendritic Cells by Probiotic Lactobacillus reuteri 17938 In Vitro

    PubMed Central

    Haileselassie, Yeneneh; Navis, Marit; Vu, Nam; Qazi, Khaleda Rahman; Rethi, Bence; Sverremark-Ekström, Eva

    2016-01-01

    Lactobacilli are widely used as probiotics with beneficial effects on infection-associated diarrhea, but also used in clinical trials of e.g., necrotizing enterocolitis and inflammatory bowel diseases. The possibility of using probiotic metabolic products, so-called postbiotics, is desirable as it could prevent possible side effects of live bacteria in individuals with a disturbed gut epithelial barrier. Here, we studied how Lactobacillus reuteri DSM 17938 cell-free supernatant (L. reuteri-CFS) influenced retinoic acid (RA)-driven mucosal-like dendritic cells (DC) and their subsequent effect on T regulatory cells (Treg) in vitro. RA clearly imprinted a mucosal-like DC phenotype with higher IL10 production, increased CD103 and CD1d expression, and a downregulated mRNA expression of several inflammatory-associated genes (NFκB1, RELB, and TNF). Treatment with L. reuteri-CFS further influenced the tolerogenic phenotype of RA-DC by downregulating most genes involved in antigen uptake, antigen presentation, and signal transduction as well as several chemokine receptors, while upregulating IL10 production. L. reuteri-CFS also augmented CCR7 expression on RA-DC. In cocultures, RA-DC increased IL10 and FOXP3 expression in Treg, but pre-treatment with L. reuteri-CFS did not further influence the Treg phenotype. In conclusion, L. reuteri-CFS modulates the phenotype and function of mucosal-like DC, implicating its potential application as postbiotic. PMID:27014275

  17. Protons Sensitize Epithelial Cells to Mesenchymal Transition

    PubMed Central

    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

  18. Respiratory epithelial cells orchestrate pulmonary innate immunity.

    PubMed

    Whitsett, Jeffrey A; Alenghat, Theresa

    2015-01-01

    The epithelial surfaces of the lungs are in direct contact with the environment and are subjected to dynamic physical forces as airway tubes and alveoli are stretched and compressed during ventilation. Mucociliary clearance in conducting airways, reduction of surface tension in the alveoli, and maintenance of near sterility have been accommodated by the evolution of a multi-tiered innate host-defense system. The biophysical nature of pulmonary host defenses are integrated with the ability of respiratory epithelial cells to respond to and 'instruct' the professional immune system to protect the lungs from infection and injury. PMID:25521682

  19. Interactions of bacterial pathogens with dendritic cells during invasion of mucosal surfaces.

    PubMed

    Granucci, Francesca; Ricciardi-Castagnoli, Paola

    2003-02-01

    Recent studies of mucosal immunity suggest a key role for dendritic cells in the regulation of gut immune responses, in both physiological and pathological conditions. Dendritic cells are widely distributed in the lamina propria of the gut and are involved in direct bacterial uptake across mucosal surfaces, which questions the role of dendritic cells in innate mucosal responses. Approximately 400 commensal microbial species are present in the gut lumen. So how do dendritic cells distinguish pathogens from luminal microflora? Are the cytokines and chemokines induced in dendritic cells tailored to the class of microbes being recognized? Several very important questions still need to be addressed. PMID:12615223

  20. Gastric mucosal cell proliferation in ethanol-induced chronic mucosal injury is related to oxidative stress and lipid peroxidation in rats.

    PubMed

    Hernández-Muñoz, R; Montiel-Ruíz, C; Vázquez-Martínez, O

    2000-08-01

    The oxygen free radicals-induced lipid peroxidation (LP) has been implicated in the pathogenesis of acute ethanol-induced gastric mucosal lesions. However, the role of LP in the generation of chronic gastric mucosal injury is unknown. We have developed a model of chronic mucosal injury induced by continuous ethanol ingestion for 5 days and characterized by marked alterations in plasma membranes from gastric mucosa. Therefore, LP was evaluated in this experimental model. Indicators of peroxidative activity, mucosal glutathione content, thymidine kinase activity (an index of cell proliferation), and histamine H2-receptor (H2R) binding constants were quantified in animals undergoing gastric mucosal damage. The effect of famotidine, a H2R antagonist that readily ameliorates the chronic mucosal injury, was also tested. Increased free radicals and LP levels were detected during gastritis; however, a second, higher peak of LP was noted in mucosal plasma membranes after ethanol withdrawal (recovery period). This further increase of LP coincided with active cell proliferation, decreased mucosal glutathione levels, and diminished specific cimetidine binding by H2R. Administration of famotidine accelerated the mucosal proliferative process, inducing the second lipoperoxidative episode sooner, and preserved the content of glutathione. In addition, LP correlated directly with cell proliferation and inversely with mucosal membrane cimetidine binding. In conclusion, LP seems to be involved in chronic ethanol-induced gastric mucosal injury. However, a further enhancement of plasma membrane LP occurred, associated with increased DNA synthesis and diminished cimetidine binding by membrane H2R. Therefore, increased LP could also participate in the compensatory mucosal proliferation initiated after ethanol withdrawal. PMID:10950107

  1. Role of Pht Proteins in Attachment of Streptococcus pneumoniae to Respiratory Epithelial Cells

    PubMed Central

    Kallio, Anna; Sepponen, Kirsi; Hermand, Philippe; Denoël, Philippe; Godfroid, Fabrice

    2014-01-01

    Pneumococcal adherence to mucosal surfaces is a critical step in nasopharyngeal colonization, but so far few pneumococcal adhesins involved in the interaction with host cells have been identified. PhtA, PhtB, PhtD, and PhtE are conserved pneumococcal surface proteins that have proven promising as vaccine candidates. One suggested virulence function of Pht proteins is to mediate adherence at the respiratory mucosa. In this study, we assessed the role of Pht proteins in pneumococcal binding to respiratory epithelial cells. Pneumococci were incubated with human nasopharyngeal epithelial cells (Detroit-562) and lung epithelial cells (A549 and NCI-H292), and the proportion of bound bacteria was measured by plating viable counts. Strains R36A (unencapsulated), D39 (serotype 2), 43 (serotype 3), 4-CDC (serotype 4), and 2737 (serotype 19F) with one or more of the four homologous Pht proteins deleted were compared with their wild-type counterparts. Also, the effect of anti-PhtD antibodies on the adherence of strain 2737 to the respiratory epithelial cells was studied. Our results suggest that Pht proteins play a role in pneumococcal adhesion to the respiratory epithelium. We also found that antibody to PhtD is able to inhibit bacterial attachment to the cells, suggesting that antibodies against PhtD present at mucosal surfaces might protect from pneumococcal attachment and subsequent colonization. However, the relative significance of Pht proteins to the ability of pneumococci to bind in vitro to epithelial cells depends on the genetic background and the capsular serotype of the strain. PMID:24491577

  2. Novel vaccine development strategies for inducing mucosal immunity

    PubMed Central

    Fujkuyama, Yoshiko; Tokuhara, Daisuke; Kataoka, Kosuke; Gilbert, Rebekah S; McGhee, Jerry R; Yuki, Yoshikazu; Kiyono, Hiroshi; Fujihashi, Kohtaro

    2012-01-01

    To develop protective immune responses against mucosal pathogens, the delivery route and adjuvants for vaccination are important. The host, however, strives to maintain mucosal homeostasis by responding to mucosal antigens with tolerance, instead of immune activation. Thus, induction of mucosal immunity through vaccination is a rather difficult task, and potent mucosal adjuvants, vectors or other special delivery systems are often used, especially in the elderly. By taking advantage of the common mucosal immune system, the targeting of mucosal dendritic cells and microfold epithelial cells may facilitate the induction of effective mucosal immunity. Thus, novel routes of immunization and antigen delivery systems also show great potential for the development of effective and safe mucosal vaccines against various pathogens. The purpose of this review is to introduce several recent approaches to induce mucosal immunity to vaccines, with an emphasis on mucosal tissue targeting, new immunization routes and delivery systems. Defining the mechanisms of mucosal vaccines is as important as their efficacy and safety, and in this article, examples of recent approaches, which will likely accelerate progress in mucosal vaccine development, are discussed. PMID:22380827

  3. Tuft-cell-derived IL-25 regulates an intestinal ILC2-epithelial response circuit.

    PubMed

    von Moltke, Jakob; Ji, Ming; Liang, Hong-Erh; Locksley, Richard M

    2016-01-14

    Parasitic helminths and allergens induce a type 2 immune response leading to profound changes in tissue physiology, including hyperplasia of mucus-secreting goblet cells and smooth muscle hypercontractility. This response, known as 'weep and sweep', requires interleukin (IL)-13 production by tissue-resident group 2 innate lymphoid cells (ILC2s) and recruited type 2 helper T cells (TH2 cells). Experiments in mice and humans have demonstrated requirements for the epithelial cytokines IL-33, thymic stromal lymphopoietin (TSLP) and IL-25 in the activation of ILC2s, but the sources and regulation of these signals remain poorly defined. In the small intestine, the epithelium consists of at least five distinct cellular lineages, including the tuft cell, whose function is unclear. Here we show that tuft cells constitutively express IL-25 to sustain ILC2 homeostasis in the resting lamina propria in mice. After helminth infection, tuft-cell-derived IL-25 further activates ILC2s to secrete IL-13, which acts on epithelial crypt progenitors to promote differentiation of tuft and goblet cells, leading to increased frequencies of both. Tuft cells, ILC2s and epithelial progenitors therefore comprise a response circuit that mediates epithelial remodelling associated with type 2 immunity in the small intestine, and perhaps at other mucosal barriers populated by these cells. PMID:26675736

  4. Tuft-cell-derived IL-25 regulates an intestinal ILC2–epithelial response circuit

    PubMed Central

    von Moltke, Jakob; Ji, Ming; Liang, Hong-Erh; Locksley, Richard M.

    2016-01-01

    Parasitic helminths and allergens induce a type 2 immune response leading to profound changes in tissue physiology, including hyperplasia of mucus-secreting goblet cells1 and smooth muscle hypercontractility2. This response, known as ‘weep and sweep’, requires interleukin (IL)-13 production by tissue-resident group 2 innate lymphoid cells (ILC2s) and recruited type 2 helper T cells (TH2 cells)3. Experiments in mice and humans have demonstrated requirements for the epithelial cytokines IL-33, thymic stromal lymphopoietin (TSLP) and IL-25 in the activation of ILC2s4–11, but the sources and regulation of these signals remain poorly defined. In the small intestine, the epithelium consists of at least five distinct cellular lineages12, including the tuft cell, whose function is unclear. Here we show that tuft cells constitutively express IL-25 to sustain ILC2 homeostasis in the resting lamina propria in mice. After helminth infection, tuft-cell-derived IL-25 further activates ILC2s to secrete IL-13, which acts on epithelial crypt progenitors to promote differentiation of tuft and goblet cells, leading to increased frequencies of both. Tuft cells, ILC2s and epithelial progenitors therefore comprise a response circuit that mediates epithelial remodelling associated with type 2 immunity in the small intestine, and perhaps at other mucosal barriers populated by these cells. PMID:26675736

  5. Endoplasmic Reticulum Stress in Intestinal Epithelial Cell Function and Inflammatory Bowel Disease

    PubMed Central

    Luo, Katherine; Cao, Stewart Siyan

    2015-01-01

    In eukaryotic cells, perturbation of protein folding homeostasis in the endoplasmic reticulum (ER) causes accumulation of unfolded and misfolded proteins in the ER lumen, which activates intracellular signaling pathways termed the unfolded protein response (UPR). Recent studies have linked ER stress and the UPR to inflammatory bowel disease (IBD). The microenvironment of the ER is affected by a myriad of intestinal luminal molecules, implicating ER stress and the UPR in proper maintenance of intestinal homeostasis. Several intestinal cell populations, including Paneth and goblet cells, require robust ER function for protein folding, maturation, and secretion. Prolonged ER stress and impaired UPR signaling may cause IBD through: (1) induction of intestinal epithelial cell apoptosis, (2) disruption of mucosal barrier function, and (3) induction of the proinflammatory response in the gut. Based on our increased understanding of ER stress in IBD, new pharmacological approaches can be developed to improve intestinal homeostasis by targeting ER protein-folding in the intestinal epithelial cells (IECs). PMID:25755668

  6. Antigen targeting to M cells for enhancing the efficacy of mucosal vaccines

    PubMed Central

    Kim, Sae-Hae; Jang, Yong-Suk

    2014-01-01

    Vaccination is one of the most successful applications of immunology and for a long time has depended on parenteral administration protocols. However, recent studies have pointed to the promise of mucosal vaccination because of its ease, economy and efficiency in inducing an immune response not only systemically, but also in the mucosal compartment where many pathogenic infections are initiated. However, successful mucosal vaccination requires the help of an adjuvant for the efficient delivery of vaccine material into the mucosa and the breaking of the tolerogenic environment, especially in oral mucosal immunization. Given that M cells are the main gateway to take up luminal antigens and initiate antigen-specific immune responses, understanding the role and characteristics of M cells is crucial for the development of successful mucosal vaccines. Especially, particular interest has been focused on the regulation of the tolerogenic mucosal microenvironment and the introduction of the luminal antigen into the lymphoid organ by exploiting the molecules of M cells. Here, we review the characteristics of M cells and the immune regulatory factors in mucosa that can be exploited for mucosal vaccine delivery and mucosal immune regulation. PMID:24626171

  7. Down-regulation of mechanisms involved in cell transport and maintenance of mucosal integrity in pigs infected with Lawsonia intracellularis

    PubMed Central

    2014-01-01

    Lawsonia intracellularis is an obligate intracellular bacterium, responsible for the disease complex known as proliferative enteropathy (PE). L. intracellularis is associated with intestinal crypt epithelial cell proliferation but the mechanisms responsible are yet to be defined. Microarray analysis was used to investigate the host-pathogen interaction in experimentally infected pigs to identify pathways that may be involved. Ileal samples originating from twenty-eight weaner pigs experimentally challenged with a pure culture of L. intracellularis (strain LR189/5/83) were subjected to microarray analysis. Microarray transcriptional signatures were validated using immunohistochemistry and quantitative real time PCR of selected genes at various time points post challenge. At peak of infection (14 days post challenge) 86% of altered transcripts were down regulated, particularly those involved in maintenance of mucosal integrity and regulation of cell transport. Among the up-regulated transcripts, CD163 and CDK1 were novel findings and considered to be important, due to their respective roles in innate immunity and cellular proliferation. Overall, targeted cellular mechanisms included those that are important in epithelial restitution, migration and protection; maintenance of stable inter-epithelial cell relationships; cell transport of nutrients and electrolytes; innate immunity; and cell cycle. PMID:24885874

  8. Down-regulation of mechanisms involved in cell transport and maintenance of mucosal integrity in pigs infected with Lawsonia intracellularis.

    PubMed

    Smith, Sionagh H; Wilson, Alison D; Van Ettinger, Imke; MacIntyre, Neil; Archibald, Alan L; Ait-Ali, Tahar

    2014-01-01

    Lawsonia intracellularis is an obligate intracellular bacterium, responsible for the disease complex known as proliferative enteropathy (PE). L. intracellularis is associated with intestinal crypt epithelial cell proliferation but the mechanisms responsible are yet to be defined. Microarray analysis was used to investigate the host-pathogen interaction in experimentally infected pigs to identify pathways that may be involved. Ileal samples originating from twenty-eight weaner pigs experimentally challenged with a pure culture of L. intracellularis (strain LR189/5/83) were subjected to microarray analysis. Microarray transcriptional signatures were validated using immunohistochemistry and quantitative real time PCR of selected genes at various time points post challenge. At peak of infection (14 days post challenge) 86% of altered transcripts were down regulated, particularly those involved in maintenance of mucosal integrity and regulation of cell transport. Among the up-regulated transcripts, CD163 and CDK1 were novel findings and considered to be important, due to their respective roles in innate immunity and cellular proliferation. Overall, targeted cellular mechanisms included those that are important in epithelial restitution, migration and protection; maintenance of stable inter-epithelial cell relationships; cell transport of nutrients and electrolytes; innate immunity; and cell cycle. PMID:24885874

  9. Mycoplasma genitalium promotes epithelial crossing and peripheral blood mononuclear cell infection by HIV-1

    PubMed Central

    Das, Kishore; De la Garza, Georgina; Siwak, Edward B.; Scofield, Virginia L.; Dhandayuthapani, Subramanian

    2014-01-01

    Summary Background Mycoplasma genitalium co-infection in HIV-infected individuals has been reported to increase the shedding of HIV in the urogenital region of females. To better understand this relationship, we investigated the influence of M. genitalium on the transmission and replication of HIV using an in vitro model. Methods The Transwell co-culture system was employed to assess the crossing of an endocervical cell barrier by HIV-1. Immunocytochemistry and confocal microscopy were used to assess the distribution of the nectin-1 molecule on M. genitalium-infected epithelial cells of the End1/E6E7 endocervical cell line, grown as monolayers in the insert wells. Peripheral blood mononuclear cells (PBMC) were cultured in the bottom wells to assess the effects of M. genitalium, passing through the semipermeable culturing membrane, on subsequent HIV infection of susceptible target cells. Results Infection of the endocervical cells with the adhesion-positive M. genitalium G37 strain (wild-type) significantly elevated the passage of HIV across the epithelial cell barrier relative to HIV transfer across endocervical cells infected with the adhesion-negative M. genitalium JB1 strain. Immunostaining of the M. genitalium-G37-infected epithelial cells disclosed capping and internalization of the junctional regulatory protein nectin-1, in association with reduced transepithelial resistance (TER) in the cell monolayer. When PBMC were cultured beneath insert wells containing M. genitalium-G37-infected epithelial cell monolayers, we observed significantly enhanced infectivity and replication of HIV added afterward to the cultures. Conclusions M. genitalium influences events on both sides of a cultured mucosal epithelial monolayer: (1) by infecting the epithelial cells and reducing the integrity of the barrier itself, and (2) by activating HIV target cells below it, thereby promoting HIV infection and progeny virus production. PMID:24661929

  10. Esophageal epithelial cells acquire functional characteristics of activated myofibroblasts after undergoing an epithelial to mesenchymal transition

    PubMed Central

    Muir, Amanda B.; Dods, Kara; Noah, Yuli; Toltzis, Sarit; Chandramouleeswaran, Prasanna Modayur; Lee, Anna; Benitez, Alain; Bedenbaugh, Adam; Falk, Gary W.; Wells, Rebecca G.; Nakagawa, Hiroshi; Wang, Mei-Lun

    2015-01-01

    Background and Aims Eosinophilic esophagitis (EoE) is an allergic inflammatory disease that leads to esophageal fibrosis and stricture. We have recently shown that in EoE, esophageal epithelial cells undergo an epithelial to mesenchymal transition (EMT), characterized by gain of mesenchymal markers and loss of epithelial gene expression. Whether epithelial cells exposed to profibrotic cytokines can also acquire the functional characteristics of activated myofibroblasts, including migration, contraction, and extracellular matrix deposition, is relevant to our understanding and treatment of EoE-associated fibrogenesis. In the current study, we characterize cell migration, contraction, and collagen production by esophageal epithelial cells that have undergone cytokine-induced EMT in vitro. Methods and Results Stimulation of human non-transformed immortalized esophageal epithelial cells (EPC2-hTERT) with profibrotic cytokines TNFα, TGFβ, and IL1β for three weeks led to acquisition of mesenchymal αSMA and vimentin, and loss of epithelial E-cadherin expression. Upon removal of the profibrotic stimulus, epithelial characteristics were partially rescued. TGFβ stimulation had a robust effect upon epithelial collagen production. Surprisingly, TNFα stimulation had the most potent effect upon cell migration and contraction, exceeding the effects of the prototypical profibrotic cytokine TGFβ. IL1β stimulation alone had minimal effect upon esophageal epithelial migration, contraction, and collagen production. Conclusions Esophageal epithelial cells that have undergone EMT acquire functional characteristics of activated myofibroblasts in vitro. Profibrotic cytokines exert differential effects upon esophageal epithelial cells, underscoring complexities of fibrogenesis in EoE, and implicating esophageal epithelial cells as effector cells in EoE-associated fibrogenesis. PMID:25183431

  11. Loss of Survivin in Intestinal Epithelial Progenitor Cells Leads to Mitotic Catastrophe and Breakdown of Gut Immune Homeostasis.

    PubMed

    Martini, Eva; Wittkopf, Nadine; Günther, Claudia; Leppkes, Moritz; Okada, Hitoshi; Watson, Alastair J; Podstawa, Eva; Backert, Ingo; Amann, Kerstin; Neurath, Markus F; Becker, Christoph

    2016-02-01

    A tightly regulated balance of proliferation and cell death of intestinal epithelial cells (IECs) is essential for maintenance of gut homeostasis. Survivin is highly expressed during embryogenesis and in several cancer types, but little is known about its role in adult gut tissue. Here, we show that Survivin is specifically expressed in transit-amplifying cells and Lgr5(+) stem cells. Genetic loss of Survivin in IECs resulted in destruction of intestinal integrity, mucosal inflammation, and death of the animals. Survivin deletion was associated with decreased epithelial proliferation due to defective chromosomal segregation. Moreover, Survivin-deficient animals showed induced phosphorylation of p53 and H2AX and increased levels of cell-intrinsic apoptosis in IECs. Consequently, induced deletion of Survivin in Lgr5(+) stem cells led to cell death. In summary, Survivin is a key regulator of gut tissue integrity by regulating epithelial homeostasis in the stem cell niche. PMID:26832409

  12. DNA repair and mutagen sensitivity of epithelial cells and lymphocytes in oropharyngeal cancer

    PubMed Central

    REITER, MAXIMILIAN; BAUMEISTER, PHILIPP; JAISER, SONJA; REISS, ANDREAS; SCHWENK-ZIEGER, SABINA; KLEINSASSER, NORBERT; HARRÉUS, ULRICH

    2012-01-01

    Tobacco-associated nitrosamines are known carcinogens causing DNA damage in epithelial cells of the head and neck. A matched case-control study was performed to evaluate the sensitivity of patients with squamous cell cancer (SCC) of the oropharynx, and controls to tobacco-associated nitrosamines. Quantitative DNA repair was evaluated following a period of 15 and 30 min. Fresh biopsies from 100 male donors of macroscopically healthy oropharyngeal cells and lymphocytes (50 SCC patients and 50 controls) were incubated with N-nitrosodiethylamine (NDEA), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) or N-nitrosonornicotine (NNN). DNA damage in epithelial cells and lymphocytes was assessed using the comet assay. Following incubation with NDEA, cells underwent a period of DNA repair. All of the nitrosamines caused equivalent genotoxic damage in mucosal cells and lymphocytes of the two groups. Lymphocyte DNA repair capacity in the control group (26.8 and 37.1% after 15 and 30 min) was comparable to the tumor group (23.6 and 40.6%). However, epithelial cell DNA repair capacity of carcinoma patients was significantly reduced to 17.1% (15 min) and 23% (30 min) compared to the DNA repair of the control group (36.2%, 15 min and 46.0%, 30 min). Mutagen sensitivity was comparable in patients and controls. Thus, reduced epithelial cell DNA repair capacity of tumor patients is a possible endogenous risk factor for the development of head and neck squamous cell cancer. PMID:22740863

  13. EDAC: Epithelial defence against cancer-cell competition between normal and transformed epithelial cells in mammals.

    PubMed

    Kajita, Mihoko; Fujita, Yasuyuki

    2015-07-01

    During embryonic development or under certain pathological conditions, viable but suboptimal cells are often eliminated from the cellular society through a process termed cell competition. Cell competition was originally identified in Drosophila where cells with different properties compete for survival; 'loser' cells are eliminated from tissues and consequently 'winner' cells become dominant. Recent studies have shown that cell competition also occurs in mammals. While apoptotic cell death is the major fate for losers in Drosophila, outcompeted cells show more variable phenotypes in mammals, such as cell death-independent apical extrusion and cellular senescence. Molecular mechanisms underlying these processes have been recently revealed. Especially, in epithelial tissues, normal cells sense and actively eliminate the neighbouring transformed cells via cytoskeletal proteins by the process named epithelial defence against cancer (EDAC). Here, we introduce this newly emerging research field: cell competition in mammals. PMID:25991731

  14. Uroepithelial cells are part of a mucosal cytokine network.

    PubMed Central

    Hedges, S; Agace, W; Svensson, M; Sjögren, A C; Ceska, M; Svanborg, C

    1994-01-01

    This study compared the cytokine production of uroepithelial cell lines in response to gram-negative bacteria and inflammatory cytokines. Human kidney (A498) and bladder (J82) epithelial cell lines were stimulated with either Escherichia coli Hu734, interleukin 1 alpha (IL-1 alpha), or tumor necrosis factor alpha (TNF-alpha). Supernatant samples were removed, and the RNA was extracted from cells at 0, 2, 6, and 24 h. The secreted cytokine levels were determined by bioassay or immunoassay; mRNA was examined by reverse transcription-PCR. The two cell lines secreted IL-6 and IL-8 constitutively. IL-6 and IL-8 mRNA were constitutively produced in both cell lines; IL-1 beta mRNA was detected in J82 cells. IL-1 alpha induced significantly higher levels of IL-6 secretion than did E. coli Hu734 or TNF-alpha. IL-1 alpha and TNF-alpha induced significantly higher levels of IL-8 secretion than did E. coli Hu734. Secreted IL-1 beta was not detected; IL-1 alpha and TNF-alpha were not detected above the levels used for stimulation. IL-1 alpha, IL-1 beta, IL-6, and IL-8 mRNAs were detected in both cell lines after exposure to the stimulants. TNF-alpha mRNA was occasionally detected in the J82 cell line after TNF-alpha stimulation. Cytokine (IL-6 and IL-8) and control (glyceraldehyde 3-phosphate dehydrogenase [G3PDH] and beta-actin) mRNA concentrations were quantitated with internal PCR standards. Cytokine mRNA levels relative to beta-actin mRNA levels were the highest in E. coli-stimulated cells. In comparison, the cytokine mRNA levels relative to G3PDH mRNA levels were the highest in IL-1 alpha-stimulated cells. beta-Actin mRNA levels decreased after bacterial stimulation but not after cytokine stimulation, while G3PDH mRNA levels increased in response to all of the stimulants tested. These results suggested that E. coli Hu734 lowered the beta-actin mRNA levels in uroepithelial cells, thus distorting the IL-6 and IL-8 mRNA levels relative to this control. In summary, E. coli IL

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

    SciTech Connect

    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.

  16. Catecholamine-Directed Epithelial Cell Interactions with Bacteria in the Intestinal Mucosa.

    PubMed

    Brown, David R

    2016-01-01

    The catecholamines epinephrine, norepinephrine and dopamine are present in or have access to mucous membranes in the digestive, respiratory and genitourinary tracts, which represent the first sites of microbial colonization and infection within the body. Epithelial cells at mucosal surfaces establish and maintain symbiotic microbial communities and serve as the initial cellular point of contact for pathogens with the animal host. These cells express receptors that are capable of detecting and responding to microbe-associated molecular patterns and in most host species express G protein-coupled receptors for catecholamines. Although it is increasingly recognized that substances produced and released from nerves and endocrine cells can exert immuno-modulatory actions at mucosal sites, there have been few investigations focused specifically on the catecholaminergic modulation of interactions between the mucosal epithelium and bacteria or other mucosa-associated microorganisms. The potential biomedical importance of this phenomenon cannot be understated. For example, psychological stress or other conditions that activate the sympathetic nervous system to release epinephrine and norepinephrine may act to produce short-term changes in luminal and mucosal microbial communities or alter the course of a bacterial infection. This chapter will briefly review this developing and important research area of mucosa-microbe interactions with a focus on intestinal host defense. PMID:26589214

  17. In vitro modeling of rat mucosal mast cell function in Trichinella spiralis infection

    PubMed Central

    Thrasher, Seana M.; Scalfone, Lisa K.; Holowka, David; Appleton, Judith A.

    2012-01-01

    Summary Intestinal infection with the parasitic nematode, Trichinella spiralis, provides a robust context for the study of mucosal mast cell function. In rats, mucosal mast cells are exposed to parasites during the earliest stage of infection, affording an opportunity for mast cells to contribute to an innate response to infection. During secondary infection, degranulation of rat mucosal mast cells coincides with expulsion of challenge larvae from the intestine. The goal of this study was to evaluate rat bone marrow-derived mast cells (BMMC) and the rat basophilic leukemia cell line (RBL-2H3) as models for mucosal mast cells, using parasite glycoproteins and antibody reagents that have been tested extensively in rats in vivo. We found that BMMC displayed a more robust mucosal phenotype. Although T. spiralis glycoproteins bound to mast cell surfaces in the absence of antibodies, they did not stimulate degranulation, nor did they inhibit degranulation triggered by immune complexes. Parasite glycoproteins complexed with specific monoclonal IgGs provoked release of RMCPII and β-hexosaminidase from both cell types in a manner that replicated results observed previously in passively immunized rats. Our results document that RBL-2H3 cells and BMMC model rat mucosal mast cells in the contexts of innate and adaptive responses to T. spiralis. PMID:23094823

  18. Annexin 2 Regulates Intestinal Epithelial Cell Spreading and Wound Closure through Rho-Related Signaling

    PubMed Central

    Babbin, Brian A.; Parkos, Charles A.; Mandell, Kenneth J.; Winfree, L. Matthew; Laur, Oskar; Ivanov, Andrei I.; Nusrat, Asma

    2007-01-01

    Epithelial cell migration is a critical event in gastrointestinal mucosal wound healing and is dependent on actin cytoskeletal reorganization. We observed increased expression of an actin regulatory protein, annexin 2, in migrating intestinal epithelial cells. Small interfering RNA (siRNA)-mediated knockdown of annexin 2 expression in Caco-2 epithelial cells resulted in significant reductions in cell spreading and wound closure associated with decreased formation of filamentous actin bundles along the base of migrating cells. Because annexin 2 has been shown to influences actin cytoskeletal remodeling through targeting signaling molecules to membrane domains, we examined the membrane association and activation status of Rho GTPases after annexin 2 knockdown. We observed Rho dissociation from membranes and decreased Rho activity following annexin 2 siRNA transfection. Inhibition of cell spreading and wound closure in annexin 2 siRNA-transfected cells was prevented by expression of constitutively active RhoA. Rho colocalized with annexin 2 in lamellipodia and along the cytoplasmic face of the plasma membrane. In addition, annexin 2 was observed to co-immunoprecipitate with endogenous Rho and constitutively active RhoA. These findings suggest that annexin 2 plays a role in targeting Rho to cellular membranes, thereby modulating Rho-related signaling events regulating cytoskeletal reorganization during epithelial cell migration. PMID:17322380

  19. Modeling mucosal candidiasis in larval zebrafish by swimbladder injection.

    PubMed

    Gratacap, Remi L; Bergeron, Audrey C; Wheeler, Robert T

    2014-01-01

    Early defense against mucosal pathogens consists of both an epithelial barrier and innate immune cells. The immunocompetency of both, and their intercommunication, are paramount for the protection against infections. The interactions of epithelial and innate immune cells with a pathogen are best investigated in vivo, where complex behavior unfolds over time and space. However, existing models do not allow for easy spatio-temporal imaging of the battle with pathogens at the mucosal level. The model developed here creates a mucosal infection by direct injection of the fungal pathogen, Candida albicans, into the swimbladder of juvenile zebrafish. The resulting infection enables high-resolution imaging of epithelial and innate immune cell behavior throughout the development of mucosal disease. The versatility of this method allows for interrogation of the host to probe the detailed sequence of immune events leading to phagocyte recruitment and to examine the roles of particular cell types and molecular pathways in protection. In addition, the behavior of the pathogen as a function of immune attack can be imaged simultaneously by using fluorescent protein-expressing C. albicans. Increased spatial resolution of the host-pathogen interaction is also possible using the described rapid swimbladder dissection technique. The mucosal infection model described here is straightforward and highly reproducible, making it a valuable tool for the study of mucosal candidiasis. This system may also be broadly translatable to other mucosal pathogens such as mycobacterial, bacterial or viral microbes that normally infect through epithelial surfaces. PMID:25490695

  20. Modeling Mucosal Candidiasis in Larval Zebrafish by Swimbladder Injection

    PubMed Central

    Gratacap, Remi L.; Bergeron, Audrey C.; Wheeler, Robert T.

    2016-01-01

    Early defense against mucosal pathogens consists of both an epithelial barrier and innate immune cells. The immunocompetency of both, and their intercommunication, are paramount for the protection against infections. The interactions of epithelial and innate immune cells with a pathogen are best investigated in vivo, where complex behavior unfolds over time and space. However, existing models do not allow for easy spatio-temporal imaging of the battle with pathogens at the mucosal level. The model developed here creates a mucosal infection by direct injection of the fungal pathogen, Candida albicans, into the swimbladder of juvenile zebrafish. The resulting infection enables high-resolution imaging of epithelial and innate immune cell behavior throughout the development of mucosal disease. The versatility of this method allows for interrogation of the host to probe the detailed sequence of immune events leading to phagocyte recruitment and to examine the roles of particular cell types and molecular pathways in protection. In addition, the behavior of the pathogen as a function of immune attack can be imaged simultaneously by using fluorescent protein-expressing C. albicans. Increased spatial resolution of the host-pathogen interaction is also possible using the described rapid swimbladder dissection technique. The mucosal infection model described here is straightforward and highly reproducible, making it a valuable tool for the study of mucosal candidiasis. This system may also be broadly translatable to other mucosal pathogens such as mycobacterial, bacterial or viral microbes that normally infect through epithelial surfaces. PMID:25490695

  1. DNA typing of epithelial cells after strangulation.

    PubMed

    Wiegand, P; Kleiber, M

    1997-01-01

    DNA typing was carried out on epithelial cells which were transferred from the hands of the suspect onto the neck of the victim. In an experimental study 16 suspect-victim combinations were investigated for estimating the typing success. Alternatively to an attack against the neck, the upper arm was used for "strangulation". PCR typing was carried out using the short tandem repeat systems (STRs) HumCD4, HumVWF31A (VWA) and Hum-FIBRA (FGA) and the success rate was > 70% for all 3 systems. In most of the cases mixed patterns containing the phenotype of the suspect and the victim were obtained. In a case where strangulation was the cause of death, epithelial cells could be removed from the neck of the victim. The DNA pattern of the suspect could be successfully amplified using four STRs, demonstrating the applicability of this approach for practical casework. PMID:9274940

  2. A distinct array of proinflammatory cytokines is expressed in human colon epithelial cells in response to bacterial invasion.

    PubMed Central

    Jung, H C; Eckmann, L; Yang, S K; Panja, A; Fierer, J; Morzycka-Wroblewska, E; Kagnoff, M F

    1995-01-01

    Pathogenic bacteria that penetrate the intestinal epithelial barrier stimulate an inflammatory response in the adjacent intestinal mucosa. The present studies asked whether colon epithelial cells can provide signals that are important for the initiation and amplification of an acute mucosal inflammatory response. Infection of monolayers of human colon epithelial cell lines (T84, HT29, Caco-2) with invasive strains of bacteria (Salmonella dublin, Shigella dysenteriae, Yersinia enterocolitica, Listeria monocytogenes, enteroinvasive Escherichia coli) resulted in the coordinate expression and upregulation of a specific array of four proinflammatory cytokines, IL-8, monocyte chemotactic protein-1, GM-CSF, and TNF alpha, as assessed by mRNA levels and cytokine secretion. Expression of the same cytokines was upregulated after TNF alpha or IL-1 stimulation of these cells. In contrast, cytokine gene expression was not altered after infection of colon epithelial cells with noninvasive bacteria or the noninvasive protozoan parasite, G. lamblia. Notably, none of the cell lines expressed mRNA for IL-2, IL-4, IL-5, IL-6, IL-12p40, IFN-gamma, or significant levels of IL-1 or IL-10 in response to the identical stimuli. The coordinate expression of IL-8, MCP-1, GM-CSF and TNF alpha appears to be a general property of human colon epithelial cells since an identical array of cytokines, as well as IL-6, also was expressed by freshly isolated human colon epithelial cells. Since the cytokines expressed in response to bacterial invasion or other proinflammatory agonists have a well documented role in chemotaxis and activation of inflammatory cells, colon epithelial cells appear to be programmed to provide a set of signals for the activation of the mucosal inflammatory response in the earliest phases after microbial invasion. Images PMID:7814646

  3. Expression of Opacity Proteins Interferes with the Transmigration of Neisseria gonorrhoeae across Polarized Epithelial Cells.

    PubMed

    Stein, Daniel C; LeVan, Adriana; Hardy, Britney; Wang, Liang-Chun; Zimmerman, Lindsey; Song, Wenxia

    2015-01-01

    Neisseria gonorrhoeae (GC) establishes infection at the mucosal surface of the human genital tract, most of which is lined with polarized epithelial cells. GC can cause localized as well as disseminated infections, leading to various complications. GC constantly change their surface structures via phase and antigenic variation, which has been implicated as a means for GC to establish infection at various anatomic locations of male and female genital tracks. However, the exact contribution of each surface molecule to bacterial infectivity remains elusive due to their phase variation. Using a GC derivative that is genetically devoid of all opa genes (MS11∆Opa), this study shows that Opa expression interferes with GC transmigration across polarized human epithelial cells. MS11∆Opa transmigrates across polarized epithelial cells much faster and to a greater extent than MS11Opa+, while adhering at a similar level as MS11Opa+. When MS11Opa+, able to phase vary Opa expression, was inoculated, only those bacteria that turn off Opa expression transmigrate across the polarized epithelial monolayer. Similar to bacteria alone or co-cultured with non-polarized epithelial cells, MS11∆Opa fails to form large microcolonies at the apical surface of polarized epithelial cells. Apical inoculation of MS11Opa+, but not MS11∆Opa, induces the recruitment of the Opa host-cell receptor carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) to the apical junction and the vicinity of bacterial adherent sites. Our results suggest that Opa expression limits gonococcal ability to invade into subepithelial tissues by forming tight interactions with neighboring bacteria and by inducing CEACAM redistribution to cell junctions. PMID:26244560

  4. Expression of Opacity Proteins Interferes with the Transmigration of Neisseria gonorrhoeae across Polarized Epithelial Cells

    PubMed Central

    Stein, Daniel C.; LeVan, Adriana; Hardy, Britney; Wang, Liang-Chun; Zimmerman, Lindsey; Song, Wenxia

    2015-01-01

    Neisseria gonorrhoeae (GC) establishes infection at the mucosal surface of the human genital tract, most of which is lined with polarized epithelial cells. GC can cause localized as well as disseminated infections, leading to various complications. GC constantly change their surface structures via phase and antigenic variation, which has been implicated as a means for GC to establish infection at various anatomic locations of male and female genital tracks. However, the exact contribution of each surface molecule to bacterial infectivity remains elusive due to their phase variation. Using a GC derivative that is genetically devoid of all opa genes (MS11∆Opa), this study shows that Opa expression interferes with GC transmigration across polarized human epithelial cells. MS11∆Opa transmigrates across polarized epithelial cells much faster and to a greater extent than MS11Opa+, while adhering at a similar level as MS11Opa+. When MS11Opa+, able to phase vary Opa expression, was inoculated, only those bacteria that turn off Opa expression transmigrate across the polarized epithelial monolayer. Similar to bacteria alone or co-cultured with non-polarized epithelial cells, MS11∆Opa fails to form large microcolonies at the apical surface of polarized epithelial cells. Apical inoculation of MS11Opa+, but not MS11∆Opa, induces the recruitment of the Opa host-cell receptor carcinoembryonic antigen–related cell adhesion molecules (CEACAMs) to the apical junction and the vicinity of bacterial adherent sites. Our results suggest that Opa expression limits gonococcal ability to invade into subepithelial tissues by forming tight interactions with neighboring bacteria and by inducing CEACAM redistribution to cell junctions. PMID:26244560

  5. Mechanisms of Innate Lymphoid Cell and Natural Killer T Cell Activation during Mucosal Inflammation

    PubMed Central

    Altmayer, Nora

    2014-01-01

    Mucosal surfaces in the airways and the gastrointestinal tract are critical for the interactions of the host with its environment. Due to their abundance at mucosal tissue sites and their powerful immunomodulatory capacities, the role of innate lymphoid cells (ILCs) and natural killer T (NKT) cells in the maintenance of mucosal tolerance has recently moved into the focus of attention. While NKT cells as well as ILCs utilize distinct transcription factors for their development and lineage diversification, both cell populations can be further divided into three polarized subpopulations reflecting the distinction into Th1, Th2, and Th17 cells in the adaptive immune system. While bystander activation through cytokines mediates the induction of ILC and NKT cell responses, NKT cells become activated also through the engagement of their canonical T cell receptors (TCRs) by (glyco)lipid antigens (cognate recognition) presented by the atypical MHC I like molecule CD1d on antigen presenting cells (APCs). As both innate lymphocyte populations influence inflammatory responses due to the explosive release of copious amounts of different cytokines, they might represent interesting targets for clinical intervention. Thus, we will provide an outlook on pathways that might be interesting to evaluate in this context. PMID:24987710

  6. Control of local immunity by airway epithelial cells.

    PubMed

    Weitnauer, M; Mijošek, V; Dalpke, A H

    2016-03-01

    The lung is ventilated by thousand liters of air per day. Inevitably, the respiratory system comes into contact with airborne microbial compounds, most of them harmless contaminants. Airway epithelial cells are known to have innate sensor functions, thus being able to detect microbial danger. To avoid chronic inflammation, the pulmonary system has developed specific means to control local immune responses. Even though airway epithelial cells can act as proinflammatory promoters, we propose that under homeostatic conditions airway epithelial cells are important modulators of immune responses in the lung. In this review, we discuss epithelial cell regulatory functions that control reactivity of professional immune cells within the microenvironment of the airways and how these mechanisms are altered in pulmonary diseases. Regulation by epithelial cells can be divided into two mechanisms: (1) mediators regulate epithelial cells' innate sensitivity in cis and (2) factors are produced that limit reactivity of immune cells in trans. PMID:26627458

  7. Generation of Mouse Lung Epithelial Cells

    PubMed Central

    Kasinski, Andrea L.; Slack, Frank J.

    2016-01-01

    Although in vivo models are excellent for assessing various facets of whole organism physiology, pathology, and overall response to treatments, evaluating basic cellular functions, and molecular events in mammalian model systems is challenging. It is therefore advantageous to perform these studies in a refined and less costly setting. One approach involves utilizing cells derived from the model under evaluation. The approach to generate such cells varies based on the cell of origin and often the genetics of the cell. Here we describe the steps involved in generating epithelial cells from the lungs of KrasLSL-G12D/+; p53LSL-R172/+ mice (Kasinski and Slack, 2012). These mice develop aggressive lung adenocarcinoma following cre-recombinase dependent removal of a stop cassette in the transgenes and subsequent expression of Kra-G12D and p53R172. While this protocol may be useful for the generation of epithelial lines from other genetic backgrounds, it should be noted that the Kras; p53 cell line generated here is capable of proliferating in culture without any additional genetic manipulation that is often needed for less aggressive backgrounds.

  8. Palifermin in Preventing Oral Mucositis Caused by Chemotherapy and/or Radiation Therapy in Young Patients Undergoing Stem Cell Transplant

    ClinicalTrials.gov

    2013-05-30

    Breast Cancer; Graft Versus Host Disease; Kidney Cancer; Leukemia; Lymphoma; Mucositis; Multiple Myeloma; Plasma Cell Neoplasm; Myelodysplastic Syndromes; Neuroblastoma; Ovarian Cancer; Sarcoma; Testicular Germ Cell Tumor

  9. Transcriptional Landscape of Glomerular Parietal Epithelial Cells

    PubMed Central

    Gharib, Sina A.; Pippin, Jeffrey W.; Ohse, Takamoto; Pickering, Scott G.; Krofft, Ronald D.; Shankland, Stuart J.

    2014-01-01

    Very little is known about the function of glomerular parietal epithelial cells (PECs). In this study, we performed genome-wide expression analysis on PEC-enriched capsulated vs. PEC-deprived decapsulated rat glomeruli to determine the transcriptional state of PECs under normal conditions. We identified hundreds of differentially expressed genes that mapped to distinct biologic modules including development, tight junction, ion transport, and metabolic processes. Since developmental programs were highly enriched in PECs, we characterized several of their candidate members at the protein level. Collectively, our findings confirm that PECs are multifaceted cells and help define their diverse functional repertoire. PMID:25127402

  10. Reduced Epithelial Na+/H+ Exchange Drives Gut Microbial Dysbiosis and Promotes Inflammatory Response in T Cell-Mediated Murine Colitis.

    PubMed

    Laubitz, Daniel; Harrison, Christy A; Midura-Kiela, Monica T; Ramalingam, Rajalakshmy; Larmonier, Claire B; Chase, John H; Caporaso, J Gregory; Besselsen, David G; Ghishan, Fayez K; Kiela, Pawel R

    2016-01-01

    Inflammatory bowel diseases (IBD) are associated with functional inhibition of epithelial Na+/H+ exchange. In mice, a selective disruption of NHE3 (Slc9a3), a major apical Na+/H+ exchanger, also promotes IBD-like symptoms and gut microbial dysbiosis. We hypothesized that disruption of Na+/H+ exchange is necessary for the development of dysbiosis, which promotes an exacerbated mucosal inflammatory response. Therefore, we performed a temporal analysis of gut microbiota composition, and mucosal immune response to adoptive T cell transfer was evaluated in Rag2-/- and NHE3-/-/Rag2-/- (DKO) mice with and without broad-spectrum antibiotics. Microbiome (16S profiling), colonic histology, T cell and neutrophil infiltration, mucosal inflammatory tone, and epithelial permeability were analyzed. In adoptive T cell transfer colitis model, Slc9a3 status was the most significant determinant of gut microbial community. In DKO mice, NHE3-deficiency and dysbiosis were associated with dramatically accelerated and exacerbated disease, with rapid body weight loss, increased mucosal T cell and neutrophil influx, increased mucosal cytokine expression, increased permeability, and expansion of CD25-FoxP3+ Tregs; this enhanced susceptibility was alleviated by oral broad-spectrum antibiotics. Based on these results and our previous work, we postulate that epithelial electrolyte homeostasis is an important modulator in the progression of colitis, acting through remodeling of the gut microbial community. PMID:27050757

  11. Reduced Epithelial Na+/H+ Exchange Drives Gut Microbial Dysbiosis and Promotes Inflammatory Response in T Cell-Mediated Murine Colitis

    PubMed Central

    Midura-Kiela, Monica T.; Ramalingam, Rajalakshmy; Larmonier, Claire B.; Chase, John H.; Caporaso, J. Gregory; Besselsen, David G.; Ghishan, Fayez K.; Kiela, Pawel R.

    2016-01-01

    Inflammatory bowel diseases (IBD) are associated with functional inhibition of epithelial Na+/H+ exchange. In mice, a selective disruption of NHE3 (Slc9a3), a major apical Na+/H+ exchanger, also promotes IBD-like symptoms and gut microbial dysbiosis. We hypothesized that disruption of Na+/H+ exchange is necessary for the development of dysbiosis, which promotes an exacerbated mucosal inflammatory response. Therefore, we performed a temporal analysis of gut microbiota composition, and mucosal immune response to adoptive T cell transfer was evaluated in Rag2-/- and NHE3-/-/Rag2-/- (DKO) mice with and without broad-spectrum antibiotics. Microbiome (16S profiling), colonic histology, T cell and neutrophil infiltration, mucosal inflammatory tone, and epithelial permeability were analyzed. In adoptive T cell transfer colitis model, Slc9a3 status was the most significant determinant of gut microbial community. In DKO mice, NHE3-deficiency and dysbiosis were associated with dramatically accelerated and exacerbated disease, with rapid body weight loss, increased mucosal T cell and neutrophil influx, increased mucosal cytokine expression, increased permeability, and expansion of CD25-FoxP3+ Tregs; this enhanced susceptibility was alleviated by oral broad-spectrum antibiotics. Based on these results and our previous work, we postulate that epithelial electrolyte homeostasis is an important modulator in the progression of colitis, acting through remodeling of the gut microbial community. PMID:27050757

  12. Mucosal immunity: its role in defense and allergy.

    PubMed

    Tlaskalová-Hogenová, Helena; Tucková, Ludmila; Lodinová-Zádniková, Rája; Stepánková, Renata; Cukrowska, Bozena; Funda, David P; Striz, Ilja; Kozáková, Hana; Trebichavský, Ilja; Sokol, Dan; Reháková, Zuzana; Sinkora, Jirí; Fundová, Petra; Horáková, Dana; Jelínková, Lenka; Sánchez, Daniel

    2002-06-01

    The interface between the organism and the outside world, which is the site of exchange of nutrients, export of products and waste components, must be selectively permeable and at the same time, it must constitute a barrier equipped with local defense mechanisms against environmental threats (e.g. invading pathogens). The boundaries with the environment (mucosal and skin surfaces) are therefore covered with special epithelial layers which support this barrier function. The immune system, associated with mucosal surfaces covering the largest area of the body (200-300 m(2)), evolved mechanisms discriminating between harmless antigens and commensal microorganisms and dangerous pathogens. The innate mucosal immune system, represented by epithelial and other mucosal cells and their products, is able to recognize the conserved pathogenic patterns on microbes by pattern recognition receptors such as Toll-like receptors, CD14 and others. As documented in experimental gnotobiotic models, highly protective colonization of mucosal surfaces by commensals has an important stimulatory effect on postnatal development of immune responses, metabolic processes (e.g. nutrition) and other host activities; these local and systemic immune responses are later replaced by inhibition, i.e. by induction of mucosal (oral) tolerance. Characteristic features of mucosal immunity distinguishing it from systemic immunity are: strongly developed mechanisms of innate defense, the existence of characteristic populations of unique types of lymphocytes, colonization of the mucosal and exocrine glands by cells originating from the mucosal organized tissues ('common mucosal system') and preferential induction of inhibition of the responses to nondangerous antigens (mucosal tolerance). Many chronic diseases, including allergy, may occur as a result of genetically based or environmentally induced changes in mechanisms regulating mucosal immunity and tolerance; this leads to impaired mucosal barrier

  13. Protective effects of Lactobacillus plantarum on epithelial barrier disruption caused by enterotoxigenic Escherichia coli in intestinal porcine epithelial cells.

    PubMed

    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) (P<0.05), inhibited the increased transcript abundances of interleukin-8 (IL-8) and tumor necrosis factor (TNF-α) (P<0.05), decreased expression of claudin-1, occludin and zonula occludens (ZO-1) (P<0.05) and protein expression of occludin (P<0.05) of IPEC-J2 cells caused by ETEC K88. Moreover, the mRNA expression of negative regulators of toll-like receptors (TLRs) [single Ig Il-1-related receptor (SIGIRR), B-cell CLL/lymphoma 3 (Bcl3), and mitogen-activated protein kinase phosphatase-1 (MKP-1)] in IPEC-J2 cells pre-treated with L. plantarum were higher (P<0.05) compared with those in cells just exposed to K88. Furthermore, L. plantarum 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. PMID:27032504

  14. Heat shock protein 70-dependent protective effect of polaprezinc on acetylsalicylic acid-induced apoptosis of rat intestinal epithelial cells.

    PubMed

    Qin, Ying; Naito, Yuji; Handa, Osamu; Hayashi, Natsuko; Kuki, Aiko; Mizushima, Katsura; Omatsu, Tatsushi; Tanimura, Yuko; Morita, Mayuko; Adachi, Satoko; Fukui, Akifumi; Hirata, Ikuhiro; Kishimoto, Etsuko; Nishikawa, Taichiro; Uchiyama, Kazuhiko; Ishikawa, Takeshi; Takagi, Tomohisa; Yagi, Nobuaki; Kokura, Satoshi; Yoshikawa, Toshikazu

    2011-11-01

    Protection of the small intestine from mucosal injury induced by nonsteroidal anti-inflammatory drugs including acetylsalicylic acid is a critical issue in the field of gastroenterology. Polaprezinc an anti-ulcer drug, consisting of zinc and L-carnosine, provides gastric mucosal protection against various irritants. In this study, we investigated the protective effect of polaprezinc on acetylsalicylic acid-induced apoptosis of the RIE1 rat intestinal epithelial cell line. Confluent rat intestinal epithelial cells were incubated with 70 µM polaprezinc for 24 h, and then stimulated with or without 15 mM acetylsalicylic acid for a further 15 h. Subsequent cellular viability was quantified by fluorometric assay based on cell lysis and staining. Acetylsalicylic acid-induced cell death was also qualified by fluorescent microscopy of Hoechst33342 and propidium iodide. Heat shock proteins 70 protein expression after adding polaprezinc or acetylsalicylic acid was assessed by western blotting. To investigate the role of Heat shock protein 70, Heat shock protein 70-specific small interfering RNA was applied. Cell viability was quantified by fluorometric assay based on cell lysis and staining and apoptosis was analyzed by fluorescence-activated cell sorting. We found that acetylsalicylic acid significantly induced apoptosis of rat intestinal epithelial cells in a dose- and time-dependent manner. Polaprezinc significantly suppressed acetylsalicylic acid-induced apoptosis of rat intestinal epithelial cells at its late phase. At the same time, polaprezinc increased Heat shock protein 70 expressions of rat intestinal epithelial cells in a time-dependent manner. However, in Heat shock protein 70-silenced rat intestinal epithelial cells, polaprezinc could not suppress acetylsalicylic acid -induced apoptosis at its late phase. We conclude that polaprezinc-increased Heat shock protein 70 expression might be an important mechanism by which polaprezinc suppresses acetylsalicylic

  15. Cryptosporidium parvum induces SIRT1 expression in host epithelial cells through downregulating let-7i

    PubMed Central

    Xie, Hongguan; Lei, Ningfei; Gong, Ai-Yu; Chen, Xian-Ming; Hu, Guoku

    2015-01-01

    Epithelial cells along human gastrointestinal mucosal surface express pathogen-recognizing receptors and actively participate in the regulation of inflammatory reactions in response to microbial infection. The NAD-dependent deacetylase sirtuin-1 (SIRT1), one member of the sirtuin family of proteins and an NAD-dependent deacetylase has been implicated in the regulation of multiple cellular processes, including inflammation, longevity, and metabolism. In this study, we demonstrated that infection of cultured human biliary epithelial cells (H69 cholangiocytes) with a parasitic protozoan, Cryptosporidium parvum, induced SIRT1 expression at the protein level without a change in SIRT1 mRNA content. Using real-time PCR and Northern blot analyses, we found that C. parvum infection decreased the expression of let-7i in infected H69 cells. Down-regulation of let-7i caused relief of miRNA-mediated translational suppression of SIRT1 and consequently, resulted in an increased SIRT1 protein level in infected H69 cell cultures. Moreover, gain- and loss-of-function studies revealed that let-7i could modulate NF-κB activation through modification of SIRT1 protein expression. Thus, our data suggest that let-7i regulates SIRT1 expression in human biliary epithelial cells in response to microbial challenge, suggesting a new role of let-7i in the regulation of NF-κB-mediated epithelial innate immune response. PMID:24862934

  16. Epithelium-Intrinsic MicroRNAs Contribute to Mucosal Immune Homeostasis by Promoting M-Cell Maturation.

    PubMed

    Nakato, Gaku; Hase, Koji; Sato, Takao; Kimura, Shunsuke; Sakakibara, Sayuri; Sugiyama, Machiko; Obata, Yuuki; Hanazato, Misaho; Iwanaga, Toshihiko; Ohno, Hiroshi

    2016-01-01

    M cells in the follicle-associated epithelium (FAE) of Peyer's patches (PPs) serve as a main portal for external antigens and function as a sentinel in mucosal immune responses. The scarcity of these cells has hampered identification of M cell-specific molecules. Recent efforts have begun to provide insight into antigen transcytosis and differentiation of M cells; however, the molecular mechanisms underlying these processes are not fully elucidated. Small non-coding RNAs including microRNA (miRNA) have been reported to regulate gene expression and control various biological processes such as cellular differentiation and function. To evaluate the expression of miRNAs in FAE, including M cells, we previously performed microarray analysis comparing intestinal villous epithelium (VE) and PP FAE. Here we confirmed FAE specific miRNA expression levels by quantitative PCR. To gain insight into miRNA function, we generated mice with intestinal epithelial cell-specific deletion of Dicer1 (DicerΔIEC) and analyzed intestinal phenotypes, including M-cell differentiation, morphology and function. DicerΔIEC mice had a marked decrease in M cells compared to control floxed Dicer mice, suggesting an essential role of miRNAs in maturation of these cells. Furthermore, transmission electron microscopic analysis revealed that depletion of miRNA caused the loss of endosomal structures in M cells. In addition, antigen uptake by M cells was impaired in DicerΔIEC mice. These results suggest that miRNAs play a significant role in M cell differentiation and help secure mucosal immune homeostasis. PMID:26930511

  17. Epithelium-Intrinsic MicroRNAs Contribute to Mucosal Immune Homeostasis by Promoting M-Cell Maturation

    PubMed Central

    Nakato, Gaku; Hase, Koji; Sato, Takao; Kimura, Shunsuke; Sakakibara, Sayuri; Sugiyama, Machiko; Obata, Yuuki; Hanazato, Misaho; Iwanaga, Toshihiko; Ohno, Hiroshi

    2016-01-01

    M cells in the follicle-associated epithelium (FAE) of Peyer’s patches (PPs) serve as a main portal for external antigens and function as a sentinel in mucosal immune responses. The scarcity of these cells has hampered identification of M cell-specific molecules. Recent efforts have begun to provide insight into antigen transcytosis and differentiation of M cells; however, the molecular mechanisms underlying these processes are not fully elucidated. Small non-coding RNAs including microRNA (miRNA) have been reported to regulate gene expression and control various biological processes such as cellular differentiation and function. To evaluate the expression of miRNAs in FAE, including M cells, we previously performed microarray analysis comparing intestinal villous epithelium (VE) and PP FAE. Here we confirmed FAE specific miRNA expression levels by quantitative PCR. To gain insight into miRNA function, we generated mice with intestinal epithelial cell-specific deletion of Dicer1 (DicerΔIEC) and analyzed intestinal phenotypes, including M-cell differentiation, morphology and function. DicerΔIEC mice had a marked decrease in M cells compared to control floxed Dicer mice, suggesting an essential role of miRNAs in maturation of these cells. Furthermore, transmission electron microscopic analysis revealed that depletion of miRNA caused the loss of endosomal structures in M cells. In addition, antigen uptake by M cells was impaired in DicerΔIEC mice. These results suggest that miRNAs play a significant role in M cell differentiation and help secure mucosal immune homeostasis. PMID:26930511

  18. Taenia solium Oncosphere Adhesion to Intestinal Epithelial and Chinese Hamster Ovary Cells In Vitro▿

    PubMed Central

    Verastegui, Manuela; Gilman, Robert H.; Arana, Yanina; Barber, Dylan; Velásquez, Jeanette; Farfán, Marilu; Chile, Nancy; Kosek, Jon C.; Kosek, Margaret; Garcia, Hector H.; Gonzalez, Armando

    2007-01-01

    The specific mechanisms underlying Taenia solium oncosphere adherence and penetration in the host have not been studied previously. We developed an in vitro adhesion model assay to evaluate the mechanisms of T. solium oncosphere adherence to the host cells. The following substrates were used: porcine intestinal mucosal scrapings (PIMS), porcine small intestinal mucosal explants (PSIME), Chinese hamster ovary cells (CHO cells), epithelial cells from ileocecal colorectal adenocarcinoma (HCT-8 cells), and epithelial cells from colorectal adenocarcinoma (Caco-2 cells). CHO cells were used to compare oncosphere adherence to fixed and viable cells, to determine the optimum time of oncosphere incubation, to determine the role of sera and monolayer cell maturation, and to determine the effect of temperature on oncosphere adherence. Light microscopy, scanning microscopy, and transmission microscopy were used to observe morphological characteristics of adhered oncospheres. This study showed in vitro adherence of activated T. solium oncospheres to PIMS, PSIME, monolayer CHO cells, Caco-2 cells, and HCT-8 cells. The reproducibility of T. solium oncosphere adherence was most easily measured with CHO cells. Adherence was enhanced by serum-binding medium with >5% fetal bovine serum, which resulted in a significantly greater number of oncospheres adhering than the number adhering when serum at a concentration less than 2.5% was used (P < 0.05). Oncosphere adherence decreased with incubation of cells at 4°C compared with the adherence at 37°C. Our studies also demonstrated that T. solium oncospheres attach to cells with elongated microvillus processes and that the oncospheres expel external secretory vesicles that have the same oncosphere processes. PMID:17698575

  19. Murine CMV infection induces the continuous production of mucosal resident T cells

    PubMed Central

    Smith, Corinne J.; Caldeira-Dantas, Sofia; Turula, Holly; Snyder, Christopher M.

    2015-01-01

    Summary Cytomegalovirus (CMV) is a herpesvirus that persists for life and maintains extremely large numbers of T cells with select specificities in circulation. However, it is unknown how viral persistence impacts T cell populations in mucosal sites. We found that many murine (M)CMV-specific CD8s in mucosal tissues became resident memory T cells (TRM). These cells adopted an intraepithelial localization in the salivary gland that correlated with, but did not depend on, expression of the integrin CD103. MCMV-specific TRM cells formed early after infection and spleen-localized cells had reduced capacities to become TRM at late times. Surprisingly however, small numbers of new TRM cells were formed from the circulating pool throughout infection, favoring populations maintained at high levels in the blood and shifting the immunodominance within the TRM populations over time. These data show that mucosal TRM populations can be dynamically maintained by a persistent infection. PMID:26526996

  20. Lack of Interleukin-10-Mediated Anti-Inflammatory Signals and Upregulated Interferon Gamma Production Are Linked to Increased Intestinal Epithelial Cell Apoptosis in Pathogenic Simian Immunodeficiency Virus Infection

    PubMed Central

    Pan, Diganta; Kenway-Lynch, Carys S.; Lala, Wendy; Veazey, Ronald S.; Lackner, Andrew A.; Das, Arpita

    2014-01-01

    ABSTRACT Interleukin-10 (IL-10) is an immunomodulatory cytokine that is important for maintenance of epithelial cell (EC) survival and anti-inflammatory responses (AIR). The majority of HIV infections occur through the mucosal route despite mucosal epithelium acting as a barrier to human immunodeficiency virus (HIV). Therefore, understanding the role of IL-10 in maintenance of intestinal homeostasis during HIV infection is of interest for better characterization of the pathogenesis of HIV-mediated enteropathy. We demonstrated here changes in mucosal IL-10 signaling during simian immunodeficiency virus (SIV) infection in rhesus macaques. Disruption of the epithelial barrier was manifested by EC apoptosis and loss of the tight-junction protein ZO-1. Multiple cell types, including a limited number of ECs, produced IL-10. SIV infection resulted in increased levels of IL-10; however, this was associated with increased production of mucosal gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α), suggesting that IL-10 was not able to regulate AIR. This observation was supported by the downregulation of STAT3, which is necessary to inhibit production of IFN-γ and TNF-α, and the upregulation of SOCS1 and SOCS3, which are important regulatory molecules in the IL-10-mediated AIR. We also observed internalization of the IL-10 receptor (IL-10R) in mucosal lymphocytes, which could limit cellular availability of IL-10 for signaling and contribute to the loss of a functional AIR. Collectively, these findings demonstrate that internalization of IL-10R with the resultant impact on IL-10 signaling and dysregulation of the IL-10-mediated AIR might play a crucial role in EC damage and subsequent SIV/HIV pathogenesis. IMPORTANCE Interleukin-10 (IL-10), an important immunomodulatory cytokine plays a key role to control inflammatory function and homeostasis of the gastrointestinal mucosal immune system. Despite recent advancements in the study of IL-10 and its role in HIV

  1. Pulmonary Epithelial Cell-Derived Cytokine TGF-β1 Is a Critical Cofactor for Enhanced Innate Lymphoid Cell Function

    PubMed Central

    Denney, Laura; Byrne, Adam J.; Shea, Thomas J.; Buckley, James S.; Pease, James E.; Herledan, Gaelle M.F.; Walker, Simone A.; Gregory, Lisa G.; Lloyd, Clare M.

    2015-01-01

    Summary Epithelial cells orchestrate pulmonary homeostasis and pathogen defense and play a crucial role in the initiation of allergic immune responses. Maintaining the balance between homeostasis and inappropriate immune activation and associated pathology is particularly complex at mucosal sites that are exposed to billions of potentially antigenic particles daily. We demonstrated that epithelial cell-derived cytokine TGF-β had a central role in the generation of the pulmonary immune response. Mice that specifically lacked epithelial cell-derived TGF-β1 displayed a reduction in type 2 innate lymphoid cells (ILCs), resulting in suppression of interleukin-13 and hallmark features of the allergic response including airway hyperreactivity. ILCs in the airway lumen were primed to respond to TGF-β by expressing the receptor TGF-βRII and ILC chemoactivity was enhanced by TGF-β. These data demonstrate that resident epithelial cells instruct immune cells, highlighting the central role of the local environmental niche in defining the nature and magnitude of immune reactions. PMID:26588780

  2. Formation of a Neurosensory Organ by Epithelial Cell Slithering.

    PubMed

    Kuo, Christin S; Krasnow, Mark A

    2015-10-01

    Epithelial cells are normally stably anchored, maintaining their relative positions and association with the basement membrane. Developmental rearrangements occur through cell intercalation, and cells can delaminate during epithelial-mesenchymal transitions and metastasis. We mapped the formation of lung neuroepithelial bodies (NEBs), innervated clusters of neuroendocrine/neurosensory cells within the bronchial epithelium, revealing a targeted mode of cell migration that we named "slithering," in which cells transiently lose epithelial character but remain associated with the membrane while traversing neighboring epithelial cells to reach cluster sites. Immunostaining, lineage tracing, clonal analysis, and live imaging showed that NEB progenitors, initially distributed randomly, downregulate adhesion and polarity proteins, crawling over and between neighboring cells to converge at diametrically opposed positions at bronchial branchpoints, where they reestablish epithelial structure and express neuroendocrine genes. There is little accompanying progenitor proliferation or apoptosis. Activation of the slithering program may explain why lung cancers arising from neuroendocrine cells are highly metastatic. PMID:26435104

  3. Ouabain modulates ciliogenesis in epithelial cells

    PubMed Central

    Larre, Isabel; Castillo, Aida; Flores-Maldonado, Catalina; Contreras, Ruben G.; Galvan, Ivan; Muñoz-Estrada, Jesus; Cereijido, Marcelino

    2011-01-01

    The exchange of substances between higher organisms and the environment occurs across transporting epithelia whose basic features are tight junctions (TJs) that seal the intercellular space, and polarity, which enables cells to transport substances vectorially. In a previous study, we demonstrated that 10 nM ouabain modulates TJs, and we now show that it controls polarity as well. We gauge polarity through the development of a cilium at the apical domain of Madin-Darby canine kidney cells (MDCK, epithelial dog kidney). Ouabain accelerates ciliogenesis in an ERK1/2-dependent manner. Claudin-2, a molecule responsible for the Na+ and H2O permeability of the TJs, is also present at the cilium, as it colocalizes and coprecipitates with acetylated α-tubulin. Ouabain modulates claudin-2 localization at the cilium through ERK1/2. Comparing wild-type and ouabain-resistant MDCK cells, we show that ouabain acts through Na+,K+-ATPase. Taken together, our previous and present results support the possibility that ouabain constitutes a hormone that modulates the transporting epithelial phenotype, thereby playing a crucial role in metazoan life. PMID:22143774

  4. Inflammation and epithelial cell injury in AIDS enteropathy: involvement of endoplasmic reticulum stress

    PubMed Central

    Maingat, Ferdinand; Halloran, Brendan; Acharjee, Shaona; van Marle, Guido; Church, Deirdre; Gill, M. John; Uwiera, Richard R. E.; Cohen, Eric A.; Meddings, Jon; Madsen, Karen; Power, Christopher

    2011-01-01

    Immunosuppressive lentivirus infections, including human, simian, and feline immunodeficiency viruses (HIV, SIV, and FIV, respectively), cause the acquired immunodeficiency syndrome (AIDS), frequently associated with AIDS enteropathy. Herein, we investigated the extent to which lentivirus infections affected mucosal integrity and intestinal permeability in conjunction with immune responses and activation of endoplasmic reticulum (ER) stress pathways. Duodenal biopsies from individuals with HIV/AIDS exhibited induction of IL-1β, CD3ε, HLA-DRA, spliced XBP-1(Xbp-1s), and CHOP expression compared to uninfected persons (P<0.05). Gut epithelial cells exposed to HIV-1 Vpr demonstrated elevated TNF-α, IL-1β, spliced Xbp-1s, and CHOP expression (P<0.05) together with calcium activation and disruption of epithelial cell monolayer permeability. In addition to reduced blood CD4+ T lymphocyte levels, viral loads in the gut and plasma were high in FIV-infected animals (P<0.05). FIV-infected animals also exhibited a failure to gain weight and increased lactulose/mannitol ratios compared with uninfected animals (P<0.05). Proinflammatory and ER stress gene expression were activated in the ileum of FIV-infected animals (P<0.05), accompanied by intestinal epithelial damage with loss of epithelial cells and leukocyte infiltration of the lamina propria. Lentivirus infections cause gut inflammation and ensuing damage to intestinal epithelial cells, likely through induction of ER stress pathways, resulting in disruption of gut functional integrity.—Maingat, F., Halloran, B., Acharjee, S., van Marle, G., Church, D., Gill, M. J., Uwiera, R. R. E., Cohen, E. A., Meddings, J., Madsen, K., Power, C. Inflammation and epithelial cell injury in AIDS enteropathy: involvement of endoplasmic reticulum stress. PMID:21427211

  5. Is the inflammasome relevant for epithelial cell function?

    PubMed

    Santana, Patricia T; Martel, Jan; Lai, Hsin-Chih; Perfettini, Jean-Luc; Kanellopoulos, Jean M; Young, John D; Coutinho-Silva, Robson; Ojcius, David M

    2016-02-01

    Inflammasomes are intracellular protein complexes that sense microbial components and damage of infected cells. Following activation by molecules released by pathogens or injured cells, inflammasomes activate caspase-1, allowing secretion of the pro-inflammatory cytokines IL-1β and IL-18 from innate immune cells. Inflammasomes are also expressed in epithelial cells, where their function has attracted less attention. Nonetheless, depending on the tissue, epithelial inflammasomes can mediate inflammation, wound healing, and pain sensitivity. We review here recent findings on inflammasomes found in epithelial tissues, highlighting the importance of these protein complexes in the response of epithelial tissues to microbial infections. PMID:26546965

  6. Impaired T-cell survival promotes mucosal inflammatory disease in SHIP1-deficient mice.

    PubMed

    Park, M Y; Srivastava, N; Sudan, R; Viernes, D R; Chisholm, J D; Engelman, R W; Kerr, W G

    2014-11-01

    T cells have a critical role in immune surveillance at mucosal surfaces. SHIP1(-/-) mice succumb to mucosal inflammatory disease that afflicts the lung and small intestine (SI). The basis of this condition has not been defined. Here we show that SHIP1 is required for the normal persistence and survival of T cells in mucosal tissues. We find that CD4 and CD8 effector T cells are reduced; however, Treg cells are increased in the SI and lungs of SHIP1(-/-) and CD4CreSHIP(flox/flox) mice. Furthermore, a subset of T cells in the SI of SHIP1(-/-) mice are FasL(+) and are more susceptible to extrinsic cell death. Mechanistic analyses showed that SHIP1 associates with the death receptor CD95/Fas and treatment with a Caspase 8 inhibitor prevents SHIP1 inhibitor-mediated T-cell death. Notably, mucosal inflammation in SHIP1(-/-) mice is reduced by treatment with a Caspase 8 inhibitor. We also find that the incidence of Crohn's disease (CD) and pneumonia is significantly increased in mice with dual T and myeloid lineage SHIP1 deletion but not in single lineage-deleted mice. Thus, by promoting survival of protective T cells, thereby preventing an inflammatory myeloid response, SHIP1 maintains an appropriate balance of innate immune function at mucosal surfaces necessary for immune homeostasis. PMID:24781051

  7. Stem cell therapy: A novel treatment approach for oral mucosal lesions.

    PubMed

    Suma, G N; Arora, Madhu Pruthi; Lakhanpal, Manisha

    2015-01-01

    Stem cells have enormous potential to alleviate sufferings of many diseases that currently have no effective therapy. The research in this field is growing at an exponential rate. Stem cells are master cells that have specialized capability for self-renewal, potency and capability to differentiate to many cell types. At present, the adult mesenchymal stem cells are being used in the head and neck region for orofacial regeneration (including enamel, dentin, pulp and alveolar bone) in lieu of their proliferative and regenerative properties, their use in the treatment of oral mucosal lesions is still in budding stages. Moreover, there is scanty literature available regarding role of stem cell therapy in the treatment of commonly seen oral mucosal lesions like oral submucous fibrosis, oral lichen planus, oral ulcers and oral mucositis. The present review will focus on the current knowledge about the role of stem cell therapies in oral mucosal lesions and could facilitate new advancements in this area (articles were obtained from electronic media like PubMed, EBSCO, Cochrane and Medline etc., from year 2000 to 2014 to review the role of stem cell therapy in oral mucosal lesions). PMID:25709329

  8. Coevolution of neoplastic epithelial cells and multilineage stroma via polyploid giant cells during immortalization and transformation of mullerian epithelial cells

    PubMed Central

    Zhang, Shiwu; Mercado-Uribe, Imelda; Sood, Anil; Bast, Robert C.; Liu, Jinsong

    2016-01-01

    Stromal cells are generally considered to be derived primarily from the host's normal mesenchymal stromal cells or bone marrow. However, the origins of stromal cells have been quite controversial. To determine the role of polyploidy in tumor development, we examined the fate of normal mullerian epithelial cells during the immortalization and transformation process by tracing the expression of SV40 large T antigen. Here we show that immortalized or HRAS-transformed mullerian epithelial cells contain a subpopulation of polyploid giant cells that grow as multicellular spheroids expressing hematopoietic markers in response to treatment with CoCl2. The immortalized or transformed epithelial cells can transdifferentiate into stromal cells when transplanted into nude mice. Immunofluorescent staining revealed expression of stem cell factors OCT4, Nanog, and SOX-2 in spheroid, whereas expression of embryonic stem cell marker SSEA1 was increased in HRAS-transformed cells compared with their immortalized isogenic counterparts. These results suggest that normal mullerian epithelial cells are intrinsically highly plastic, via the formation of polyploid giant cells and activation of embryonic stem-like program, which work together to promote the coevolution of neoplastic epithelial cells and multiple lineage stromal cells. PMID:27382431

  9. Establishment of Hertwig's epithelial root sheath/epithelial rests of Malassez cell line from human periodontium.

    PubMed

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

    2014-07-01

    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 population in periodontium and the primary isolation of them is considered to be difficult. To overcome these problems, we immortalized primary HERS/ERM cells isolated from human periodontium using SV40 large T antigen (SV40 LT) and performed a characterization of the immortalized cell line. Primary HERS/ERM cells could not be maintained for more than 6 passages; however, immortalized HERS/ERM cells were maintained for more than 20 passages. There were no differences in the morphological and immunophenotypic characteristics of HERS/ERM cells and immortalized HERS/ERM cells. The expression of epithelial stem cell and embryonic stem cell markers was maintained in immortalized HERS/ERM cells. Moreover, immortalized HERS/ERM cells could acquire mesenchymal phenotypes through the epithelial-mesenchymal transition via TGF-β1. In conclusion, we established an immortalized human HERS/ERM cell line with SV40 LT and expect this cell line to contribute to the understanding of the functional roles of HERS/ERM cells and the tissue engineering of teeth. PMID:25081036

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

    SciTech Connect

    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.

  11. Stromal-epithelial interactions in aging and cancer: senescent fibroblasts alter epithelial cell differentiation

    PubMed Central

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

    2016-01-01

    Summary Cellular senescence suppresses cancer by arresting cells at risk of 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. Furthermore, 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. PMID:15657080

  12. Mucosal Immunity and B Cells in Teleosts: Effect of Vaccination and Stress

    PubMed Central

    Parra, David; Reyes-Lopez, Felipe E.; Tort, Lluis

    2015-01-01

    Fish are subjected to several insults from the environment, which may endanger animal survival. Mucosal surfaces are the first line of defense against these threats, acting as a physical barrier to protect the animal but also functioning as an active immune tissue. Thus, four mucosal-associated lymphoid tissues (MALTs), which lead the immune responses in gut, skin, gills, and nose, have been described in fish. Humoral and cellular immunity, as well as their regulation and the factors that influence the response in these mucosal lymphoid tissues, are still not well known in most fish species. Mucosal B-lymphocytes and immunoglobulins (Igs) are key players in the immune response that takes place in those MALTs. The existence of IgT as a mucosal specialized Ig gives us the opportunity of measuring specific responses after infection or vaccination, a fact that was not possible until recently in most fish species. The vaccination process is influenced by several factors, being stress one of the main stimuli determining the success of the vaccine. Thus, one of the major goals in a vaccination process is to avoid possible situations of stress, which might interfere with fish immune performance. However, interaction between immune and neuroendocrine systems at mucosal tissues is still unknown. In this review, we will summarize the latest findings about B-lymphocytes and Igs in mucosal immunity and the effect of stress and vaccination on B-cell response at mucosal sites. It is important to point out that a limited number of studies have been published regarding stress in mucosa and very few about the influence of stress over mucosal B-lymphocytes. PMID:26236311

  13. Henipavirus Pathogenesis in Human Respiratory Epithelial Cells

    PubMed Central

    Escaffre, Olivier; Borisevich, Viktoriya; Carmical, J. Russ; Prusak, Deborah; Prescott, Joseph; Feldmann, Heinz

    2013-01-01

    Hendra virus (HeV) and Nipah virus (NiV) are deadly zoonotic viruses for which no vaccines or therapeutics are licensed for human use. Henipavirus infection causes severe respiratory illness and encephalitis. Although the exact route of transmission in human is unknown, epidemiological studies and in vivo studies suggest that the respiratory tract is important for virus replication. However, the target cells in the respiratory tract are unknown, as are the mechanisms by which henipaviruses can cause disease. In this study, we characterized henipavirus pathogenesis using primary cells derived from the human respiratory tract. The growth kinetics of NiV-Malaysia, NiV-Bangladesh, and HeV were determined in bronchial/tracheal epithelial cells (NHBE) and small airway epithelial cells (SAEC). In addition, host responses to infection were assessed by gene expression analysis and immunoassays. Viruses replicated efficiently in both cell types and induced large syncytia. The host response to henipavirus infection in NHBE and SAEC highlighted a difference in the inflammatory response between HeV and NiV strains as well as intrinsic differences in the ability to mount an inflammatory response between NHBE and SAEC. These responses were highest during HeV infection in SAEC, as characterized by the levels of key cytokines (interleukin 6 [IL-6], IL-8, IL-1α, monocyte chemoattractant protein 1 [MCP-1], and colony-stimulating factors) responsible for immune cell recruitment. Finally, we identified virus strain-dependent variability in type I interferon antagonism in NHBE and SAEC: NiV-Malaysia counteracted this pathway more efficiently than NiV-Bangladesh and HeV. These results provide crucial new information in the understanding of henipavirus pathogenesis in the human respiratory tract at an early stage of infection. PMID:23302882

  14. Henipavirus pathogenesis in human respiratory epithelial cells.

    PubMed

    Escaffre, Olivier; Borisevich, Viktoriya; Carmical, J Russ; Prusak, Deborah; Prescott, Joseph; Feldmann, Heinz; Rockx, Barry

    2013-03-01

    Hendra virus (HeV) and Nipah virus (NiV) are deadly zoonotic viruses for which no vaccines or therapeutics are licensed for human use. Henipavirus infection causes severe respiratory illness and encephalitis. Although the exact route of transmission in human is unknown, epidemiological studies and in vivo studies suggest that the respiratory tract is important for virus replication. However, the target cells in the respiratory tract are unknown, as are the mechanisms by which henipaviruses can cause disease. In this study, we characterized henipavirus pathogenesis using primary cells derived from the human respiratory tract. The growth kinetics of NiV-Malaysia, NiV-Bangladesh, and HeV were determined in bronchial/tracheal epithelial cells (NHBE) and small airway epithelial cells (SAEC). In addition, host responses to infection were assessed by gene expression analysis and immunoassays. Viruses replicated efficiently in both cell types and induced large syncytia. The host response to henipavirus infection in NHBE and SAEC highlighted a difference in the inflammatory response between HeV and NiV strains as well as intrinsic differences in the ability to mount an inflammatory response between NHBE and SAEC. These responses were highest during HeV infection in SAEC, as characterized by the levels of key cytokines (interleukin 6 [IL-6], IL-8, IL-1α, monocyte chemoattractant protein 1 [MCP-1], and colony-stimulating factors) responsible for immune cell recruitment. Finally, we identified virus strain-dependent variability in type I interferon antagonism in NHBE and SAEC: NiV-Malaysia counteracted this pathway more efficiently than NiV-Bangladesh and HeV. These results provide crucial new information in the understanding of henipavirus pathogenesis in the human respiratory tract at an early stage of infection. PMID:23302882

  15. Nuclear microscopy of rat colon epithelial cells

    NASA Astrophysics Data System (ADS)

    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.

  16. Regulation of intestinal epithelial cells transcriptome by enteric glial cells: impact on intestinal epithelial barrier functions

    PubMed Central

    2009-01-01

    Background Emerging evidences suggest that enteric glial cells (EGC), a major constituent of the enteric nervous system (ENS), are key regulators of intestinal epithelial barrier (IEB) functions. Indeed EGC inhibit intestinal epithelial cells (IEC) proliferation and increase IEB paracellular permeability. However, the role of EGC on other important barrier functions and the signalling pathways involved in their effects are currently unknown. To achieve this goal, we aimed at identifying the impact of EGC upon IEC transcriptome by performing microarray studies. Results EGC induced significant changes in gene expression profiling of proliferating IEC after 24 hours of co-culture. 116 genes were identified as differentially expressed (70 up-regulated and 46 down-regulated) in IEC cultured with EGC compared to IEC cultured alone. By performing functional analysis of the 116 identified genes using Ingenuity Pathway Analysis, we showed that EGC induced a significant regulation of genes favoring both cell-to-cell and cell-to-matrix adhesion as well as cell differentiation. Consistently, functional studies showed that EGC induced a significant increase in cell adhesion. EGC also regulated genes involved in cell motility towards an enhancement of cell motility. In addition, EGC profoundly modulated expression of genes involved in cell proliferation and cell survival, although no clear functional trend could be identified. Finally, important genes involved in lipid and protein metabolism of epithelial cells were shown to be differentially regulated by EGC. Conclusion This study reinforces the emerging concept that EGC have major protective effects upon the IEB. EGC have a profound impact upon IEC transcriptome and induce a shift in IEC phenotype towards increased cell adhesion and cell differentiation. This concept needs to be further validated under both physiological and pathophysiological conditions. PMID:19883504

  17. Influence of oral health on mucositis in patients undergoing hematopoietic progenitor cell transplantation (HPCT)

    PubMed Central

    Oñate-Sánchez, Ricardo E.; Cabrerizo-Merino, María C.; de Arriba de la Fuente, Felipe; Heras Fernando, Inmaculada; Vicente García, Vicente

    2012-01-01

    Aims: To establish whether or not the state of patient oral health can influence the occurrence and/or severity of oral mucositis during hematopoietic progenitor cell transplantation (HPCT). Materials and Methods: The study included 72 patients awaiting HPCT. Prior to transplantation, clinical exploration and radiology were carried out and oral photographs were taken. This evaluated the extent of caries present, the number of missing teeth and the number of dental fillings in each patient; CAO (Caries and Obturations Index) DMFS (Decayed, Missing, and Filled Surfaces) and Restoration Indices were calculated. Gingival pathology was also examined by means of the Ainamo and Bay Gingival Bleeding Index. O’Leary’s Plaque Index was used to evaluate the level of patient oral hygiene. This data was analyzed to see if it exercised any influence on the mucositis grade suffered during HPCT. Results: 96,87% of patients suffered some degree of mucositis during their treatment by the Transplant Unit. The grade of mucositis was seen to be influenced by the number of missing teeth (ANOVA p<0.016) and by the DMFS Index (ANOVA p< 0.038). Although this was not one of the aims of this study, patient age and the administration of colony-stimulating factors were also seen to influence these clinical manifestations. Conclusions: The state of prior oral health can influence decisively the mucositis suffered during transplantation. Key words: Hematopoietic progenitor cell transplantation, mucositis, state of oral health. PMID:22157660

  18. Increased Mucosal CD4+ T Cell Activation in Rhesus Macaques following Vaccination with an Adenoviral Vector

    PubMed Central

    Bukh, Irene; Calcedo, Roberto; Roy, Soumitra; Carnathan, Diane G.; Grant, Rebecca; Qin, Qiuyue; Boyd, Surina; Ratcliffe, Sarah J.; Veeder, Christin L.; Bellamy, Scarlett L.; Betts, Michael R.

    2014-01-01

    ABSTRACT The possibility that vaccination with adenovirus (AdV) vectors increased mucosal T cell activation remains a central hypothesis to explain the potential enhancement of HIV acquisition within the Step trial. Modeling this within rhesus macaques is complicated because human adenoviruses, including human adenovirus type 5 (HAdV-5), are not endogenous to macaques. Here, we tested whether vaccination with a rhesus macaque-derived adenoviral vector (simian adenovirus 7 [SAdV-7]) enhances mucosal T cell activation within rhesus macaques. Following intramuscular SAdV-7 vaccination, we observed a pronounced increase in SAdV-7-specific CD4+ T cell responses in peripheral blood and, more dramatically, in rectal mucosa tissue. Vaccination also induced a significant increase in the frequency of activated memory CD4+ T cells in SAdV-7- and HAdV-5-vaccinated animals in the rectal mucosa but not in peripheral blood. These fluctuations within the rectal mucosa were also associated with a pronounced decrease in the relative frequency of naive resting CD4+ T cells. Together, these results indicate that peripheral vaccination with an AdV vector can increase the activation of mucosal CD4+ T cells, potentially providing an experimental model to further evaluate the role of host-vector interactions in increased HIV acquisition after AdV vector vaccination. IMPORTANCE The possibility that vaccination with a human adenovirus 5 vector increased mucosal T cell activation remains a central hypothesis to explain the potential enhancement of human immunodeficiency virus (HIV) acquisition within the Step trial. In this study, we tested whether vaccination with a rhesus macaque-derived adenoviral vector in rhesus macaques enhances mucosal CD4+ T cell activation, the main cell target of simian immunodeficiency virus (SIV)/HIV. The results showed that vaccination with an adenoviral vector indeed increases activation of mucosal CD4+ T cells and potentially increases susceptibility to SIV

  19. Epithelial-mesenchymal transition can suppress major attributes of human epithelial tumor-initiating cells

    PubMed Central

    Celià-Terrassa, Toni; Meca-Cortés, Óscar; Mateo, Francesca; Martínez de Paz, Alexia; Rubio, Nuria; Arnal-Estapé, Anna; Ell, Brian J.; Bermudo, Raquel; Díaz, Alba; Guerra-Rebollo, Marta; Lozano, Juan José; Estarás, Conchi; Ulloa, Catalina; ρlvarez-Simón, Daniel; Milà, Jordi; Vilella, Ramón; Paciucci, Rosanna; Martínez-Balbás, Marian; García de Herreros, Antonio; Gomis, Roger R.; Kang, Yibin; Blanco, Jerónimo; Fernández, Pedro L.; Thomson, Timothy M.

    2012-01-01

    Malignant progression in cancer requires populations of tumor-initiating cells (TICs) endowed with unlimited self renewal, survival under stress, and establishment of distant metastases. Additionally, the acquisition of invasive properties driven by epithelial-mesenchymal transition (EMT) is critical for the evolution of neoplastic cells into fully metastatic populations. Here, we characterize 2 human cellular models derived from prostate and bladder cancer cell lines to better understand the relationship between TIC and EMT programs in local invasiveness and distant metastasis. The model tumor subpopulations that expressed a strong epithelial gene program were enriched in highly metastatic TICs, while a second subpopulation with stable mesenchymal traits was impoverished in TICs. Constitutive overexpression of the transcription factor Snai1 in the epithelial/TIC-enriched populations engaged a mesenchymal gene program and suppressed their self renewal and metastatic phenotypes. Conversely, knockdown of EMT factors in the mesenchymal-like prostate cancer cell subpopulation caused a gain in epithelial features and properties of TICs. Both tumor cell subpopulations cooperated so that the nonmetastatic mesenchymal-like prostate cancer subpopulation enhanced the in vitro invasiveness of the metastatic epithelial subpopulation and, in vivo, promoted the escape of the latter from primary implantation sites and accelerated their metastatic colonization. Our models provide new insights into how dynamic interactions among epithelial, self-renewal, and mesenchymal gene programs determine the plasticity of epithelial TICs. PMID:22505459

  20. Documentation of angiotensin II receptors in glomerular epithelial cells

    NASA Technical Reports Server (NTRS)

    Sharma, M.; Sharma, R.; Greene, A. S.; McCarthy, E. T.; Savin, V. J.; Cowley, A. W. (Principal Investigator)

    1998-01-01

    Angiotensin II decreases glomerular filtration rate, renal plasma flow, and glomerular capillary hydraulic conductivity. Although angiotensin II receptors have been demonstrated in mesangial cells and proximal tubule cells, the presence of angiotensin II receptors in glomerular epithelial cells has not previously been shown. Previously, we have reported that angiotensin II caused an accumulation of cAMP and a reorganization of the actin cytoskeleton in cultured glomerular epithelial cells. Current studies were conducted to verify the presence of angiotensin II receptors by immunological and non-peptide receptor ligand binding techniques and to ascertain the activation of intracellular signal transduction in glomerular epithelial cells in response to angiotensin II. Confluent monolayer cultures of glomerular epithelial cells were incubated with angiotensin II, with or without losartan and/or PD-123,319 in the medium. Membrane vesicle preparations were obtained by homogenization of washed cells followed by centrifugation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of membrane proteins followed by multiscreen immunoblotting was used to determine the presence of angiotensin II receptor type 1 (AT1) or type 2 (AT2). Angiotensin II-mediated signal transduction in glomerular epithelial cells was studied by measuring the levels of cAMP, using radioimmunoassay. Results obtained in these experiments showed the presence of both AT1 and AT2 receptor types in glomerular epithelial cells. Angiotensin II was found to cause an accumulation of cAMP in glomerular epithelial cells, which could be prevented only by simultaneous use of losartan and PD-123,319, antagonists for AT1 and AT2, respectively. The presence of both AT1 and AT2 receptors and an increase in cAMP indicate that glomerular epithelial cells respond to angiotensin II in a manner distinct from that of mesangial cells or proximal tubular epithelial cells. Our results suggest that glomerular epithelial

  1. Applications of mouse airway epithelial cell culture for asthma research.

    PubMed

    Horani, Amjad; Dickinson, John D; Brody, Steven L

    2013-01-01

    Primary airway epithelial cell culture provides a valuable tool for studying cell differentiation, cell-cell interactions, and the role of immune system factors in asthma pathogenesis. In this chapter, we discuss the application of mouse tracheal epithelial cell cultures for the study of asthma biology. A major advantage of this system is the ability to use airway epithelial cells from mice with defined genetic backgrounds. The in vitro proliferation and differentiation of mouse airway epithelial cells uses the air-liquid interface condition to generate well-differentiated epithelia with characteristics of native airways. Protocols are provided for manipulation of differentiation, induction of mucous cell metaplasia, genetic modification, and cell and pathogen coculture. Assays for the assessment of gene expression, responses of cells, and analysis of specific cell subpopulations within the airway epithelium are included. PMID:23943446

  2. Klebsiella pneumoniae Is Able to Trigger Epithelial-Mesenchymal Transition Process in Cultured Airway Epithelial Cells

    PubMed Central

    Leone, Laura; Mazzetta, Francesca; Martinelli, Daniela; Valente, Sabatino; Alimandi, Maurizio; Raffa, Salvatore; Santino, Iolanda

    2016-01-01

    The ability of some bacterial pathogens to activate Epithelial-Mesenchymal Transition normally is a consequence of the persistence of a local chronic inflammatory response or depends on a direct interaction of the pathogens with the host epithelial cells. In this study we monitored the abilities of the K. pneumoniae to activate the expression of genes related to EMT-like processes and the occurrence of phenotypic changes in airway epithelial cells during the early steps of cell infection. We describe changes in the production of intracellular reactive oxygen species and increased HIF-1α mRNA expression in cells exposed to K. pneumoniae infection. We also describe the upregulation of a set of transcription factors implicated in the EMT processes, such as Twist, Snail and ZEB, indicating that the morphological changes of epithelial cells already appreciable after few hours from the K. pneumoniae infection are tightly regulated by the activation of transcriptional pathways, driving epithelial cells to EMT. These effects appear to be effectively counteracted by resveratrol, an antioxidant that is able to exert a sustained scavenging of the intracellular ROS. This is the first report indicating that strains of K. pneumoniae may promote EMT-like programs through direct interaction with epithelial cells without the involvement of inflammatory cells. PMID:26812644

  3. Differential crosstalk between epithelial cells, dendritic cells and bacteria in a co-culture model.

    PubMed

    Zoumpopoulou, Georgia; Tsakalidou, Effie; Dewulf, Joelle; Pot, Bruno; Grangette, Corinne

    2009-04-30

    Intestinal epithelial cells (IECs) provide a primary physical barrier against commensal and pathogenic bacteria, but the influence of IECs in the regulation of the associated mucosal immune system remains largely unknown. The network of dendritic cells (DCs) in the vicinity of IECs is known to play a crucial role in the regulation of gut homeostasis. We investigated the cross-talk between murine IECs (m-IC(cl2) cell line), bone marrow derived DCs and different bacteria using an in vitro Transwell co-culture model. IECs responded poorly to different gram-positive lactic acid bacteria (LAB) and to a Staphylococcus aureus strain. In contrast two Escherichia coli strains, including the probiotic strain Nissle 1917, strongly activated IECs, as evidenced by the induction of different chemokines. While a differential maturation of DCs is observed upon direct stimulation with the various bacteria, DC maturation across the epithelial barrier was only observed upon challenge of the apical surface of the IECs with both E. coli strains and LPS. These results suggested that the capacity of bacteria to induce pro-inflammatory signals through the epithelial barrier is not linked to their pathogenic or commensal status, but seem to be dependent on the presence of specific surface factors. As already reported, we confirmed that m-IC(cl2) cells are highly susceptible to LPS. It is highly possible, at least in this model, that free LPS is the "specific factor" key to activate IEC or BMDC. Moreover, IECs are broadly unresponsive to gram-positive bacterial components, notably TLR-2 ligands, in contrast to gram-negative bacterial components. These results suggest that the gut epithelium will sense the commensal bacteria in a different way, and seems to be unresponsive to gram positive bacteria in particular to LAB. PMID:19264370

  4. HIV-associated disruption of tight and adherens junctions of oral epithelial cells facilitates HSV-1 infection and spread.

    PubMed

    Sufiawati, Irna; Tugizov, Sharof M

    2014-01-01

    Herpes simplex virus (HSV) types 1 and 2 are the most common opportunistic infections in HIV/AIDS. In these immunocompromised individuals, HSV-1 reactivates and replicates in oral epithelium, leading to oral disorders such as ulcers, gingivitis, and necrotic lesions. Although the increased risk of HSV infection may be mediated in part by HIV-induced immune dysfunction, direct or indirect interactions of HIV and HSV at the molecular level may also play a role. In this report we show that prolonged interaction of the HIV proteins tat and gp120 and cell-free HIV virions with polarized oral epithelial cells leads to disruption of tight and adherens junctions of epithelial cells through the mitogen-activated protein kinase signaling pathway. HIV-induced disruption of oral epithelial junctions facilitates HSV-1 paracellular spread between the epithelial cells. Furthermore, HIV-associated disruption of adherens junctions exposes sequestered nectin-1, an adhesion protein and critical receptor for HSV envelope glycoprotein D (gD). Exposure of nectin-1 facilitates binding of HSV-1 gD, which substantially increases HSV-1 infection of epithelial cells with disrupted junctions over that of cells with intact junctions. Exposed nectin-1 from disrupted adherens junctions also increases the cell-to-cell spread of HSV-1 from infected to uninfected oral epithelial cells. Antibodies to nectin-1 and HSV-1 gD substantially reduce HSV-1 infection and cell-to-cell spread, indicating that HIV-promoted HSV infection and spread are mediated by the interaction of HSV gD with HIV-exposed nectin-1. Our data suggest that HIV-associated disruption of oral epithelial junctions may potentiate HSV-1 infection and its paracellular and cell-to-cell spread within the oral mucosal epithelium. This could be one of the possible mechanisms of rapid development of HSV-associated oral lesions in HIV-infected individuals. PMID:24586397

  5. HIV-Associated Disruption of Tight and Adherens Junctions of Oral Epithelial Cells Facilitates HSV-1 Infection and Spread

    PubMed Central

    Sufiawati, Irna; Tugizov, Sharof M.

    2014-01-01

    Herpes simplex virus (HSV) types 1 and 2 are the most common opportunistic infections in HIV/AIDS. In these immunocompromised individuals, HSV-1 reactivates and replicates in oral epithelium, leading to oral disorders such as ulcers, gingivitis, and necrotic lesions. Although the increased risk of HSV infection may be mediated in part by HIV-induced immune dysfunction, direct or indirect interactions of HIV and HSV at the molecular level may also play a role. In this report we show that prolonged interaction of the HIV proteins tat and gp120 and cell-free HIV virions with polarized oral epithelial cells leads to disruption of tight and adherens junctions of epithelial cells through the mitogen-activated protein kinase signaling pathway. HIV-induced disruption of oral epithelial junctions facilitates HSV-1 paracellular spread between the epithelial cells. Furthermore, HIV-associated disruption of adherens junctions exposes sequestered nectin-1, an adhesion protein and critical receptor for HSV envelope glycoprotein D (gD). Exposure of nectin-1 facilitates binding of HSV-1 gD, which substantially increases HSV-1 infection of epithelial cells with disrupted junctions over that of cells with intact junctions. Exposed nectin-1 from disrupted adherens junctions also increases the cell-to-cell spread of HSV-1 from infected to uninfected oral epithelial cells. Antibodies to nectin-1 and HSV-1 gD substantially reduce HSV-1 infection and cell-to-cell spread, indicating that HIV-promoted HSV infection and spread are mediated by the interaction of HSV gD with HIV-exposed nectin-1. Our data suggest that HIV-associated disruption of oral epithelial junctions may potentiate HSV-1 infection and its paracellular and cell-to-cell spread within the oral mucosal epithelium. This could be one of the possible mechanisms of rapid development of HSV-associated oral lesions in HIV-infected individuals. PMID:24586397

  6. Epithelial in vitro cell systems in carcinogenesis studies

    SciTech Connect

    Borek, C.

    1983-01-01

    The development of epithelial cells systems to study oncogenic transformation has presented a major challenge in the field of carcinogenesis. Because there exists in man a preponderance of carcinomas over sarcomas, the importance of studying oncogenic transformation in epithelial cells is of great relevance to human disease. The difficulty lies in the fact that different tissues contain epithelial cells with singular differentiated characteristics, which must be defined to assert the different nature of the cells being used. Liver cells in culture are a case in point. By careful maintenance and optimal culture conditions, one can maintain many of the differentiated characteristics of the cells for prolonged periods of time.

  7. A novel cholinergic epithelial cell with chemosensory traits in the murine conjunctiva.

    PubMed

    Wiederhold, Stephanie; Papadakis, Tamara; Chubanov, Vladimir; Gudermann, Thomas; Krasteva-Christ, Gabriela; Kummer, Wolfgang

    2015-11-01

    We recently identified a specialized cholinergic cell type in tracheal and urethral epithelium that utilizes molecules of the canonical taste transduction signaling cascade to sense potentially harmful substances in the luminal content. Upon stimulation, this cell initiates protective reflexes. Assuming a sentinel role of such cells at mucosal surfaces exposed to bacteria, we hypothesized their occurrence also in ocular mucosal surfaces. Utilizing a mouse strain expressing eGFP under the promoter of the acetylcholine synthesizing enzyme, choline acetyltransferase (ChAT-eGFP), we observed a cholinergic cell in the murine conjunctiva. Singular cholinergic cells reaching the epithelial surface with slender processes were detected in fornical, but neither in bulbar nor palpebral epithelia. These cells were found neither in the lacrimal canaliculi, nor in the lacrimal sac and the nasolacrimal duct. Cholinergic conjunctival epithelial cells were immunoreactive for components of the canonical taste transduction signaling cascade, i.e. α-gustducin, phospholipase Cβ2 and the monovalent cation channel TRPM5. Calcitonin gene-related peptide- and substance P-immunoreactive sensory nerve fibers were observed extending into the conjunctival epithelium approaching slender ChAT-eGFP-positive cells. In addition, we noted both ChAT-eGFP expression and α-gustducin-immunoreactivity, albeit in different cell populations, in occasionally occurring lymphoid follicles of the nictitating membrane. The data show a previously unidentified cholinergic cell in murine conjunctiva with chemosensory traits that presumably utilizes acetylcholine for signaling. In analogy to similar cells described in the respiratory and urethral epithelium, it might serve to detect bacterial products and to initiate protective reflexes. PMID:26119492

  8. Heterogeneity and stochastic growth regulation of biliary epithelial cells dictate dynamic epithelial tissue remodeling.

    PubMed

    Kamimoto, Kenji; Kaneko, Kota; Kok, Cindy Yuet-Yin; Okada, Hajime; Miyajima, Atsushi; Itoh, Tohru

    2016-01-01

    Dynamic remodeling of the intrahepatic biliary epithelial tissue plays key roles in liver regeneration, yet the cellular basis for this process remains unclear. We took an unbiased approach based on in vivo clonal labeling and tracking of biliary epithelial cells in the three-dimensional landscape, in combination with mathematical simulation, to understand their mode of proliferation in a mouse liver injury model where the nascent biliary structure formed in a tissue-intrinsic manner. An apparent heterogeneity among biliary epithelial cells was observed: whereas most of the responders that entered the cell cycle upon injury exhibited a limited and tapering growth potential, a select population continued to proliferate, making a major contribution in sustaining the biliary expansion. Our study has highlighted a unique mode of epithelial tissue dynamics, which depends not on a hierarchical system driven by fixated stem cells, but rather, on a stochastically maintained progenitor population with persistent proliferative activity. PMID:27431614

  9. Differential effects of human papillomavirus type 6, 16, and 18 DNAs on immortalization and transformation of human cervical epithelial cells

    SciTech Connect

    Pecoraro, G.; Morgan, D.; Defendi, V. )

    1989-01-01

    The human papillomaviruses (HPVs) are associated with specific benign and malignant lesions of the skin and mucosal epithelia. Cloned viral DNAs from HPV types 6b, 16, and 18 associated with different pathological manifestations of genital neoplasia in vivo were introduced into primary human cervical epithelial cells by electroporation. Cells transfected with HPV16 or HPV18 DNA acquired indefinite lifespans, distinct morphological alterations, and anchorage-independent growth (HPV18), and contain integrated transcriptionally active viral genomes. HPV6b or plasmid electroporated cells senesced at low passage. The alterations in growth and differentiation of the cells appear to reflect the progressive oncogenic processes that result in cervical carcinoma in vivo.

  10. Epidemic Keratoconjunctivitis-Causing Adenoviruses Induce MUC16 Ectodomain Release To Infect Ocular Surface Epithelial Cells.

    PubMed

    Menon, Balaraj B; Zhou, Xiaohong; Spurr-Michaud, Sandra; Rajaiya, Jaya; Chodosh, James; Gipson, Ilene K

    2016-01-01

    Human adenoviruses (HAdV), species D in particular (HAdV-D), are frequently associated with epidemic keratoconjunctivitis (EKC). Although the infection originates at the ocular surface epithelium, the mechanisms by which HAdV-Ds bypass the membrane-associated mucin (MAM)-rich glycocalyx of the ocular surface epithelium to trigger infection and inflammation remain unknown. Here, we report that an EKC-causing adenovirus (HAdV-D37), but not a non-EKC-causing one (HAdV-D19p), induces ectodomain release of MUC16-a MAM with barrier functions at the ocular surface-from cultured human corneal and conjunctival epithelial cells. HAdV-D37, but not HAdV-D19p, is also found to decrease the glycocalyx barrier function of corneal epithelial cells, as determined by rose bengal dye penetrance assays. Furthermore, results from quantitative PCR (qPCR) amplification of viral genomic DNA using primers specific to a conserved region of the E1B gene show that, in comparison to infection by HAdV-D19p, infection by HAdV-D37 is significantly increased in corneal epithelial cells. Collectively, these results point to a MUC16 ectodomain release-dependent mechanism utilized by the EKC-causing HAdV-D37 to initiate infection at the ocular surface. These findings are important in terms of understanding the pathogenesis of adenoviral keratoconjunctivitis. Similar MAM ectodomain release mechanisms may be prevalent across other mucosal epithelia in the body (e.g., the airway epithelium) that are prone to adenoviral infection. IMPORTANCE Human adenoviruses (HAdVs) are double-stranded DNA viruses that cause infections across all mucosal tissues in the body. At the ocular surface, HAdVs cause keratoconjunctivitis (E. Ford, K. E. Nelson, and D. Warren, Epidemiol Rev 9:244-261, 1987, and C. M. Robinson, D. Seto, M. S. Jones, D. W. Dyer, and J. Chodosh, Infect Genet Evol 11:1208-1217, 2011, doi:10.1016/j.meegid.2011.04.031)-a highly contagious infection that accounts for nearly 60% of conjunctivitis cases

  11. Epidemic Keratoconjunctivitis-Causing Adenoviruses Induce MUC16 Ectodomain Release To Infect Ocular Surface Epithelial Cells

    PubMed Central

    Zhou, Xiaohong; Spurr-Michaud, Sandra; Rajaiya, Jaya; Chodosh, James; Gipson, Ilene K.

    2016-01-01

    ABSTRACT Human adenoviruses (HAdV), species D in particular (HAdV-D), are frequently associated with epidemic keratoconjunctivitis (EKC). Although the infection originates at the ocular surface epithelium, the mechanisms by which HAdV-Ds bypass the membrane-associated mucin (MAM)-rich glycocalyx of the ocular surface epithelium to trigger infection and inflammation remain unknown. Here, we report that an EKC-causing adenovirus (HAdV-D37), but not a non-EKC-causing one (HAdV-D19p), induces ectodomain release of MUC16—a MAM with barrier functions at the ocular surface—from cultured human corneal and conjunctival epithelial cells. HAdV-D37, but not HAdV-D19p, is also found to decrease the glycocalyx barrier function of corneal epithelial cells, as determined by rose bengal dye penetrance assays. Furthermore, results from quantitative PCR (qPCR) amplification of viral genomic DNA using primers specific to a conserved region of the E1B gene show that, in comparison to infection by HAdV-D19p, infection by HAdV-D37 is significantly increased in corneal epithelial cells. Collectively, these results point to a MUC16 ectodomain release-dependent mechanism utilized by the EKC-causing HAdV-D37 to initiate infection at the ocular surface. These findings are important in terms of understanding the pathogenesis of adenoviral keratoconjunctivitis. Similar MAM ectodomain release mechanisms may be prevalent across other mucosal epithelia in the body (e.g., the airway epithelium) that are prone to adenoviral infection. IMPORTANCE Human adenoviruses (HAdVs) are double-stranded DNA viruses that cause infections across all mucosal tissues in the body. At the ocular surface, HAdVs cause keratoconjunctivitis (E. Ford, K. E. Nelson, and D. Warren, Epidemiol Rev 9:244–261, 1987, and C. M. Robinson, D. Seto, M. S. Jones, D. W. Dyer, and J. Chodosh, Infect Genet Evol 11:1208–1217, 2011, doi:10.1016/j.meegid.2011.04.031)—a highly contagious infection that accounts for nearly 60% of

  12. Isolation of Cancer Epithelial Cells from Mouse Mammary Tumors

    PubMed Central

    Johnson, Sara; Chen, Hexin; Lo, Pang-Kuo

    2016-01-01

    The isolation of cancer epithelial cells from mouse mammary tumor is accomplished by digestion of the solid tumor. Red blood cells and other contaminates are removed using several washing techniques such that primary epithelial cells can further enriched. This procedure yields primary tumor cells that can be used for in vitro tissue culture, fluorescence-activated cell sorting (FACS) and a wide variety of other experiments (Lo et al., 2012).

  13. Multi-functionality and plasticity characterize epithelial cells in Hydra

    PubMed Central

    Buzgariu, W; Al Haddad, S; Tomczyk, S; Wenger, Y; Galliot, B

    2015-01-01

    Epithelial sheets, a synapomorphy of all metazoans but porifers, are present as 2 layers in cnidarians, ectoderm and endoderm, joined at their basal side by an extra-cellular matrix named mesoglea. In the Hydra polyp, epithelial cells of the body column are unipotent stem cells that continuously self-renew and concomitantly express their epitheliomuscular features. These multifunctional contractile cells maintain homeostasis by providing a protective physical barrier, by digesting nutrients, by selecting a stable microbiota, and by rapidly closing wounds. In addition, epithelial cells are highly plastic, supporting the adaptation of Hydra to physiological and environmental changes, such as long starvation periods where survival relies on a highly dynamic autophagy flux. Epithelial cells also play key roles in developmental processes as evidenced by the organizer activity they develop to promote budding and regeneration. We propose here an integrative view of the homeostatic and developmental aspects of epithelial plasticity in Hydra. PMID:26716072

  14. Multi-functionality and plasticity characterize epithelial cells in Hydra.

    PubMed

    Buzgariu, W; Al Haddad, S; Tomczyk, S; Wenger, Y; Galliot, B

    2015-01-01

    Epithelial sheets, a synapomorphy of all metazoans but porifers, are present as 2 layers in cnidarians, ectoderm and endoderm, joined at their basal side by an extra-cellular matrix named mesoglea. In the Hydra polyp, epithelial cells of the body column are unipotent stem cells that continuously self-renew and concomitantly express their epitheliomuscular features. These multifunctional contractile cells maintain homeostasis by providing a protective physical barrier, by digesting nutrients, by selecting a stable microbiota, and by rapidly closing wounds. In addition, epithelial cells are highly plastic, supporting the adaptation of Hydra to physiological and environmental changes, such as long starvation periods where survival relies on a highly dynamic autophagy flux. Epithelial cells also play key roles in developmental processes as evidenced by the organizer activity they develop to promote budding and regeneration. We propose here an integrative view of the homeostatic and developmental aspects of epithelial plasticity in Hydra. PMID:26716072

  15. Sonic Hedgehog regulates thymic epithelial cell differentiation

    PubMed Central

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

    2016-01-01

    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. PMID

  16. Sonic Hedgehog regulates thymic epithelial cell differentiation.

    PubMed

    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. PMID

  17. Collective Epithelial Migration and Cell Rearrangements Drive Mammary Branching Morphogenesis

    PubMed Central

    Ewald, Andrew J.; Brenot, Audrey; Duong, Myhanh; Chan, Bianca S.; Werb, Zena

    2009-01-01

    Summary Epithelial organs are built through the movement of groups of interconnected cells. We observed cells in elongating mammary ducts reorganize into a multilayered epithelium, migrate collectively, and rearrange dynamically, all without forming leading cellular extensions. Duct initiation required proliferation, Rac, and myosin light-chain kinase, whereas repolarization to a bilayer depended on Rho kinase. We observed that branching morphogenesis results from the active motility of both luminal and myoepithelial cells. Luminal epithelial cells advanced collectively, whereas myoepithelial cells appeared to restrain elongating ducts. Significantly, we observed that normal epithelium and neoplastic hyperplasias are organized similarly during morphogenesis, suggesting common mechanisms of epithelial growth. PMID:18410732

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

    PubMed Central

    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. PMID:25837385

  19. Prominent role for plasmacytoid dendritic cells in mucosal T cell-independent IgA induction.

    PubMed

    Tezuka, Hiroyuki; Abe, Yukiko; Asano, Jumpei; Sato, Taku; Liu, Jiajia; Iwata, Makoto; Ohteki, Toshiaki

    2011-02-25

    Although both conventional dendritic cells (cDCs) and plasmacytoid dendritic cells (pDCs) are present in the gut-associated lymphoid tissues (GALT), the roles of pDCs in the gut remain largely unknown. Here we show a critical role for pDCs in T cell-independent (TI) IgA production by B cells in the GALT. When pDCs of the mesenteric lymph nodes (MLNs) and Peyer's patches (PPs) (which are representative GALT) were cultured with naive B cells to induce TI IgA class switch recombination (CSR), IgA production was substantially higher than in cocultures of these cells with cDCs. IgA production was dependent on APRIL and BAFF production by pDCs. Importantly, pDC expression of APRIL and BAFF was dependent on stromal cell-derived type I IFN signaling under steady-state conditions. Our findings provide insight into the molecular basis of pDC conditioning to induce mucosal TI IgA production, which may lead to improvements in vaccination strategies and treatment for mucosal-related disorders. PMID:21333555

  20. Antiviral factors and type I/III interferon expression associated with regulatory factors in the oral epithelial cells from HIV-1-serodiscordant couples.

    PubMed

    Cervantes, Cesar A C; Oliveira, Luanda M S; Manfrere, Kelly C G; Lima, Josenilson F; Pereira, Natalli Z; Duarte, Alberto J S; Sato, Maria N

    2016-01-01

    Individuals who remain HIV-seronegative despite repeated unprotected exposure to the virus are defined as exposed seronegative (ESN) individuals. Innate and adaptive immunity, as well as genetic factors, provide ESNs with important advantages that allow for low infection susceptibility. The majority of HIV-1-infected individuals undergo antiretroviral therapy, which can decrease the level of HIV-1 exposure in ESNs. We analyzed type I interferon (IFN)-related antiviral and regulatory factors in peripheral blood mononuclear cells (PBMCs) and oral epithelial cells from serodiscordant couples. Our findings revealed that ESNs did not induce the expression of antiviral factors (APOBEC-3G, TRIM5-α, SAMDH1, STING, TBk1) or regulatory factors (Trex, Foxo3, Socs3, IL-10) in PBMCs, unlike their HIV-1-infected partners. In contrast, ESNs upregulated APOBEC-3G and type I/III IFNs (IFNs-α,-β/-λ) in oral mucosal epithelial cells similar to their HIV-infected partners. The serodiscordant groups exhibited an increased expression of type I IFN-induced regulators, such as Trex and Foxo3, in oral epithelial cells. TLR7, TLR8 and TLR9 were expressed in oral epithelial cells of both ESNs and HIV-1-infected subjects. These findings revealed evidence of antiviral factors, type I/III interferon and regulatory factor expression only in the oral mucosal compartment of ESNs, while HIV-1-infected partners systemically and oral mucosal expressed the antiviral profile. PMID:27168019

  1. Antiviral factors and type I/III interferon expression associated with regulatory factors in the oral epithelial cells from HIV-1-serodiscordant couples

    PubMed Central

    Cervantes, Cesar A. C.; Oliveira, Luanda M. S.; Manfrere, Kelly C. G.; Lima, Josenilson F.; Pereira, Natalli Z.; Duarte, Alberto J. S.; Sato, Maria N.

    2016-01-01

    Individuals who remain HIV-seronegative despite repeated unprotected exposure to the virus are defined as exposed seronegative (ESN) individuals. Innate and adaptive immunity, as well as genetic factors, provide ESNs with important advantages that allow for low infection susceptibility. The majority of HIV-1-infected individuals undergo antiretroviral therapy, which can decrease the level of HIV-1 exposure in ESNs. We analyzed type I interferon (IFN)-related antiviral and regulatory factors in peripheral blood mononuclear cells (PBMCs) and oral epithelial cells from serodiscordant couples. Our findings revealed that ESNs did not induce the expression of antiviral factors (APOBEC-3G, TRIM5-α, SAMDH1, STING, TBk1) or regulatory factors (Trex, Foxo3, Socs3, IL-10) in PBMCs, unlike their HIV-1-infected partners. In contrast, ESNs upregulated APOBEC-3G and type I/III IFNs (IFNs-α,-β/-λ) in oral mucosal epithelial cells similar to their HIV-infected partners. The serodiscordant groups exhibited an increased expression of type I IFN-induced regulators, such as Trex and Foxo3, in oral epithelial cells. TLR7, TLR8 and TLR9 were expressed in oral epithelial cells of both ESNs and HIV-1-infected subjects. These findings revealed evidence of antiviral factors, type I/III interferon and regulatory factor expression only in the oral mucosal compartment of ESNs, while HIV-1-infected partners systemically and oral mucosal expressed the antiviral profile. PMID:27168019

  2. Mechanistic roles of epithelial and immune cell signaling during the development of colitis-associated cancer

    PubMed Central

    Subramaniam, Renuka; Mizoguchi, Atsushi; Mizoguchi, Emiko

    2016-01-01

    To date, substantial evidence has shown a significant association between inflammatory bowel diseases (IBD) and development of colitis-associated cancer (CAC). The incidence/prevalence of IBD is higher in western countries including the US, Australia, and the UK. Although CAC development is generally characterized by stepwise accumulation of genetic as well as epigenetic changes, precise mechanisms of how chronic inflammation leads to the development of CAC are largely unknown. Preceding intestinal inflammation is one of the major influential factors for CAC tumorigenesis. Mucosal immune responses including activation of aberrant signaling pathways both in innate and adaptive immune cells play a pivotal role in CAC. Tumor progression and metastasis are shaped by a tightly controlled tumor microenvironment which is orchestrated by several immune cells and stromal cells including macrophages, neutrophils, dendritic cells, myeloid derived suppressor cells, T cells, and myofibroblasts. In this article, we will discuss the contributing factors of epithelial as well as immune cell signaling in initiation of CAC tumorigenesis and mucosal immune regulatory factors in the colonic tumor microenvironment. In depth understanding of these factors is necessary to develop novel anti-inflammatory and anti-cancer therapies for CAC in the near future. PMID:27110580

  3. Epithelial discrimination of commensal and pathogenic Candida albicans.

    PubMed

    Tang, S X; Moyes, D L; Richardson, J P; Blagojevic, M; Naglik, J R

    2016-04-01

    All mucosal surfaces are lined by epithelial cells and are colonised by opportunistic microbes. In health, these opportunistic microbes remain commensal and are tolerated by the immune system. However, when the correct environmental conditions arise, these microbes can become pathogenic and need to be controlled or cleared by the immune system to prevent disease. The mechanisms that enable epithelial cells to initiate the 'danger' signals activated specifically by pathogenic microbes are critical to mucosal defence and homeostasis but are not well understood. Deciphering these mechanisms will provide essential understanding to how mucosal tissues maintain health and activate immunity, as well as how pathogens promote disease. This review focuses on the interaction of the human fungal pathogen Candida albicans with epithelial cells and the epithelial mechanisms that enable mucosal tissues to discriminate between the commensal and pathogenic state of this medically important fungus. PMID:26843519

  4. Duodenal mucosal T cell subpopulation and bacterial cultures in acquired immune deficiency syndrome.

    PubMed

    Budhraja, M; Levendoglu, H; Kocka, F; Mangkornkanok, M; Sherer, R

    1987-05-01

    Enteric infections, chronic diarrhea frequently with no obvious etiology, and weight loss cause major morbidity and mortality in acquired immune deficiency syndrome (AIDS). Alterations in mucosal immunity may explain the increased incidence of enteric infections, and contamination of the upper small intestine with bacteria may be the cause of weight loss observed in these patients. To test this hypothesis we studied the mucosal T lymphocyte subset in duodenal mucosal biopsies in 14 AIDS and seven control patients. Duodenal fluid was also cultured for aerobic and anaerobic bacteria. There was a significant decrease among leu-3a T cells (helper/inducer) subset in AIDS. The proportion of mucosal T cells reacting with leu-2a (cytotoxic/suppressor) was significantly increased in AIDS patients. These patients also had a significant reversal of the normal mucosal helper/suppressor T cell ratio. There was no change in the number of leu-7 cells (cells mediate natural killer and antibody-dependent cellular cytotoxicity) as compared to controls. All patients with diarrhea and three of five patients without diarrhea had bacteria in their duodenal fluid. Mean number of organisms was 4.5 X 10(4)/ml. Cultures were negative in all control subjects. The results reveal that the abnormalities of T cell subpopulation in the blood of AIDS patients also occur in their duodenal mucosa. This immunological abnormality is associated with the bacterial colonization of upper gastrointestinal tract which may explain the diarrhea and weight loss observed in majority of our patients. The results also indicate that increased incidence of enteric infections in AIDS may be explained on the basis of altered mucosal immunity. PMID:2953237

  5. Technical note: Isolation and characterization of porcine mammary epithelial cells.

    PubMed

    Dahanayaka, S; Rezaei, R; Porter, W W; Johnson, G A; Burghardt, R C; Bazer, F W; Hou, Y Q; Wu, Z L; Wu, G

    2015-11-01

    Within the mammary gland, functional synthesis of milk is performed by its epithelial (alveolar) cells. The availability of a stable mammary epithelial cell line is essential for biochemical studies to elucidate cellular and molecular mechanisms responsible for nutritional regulation of lactation. Therefore, porcine mammary epithelial cells (PMEC) were isolated from mammary glands of a 9-mo-old nonpregnant and nonlactating gilt and cultured to establish a nonimmortalized cell line. These cells were characterized by expression of cytokeratin-18 (an intermediate filament specific for epithelial cells), β-casein (a specific marker for mammary epithelial cells), and α-lactalbumin. In culture, the PMEC doubled in number every 24 h and maintained a cobblestone morphology, typical for cultured epithelial cells, for at least 15 passages. Addition of 0.2 to 2 μg/mL prolactin to culture medium for 3 d induced the production of β-casein and α-lactalbumin by PMEC in a dose-dependent manner. Thus, we have successfully developed a useful PMEC line for future studies of cellular and molecular regulation of milk synthesis by mammary epithelial cells of the sow. PMID:26641038

  6. Transforming growth factor-beta mediates intestinal healing and susceptibility to injury in vitro and in vivo through epithelial cells.

    PubMed

    Beck, Paul L; Rosenberg, Ian M; Xavier, Ramnik J; Koh, Theodore; Wong, Josée F; Podolsky, Daniel K

    2003-02-01

    In vitro studies suggest that transforming growth factor (TGF)-beta has potent effects on gastrointestinal mucosal integrity, wound repair, and neoplasia. However, the multiplicity of actions of this peptide on many different cell types confounds efforts to define the role of TGF-beta within the intestinal epithelium in vivo. To delineate these effects selective blockade of intestinal epithelial TGF-beta activity was undertaken through targeted expression of a dominant-negative (DN) TGF-beta RII to intestinal epithelial cells in vitro and in vivo. Stable intestinal epithelial cell (IEC)-6 lines overexpressing TGF-beta RII-DN (nucleotides -7 to 573) were established. Transgenic mice overexpressing TGF-beta RII-DN under the regulation of a modified liver fatty acid-binding promoter (LFABP-PTS4) were constructed. In vitro healing was assessed by wounding of confluent monolayers. Colitis was induced by the addition of dextran sodium sulfate (2.5 to 7.5% w/v) to their drinking water. Overexpression of TGF-beta RII-DN in intestinal epithelial cell-6 cells resulted in a marked reduction in cell migration and TGF-beta-stimulated wound healing in vitro. TGF-beta RII-DN transgenic mice did not exhibit baseline intestinal inflammation or changes in survival, body weight, epithelial cell proliferation, aberrant crypt foci, or tumor formation. TGF-beta RII-DN mice were markedly more susceptible to dextran sodium sulfate-induced colitis and exhibited impaired recovery after colonic injury. TGF-beta is required for intestinal mucosal healing and TGF-beta modulation of the intestinal epithelium plays a central role in determining susceptibility to injury. PMID:12547717

  7. Mesenchymal-epithelial interactions during digestive tract development and epithelial stem cell regeneration.

    PubMed

    Le Guen, Ludovic; Marchal, Stéphane; Faure, Sandrine; de Santa Barbara, Pascal

    2015-10-01

    The gastrointestinal tract develops from a simple and uniform tube into a complex organ with specific differentiation patterns along the anterior-posterior and dorso-ventral axes of asymmetry. It is derived from all three germ layers and their cross-talk is important for the regulated development of fetal and adult gastrointestinal structures and organs. Signals from the adjacent mesoderm are essential for the morphogenesis of the overlying epithelium. These mesenchymal-epithelial interactions govern the development and regionalization of the different gastrointestinal epithelia and involve most of the key morphogens and signaling pathways, such as the Hedgehog, BMPs, Notch, WNT, HOX, SOX and FOXF cascades. Moreover, the mechanisms underlying mesenchyme differentiation into smooth muscle cells influence the regionalization of the gastrointestinal epithelium through interactions with the enteric nervous system. In the neonatal and adult gastrointestinal tract, mesenchymal-epithelial interactions are essential for the maintenance of the epithelial regionalization and digestive epithelial homeostasis. Disruption of these interactions is also associated with bowel dysfunction potentially leading to epithelial tumor development. In this review, we will discuss various aspects of the mesenchymal-epithelial interactions observed during digestive epithelium development and differentiation and also during epithelial stem cell regeneration. PMID:26126787

  8. Cell Chirality Induces Collective Cell Migration in Epithelial Sheets

    NASA Astrophysics Data System (ADS)

    Sato, Katsuhiko; Hiraiwa, Tetsuya; Shibata, Tatsuo

    2015-10-01

    During early development, epithelial cells form a monolayer sheet and migrate in a uniform direction. Here, we address how this collective migration can occur without breaking the cell-to-cell attachments. Repeated contraction and expansion of the cell-to-cell interfaces enables the cells to rearrange their positions autonomously within the sheet. We show that when the interface tension is strengthened in a direction that is tilted from the body axis, cell rearrangements occur in such a way that unidirectional movement is induced. We use a vertex model to demonstrate that such anisotropic tension can generate the unidirectional motion of cell sheets. Our results suggest that cell chirality facilitates collective cell migration during tissue morphogenesis.

  9. Fungal glycan interactions with epithelial cells in allergic airway disease

    PubMed Central

    Roy, René M.; Klein, Bruce S.

    2014-01-01

    Human exposure to fungi results in a wide range of health outcomes, from invasive disease or allergy to immune tolerance. Inhaled fungi contact airway epithelial cells as an early event, and this host:fungal interaction can shape the eventual immunological outcome. Emerging evidence points to exposure to fungal cell wall carbohydrates in the development of allergic airway disease. Herein, we describe determinants of fungal allergenicity, and review the responses of airway epithelial cells to fungal carbohydrates. A greater understanding of the recognition of and response to fungal carbohydrates by airway epithelial cells may lead to the development of targeted therapies that ameliorate allergic airway disease. PMID:23602359

  10. Quantitative Assessment of Cytosolic Salmonella in Epithelial Cells

    PubMed Central

    Knodler, Leigh A.; Nair, Vinod; Steele-Mortimer, Olivia

    2014-01-01

    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. PMID:24400108

  11. Allosteric properties of phosphofructokinase from the epithelial cells of thermally injured rat small intestine.

    PubMed

    Khoja, S M; Salleh, M; Ardawi, M

    1987-01-01

    1. The allosteric properties of phosphofructokinase from the epithelial cells of thermally injured rat small intestine were studied and compared with those properties of the normal rats. 2. The fructose 6-phosphate saturation curve of mucosal phosphofructokinase from thermally injured rats (3 days post injury, 33% of body surface area) displayed cooperatively; the ratio of the activity observed at pH 7.0 in the presence of 0.5 mM fructose 6-phosphate and 2.5 mM-ATP to the optimal activity at pH 8.0, v 0.5/V, was 0.42 +/- 0.02 in the normal rats and 0.22 +/- 0.03 in the injured rats. 3. The enzyme from thermally injured rats was very sensitive to inhibition by ATP as compared to that from normal rats. 4. The enzyme from thermally injured rats was inhibited by citrate and phosphocreatine in a synergistic manner with ATP. 5. Activation under nearly cellular conditions was produced by ADP, AMP and glucose-1,6-biphosphate. 6. In general, the mucosal enzyme of thermally injured rats was more susceptible to inhibition or activation by various metabolites than the enzyme of the normal rats. 7. These results may suggest that mucosal phosphofructokinase of thermally injured rats may not be subject to the same control mechanism as the normal rats in vivo due to changes in the concentrations of fructose-2,6-biphosphate. PMID:2957148

  12. [Values of the micronucleus test on animal epithelial cells exposed to titanium dioxide].

    PubMed

    Iurchenko, V V; Krivtsova, E K; Iuretseva, N A; Tul'skaia, E A; Mamonov, R A; Zholdakova, Z I; Sinitsyna, O O; Mal'tseva, M M; Pankratova, G P; Sycheva, L P

    2011-01-01

    The genetic safety of titanium dioxide (TD)-containing foods and cosmetic products has been little investigated. The study evaluated the mutagenic activity of TD in the micronucleus test with animal visceral mucosal epithelial cells. Two simethicone-coated anatase samples (mean size 160 and 33.2 nm) were inserted into the mouse stomach in doses of 40-200-1000 mg/kg seven times and applied as an ingredient of 10 and 25% cream (doses 250 and 625 mg/kg, respectively) to the hair-sheared rat skin once for 4 hours. Analysis of cytogenetic disorders (micronuclei, protrusions, and the atypical form of the nucleus) revealed no mutagenic properties of TD on the mucosal epithelium of the mouse and rat intestine, mouse prostomach, and rat uterine bladder. Enhanced mitotic activity was observed in all the study tissues after exposure of both samples to TD given in some or in all (in the rat urinary bladder mucosal epithelium) doses. PMID:22185006

  13. Lung epithelial cells modulate the inflammatory response of alveolar macrophages.

    PubMed

    Rubovitch, Vardit; Gershnabel, Shoham; Kalina, Moshe

    2007-12-01

    The goal of this study was to examine the effect of alveolar epithelial cells on inflammatory responses in macrophages. Lung epithelial cells (either rat RLE-6TN or human A549 cells) reduced LPS-induced NO production in alveolar macrophages (AM) in a contact-independent mechanism. The inhibitory effect of the epithelial cells was present already at the transcriptional level: LPS-induced inducible NO synthase (iNOS) expression was significantly smaller. Surfactant protein A (SP-A)-induced NO production by alveolar macrophages was also reduced in the presence of A549 cells, though, by a different kinetics. LPS-induced interleukin-6 (IL-6) production (another inflammatory pathway) by alveolar macrophages was also reduced in the presence of RLE-6TN cells. These data suggest a role for lung epithelial cells in the complicated modulation of inflammatory processes, and provide an insight into the mechanism underlying. PMID:17851743

  14. Epithelial cells as alternative human biomatrices for comet assay

    PubMed Central

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

    2014-01-01

    The comet assay is a valuable experimental tool aimed at mapping DNA damage in human cells in vivo for environmental and occupational monitoring, as well as for therapeutic purposes, such as storage prior to transplant, during tissue engineering, and in experimental ex vivo assays. Furthermore, due to its great versatility, the comet assay allows to explore the use of alternative cell types to assess DNA damage, such as epithelial cells. Epithelial cells, as specialized components of many organs, have the potential to serve as biomatrices that can be used to evaluate genotoxicity and may also serve as early effect biomarkers. Furthermore, 80% of solid cancers are of epithelial origin, which points to the importance of studying DNA damage in these tissues. Indeed, studies including comet assay in epithelial cells have either clear clinical applications (lens and corneal epithelial cells) or examine genotoxicity within human biomonitoring and in vitro studies. We here review improvements in determining DNA damage using the comet assay by employing lens, corneal, tear duct, buccal, and nasal epithelial cells. For some of these tissues invasive sampling procedures are needed. Desquamated epithelial cells must be obtained and dissociated prior to examination using the comet assay, and such procedures may induce varying amounts of DNA damage. Buccal epithelial cells require lysis enriched with proteinase K to obtain free nucleosomes. Over a 30 year period, the comet assay in epithelial cells has been little employed, however its use indicates that it could be an extraordinary tool not only for risk assessment, but also for diagnosis, prognosis of treatments and diseases. PMID:25506353

  15. Intrinsic epithelial cells repair the kidney after injury.

    PubMed

    Humphreys, Benjamin D; Valerius, M Todd; Kobayashi, Akio; Mugford, Joshua W; Soeung, Savuth; Duffield, Jeremy S; McMahon, Andrew P; Bonventre, Joseph V

    2008-03-01

    Understanding the mechanisms of nephron repair is critical for the design of new therapeutic approaches to treat kidney disease. The kidney can repair after even a severe insult, but whether adult stem or progenitor cells contribute to epithelial renewal after injury and the cellular origin of regenerating cells remain controversial. Using genetic fate-mapping techniques, we generated transgenic mice in which 94%-95% of tubular epithelial cells, but no interstitial cells, were labeled with either beta-galactosidase (lacZ) or red fluorescent protein (RFP). Two days after ischemia-reperfusion injury (IRI), 50.5% of outer medullary epithelial cells coexpress Ki67 and RFP, indicating that differentiated epithelial cells that survived injury undergo proliferative expansion. After repair was complete, 66.9% of epithelial cells had incorporated BrdU, compared to only 3.5% of cells in the uninjured kidney. Despite this extensive cell proliferation, no dilution of either cell-fate marker was observed after repair. These results indicate that regeneration by surviving tubular epithelial cells is the predominant mechanism of repair after ischemic tubular injury in the adult mammalian kidney. PMID:18371453

  16. Clinical implications of epithelial cell plasticity in cancer progression.

    PubMed

    Aparicio, Luis A; Blanco, Moisés; Castosa, Raquel; Concha, Ángel; Valladares, Manuel; Calvo, Lourdes; Figueroa, Angélica

    2015-09-28

    In the last few years, the role of epithelial cell plasticity in cancer biology research has gained increasing attention. This concept refers to the ability of the epithelial cells to dynamically switch between different phenotypic cellular states. This programme is particularly relevant during the epithelial-to-mesenchymal transition (EMT) in cancer progression. During colonization, epithelial cells first activate the EMT programme to disseminate from a primary tumour to reach a distant tissue site. During this process, cells are transported into the circulation and are able to escape the immune system of the host. Then, a reverse process called mesenchymal-to-epithelial transition (MET) occurs on cells that settle in the distant organs. Although epithelial cell plasticity has an important impact on tumour biology, the clinical relevance of this concept remains to be recapitulated. In this review, we will update the current state of epithelial cell plasticity in cancer progression and its clinical implications for the design of therapeutic strategies, the acquisition of multidrug resistance, and future perspectives for the management of cancer patients. PMID:26099173

  17. Tsr Chemoreceptor Interacts With IL-8 Provoking E. coli Transmigration Across Human Lung Epithelial Cells

    PubMed Central

    Han, Bing; Li, Manshu; Xu, Yonghao; Islam, Diana; Khang, Julie; Del Sorbo, Lorenzo; Lee, Warren; Szaszi, Katalin; Zhong, Nanshan; Slutsky, Arthur S.; Li, Yimin; Zhang, Haibo

    2016-01-01

    Bacterial colonization of epithelial surfaces and subsequent transmigration across the mucosal barrier are essential for the development of infection. We hypothesized that the methyl-accepting proteins (MCPs), known as chemoreceptors expressed on Escherichia coli (E. coli) bacterial surface, play an important role in mediating bacterial transmigration. We demonstrated a direct interaction between human interleukin-8 (IL-8) and Tsr receptor, a major MCP chemoreceptor. Stimulation of human lung epithelial cell monolayer with IL-8 resulted in increased E. coli adhesion and transmigration of the native strain (RP437) and a strain expressing only Tsr (UU2373), as compared to a strain (UU2599) with Tsr truncation. The augmented E. coli adhesion and migration was associated with a higher expression of carcinoembryonic antigen-related cell adhesion molecule 6 and production of inflammatory cytokines/chemokines, and a lower expression of the tight junction protein claudin-1 and the plasma membrane protein caveolin-1 in lung epithelial cells. An increased E. coli colonization and pulmonary cytokine production induced by the RP437 and UU2373 strains was attenuated in mice challenged with the UU2599 strain. Our results suggest a critical role of the E. coli Tsr chemoreceptor in mediating bacterial colonization and transmigration across human lung epithelium during development of pulmonary infections. PMID:27506372

  18. Free and complexed-secretory immunoglobulin A triggers distinct intestinal epithelial cell responses.

    PubMed

    Salerno-Goncalves, R; Safavie, F; Fasano, A; Sztein, M B

    2016-09-01

    Secretory immunoglobulin A (SIgA) antibodies play an important role in protecting the mucosal surfaces against pathogens and maintaining homeostasis with the commensal microbiota. Because a substantial portion of the gut microbiota is coated with SIgA, we hypothesized that microbiota-SIgA complexes are important for the maintenance of gut homeostasis. Here we investigated the relationship between microbiota-SIgA complexes and inflammatory epithelial cell responses. We used a multi-cellular three-dimensional (3D) organotypical model of the human intestinal mucosa composed of an intestinal epithelial cell line and primary human lymphocytes/monocytes, endothelial cells and fibroblasts. We also used human SIgA from human colostrum, and a prominent bacterial member of the first colonizers, Escherichia coli, as a surrogate commensal. We found that free and microbiota-complexed SIgA triggered different epithelial responses. While free SIgA up-regulated mucus production, expression of polymeric immunoglobulin receptor (pIgR) and secretion of interleukin-8 and tumoir necrosis factor-α, microbiota-complexed SIgA mitigated these responses. These results suggest that free and complexed SIgA have different functions as immunoregulatory agents in the gut and that an imbalance between the two may affect gut homeostasis. PMID:27084834

  19. Tsr Chemoreceptor Interacts With IL-8 Provoking E. coli Transmigration Across Human Lung Epithelial Cells.

    PubMed

    Han, Bing; Li, Manshu; Xu, Yonghao; Islam, Diana; Khang, Julie; Del Sorbo, Lorenzo; Lee, Warren; Szaszi, Katalin; Zhong, Nanshan; Slutsky, Arthur S; Li, Yimin; Zhang, Haibo

    2016-01-01

    Bacterial colonization of epithelial surfaces and subsequent transmigration across the mucosal barrier are essential for the development of infection. We hypothesized that the methyl-accepting proteins (MCPs), known as chemoreceptors expressed on Escherichia coli (E. coli) bacterial surface, play an important role in mediating bacterial transmigration. We demonstrated a direct interaction between human interleukin-8 (IL-8) and Tsr receptor, a major MCP chemoreceptor. Stimulation of human lung epithelial cell monolayer with IL-8 resulted in increased E. coli adhesion and transmigration of the native strain (RP437) and a strain expressing only Tsr (UU2373), as compared to a strain (UU2599) with Tsr truncation. The augmented E. coli adhesion and migration was associated with a higher expression of carcinoembryonic antigen-related cell adhesion molecule 6 and production of inflammatory cytokines/chemokines, and a lower expression of the tight junction protein claudin-1 and the plasma membrane protein caveolin-1 in lung epithelial cells. An increased E. coli colonization and pulmonary cytokine production induced by the RP437 and UU2373 strains was attenuated in mice challenged with the UU2599 strain. Our results suggest a critical role of the E. coli Tsr chemoreceptor in mediating bacterial colonization and transmigration across human lung epithelium during development of pulmonary infections. PMID:27506372

  20. Infection of human fallopian tube epithelial cells with Neisseria gonorrhoeae protects cells from tumor necrosis factor alpha-induced apoptosis.

    PubMed

    Morales, Priscilla; Reyes, Paz; Vargas, Macarena; Rios, Miguel; Imarai, Mónica; Cardenas, Hugo; Croxatto, Horacio; Orihuela, Pedro; Vargas, Renato; Fuhrer, Juan; Heckels, John E; Christodoulides, Myron; Velasquez, Luis

    2006-06-01

    Following infection with Neisseria gonorrhoeae, bacteria may ascend into the Fallopian tubes (FT) and induce salpingitis, a major cause of infertility. In the FT, interactions between mucosal epithelial cells and gonococci are pivotal events in the pathogen's infection cycle and the inflammatory response. In the current study, primary FT epithelial cells were infected in vitro with different multiplicities of infection (MOI) of Pil+ Opa+ gonococci. Bacteria showed a dose-dependent association with cells and induced the secretion of tumor necrosis factor alpha (TNF-alpha). A significant finding was that gonococcal infection (MOI = 1) induced apoptosis in approximately 30% of cells, whereas increasing numbers of bacteria (MOI = 10 to 100) did not induce apoptosis. Apoptosis was observed in only 11% of cells with associated bacteria, whereas >84% of cells with no adherent bacteria were apoptotic. TNF-alpha was a key contributor to apoptosis, since (i) culture supernatants from cells infected with gonococci (MOI = 1) induced apoptosis in naïve cultures, suggesting that a soluble factor was responsible; (ii) gonococcal infection-induced apoptosis was inhibited with anti-TNF-alpha antibodies; and (iii) the addition of exogenous TNF-alpha induced apoptosis, which was inhibited by the presence of increasing numbers of bacteria (MOI = 10 to 100). These data suggest that TNF-alpha-mediated apoptosis of FT epithelial cells is likely a primary host defense mechanism to prevent pathogen colonization. However, epithelial cell-associated gonococci have evolved a mechanism to protect the cells from undergoing TNF-alpha-mediated apoptosis, and this modulation of the host innate response may contribute to establishment of infection. Understanding the antiapoptotic mechanisms used by Neisseria gonorrhoeae will inform the pathogenesis of salpingitis and could suggest new intervention strategies for prevention and treatment of the disease. PMID:16714596

  1. Immunomodulatory effect of cathelicidins in response to a β-glucan in intestinal epithelial cells from rainbow trout.

    PubMed

    Schmitt, Paulina; Wacyk, Jurij; Morales-Lange, Byron; Rojas, Verónica; Guzmán, Fanny; Dixon, Brian; Mercado, Luis

    2015-07-01

    The aim of the present study was to characterize intestinal immune mechanisms involved in the response to β-glucans in rainbow trout. Among the immune effectors regulated in response to immunostimulants, host defense peptides (HDPs) are abundantly expressed in epithelial linings, suggesting their important role in the mucosal immune response. Therefore, the immunomodulatory properties of expressed HDPs in the epithelial intestinal cells of rainbow trout in response to the β-glucan, zymosan, were assessed. The results showed that zymosan increased the production of the HDP, cathelicidin, and the cytokine, IL-1β, in the intestinal epithelial RTgutGC cell line at the transcript and protein levels. Thus, cathelicidin-2 variants were produced and were shown to (i) induce the production of IL-1β in RTgutGC cells and (ii) display a synergic effect with zymosan in IL-1β upregulation. Importantly, the colocalization of both rtCATH-2 and IL-1β was detected in the intestinal epithelial cells of rainbow trout fed with a 0.3% zymosan-supplemented diet. We propose that trout cathelicidins are expressed by intestinal epithelial cells and exert immunomodulatory effects to improve the local intestinal immune response triggered by immunostimulants. PMID:25818364

  2. Liver epithelial cells inhibit proliferation and invasiveness of hepatoma cells.

    PubMed

    Jeng, Kuo-Shyang; Jeng, Chi-Juei; Jeng, Wen-Juei; Sheen, I-Shyan; Li, Shih-Yun; Hung, Zih-Hang; Hsiau, Hsin-I; Yu, Ming-Che; Chang, Chiung-Fang

    2016-03-01

    Hepatocellular carcinoma (HCC) is a worldwide malignancy with poor prognosis. Liver progenitors or stem cells could be a potential therapy for HCC treatment since they migrate toward tumors. Rat liver epithelial (RLE) cells have both progenitor and stem cell-like properties. Therefore, our study elucidated the therapeutic effect of RLE cells in rat hepatoma cells. RLE cells were isolated from 10-day old rats and characterized for stem cell marker expression. RLE cells and rat hepatoma cells (H4-IIE-C3 cells) were co-cultured and divided into four groups with different ratios of RLE and hepatoma cells. Group A had only rat hepatoma cells as a control group. The ratios of rat hepatoma and RLE cells in group B, C and D were 5:1, 1:1 and 1:5, respectively. Effective inhibition of cell proliferation and migration was found in group D when compared to group A. There was a significant decrease in Bcl2 expression and increase in late apoptosis of rat hepatoma cells when adding more RLE cells. RLE cells reduced cell proliferation and migration of rat hepatoma cells. These results suggested that RLE cells could be used as a potential cell therapy. PMID:26647726

  3. Neisseria gonorrhoeae infection protects human endocervical epithelial cells from apoptosis via expression of host antiapoptotic proteins.

    PubMed

    Follows, S A; Murlidharan, J; Massari, P; Wetzler, L M; Genco, C A

    2009-09-01

    Several microbial pathogens can modulate the host apoptotic response to infection, which may contribute to immune evasion. Various studies have reported that infection with the sexually transmitted disease pathogen Neisseria gonorrhoeae can either inhibit or induce apoptosis. N. gonorrhoeae infection initiates at the mucosal epithelium, and in women, cells from the ectocervix and endocervix are among the first host cells encountered by this pathogen. In this study, we defined the antiapoptotic effect of N. gonorrhoeae infection in human endocervical epithelial cells (End/E6E7 cells). We first established that N. gonorrhoeae strain FA1090B failed to induce cell death in End/E6E7 cells. Subsequently, we demonstrated that stimulation with N. gonorrhoeae protected these cells from staurosporine (STS)-induced apoptosis. Importantly, only End/E6E7 cells incubated with live bacteria and in direct association with N. gonorrhoeae were protected from STS-induced apoptosis, while heat-killed and antibiotic-killed bacteria failed to induce protection. Stimulation of End/E6E7 cells with live N. gonorrhoeae induced NF-kappaB activation and resulted in increased gene expression of the NF-kappaB-regulated antiapoptotic genes bfl-1, cIAP-2, and c-FLIP. Furthermore, cIAP-2 protein levels also increased in End/E6E7 cells incubated with gonococci. Collectively, our results indicate that the antiapoptotic effect of N. gonorrhoeae in human endocervical epithelial cells results from live infection via expression of host antiapoptotic proteins. Securing an intracellular niche through the inhibition of apoptosis may be an important mechanism utilized by N. gonorrhoeae for microbial survival and immune evasion in cervical epithelial cells. PMID:19546192

  4. The mucosal immune system for vaccine development.

    PubMed

    Lamichhane, Aayam; Azegamia, Tatsuhiko; Kiyonoa, Hiroshi

    2014-11-20

    Mucosal surfaces are continuously exposed to the external environment and therefore represent the largest lymphoid organ of the body. In the mucosal immune system, gut-associated lymphoid tissues (GALTs), including Peyer's patches and isolated lymphoid follicles, play an important role in the induction of antigen-specific immune responses in the gut. GALTs have unique organogenesis characteristics and interact with the network of dendritic cells and T cells for the simultaneous induction and regulation of IgA responses and oral tolerance. In these lymphoid tissues, antigens are up taken by M cells in the epithelial layer, and antigen-specific immune responses are subsequently initiated by GALT cells. Nasopharynx- and tear-duct-associated lymphoid tissues (NALTs and TALTs) are key organized lymphoid structures in the respiratory tract and ocular cavities, respectively, and have been shown to interact with each other. Mucosal surfaces are also characterized by host-microbe interactions that affect the genesis and maturation of mucosa-associated lymphoid tissues and the induction and regulation of innate and acquired mucosal immune responses. Because most harmful pathogens enter the body through mucosal surfaces by ingestion, inhalation, or sexual contact, the mucosa is a candidate site for vaccination. Mucosal vaccination has some physiological and practical advantages, such as decreased costs and reduced risk of needle-stick injuries and transmission of bloodborne diseases, and it is painless. Recently, the application of modern bioengineering and biochemical engineering technologies, including gene transformation and manipulation systems, resulted in the development of systems to express vaccine antigens in transgenic plants and nanogels, which will usher in a new era of delivery systems for mucosal vaccine antigens. In this review, based on some of our research group's thirty seven years of progress and effort, we highlight the unique features of mucosal immune

  5. Strong mucosal immune responses in SIV infected macaques contribute to viral control and preserved CD4+ T-cell levels in blood and mucosal tissues

    PubMed Central

    2011-01-01

    Background Since there is still no protective HIV vaccine available, better insights into immune mechanism of persons effectively controlling HIV replication in the absence of any therapy should contribute to improve further vaccine designs. However, little is known about the mucosal immune response of this small unique group of patients. Using the SIV-macaque-model for AIDS, we had the rare opportunity to analyze 14 SIV-infected rhesus macaques durably controlling viral replication (controllers). We investigated the virological and immunological profile of blood and three different mucosal tissues and compared their data to those of uninfected and animals progressing to AIDS-like disease (progressors). Results Lymphocytes from blood, bronchoalveolar lavage (BAL), and duodenal and colonic biopsies were phenotypically characterized by polychromatic flow cytometry. In controllers, we observed higher levels of CD4+, CD4+CCR5+ and Gag-specific CD8+ T-cells as well as lower immune activation in blood and all mucosal sites compared to progressors. However, we could also demonstrate that immunological changes are distinct between these three mucosal sites. Intracellular cytokine staining demonstrated a significantly higher systemic and mucosal CD8+ Gag-specific cellular immune response in controllers than in progressors. Most remarkable was the polyfunctional cytokine profile of CD8+ lymphocytes in BAL of controllers, which significantly dominated over their blood response. The overall suppression of viral replication in the controllers was confirmed by almost no detectable viral RNA in blood and all mucosal tissues investigated. Conclusion A strong and complex virus-specific CD8+ T-cell response in blood and especially in mucosal tissue of SIV-infected macaques was associated with low immune activation and an efficient suppression of viral replication. This likely afforded a repopulation of CD4+ T-cells in different mucosal compartments to almost normal levels. We

  6. Sodium selectivity of Reissner's membrane epithelial cells

    PubMed Central

    2011-01-01

    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 media. PMID:21284860

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

    PubMed

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

    2015-06-01

    Epithelial mesenchymal transition (EMT) of lens epithelial cells (LECs) may contribute to the development of posterior capsular opacification (PCO), which leads to visual impairment. Andrographolide has been shown to have therapeutic potential against various cancers. However, its effect on human LECs is still unknown. The purpose of this study is to evaluate the effect of andrographolide on EMT induced by growth factors in the fetal human lens epithelial cell line (FHL 124). Initially the LECs were treated with growth factors (TGF-beta 2 and bFGF) to induce EMT. Subsequently these EMT-induced cells were treated with andrographolide at 100 and 500 nM concentrations for 24 h. Our results showed that FHL 124 cells treated with growth factors had a significant decrease in protein and m-RNA levels of epithelial markers pax6 and E-Cadherin. After administering andrographolide, these levels significantly increased. It was noticed that EMT markers alpha-SMA, fibronectin and collagen IV significantly decreased after treatment with andrographolide when compared to the other group. Treatment with andrographolide significantly inhibited phosphorylation of ERK and JNK. Cell cycle analysis showed that andrographolide did not arrest cells at G0/G1 or G2/M at tested concentrations. Our findings suggest that andrographolide helps sustain epithelial characteristics by modulating EMT markers and inhibiting the mitogen-activated protein kinase (MAPK) signalling pathway in LECs. Hence it can prove to be useful in curbing EMT-mediated PCO. PMID:25963259

  8. Human mucosal-associated invariant T cells contribute to antiviral influenza immunity via IL-18-dependent activation.

    PubMed

    Loh, Liyen; Wang, Zhongfang; Sant, Sneha; Koutsakos, Marios; Jegaskanda, Sinthujan; Corbett, Alexandra J; Liu, Ligong; Fairlie, David P; Crowe, Jane; Rossjohn, Jamie; Xu, Jianqing; Doherty, Peter C; McCluskey, James; Kedzierska, Katherine

    2016-09-01

    Mucosal-associated invariant T (MAIT) cells are innate-like T lymphocytes known to elicit potent immunity to a broad range of bacteria, mainly via the rapid production of inflammatory cytokines. Whether MAIT cells contribute to antiviral immunity is less clear. Here we asked whether MAIT cells produce cytokines/chemokines during severe human influenza virus infection. Our analysis in patients hospitalized with avian H7N9 influenza pneumonia showed that individuals who recovered had higher numbers of CD161(+)Vα7.2(+) MAIT cells in peripheral blood compared with those who succumbed, suggesting a possible protective role for this lymphocyte population. To understand the mechanism underlying MAIT cell activation during influenza, we cocultured influenza A virus (IAV)-infected human lung epithelial cells (A549) and human peripheral blood mononuclear cells in vitro, then assayed them by intracellular cytokine staining. Comparison of influenza-induced MAIT cell activation with the profile for natural killer cells (CD56(+)CD3(-)) showed robust up-regulation of IFNγ for both cell populations and granzyme B in MAIT cells, although the individual responses varied among healthy donors. However, in contrast to the requirement for cell-associated factors to promote NK cell activation, the induction of MAIT cell cytokine production was dependent on IL-18 (but not IL-12) production by IAV-exposed CD14(+) monocytes. Overall, this evidence for IAV activation via an indirect, IL-18-dependent mechanism indicates that MAIT cells are protective in influenza, and also possibly in any human disease process in which inflammation and IL-18 production occur. PMID:27543331

  9. Serum-Induced Differentiation of Human Meibomian Gland Epithelial Cells

    PubMed Central

    Sullivan, David A.; Liu, Yang; Kam, Wendy R.; Ding, Juan; Green, Karin M.; Shaffer, Scott A.; Hatton, Mark P.; Liu, Shaohui

    2014-01-01

    Purpose. We hypothesize that culturing immortalized human meibomian gland epithelial cells in serum-containing medium will induce their differentiation. The purpose of this investigation was to begin to test our hypothesis, and explore the impact of serum on gene expression and lipid accumulation in human meibomian gland epithelial cells. Methods. Immortalized and primary human meibomian gland epithelial cells were cultured in the presence or absence of serum. Cells were evaluated for lysosome and lipid accumulation, polar and neutral lipid profiles, and gene expression. Results. Our results support our hypothesis that serum stimulates the differentiation of human meibomian gland epithelial cells. This serum-induced effect is associated with a significant increase in the expression of genes linked to cell differentiation, epithelium development, the endoplasmic reticulum, Golgi apparatus, vesicles, and lysosomes, and a significant decrease in gene activity related to the cell cycle, mitochondria, ribosomes, and translation. These cellular responses are accompanied by an accumulation of lipids within lysosomes, as well as alterations in the fatty acid content of polar and nonpolar lipids. Of particular importance, our results show that the molecular and biochemical changes of immortalized human meibomian gland epithelial cells during differentiation are analogous to those of primary cells. Conclusions. Overall, our findings indicate that immortalized human meibomian gland epithelial cells may serve as an ideal preclinical model to identify factors that control cellular differentiation in the meibomian gland. PMID:24867579

  10. Sepsis-associated AKI: epithelial cell dysfunction.

    PubMed

    Emlet, David R; Shaw, Andrew D; Kellum, John A

    2015-01-01

    Acute kidney injury (AKI) occurs frequently in critically ill patients with sepsis, in whom it doubles the mortality rate and half of the survivors suffer permanent kidney damage or chronic kidney disease. Failure in the development of viable therapies has prompted studies to better elucidate the cellular and molecular etiologies of AKI, which have generated novel theories and paradigms for the mechanisms of this disease. These studies have shown multifaceted origins and elements of AKI that, in addition to/in lieu of ischemia, include the generation of damage-associated molecular patterns and pathogen-associated molecular patterns, the inflammatory response, humoral and cellular immune activation, perturbation of microvascular flow and oxidative stress, bioenergetic alterations, cell-cycle alterations, and cellular de-differentiation/re-differentiation. It is becoming clear that a major etiologic effector of all these inputs is the renal tubule epithelial cell (RTEC). This review discusses these elements and their effects on RTECs, and reviews the current hypotheses of how these effects may determine the fate of RTECs during sepsis-induced AKI. PMID:25795502

  11. Genetics and epithelial cell dysfunction in cystic fibrosis

    SciTech Connect

    Riordan, J.R.; Buchwald, M.

    1987-01-01

    This book examines the advances being made in the study of the physiology, cell biology, and molecular genetics of cystic fibrosis. Emphasis is placed on various areas of research that involve epithelial cells (e.g., the CF-specific phenotypes exhibited by epithelial cells, abnormalities in epithelium ion transport, chloride channel regulation in CF epithelial.) Coverage is presented on the current status of CF, including data on the incidence of the disease, its mode of inheritance, chromosomal localization, genetic heterogeneity, and screening and management.

  12. Alignment of cell division axes in directed epithelial cell migration

    NASA Astrophysics Data System (ADS)

    Marel, Anna-Kristina; Podewitz, Nils; Zorn, Matthias; Oskar Rädler, Joachim; Elgeti, Jens

    2014-11-01

    Cell division is an essential dynamic event in tissue remodeling during wound healing, cancer and embryogenesis. In collective migration, tensile stresses affect cell shape and polarity, hence, the orientation of the cell division axis is expected to depend on cellular flow patterns. Here, we study the degree of orientation of cell division axes in migrating and resting epithelial cell sheets. We use microstructured channels to create a defined scenario of directed cell invasion and compare this situation to resting but proliferating cell monolayers. In experiments, we find a strong alignment of the axis due to directed flow while resting sheets show very weak global order, but local flow gradients still correlate strongly with the cell division axis. We compare experimental results with a previously published mesoscopic particle based simulation model. Most of the observed effects are reproduced by the simulations.

  13. Microfluidic approaches for epithelial cell layer culture and characterisation

    PubMed Central

    Thuenauer, Roland; Rodriguez-Boulan, Enrique; Römer, Winfried

    2014-01-01

    In higher eukaryotes, epithelial cell layers line most body cavities and form selective barriers that regulate the exchange of solutes between compartments. In order to fulfil these functions, the cells assume a polarised architecture and maintain two distinct plasma membrane domains, the apical domain facing the lumen and the basolateral domain facing other cells and the extracellular matrix. Microfluidic biochips offer the unique opportunity to establish novel in vitro models of epithelia in which the in vivo microenvironment of epithelial cells is precisely reconstituted. In addition, analytical tools to monitor biologically relevant parameters can be directly integrated on-chip. In this review we summarise recently developed biochip designs for culturing epithelial cell layers. Since endothelial cell layers, which line blood vessels, have similar barrier functions and polar organisation as epithelial cell layers, we also discuss biochips for culturing endothelial cell layers. Furthermore, we review approaches to integrate tools to analyse and manipulate epithelia and endothelia in microfluidic biochips, including methods to perform electrical impedance spectroscopy, methods to detect substances undergoing trans-epithelial transport via fluorescence, spectrophotometry, and mass spectrometry, techniques to mechanically stimulate cells via stretching and fluid flow-induced shear stress, and methods to carry out high-resolution imaging of vesicular trafficking with light microscopy. Taken together, this versatile microfluidic toolbox enables novel experimental approaches to characterise epithelial monolayers. PMID:24668405

  14. Microfluidic approaches for epithelial cell layer culture and characterisation.

    PubMed

    Thuenauer, Roland; Rodriguez-Boulan, Enrique; Römer, Winfried

    2014-07-01

    In higher eukaryotes, epithelial cell layers line most body cavities and form selective barriers that regulate the exchange of solutes between compartments. In order to fulfil these functions, the cells assume a polarised architecture and maintain two distinct plasma membrane domains, the apical domain facing the lumen and the basolateral domain facing other cells and the extracellular matrix. Microfluidic biochips offer the unique opportunity to establish novel in vitro models of epithelia in which the in vivo microenvironment of epithelial cells is precisely reconstituted. In addition, analytical tools to monitor biologically relevant parameters can be directly integrated on-chip. In this review we summarise recently developed biochip designs for culturing epithelial cell layers. Since endothelial cell layers, which line blood vessels, have similar barrier functions and polar organisation as epithelial cell layers, we also discuss biochips for culturing endothelial cell layers. Furthermore, we review approaches to integrate tools to analyse and manipulate epithelia and endothelia in microfluidic biochips; including methods to perform electrical impedance spectroscopy; methods to detect substances undergoing trans-epithelial transport via fluorescence, spectrophotometry, and mass spectrometry; techniques to mechanically stimulate cells via stretching and fluid flow-induced shear stress; and methods to carry out high-resolution imaging of vesicular trafficking using light microscopy. Taken together, this versatile microfluidic toolbox enables novel experimental approaches to characterise epithelial monolayers. PMID:24668405

  15. Molecular responses of rat tracheal epithelial cells to transmembrane pressure.

    PubMed

    Ressler, B; Lee, R T; Randell, S H; Drazen, J M; Kamm, R D

    2000-06-01

    Smooth muscle constriction in asthma causes the airway to buckle into a rosette pattern, folding the epithelium into deep crevasses. The epithelial cells in these folds are pushed up against each other and thereby experience compressive stresses. To study the epithelial cell response to compressive stress, we subjected primary cultures of rat tracheal epithelial cells to constant elevated pressures on their apical surface (i.e., a transmembrane pressure) and examined changes in the expression of genes that are important for extracellular matrix production and maintenance of smooth muscle activation. Northern blot analysis of RNA extracted from cells subjected to transmembrane pressure showed induction of early growth response-1 (Egr-1), endothelin-1, and transforming growth factor-beta1 in a pressure-dependent and time-dependent manner. Increases in Egr-1 protein were detected by immunohistochemistry. Our results demonstrate that airway epithelial cells respond rapidly to compressive stresses. Potential transduction mechanisms of transmembrane pressure were also investigated. PMID:10835333

  16. Alveolar Epithelial Cells Undergo Epithelial-to-Mesenchymal Transition in Response to Endoplasmic Reticulum Stress*

    PubMed Central

    Tanjore, Harikrishna; Cheng, Dong-Sheng; Degryse, Amber L.; Zoz, Donald F.; Abdolrasulnia, Rasul; Lawson, William E.; Blackwell, Timothy S.

    2011-01-01

    Expression of mutant surfactant protein C (SFTPC) results in endoplasmic reticulum (ER) stress in type II alveolar epithelial cells (AECs). AECs have been implicated as a source of lung fibroblasts via epithelial-to-mesenchymal transition (EMT); therefore, we investigated whether ER stress contributes to EMT as a possible mechanism for fibrotic remodeling. ER stress was induced by tunicamyin administration or stable expression of mutant (L188Q) SFTPC in type II AEC lines. Both tunicamycin treatment and mutant SFTPC expression induced ER stress and the unfolded protein response. With tunicamycin or mutant SFTPC expression, phase contrast imaging revealed a change to a fibroblast-like appearance. During ER stress, expression of epithelial markers E-cadherin and Zonula occludens-1 decreased while expression of mesenchymal markers S100A4 and α-smooth muscle actin increased. Following induction of ER stress, we found activation of a number of pathways, including MAPK, Smad, β-catenin, and Src kinase. Using specific inhibitors, the combination of a Smad2/3 inhibitor (SB431542) and a Src kinase inhibitor (PP2) blocked EMT with maintenance of epithelial appearance and epithelial marker expression. Similar results were noted with siRNA targeting Smad2 and Src kinase. Together, these studies reveal that induction of ER stress leads to EMT in lung epithelial cells, suggesting possible cross-talk between Smad and Src kinase pathways. Dissecting pathways involved in ER stress-induced EMT may lead to new treatment strategies to limit fibrosis. PMID:21757695

  17. Minor Salivary Gland Changes in Oral Epithelial Dysplasia and Oral Squamous Cell Carcinoma - A Histopathological Study

    PubMed Central

    Chitturi, Ravi Teja; Ragunathan, Yoithapprabhunath Thukanayakanpalayam; Lakshmi, Suman Jhansi; Nallusamy, Jaisanghar; Joseph, Isaac

    2016-01-01

    Introduction The most common etiology for Oral Squamous Cell Carcinoma (OSCC) is tobacco and tobacco related products which cause nuclear damage to the keratinocytes. The chemical carcinogens not only affect the lining of oral epithelium but also affect the lining epithelium of the excretory ducts of the salivary glands. Thus, there is a possibility of epithelial dysplasia of the salivary duct epithelium which may lead to potential malignant transformation. Aim The study was performed to see the changes in the minor salivary glands and excretory ducts in cases of oral epithelial dysplasia and OSCC. Materials and Methods A total of 278 archival cases of mild, moderate and severe epithelial dysplasia, carcinoma in situ, OSCC including verrucous carcinoma were histopathologically evaluated to observe changes in the excretory ducts and the minor salivary glands. Results In the study there were 56.5% males and 43.5% females. The age group that was most commonly affected in both the sexes was 50-60 yr old. Buccal mucosa was the most common site of involvement. Ductal changes observed in the excretory duct include simple hyperplasia, metaplastic changes such as mucous, oncocytic & squamous, and infiltration of inflammatory cells and malignant cells. Acinar changes observed were degeneration, squamous metaplasia, myoepithelial cell proliferation and inflammatory cell infiltration. Both the excretory ducts and ducts within the gland showed dysplasia. Conclusion According to observations in our study it is suggested that histopathological interpretation for oral mucosal lesions especially oral epithelial dysplasias and OSCC should also include changes related to salivary gland tissue to provide a better treatment plan and prevent recurrence of the malignant tumours.

  18. Flow Cytometry Analysis of Thymic Epithelial Cells and Their Subpopulations.

    PubMed

    Ohigashi, Izumi; Takahama, Yousuke

    2016-01-01

    The parenchyma of the thymus is compartmentalized into the cortex and the medulla, which are constructed by cortical thymic epithelial cells (cortical TECs, cTECs) and medullary thymic epithelial cells (mTECs), respectively. cTECs and mTECs essentially and differentially regulate the development and repertoire selection of T cells. Consequently, the biology of T cell development and selection includes the study of TECs in addition to the study of developing T cells and other hematopoietic cells including dendritic cells. In this chapter, we describe the methods for flow cytometric analysis and sorting of TECs and their subpopulations, including cTECs and mTECs. PMID:26294398

  19. Diversity of epithelial stem cell types in adult lung.

    PubMed

    Li, Feng; He, Jinxi; Wei, Jun; Cho, William C; Liu, Xiaoming

    2015-01-01

    Lung is a complex organ lined with epithelial cells. In order to maintain its homeostasis and normal functions following injuries caused by varied extraneous and intraneous insults, such as inhaled environmental pollutants and overwhelming inflammatory responses, the respiratory epithelium normally undergoes regenerations by the proliferation and differentiation of region-specific epithelial stem/progenitor cells that resided in distinct niches along the airway tree. The importance of local epithelial stem cell niches in the specification of lung stem/progenitor cells has been recently identified. Studies using cell differentiating and lineage tracing assays, in vitro and/or ex vivo models, and genetically engineered mice have suggested that these local epithelial stem/progenitor cells within spatially distinct regions along the pulmonary tree contribute to the injury repair of epithelium adjacent to their respective niches. This paper reviews recent findings in the identification and isolation of region-specific epithelial stem/progenitor cells and local niches along the airway tree and the potential link of epithelial stem cells for the development of lung cancer. PMID:25810726

  20. Diversity of Epithelial Stem Cell Types in Adult Lung

    PubMed Central

    Li, Feng; He, Jinxi; Wei, Jun; Cho, William C.; Liu, Xiaoming

    2015-01-01

    Lung is a complex organ lined with epithelial cells. In order to maintain its homeostasis and normal functions following injuries caused by varied extraneous and intraneous insults, such as inhaled environmental pollutants and overwhelming inflammatory responses, the respiratory epithelium normally undergoes regenerations by the proliferation and differentiation of region-specific epithelial stem/progenitor cells that resided in distinct niches along the airway tree. The importance of local epithelial stem cell niches in the specification of lung stem/progenitor cells has been recently identified. Studies using cell differentiating and lineage tracing assays, in vitro and/or ex vivo models, and genetically engineered mice have suggested that these local epithelial stem/progenitor cells within spatially distinct regions along the pulmonary tree contribute to the injury repair of epithelium adjacent to their respective niches. This paper reviews recent findings in the identification and isolation of region-specific epithelial stem/progenitor cells and local niches along the airway tree and the potential link of epithelial stem cells for the development of lung cancer. PMID:25810726

  1. Characterization and regulation of adenosine transport in T84 intestinal epithelial cells.

    PubMed

    Mun, E C; Tally, K J; Matthews, J B

    1998-02-01

    Adenosine release from mucosal sources during inflammation and ischemia activates intestinal epithelial Cl- secretion. Previous data suggest that A2b receptor-mediated Cl- secretory responses may be dampened by epithelial cell nucleoside scavenging. The present study utilizes isotopic flux analysis and nucleoside analog binding assays to directly characterize the nucleoside transport system of cultured T84 human intestinal epithelial cells and to explore whether adenosine transport is regulated by secretory agonists, metabolic inhibition, or phorbol ester. Uptake of adenosine across the apical membrane displayed characteristics of simple diffusion. Kinetic analysis of basolateral uptake revealed a Na(+)-independent, nitrobenzylthioinosine (NBTI)-sensitive facilitated-diffusion system with low affinity but high capacity for adenosine. NBTI binding studies indicated a single population of high-affinity binding sites basolaterally. Neither forskolin, 5'-(N-ethylcarboxamido)-adenosine, nor metabolic inhibition significantly altered adenosine transport. However, phorbol 12-myristate 13-acetate significantly reduced both adenosine transport and the number of specific NBTI binding sites, suggesting that transporter number may be decreased through activation of protein kinase C. This basolateral facilitated adenosine transporter may serve a conventional function in nucleoside salvage and a novel function as a regulator of adenosine-dependent Cl- secretory responses and hence diarrheal disorders. PMID:9486178

  2. HIV is inactivated after transepithelial migration via adult oral epithelial cells but not fetal epithelial cells

    PubMed Central

    Tugizov, Sharof M.; Herrera, Rossana; Veluppillai, Piri; Greenspan, Deborah; Soros, Vanessa; Greene, Warner C.; Levy, Jay A.; Palefsky, Joel M.

    2010-01-01

    Oral transmission of human immunodeficiency virus (HIV) in adult populations is rare. However, HIV spread across fetal/neonatal oropharyngeal epithelia could be important in mother-to-child transmission. Analysis of HIV transmission across polarized adult and fetal oral epithelial cells revealed that HIV transmigrates through both adult and fetal cells. However, only virions that passed through the fetal cells – and not those that passed through the adult cells – remained infectious. Analysis of expression of anti-HIV innate proteins beta-defensins 2 and 3, and secretory leukocyte protease inhibitor in adult, fetal, and infant oral epithelia showed that their expression is predominantly in the adult oral epithelium. Retention of HIV infectivity after transmigration correlated inversely with the expression of these innate proteins. Inactivation of innate proteins in adult oral keratinocytes restored HIV infectivity. These data suggest that high-level innate protein expression may contribute to the resistance of the adult oral epithelium to HIV transmission. PMID:21056450

  3. Increased sulfation of glycoconjugates by cultured nasal epithelial cells from patients with cystic fibrosis.

    PubMed Central

    Cheng, P W; Boat, T F; Cranfill, K; Yankaskas, J R; Boucher, R C

    1989-01-01

    Cystic fibrosis (CF) respiratory epithelia exhibit abnormal anion transport that may be linked to abnormal lung defense. In these studies, we investigated whether primary cultures of CF respiratory epithelial cells regulate abnormally the sulfate content of high molecular weight glycoconjugates (HMG) participating in airways' mucosal defense. HMG, including glycosaminoglycans and mucin-type glycoproteins released spontaneously into medium and HMG released from cell surfaces by trypsin, were metabolically labeled with 35SO4- and [6-3H]-glucosamine (GlcN) or 35SO4- and [3H]serine. All three classes of HMG from CF cells exhibited 35S/3H labeling ratios 1.5-4-fold greater than HMG from normal or disease control cells. Differences for labeling ratios of HMG from CF cells were shown to be the consequence of increased 35SO4- incorporation rather than decreased peptide synthesis and release or HMG glycosylation. The buoyant density of CF mucin-type HMG also was increased, consistent with increased sulfation. These observations suggest that oversulfation of a spectrum of HMG is a genetically determined characteristic of CF epithelial cells and may play an important pathophysiological role by altering the properties of mucous secretions and/or the interactions between selected bacteria and HMG at the airways' surface. Images PMID:2738159

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

    SciTech Connect

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

    2009-07-01

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

  5. Role of urokinase and its receptor in basal and stimulated colonic epithelial cell migration in vitro

    PubMed Central

    Wilson, A; Gibson, P

    2000-01-01

    BACKGROUND—Migration of colonic epithelial cells is important for mucosal repair following injury. The urokinase (u-PA) system regulates migration in other cell types.
AIM—To examine the role of u-PA and its receptor (u-PAR) in colonic epithelial cell migration.
METHODS—Migration was assessed over 24 hours in circular wounds made in confluent monolayers of LIM1215 and Caco-2 human colon cancer cells. The function of u-PA and u-PAR was ablated with antisense oligonucleotides to block expression, with synthetic u-PA peptides to block interaction, and with aprotinin to block u-PA mediated proteolysis.
RESULTS—Migration was stimulated two to threefold by exogenous u-PA, an effect dependent on u-PAR binding but independent of u-PA mediated mitogenesis and proteolysis. Expression of u-PA and u-PAR was inhibited by 80% by the appropriate antisense oligonucleotide. Basal migration and the motogenic effects of butyrate, epidermal growth factor, and phorbol-12-myristate-13-acetate were suppressed by the u-PAR antisense oligonucleotide (40-60%) but were at best minimally affected following inhibition of u-PA expression and binding. 
CONCLUSIONS—In an in vitro model of wounded colonic epithelium, u-PAR promotes cell migration through mechanisms that are not exclusively dependent on u-PA binding. Therefore, u-PA and u-PAR may contribute to colonic mucosal repair in vivo.


Keywords: colon; migration; urokinase; urokinase receptor; epidermal growth factor; butyrate; protein kinase C PMID:10861271

  6. Parvalbumin in cortical epithelial cells of the pigeon thymus

    PubMed Central

    ATOJI, YASURO; YAMAMOTO, YOSHIO; SUZUKI, YOSHITAKA

    2000-01-01

    We examined the distribution of parvalbumin in the pigeon thymus by light and electron microscopic immunohistochemistry. Tissues were also examined by conventional electron microscopy to determine the ultrastructure of immunoreactive cells. Parvalbumin immunoreaction was located in epithelial cells of the cortex, which formed dense mesh-like structures. Parvalbumin-positive epithelial cells were classified into 2 types. The first comprised elongated cells. In these, the nucleus was spindle-shaped, oval, or triangular, with a slightly irregular contour and contained rich heterochromatin peripherally. The cytoplasm was pale and processes extended laterally or ramified among the surrounding thymocytes. This type of cell formed the majority of immunoreactive cells. The other cell type consisted of polygonal epithelial cells. The nucleus was oval with deep indentations. Euchromatin occupied a large part of the nucleus. The cytoplasm contained numerous cell organelles compared with the elongated type, in particular, electron-dense vacuoles of various sizes and often bundles of tonofilaments. Both types of epithelial cell were interconnected by desmosomes. No secretory granules were found in the cytoplasm of elongated or polygonal cells. These results indicate the presence of heterogeneous group of parvalbumin-immunoreactive epithelial cells and suggest the likelihood of different functional roles for parvalbumin in the pigeon thymus. PMID:10853953

  7. Stochastic Terminal Dynamics in Epithelial Cell Intercalation

    NASA Astrophysics Data System (ADS)

    Eule, Stephan; Metzger, Jakob; Reichl, Lars; Kong, Deqing; Zhang, Yujun; Grosshans, Joerg; Wolf, Fred

    2015-03-01

    We found that the constriction of epithelial cell contacts during intercalation in germ band extension in Drosophila embryos follows intriguingly simple quantitative laws. The mean contact length < L > follows < L > (t) ~(T - t) α , where T is the finite collapse time; the time dependent variance of contact length is proportional to the square of the mean; finally the time dependent probability density of the contact lengths remains close to Gaussian during the entire process. These observations suggest that the dynamics of contact collapse can be captured by a stochastic differential equation analytically tractable in small noise approximation. Here, we present such a model, providing an effective description of the non-equilibrium statistical mechanics of contact collapse. All model parameters are fixed by measurements of time dependent mean and variance of contact lengths. The model predicts the contact length covariance function that we obtain in closed form. The contact length covariance function closely matches experimental observations suggesting that the model well captures the dynamics of contact collapse.

  8. VACCINES. A mucosal vaccine against Chlamydia trachomatis generates two waves of protective memory T cells.

    PubMed

    Stary, Georg; Olive, Andrew; Radovic-Moreno, Aleksandar F; Gondek, David; Alvarez, David; Basto, Pamela A; Perro, Mario; Vrbanac, Vladimir D; Tager, Andrew M; Shi, Jinjun; Yethon, Jeremy A; Farokhzad, Omid C; Langer, Robert; Starnbach, Michael N; von Andrian, Ulrich H

    2015-06-19

    Genital Chlamydia trachomatis (Ct) infection induces protective immunity that depends on interferon-γ-producing CD4 T cells. By contrast, we report that mucosal exposure to ultraviolet light (UV)-inactivated Ct (UV-Ct) generated regulatory T cells that exacerbated subsequent Ct infection. We show that mucosal immunization with UV-Ct complexed with charge-switching synthetic adjuvant particles (cSAPs) elicited long-lived protection in conventional and humanized mice. UV-Ct-cSAP targeted immunogenic uterine CD11b(+)CD103(-) dendritic cells (DCs), whereas UV-Ct accumulated in tolerogenic CD11b(-)CD103(+) DCs. Regardless of vaccination route, UV-Ct-cSAP induced systemic memory T cells, but only mucosal vaccination induced effector T cells that rapidly seeded uterine mucosa with resident memory T cells (T(RM) cells). Optimal Ct clearance required both T(RM) seeding and subsequent infection-induced recruitment of circulating memory T cells. Thus, UV-Ct-cSAP vaccination generated two synergistic memory T cell subsets with distinct migratory properties. PMID:26089520

  9. Characteristics and EGFP expression of porcine mammary gland epithelial cells.

    PubMed

    Zheng, Yue-Mao; He, Xiao-Ying

    2010-12-01

    The aims of this study were to establish a porcine mammary gland epithelial (PMGE) cell line, and to determine if these PMGE cells could be maintained long-term in culture by continuous subculturing following transfection with a reporter gene, enhanced green fluorescence protein (EGFP). Primary culture of PMGE cells was achieved by outgrowth of migrating cells from the fragments of the mammary gland tissue of a lactating pig. The passage sixteen PMGE cells were transfected with EGFP gene using lipofection. The expression of Cell keratins of epithelial cells in PMGE cells was tested by immunofluorescence. Βeta-Casein gene mRNA was tested for PMGE cells by RT-PCR. The results showed that PMGE cells could form dome-like structure which looked like nipple, and the cells contained different cell types. The expression of Cell keratins demonstrated the property of epithelial cells, and the PMGE cells could express transcript encoding a Βeta-Casein protein. EGFP gene was successfully transferred into the PMGE cells, and the transfected cells could be maintained long-term in culture by continuous subculturing. In conclusion, we have established a EGFP gene transfected porcine mammary gland epithelial (ET-PMGE) cell line. PMID:20400167

  10. Cell volume regulation in epithelial physiology and cancer

    PubMed Central

    Pedersen, Stine F.; Hoffmann, Else K.; 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 regulation both rely on the spatially and temporally coordinated function of ion channels and transporters. In healthy epithelia, specific ion channels/transporters localize to the luminal and basolateral membranes, contributing to functional epithelial polarity. In pathophysiological processes 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 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. PMID:24009588

  11. MFGE8 regulates TGF-β-induced epithelial mesenchymal transition in endometrial epithelial cells in vitro.

    PubMed

    Yu, Liang; Hu, Rong; Sullivan, Claretta; Swanson, R James; Oehninger, Sergio; Sun, Ying-Pu; Bocca, Silvina

    2016-09-01

    This study investigated the role of milk fat globule-epidermal growth factor-factor 8 (MFGE8) in TGF-β-induced epithelial-mesenchymal transition (EMT) of endometrial epithelial cells. These were in vitro studies using human endometrial epithelial cells and mouse blastocysts. We investigated the ability of TGF-β to induce EMT in endometrial epithelial cells (HEC-1A) by assessment of cytological phenotype (by light and atomic force microscopy), changes in expression of the markers of cell adhesion/differentiation E- and N-cadherin, and of the transcription factor Snail (by immunofluorescence and immunoblotting), and competence to support embryo attachment in a mouse blastocyst outgrowth assay. We also studied the effects of E-cadherin expression in cells transfected by retroviral shRNA vectors specifically silencing MFGE8. Results demonstrated that TGF-β induced EMT as demonstrated by phenotypic cell changes, by a switch of cadherin expression as well as by upregulation of the expression of the mesenchymal markers Snail and Vimentin. Upon MFGE8 knockdown, these processes were interfered with, suggesting that MFGE8 and TGF-β together may participate in regulation of EMT. This study demonstrated for the first time that endometrial MFGE8 modulates TGF-β-induced EMT in human endometrium cells. PMID:27340235

  12. Morphological appearances of human lens epithelial cells in culture.

    PubMed

    Power, W; Neylan, D; Collum, L

    1993-01-01

    A system for culturing human lens epithelial cells in the laboratory was developed. The morphological appearances of the cells was studied using phase contrast, scanning and transmission electron microscopy. Cell marker studies using monoclonal antibodies to cytokeratin, vimentin and epithelial membrane antigen were also performed. There was a marked increase in cell size as a function of time in culture. After 3 to 4 weeks cells showed early signs of ageing. By 6 to 8 weeks the majority of the cells had become very irregular in shape and demonstrated irregularities of the plasma membrane and intra-cytoplasmic vacuole formation. The cells stained strongly for vimentin and epithelial membrane antigen. Staining with cytokeratin was somewhat weaker. This culture technique provides us with a suitable model for studying the growth behavior of these cells. PMID:7512459

  13. Regulated Mucin Secretion from Airway Epithelial Cells

    PubMed Central

    Adler, Kenneth B.; Tuvim, Michael J.; Dickey, Burton F.

    2013-01-01

    Secretory epithelial cells of the proximal airways synthesize and secrete gel-forming polymeric mucins. The secreted mucins adsorb water to form mucus that is propelled by neighboring ciliated cells, providing a mobile barrier which removes inhaled particles and pathogens from the lungs. Several features of the intracellular trafficking of mucins make the airway secretory cell an interesting comparator for the cell biology of regulated exocytosis. Polymeric mucins are exceedingly large molecules (up to 3 × 106 Da per monomer) whose folding and initial polymerization in the ER requires the protein disulfide isomerase Agr2. In the Golgi, mucins further polymerize to form chains and possibly branched networks comprising more than 20 monomers. The large size of mucin polymers imposes constraints on their packaging into transport vesicles along the secretory pathway. Sugar side chains account for >70% of the mass of mucins, and their attachment to the protein core by O-glycosylation occurs in the Golgi. Mature polymeric mucins are stored in large secretory granules ∼1 μm in diameter. These are translocated to the apical membrane to be positioned for exocytosis by cooperative interactions among myristoylated alanine-rich C kinase substrate, cysteine string protein, heat shock protein 70, and the cytoskeleton. Mucin granules undergo exocytic fusion with the plasma membrane at a low basal rate and a high stimulated rate. Both rates are mediated by a regulated exocytic mechanism as indicated by phenotypes in both basal and stimulated secretion in mice lacking Munc13-2, a sensor of the second messengers calcium and diacylglycerol (DAG). Basal secretion is induced by low levels of activation of P2Y2 purinergic and A3 adenosine receptors by extracellular ATP released in paracrine fashion and its metabolite adenosine. Stimulated secretion is induced by high levels of the same ligands, and possibly by inflammatory mediators as well. Activated receptors are coupled to

  14. Effect of freezing on lens epithelial cell growth.

    PubMed

    Fukaya, Y; Hara, T; Hara, T; Iwata, S

    1988-05-01

    The effect of freezing on the growth of rat lens epithelial cells was studied in vitro. We found that 80% of the lens epithelial cells died after freezing at -45 degrees C for two hours and that the surviving cells could grow with the addition of growth factors or when placed on a sheet of type 4 collagen, but not when placed on a plain plastic culture dish. These results suggest that the surviving cells are at the Go phase of the cell cycle and that type 4 collagen or growth factors can initiate cell division. PMID:3294380

  15. Adipose-Derived Mesenchymal Stem Cells Restore Impaired Mucosal Immune Responses in Aged Mice

    PubMed Central

    Aso, Kazuyoshi; Tsuruhara, Akitoshi; Takagaki, Kentaro; Oki, Katsuyuki; Ota, Megumi; Nose, Yasuhiro; Tanemura, Hideki; Urushihata, Naoki; Sasanuma, Jinichi; Sano, Masayuki; Hirano, Atsuyuki; Aso, Rio; McGhee, Jerry R.; Fujihashi, Kohtaro

    2016-01-01

    It has been shown that adipose-derived mesenchymal stem cells (AMSCs) can differentiate into adipocytes, chondrocytes and osteoblasts. Several clinical trials have shown the ability of AMSCs to regenerate these differentiated cell types. Age-associated dysregulation of the gastrointestinal (GI) immune system has been well documented. Our previous studies showed that impaired mucosal immunity in the GI tract occurs earlier during agingthan is seen in the systemic compartment. In this study, we examined the potential of AMSCs to restore the GI mucosal immune system in aged mice. Aged (>18 mo old) mice were adoptively transferred with AMSCs. Two weeks later, mice were orally immunized with ovalbumin (OVA) plus cholera toxin (CT) three times at weekly intervals. Seven days after the final immunization, when fecal extract samples and plasma were subjected to OVA- and CT-B-specific ELISA, elevated levels of mucosal secretory IgA (SIgA) and plasma IgG antibody (Ab) responses were noted in aged mouse recipients. Similar results were also seen aged mice which received AMSCs at one year of age. When cytokine production was examined, OVA-stimulated Peyer’s patch CD4+ T cells produced increased levels of IL-4. Further, CD4+ T cells from the lamina propria revealed elevated levels of IL-4 and IFN-γ production. In contrast, aged mice without AMSC transfer showed essentially no OVA- or CT-B-specific mucosal SIgA or plasma IgG Ab or cytokine responses. Of importance, fecal extracts from AMSC transferred aged mice showed neutralization activity to CT intoxication. These results suggest that AMSCs can restore impaired mucosal immunity in the GI tract of aged mice. PMID:26840058

  16. Adipose-Derived Mesenchymal Stem Cells Restore Impaired Mucosal Immune Responses in Aged Mice.

    PubMed

    Aso, Kazuyoshi; Tsuruhara, Akitoshi; Takagaki, Kentaro; Oki, Katsuyuki; Ota, Megumi; Nose, Yasuhiro; Tanemura, Hideki; Urushihata, Naoki; Sasanuma, Jinichi; Sano, Masayuki; Hirano, Atsuyuki; Aso, Rio; McGhee, Jerry R; Fujihashi, Kohtaro

    2016-01-01

    It has been shown that adipose-derived mesenchymal stem cells (AMSCs) can differentiate into adipocytes, chondrocytes and osteoblasts. Several clinical trials have shown the ability of AMSCs to regenerate these differentiated cell types. Age-associated dysregulation of the gastrointestinal (GI) immune system has been well documented. Our previous studies showed that impaired mucosal immunity in the GI tract occurs earlier during agingthan is seen in the systemic compartment. In this study, we examined the potential of AMSCs to restore the GI mucosal immune system in aged mice. Aged (>18 mo old) mice were adoptively transferred with AMSCs. Two weeks later, mice were orally immunized with ovalbumin (OVA) plus cholera toxin (CT) three times at weekly intervals. Seven days after the final immunization, when fecal extract samples and plasma were subjected to OVA- and CT-B-specific ELISA, elevated levels of mucosal secretory IgA (SIgA) and plasma IgG antibody (Ab) responses were noted in aged mouse recipients. Similar results were also seen aged mice which received AMSCs at one year of age. When cytokine production was examined, OVA-stimulated Peyer's patch CD4+ T cells produced increased levels of IL-4. Further, CD4+ T cells from the lamina propria revealed elevated levels of IL-4 and IFN-γ production. In contrast, aged mice without AMSC transfer showed essentially no OVA- or CT-B-specific mucosal SIgA or plasma IgG Ab or cytokine responses. Of importance, fecal extracts from AMSC transferred aged mice showed neutralization activity to CT intoxication. These results suggest that AMSCs can restore impaired mucosal immunity in the GI tract of aged mice. PMID:26840058

  17. ONCOGENE ALTERNATIONS IN IN VITRO TRANSFORMED RAT TRACHEAL EPITHELIAL CELLS

    EPA Science Inventory

    Ten derivations of rat tracheal epithelial (RTE) cells, including normal cells, normal primary cultures, 7 tumorigenic cell lines and 1 non-tumorigenic cell line transformed by treatment with 7,12-dimethylbenz(a)anthracene (DMBA), benzo(a)pyrene (BP) and/or 12-0-tetradecanoylphor...

  18. Left-right asymmetric cell intercalation drives directional collective cell movement in epithelial morphogenesis

    NASA Astrophysics Data System (ADS)

    Sato, Katsuhiko; Hiraiwa, Tetsuya; Maekawa, Emi; Isomura, Ayako; Shibata, Tatsuo; Kuranaga, Erina

    2015-12-01

    Morphogenetic epithelial movement occurs during embryogenesis and drives complex tissue formation. However, how epithelial cells coordinate their unidirectional movement while maintaining epithelial integrity is unclear. Here we propose a novel mechanism for collective epithelial cell movement based on Drosophila genitalia rotation, in which epithelial tissue rotates clockwise around the genitalia. We found that this cell movement occurs autonomously and requires myosin II. The moving cells exhibit repeated left-right-biased junction remodelling, while maintaining adhesion with their neighbours, in association with a polarized myosin II distribution. Reducing myosinID, known to cause counter-clockwise epithelial-tissue movement, reverses the myosin II distribution. Numerical simulations revealed that a left-right asymmetry in cell intercalation is sufficient to induce unidirectional cellular movement. The cellular movement direction is also associated with planar cell-shape chirality. These findings support a model in which left-right asymmetric cell intercalation within an epithelial sheet drives collective cellular movement in the same direction.

  19. Modulation of Intestinal Epithelial Cell Proliferation, Migration, and Differentiation In Vitro by Astragalus Polysaccharides

    PubMed Central

    Zhang, Chun Li; Ren, Hui Jun; Liu, Meng Meng; Li, Xiao Gai; Sun, De Li; Li, Nan; Ming, Liang

    2014-01-01

    Previous studies have shown that Astragalus polysaccharides (APS) can be used to treat general gastrointestinal disturbances including intestinal mucosal injury. However, the mechanism by which APS mediate this effect is unclear. In the present study, the effects of APS on proliferation, migration, and differentiation of intestinal epithelial cells (IEC-6) were assessed using an in vitro wounding model and colorimetric thiazolyl blue (MTT) assays. The effect of APS on IEC-6 cell differentiation was observed using a light microscope and scanning electron microscope, and the expression of differentiation-specific markers of IEC-6 cells, such as cytokeratin 18 (CK18), alkaline phosphatase (ALP), tight junction protein ZO-2, and sucrase-isomaltase (SI), was determined by immunofluorescence assay (IFA) and real-time PCR. In addition, APS-induced signaling pathways in IEC-6 cells were characterized. Our results indicated that APS significantly enhance migration and proliferation of IEC-6 cells in vitro. APS-treated IEC-6 cells have numerous microvilli on their apical surface and also highly express CK18, ALP, ZO-2, and SI. Moreover, APS-treated IEC-6 cells, in which the activity and expression level of ornithine decarboxylase (ODC) were significantly elevated, also exhibited an increase in cellular putrescine, whereas no significant increase in TGF-β levels was observed. These findings suggest that APS may enhance intestinal epithelial cell proliferation, migration, and differentiation in vitro by stimulating ODC gene expression and activity and putrescine production, independent of TGF-β. Exogenous administration of APS may provide a new approach for modulating intestinal epithelial wound restitution in vivo. PMID:25157577

  20. Hypo-responsiveness of interleukin-8 production in human embryonic epithelial intestine 407 cells independent of NF-{kappa}B pathway: New lessons from endotoxin and ribotoxic deoxynivalenol

    SciTech Connect

    Moon, Yuseok Yang, Hyun; Park, Seung-Hwan

    2008-08-15

    Mucosal epithelium senses external toxic insults and transmits the danger signals into the epithelial cells in order to activate a broad range of inflammatory responses. However, pre-exposure to the commensal endotoxins can induce inflammatory tolerance and maintain the homeostasis without excessive immune responses. We recently reported that ribotoxin deoxynivalenol (DON) and its derivatives elicited the pro-inflammatory response as the mucosal insults in human epithelial cells. Taking the knowledge into consideration, we tested the hypothesis that endotoxin pre-exposure can attenuate ribotoxin-induced epithelial interleukin-8 (IL-8) production via a tolerance mechanism. Pre-exposure to endotoxin repressed IL-8 release and its gene expression. However, inflammatory tolerance was not mediated by the attenuated NF-{kappa}B activation which has been generally recognized as the major mediator of LPS-mediated toll-like receptor (TLR) signaling pathway. Instead, pre-exposure to endotoxin was observed to trigger the delayed induction of peroxisome proliferator-activated receptor gamma (PPAR-{gamma}) which contributed to the diminished IL-8 production in the human epithelial cells. Moreover, endogenous PPAR-{gamma} agonist suppressed toxicant-mediated interleukin-8 production and IL-8 mRNA stability. Taken together, endotoxin induced hypo-production of pro-inflammatory cytokine IL-8 in the human epithelial cells, which was associated with the delayed activation of PPAR-{gamma} expression by pre-existing endotoxin.

  1. Control of Francisella tularensis Intracellular Growth by Pulmonary Epithelial Cells

    PubMed Central

    Maggio, Savannah; Takeda, Kazuyo; Stark, Felicity; Meierovics, Anda I.; Yabe, Idalia; Cowley, Siobhan C.

    2015-01-01

    The virulence of F. tularensis is often associated with its ability to grow in macrophages, although recent studies show that Francisella proliferates in multiple host cell types, including pulmonary epithelial cells. Thus far little is known about the requirements for killing of F. tularensis in the non-macrophage host cell types that support replication of this organism. Here we sought to address this question through the use of a murine lung epithelial cell line (TC-1 cells). Our data show that combinations of the cytokines IFN-γ, TNF, and IL-17A activated murine pulmonary epithelial cells to inhibit the intracellular growth of the F. tularensis Live Vaccine Strain (LVS) and the highly virulent F. tularensis Schu S4 strain. Although paired combinations of IFN-γ, TNF, and IL-17A all significantly controlled LVS growth, simultaneous treatment with all three cytokines had the greatest effect on LVS growth inhibition. In contrast, Schu S4 was more resistant to cytokine-induced growth effects, exhibiting significant growth inhibition only in response to all three cytokines. Since one of the main antimicrobial mechanisms of activated macrophages is the release of reactive nitrogen intermediates (RNI) via the activity of iNOS, we investigated the role of RNI and iNOS in Francisella growth control by pulmonary epithelial cells. NOS2 gene expression was significantly up-regulated in infected, cytokine-treated pulmonary epithelial cells in a manner that correlated with LVS and Schu S4 growth control. Treatment of LVS-infected cells with an iNOS inhibitor significantly reversed LVS killing in cytokine-treated cultures. Further, we found that mouse pulmonary epithelial cells produced iNOS during in vivo respiratory LVS infection. Overall, these data demonstrate that lung epithelial cells produce iNOS both in vitro and in vivo, and can inhibit Francisella intracellular growth via reactive nitrogen intermediates. PMID:26379269

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

    SciTech Connect

    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.

  3. Interferons Mediate Terminal Differentiation of Human Cortical Thymic Epithelial Cells

    PubMed Central

    Vidalain, Pierre-Olivier; Laine, David; Zaffran, Yona; Azocar, Olga; Servet-Delprat, Christine; Wild, T. Fabian; Rabourdin-Combe, Chantal; Valentin, Hélène

    2002-01-01

    In the thymus, epithelial cells comprise a heterogeneous population required for the generation of functional T lymphocytes, suggesting that thymic epithelium disruption by viruses may compromise T-cell lymphopoiesis in this organ. In a previous report, we demonstrated that in vitro, measles virus induced differentiation of cortical thymic epithelial cells as characterized by (i) cell growth arrest, (ii) morphological and phenotypic changes, and (iii) apoptotis as a final step of this process. In the present report, we have analyzed the mechanisms involved. First, measles virus-induced differentiation of thymic epithelial cells is shown to be strictly dependent on beta interferon (IFN-β) secretion. In addition, transfection with double-stranded RNA, a common intermediate of replication for a broad spectrum of viruses, is reported to similarly mediate thymic epithelial cell differentiation through IFN-β induction. Finally, we demonstrated that recombinant IFN-α, IFN-β, or IFN-γ was sufficient to induce differentiation and apoptosis of uninfected thymic epithelial cells. These observations suggested that interferon secretion by either infected cells or activated leukocytes, such as plasmacytoid dendritic cells or lymphocytes, may induce thymic epithelium disruption in a pathological context. Thus, we have identified a new mechanism that may contribute to thymic atrophy and altered T-cell lymphopoiesis associated with many infections. PMID:12050353

  4. Interleukin-8 stimulates the migration of human colonic epithelial cells in vitro.

    PubMed

    Wilson, A J; Byron, K; Gibson, P R

    1999-09-01

    The migration of colonic epithelial cells (restitution) is an important event in the repair of mucosal injuries. Interleukin-8 (IL-8) is a physiological initiator of the chemotactic migration of leucocytes. This study aimed to determine whether IL-8 had a similar effect on migration in an in vitro model of wounded colonic epithelium. Cell migration over 24 h was assessed in circular wounds made in confluent monolayers of the human colon cancer cell line LIM1215. This migration was stimulated in a concentration-dependent manner by IL-8, with maximal effects of approx. 1.75-fold above basal migration. The motogenic effect of IL-8 was mediated independently of effects on cell proliferation. In contrast, it was partially dependent upon gene transcription and protein synthesis and involved the activation of pertussis-toxin-sensitive G-proteins. The short-chain fatty acids, acetate, propionate, butyrate and valerate, the activator of protein kinase C (phorbol-12-myristate-13-acetate) and tumour necrosis factor-alpha (TNF-alpha) all stimulated the secretion of IL-8. However, only the motogenic effect of TNF-alpha was dependent upon IL-8. In conclusion, IL-8 stimulated cell migration in an in vitro model of colonic epithelium, whereas the motogenic effect of at least one physiologically relevant factor was dependent upon an increase in its endogenous levels. If IL-8 stimulates colonic epithelial restitution in vivo, this would have ramifications for the control of repair processes following wounding of the colonic mucosa. PMID:10464065

  5. Commensal Streptococcus salivarius Modulates PPARγ Transcriptional Activity in Human Intestinal Epithelial Cells

    PubMed Central

    Couvigny, Benoît; de Wouters, Tomas; Kaci, Ghalia; Jacouton, Elsa; Delorme, Christine; Doré, Joël; Renault, Pierre; Blottière, Hervé M.

    2015-01-01

    The impact of commensal bacteria in eukaryotic transcriptional regulation has increasingly been demonstrated over the last decades. A multitude of studies have shown direct effects of commensal bacteria from local transcriptional activity to systemic impact. The commensal bacterium Streptococcus salivarius is one of the early bacteria colonizing the oral and gut mucosal surfaces. It has been shown to down-regulate nuclear transcription factor (NF-кB) in human intestinal cells, a central regulator of the host mucosal immune system response to the microbiota. In order to evaluate its impact on a further important transcription factor shown to link metabolism and inflammation in the intestine, namely PPARγ (peroxisome proliferator-activated receptor), we used human intestinal epithelial cell-lines engineered to monitor PPARγ transcriptional activity in response to a wide range of S. salivarius strains. We demonstrated that different strains from this bacterial group share the property to inhibit PPARγ activation independently of the ligand used. First attempts to identify the nature of the active compounds showed that it is a low-molecular-weight, DNase-, proteases- and heat-resistant metabolite secreted by S. salivarius strains. Among PPARγ-targeted metabolic genes, I-FABP and Angptl4 expression levels were dramatically reduced in intestinal epithelial cells exposed to S. salivarius supernatant. Both gene products modulate lipid accumulation in cells and down-regulating their expression might consequently affect host health. Our study shows that species belonging to the salivarius group of streptococci impact both host inflammatory and metabolic regulation suggesting a possible role in the host homeostasis and health. PMID:25946041

  6. Commensal Streptococcus salivarius Modulates PPARγ Transcriptional Activity in Human Intestinal Epithelial Cells.

    PubMed

    Couvigny, Benoît; de Wouters, Tomas; Kaci, Ghalia; Jacouton, Elsa; Delorme, Christine; Doré, Joël; Renault, Pierre; Blottière, Hervé M; Guédon, Eric; Lapaque, Nicolas

    2015-01-01

    The impact of commensal bacteria in eukaryotic transcriptional regulation has increasingly been demonstrated over the last decades. A multitude of studies have shown direct effects of commensal bacteria from local transcriptional activity to systemic impact. The commensal bacterium Streptococcus salivarius is one of the early bacteria colonizing the oral and gut mucosal surfaces. It has been shown to down-regulate nuclear transcription factor (NF-кB) in human intestinal cells, a central regulator of the host mucosal immune system response to the microbiota. In order to evaluate its impact on a further important transcription factor shown to link metabolism and inflammation in the intestine, namely PPARγ (peroxisome proliferator-activated receptor), we used human intestinal epithelial cell-lines engineered to monitor PPARγ transcriptional activity in response to a wide range of S. salivarius strains. We demonstrated that different strains from this bacterial group share the property to inhibit PPARγ activation independently of the ligand used. First attempts to identify the nature of the active compounds showed that it is a low-molecular-weight, DNase-, proteases- and heat-resistant metabolite secreted by S. salivarius strains. Among PPARγ-targeted metabolic genes, I-FABP and Angptl4 expression levels were dramatically reduced in intestinal epithelial cells exposed to S. salivarius supernatant. Both gene products modulate lipid accumulation in cells and down-regulating their expression might consequently affect host health. Our study shows that species belonging to the salivarius group of streptococci impact both host inflammatory and metabolic regulation suggesting a possible role in the host homeostasis and health. PMID:25946041

  7. Inhibition of corneal epithelial cell migration by cadmium and mercury

    SciTech Connect

    Ubels, J.L.; Osgood, T.B. Medical Coll. of Wisconsin, Milwaukee )

    1991-02-01

    In a previous comparative study of corneal healing in fish, the authors observed that corneal epithelial healing occurs very rapidly in vivo in the marine teleost Myoxocephalus octodecimspinosus (longhorn sculpin) with a 6-mm diameter wound on the mammalian cornea. This rapid healing which permits prompt restoration of the epithelial barrier is apparently an adaptation to the large ionic and osmotic gradients between the environment and the intraocular fluids of the fish. These observations suggested that epithelial healing in the sculpin cornea might be useful model in aquatic biomedical toxicology if an in vitro method for measurement of healing rates could be developed. In this report the authors demonstrate that sculpin eyes maintained in short-term organ culture have a rapid corneal epithelial healing response and that this model can be used to demonstrate the toxic effects of heavy metals on epithelial cell migration.

  8. Inhibition of T-cell Response by Escherichia coli Heat-Labile Enterotoxin-Treated Epithelial Cells

    PubMed Central

    Lopes, Luciene M.; Maroof, Asher; Dougan, Gordon; Chain, Benjamin M.

    2000-01-01

    Escherichia coli heat-labile enterotoxin (LT) is an extensively studied adjuvant of mucosal responses. Nevertheless, its mode of action as an adjuvant remains incompletely understood. In this study, we describe a simplified in vitro model with which to look at some aspects of immunoregulation by LT. The interaction of LT with the apical surface of a monolayer of CaCo-2 epithelial cells induces the release of a soluble factor which inhibits the antigen-induced release of interleukin-2 by T cells cultured at the basolateral side of the cells. The release of this factor requires the ADP-ribosylating activity of LT since the isolated B subunit, as well as an enzymatically silent LT mutant, loses biological activity in this model. The inhibitory activity is likely to be due to prostaglandin release, since it is blocked by indomethacin. The contribution of LT-induced prostaglandin release to the complex immunoregulatory activity of LT is discussed. PMID:11083810

  9. Protective role of intracellular glutathione against ethanol-induced damage in cultured rat gastric mucosal cells

    SciTech Connect

    Mutoh, H.; Hiraishi, H.; Ota, S.; Yoshida, H.; Ivey, K.J.; Terano, A.; Sugimoto, T. )

    1990-06-01

    This study investigated whether intracellular glutathione is cytoprotective against ethanol-induced injury to cultured rat gastric mucosal cells in vitro. Secondly, it investigated whether reduced glutathione or oxidized glutathione is responsible for this cytoprotection. Cytolysis was quantified by measuring 51Cr release from prelabeled cells. Concentrations of ethanol greater than 12% caused cell damage and increased 51Cr release in a dose-dependent and time-related fashion. When a substrate for glutathione synthesis, N-acetyl-L-cysteine, was provided to cultured cells for 4 h before challenge with ethanol, cytolysis was significantly decreased corresponding with an increase in cellular glutathione content. Pretreatment with diethyl maleate, which depletes reduced glutathione without forming oxidized glutathione, potentiated ethanol-induced cell damage in a dose-dependent manner with the decrease of cellular glutathione content. The administration of tert-butyl hydroperoxide (which is specifically reduced by glutathione peroxidase to generate oxidized glutathione from reduced glutathione) or diamide (which nonenzymatically oxidizes reduced glutathione to oxidized glutathione) enhanced ethanol injury. We conclude that in cultured gastric mucosal cells, (a) intracellular glutathione maintains integrity of gastric mucosal cells against ethanol in vitro; and (b) reduced glutathione rather than oxidized glutathione is responsible for this cytoprotection. We postulate that the presence of reduced glutathione is essential to allow glutathione peroxidase to catalyze the ethanol-generated toxic oxygen radical, hydrogen peroxide.

  10. An NK Cell Perforin Response Elicited via IL-18 Controls Mucosal Inflammation Kinetics during Salmonella Gut Infection.

    PubMed

    Müller, Anna A; Dolowschiak, Tamas; Sellin, Mikael E; Felmy, Boas; Verbree, Carolin; Gadient, Sandra; Westermann, Alexander J; Vogel, Jörg; LeibundGut-Landmann, Salome; Hardt, Wolf-Dietrich

    2016-06-01

    Salmonella Typhimurium (S.Tm) is a common cause of self-limiting diarrhea. The mucosal inflammation is thought to arise from a standoff between the pathogen's virulence factors and the host's mucosal innate immune defenses, particularly the mucosal NAIP/NLRC4 inflammasome. However, it had remained unclear how this switches the gut from homeostasis to inflammation. This was studied using the streptomycin mouse model. S.Tm infections in knockout mice, cytokine inhibition and -injection experiments revealed that caspase-1 (not -11) dependent IL-18 is pivotal for inducing acute inflammation. IL-18 boosted NK cell chemoattractants and enhanced the NK cells' migratory capacity, thus promoting mucosal accumulation of mature, activated NK cells. NK cell depletion and Prf-/- ablation (but not granulocyte-depletion or T-cell deficiency) delayed tissue inflammation. Our data suggest an NK cell perforin response as one limiting factor in mounting gut mucosal inflammation. Thus, IL-18-elicited NK cell perforin responses seem to be critical for coordinating mucosal inflammation during early infection, when S.Tm strongly relies on virulence factors detectable by the inflammasome. This may have broad relevance for mucosal defense against microbial pathogens. PMID:27341123

  11. An NK Cell Perforin Response Elicited via IL-18 Controls Mucosal Inflammation Kinetics during Salmonella Gut Infection

    PubMed Central

    Müller, Anna A.; Dolowschiak, Tamas; Sellin, Mikael E.; Felmy, Boas; Verbree, Carolin; Gadient, Sandra; Westermann, Alexander J.; Vogel, Jörg; LeibundGut-Landmann, Salome; Hardt, Wolf-Dietrich

    2016-01-01

    Salmonella Typhimurium (S.Tm) is a common cause of self-limiting diarrhea. The mucosal inflammation is thought to arise from a standoff between the pathogen's virulence factors and the host's mucosal innate immune defenses, particularly the mucosal NAIP/NLRC4 inflammasome. However, it had remained unclear how this switches the gut from homeostasis to inflammation. This was studied using the streptomycin mouse model. S.Tm infections in knockout mice, cytokine inhibition and –injection experiments revealed that caspase-1 (not -11) dependent IL-18 is pivotal for inducing acute inflammation. IL-18 boosted NK cell chemoattractants and enhanced the NK cells' migratory capacity, thus promoting mucosal accumulation of mature, activated NK cells. NK cell depletion and Prf-/- ablation (but not granulocyte-depletion or T-cell deficiency) delayed tissue inflammation. Our data suggest an NK cell perforin response as one limiting factor in mounting gut mucosal inflammation. Thus, IL-18-elicited NK cell perforin responses seem to be critical for coordinating mucosal inflammation during early infection, when S.Tm strongly relies on virulence factors detectable by the inflammasome. This may have broad relevance for mucosal defense against microbial pathogens. PMID:27341123

  12. Mucosally transplanted mesenchymal stem cells stimulate intestinal healing by promoting angiogenesis

    PubMed Central

    Manieri, Nicholas A.; Mack, Madison R.; Himmelrich, Molly D.; Worthley, Daniel L.; Hanson, Elaine M.; Eckmann, Lars; Wang, Timothy C.; Stappenbeck, Thaddeus S.

    2015-01-01

    Mesenchymal stem cell (MSC) therapy is an emerging field of regenerative medicine; however, it is often unclear how these cells mediate repair. Here, we investigated the use of MSCs in the treatment of intestinal disease and modeled abnormal repair by creating focal wounds in the colonic mucosa of prostaglandin-deficient mice. These wounds developed into ulcers that infiltrated the outer intestinal wall. We determined that penetrating ulcer formation in this model resulted from increased hypoxia and smooth muscle wall necrosis. Prostaglandin I2 (PGI2) stimulated VEGF-dependent angiogenesis to prevent penetrating ulcers. Treatment of mucosally injured WT mice with a VEGFR inhibitor resulted in the development of penetrating ulcers, further demonstrating that VEGF is critical for mucosal repair. We next used this model to address the role of transplanted colonic MSCs (cMSCs) in intestinal repair. Compared with intravenously injected cMSCs, mucosally injected cMSCs more effectively prevented the development of penetrating ulcers, as they were more efficiently recruited to colonic wounds. Importantly, mucosally injected cMSCs stimulated angiogenesis in a VEGF-dependent manner. Together, our results reveal that penetrating ulcer formation results from a reduction of local angiogenesis and targeted injection of MSCs can optimize transplantation therapy. Moreover, local MSC injection has potential for treating diseases with features of abnormal angiogenesis and repair. PMID:26280574

  13. Role of autophagy in the regulation of epithelial cell junctions.

    PubMed

    Nighot, Prashant; Ma, Thomas

    2016-01-01

    Autophagy is a cell survival mechanism by which bulk cytoplasmic material, including soluble macromolecules and organelles, is targeted for lysosomal degradation. The role of autophagy in diverse cellular processes such as metabolic stress, neurodegeneration, cancer, aging, immunity, and inflammatory diseases is being increasingly recognized. Epithelial cell junctions play an integral role in the cell homeostasis via physical binding, regulating paracellular pathways, integrating extracellular cues into intracellular signaling, and cell-cell communication. Recent data indicates that cell junction composition is very dynamic. The junctional protein complexes are actively regulated in response to various intra- and extra-cellular clues by intracellular trafficking and degradation pathways. This review discusses the recent and emerging information on how autophagy regulates various epithelial cell junctions. The knowledge of autophagy regulation of epithelial junctions will provide further rationale for targeting autophagy in a wide variety of human disease conditions. PMID:27583189

  14. Characteristics and EGFP expression of goat mammary gland epithelial cells.

    PubMed

    Zheng, Y-M; He, X-Y; Zhang, Y

    2010-12-01

    The aims of this study were (i) to establish a goat mammary gland epithelial (GMGE) cell line, and (ii) to determine if these GMGE cells could be maintained long-term in culture by continuous subculturing following transfection with a reporter gene, enhanced green fluorescence protein (EGFP). Primary culture of GMGE cells was achieved by outgrowth of migrating cells from the fragments of the mammary gland tissue of a lactating goat. The passage 16 GMGE cells were transfected with EGFP gene using lipofection. The expression of Cell keratins of epithelial cells in GMGE cells was test by immunofluorescence. Βeta-Casein gene mRNA was test for GMGE cells by RT-PCR. The results showed that when grown at low density on a plastic substratum, the GMGE cells formed islands, and when grown to confluency, the cells formed a monolayer and aggregated with the characteristic cobble-stone morphology of epithelial cells. GMGE cells could form dome-like structure which looked like nipple, and the lumen-like structures formed among the cells. Several blister-like structures appeared in the appearance of the cells. The GMGE cells contained different cell types, majority of the cells were short shuttle-like or polygon which were beehive-like. A part of cells were round and flat, a small number of cells were elongated. Some of the GMGE cells contained milk drops. The cell nuclei were round which had 2-4 obvious cores. The expression of Cell keratins demonstrated the property of epithelial cells in GMGE cells by immunofluorescence. The GMGE cells could express transcript encoding a Βeta-Casein protein. EGFP gene was successfully transferred into the GMGE cells, and the transfected cells could be maintained long-term in culture by continuous subculturing. In conclusion, we have established a EGFP gene transfected GMGE (ET-GMGE) cell line and maintained it long-term in culture by continuous subculturing. PMID:20113446

  15. The Epithelial Cell in Lung Health and Emphysema Pathogenesis

    PubMed Central

    Mercer, Becky A.; Lemaître, Vincent; Powell, Charles A.; D’Armiento, Jeanine

    2009-01-01

    Cigarette smoking is the primary cause of the irreversible lung disease emphysema. Historically, inflammatory cells such as macrophages and neutrophils have been studied for their role in emphysema pathology. However, recent studies indicate that the lung epithelium is an active participant in emphysema pathogenesis and plays a critical role in the lung’s response to cigarette smoke. Tobacco smoke increases protease production and alters cytokine expression in isolated epithelial cells, suggesting that these cells respond potently even in the absence of a complete inflammatory program. Tobacco smoke also acts as an immunosuppressant, reducing the defense function of airway epithelial cells and enhancing colonization of the lower airways. Thus, the paradigm that emphysema is strictly an inflammatory-cell based disease is shifting to consider the involvement of resident epithelial cells. Here we review the role of epithelial cells in lung development and emphysema. To better understand tobacco-epithelial interactions we performed microarray analyses of RNA from human airway epithelial cells exposed to smoke extract for 24 hours. These studies identified differential regulation of 425 genes involved in diverse biological processes, such as apoptosis, immune function, cell cycle, signal transduction, proliferation, and antioxidants. Some of these genes, including VEGF, glutathione peroxidase, IL-13 receptor, and cytochrome P450, have been previously reported to be altered in the lungs of smokers. Others, such as pirin, cathepsin L, STAT1, and BMP2, are shown here for the first time to have a potential role in smoke-associated injury. These data broaden our understanding of the importance of epithelial cells in lung health and cigarette smoke-induced emphysema. PMID:19662102

  16. Isolation, immortalization, and characterization of a human breast epithelial cell line with stem cell properties

    PubMed Central

    Gudjonsson, Thorarinn; Villadsen, René; Nielsen, Helga Lind; Rønnov-Jessen, Lone; Bissell, Mina J.; Petersen, Ole William

    2002-01-01

    The epithelial compartment of the human breast comprises two distinct lineages: the luminal epithelial and the myoepithelial lineage. We have shown previously that a subset of the luminal epithelial cells could convert to myoepithelial cells in culture signifying the possible existence of a progenitor cell. We therefore set out to identify and isolate the putative precursor in the luminal epithelial compartment. Using cell surface markers and immunomagnetic sorting, we isolated two luminal epithelial cell populations from primary cultures of reduction mammoplasties. The major population coexpresses sialomucin (MUC+) and epithelial-specific antigen (ESA+) whereas the minor population has a suprabasal position and expresses epithelial specific antigen but no sialomucin (MUC−/ESA+). Two cell lines were further established by transduction of the E6/E7 genes from human papilloma virus type 16. Both cell lines maintained a luminal epithelial phenotype as evidenced by expression of the tight junction proteins, claudin-1 and occludin, and by generation of a high transepithelial electrical resistance on semipermeable filters. Whereas in clonal cultures, the MUC+/ESA+ epithelial cell line was luminal epithelial restricted in its differentiation repertoire, the suprabasal-derived MUC−/ESA+ epithelial cell line was able to generate itself as well as MUC+/ESA+ epithelial cells and Thy-1+/α-smooth muscle actin+ (ASMA+) myoepithelial cells. The MUC−/ESA+ epithelial cell line further differed from the MUC+/ESA+ epithelial cell line by the expression of keratin K19, a feature of a subpopulation of epithelial cells in terminal duct lobular units in vivo. Within a reconstituted basement membrane, the MUC+/ESA+ epithelial cell line formed acinus-like spheres. In contrast, the MUC−/ESA+ epithelial cell line formed elaborate branching structures resembling uncultured terminal duct lobular units both by morphology and marker expression. Similar structures were obtained by

  17. Phototoxic aptamers selectively enter and kill epithelial cancer cells

    PubMed Central

    Ferreira, Cátia S. M.; Cheung, Melissa C.; Missailidis, Sotiris; Bisland, Stuart; Gariépy, Jean

    2009-01-01

    The majority of cancers arise from malignant epithelial cells. We report the design of synthetic oligonucleotides (aptamers) that are only internalized by epithelial cancer cells and can be precisely activated by light to kill such cells. Specifically, phototoxic DNA aptamers were selected to bind to unique short O-glycan-peptide signatures on the surface of breast, colon, lung, ovarian and pancreatic cancer cells. These surface antigens are not present on normal epithelial cells but are internalized and routed through endosomal and Golgi compartments by cancer cells, thus providing a focused mechanism for their intracellular delivery. When modified at their 5′ end with the photodynamic therapy agent chlorin e6 and delivered to epithelial cancer cells, these aptamers exhibited a remarkable enhancement (>500-fold increase) in toxicity upon light activation, compared to the drug alone and were not cytotoxic towards cell types lacking such O-glycan-peptide markers. Our findings suggest that these synthetic oligonucleotide aptamers can serve as delivery vehicles in precisely routing cytotoxic cargoes to and into epithelial cancer cells. PMID:19103663

  18. A technique to harvest viable tracheobronchial epithelial cells from living human donors.

    PubMed

    Kelsen, S G; Mardini, I A; Zhou, S; Benovic, J L; Higgins, N C

    1992-07-01

    The ability to obtain airway epithelial cells from the lower respiratory tract in living human donors will facilitate study of the biologic properties of these cells. We report our experience harvesting tracheobronchial epithelial cells from living human donors by brushing the mucosal surface of the trachea and mainstem bronchi. Cells were obtained on 21 occasions from 18 healthy adult subjects under direct vision with a brush-tipped catheter during fiberoptic bronchoscopy. The average number of cells harvested per subject was 14 +/- 2 x 10(6), and cell viability determined by trypan blue exclusion averaged 36 +/- 4%. Of note, cell viability was significantly enhanced when lidocaine was confined to the nares. Lidocaine was also observed to diminish cell viability in vitro in a dose-dependent fashion. Morphologic and staining properties were used to classify harvested cells into the three major cell types present in the mucosa (i.e., ciliated, secretory, and basal cells). All three subtypes were obtained. The percentage of ciliated, secretory, and basal-like cells was 24 +/- 2%, 11 +/- 1%, 29 +/- 1%, respectively, while the remaining 36% were difficult to type. In one subject in whom brushing was performed on three occasions over a 7-wk period, the percentage of each of the three subtypes was similar across procedures. Harvested cells could be successfully placed in primary culture with a plating efficiency of 50 to 60% and could be subcultured for up to seven passages. Acutely dissociated cells could be used to study the beta-adrenergic receptor adenylyl cyclase system since they produced cAMP in response to isoproterenol.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1320903

  19. IL-17A induces Pendrin expression and chloride-bicarbonate exchange in human bronchial epithelial cells.

    PubMed

    Adams, Kelly M; Abraham, Valsamma; Spielman, Daniel; Kolls, Jay K; Rubenstein, Ronald C; Conner, Gregory E; Cohen, Noam A; Kreindler, James L

    2014-01-01

    The epithelium plays an active role in the response to inhaled pathogens in part by responding to signals from the immune system. Epithelial responses may include changes in chemokine expression, increased mucin production and antimicrobial peptide secretion, and changes in ion transport. We previously demonstrated that interleukin-17A (IL-17A), which is critical for lung host defense against extracellular bacteria, significantly raised airway surface pH in vitro, a finding that is common to a number of inflammatory diseases. Using microarray analysis of normal human bronchial epithelial (HBE) cells treated with IL-17A, we identified the electroneutral chloride-bicarbonate exchanger Pendrin (SLC26A4) as a potential mediator of this effect. These data were verified by real-time, quantitative PCR that demonstrated a time-dependent increase in Pendrin mRNA expression in HBE cells treated with IL-17A up to 48 h. Using immunoblotting and immunofluorescence, we confirmed that Pendrin protein expression is increased in IL-17 treated HBE cells and that it is primarily localized to the mucosal surface of the cells. Functional studies using live-cell fluorescence to measure intracellular pH demonstrated that IL-17A induced chloride-bicarbonate exchange in HBE cells that was not present in the absence of IL-17A. Furthermore, HBE cells treated with short interfering RNA against Pendrin showed substantially reduced chloride-bicarbonate exchange. These data suggest that Pendrin is part of IL-17A-dependent epithelial changes and that Pendrin may therefore be a therapeutic target in IL-17A-dependent lung disease. PMID:25141009

  20. Intestinal inflammation and mucosal barrier function.

    PubMed

    Sánchez de Medina, Fermín; Romero-Calvo, Isabel; Mascaraque, Cristina; Martínez-Augustin, Olga

    2014-12-01

    Intestinal mucosal barrier function is the capacity of the intestine to provide adequate containment of luminal microorganisms and molecules while preserving the ability to absorb nutrients. The central element is the epithelial layer, which physically separates the lumen and the internal milieu and is in charge of vectorial transport of ions, nutrients, and other substances. The secretion of mucus-forming mucins, sIgA, and antimicrobial peptides reinforces the mucosal barrier on the extraepithelial side, while a variety of immune cells contributes to mucosal defense in the inner side. Thus, the mucosal barrier is of physical, biochemical, and immune nature. In addition, the microbiota may be viewed as part of this system because of the mutual influence occurring between the host and the luminal microorganisms. Alteration of the mucosal barrier function with accompanying increased permeability and/or bacterial translocation has been linked with a variety of conditions, including inflammatory bowel disease. Genetic and environmental factors may converge to evoke a defective function of the barrier, which in turn may lead to overt inflammation of the intestine as a result of an exacerbated immune reaction toward the microbiota. According to this hypothesis, inflammatory bowel disease may be both precipitated and treated by either stimulation or downregulation of the different elements of the mucosal barrier, with the outcome depending on timing, the cell type affected, and other factors. In this review, we cover briefly the elements of the barrier and their involvement in functional defects and the resulting phenotype. PMID:25222662

  1. Probiotics promote endocytic allergen degradation in gut epithelial cells

    SciTech Connect

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

    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.

  2. Research controversies in management of oral mucositis.

    PubMed

    Biron, P; Sebban, C; Gourmet, R; Chvetzoff, G; Philip, I; Blay, J Y

    2000-01-01

    The management of mucositis is the subject of many controversies, and the optimal treatment is still not known. Several evaluation scoring systems have been described, but no one of these is appropriate to all clinical situations: a simple scale such as that devised by the WHO can be used routinely, and more sophisticated ones can be implemented by trained experimenters working in research. We have considered the impact of each of the treatments currently available on each stage of mucositis. In attempts at prevention, self-care, in the sense of oral hygiene, must remain atraumatic. It is probably advisable to differentiate patients with good previous oral care, in whom tooth brushing is beneficial, from others, in whom the risk of hemorrhage and infection excludes any brushing. Before the dosage of chemotherapy is reduced, the curative or palliative intent of the strategy must be carefully evaluated. In the vascular phase protection of the proliferating cells is attempted by means of vasoconstriction (cryotherapy), cytoprotection (prostaglandin E2 and other antioxidants) or epithelial cell-inhibiting factors such as TGF-B3. Treatments applied in the epithelial phase are directed at increasing the cell proliferation to accelerate epithelial restoration by sucralfate and several growth factors: hematopoietic GF, which has demonstrated a direct effect on the mucosa (GM-CSF), or epithelial growth factors such as keratinocyte GF. In the ulcerative and bacteriological phase attempts are made to attenuate sepsis by means of antiseptics (chlorhexidine), amphotericin B and antiviral agents or antibiotic lozenges. In the healing phase application of the low-energy helium-neon laser has demonstrably been followed by a later time of onset, less pronounced peak severity and shorter duration of oral mucositis. After cancer treatment, oral hygiene, inhibition of oral flora, and pain relief are the main goals. Physiopathogen-specific treatment is the next step, with the emphasis

  3. Lateral adhesion drives reintegration of misplaced cells into epithelial monolayers.

    PubMed

    Bergstralh, Dan T; Lovegrove, Holly E; St Johnston, Daniel

    2015-11-01

    Cells in simple epithelia orient their mitotic spindles in the plane of the epithelium so that both daughter cells are born within the epithelial sheet. This is assumed to be important to maintain epithelial integrity and prevent hyperplasia, because misaligned divisions give rise to cells outside the epithelium. Here we test this assumption in three types of Drosophila epithelium; the cuboidal follicle epithelium, the columnar early embryonic ectoderm, and the pseudostratified neuroepithelium. Ectopic expression of Inscuteable in these tissues reorients mitotic spindles, resulting in one daughter cell being born outside the epithelial layer. Live imaging reveals that these misplaced cells reintegrate into the tissue. Reducing the levels of the lateral homophilic adhesion molecules Neuroglian or Fasciclin 2 disrupts reintegration, giving rise to extra-epithelial cells, whereas disruption of adherens junctions has no effect. Thus, the reinsertion of misplaced cells seems to be driven by lateral adhesion, which pulls cells born outside the epithelial layer back into it. Our findings reveal a robust mechanism that protects epithelia against the consequences of misoriented divisions. PMID:26414404

  4. Lingual Epithelial Stem Cells and Organoid Culture of Them

    PubMed Central

    Hisha, Hiroko; Tanaka, Toshihiro; Ueno, Hiroo

    2016-01-01

    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. PMID:26828484

  5. Effects of ethanol on an intestinal epithelial cell line

    SciTech Connect

    Nano, J.L.; Cefai, D.; Rampal, P. )

    1990-02-01

    The effect of exposure of an intestinal epithelial cell line to various concentrations of ethanol (217 mM (1%) to 652 mM (3%)) during 24, 48, and 72 hr was investigated in vitro using a rat intestinal epithelial cell line (IRD 98). Incubation of these cells in the presence of ethanol significantly decreased cell growth. This inhibition was accompanied by a strong increase in cellular protein. Stimulation of specific disaccharidases, gamma-glutamyl transferase, and aminopeptidase activities by ethanol was dose- and time-dependent. Ethanol induces a change in the relative proportions of the different lipid classes synthesized; triglycerides, fatty acids, and cholesterol esters were preferentially synthethysed. Our findings show that cell lines are good models for investigation of the effects of ethanol, and that alcohol considerably modifies the functions of intestinal epithelial cells.

  6. Development of human epithelial cell systems for radiation risk assessment

    NASA Technical Reports Server (NTRS)

    Yang, C. H.; Craise, L. M.

    1994-01-01

    The most important health effect of space radiation for astronauts is cancer induction. For radiation risk assessment, an understanding of carcinogenic effect of heavy ions in human cells is most essential. In our laboratory, we have successfully developed a human mammary epithelial cell system for studying the neoplastic transformation in vitro. Growth variants were obtained from heavy ion irradiated immortal mammary cell line. These cloned growth variants can grow in regular tissue culture media and maintain anchorage dependent growth and density inhibition property. Upon further irradiation with high-Linear Energy Transfer (LET) radiation, transformed foci were found. Experimental results from these studies suggest that multiexposure of radiation is required to induce neoplastic tranformation of human epithelial cells. This multihits requirement may be due to high genomic stability of human cells. These growth variants can be useful model systems for space flight experiments to determine the carcinogenic effect of space radiation in human epithelial cells.

  7. Medullary thymic epithelial stem cells: role in thymic epithelial cell maintenance and thymic involution.

    PubMed

    Hamazaki, Yoko; Sekai, Miho; Minato, Nagahiro

    2016-05-01

    The thymus consists of two distinct anatomical regions, the cortex and the medulla; medullary thymic epithelial cells (mTECs) play a crucial role in establishing central T-cell tolerance for self-antigens. Although the understanding of mTEC development in thymic organogenesis as well as the regulation of their differentiation and maturation has improved, the mechanisms of postnatal maintenance remain poorly understood. This issue has a central importance in immune homeostasis and physiological thymic involution as well as autoimmune disorders in various clinicopathological settings. Recently, several reports have demonstrated the existence of TEC stem or progenitor cells in the postnatal thymus, which are either bipotent or unipotent. We identified stem cells specified for mTEC-lineage that are generated in the thymic ontogeny and may sustain mTEC regeneration and lifelong central T-cell self-tolerance. This finding suggested that the thymic medulla is maintained autonomously by its own stem cells. Although several issues, including the relationship with other putative TEC stem/progenitors, remain unclear, further examination of mTEC stem cells (mTECSCs) and their regulatory mechanisms may contribute to the understanding of postnatal immune homeostasis. Possible relationships between decline of mTECSC activity and early thymic involution as well as various autoimmune disorders are discussed. PMID:27088906

  8. Assessing the adhesion of putative indigenous probiotic lactobacilli to human colonic epithelial cells

    PubMed Central

    Duary, Raj Kumar; Rajput, Yudhishthir Singh; Batish, Virender Kumar; Grover, Sunita

    2011-01-01

    Background & objectives: Adherence of bacteria to epithelial cells and mucosal surfaces is a key criterion for selection of probiotic. We assessed the adhesion property of selected indigenous probiotic Lactobacillus strains based on their hydrophobicity and ability to adhere to human epithelial cells. Methods: Five human faecal Lactobacillus isolates, one from buffalo milk and one from cheese were assessed for hydrophobicity following the microbial adhesion to hydrocarbons (MATH) method and colonization potentials based on their adherence to Caco2 and HT-29 colonic adenocarcinomal human intestinal epithelial cell lines. Lactobacillus strains that adhered to Caco2 and HT-29 cell lines were quantified by plating after trypsinization and simultaneously the adhered bacteria were also examined microscopically after staining with Geimsa stain and counted in different fields. Results: Among the tested faecal isolates, L. plantarum Lp91 showed maximum percentage hydrophobicity (35.73±0.40 for n-hexadecane and 34.26±0.63 for toluene) closely followed by L. plantarum Lp9 (35.53±0.29 for n-hexadecane and 33.00±0.57 for toluene). Based on direct adhesion to epithelial cells, L. plantarum Lp91 was the most adhesive strain to HT-29 and Caco2 cell lines with per cent adhesion values of 12.8 ± 1.56 and 10.2 ± 1.09, respectively. L. delbrukeii CH4, was the least adhesive with corresponding figures of 2.5 ± 0.37 and 2.6 ± 0.20 per cent on HT-29 and Caco2 cell lines. Adhesion of the six isolated Lactobacillus strain to HT-29 cell and Caco2 lines as recorded under microscope varied between 131.0 ± 13.9 (Lp75) to 342.7 ± 50.52 (Lp91) and 44.7 ± 9.29 (CH4) to 315.7± 35.4 (Lp91), respectively. Interpretation & conclusions: Two Indigenous probiotic Lactobacillus strains (Lp9, Lp91) demonstrated their ability to adhere to epithelial cell and exhibited strong hydrophobicity under in vitro conditions, and thus could have better prospects to colonize the gut with extended transit

  9. Middle ear mucosal regeneration with three-dimensionally tissue-engineered autologous middle ear cell sheets in rabbit model.

    PubMed

    Yaguchi, Yuichiro; Murakami, Daisuke; Yamato, Masayuki; Hama, Takanori; Yamamoto, Kazuhisa; Kojima, Hiromi; Moriyama, Hiroshi; Okano, Teruo

    2016-03-01

    The likelihood of recurrent retraction and adhesion of newly formed tympanic membrane is high when middle ear mucosa is extensively lost during cholesteatoma and adhesive otitis media surgery. If rapid postoperative regeneration of the mucosa on the exposed bone surface can be achieved, prevention of recurrent eardrum adhesion and cholesteatoma formation, for which there has been no definitive treatment, can be expected. Suture-less transplantation of tissue-engineered mucosal cell sheets was examined immediately after the operation of otitis media surgery in order to quickly regenerate middle ear mucosa lost during surgery in a rabbit model. Transplantable middle ear mucosal cell sheets with a three-dimensional tissue architecture very similar to native middle ear mucosa were fabricated from middle ear mucosal tissue fragments obtained in an autologous manner from middle ear bulla on temperature-responsive culture surfaces. Immediately after the mucosa was resected from middle ear bone bulla inner cavity, mucosal cell sheets were grafted at the resected site. Both bone hyperplasia and granulation tissue formation were inhibited and early mucosal regeneration was observed in the cell sheet-grafted group, compared with the control group in which only mucosal removal was carried out and the bone surface exposed. This result indicates that tissue engineered mucosal cell sheets would be useful to minimize complications after the surgical operation on otitis media and future clinical application is expected. Copyright © 2013 John Wiley & Sons, Ltd. PMID:23894137

  10. Osteonecrosis of the Jaw Developed in Mice: DISEASE VARIANTS REGULATED BY γδ T CELLS IN ORAL MUCOSAL BARRIER IMMUNITY.

    PubMed

    Park, Sil; Kanayama, Keiichi; Kaur, Kawaljit; Tseng, Han-Ching Helen; Banankhah, Sina; Quje, Davood Talebi; Sayre, James W; Jewett, Anahid; Nishimura, Ichiro

    2015-07-10

    Osteonecrosis of the jaw (ONJ), an uncommon co-morbidity in patients treated with bisphosphonates (BP), occurs in the segment of jawbone interfacing oral mucosa. This study aimed to investigate a role of oral mucosal barrier γδ T cells in the pathogenesis of ONJ. Female C57Bl/6J (B6) mice received a bolus zoledronate intravenous injection (ZOL, 540 μg/kg), and their maxillary left first molars were extracted 1 week later. ZOL-treated mice (WT ZOL) delayed oral wound healing with patent open wounds 4 weeks after tooth extraction with characteristic oral epithelial hyperplasia. γδ T cells appeared within the tooth extraction site and hyperplastic epithelium in WT ZOL mice. In ZOL-treated γδ T cell null (Tcrd(-/-) ZOL) mice, the tooth extraction open wound progressively closed; however, histological ONJ-like lesions were identified in 75 and 60% of WT ZOL and Tcrd(-/-) ZOL mice, respectively. Although the bone exposure phenotype of ONJ was predominantly observed in WT ZOL mice, Tcrd(-/-) ZOL mice developed the pustule/fistula disease phenotype. We further addressed the role of γδ T cells from human peripheral blood (h-γδ T cells). When co-cultured with ZOL-pretreated human osteoclasts in vitro, h-γδ T cells exhibited rapid expansion and robust IFN-γ secretion. When h-γδ T cells were injected into ZOL-treated immunodeficient (Rag2(-/-) ZOL) mice, the oral epithelial hyperplasia developed. However, Rag2(-/-) ZOL mice did not develop osteonecrosis. The results indicate that γδ T cells are unlikely to influence the core osteonecrosis mechanism; however, they may serve as a critical modifier contributing to the different oral mucosal disease variations of ONJ. PMID:26013832

  11. Heterogeneity and stochastic growth regulation of biliary epithelial cells dictate dynamic epithelial tissue remodeling

    PubMed Central

    Kamimoto, Kenji; Kaneko, Kota; Kok, Cindy Yuet-Yin; Okada, Hajime; Miyajima, Atsushi; Itoh, Tohru

    2016-01-01

    Dynamic remodeling of the intrahepatic biliary epithelial tissue plays key roles in liver regeneration, yet the cellular basis for this process remains unclear. We took an unbiased approach based on in vivo clonal labeling and tracking of biliary epithelial cells in the three-dimensional landscape, in combination with mathematical simulation, to understand their mode of proliferation in a mouse liver injury model where the nascent biliary structure formed in a tissue-intrinsic manner. An apparent heterogeneity among biliary epithelial cells was observed: whereas most of the responders that entered the cell cycle upon injury exhibited a limited and tapering growth potential, a select population continued to proliferate, making a major contribution in sustaining the biliary expansion. Our study has highlighted a unique mode of epithelial tissue dynamics, which depends not on a hierarchical system driven by fixated stem cells, but rather, on a stochastically maintained progenitor population with persistent proliferative activity. DOI: http://dx.doi.org/10.7554/eLife.15034.001 PMID:27431614

  12. CHARACTERIZATION OF ALVEOLAR EPITHELIAL CELLS CULTURED IN SEMIPERMEABLE HOLLOW FIBERS

    PubMed Central

    Grek, Christina L.; Newton, Danforth A.; Qiu, Yonhzhi; Wen, Xuejun; Spyropoulos, Demetri D.; Baatz, John E.

    2012-01-01

    Cell culture methods commonly used to represent alveolar epithelial cells in vivo have lacked airflow, a 3-dimensional air-liquid interface, and dynamic stretching characteristics of native lung tissue—physiological parameters critical for normal phenotypic gene expression and cellular function. Here the authors report the development of a selectively semipermeable hollow fiber culture system that more accurately mimics the in vivo microenvironment experienced by mammalian distal airway cells than in conventional or standard air-liquid interface culture. Murine lung epithelial cells (MLE-15) were cultured within semipermeable polyurethane hollow fibers and introduced to controlled airflow through the microfiber interior. Under these conditions, MLE-15 cells formed confluent monolayers, demonstrated a cuboidal morphology, formed tight junctions, and produced and secreted surfactant proteins. Numerous lamellar bodies and microvilli were present in MLE-15 cells grown in hollow fiber culture. Conversely, these alveolar type II cell characteristics were reduced in MLE-15 cells cultured in conventional 2D static culture systems. These data support the hypothesis that MLE-15 cells grown within our microfiber culture system in the presence of airflow maintain the phenotypic characteristics of type II cells to a higher degree than those grown in standard in vitro cell culture models. Application of our novel model system may prove advantageous for future studies of specific gene and protein expression involving alveolar epithelial or bronchiolar epithelial cells. PMID:19263283

  13. Apoptotic epithelial cells control the abundance of Treg cells at barrier surfaces.

    PubMed

    Nakahashi-Oda, Chigusa; Udayanga, Kankanam Gamage Sanath; Nakamura, Yoshiyuki; Nakazawa, Yuta; Totsuka, Naoya; Miki, Haruka; Iino, Shuichi; Tahara-Hanaoka, Satoko; Honda, Shin-ichiro; Shibuya, Kazuko; Shibuya, Akira

    2016-04-01

    Epithelial tissues continually undergo apoptosis. Commensal organisms that inhabit the epithelium influence tissue homeostasis, in which regulatory T cells (Treg cells) have a central role. However, the physiological importance of epithelial cell apoptosis and how the number of Treg cells is regulated are both incompletely understood. Here we found that apoptotic epithelial cells negatively regulated the commensal-stimulated proliferation of Treg cells. Gut commensals stimulated CX3CR1(+)CD103(-)CD11b(+) dendritic cells (DCs) to produce interferon-β (IFN-β), which augmented the proliferation of Treg cells in the intestine. Conversely, phosphatidylserine exposed on apoptotic epithelial cells suppressed IFN-β production by the DCs via inhibitory signaling mediated by the cell-surface glycoprotein CD300a and thus suppressed Treg cell proliferation. Our findings reveal a regulatory role for apoptotic epithelial cells in maintaining the number of Treg cell and tissue homeostasis. PMID:26855029

  14. Mechanobiology in Lung Epithelial Cells: Measurements, Perturbations, and Responses

    PubMed Central

    Waters, Christopher M.; Roan, Esra; Navajas, Daniel

    2015-01-01

    Epithelial cells of the lung are located at the interface between the environment and the organism and serve many important functions including barrier protection, fluid balance, clearance of particulate, initiation of immune responses, mucus and surfactant production, and repair following injury. Because of the complex structure of the lung and its cyclic deformation during the respiratory cycle, epithelial cells are exposed to continuously varying levels of mechanical stresses. While normal lung function is maintained under these conditions, changes in mechanical stresses can have profound effects on the function of epithelial cells and therefore the function of the organ. In this review, we will describe the types of stresses and strains in the lungs, how these are transmitted, and how these may vary in human disease or animal models. Many approaches have been developed to better understand how cells sense and respond to mechanical stresses, and we will discuss these approaches and how they have been used to study lung epithelial cells in culture. Understanding how cells sense and respond to changes in mechanical stresses will contribute to our understanding of the role of lung epithelial cells during normal function and development and how their function may change in diseases such as acute lung injury, asthma, emphysema, and fibrosis. PMID:23728969

  15. Cell phone radiation effects on cytogenetic abnormalities of oral mucosal cells.

    PubMed

    Daroit, Natália Batista; Visioli, Fernanda; Magnusson, Alessandra Selinger; Vieira, Geila Radunz; Rados, Pantelis Varvaki

    2015-01-01

    The aim of this study was to evaluate the effects of exposure to cell phone electromagnetic radiation on the frequency of micronuclei, broken eggs cells, binucleated cells, and karyorrhexis in epithelial cells of the oral mucosa. The sample was composed of 60 cell phone users, who were non-smokers and non-drinkers, and had no clinically visible oral lesions. Cells were obtained from anatomical sites with the highest incidence of oral cancer: lower lip, border of the tongue, and floor of the mouth. The Feulgen reaction was used for quantification of nuclear anomalies in 1,000 cells/slide. A slightly increase in the number of micronucleated cells in the lower lip and in binucleated cells on the floor of the mouth was observed in individuals who used their phones > 60 minutes/week. The analysis also revealed an increased number of broken eggs in the tongue of individuals owning a cell phone for over eight years. Results suggest that exposure to electromagnetic waves emitted by cell phones can increase nuclear abnormalities in individuals who use a cell phone for more than 60 minutes per week and for over eight years. Based on the present findings, we suggest that exposure to electromagnetic radiation emitted by cell phones may interfere with the development of metanuclear anomalies. Therefore, it is demonstrated that, despite a significant increase in these anomalies, the radiation emitted by cell phones among frequent users is within acceptable physiological limits. PMID:26486771

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

    SciTech Connect

    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.

  17. DNA damage, apoptosis and cell cycle changes induced by fluoride in rat oral mucosal cells and hepatocytes

    PubMed Central

    He, Ling-Fei; Chen, Jian-Gang

    2006-01-01

    AIM: To study the effect of fluoride on oxidative stress, DNA damage and apoptosis as well as cell cycle of rat oral mucosal cells and hepatocytes. METHODS: Ten male SD rats weighing 80~120 g were randomly divided into control group and fluoride group, 5 animals each group. The animals in fluoride group had free access to deionized water containing 150 mg/L sodium fluoride (NaF). The animals in control group were given distilled water. Four weeks later, the animals were killed. Reactive oxygen species (ROS) in oral mucosa and liver were measured by Fenton reaction, lipid peroxidation product, malondialdehyde (MDA), was detected by thiobarbituric acid (TBA) reaction, reduced glutathione (GSH) was assayed by dithionitrobenzoic acid (DTNB) reaction. DNA damage in oral mucosal cells and hepatocytes was determined by single cell gel (SCG) electrophoresis or comet assay. Apoptosis and cell cycle in oral mucosal cells and hepatocytes were detected by flow cytometry. RESULTS: The contents of ROS and MDA in oral mucosa and liver tissue of fluoride group were significantly higher than those of control group (P < 0.01), but the level of GSH was markedly decreased (P < 0.01). The contents of ROS, MDA and GSH were (134.73 ± 12.63) U/mg protein, (1.48 ± 0.13) mmol/mg protein and (76.38 ± 6.71) mmol/mg protein in oral mucosa respectively, and (143.45 ±11.76) U/mg protein, (1.44 ± 0.12) mmol/mg protein and (78.83 ± 7.72) mmol/mg protein in liver tissue respectively. The DNA damage rate in fluoride group was 50.20% in oral mucosal cells and 44.80% in hepatocytes, higher than those in the control group (P < 0.01). The apoptosis rate in oral mucosal cells was (13.63 ± 1.81) % in fluoride group, and (12.76 ± 1.67) % in hepatocytes, higher than those in control group. Excess fluoride could differently lower the number of oral mucosal cells and hepatocytes at G0/G1 and S G2/M phases (P < 0.05). CONCLUSION: Excess fluoride can induce

  18. Detection of Epstein-Barr virus genome and latent infection gene expression in normal epithelia, epithelial dysplasia, and squamous cell carcinoma of the oral cavity.

    PubMed

    Kikuchi, Kentaro; Noguchi, Yoshihiro; de Rivera, Michelle Wendoline Garcia-Niño; Hoshino, Miyako; Sakashita, Hideaki; Yamada, Tsutomu; Inoue, Harumi; Miyazaki, Yuji; Nozaki, Tadashige; González-López, Blanca Silvia; Ide, Fumio; Kusama, Kaoru

    2016-03-01

    A relationship between Epstein-Barr virus (EBV) infection and cancer of lymphoid and epithelial tissues such as Burkitt's lymphoma, Hodgkin's disease, nasopharyngeal carcinoma (NPC), gastric carcinoma, and oral cancer has been reported. EBV is transmitted orally and infects B cells and epithelial cells. However, it has remained uncertain whether EBV plays a role in carcinogenesis of oral mucosal tissue. In the present study, we detected the EBV genome and latent EBV gene expression in normal mucosal epithelia, epithelial dysplasia, and oral squamous cell carcinoma (OSCC) to clarify whether EBV is involved in carcinogenesis of the oral cavity. We examined 333 formalin-fixed, paraffin-embedded tissue samples (morphologically normal oral mucosa 30 samples, gingivitis 32, tonsillitis 17, oral epithelial dysplasia 83, OSCC 150, and NPC 21). EBV latent infection genes (EBNA-2, LMP-1) were detected not only in OSCC (50.2 %, 10.7 %) but also in severe epithelial dysplasia (66.7 %, 44.4 %), mild to moderate epithelial dysplasia (43.1 %, 18.5 %), gingivitis (78.1 %, 21.9 %), and normal mucosa (83.3 %, 23.3 %). Furthermore, the intensity of EBV latent infection gene expression (EBER, LMP-1) was significantly higher in severe epithelial dysplasia (94.4 %, 72.2 %) than in OSCC (34.7 %, 38.7 %). These results suggest that EBV latent infection genes and their increased expression in severe epithelial dysplasia might play an important role in the dysplasia-carcinoma sequence in the oral cavity. PMID:26449822

  19. Porphyromonas gingivalis invades oral epithelial cells in vitro.

    PubMed

    Sandros, J; Papapanou, P; Dahlén, G

    1993-05-01

    The aim of the present study was to analyze the adhesive and invasive potential of a number of P. gingivalis strains, in an in vitro system utilizing cultures of human oral epithelial cells (KB cell line, ATCC CCL 17). P. gingivalis strains W50 and FDC 381 (laboratory strains) and OMGS 1738, 1743 and 1439 (clinical isolates) as well as E. coli strain HB 101 (non-adhering, non-invasive control) were used. Adherence was assessed by means of scintillation counting and light microscopy, after incubation of radiolabelled bacteria with epithelial cells. In the invasion assay, monolayers were infected with the P. gingivalis and E. coli strains and further incubated with an antibiotic mixture (metronidazole 0.1 mg/ml and gentamicin 0.5 mg/ml). Invasion was evaluated by (i) assessing presence of bacteria surviving the antibiotic treatment, and (ii) electron microscopy. All P. gingivalis strains adhered to and entered into the oral epithelial cells. After 3 hours of incubation, bacteria were frequently identified intracellularly by means of electron microscopy. The cellular membranes, encapsulating the microorganisms in early stages of the invasive process, appeared later to disintegrate. The presence of coated pits on the epithelial cell surfaces suggested that internalization of P. gingivalis was associated with receptor-mediated endocytosis (RME). Formation of outer membrane vesicles (blebs) by intracellular bacteria indicated that internalized P. gingivalis was able to retain its viability. E. coli strain HB 101 neither adhered to nor invaded epithelial cells. PMID:8388449

  20. Porphyromonas gingivalis Fimbriae Bind to Cytokeratin of Epithelial Cells

    PubMed Central

    Sojar, Hakimuddin T.; Sharma, Ashu; Genco, Robert J.

    2002-01-01

    The adherence of Porphyromonas gingivalis to host cells is likely a prerequisite step in the pathogenesis of P. gingivalis-induced periodontal disease. P. gingivalis binds to and invades epithelial cells, and fimbriae are shown to be involved in this process. Little is known regarding epithelial receptor(s) involved in binding of P. gingivalis fimbriae. Using an overlay assay with purified P. gingivalis fimbriae as a probe, two major epithelial cell proteins with masses of 50 and 40 kDa were identified by immunoblotting with fimbria-specific antibodies. Iodinated purified fimbriae also bound to the same two epithelial cell proteins. An affinity chromatography technique was utilized to isolate and purify the epithelial components to which P. gingivalis fimbriae bind. Purified fimbriae were coupled to CNBr-activated Sepharose-4B, and the solubilized epithelial cell extract proteins bound to the immobilized fimbriae were isolated from the column. A major 50-kDa component and a minor 40-kDa component were purified and could be digested with trypsin, suggesting that they were proteins. These affinity-eluted 50- and 40-kDa proteins were then subjected to amino-terminal sequencing, and no sequence could be determined, suggesting that these proteins have blocked amino-terminal residues. CNBr digestion of the 50-kDa component resulted in an internal sequence homologous to that of Keratin I molecules. Further evidence that P. gingivalis fimbriae bind to cytokeratin molecule(s) comes from studies showing that multicytokeratin rabbit polyclonal antibodies cross-react with the affinity-purified 50-kDa epithelial cell surface component. Also, binding of purified P. gingivalis fimbriae to epithelial components can be inhibited in an overlay assay by multicytokeratin rabbit polyclonal antibodies. Furthermore, we showed that biotinylated purified fimbriae bind to purified human epidermal keratin in an overlay assay. These studies suggest that the surface-accessible epithelial

  1. Differentiation of porcine mesenchymal stem cells into epithelial cells as a potential therapeutic application to facilitate epithelial regeneration.

    PubMed

    Kokubun, Kelsey; Pankajakshan, Divya; Kim, Min-Jung; Agrawal, Devendra K

    2016-02-01

    Epithelial denudation is one of the characteristics of chronic asthma. To restore its functions, the airway epithelium has to rapidly repair the injuries and regenerate its structure and integrity. Mesenchymal stem cells (MSCs) have the ability to differentiate into many cell lineages. However, the differentiation of MSCs into epithelial cells has not been fully studied. Here, we examined the differentiation of MSCs into epithelial cells using three different media compositions with various growth supplementations. The MSCs were isolated from porcine bone marrow by density gradient centrifugation. The isolated MSCs were CD11(-) CD34(-) CD45(-) CD44(+) CD90(+) and CD105(+) by immunostaining and flow cytometry. MSCs were stimulated with EpiGRO (Millipore), BEpiCM (ScienCell) and AECGM (PromoCell) media for 5 and 10 days, and epithelial differentiation was assessed by qPCR (keratin 14, 18 and EpCAM), fluorometry (cytokeratin 7-8, cytokeratin 14-15-16-19 and EpCAM), western blot analysis (pancytokeratin, EpCAM) and flow cytometry (cytokeratin 7-8, cytokeratin 14-15-16-19 and EpCAM). The functional marker MUC1 was also assessed after 10 days of air-liquid interface (ALI) culture in optimized media. Cells cultured in BEpiCM containing fibroblast growth factor and prostaglandin E2 showed the highest expression of the epithelial markers: CK7-8 (85.90%); CK-14-15-16-19 (10.14%); and EpCAM (64.61%). The cells also expressed functional marker MUC1 after ALI culture. The differentiated MSCs when cultured in BEpiCM medium ex vivo in a bioreactor on a decellularized trachea for 10 days retained the epithelial-like phenotype. In conclusion, porcine bone marrow-derived MSCs demonstrate commitment to the epithelial lineage and might be a potential therapy for facilitating the repair of denuded airway epithelium. PMID:23696537

  2. Sphingosine-1-Phosphate Protects Intestinal Epithelial Cells from Apoptosis Through the Akt Signaling Pathway

    PubMed Central

    Greenspon, Jose; Li, Ruiyun; Xiao, Lan; Rao, Jaladanki N.; Marasa, Bernard S.; Strauch, Eric D.; Wang, Jian-Ying; Turner, Douglas J.

    2009-01-01

    Objective The regulation of apoptosis of intestinal mucosal cells is important in maintenance of normal intestinal physiology. Summary Sphingosine-1-phosphate (S1P) has been shown to play a critical role in cellular protection to otherwise lethal stimuli in several nonintestinal tissues. Methods The current study determines whether S1P protected normal intestinal epithelial cells (IECs) from apoptosis and whether Akt activation was the central pathway for this effect. Results S1P demonstrated significantly reduced levels of apoptosis induced by tumor necrosis factor-alpha (TNF-α)/cycloheximide (CHX). S1P induced increased levels of phosphorylated Akt and increased Akt activity, but did not affect total amounts of Akt. This activation of Akt was associated with decreased levels of both caspase-3 protein levels and of caspase-3 activity. Inactivation of Akt by treatment with the PI3K chemical inhibitor LY294002 or by overexpression of the dominant negative mutant of Akt (DNMAkt) prevented the protective effect of S1P on apoptosis. Additionally, silencing of the S1P-1 receptor by specific siRNA demonstrated a lesser decrease in apoptosis to S1P exposure. Conclusion These results indicate that S1P protects intestinal epithelial cells from apoptosis via an Akt-dependent pathway. PMID:18654850

  3. Isolation and Characterization of Intestinal Epithelial Cells from Normal and SIV-Infected Rhesus Macaques

    PubMed Central

    Pan, Diganta; Das, Arpita; Liu, David; Veazey, Ronald S.; Pahar, Bapi

    2012-01-01

    Impairment of intestinal epithelial barriers contributes to the progression of HIV/SIV infection and leads to generalized HIV-induced immune-cell activation during chronic infection. Rhesus macaques are the major animal model for studying HIV pathogenesis. However, detailed characterization of isolated rhesus epithelial cells (ECs) from intestinal tissues is not well defined. It is also not well documented whether isolated ECs had any other cell contaminants from intestinal tissues during the time of processing that might hamper interpretation of EC preparations or cultures. In this study, we identify and characterize ECs based on flow cytometry and immunohistochemistry methods using various enzymatic and mechanical isolation techniques to enrich ECs from intestinal tissues. This study shows that normal healthy ECs differentially express HLA-DR, CD23, CD27, CD90, CD95 and IL-10R markers. Early apoptosis and upregulation of ICAM-1 and HLA-DR in intestinal ECs are thought to be the key features in SIV mediated enteropathy. The data suggest that intestinal ECs might be playing an important role in mucosal immune responses by regulating the expression of different important regulatory and adhesion molecules and their function. PMID:22291924

  4. Amniotic epithelial cells promote wound healing in mice through high epithelialization and engraftment.

    PubMed

    Jin, Enze; Kim, Tae-Hee; Han, Seongho; Kim, Sung-Whan

    2016-07-01

    Although human amniotic epithelial cells (AMEs) are an attractive source of stem cells, their therapeutic potential in wound healing has not been fully investigated. We evaluated the therapeutic potential of AMEs for wound healing. Real-time PCR showed that the epithelialization growth factors epidermal growth factor (EGF), platelet-derived growth factor (PDGF)-B and chemotactic factors interleukin-8 (IL-8 or CXCL8) and neutrophil-activating protein-2 (NAP-2 or CXCL7) were upregulated in AMEs compared with adipose-derived mesenchymal stem cells (ADMs). In vitro scratch wound assays revealed that AME-derived conditioned medium substantially accelerated wound closure. Wounds in NOD/SCID mice were created by skin excision, followed by AME transplantation. AMEs implantation significantly accelerated wound healing and increased cellularity and re-epithelialization. Transplanted AMEs exhibited high engraftment rates and expressed keratinocyte-specific proteins and cytokeratin in the wound area, suggesting direct benefits for cutaneous closure. Taken together, these data indicate that AMEs possess therapeutic capability for wound healing through the secretion of epithelialization growth factors and enhanced engraftment properties. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26174407

  5. Aggressive Calcifying Epithelial Odontogenic Tumor of the Maxillary Sinus with Extraosseous Oral Mucosal Involvement: A Case Report

    PubMed Central

    Rani, Vidya; Masthan, Mahaboob Kadar; Aravindha, Babu; Leena, Sankari

    2016-01-01

    Calcifying epithelial odontogenic tumors are benign odontogenic neoplasms whose occurrence in the maxillary sinus is rare. Maxillary tumors tend to be locally aggressive and may rapidly involve the surrounding vital structures. We report a case of a large calcifying epithelial odontogenic tumor of the maxilla, involving the maxillary sinus in a 48-year-old woman. The tumor was largely intraosseous. In the canine and first premolar regions, the loss of bone could be palpated but the oral mucosa appeared normal. Histologically, the tumor tissue could be seen in the connective tissue below the oral epithelium. The most significant finding was the presence of an intraosseous tumor with an extraosseous involvement in a single tumor, indicating aggressive behavior and warranting aggressive treatment. In this article, we discuss the rare presentation of the tumor and its radiological appearance and histological features. We also highlight the importance of a detailed histopathological examination of the excised specimen. PMID:26989286

  6. Aggressive Calcifying Epithelial Odontogenic Tumor of the Maxillary Sinus with Extraosseous Oral Mucosal Involvement: A Case Report.

    PubMed

    Rani, Vidya; Masthan, Mahaboob Kadar; Aravindha, Babu; Leena, Sankari

    2016-03-01

    Calcifying epithelial odontogenic tumors are benign odontogenic neoplasms whose occurrence in the maxillary sinus is rare. Maxillary tumors tend to be locally aggressive and may rapidly involve the surrounding vital structures. We report a case of a large calcifying epithelial odontogenic tumor of the maxilla, involving the maxillary sinus in a 48-year-old woman. The tumor was largely intraosseous. In the canine and first premolar regions, the loss of bone could be palpated but the oral mucosa appeared normal. Histologically, the tumor tissue could be seen in the connective tissue below the oral epithelium. The most significant finding was the presence of an intraosseous tumor with an extraosseous involvement in a single tumor, indicating aggressive behavior and warranting aggressive treatment. In this article, we discuss the rare presentation of the tumor and its radiological appearance and histological features. We also highlight the importance of a detailed histopathological examination of the excised specimen. PMID:26989286

  7. Determination of tolerable fatty acids and cholera toxin concentrations using human intestinal epithelial cells and BALB/c mouse macrophages.

    PubMed

    Tamari, Farshad; Tychowski, Joanna; Lorentzen, Laura

    2013-01-01

    The positive role of fatty acids in the prevention and alleviation of non-human and human diseases have been and continue to be extensively documented. These roles include influences on infectious and non-infectious diseases including prevention of inflammation as well as mucosal immunity to infectious diseases. Cholera is an acute intestinal illness caused by the bacterium Vibrio cholerae. It occurs in developing nations and if left untreated, can result in death. While vaccines for cholera exist, they are not always effective and other preventative methods are needed. We set out to determine tolerable concentrations of three fatty acids (oleic, linoleic and linolenic acids) and cholera toxin using mouse BALB/C macrophages and human intestinal epithelial cells, respectively. We solubilized the above fatty acids and used cell proliferation assays to determine the concentration ranges and specific concentrations of the fatty acids that are not detrimental to human intestinal epithelial cell viability. We solubilized cholera toxin and used it in an assay to determine the concentration ranges and specific concentrations of cholera toxin that do not statistically decrease cell viability in BALB/C macrophages. We found the optimum fatty acid concentrations to be between 1-5 ng/μl, and that for cholera toxin to be < 30 ng per treatment. This data may aid future studies that aim to find a protective mucosal role for fatty acids in prevention or alleviation of cholera infections. PMID:23748896

  8. High mobility group box 1-induced epithelial mesenchymal transition in human airway epithelial cells.

    PubMed

    Chen, Yu-Ching; Statt, Sarah; Wu, Reen; Chang, Hao-Teng; Liao, Jiunn-Wang; Wang, Chien-Neng; Shyu, Woei-Cherng; Lee, Chen-Chen

    2016-01-01

    Epithelial-mesenchymal transition (EMT) is implicated in bronchial remodeling and loss of lung function in chronic inflammatory airway diseases. Previous studies showed the involvement of the high mobility group box 1 (HMGB1) protein in the pathology of chronic pulmonary inflammatory diseases. However, the role of HMGB1 in EMT of human airway epithelial cells is still unclear. In this study, we used RNA sequencing to show that HMGB1 treatment regulated EMT-related gene expression in human primary-airway epithelial cells. The top five upregulated genes were SNAI2, FGFBP1, VIM, SPARC (osteonectin), and SERPINE1, while the downregulated genes included OCLN, TJP1 (ZO-1), FZD7, CDH1 (E-cadherin), and LAMA5. We found that HMGB1 induced downregulation of E-cadherin and ZO-1, and upregulation of vimentin mRNA transcription and protein translation in a dose-dependent manner. Additionally, we observed that HMGB1 induced AKT phosphorylation, resulting in GSK3β inactivation, cytoplasmic accumulation, and nuclear translocation of β-catenin to induce EMT in human airway epithelial cells. Treatment with PI3K inhibitor (LY294006) and β-catenin shRNA reversed HMGB1-induced EMT. Moreover, HMGB1 induced expression of receptor for advanced glycation products (RAGE), but not that of Toll-like receptor (TLR) 2 or TLR4, and RAGE shRNA inhibited HMGB1-induced EMT in human airway epithelial cells. In conclusion, we found that HMGB1 induced EMT through RAGE and the PI3K/AKT/GSK3β/β-catenin signaling pathway. PMID:26739898

  9. Salivary epithelial cells: an unassuming target site for gene therapeutics

    PubMed Central

    Perez, Paola; Rowzee, Anne M.; Zheng, Changyu; Adriaansen, Janik; Baum, Bruce J.

    2010-01-01

    Salivary glands are classical exocrine glands whose external secretions result in the production of saliva. However, in addition to the secretion of exocrine proteins, salivary epithelial cells are also capable of secreting proteins internally, into the bloodstream. This brief review examines the potential for using salivary epithelial cells as a target site for in situ gene transfer, with an ultimate goal of producing therapeutic proteins for treating both systemic and upper gastrointestinal tract disorders. The review discusses the protein secretory pathways reported to be present in salivary epithelial cells, the viral gene transfer vectors shown useful for transducing these cells, model transgenic secretory proteins examined, and some clinical conditions that might benefit from such salivary gland gene transfer. PMID:20219693

  10. Comparative analysis of mononuclear cells isolated from mucosal lymphoid follicles of the human ileum and colon

    PubMed Central

    Junker, Y; Bode, H; Wahnschaffe, U; Kroesen, A; Loddenkemper, C; Duchmann, R; Zeitz, M; Ullrich, R

    2009-01-01

    Studies of human mucosal lymphoid follicles are rare and have been limited to children's Peyer's patches, which are visible at endoscopy. We investigated lymphoid follicles in ileum biopsies of 87 patients and surgical colon specimens from 66 cancer patients, and examined phenotype and function of isolated follicular immune cells. Two (0–10) and 12 (0–117) follicles per patient were found in ileum and colon samples respectively (P < 0·001). The number of lymphoid follicles mononuclear cells (LFMC) that could be isolated per patient was higher from colon compared with ileum specimens [725 000 (0–23 Mio) versus 100 000 (0–1·3 Mio), P < 0·001]. T cells were predominant in both LFMC and lamina propria mononuclear cells (LPMC), but B cells were more and plasma cells less frequent in LFMC. T cells from mucosal follicles were more frequently CD4-positive and CD62L-positive, but less frequently CD8-positive, CD103-positive and CD69-positive than lamina propria T cells. LFMC from ileum compared with colon showed no differences in mononuclear cell composition. Anti-CD3/CD28 stimulation induced similar proliferation of LFMC and LPMC from ileum and colon, as well as secretion of high levels of interferon-γ, tumour necrosis factor-α and interleukin (IL)-2, but lower levels of IL-4, IL-6 and IL-10. LFMC from colon secreted more IL-2 than those from ileum. Our study shows that mucosal lymphoid follicles can be identified clearly in adult human colon and yield viable immune cells sufficient for phenotypical and functional analysis. The cellular composition of LFMC from ileum and colon is similar, and both secrete predominantly T helper type 1 cytokines. PMID:19250280

  11. The mucosal immune system: From dentistry to vaccine development

    PubMed Central

    KIYONO, Hiroshi; AZEGAMI, Tatsuhiko

    2015-01-01

    The oral cavity is the beginning of the aero-digestive tract, which is covered by mucosal epithelium continuously under the threat of invasion of pathogens, it is thus protected by the mucosal immune system. In the early phase of our scientific efforts for the demonstration of mucosal immune system, dental science was one of major driving forces due to their foreseeability to use oral immunity for the control of oral diseases. The mucosal immune system is divided functionally into, but interconnected inductive and effector sites. Intestinal Peyer’s patches (PPs) are an inductive site containing antigen-sampling M cells and immunocompetent cells required to initiate antigen-specific immune responses. At effector sites, PP-originated antigen-specific IgA B cells become plasma cells to produce polymeric IgA and form secretory IgA by binding to poly-Ig receptor expressed on epithelial cells for protective immunity. The development of new-generation mucosal vaccines, including the rice-based oral vaccine MucoRice, on the basis of the coordinated mucosal immune system is a promising strategy for the control of mucosal infectious diseases. PMID:26460320

  12. Establishment and Characterization of Immortalized Human Amniotic Epithelial Cells

    PubMed Central

    Zhou, Kaixuan; Koike, Chika; Yoshida, Toshiko; Okabe, Motonori; Fathy, Moustafa; Kyo, Satoru; Kiyono, Tohru; Saito, Shigeru

    2013-01-01

    Abstract Human amniotic epithelial cells (HAEs) have a low immunogenic profile and possess potent immunosuppressive properties. HAEs also have several characteristics similar to stem cells, and they are discarded after parturition. Thus, they could potentially be used in cell therapy with fewer ethical problems. HAEs have a short life, so our aim is to establish and characterize immortalized human amniotic epithelial cells (iHAEs). HAEs were introduced with viral oncogenes E6/E7 and with human telomerase reverse transcriptase (hTERT) to create iHAEs. These iHAEs have proliferated around 200 population doublings (PDs) for at least 12 months. High expression of stem cell markers (Oct 3/4, Nanog, Sox2, Klf4) and epithelial markers (CK5, CK18) were detected by immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). These iHAEs were expanded in ultra-low-attachment dishes to form spheroids similarly to epithelial stem/precursor cells. High expression of mesenchymal (CD44, CD73, CD90, CD105) and somatic (CD24, CD29, CD271, Nestin) stem cell markers was detected by flow cytometry. The iHAEs showed adipogenic, osteogenic, neuronal, and cardiac differentiation abilities. In conclusion, the immortalization of HAEs with the characteristics of stem cells has been established, allowing these iHAEs to become useful for cell therapy and regenerative medicine. PMID:23298399

  13. Dendritic cell CD83 homotypic interactions regulate inflammation and promote mucosal homeostasis

    PubMed Central

    Bates, J M; Flanagan, K; Mo, L; Ota, N; Ding, J; Ho, S; Liu, S; Roose-Girma, M; Warming, S; Diehl, L

    2015-01-01

    Dendritic cells (DCs) form an extensive network in the intestinal lamina propria, which orchestrates the mucosal immune response. Alterations in DC function can predispose to inflammatory bowel disease, although by unknown mechanisms. We show that CD83, a highly regulated DC cell surface protein, modulates the immune response to prevent colitis. Mice with a conditional knockout of CD83 in DCs develop exacerbated colitis following dextran sodium sulfate challenge, whereas mucosal overexpression of CD83 inhibits DC inflammatory response and protects against colitis. These CD83 perturbations can be modeled in vitro where we show that CD83 homotypic interaction occurs via cell–cell contact and inhibits pro-inflammatory responses. CD83 knockdown or cytoplasmic truncation abrogates the effects of homotypic binding. We demonstrate that CD83 homotypic interaction regulates DC activation via the mitogen-activated protein kinase pathway by inhibiting p38α phosphorylation. Our findings indicate that CD83 homotypic interactions regulate DC activation and promote mucosal homeostasis. PMID:25204675

  14. Mucosal-associated invariant T cell alterations in obese and type 2 diabetic patients

    PubMed Central

    Magalhaes, Isabelle; Pingris, Karine; Poitou, Christine; Bessoles, Stéphanie; Venteclef, Nicolas; Kiaf, Badr; Beaudoin, Lucie; Da Silva, Jennifer; Allatif, Omran; Rossjohn, Jamie; Kjer-Nielsen, Lars; McCluskey, James; Ledoux, Séverine; Genser, Laurent; Torcivia, Adriana; Soudais, Claire; Lantz, Olivier; Boitard, Christian; Aron-Wisnewsky, Judith; Larger, Etienne; Clément, Karine; Lehuen, Agnès

    2015-01-01

    Obesity and type 2 diabetes (T2D) are associated with low-grade inflammation, activation of immune cells, and alterations of the gut microbiota. Mucosal-associated invariant T (MAIT) cells, which are innate-like T cells that recognize bacterial ligands, are present in blood and enriched in mucosal and inflamed tissues. Here, we analyzed MAIT cells in the blood and adipose tissues of patients with T2D and/or severe obesity. We determined that circulating MAIT cell frequency was dramatically decreased in both patient groups, and this population was even undetectable in some obese patients. Moreover, in both patient groups, circulating MAIT cells displayed an activated phenotype that was associated with elevated Th1 and Th17 cytokine production. In obese patients, MAIT cells were more abundant in adipose tissue than in the blood and exhibited a striking IL-17 profile. Bariatric surgery in obese patients not only improved their metabolic parameters but also increased circulating MAIT cell frequency at 3 months after surgery. Similarly, cytokine production by blood MAIT cells was strongly decreased after surgery. This study reveals profound MAIT cell abnormalities in patients harboring metabolic disorders, suggesting their potential role in these pathologies. PMID:25751065

  15. Gastrointestinal mucosal barrier function and diseases.

    PubMed

    Oshima, Tadayuki; Miwa, Hiroto

    2016-08-01

    The gastrointestinal mucosal barrier plays an essential role in the separation of the inside of the body from the outside environment. Tight junctions (TJs) are the most important component for construction of a constitutive barrier of epithelial cells, and they regulate the permeability of the barrier by tightly sealing the cell-cell junctions. TJ proteins are represented by claudins, occludin, junctional adhesion molecules, and scaffold protein zonula occludens. Among these TJ proteins, claudins are the major components of TJs and are responsible for the barrier and the polarity of the epithelial cells. Gastrointestinal diseases including reflux esophagitis, inflammatory bowel disease, functional gastrointestinal disorders, and cancers may be regulated by these molecules, and disruption of their functions leads to chronic inflammatory conditions and chronic or progressive disease. Therefore, regulation of the barrier function of epithelial cells by regulating the expression and localization of TJ proteins is a potential new target for the treatment of these diseases. Treatment strategies for these diseases might thus be largely altered if symptom generation and/or immune dysfunction could be regulated through improvement of mucosal barrier function. Since TJ proteins may also modify tumor infiltration and metastasis, other important goals include finding a good TJ biomarker of cancer progression and patient prognosis, and developing TJ protein-targeted therapies that can modify patient prognosis. This review summarizes current understanding of gastrointestinal barrier function, TJ protein expression, and the mechanisms underlying epithelial barrier dysregulation in gastrointestinal diseases. PMID:27048502

  16. Diffusion of Immunoglobulin G in Shed Vaginal Epithelial Cells and in Cell-Free Regions of Human Cervicovaginal Mucus

    PubMed Central

    Wang, Ying-Ying; Schroeder, Holly A.; Nunn, Kenetta L.; Woods, Karen; Anderson, Deborah J.; Cone, Richard A.

    2016-01-01

    Human cervicovaginal mucus (CVM) is a viscoelastic gel containing a complex mixture of mucins, shed epithelial cells, microbes and macromolecules, such as antibodies, that together serve as the first line of defense against invading pathogens. Here, to investigate the affinity between IgG and different mucus constituents, we used Fluorescence Recovery After Photobleaching (FRAP) to measure the diffusion of IgG in fresh, minimally modified CVM. We found that CVM exhibits substantial spatial variations that necessitate careful selection of the regions in which to perform FRAP. In portions of CVM devoid of cells, FRAP measurements using different IgG antibodies and labeling methods consistently demonstrate that both exogenous and endogenous IgG undergo rapid diffusion, almost as fast as in saline, in good agreement with the rapid diffusion of IgG in mid-cycle endocervical mucus that is largely devoid of cells. This rapid diffusion indicates the interactions between secreted mucins and IgG must be very weak and transient. IgG also accumulated in cellular debris and shed epithelial cells that had become permeable to IgG, which may allow shed epithelial cells to serve as reservoirs of secreted IgG. Interestingly, in contrast to cell-free regions of CVM, the diffusion of cell-associated IgG was markedly slowed, suggesting greater affinity between IgG and cellular constituents. Our findings contribute to an improved understanding of the role of IgG in mucosal protection against infectious diseases, and may also provide a framework for using FRAP to study molecular interactions in mucus and other complex biological environments. PMID:27362256

  17. Mucosal Mast Cell Count Is Associated With Intestinal Permeability in Patients With Diarrhea Predominant Irritable Bowel Syndrome

    PubMed Central

    Lee, Hyuk; Park, Dong Il; Kim, Hong Joo; Cho, Yong Kyun; Sohn, Chong Il; Jeon, Woo Kyu; Kim, Byung Ik; Chae, Seoung Wan

    2013-01-01

    Background/Aims Although mucosal mast cell tryptase is known to significantly increase intestinal permeability, the relationship between mucosal mast cells and intestinal permeability remains unclear. The objective of this study was to evaluate the correlation among intestinal permeability, tryptase activity and mucosal mast cell count. Methods Rectal biopsies from 16 patients with diarrhea-predominant irritable bowel syndrome (IBS-D) and 7 normal subjects were assessed for tryptase activity and macromolecular permeability using horseradish peroxidase in Ussing chambers. In addition, mucosal mast cell levels were immunohistochemically quantified via image analysis. Results Rectal biopsy of tissues from IBS-D patients showed significantly increased permeability compared with those from normal controls (0.644 ± 0.08 and 0.06 ± 0.00 ng/2 hr/mm2, P < 0.01). Tryptase activity was also substantially higher in rectal biopsy samples from IBS-D patients than those from normal controls (0.86 ± 0.18 and 0.28 ± 0.04 mU/mg protein, P < 0.05). Mucosal mast cell counts were not significantly different between the 2 groups (P > 0.05). However, correlation analysis revealed that only mucosal mast cell count was significantly correlated with intestinal permeability in IBS-D patients (r = 0.558, P < 0.05). Conclusions This study demonstrated a positive correlation between the number of mucosal mast cells and intestinal permeability, suggesting that mucosal mast cells play an important role for increased intestinal permeability in patients with IBS-D. PMID:23667756

  18. Commensal bacteria can enter colonic epithelial cells and induce proinflammatory cytokine secretion: a possible pathogenic mechanism of ulcerative colitis.

    PubMed

    Ohkusa, Toshifumi; Yoshida, Tsutomu; Sato, Nobuhiro; Watanabe, Sumio; Tajiri, Hisao; Okayasu, Isao

    2009-05-01

    Interleukin 2 (IL-2)- and IL-10-knockout mice develop spontaneous colitis under conventional but not germ-free conditions, suggesting that commensal bacteria play an important role in the pathogenesis of colitis. However, interactions between commensal bacteria and colonic epithelial cells have not been fully investigated. We therefore assessed the ability of various commensal bacteria and probiotics to adhere to and invade colonic epithelial cells. Effects of the bacteria on production of proinflammatory cytokines were also measured. Commensal bacteria, including mucosal organisms isolated from ulcerative colitis (UC) patients, such as Fusobacterium varium, reported as a possible pathogen in UC, Bacteroides vulgatus, Escherichia coli and Clostridium clostridioforme, as well as their type strains and probiotics, were assessed for their ability to adhere to and invade colonic epithelial cells using two cell lines, SW-480 and HT-29. Our experiments employed co-incubation, a combination of scanning and transmission electron microscopy and recovery of bacteria from infected-cell lysates. F. varium and several other commensal bacteria, but not probiotics, adhered to colonic epithelial cells and invaded their cytoplasm. ELISA and real-time PCR revealed that the host cells, particularly those invaded by F. varium, showed significant increases in IL-8 and TNF-alpha concentrations in supernatants, with elevation of IL-8, TNF-alpha, MCP-1 and IL-6 mRNAs. Furthermore, IL-8 and TNF-alpha expression and nuclear phosphorylated NF-kappaB p65 expression could be immunohistochemically confirmed in inflamed epithelium with cryptitis or crypt abscess in UC patients. Certain commensal bacteria can invade colonic epithelial cells, activating early intracellular signalling systems to trigger host inflammatory reactions. PMID:19369513

  19. N-acetylcysteine inhibits Na+ absorption across human nasal epithelial cells.

    PubMed

    Rochat, Thierry; Lacroix, Jean-Silvain; Jornot, Lan

    2004-10-01

    N-acetylcysteine (NAC) is a widely used mucolytic drug in patients with a variety of respiratory disorders. The mechanism of action is based on rupture of the disulfide bridges of the high molecular glycoproteins present in the mucus, resulting in smaller subunits of the glycoproteins and reduced viscosity of the mucus. Because Na(+) absorption regulates airway surface liquid volume and thus the efficiency of mucociliary clearance, we asked whether NAC affects the bioelectric properties of human nasal epithelial cells. A 24-h basolateral treatment with 10 mM of NAC decreased the transepithelial potential difference and short-circuit current (I(SC)) by 40%, and reduced the amiloride-sensitive current by 50%, without affecting the transepithelial resistance. After permeabilization of the basolateral membranes of cells with amphotericin B in the presence of a mucosal-to-serosal Na(+) gradient (135:25 mM), NAC inhibited 45% of the amiloride-sensitive current. The Na(+)-K(+)-ATPase pump activity and the basolateral K(+) conductance were not affected by NAC treatment. NAC did not alter total cell mRNA and protein levels of alpha-epithelial Na(+) channel (EnaC) subunit, but reduced abundance of alpha-ENaC subunits in the apical cell membrane as quantified by biotinylation. This effect can be ascribed to the sulphydryl (SH) group of NAC, since N-acetylserine and S-carboxymethyl-l-cysteine were ineffective. Given the importance of epithelial Na(+) channels in controlling the thin layer of fluid that covers the surface of the airways, the increase in the fluidity of the airway mucus following NAC treatment in vivo might be in part related to downregulation of Na(+) absorption and consequently water transport. PMID:15281093

  20. Transcriptional PROFILING OF MUCOCILIARY DIFFERENTIATION IN HUMAN AIRWAY EPITHELIAL CELLS

    EPA Science Inventory

    When cultured at an air-liquid interface (ALI) in the appropriate medium, primary human airway epithelial cells form a polarized, pseudostratified epithelium composed of ciliated and mucus-secreting cells. This culture system provides a useful tool for the in vitro study of...

  1. Lateral adhesion drives reintegration of misplaced cells into epithelial monolayers

    PubMed Central

    St Johnston, Daniel

    2016-01-01

    Cells in simple epithelia orient their mitotic spindles in the plane of the epithelium so that both daughter cells are born within the epithelial sheet. This is assumed to be important to maintain epithelial integrity and prevent hyperplasia, because misaligned divisions give rise to cells outside the epithelium1,2. Here we test this assumption in three types of Drosophila epithelia; the cuboidal follicle epithelium, the columnar early embryonic ectoderm, and the pseudostratified neuroepithelium. Ectopic expression of Inscuteable in these tissues reorients mitotic spindles, resulting in one daughter cell being born outside of the epithelial layer. Live imaging reveals that these misplaced cells reintegrate into the tissue. Reducing the levels of the lateral homophilic adhesion molecules Neuroglian or Fasciclin 2 disrupts reintegration, giving rise to extra-epithelial cells, whereas disruption of adherens junctions has no effect. Thus, the reinsertion of misplaced cells appears to be driven by lateral adhesion, which pulls cells born outside the epithelia layer back into it. Our findings reveal a robust mechanism that protects epithelia against the consequences of misoriented divisions. PMID:26414404

  2. Epithelial stem cells and implications for wound repair.

    PubMed

    Plikus, Maksim V; Gay, Denise L; Treffeisen, Elsa; Wang, Anne; Supapannachart, Rarinthip June; Cotsarelis, George

    2012-12-01

    Activation of epithelial stem cells and efficient recruitment of their proliferating progeny plays a critical role in cutaneous wound healing. The reepithelialized wound epidermis has a mosaic composition consisting of progeny that can be traced back both to epidermal and several types of hair follicle stem cells. The contribution of hair follicle stem cells to wound epidermis is particularly intriguing as it involves lineage identity change from follicular to epidermal. Studies from our laboratory show that hair follicle-fated bulge stem cells commit only transient amplifying epidermal progeny that participate in the initial wound re-epithelialization, but eventually are outcompeted by other epidermal clones and largely disappear after a few months. Conversely, recently described stem cell populations residing in the isthmus portion of hair follicle contribute long-lasting progeny toward wound epidermis and, arguably, give rise to new interfollicular epidermal stem cells. The role of epithelial stem cells during wound healing is not limited to regenerating stratified epidermis. By studying regenerative response in large cutaneous wounds, our laboratory uncovered that epithelial cells in the center of the wound can acquire greater morphogenetic plasticity and, together with the underlying wound dermis, can engage in an embryonic-like process of hair follicle neogenesis. Future studies should uncover the cellular and signaling basis of this remarkable adult wound regeneration phenomenon. PMID:23085626

  3. Epithelial Stem Cells and Implications for Wound Repair

    PubMed Central

    Plikus, Maksim V.; Gay, Denise L.; Treffeisen, Elsa; Wang, Anne; Supapannachart, Rarinthip June; Cotsarelis, George

    2012-01-01

    Activation of epithelial stem cells and efficient recruitment of their proliferating progeny plays a critical role in cutaneous wound healing. The reepithelialized wound epidermis hasa mosaic composition consisting of progeny that can be traced back both to epidermal and several types of hair follicle stem cells. The contribution of hair follicle stem cells to wound epidermis is particularly intriguing as it involves lineage identity change from follicular to epidermal. Studies from our laboratory show that hair follicle-fated bulge stem cells commit only transient amplifying epidermal progeny that participate in the initial wound re-epithelialization, but eventually are outcompeted by other epidermal clones and largely disappear after a few months. Conversely, recently described stem cell populations residing in the isthmus portion of hair follicle contribute long-lasting progeny toward wound epidermis and, arguably, give rise to new inter-follicular epidermal stem cells. The role of epithelial stem cells during wound healing is not limited to regenerating stratified epidermis. By studying regenerative response in large cutaneous wounds, our laboratory uncovered that epithelial cells in the center of the wound can acquire greater morphogenetic plasticity and, together with the underlying wound dermis, can engage in an embryonic-like process of hair follicle neogenesis. Future studies should uncover cellular and signaling basis of this remarkable adult wound regeneration phenomenon. PMID:23085626

  4. AN IN VITRO MODEL FOR MURINE URETERIC EPITHELIAL CELLS

    EPA Science Inventory

    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...

  5. [Isolation, purification and identification of epithelial cells derived from fetal islet-like cell clusters].

    PubMed

    Qiao, Hai; Zhao, Ting; Wang, Yun; Yang, Chun-Rong; Xiao, Mei; Dou, Zhong-Ying

    2007-03-01

    The aim of this article is to provide methods for the isolation and identification of pancreatic stem cells and cell source for research and therapy of diabetes. ICCs were isolated by collagenase IV digesting and then cultured; epithelial cells were purified from monolayer cultured ICCs. The growth curve of the epithelial cells was measured by MTT. The expression of molecular markers in the cells was identified by immunohistochemical staining. The surface markers in the epithelial cells were analyzed by FACS. Epithelial cells were purified from isolated human fetal ICCs and passaged 40 times, and 10(6) - 10(8) cells were cryopreservated per passage. The growth curve demonstrated that the epithelial cells proliferated rapidly. The epithelial cells expressed PDX-1, PCNA, CK-7, CK-19, Nestin, Glut2, and Vimentin, but Insulin was undetected. The cells expressed CD29, CD44, and CD166, but did not express CD11a, CD14, CD34, CD45, CD90, CD105, and CD117. Taken together, these results indicate that self-renewable epithelial cells can be isolated and purified from human fetal pancreas. These also show that the epithelial cells originate from ducts and have the characteristics of pancreatic stem cells. PMID:17460896

  6. The syncytial nature of epithelial cells in the thymic cortex.

    PubMed Central

    Kendall, M D

    1986-01-01

    The epithelial cells of the cortex of human and rodent thymus glands were examined by light and electron microscopy, and the intracellular membrane potentials measured from the subcapsular, cortical and medullary regions. In the human thymus cortex, there is a highly correlated age-independent relationship (r = 0.78) between the distance in micron from one adjacent Type 2/3 epithelial nucleus to another, and the number of thymocytes between them. In rodent glands that had undergone some degree of involution due to hypoxia simulating an altitude of 17 000 feet or following the injection of phenylhydrazine, Type 2/3 epithelial cells were often found to be bi- or multinucleated. Electrophysiological studies of 10 mouse thymus lobes using 0.2 micron tipped electrodes showed that there were highly significant differences (P less than 0.0001) between the intracellular membrane potentials of the subcapsular zone, the cortex and the medulla. When dyes were injected intracellularly (through 0.5 micron tipped electrodes) into individual epithelial cells, methylene blue remained within the cytoplasm, but procion yellow passed in 30 minutes into the nuclei of all the epithelial cells of the cortex but not those of the subcapsular zone, nor the medulla. This indicates that the cortex must be a functional syncytium and it differs in this respect from the rest of the gland. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 Fig. 9 Fig. 10 PMID:3319999

  7. Stimulation of Mucosal Mast Cell Growth in Normal and Nude Rat Bone Marrow Cultures

    NASA Astrophysics Data System (ADS)

    Haig, David M.; McMenamin, Christine; Gunneberg, Christian; Woodbury, Richard; Jarrett, Ellen E. E.

    1983-07-01

    Mast cells with the morphological and biochemical properties of mucosal mast cells (MMC) appear and proliferate to form the predominant cell type in rat bone marrow cultures stimulated with factors from antigen- or mitogen-activated lymphocytes. Conditioned media causing a selective proliferation of MMC were derived from mesenteric lymph node cells of Nippostrongylus brasiliensis-infected rats restimulated in vitro with specific antigen or from normal or infected rat mesenteric lymph node cells stimulated with concanavalin A. MMC growth factor is not produced by T-cell-depleted mesenteric lymph node cells or by the mesenteric lymph node cells of athymic rats. By contrast, MMC precursors are present in the bone marrow of athymic rats and are normally receptive to the growth factor produced by the lymphocytes of thymus-intact rats. The thymus dependence of MMC hyperplasia is thus based on the requirement of a thymus-independent precursor for a T-cell-derived growth promoter.

  8. Protrusive Activity Guides Changes in Cell-Cell Tension during Epithelial Cell Scattering

    PubMed Central

    Maruthamuthu, Venkat; Gardel, Margaret L.

    2014-01-01

    Knowing how epithelial cells regulate cell-matrix and cell-cell adhesions is essential to understand key events in morphogenesis as well as pathological events such as metastasis. During epithelial cell scattering, epithelial cell islands rupture their cell-cell contacts and migrate away as single cells on the extracellular matrix (ECM) within hours of growth factor stimulation, even as adhesion molecules such as E-cadherin are present at the cell-cell contact. How the stability of cell-cell contacts is modulated to effect such morphological transitions is still unclear. Here, we report that in the absence of ECM, E-cadherin adhesions continue to sustain substantial cell-generated forces upon hepatocyte growth factor (HGF) stimulation, consistent with undiminished adhesion strength. In the presence of focal adhesions, constraints that preclude the spreading and movement of cells at free island edges also prevent HGF-mediated contact rupture. To explore the role of cell motion and cell-cell contact rupture, we examine the biophysical changes that occur during the scattering of cell pairs. We show that the direction of cell movement with respect to the cell-cell contact is correlated with changes in the average intercellular force as well as the initial direction of cell-cell contact rupture. Our results suggest an important role for protrusive activity resulting in cell displacement and force redistribution in guiding cell-cell contact rupture during scattering. PMID:25099795

  9. Evaluating alternative stem cell hypotheses for adult corneal epithelial maintenance

    PubMed Central

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

    2015-01-01

    In this review we evaluate evidence for three different hypotheses that explain how the corneal epithelium is maintained. The limbal epithelial stem cell (LESC) hypothesis is most widely accepted. This proposes that stem cells in the basal layer of the limbal epithelium, at the periphery of the cornea, maintain themselves and also produce transient (or transit) amplifying cells (TACs). TACs then move centripetally to the centre of the cornea in the basal layer of the corneal epithelium and also replenish cells in the overlying suprabasal layers. The LESCs maintain the corneal epithelium during normal homeostasis and become more active to repair significant wounds. Second, the corneal epithelial stem cell (CESC) hypothesis postulates that, during normal homeostasis, stem cells distributed throughout the basal corneal epithelium, maintain the tissue. According to this hypothesis, LESCs are present in the limbus but are only active during wound healing. We also consider a third possibility, that the corneal epithelium is maintained during normal homeostasis by proliferation of basal corneal epithelial cells without any input from stem cells. After reviewing the published evidence, we conclude that the LESC and CESC hypotheses are consistent with more of the evidence than the third hypothesis, so we do not consider this further. The LESC and CESC hypotheses each have difficulty accounting for one main type of evidence so we evaluate the two key lines of evidence that discriminate between them. Finally, we discuss how lineage-tracing experiments have begun to resolve the debate in favour of the LESC hypothesis. Nevertheless, it also seems likely that some basal corneal epithelial cells can act as long-term progenitors if limbal stem cell function is compromised. Thus, this aspect of the CESC hypothesis may have a lasting impact on our understanding of corneal epithelial maintenance, even if it is eventually shown that stem cells are restricted to the limbus as proposed

  10. Commensal Bacteria Modulate Innate Immune Responses of Vaginal Epithelial Cell Multilayer Cultures

    PubMed Central

    Rose, William A.; McGowin, Chris L.; Spagnuolo, Rae Ann; Eaves-Pyles, Tonyia D.; Popov, Vsevolod L.; Pyles, Richard B.

    2012-01-01

    The human vaginal microbiome plays a critical but poorly defined role in reproductive health. Vaginal microbiome alterations are associated with increased susceptibility to sexually-transmitted infections (STI) possibly due to related changes in innate defense responses from epithelial cells. Study of the impact of commensal bacteria on the vaginal mucosal surface has been hindered by current vaginal epithelial cell (VEC) culture systems that lack an appropriate interface between the apical surface of stratified squamous epithelium and the air-filled vaginal lumen. Therefore we developed a reproducible multilayer VEC culture system with an apical (luminal) air-interface that supported colonization with selected commensal bacteria. Multilayer VEC developed tight-junctions and other hallmarks of the vaginal mucosa including predictable proinflammatory cytokine secretion following TLR stimulation. Colonization of multilayers by common vaginal commensals including Lactobacillus crispatus, L. jensenii, and L. rhamnosus led to intimate associations with the VEC exclusively on the apical surface. Vaginal commensals did not trigger cytokine secretion but Staphylococcus epidermidis, a skin commensal, was inflammatory. Lactobacilli reduced cytokine secretion in an isolate-specific fashion following TLR stimulation. This tempering of inflammation offers a potential explanation for increased susceptibility to STI in the absence of common commensals and has implications for testing of potential STI preventatives. PMID:22412914

  11. Lactobacillus acidophilus binds to MUC3 component of cultured intestinal epithelial cells with highest affinity.

    PubMed

    Das, Jugal Kishore; Mahapatra, Rajani Kanta; Patro, Shubhransu; Goswami, Chandan; Suar, Mrutyunjay

    2016-04-01

    Lactobacillus strains have been shown to adhere to the mucosal components of intestinal epithelial cells. However, established in vitro adhesion assays have several drawbacks in assessing the adhesion of new Lactobacillus strains. The present study aimed to compare the adhesion of four different Lactobacillus strains and select the most adherent microbe, based on in silico approach supported by in vitro results. The mucus-binding proteins in Lactobacillus acidophilus, L. plantarum, L. brevis and L. fermentum were identified and their capacities to interact with intestinal mucin were compared by molecular docking analysis. Lactobacillus acidophilus had the maximal affinity of binding to mucin with predicted free energy of -6.066 kcal mol(-1) Further, in vitro experimental assay of adhesion was performed to validate the in silico results. The adhesion of L. acidophilus to mucous secreting colon epithelial HT-29 MTX cells was highest at 12%, and it formed biofilm with maximum depth (Z = 84 μm). Lactobacillus acidophilus was determined to be the most adherent strain in the study. All the Lactobacillus strains tested in this study, displayed maximum affinity of binding to MUC3 component of mucus as compared to other gastrointestinal mucins. These findings may have importance in the design of probiotics and health care management. PMID:26946538

  12. [Epithelial cell in intestinal homeostasis and inflammatory bowel diseases].

    PubMed

    Zouiten-Mekki, Lilia; Serghini, Meriem; Fekih, Monia; Kallel, Lamia; Matri, Samira; Ben Mustapha, Nadia; Boubaker, Jalel; Filali, Azza

    2013-12-01

    Crohn's disease (CD) and ulcerative colitis (UC) are the principal inflammatory bowel diseases (IBD) which physiopathology is currently poorly elucidated. During these diseases, the participation of the epithelial cell in the installation and the perpetuation of the intestinal inflammation is now clearly implicated. In fact, the intestinal epithelium located at the interface between the internal environment and the intestinal luminal, is key to the homeostatic regulation of the intestinal barrier. This barrier can schematically be regarded as being three barriers in one: a physical, chemical and immune barrier. The barrier function of epithelial cell can be altered by various mechanisms as occurs in IBD. The goal of this article is to review the literature on the role of the epithelial cell in intestinal homeostasis and its implication in the IBD. PMID:24356146

  13. Infrared micro-spectroscopic studies of epithelial cells

    PubMed Central

    Romeo, Melissa; Mohlenhoff, Brian; Jennings, Michael; Diem, Max

    2009-01-01

    We report results from a study of human and canine mucosal cells, investigated by infrared micro-spectroscopy, and analyzed by methods of multivariate statistics. We demonstrate that the infrared spectra of individual cells are sensitive to the stage of maturation, and that a distinction between healthy and diseased cells will be possible. Since this report is written for an audience not familiar with infrared micro-spectroscopy, a short introduction into this field is presented along with a summary of principal component analysis. PMID:16797481

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

    PubMed Central

    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

  15. Reprogramming tumor-infiltrating dendritic cells for CD103+CD8+ mucosal T cell differentiation and breast cancer rejection

    PubMed Central

    Wu, Te-Chia; Xu, Kangling; Banchereau, Romain; Marches, Florentina; Yu, Chun I; Martinek, Jan; Anguiano, Esperanza; Pedroza-Gonzalez, Alexander; Snipes, G. Jackson; O’Shaughnessy, Joyce; Nishimura, Stephen; Liu, Yong-Jun; Pascual, Virginia; Banchereau, Jacques; Oh, Sangkon; Palucka, Karolina

    2014-01-01

    Our studies showed that tumor-infiltrating dendritic cells (DC) in breast cancer drive inflammatory T helper 2 (iTh2) cells and protumor inflammation. Here we show that intratumoral delivery of the β-glucan curdlan, a ligand of dectin-1, blocks the generation of iTh2 cells, and prevents breast cancer progression in vivo. Curdlan reprograms tumor-infiltrating DC via the ligation of dectin-1, enabling the DC to become resistant to cancer-derived thymic stromal lymphopoietin (TSLP), to produce IL12p70, and to favor the generation of T helper 1 (Th1) cells. DC activated via dectin-1, but not those activated with TLR-7/8 ligand or poly IC, induce CD8+ T cells to express CD103 (αE integrin), a ligand for cancer cells E-cadherin. Generation of these mucosal CD8+ T cells is regulated by DC-derived integrin αvβ8 and TGF-β activation in a dectin-1-dependent fashion. These CD103+CD8+ mucosal T cells accumulate in the tumors thereby increasing cancer necrosis and inhibiting cancer progression in vivo in a humanized mouse model of breast cancer. Importantly, CD103+CD8+ mucosal T cells elicited by reprogrammed DC can reject established cancer. Thus, reprogramming tumor-infiltrating DC represents a new strategy for cancer rejection. PMID:24795361

  16. Interleukin-8 response of gastric epithelial cell lines to Helicobacter pylori stimulation in vitro.

    PubMed Central

    Sharma, S A; Tummuru, M K; Miller, G G; Blaser, M J

    1995-01-01

    Gastric infection with Helicobacter pylori activates a mucosal inflammatory response by mononuclear cells and neutrophils that includes expression of cytokines interleukin-1 beta (IL-1 beta), IL-6, tumor necrosis factor alpha, and IL-8. In this study, we analyzed the IL-8 response of human gastric cancer cell lines (Kato III, AGS, and MKN28) to H. pylori infection in vitro. IL-8 mRNA expression was detected by reverse transcription-PCR amplification of RNA extracted from epithelial cells after incubation with different H. pylori wild-type and mutant strains, and IL-8 secretion was measured by an enzyme-linked immunosorbent assay. Exposure to viable H. pylori induced IL-8 mRNA and protein synthesis in all three gastric cell lines but not in nongastric epithelial cell lines. Heat-killed H. pylori and a crude cytotoxin preparation did not induce significant IL-8 secretion. IL-8 mRNA peaked between 2 and 4 h postinfection, and IL-8 protein production was maximal 24 h postinfection. Exposure of gastric carcinoma cells to other gastrointestinal bacteria, such as Pseudomonas aeruginosa, Campylobacter jejuni, and Escherichia coli, but not Campylobacter fetus, induced IL-8 synthesis. Wild-type strains that expressed the vacuolating cytotoxin (Tox+) and a cytotoxin-associated gene (cagA) product (CagA+) induced significantly more IL-8 than did CagA- Tox- strains. However, there was no decrease in IL-8 induction by isogenic mutants of CagA-, Tox-, or Cag- Tox- strains or by a mutant lacking the urease subunits. These results indicate that exposure to H. pylori and other gram-negative organisms that do not colonize the gastric mucosa induces IL-8 production by gastric carcinoma cells in vitro. Although the CagA+ Tox+ phenotype of H. pylori is associated with enhanced IL-8 production by gastric cell lines, other bacterial constituents are clearly essential. PMID:7729872

  17. Microbial exposure alters HIV-1-induced mucosal CD4+ T cell death pathways Ex vivo

    PubMed Central

    2014-01-01

    Background Early HIV-1 infection causes massive CD4+ T cell death in the gut and translocation of bacteria into the circulation. However, the programmed cell death (PCD) pathways used by HIV-1 to kill CD4+ T cells in the gut, and the impact of microbial exposure on T cell loss, remain unclear. Understanding mucosal HIV-1 triggered PCD could be advanced by an ex vivo system involving lamina propria mononuclear cells (LPMCs). We therefore modeled the interactions of gut LPMCs, CCR5-tropic HIV-1 and a commensal gut bacterial species, Escherichia coli. In this Lamina Propria Aggregate Culture (LPAC) model, LPMCs were infected with HIV-1BaL by spinoculation and cultured in the presence or absence of heat killed E.coli. CD4+ T cell numbers derived from flow cytometry and viable cell counts were reported relative to mock infection. Viable cells were identified by viability dye exclusion (AqVi), and intracellular HIV-1 Gag p24 protein was used to identify infected cells. Annexin V and AqVi were used to identify apoptotic versus necrotic cells. Caspase-1 and Caspase-3 activities were blocked using specific inhibitors YVAD and DEVD, respectively. Results CD4+ T cell depletion following HIV-1 infection was reproducibly observed by 6 days post infection (dpi). Depletion at 6 dpi strongly correlated with infection frequency at 4 dpi, was significantly blocked by Efavirenz treatment, and was primarily driven by p24-negative cells that were predominantly necrotic. HIV-1 infection significantly induced CD4+ T-cell intrinsic Caspase-1 activity, whereas Caspase-1 inhibition, but not Caspase-3 inhibition, significantly blocked CD4+ T cell depletion. Exposure to E.coli enhanced HIV-1 infection and CD4+ T depletion, and significantly increased the number of apoptotic p24+ cells. Notably, CD4+ T cell depletion in the presence of E.coli was partially blocked by Caspase-3, but not by Caspase-1 inhibition. Conclusions In the LPAC model, HIV-1 induced Caspase-1 mediated pyroptosis in

  18. Epithelial cell cultures from normal and cancerous human tissues.

    PubMed

    Owens, R B; Smith, H S; Nelson-Rees, W A; Springer, E L

    1976-04-01

    Thirty epithelial cell strains were isolated from human carcinomas and normal epithelial tissues by collagenase digestion and selective removal of fibroblasts with trypsin-Versene. Most strains were obtained from metastatic carcinomas or epithelia of the urinary and intestinal tracts. The success rate for growth of both neoplastic and normal tissues (excluding skin) was 38%. Six of these strains showed gross morphologic and chromosome changes typical of malignant cells. Nine resembled normal epithelium. The other 15 exhibited some degree of morphologic change from normal. PMID:176412

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

    PubMed

    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. PMID:27189858

  20. Immediate early responses of avian tracheal epithelial cells to infection with highly pathogenic avian invluenza virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Highly pathogenic (HP) avian influenza viruses (AIV) present an ongoing threat to the world poultry industry. In order to develop new AIV control strategies it is necessary to understand the underlying mechanism of viral infection at mucosal respiratory sites. Chicken and duck tracheal epithelial ...

  1. Expression of the FGFR2 mesenchymal splicing variant in epithelial cells drives epithelial-mesenchymal transition

    PubMed Central

    Ranieri, Danilo; Rosato, Benedetta; Nanni, Monica; Magenta, Alessandra

    2016-01-01

    The FGFRs are receptor tyrosine kinases expressed by tissue-specific alternative splicing in epithelial IIIb or mesenchymal IIIc isoforms. Deregulation of FGF/FGFR signaling unbalances the epithelial-stromal homeostasis and may lead to cancer development. In the epithelial-context, while FGFR2b/KGFR acts as tumor suppressor, FGFR2c appears to play an oncogenic role. Based on our recent observation that the switching of FGFR2b versus FGFR2c induces EMT, here we investigated the biological outcome of the ectopic expression of FGFR2c in normal human keratinocytes. Morphological analysis showed that, differently from FGFR2b overexpression, the forced expression and activation of FGFR2c drive the epithelial cells to acquire a mesenchymal-like shape and actin reorganization. Moreover, the appearance of invasiveness and anchorage-independent growth ability in FGFR2c transfected keratinocytes was consistent with the potential tumorigenic role proposed for this receptor variant. Biochemical and molecular approaches revealed that the observed phenotypic changes were accompanied by modulation of EMT biomarkers and indicated the involvement of EMT transcription factors and miRs. Finally, the analysis of the expression pattern of discriminating markers strongly suggested that activation of FGFR2c triggers a process corresponding to the initiation of the pathological type III EMT, but not to the more physiological type II EMT occurring during FGFR2b-mediated wound healing. PMID:26713601

  2. Sp3 regulates fas expression in lung epithelial cells.

    PubMed Central

    Pang, H; Miranda, K; Fine, A

    1998-01-01

    By transducing an apoptotic signal in immune effector cells, Fas has been directly implicated in the control of immunological activity. Expression and functional results, however, have also suggested a role for Fas in regulating cell turnover in specific epithelial populations. To characterize factors responsible for Fas expression in epithelial cells, approximately 3 kb of the 5' flanking region of the mouse Fas gene was isolated. By rapid amplification of cDNA ends and primer extension, transcriptional start sites were identified within 50 bp upstream of the translation start site. Transient transfection of promoter-luciferase constructs in a mouse lung epithelial cell line, MLE-15, localized promoter activity to the first 77 bp of upstream sequence. By using a 60 bp DNA probe (-18 to -77) in electrophoretic mobility-shift assays, three shifted complexes were found. Incubation with excess cold Sp1 oligonucleotide or an anti-Sp3 antibody inhibited complex formation. Site-directed mutagenesis of the Sp1 site resulted in 60-70% loss of promoter activity. In Drosophila SL-2 cells, promoter activity was markedly increased by co-transfection of an Sp3 expression construct. These results show that the Sp3 protein is involved in regulating Fas gene expression in lung epithelial cells. PMID:9639581

  3. Gut Mucosal FOXP3+ Regulatory CD4+ T Cells and Nonregulatory CD4+ T Cells Are Differentially Affected by Simian Immunodeficiency Virus Infection in Rhesus Macaques▿

    PubMed Central

    Allers, Kristina; Loddenkemper, Christoph; Hofmann, Jörg; Unbehaun, Anett; Kunkel, Désirée; Moos, Verena; Kaup, Franz-Josef; Stahl-Hennig, Christiane; Sauermann, Ulrike; Epple, Hans-Jörg; Schneider, Thomas

    2010-01-01

    The gastrointestinal tract represents a major site for human and simian immunodeficiency virus (HIV and SIV) replication and CD4+ T-cell depletion. Despite severe depletion of mucosal CD4+ T cells, FOXP3+ regulatory CD4+ T cells (Treg) are highly increased in the gut mucosa of chronically HIV-infected individuals and may contribute to HIV pathogenesis, either by their immunosuppressive function or as a significant target cell population for virus production. Little is known about the susceptibility of mucosal Treg to viral infection and the longitudinal effect of HIV/SIV infection on Treg dynamics. In this study, we determined the level of SIV infection in Treg and nonregulatory CD4+ T cells (non-Treg) isolated from the colon of SIV-infected rhesus macaques. The dynamics of mucosal Treg and alterations in the mucosal CD4+ T-cell pool were examined longitudinally. Our findings indicate that mucosal Treg were less susceptible to productive SIV infection than non-Treg and thus were selectively spared from SIV-mediated cell death. In addition to improved survival, local expansion of Treg by SIV-induced proliferation of the mucosal CD4+ T-cell pool facilitated the accumulation of mucosal Treg during the course of infection. High frequency of mucosal Treg in chronic SIV infection was strongly related to a reduction of perforin-expressing cells. In conclusion, this study suggests that mucosal Treg are less affected by productive SIV infection than non-Treg and therefore spared from depletion. Although SIV production is limited in mucosal Treg, Treg accumulation may indirectly contribute to viral persistence by suppressing antiviral immune responses. PMID:20071575

  4. Induction of mucosal immunity through systemic immunization: Phantom or reality?

    PubMed

    Su, Fei; Patel, Girishchandra B; Hu, Songhua; Chen, Wangxue

    2016-04-01

    Generation of protective immunity at mucosal surfaces can greatly assist the host defense against pathogens which either cause disease at the mucosal epithelial barriers or enter the host through these surfaces. Although mucosal routes of immunization, such as intranasal and oral, are being intensely explored and appear promising for eliciting protective mucosal immunity in mammals, their application in clinical practice has been limited due to technical and safety related challenges. Most of the currently approved human vaccines are administered via systemic (such as intramuscular and subcutaneous) routes. Whereas these routes are acknowledged as being capable to elicit antigen-specific systemic humoral and cell-mediated immune responses, they are generally perceived as incapable of generating IgA responses or protective mucosal immunity. Nevertheless, currently licensed systemic vaccines do provide effective protection against mucosal pathogens such as influenza viruses and Streptococcus pneumoniae. However, whether systemic immunization induces protective mucosal immunity remains a controversial topic. Here we reviewed the current literature and discussed the potential of systemic routes of immunization for the induction of mucosal immunity. PMID:26752023

  5. A molecular basis underpinning the T cell receptor heterogeneity of mucosal-associated invariant T cells

    PubMed Central

    Eckle, Sidonia B.G.; Birkinshaw, Richard W.; Kostenko, Lyudmila; Corbett, Alexandra J.; McWilliam, Hamish E.G.; Reantragoon, Rangsima; Chen, Zhenjun; Gherardin, Nicholas A.; Beddoe, Travis; Liu, Ligong; Patel, Onisha; Meehan, Bronwyn; Fairlie, David P.; Villadangos, Jose A.; Godfrey, Dale I.

    2014-01-01

    Mucosal-associated invariant T (MAIT) cells express an invariant T cell receptor (TCR) α-chain (TRAV1-2 joined to TRAJ33, TRAJ20, or TRAJ12 in humans), which pairs with an array of TCR β-chains. MAIT TCRs can bind folate- and riboflavin-based metabolites restricted by the major histocompatibility complex (MHC)-related class I−like molecule, MR1. However, the impact of MAIT TCR and MR1-ligand heterogeneity on MAIT cell biology is unclear. We show how a previously uncharacterized MR1 ligand, acetyl-6-formylpterin (Ac-6-FP), markedly stabilized MR1, potently up-regulated MR1 cell surface expression, and inhibited MAIT cell activation. These enhanced properties of Ac-6-FP were attributable to structural alterations in MR1 that subsequently affected MAIT TCR recognition via conformational changes within the complementarity-determining region (CDR) 3β loop. Analysis of seven TRBV6-1+ MAIT TCRs demonstrated how CDR3β hypervariability impacted on MAIT TCR recognition by altering TCR flexibility and contacts with MR1 and the Ag itself. Ternary structures of TRBV6-1, TRBV6-4, and TRBV20+ MAIT TCRs in complex with MR1 bound to a potent riboflavin-based antigen (Ag) showed how variations in TRBV gene usage exclusively impacted on MR1 contacts within a consensus MAIT TCR-MR1 footprint. Moreover, differential TRAJ gene usage was readily accommodated within a conserved MAIT TCR-MR1-Ag docking mode. Collectively, MAIT TCR heterogeneity can fine-tune MR1 recognition in an Ag-dependent manner, thereby modulating MAIT cell recognition. PMID:25049336

  6. CsrRS and environmental pH regulate group B streptococcus adherence to human epithelial cells and extracellular matrix.

    PubMed

    Park, Su Eun; Jiang, Shengmei; Wessels, Michael R

    2012-11-01

    Streptococcus agalactiae (group B Streptococcus or GBS) is a common colonizer of the gastrointestinal and genital tracts and an important cause of invasive infections in newborn infants and in adults with predisposing chronic conditions or advanced age. Attachment to epithelial surfaces at mucosal sites is a critical step in the successful colonization of a human host, and regulation of this process is likely to play an important role in both commensalism and dissemination to cause invasive disease. We found that inactivation of the CsrRS (or CovRS) two-component system increased GBS adherence to epithelial cells derived from human vaginal, cervical, and respiratory epithelium, as well as increasing adherence to extracellular matrix proteins and increasing biofilm formation on polystyrene. Neutral (as opposed to acidic) pH enhanced GBS binding to vaginal epithelial cells and to fibrinogen and fibronectin, effects that were partially dependent on CsrRS. The regulatory effects of CsrRS and environmental pH on bacterial adherence correlated with their effects on the expression of multiple surface adhesins, as assessed by quantitative reverse transcription-PCR. We conclude that GBS adherence to epithelial and abiotic surfaces is regulated by the CsrRS two-component system and by environmental pH through their regulatory effects on the expression of bacterial surface adhesins. Dynamic regulation of GBS adherence enhances the organism's adaptability to survival in multiple niches in the human host. PMID:22949550

  7. CsrRS and Environmental pH Regulate Group B Streptococcus Adherence to Human Epithelial Cells and Extracellular Matrix

    PubMed Central

    Park, Su Eun; Jiang, Shengmei

    2012-01-01

    Streptococcus agalactiae (group B Streptococcus or GBS) is a common colonizer of the gastrointestinal and genital tracts and an important cause of invasive infections in newborn infants and in adults with predisposing chronic conditions or advanced age. Attachment to epithelial surfaces at mucosal sites is a critical step in the successful colonization of a human host, and regulation of this process is likely to play an important role in both commensalism and dissemination to cause invasive disease. We found that inactivation of the CsrRS (or CovRS) two-component system increased GBS adherence to epithelial cells derived from human vaginal, cervical, and respiratory epithelium, as well as increasing adherence to extracellular matrix proteins and increasing biofilm formation on polystyrene. Neutral (as opposed to acidic) pH enhanced GBS binding to vaginal epithelial cells and to fibrinogen and fibronectin, effects that were partially dependent on CsrRS. The regulatory effects of CsrRS and environmental pH on bacterial adherence correlated with their effects on the expression of multiple surface adhesins, as assessed by quantitative reverse transcription-PCR. We conclude that GBS adherence to epithelial and abiotic surfaces is regulated by the CsrRS two-component system and by environmental pH through their regulatory effects on the expression of bacterial surface adhesins. Dynamic regulation of GBS adherence enhances the organism's adaptability to survival in multiple niches in the human host. PMID:22949550

  8. High mobility group box 1-induced epithelial mesenchymal transition in human airway epithelial cells

    PubMed Central

    Chen, Yu-Ching; Statt, Sarah; Wu, Reen; Chang, Hao-Teng; Liao, Jiunn-Wang; Wang, Chien-Neng; Shyu, Woei-Cherng; Lee, Chen-Chen

    2016-01-01

    Epithelial–mesenchymal transition (EMT) is implicated in bronchial remodeling and loss of lung function in chronic inflammatory airway diseases. Previous studies showed the involvement of the high mobility group box 1 (HMGB1) protein in the pathology of chronic pulmonary inflammatory diseases. However, the role of HMGB1 in EMT of human airway epithelial cells is still unclear. In this study, we used RNA sequencing to show that HMGB1 treatment regulated EMT-related gene expression in human primary-airway epithelial cells. The top five upregulated genes were SNAI2, FGFBP1, VIM, SPARC (osteonectin), and SERPINE1, while the downregulated genes included OCLN, TJP1 (ZO-1), FZD7, CDH1 (E-cadherin), and LAMA5. We found that HMGB1 induced downregulation of E-cadherin and ZO-1, and upregulation of vimentin mRNA transcription and protein translation in a dose-dependent manner. Additionally, we observed that HMGB1 induced AKT phosphorylation, resulting in GSK3β inactivation, cytoplasmic accumulation, and nuclear translocation of β-catenin to induce EMT in human airway epithelial cells. Treatment with PI3K inhibitor (LY294006) and β-catenin shRNA reversed HMGB1-induced EMT. Moreover, HMGB1 induced expression of receptor for advanced glycation products (RAGE), but not that of Toll-like receptor (TLR) 2 or TLR4, and RAGE shRNA inhibited HMGB1-induced EMT in human airway epithelial cells. In conclusion, we found that HMGB1 induced EMT through RAGE and the PI3K/AKT/GSK3β/β-catenin signaling pathway. PMID:26739898

  9. Induction of apoptosis of lymphocytes in rat mucosal immune system

    PubMed Central

    Chen, Xue-Qing; Zhang, Wan-Dai; Song, Yu-Gang; Zhou, Dian-Yuan

    1998-01-01

    AIM: To undergo apoptosis during negative and positive selection processes in rat mucosal immune system which are implicated in the pathogenesis of various mucosal diseases. METHODS: Female Sprague-Dawley rats were given protein synthesis inhibitor, cycloheximide, intravenously or intraperitoneally, an apoptosis was recognized by morphological hallmark under light and electronmicroscopy, and the expression of proliferating cell nuclear antigen was visualized immunohistochemically. RESULTS: The apoptosis of mucosal lymphocytes in the digestive tract, as well as in trachea, uterus and lacrimal gland was induced by cycloheximide ( > 1.0 mg·kg-1 body weight), which were located mainly in lamina propria and germinal centers of lymphoid nodules. At the same time, a portion of crypt epithelial cells of proliferating zone in small and large intestine, and the epithelial cells in genital tract were also found to undergo apoptosis. Immunostainings showed that apoptotic cells expressed proliferating cell nuclear antigen. CONCLUSION: Apoptosis of lymphocytes in mucosal immune system can be induced by cycloheximide. This model will facilitate the understanding of normal mucosal immune system and its role in the pathogenesis of related diseases such as inflammatory bowel diseases. PMID:11819221

  10. Mucosal Immune Development in Early Life: Setting the Stage.

    PubMed

    Brugman, Sylvia; Perdijk, Olaf; van Neerven, R J Joost; Savelkoul, Huub F J

    2015-08-01

    Our environment poses a constant threat to our health. To survive, all organisms must be able to discriminate between good (food ingredients and microbes that help digest our food) and bad (pathogenic microbes, viruses and toxins). In vertebrates, discrimination between beneficial and harmful antigens mainly occurs at the mucosal surfaces of the respiratory, digestive, urinary and genital tract. Here, an extensive network of cells and organs form the basis of what we have come to know as the mucosal immune system. The mucosal immune system is composed of a single epithelial cell layer protected by a mucus layer. Different immune cells monitor the baso-lateral side of the epithelial cells and dispersed secondary lymphoid organs, such as Peyer's patches and isolated lymphoid follicles are equipped with immune cells able to mount appropriate and specific responses. This review will focus on the current knowledge on host, dietary and bacterial-derived factors that shape the mucosal immune system before and after birth. We will discuss current knowledge on fetal immunity (both responsiveness and lymphoid organ development) as well as the impact of diet and microbial colonization on neonatal immunity and disease susceptibility. Lastly, inflammatory bowel disease will be discussed as an example of how the composition of the microbiota might predispose to disease later in life. A fundamental understanding of the mechanisms involved in mucosal immune development and tolerance will aid nutritional intervention strategies to improve health in neonatal and adult life. PMID:25666708

  11. Perturbations of mucosal homeostasis through interactions of intestinal microbes with myeloid cells.

    PubMed

    Schey, Regina; Danzer, Claudia; Mattner, Jochen

    2015-02-01

    Mucosal surfaces represent the largest areas of interactions of the host with its environment. Subsequently, the mucosal immune system has evolved complex strategies to maintain the integrity of the host by inducing protective immune responses against pathogenic and tolerance against dietary and commensal microbial antigens within the broad range of molecules the intestinal epithelium is exposed to. Among many other specialized cell subsets, myeloid cell populations - due to their strategic location in the subepithelial lamina propria - are the first ones to scavenge and process these intestinal antigens and to send consecutive signals to other immune and non-immune cell subsets. Thus, myeloid cell populations represent attractive targets for clinical intervention in chronic inflammatory bowel diseases (IBDs) such as ulcerative colitis (UC) and Crohn's disease (CD) as they initiate and modulate inflammatory or regulatory immune response and shape the intestinal T cell pool. Here, we discuss the interactions of the intestinal microbiota with dendritic cell and macrophage populations and review in this context the literature on four promising candidate molecules that are critical for the induction and maintenance of intestinal homeostasis on the one hand, but also for the initiation and propagation of chronic intestinal inflammation on the other. PMID:25466587

  12. Experiment K-7-17: Effects of Spaceflight on the Proliferation of Jejunal Mucosal Cells

    NASA Technical Reports Server (NTRS)

    Phillips, R. W.; Moeller, C. L.; Sawyer, H. R.; Smirnov, K. L.

    1994-01-01

    The purpose of this project was to test the hypothesis that the generalized, whole body decrease in synthetic activity due to microgravity conditions encountered during spaceflight would be demonstrable in cells and tissues characterized by a rapid rate of turnover. Jejunal mucosal cells were chosen as a model since these cells are among the most rapidly proliferating in the body. Accordingly, the percentage of mitotic cells present in the crypts of Lieberkuhn in each of 5 rats flown on the COSMOS 2044 mission were compared to the percentage of mitotic cells present in the crypts in rats included in each of 3 ground control groups (i.e., vivarium, synchronous and caudal-elevated). No significant difference (p greater than .05) was detected in mitotic indices between the flight and vivarium group. Although the ability of jejunal mucosal cells to divide by mitosis was not impaired in flight group, there was, however, a reduction in the length of villi and depth of crypts. The concommitant reduction in villus length and crypth depth in the flight group probably reflects changes in connective tissue components within the core of villi.

  13. Perturbations of mucosal homeostasis through interactions of intestinal microbes with myeloid cells

    PubMed Central

    Schey, Regina; Danzer, Claudia; Mattner, Jochen

    2014-01-01

    Mucosal surfaces represent the largest areas of interactions of the host with its environment. Subsequently, the mucosal immune system has evolved complex strategies to maintain the integrity of the host by inducing protective immune responses against pathogenic and tolerance against dietary and commensal microbial antigens within the broad range of molecules the intestinal epithelium is exposed to. Among many other specialized cell subsets, myeloid cell populations - due to their strategic location in the subepithelial lamina propria - are the first ones to scavenge and process these intestinal antigens and to send consecutive signals to other immune and non-immune cell subsets. Thus, myeloid cell populations represent attractive targets for clinical intervention in chronic inflammatory bowel diseases (IBDs) such as ulcerative colitis (UC) and Crohn's disease (CD) as they initiate and modulate inflammatory or regulatory immune response and shape the intestinal T cell pool. Here, we discuss the interactions of the intestinal microbiota with dendritic cell and macrophage populations and review in this context the literature on four promising candidate molecules that are critical for the induction and maintenance of intestinal homeostasis on the one hand, but also for the initiation and propagation of chronic intestinal inflammation on the other. PMID:25466587

  14. Airway epithelial cell response to human metapneumovirus infection

    SciTech Connect

    Bao, X.; Liu, T.; Spetch, L.; Kolli, D.; Garofalo, R.P.; Casola, A.

    2007-11-10

    Human metapneumovirus (hMPV) is a major cause of lower respiratory tract infections (LRTIs) in infants, elderly and immunocompromised patients. In this study, we show that hMPV can infect in a similar manner epithelial cells representative of different tracts of the airways. hMPV-induced expression of chemokines IL-8 and RANTES in primary small alveolar epithelial cells (SAE) and in a human alveolar type II-like epithelial cell line (A549) was similar, suggesting that A549 cells can be used as a model to study lower airway epithelial cell responses to hMPV infection. A549 secreted a variety of CXC and CC chemokines, cytokines and type I interferons, following hMPV infection. hMPV was also a strong inducer of transcription factors belonging to nuclear factor (NF)-{kappa}B, interferon regulatory factors (IRFs) and signal transducers and activators of transcription (STATs) families, which are known to orchestrate the expression of inflammatory and immunomodulatory mediators.

  15. Cigarette smoke extract affects mitochondrial function in alveolar epithelial cells.

    PubMed

    Ballweg, Korbinian; Mutze, Kathrin; Königshoff, Melanie; Eickelberg, Oliver; Meiners, Silke

    2014-12-01

    Cigarette smoke is the main risk factor for chronic obstructive pulmonary disease (COPD). Exposure of cells to cigarette smoke induces an initial adaptive cellular stress response involving increased oxidative stress and induction of inflammatory signaling pathways. Exposure of mitochondria to cellular stress alters their fusion/fission dynamics. Whereas mild stress induces a prosurvival response termed stress-induced mitochondrial hyperfusion, severe stress results in mitochondrial fragmentation and mitophagy. In the present study, we analyzed the mitochondrial response to mild and nontoxic doses of cigarette smoke extract (CSE) in alveolar epithelial cells. We characterized mitochondrial morphology, expression of mitochondrial fusion and fission genes, markers of mitochondrial proteostasis, as well as mitochondrial functions such as membrane potential and oxygen consumption. Murine lung epithelial (MLE)12 and primary mouse alveolar epithelial cells revealed pronounced mitochondrial hyperfusion upon treatment with CSE, accompanied by increased expression of the mitochondrial fusion protein mitofusin 2 and increased metabolic activity. We did not observe any alterations in mitochondrial proteostasis, i.e., induction of the mitochondrial unfolded protein response or mitophagy. Therefore, our data indicate an adaptive prosurvival response of mitochondria of alveolar epithelial cells to nontoxic concentrations of CSE. A hyperfused mitochondrial network, however, renders the cell more vulnerable to additional stress, such as sustained cigarette smoke exposure. As such, cigarette smoke-induced mitochondrial hyperfusion, although part of a beneficial adaptive stress response in the first place, may contribute to the pathogenesis of COPD. PMID:25326581

  16. AIRWAY EPITHELIAL CELL RESPONSE TO HUMAN METAPNEUMOVIRUS INFECTION

    PubMed Central

    X, Bao; T, Liu; L, Spetch; D, Kolli; R.P, Garofalo; A, Casola

    2007-01-01

    Human metapneumovirus (hMPV) is a major cause of lower respiratory tract infections (LRTIs) in infants, elderly and immunocompromised patients. In this study, we show that hMPV can infect in a similar manner epithelial cells representative of different tracts of the airways. hMPV-induced expression of chemokines IL-8 and RANTES in primary small alveolar epithelial cells (SAE) and in a human alveolar type II-like epithelial cell line (A549) was similar, suggesting that A549 cells can be used as a model to study lower airway epithelial cell responses to hMPV infection. A549 secreted a variety of CXC and CC chemokines, cytokines and type I interferons, following hMPV infection. hMPV was also a strong inducer of transcription factors belonging to nuclear factor (NF)-κB, interferon regulatory factors (IRFs) and signal transducers and activators of transcription (STATs) families, which are known to orchestrate the expression of inflammatory and immuno-modulatory mediators. PMID:17655903

  17. Apicobasal polarity controls lymphocyte adhesion to hepatic epithelial cells.

    PubMed

    Reglero-Real, Natalia; Alvarez-Varela, Adrián; Cernuda-Morollón, Eva; Feito, Jorge; Marcos-Ramiro, Beatriz; Fernández-Martín, Laura; Gómez-Lechón, Maria José; Muntané, Jordi; Sandoval, Pilar; Majano, Pedro L; Correas, Isabel; Alonso, Miguel A; Millán, Jaime

    2014-09-25

    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. PMID:25242329

  18. IL-17-producing innate lymphoid cells are restricted to mucosal tissues and are depleted in SIV-infected macaques

    PubMed Central

    Xu, Huanbin; Wang, Xiaolei; Liu, David X.; Moroney-Rasmussen, Terri; Lackner, Andrew A.; Veazey, Ronald S.

    2013-01-01

    SUMMARY Innate lymphoid cells (ILCs) are an emerging subset of lymphocytes involved in surveillance against virally infected cells. Here we show CD3−CD8high lymphocytes in macaque blood include major subsets of ILCs including NK cells expressing CD16, NKp46 and NKG2A, but also populations of ILCs in mucosal tissues having different properties. One ILC subset secreted IL-17 (ILC17), but these were restricted to mucosal tissues. Some mucosal ILC17 cells expressed classical NK-cell markers, but little NKG2A or NKG2D. Some ILC17 cells secreted IL-22 and TNF-α, but few produced IFN-γ or contained granzyme B. IL-17 production by ILCs was induced by IL-6, TGF-β and IL-23. Further, SIV infection resulted in a significant loss of ILC17 cells, especially in the jejunum, which persisted throughout SIV infection. These findings ILC17 cells may be involved in innate mucosal immune responses, and their loss may contribute to loss of intestinal mucosal integrity and disease progression in HIV/SIV infection. PMID:22669579

  19. Low direct cytotoxicity of nabumetone on gastric mucosal cells.

    PubMed

    Arai, Yasuhiro; Tanaka, Ken-Ichiro; Ushijima, Hironori; Tomisato, Wataru; Tsutsumi, Shinji; Aburaya, Mayuko; Hoshino, Tatsuya; Yokomizo, Kazumi; Suzuki, Keitarou; Katsu, Takashi; Tsuchiya, Tomofusa; Mizushima, Tohru

    2005-09-01

    Prodrugs of non-steroidal anti-inflammatory drugs (NSAIDs) are widely used for clinical purposes because they are not harmful to the gastrointestinal mucosa. We recently showed that NSAIDs have direct cytotoxicity in NSAID-induced gastric lesions. We show here that under conditions where the NSAIDs indomethacin and celecoxib clearly induce cell death, an NSAID prodrug, nabumetone, and its active metabolite 6-methoxy-2-naphthylacetic acid (6MNA), did not have such effects. Moreover, nabumetone and 6MNA exhibited much lower membrane permeabilizing activities than did indomethacin and celecoxib. We recently reported that when an orally administered NSAID was used in combination with a low dose of intravenously administered indomethacin, the severity of gastric lesions produced in rats depended on the cytotoxicity of the orally administered NSAID. Using a similar protocol, we show here that gastric lesions were produced when the orally administered NSAID was celecoxib, but not when nabumetone was used. We thus propose that the low direct cytotoxicity of nabumetone observed in vitro is maintained in vivo, and that the use of nabumetone does not harm the gastric mucosa. PMID:16133963

  20. Direct contact between dendritic cells and bronchial epithelial cells inhibits T cell recall responses towards mite and pollen allergen extracts in vitro.

    PubMed

    Papazian, D; Wagtmann, V R; Hansen, S; Würtzen, P A

    2015-08-01

    Airway epithelial cells (AECs) form a polarized barrier along the respiratory tract. They are the first point of contact with airborne antigens and are able to instruct resident immune cells to mount appropriate immune responses by either soluble or contact-dependent mechanisms. We hypothesize that a healthy, polarized epithelial cell layer inhibits inflammatory responses towards allergens to uphold homeostasis. Using an in-vitro co-culture model of the airway epithelium, where a polarized cell layer of bronchial epithelial cells can interact with dendritic cells (DCs), we have investigated recall T cell responses in allergic patients sensitized to house dust mite, grass and birch pollen. Using allergen extract-loaded DCs to stimulate autologous allergen-specific T cell lines, we show that AEC-imprinted DCs inhibit T cell proliferation significantly of Bet v 1-specific T cell lines as well as decrease interleukin (IL)-5 and IL-13 production, whereas inhibition of Phl p 5-specific T cells varied between different donors. Stimulating autologous CD4(+) T cells from allergic patients with AEC-imprinted DCs also inhibited proliferation significantly and decreased production of both T helper type 1 (Th1) and Th2 cytokines upon rechallenge. The inhibitory effects of AECs' contact with DCs were absent when allergen extract-loaded DCs had been exposed only to AECs supernatants, but present after direct contact with AECs. We conclude that direct contact between DCs and AECs inhibits T cell recall responses towards birch, grass and house dust mite allergens in vitro, suggesting that AECs-DC contact in vivo constitute a key element in mucosal homeostasis in relation to allergic sensitisation. PMID:25707463

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

    PubMed

    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. PMID:25239531

  2. Epithelial-intrinsic IKKα expression regulates group 3 innate lymphoid cell responses and antibacterial immunity

    PubMed Central

    Giacomin, Paul R.; Moy, Ryan H.; Noti, Mario; Osborne, Lisa C.; Siracusa, Mark C.; Alenghat, Theresa; Liu, Bigang; McCorkell, Kelly A.; Troy, Amy E.; Rak, Gregory D.; Hu, Yinling; May, Michael J.; Ma, Hak-Ling; Fouser, Lynette A.; Sonnenberg, Gregory F.

    2015-01-01

    Innate lymphoid cells (ILCs) are critical for maintaining epithelial barrier integrity at mucosal surfaces; however, the tissue-specific factors that regulate ILC responses remain poorly characterized. Using mice with intestinal epithelial cell (IEC)–specific deletions in either inhibitor of κB kinase (IKK)α or IKKβ, two critical regulators of NFκB activation, we demonstrate that IEC-intrinsic IKKα expression selectively regulates group 3 ILC (ILC3)–dependent antibacterial immunity in the intestine. Although IKKβΔIEC mice efficiently controlled Citrobacter rodentium infection, IKKαΔIEC mice exhibited severe intestinal inflammation, increased bacterial dissemination to peripheral organs, and increased host mortality. Consistent with weakened innate immunity to C. rodentium, IKKαΔIEC mice displayed impaired IL-22 production by RORγt+ ILC3s, and therapeutic delivery of rIL-22 or transfer of sort-purified IL-22–competent ILCs from control mice could protect IKKαΔIEC mice from C. rodentium–induced morbidity. Defective ILC3 responses in IKKαΔIEC mice were associated with overproduction of thymic stromal lymphopoietin (TSLP) by IECs, which negatively regulated IL-22 production by ILC3s and impaired innate immunity to C. rodentium. IEC-intrinsic IKKα expression was similarly critical for regulation of intestinal inflammation after chemically induced intestinal damage and colitis. Collectively, these data identify a previously unrecognized role for epithelial cell–intrinsic IKKα expression and TSLP in regulating ILC3 responses required to maintain intestinal barrier immunity. PMID:26371187

  3. MR1-restricted mucosal associated invariant T (MAIT) cells in the immune response to Mycobacterium tuberculosis

    PubMed Central

    Gold, Marielle C.; Napier, Ruth J.; Lewinsohn, David M.

    2014-01-01

    Summary The intracellular pathogen Mycobacterium tuberculosis (Mtb) and its human host have long co-evolved. Although the host cellular immune response is critical to the control of the bacterium information on the specific contribution of different immune cell subsets in humans is incomplete. Mucosal associated invariant T (MAIT) cells are a prevalent and unique T-cell population in humans with the capacity to detect intracellular infection with bacteria including Mtb. MAIT cells detect bacterially derived metabolites presented by the evolutionarily conserved major histocompatibility complex-like molecule MR1. Here we review recent advances in our understanding of this T-cell subset and address the potential roles for MR1-restricted T cells in the control, diagnosis, and therapy of tuberculosis. PMID:25703558

  4. Mucosal Immunology of Food Allergy

    PubMed Central

    Berin, M. Cecilia; Sampson, Hugh A.

    2013-01-01

    Food allergies are increasing in prevalence at a higher rate than can be explained by genetic factors, suggesting a role for as yet unidentified environmental factors. In this review, we summarize the state of knowledge about the healthy immune response to antigens in the diet and the basis of immune deviation that results in IgE sensitization and allergic reactivity to foods. The intestinal epithelium forms the interface between the external environment and the mucosal immune system, and emerging data suggest that the interaction between intestinal epithelial cells and mucosal dendritic cells is of particular importance in determining the outcome of immune responses to dietary antigens. Exposure to food allergens through non-oral routes, in particular through the skin, is increasingly recognized as a potentially important factor in the increasing rate of food allergy. There are many open questions on the role of environmental factors such as dietary factors and microbiota in the development of food allergy, but data suggest that both have an important modulatory effect on the mucosal immune system. Finally, we discuss recent developments in our understanding of immune mechanisms of clinical manifestations of food allergy. New experimental tools, particularly in the field of genomics and microbiome, are likely to shed light on factors responsible for the growing clinical problem of food allergy. PMID:23660362

  5. Modulation of Candida albicans attachment to human epithelial cells by bacteria and carbohydrates.

    PubMed Central

    Centeno, A; Davis, C P; Cohen, M S; Warren, M M

    1983-01-01

    The effects of carbohydrates (mannose and dextrose). Escherichia coli 07KL. and Klebsiella pneumoniae on Candida albicans attachment to epithelial cells was studied. Dextrose had no effect on yeast attachment to epithelial cells. Conversely, mannose significantly decreased both yeast and piliated bacterial attachment (E. coli 07KL, heavily piliated K. pneumoniae) whereas having no effect on nonpiliated K. pneumoniae attachment to epithelial cells. The number of yeasts attaching to epithelial cells was enhanced by preincubation of epithelial cells with piliated strains of bacteria, whereas preincubation with nonpiliated strains of bacteria had no effect on yeast attachment. Scanning electron microscopy showed that piliated bacteria and yeasts were juxtaposed on the epithelial cell surface. These data suggest that certain piliated strains of bacteria can enhance C. albicans attachment to epithelial cells and that type 1 pili of bacteria can be a factor in the enhanced attachment of C. albicans to epithelial cells. Images PMID:6132878

  6. Renal epithelial cells can release ATP by vesicular fusion

    PubMed Central

    Bjaelde, Randi G.; Arnadottir, Sigrid S.; Overgaard, Morten T.; Leipziger, Jens; Praetorius, Helle A.

    2013-01-01

    Renal epithelial cells have the ability to release nucleotides as paracrine factors. In the intercalated cells of the collecting duct, ATP is released by connexin30 (cx30), which is selectively expressed in this cell type. However, ATP is released by virtually all renal epithelia and the aim of the present study was to identify possible alternative nucleotide release pathways in a renal epithelial cell model. We used MDCK (type1) cells to screen for various potential ATP release pathways. In these cells, inhibition of the vesicular H+-ATPases (bafilomycin) reduced both the spontaneous and hypotonically (80%)-induced nucleotide release. Interference with vesicular fusion using N-ethylamide markedly reduced the spontaneous nucleotide release, as did interference with trafficking from the endoplasmic reticulum to the Golgi apparatus (brefeldin A1) and vesicular transport (nocodazole). These findings were substantiated using a siRNA directed against SNAP-23, which significantly reduced spontaneous ATP release. Inhibition of pannexin and connexins did not affect the spontaneous ATP release in this cell type, which consists of ~90% principal cells. TIRF-microscopy of either fluorescently-labeled ATP (MANT-ATP) or quinacrine-loaded vesicles, revealed that spontaneous release of single vesicles could be promoted by either hypoosmolality (50%) or ionomycin. This vesicular release decreased the overall cellular fluorescence by 5.8 and 7.6% respectively. In summary, this study supports the notion that spontaneous and induced ATP release can occur via exocytosis in renal epithelial cells. PMID:24065923

  7. Lung epithelial cell death induced by oil-dispersant mixtures.

    PubMed

    Wang, He; Shi, Yongli; Major, Danielle; Yang, Zhanjun

    2012-08-01

    The dispersants used in oil spill disasters are claimed to be safe, but increased solubility of high-molecular-weight components in crude oil is of public health concern. The water-accommodated fractions (WAF) of crude oil mixed with dispersants may become airborne and cause lung epithelial damage when inhaled. This study was designed to examine the cell death and related death pathways of lung epithelial cells in response to WAF. Cultured A549 cells were treated for 2 or 24h with different concentrations of WAF. The WAF was prepared by mixing each of the dispersants (Corexit EC9527A, Corexit EC9500A and Corexit EC9580A) with crude oil for extraction with PBS. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide MTT assay, lactate dehydrogenase assay, morphology and cleaved caspase 9 protein, and microtubule-associated protein 1 light chain 3 were all used to measure cell viability, necrosis, apoptosis and autophagy quantitation, respectively. Results showed that the WAF of oil-dispersant mixtures caused cell death in the lung epithelial cells, in a dose-dependent manner, with the major cellular pathways of necrosis and apoptosis involved. Autophagy also occurred in cells exposed to WAF mixtures at lower concentrations before any detectable cell death, indicating greater sensitivity to WAF exposure. The three types of cell behavior, namely necrosis, apoptosis and autophagy, may play different roles in oil spill-related respiratory disorders. PMID:22504303

  8. Effect of dietary fat on the distribution of mucosal mass and cell proliferation along the small intestine.

    PubMed Central

    Jenkins, A P; Thompson, R P

    1992-01-01

    This study investigated how substitution of long chain triglycerides for glucose in a mixed diet affects the overall small intestinal mucosal mass and the distribution of mucosal mass and cell proliferation along the small intestine. Four groups of eight female Wistar rats (180-200 g) were isocalorically fed mixed diets containing the essential fatty acid rich oil Efamol substituted for glucose at concentrations of 1.2%, 10%, 25%, and 50% total calories for 20 to 23 days. The small intestine was divided into three equal length segments and whole gut weights, mucosal weights, protein and DNA determined. Cell proliferation was estimated from the two hour accumulation of vincristine arrested metaphases in microdissected crypts at points 0%, 17%, 33%, 50%, 66%, and 100% small intestinal length. There were no differences between groups in parameters of overall small intestinal or distal segment mucosal mass. With increasing levels of fat, however, there was a significant trend for the mucosal mass of the proximal segment to fall and that of the middle segment to rise. The pattern of two hour metaphase accumulation reflected these changes. These regional changes in mucosal mass and cell proliferation may reflect differences in the sites of absorption of fat and glucose. PMID:1541418

  9. Differentiation of human bronchial epithelial cells: role of hydrocortisone in development of ion transport pathways involved in mucociliary clearance.

    PubMed

    Zaidman, Nathan A; Panoskaltsis-Mortari, Angela; O'Grady, Scott M

    2016-08-01

    Glucocorticoids strongly influence the mucosal-defense functions performed by the bronchial epithelium, and inhaled corticosteroids are critical in the treatment of patients with inflammatory airway diseases such as asthma, chronic obstructive pulmonary disease, and cystic fibrosis. A common pathology associated with these diseases is reduced mucociliary clearance, a defense mechanism involving the coordinated transport of salt, water, and mucus by the bronchial epithelium, ultimately leading to retention of pathogens and particles in the airways and to further disease progression. In the present study we investigated the role of hydrocortisone (HC) in differentiation and development of the ion transport phenotype of normal human bronchial epithelial cells under air-liquid interface conditions. Normal human bronchial epithelial cells differentiated in the absence of HC (HC0) showed significantly less benzamil-sensitive short-circuit current than controls, as well as a reduced response after stimulation with the selective β2-adrenergic receptor agonist salbutamol. Apical membrane localization of epithelial Na(+) channel α-subunits was similarly reduced in HC0 cells compared with controls, supporting a role of HC in the trafficking and density of Na(+) channels in the plasma membrane. Additionally, glucocorticoid exposure during differentiation regulated the transcription of cystic fibrosis transmembrane conductance regulator and β2-adrenergic receptor mRNAs and appeared to be necessary for the expression of cystic fibrosis transmembrane conductance regulator-dependent anion secretion in response to β2-agonists. HC had no significant effect on surface cell differentiation but did modulate the expression of mucin mRNAs. These findings indicate that glucocorticoids support mucosal defense by regulating critical transport pathways essential for effective mucociliary clearance. PMID:27306366

  10. Functional role of mucosal-associated invariant T cells in HIV infection.

    PubMed

    Saeidi, Alireza; Ellegård, Rada; Yong, Yean K; Tan, Hong Y; Velu, Vijayakumar; Ussher, James E; Larsson, Marie; Shankar, Esaki M

    2016-08-01

    MAIT cells represent an evolutionarily conserved, MR1-restricted, innate-like cell subset that express high levels of CD161; have a canonical semi-invariant TCR iVα7.2; and may have an important role in mucosal immunity against various bacterial and fungal pathogens. Mature MAIT cells are CD161(hi)PLZF(hi)IL-18Rα(+)iVα7.2(+)γδ-CD3(+)CD8(+) T cells and occur in the peripheral blood, liver, and mucosa of humans. MAIT cells are activated by a metabolic precursor of riboflavin synthesis presented by MR1 and, therefore, respond to many bacteria and some fungi. Despite their broad antibacterial properties, their functional role in persistent viral infections is poorly understood. Although there is an increasing line of evidence portraying the depletion of MAIT cells in HIV disease, the magnitude and the potential mechanisms underlying such depletion remain unclear. Recent studies suggest that MAIT cells are vulnerable to immune exhaustion as a consequence of HIV and hepatitis C virus infections and HIV/tuberculosis coinfections. HIV infection also appears to cause functional depletion of MAIT cells resulting from abnormal expression of T-bet and EOMES, and effective ART is unable to completely salvage functional MAIT cell loss. Depletion and exhaustion of peripheral MAIT cells may affect mucosal immunity and could increase susceptibility to opportunistic infections during HIV infection. Here, we review some of the important mechanisms associated with depletion and functional loss of MAIT cells and also suggest potential immunotherapeutic strategies to restore MAIT cell functions, including the use of IL-7 to restore effector functions in HIV disease. PMID:27256572

  11. Calcium signaling is involved in ethanol-induced volume decrease and gap junction closure in cultured rat gastric mucosal cells.

    PubMed

    Mustonen, Harri; Kiviluoto, Tuula; Paimela, Hannu; Puolakkainen, Pauli; Kivilaakso, Eero

    2005-01-01

    Ethanol is a well-established "barrier breaker" in gastric mucosa, but its detailed effects at the cellular level remain unclear. We have previously shown that the intracellular free calcium concentration is increased, gap junctions are closed, and cell volume is decreased after exposure to 5% (v/v) ethanol in primarily cultured rabbit gastric epithelial cells. Rat gastric mucosal (RGM) cells were grown to confluence on a coverslip or on a filter membrane. Gap junctional diffusion was measured in 5-carboxyfluorescein-loaded cells by bleaching a small area with a laser and measuring the recovery with confocal microscope. Intracellular calcium was measured spectrofluorometrically in fura-2-loaded cells. For cell volume measurements the cell monolayer was loaded with calcein and imaged along the Z-axis with a confocal microscope. The changes in fluorescence intensity were intercepted as a measure of cell volume change. TMB-8 was used to inhibit intracellular calcium release and lanthanum to block plasma membrane calcium selective ion channels, while BABTA served as an intracellular calcium chelating agent. Results showed that ethanol (7.5%, v/v) exposure increased intracellular calcium from 69 +/- 7 to 142 +/- 11 nM (N = 5; P < 0.05), decreased cell volume by -23 +/- 5% (N = 8; P < 0.05), and induced gap junction closure (fluorescence recovery from 37 +/- 9 to 15 +/- 3%; N = 6; P < 0.05). A serosal potassium channel blocker, quinine, almost completely prevented the ethanol-induced cell volume decrease (from -23 +/- 5 to -3 +/- 3%), suggesting that opening of basolateral potassium channels underlies cell shrinkage. BABTA inhibited completely (from 35 +/- 3 to 39 +/- 4 nM; N = 6; P < 0.05), and TMB-8 + lanthanum partially (from 60 +/- 6 to 92 +/- 12 nM; N = 6; P < 0.05), the ethanol-induced intracellular calcium increase. BABTA also abolished the ethanol-induced volume decrease (from -23 +/- 5 to 1 +/- 4%; N = 6; P < 0.05), while TMB-8 + lanthanum had a lesser effect on

  12. Metabolic cooperativity between epithelial cells and adipocytes of mice

    SciTech Connect

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

    1981-01-01

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

  13. NITROTYROSINE ATTENUATES RSV-INDUCED INFLAMMATION IN AIRWAY EPITHELIAL CELLS

    EPA Science Inventory

    Nitrotyrosine attenuates RSV-induced inflammation in airway epithelial cells. Joleen Soukup, Zuowei Li, Susanne Becker and Yuh-Chin Huang. NHEERL, ORD, USEPA, RTP, North Carolina, CEMALB, University of North Carolina, Chapel Hill, North Carolina

    Nitrotyrosine (NO2Tyr) is a...

  14. Interactions between Periodontal Bacteria and Human Oral Epithelial Cells: Fusobacterium nucleatum Adheres to and Invades Epithelial Cells

    PubMed Central

    Han, Yiping W.; Shi, Wenyuan; Huang, George T.-J.; Kinder Haake, Susan; Park, No-Hee; Kuramitsu, Howard; Genco, Robert J.

    2000-01-01

    Bacteria are causative agents of periodontal diseases. Interactions between oral bacteria and gingival epithelial cells are essential aspects of periodontal infections. Using an in vitro tissue culture model, a selected group of gram-negative anaerobic bacteria frequently associated with periodontal diseases, including Bacteroides forsythus, Campylobacter curvus, Eikenella corrodens, Fusobacterium nucleatum, Porphyromonas gingivalis, and Prevotella intermedia, were examined for their ability to adhere to and invade primary cultures of human gingival epithelial cells (HGEC). The effects of these bacteria on the production of interleukin-8 (IL-8), a proinflammatory chemokine, were also measured. These studies provided an initial demonstration that F. nucleatum adhered to and invaded HGEC and that this was accompanied by high levels of IL-8 secretion from the epithelial cells. The attachment and invasion characteristics of F. nucleatum were also tested using KB cells, an oral epithelial cell line. The invasion was verified by transmission electron microscopy and with metabolic inhibitors. Invasion appeared to occur via a “zipping” mechanism and required the involvement of actins, microtubules, signal transduction, protein synthesis, and energy metabolism of the epithelial cell, as well as protein synthesis by F. nucleatum. A spontaneous mutant, lam, of F. nucleatum, isolated as defective in autoagglutination, was unable to attach to or invade HGEC or KB cells, further indicating the requirement of bacterial components in these processes. Sugar inhibition assays indicated that lectin-like interactions were involved in the attachment of F. nucleatum to KB cells. Investigation of these new virulence phenotypes should improve our understanding of the role of F. nucleatum in periodontal infections. PMID:10816455

  15. Culture and characterization of human junctional epithelial cells.

    PubMed

    Matsuyama, T; Izumi, Y; Sueda, T

    1997-03-01

    This study was undertaken to establish a culture of junctional epithelial cells derived from gingival tissue attached to the tooth surface and to characterize these cells immunocytochemically and ultrastructurally. Primary cultures of cells were obtained from the junctional tissue explanted on type I collagen-coated dishes and immersed in Dulbecco's modified Eagle's medium containing 10% fetal bovine serum (FBS). Cells were subcultured with conditioned serum-free keratinocyte medium (keratinocyte-SFM + 5% FBS) on dishes coated with solubilized extract of the basement membrane. After 24 hours, the medium was changed to keratinocyte-SFM (0.09 mM Ca2+). The cell-doubling time was 40.5 hours. As a control, cells from gingival tissue were cultured by the same method. Cells from junctional tissue and gingival tissue were compared immunocytochemically using monoclonal antibodies to keratin, vimentin, and desmoplakins I and II and using Dolichos biflorus agglutinin (DBA). The keratin AE1 and AE3 was expressed by all of culture cells. The vimentin (specific for the intermediate filament of mesenchymal cells) was also expressed by all cells. The expression pattern of keratin 19 was observed not only by cells from junctional tissue but also by cells from gingival tissue. All keratin peptides were expressed in both cells. However, DBA reacted only with cells from the junctional tissue. Anti-desmoplakin I and II reacted with both cells, however, the staining patterns differed. DBA-positive cultured epithelial cells from the junctional tissue showed poor tonofilament bundles and were rich in cytoplasmic organelles. These findings suggest that junctional epithelial cells can be isolated from junctional tissue and cultured under improved conditions. PMID:9100198

  16. Epithelial adhesion molecules and the regulation of intestinal homeostasis during neutrophil transepithelial migration.

    PubMed

    Sumagin, Ronen; Parkos, Charles A

    2015-01-01

    Epithelial adhesion molecules play essential roles in regulating cellular function and maintaining mucosal tissue homeostasis. Some form epithelial junctional complexes to provide structural support for epithelial monolayers and act as a selectively permeable barrier separating luminal contents from the surrounding tissue. Others serve as docking structures for invading viruses and bacteria, while also regulating the immune response. They can either obstruct or serve as footholds for the immune cells recruited to mucosal surfaces. Currently, it is well appreciated that adhesion molecules collectively serve as environmental cue sensors and trigger signaling events to regulate epithelial function through their association with the cell cytoskeleton and various intracellular adapter proteins. Immune cells, particularly neutrophils (PMN) during transepithelial migration (TEM), can modulate adhesion molecule expression, conformation, and distribution, significantly impacting epithelial function and tissue homeostasis. This review discusses the roles of key intestinal epithelial adhesion molecules in regulating PMN trafficking and outlines the potential consequences on epithelial function. PMID:25838976

  17. Epithelial adhesion molecules and the regulation of intestinal homeostasis during neutrophil transepithelial migration

    PubMed Central

    Sumagin, Ronen; Parkos, Charles A

    2014-01-01

    Epithelial adhesion molecules play essential roles in regulating cellular function and maintaining mucosal tissue homeostasis. Some form epithelial junctional complexes to provide structural support for epithelial monolayers and act as a selectively permeable barrier separating luminal contents from the surrounding tissue. Others serve as docking structures for invading viruses and bacteria, while also regulating the immune response. They can either obstruct or serve as footholds for the immune cells recruited to mucosal surfaces. Currently, it is well appreciated that adhesion molecules collectively serve as environmental cue sensors and trigger signaling events to regulate epithelial function through their association with the cell cytoskeleton and various intracellular adapter proteins. Immune cells, particularly neutrophils (PMN) during transepithelial migration (TEM), can modulate adhesion molecule expression, conformation, and distribution, significantly impacting epithelial function and tissue homeostasis. This review discusses the roles of key intestinal epithelial adhesion molecules in regulating PMN trafficking and outlines the potential consequences on epithelial function. PMID:25838976

  18. Lactobacillus equigenerosi Strain Le1 Invades Equine Epithelial Cells

    PubMed Central

    Botha, Marlie; Botes, Marelize; Loos, Ben; Smith, Carine

    2012-01-01

    Lactobacillus equigenerosi strain Le1, a natural inhabitant of the equine gastrointestinal tract, survived pH 3.0 and incubation in the presence of 1.5% (wt/vol) bile salts for at least 2 h. Strain Le1 showed 8% cell surface hydrophobicity, 60% auto-aggregation, and 47% coaggregation with Clostridium difficile C6. Only 1% of the cells adhered to viable buccal epithelial cells and invaded the cells within 20 min after contact. Preincubation of strain Le1 in a buffer containing pronase prevented adhesion to viable epithelial cells. Preincubation in a pepsin buffer delayed invasion from 20 min to 1 h. Strain Le1 did not adhere to nonviable epithelial cells. Administration of L. equigenerosi Le1 (1 × 109 CFU per 50 kg body weight) to healthy horses did not increase white blood cell numbers. Differential white blood cell counts and aspartate aminotransferase levels remained constant. Glucose, lactate, cholesterol, and urea levels remained constant during administration with L. equigenerosi Le1 but decreased during the week after administration. PMID:22504808

  19. Differentiation of cultured epithelial cells: Response to toxic agents

    SciTech Connect

    Rice, R.H.; LaMontagne, A.D.; Petito, C.T.; Rong, Xianhui )

    1989-03-01

    Cell culture systems are instrumental in elucidating regulation of normal function and mechanisms of its perturbation by toxic substances. To this end, three applications of epithelial cells cultured with 3T3 feeder layer support are described. First, treatment of the premalignant human epidermal keratinocyte line SCC-12F2 with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate suppressed cell growth and differentiation. This agent produced a biphasic growth response greatly inhibiting cell growth at 1 to 10 nM, but much less above 100 nM. Expression of the differentiated functions involucrin and transglutaminase was found to be inhibited markedly at concentrations above 10 nM. Second, 3-methylcholanthrene toxicity was surveyed in a variety of rat epithelial cell types. The two most sensitive to growth inhibition were epidermal and mammary epithelial cells, while those from bladder, prostate, thyroid, and endometrium were insensitive to growth inhibition. Finally, expression of estrogen receptors in rat endometrial cells was shown to be stimulated by the cAmP-elevating agent forskolin. Maximal stimulation of 3- to 6-fold occurred in 6 hr, compatible with a requirement for protein synthesis. Pursuit of such results will aid in understanding differences in response among cell types and species, in elucidating mechanisms of action of known toxic substances and, ultimately, in predicting toxicity of less well understood agents.

  20. Collective Epithelial and Mesenchymal Cell Migration During Gastrulation

    PubMed Central

    Chuai, Manli; Hughes, David; Weijer, Cornelis J

    2012-01-01

    Gastrulation, the process that puts the three major germlayers, the ectoderm, mesoderm and endoderm in their correct topological position in the developing embryo, is characterised by extensive highly organised collective cell migration of epithelial and mesenchymal cells. We discuss current knowledge and insights in the mechanisms controlling these cell behaviours during gastrulation in the chick embryo. We discuss several ideas that have been proposed to explain the observed large scale vortex movements of epithelial cells in the epiblast during formation of the primitive streak. We review current insights in the control and execution of the epithelial to mesenchymal transition (EMT) underlying the formation of the hypoblast and the ingression of the mesendoderm cells through the streak. We discuss the mechanisms by which the mesendoderm cells move, the nature and dynamics of the signals that guide these movements, as well as the interplay between signalling and movement that result in tissue patterning and morphogenesis. We argue that instructive cell-cell signaling and directed chemotactic movement responses to these signals are instrumental in the execution of all phases of gastrulation. PMID:23204916

  1. Candidalysin is a fungal peptide toxin critical for mucosal infection.

    PubMed

    Moyes, David L; Wilson, Duncan; Richardson, Jonathan P; Mogavero, Selene; Tang, Shirley X; Wernecke, Julia; Höfs, Sarah; Gratacap, Remi L; Robbins, Jon; Runglall, Manohursingh; Murciano, Celia; Blagojevic, Mariana; Thavaraj, Selvam; Förster, Toni M; Hebecker, Betty; Kasper, Lydia; Vizcay, Gema; Iancu, Simona I; Kichik, Nessim; Häder, Antje; Kurzai, Oliver; Luo, Ting; Krüger, Thomas; Kniemeyer, Olaf; Cota, Ernesto; Bader, Oliver; Wheeler, Robert T; Gutsmann, Thomas; Hube, Bernhard; Naglik, Julian R

    2016-04-01

    Cytolytic proteins and peptide toxins are classical virulence factors of several bacterial pathogens which disrupt epithelial barrier function, damage cells and activate or modulate host immune responses. Such toxins have not been identified previously in human pathogenic fungi. Here we identify the first, to our knowledge, fungal cytolytic peptide toxin in the opportunistic pathogen Candida albicans. This secreted toxin directly damages epithelial membranes, triggers a danger response signalling pathway and activates epithelial immunity. Membrane permeabilization is enhanced by a positive charge at the carboxy terminus of the peptide, which triggers an inward current concomitant with calcium influx. C. albicans strains lacking this toxin do not activate or damage epithelial cells and are avirulent in animal models of mucosal infection. We propose the name 'Candidalysin' for this cytolytic peptide toxin; a newly identified, critical molecular determinant of epithelial damage and host recognition of the clinically important fungus, C. albicans. PMID:27027296

  2. Ciliary neurotrophic factor promotes the activation of corneal epithelial stem/progenitor cells and accelerates corneal epithelial wound healing.

    PubMed

    Zhou, Qingjun; Chen, Peng; Di, Guohu; Zhang, Yangyang; Wang, Yao; Qi, Xia; Duan, Haoyun; Xie, Lixin

    2015-05-01

    Ciliary neurotrophic factor (CNTF), a well-known neuroprotective cytokine, has been found to play an important role in neurogenesis and functional regulations of neural stem cells. As one of the most innervated tissue, however, the role of CNTF in cornea epithelium remains unclear. This study was to explore the roles and mechanisms of CNTF in the activation of corneal epithelial stem/progenitor cells and wound healing of both normal and diabetic mouse corneal epithelium. In mice subjecting to mechanical removal of corneal epithelium, the corneal epithelial stem/progenitor cell activation and wound healing were promoted by exogenous CNTF application, while delayed by CNTF neutralizing antibody. In cultured corneal epithelial stem/progenitor cells, CNTF enhanced the colony-forming efficiency, stimulated the mitogenic proliferation, and upregulated the expression levels of corneal epithelial stem/progenitor cell-associated transcription factors. Furthermore, the promotion of CNTF on the corneal epithelial stem/progenitor cell activation and wound healing was mediated by the activation of STAT3. Moreover, in diabetic mice, the content of CNTF in corneal epithelium decreased significantly when compared with that of normal mice, and the supplement of CNTF promoted the diabetic corneal epithelial wound healing, accompanied with the advanced activation of corneal epithelial stem/progenitor cells and the regeneration of corneal nerve fibers. Thus, the capability of expanding corneal epithelial stem/progenitor cells and promoting corneal epithelial wound healing and nerve regeneration indicates the potential application of CNTF in ameliorating limbal stem cell deficiency and treating diabetic keratopathy. PMID:25546438

  3. Radical-containing ultrafine particulate matter initiates epithelial-to-mesenchymal transitions in airway epithelial cells.

    PubMed

    Thevenot, Paul T; Saravia, Jordy; Jin, Nili; Giaimo, Joseph D; Chustz, Regina E; Mahne, Sarah; Kelley, Matthew A; Hebert, Valeria Y; Dellinger, Barry; Dugas, Tammy R; Demayo, Francesco J; Cormier, Stephania A

    2013-02-01

    Environmentally persistent free radicals (EPFRs) in combustion-generated particulate matter (PM) are capable of inducing pulmonary pathologies and contributing to the development of environmental asthma. In vivo exposure of infant rats to EPFRs demonstrates their ability to induce airway hyperresponsiveness to methacholine, a hallmark of asthma. However, the mechanisms by which combustion-derived EPFRs elicit in vivo responses remain elusive. In this study, we used a chemically defined EPFR consisting of approximately 0.2 μm amorphrous silica containing 3% cupric oxide with the organic pollutant 1,2-dichlorobenzene (DCB-230). DCB-230 possesses similar radical content to urban-collected EPFRs but offers several advantages, including lack of contaminants and chemical uniformity. DCB-230 was readily taken up by BEAS-2B and at high doses (200 μg/cm(2)) caused substantial necrosis. At low doses (20 μg/cm(2)), DCB-230 particles caused lysosomal membrane permeabilization, oxidative stress, and lipid peroxidation within 24 hours of exposure. During this period, BEAS-2B underwent epithelial-to-mesenchymal transition (EMT), including loss of epithelial cell morphology, decreased E-cadherin expression, and increased α-smooth muscle actin (α-SMA) and collagen I production. Similar results were observed in neonatal air-liquid interface culture (i.e., disruption of epithelial integrity and EMT). Acute exposure of infant mice to DCB-230 resulted in EMT, as confirmed by lineage tracing studies and evidenced by coexpression of epithelial E-cadherin and mesenchymal α-SMA proteins in airway cells and increased SNAI1 expression in the lungs. EMT in neonatal mouse lungs after EPFR exposure may provide an explanation for epidemiological evidence supporting PM exposure and increased risk of asthma. PMID:23087054

  4. The effect of lysolecithin on prostanoid and platelet-activating factor formation by human gall-bladder mucosal cells

    PubMed Central

    Nag, M. K.; Deshpande, Y. G.; Beck, D.; Li, A.

    1995-01-01

    It has been demonstrated that lysolecithin (lysophosphatidyl choline, LPC) produces experimental cholecystitis in cats mediated by arachidonic acid metabolites. LPC is a cytolytic agent that has been postulated as a contributing factor in the development of cholecystitis in humans. The purpose of this research was to evaluate the effect of LPC on human gall-bladder mucosal cell phospholipase A2 and cyclooxygenase activity. Gall-bladder mucosal cells were isolated from the gall-bladders of patients undergoing routine cholecystectomy. Fresh, isolated cells were maintained in tissue culture and stimulated with varying doses of LPC. Platelet-activating factor concentration was quantitated as an index of phospholipase A2 activity and prostanoids were measured as an index of cyclooxygenase activity. Also, the effect of LPC on cyclooxygenase 1 and 2 expression in microsomal protein was evaluated. LPC caused dose related increases in 6-keto-PGF1α and PAF produced by human gall-bladder mucosal cells. Exposure of human gall-bladder mucosal cells to LPC failed to elicit expression of constitutive cyclooxygenase-1, while the expression of inducible cyclooxygenase-2 was increased. The results of this study indicate that LPC induces the formation of prostanoids and PAF by human gall-bladder mucosal cells, suggesting that this substance may promote the development of gall-bladder inflammation. PMID:18475621

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

    NASA Astrophysics Data System (ADS)

    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

  6. Recognition of Vitamin B Precursors and Byproducts by Mucosal Associated Invariant T Cells.

    PubMed

    Eckle, Sidonia B G; Corbett, Alexandra J; Keller, Andrew N; Chen, Zhenjun; Godfrey, Dale I; Liu, Ligong; Mak, Jeffrey Y W; Fairlie, David P; Rossjohn, Jamie; McCluskey, James

    2015-12-18

    Vitamin B2 (riboflavin) is essential for metabolic functions and is synthesized by many bacteria, yeast, and plants, but not by mammals and other animals, which must acquire it from the diet. In mammals, modified pyrimidine intermediates from the microbial biosynthesis of riboflavin are recognized as signature biomarkers of microbial infection. This recognition occurs by specialized lymphocytes known as mucosal associated invariant T (MAIT) cells. The major histocompatibility class I-like antigen-presenting molecule, MR1, captures these pyrimidine intermediates, but only after their condensation with small molecules derived from glycolysis and other metabolic pathways to form short-lived antigens. The resulting MR1-Ag complexes are recognized by MAIT cell antigen receptors (αβ T cell receptors (TCRs)), and the subsequent MAIT cell immune responses are thought to protect the host from pathogens at mucosal surfaces. Here, we review our understanding of how these novel antigens are generated and discuss their interactions with MR1 and MAIT TCRs. PMID:26468291

  7. Investigation of lectinized liposomes as M-cell targeted carrier-adjuvant for mucosal immunization.

    PubMed

    Gupta, Prem N; Vyas, Suresh P

    2011-01-01

    In the present investigation hepatitis B surface antigen (HBsAg) encapsulated liposomes were developed and coupled with Ulex europaeus agglutinin 1 (UEA-1) to increase transmucosal uptake by M-cells of the Peyer's patches. The liposomes were characterized for shape, size, polydispersity and encapsulation efficiency. Bovine submaxillary mucin (BSM) was used as a biological model for the in vitro determination of lectin activity and specificity. Dual staining technique was used to investigate targeting of lectinized liposomes to the M-cells. Anti-HBsAg IgG response in serum and anti-HBsAg sIgA level in various mucosal fluids was estimated by using ELISA, following oral immunization with lectinized and non-lectinized liposomes in Balb/c mice. Additionally, interleukin-2 (IL-2) and interferon-γ (IFN-γ) level in the spleen homogenates was determined. The results suggest that lectinized liposomes were successfully developed, exhibited increased activity with BSM as compared to non-lectinized liposomes and α-l-fucose specificity of the lectinized liposomes was also maintained. The lectinized liposomes were predominantly targeted to the M-cells. The serum anti-HBsAg IgG titre obtained after 3 consecutive days oral immunizations with HBsAg encapsulated lectinized liposomes and boosting after third week was comparable with the titre recorded after single intramuscular prime and third week boosting with alum-HBsAg. Moreover, lectinized liposomes induced higher sIgA level in mucosal secretions and cytokines level in the spleen homogenates. The results showed that the developed surface modified liposomes could be a potential module for the development of effective mucosal vaccines. PMID:20843665

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

    SciTech Connect

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

    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.

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

    PubMed

    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. PMID:27398446

  10. Differentiated kidney epithelial cells repair injured proximal tubule.

    PubMed

    Kusaba, Tetsuro; Lalli, Matthew; Kramann, Rafael; Kobayashi, Akio; Humphreys, Benjamin D

    2014-01-28

    Whether kidney proximal tubule harbors a scattered population of epithelial stem cells is a major unsolved question. Lineage-tracing studies, histologic characterization, and ex vivo functional analysis results conflict. To address this controversy, we analyzed the lineage and clonal behavior of fully differentiated proximal tubule epithelial cells after injury. A CreER(T2) cassette was knocked into the sodium-dependent inorganic phosphate transporter SLC34a1 locus, which is expressed only in differentiated proximal tubule. Tamoxifen-dependent recombination was absolutely specific to proximal tubule. Clonal analysis after injury and repair showed that the bulk of labeled cells proliferate after injury with increased clone size after severe compared with mild injury. Injury to labeled proximal tubule epithelia induced expression of CD24, CD133, vimentin, and kidney-injury molecule-1, markers of putative epithelial stem cells in the human kidney. Similar results were observed in cultured proximal tubules, in which labeled clones proliferated and expressed dedifferentiation and injury markers. When mice with completely labeled kidneys were subject to injury and repair there was no dilution of fate marker despite substantial proliferation, indicating that unlabeled progenitors do not contribute to kidney repair. During nephrogenesis and early kidney growth, single proximal tubule clones expanded, suggesting that differentiated cells also contribute to tubule elongation. These findings provide no evidence for an intratubular stem-cell population, but rather indicate that terminally differentiated epithelia reexpress apparent stem-cell markers during injury-induced dedifferentiation and repair. PMID:24127583

  11. Interleukin-22 Promotes Intestinal Stem Cell-Mediated Epithelial Regeneration

    PubMed Central

    Dudakov, Jarrod A.; Jenq, Robert R.; Velardi, Enrico; Young, Lauren F.; Smith, Odette M.; Lawrence, Gillian; Ivanov, Juliet A.; Fu, Ya-Yuan; Takashima, Shuichiro; Hua, Guoqiang; Martin, Maria L.; O'Rourke, Kevin P.; Lo, Yuan-Hung; Mokry, Michal; Romera-Hernandez, Monica; Cupedo, Tom; Dow, Lukas; Nieuwenhuis, Edward E.; Shroyer, Noah F.; Liu, Chen; Kolesnick, Richard

    2015-01-01

    Epithelial regeneration is critical for barrier maintenance and organ function after intestinal injury. The intestinal stem cell (ISC) niche provides Wnt, Notch, and epidermal growth factor (EGF) signals supporting Lgr5+ crypt base columnar ISCs for normal epithelial maintenance1,2. However, little is known about the regulation of the ISC compartment after tissue damage. Utilizing ex vivo organoid cultures, we provide evidence that innate lymphoid cells (ILCs), potent producers of Interleukin-22 (IL-22) after intestinal injury3,4, increased the growth of murine small intestine (SI) organoids in an IL-22-dependent fashion. Recombinant IL-22 directly targeted ISCs, augmenting the growth of both murine and human intestinal organoids, increasing proliferation, and promoting ISC expansion. IL-22 induced Stat3 phosphorylation in Lgr5+ ISCs, and Stat3 was critical for both organoid formation and IL-22-mediated regeneration. Treatment with IL-22 in vivo after murine allogeneic bone marrow transplantation (BMT) enhanced recovery of ISCs, increased epithelial regeneration, and reduced intestinal pathology and mortality from graft vs. host disease (GVHD). Atoh1-deficient organoid culture demonstrated that IL-22 induced epithelial regeneration independent of the Paneth cell niche. Our findings reveal a fundamental mechanism by which the immune system is able to support intestinal epithelium, activating ISCs to promote regeneration. PMID:26649819

  12. Notch Signaling in Meibomian Gland Epithelial Cell Differentiation

    PubMed Central

    Gidfar, Sanaz; Afsharkhamseh, Neda; Sanjari, Sara; Djalilian, Ali R.

    2016-01-01

    Purpose Notch1 was previously shown to play a critical role in murine meibomian gland function and maintenance. In this study, we have examined the expression and activation of Notch pathway in human meibomian gland epithelial cells in vitro. Methods An immortalized human meibomian gland epithelial cell (HMGEC) line was cultured under proliferative and differentiative conditions. Expression of Notch receptors and ligands were evaluated by quantitative PCR and Western blot. The effect of Notch inhibition and induction on oil production was also assessed. Results Human meibomian gland epithelial cell expressed Notch1, Notch2, Notch3, Jagged1, Jagged2, Delta-like 1, and Delta-like 3. The level of cleaved (activated) Notch1 strongly increased with differentiation. The expression of Notch3 was inversely correlated with proliferation. Induction and inhibition of Notch1 led to an increase and decrease in the amount of oil production, respectively. Conclusions Notch signaling appears to play an important role in human meibomian gland epithelial differentiation and oil production. This may provide a potential therapeutic pathway for treating meibomian gland dysfunction. PMID:26943148

  13. Interleukin-22 promotes intestinal-stem-cell-mediated epithelial regeneration.

    PubMed

    Lindemans, Caroline A; Calafiore, Marco; Mertelsmann, Anna M; O'Connor, Margaret H; Dudakov, Jarrod A; Jenq, Robert R; Velardi, Enrico; Young, Lauren F; Smith, Odette M; Lawrence, Gillian; Ivanov, Juliet A; Fu, Ya-Yuan; Takashima, Shuichiro; Hua, Guoqiang; Martin, Maria L; O'Rourke, Kevin P; Lo, Yuan-Hung; Mokry, Michal; Romera-Hernandez, Monica; Cupedo, Tom; Dow, Lukas E; Nieuwenhuis, Edward E; Shroyer, Noah F; Liu, Chen; Kolesnick, Richard; van den Brink, Marcel R M; Hanash, Alan M

    2015-12-24

    Epithelial regeneration is critical for barrier maintenance and organ function after intestinal injury. The intestinal stem cell (ISC) niche provides Wnt, Notch and epidermal growth factor (EGF) signals supporting Lgr5(+) crypt base columnar ISCs for normal epithelial maintenance. However, little is known about the regulation of the ISC compartment after tissue damage. Using ex vivo organoid cultures, here we show that innate lymphoid cells (ILCs), potent producers of interleukin-22 (IL-22) after intestinal injury, increase the growth of mouse small intestine organoids in an IL-22-dependent fashion. Recombinant IL-22 directly targeted ISCs, augmenting the growth of both mouse and human intestinal organoids, increasing proliferation and promoting ISC expansion. IL-22 induced STAT3 phosphorylation in Lgr5(+) ISCs, and STAT3 was crucial for both organoid formation and IL-22-mediated regeneration. Treatment with IL-22 in vivo after mouse allogeneic bone marrow transplantation enhanced the recovery of ISCs, increased epithelial regeneration and reduced intestinal pathology and mortality from graft-versus-host disease. ATOH1-deficient organoid culture demonstrated that IL-22 induced epithelial regeneration independently of the Paneth cell niche. Our findings reveal a fundamental mechanism by which the immune system is able to support the intestinal epithelium, activating ISCs to promote regeneration. PMID:26649819

  14. Structural and functional analysis of endosomal compartments in epithelial cells.

    PubMed

    Bay, Andres E Perez; Schreiner, Ryan; Rodriguez-Boulan, Enrique

    2015-01-01

    Epithelial cells display segregated early endosomal compartments, termed apical sorting endosomes and basolateral sorting endosomes, that converge into a common late endosomal-lysosomal degradative compartment and common recycling endosomes (CREs). Unlike recycling endosomes of nonpolarized cells, CREs have the ability to sort apical and basolateral plasma membrane proteins into distinct apical and basolateral recycling routes, utilizing mechanisms similar to those employed by the trans Golgi network in the biosynthetic pathway. The apical recycling route includes an additional compartment, the apical recycling endosomes, consisting of multiple vesicles bundled around the basal body. Recent evidence indicates that, in addition to their role in internalizing ligands and recycling their receptors back to the cell surface, endosomal compartments act as intermediate stations in the biosynthetic routes to the plasma membrane. Here we review methods employed by our laboratory to study the endosomal compartments of epithelial cells and their multiple trafficking roles. PMID:26360040

  15. Epithelial-Mesenchymal Transition and Cell Cooperativity in Metastasis

    PubMed Central

    Tsuji, Takanori; Ibaragi, Soichiro; Hu, Guo-fu

    2009-01-01

    The role of epithelial-mesenchymal transition (EMT) in metastasis remains to be controversial. EMT has been postulated as an absolute requirement for tumor invasion and metastasis. Three different models including incomplete EMT, mesenchymal-epithelial transition (MET), and collective migration have been proposed for the role of EMT in cancer invasion and metastasis. However, skepticism remains as to whether EMT truly occurs during caner progression, and if it does, whether it plays an indispensible role in metastasis. Our recent findings suggest that EMT cells are responsible for degrading the surrounding matrix to enable invasion and intravasation of both EMT and non-EMT cells. Only non-EMT cell that have entered the blood stream are able to reestablish colonies in the secondary sites. Here we discuss an alternative model for the role of EMT in cancer metastasis in which EMT and non-EMT cells cooperate to complete the entire process of spontaneous metastasis. PMID:19738043

  16. Culture and immortalization of pancreatic ductal epithelial cells.

    PubMed

    Lawson, Terence; Ouellette, Michel; Kolar, Carol; Hollingsworth, Michael

    2005-01-01

    Some populations of the epithelial cells from the duct and ductular network of the mammalian pancreas have been isolated and maintained in vitro for up to 3 mo. These cells express many of the surface factors that are unique to them in vivo. They also retain significant drug- and carcinogen-metabolizing capacity in vitro. In this chapter we review the progression of the methods for the isolation, culture and maintenance in vitro for these cells from the earliest when only duct/ductular fragments were obtainable to the current ones which provide epithelial cells. The critical steps in the isolation process are identified and strategies are provided to facilitate these steps. These include the selection of tissue digestive enzymes, the importance of extensive mincing before culture and the importance of roles of some co-factors used in the culture medium. PMID:15542901

  17. Inactivation of Rb in stromal fibroblasts promotes epithelial cell invasion.

    PubMed

    Pickard, Adam; Cichon, Ann-Christin; Barry, Anna; Kieran, Declan; Patel, Daksha; Hamilton, Peter; Salto-Tellez, Manuel; James, Jacqueline; McCance, Dennis J

    2012-07-18

    Stromal-derived growth factors are required for normal epithelial growth but are also implicated in tumour progression. We have observed inactivation of the retinoblastoma protein (Rb), through phosphorylation, in cancer-associated fibroblasts in oro-pharyngeal cancer specimens. Rb is well known for its cell-autonomous effects on cancer initiation and progression; however, cell non-autonomous functions of Rb are not well described. We have identified a cell non-autonomous role of Rb, using three-dimensional cultures, where depletion of Rb in stromal fibroblasts enhances invasive potential of transformed epithelia. In part, this is mediated by upregulation of keratinocyte growth factor (KGF), which is produced by the depleted fibroblasts. KGF drives invasion of epithelial cells through induction of MMP1 expression in an AKT- and Ets2-dependent manner. Our data identify that stromal fibroblasts can alter the invasive behaviour of the epithelium, and we show that altered expression of KGF can mediate these functions. PMID:22643222

  18. Cytotoxic Action of Serratia marcescens Hemolysin on Human Epithelial Cells

    PubMed Central

    Hertle, Ralf; Hilger, Martina; Weingardt-Kocher, Sandra; Walev, Iwan

    1999-01-01

    Incubation of human epithelial cells with nanomolar concentrations of chromatographically purified Serratia marcescens hemolysin (ShlA) caused irreversible vacuolation and subsequent lysis of the cells. Vacuolation differed from vacuole formation by Helicobacter pylori VacA. Sublytic doses of ShlA led to a reversible depletion of intracellular ATP. Restoration to the initial ATP level was presumably due to the repair of the toxin damage and was inhibited by cycloheximide. Pores formed in epithelial cells and fibroblasts without disruption of the plasma membrane, and the pores appeared to be considerably smaller than those observed in artificial lipid membranes and in erythrocytes and did not allow the influx of propidium iodide or trypan blue. All cytotoxic effects induced by isolated recombinant ShlA were also obtained with exponentially growing S. marcescens cells. The previously suggested role of the hemolysin in the pathogenicity of S. marcescens is supported by these data. PMID:9916096

  19. COMPARISON OF PM-INDUCED GENE EXPRESSION PROFILES BETWEEN BRONCHIAL EPITHELIAL CELLS AND NASAL EPITHELIAL CELLS IN HUMAN

    EPA Science Inventory

    Epidemiologic studies have linked exposures to particulate matter (PM) and increased pulmonary mortality and morbidity. Bronchial epithelial cells (BEC) are the primary target of PM. PM exposure induces a wide array of biological responses in BEC. Primary human BEC, however, need...

  20. Circulating progenitor epithelial cells traffic via CXCR4/CXCL12 in response to airway injury.

    PubMed

    Gomperts, Brigitte N; Belperio, John A; Rao, P Nagesh; Randell, Scott H; Fishbein, Michael C; Burdick, Marie D; Strieter, Robert M

    2006-02-01

    Recipient airway epithelial cells are found in human sex-mismatched lung transplants, implying that circulating progenitor epithelial cells contribute to the repair of the airway epithelium. Markers of circulating progenitor epithelial cells and mechanisms for their trafficking remain to be elucidated. We demonstrate that a population of progenitor epithelial cells exists in the bone marrow and the circulation of mice that is positive for the early epithelial marker cytokeratin 5 (CK5) and the chemokine receptor CXCR4. We used a mouse model of sex-mismatched tracheal transplantation and found that CK5+ circulating progenitor epithelial cells contribute to re-epithelialization of the airway and re-establishment of the pseudostratified epithelium. The presence of CXCL12 in tracheal transplants provided a mechanism for CXCR4+ circulating progenitor epithelial cell recruitment to the airway. Depletion of CXCL12 resulted in the epithelium defaulting to squamous metaplasia, which was derived solely from the resident tissue progenitor epithelial cells. Our findings demonstrate that CK5+CXCR4+ cells are markers of circulating progenitor epithelial cells in the bone marrow and circulation and that CXCR4/CXCL12-mediated recruitment of circulating progenitor epithelial cells is necessary for the re-establishment of a normal pseudostratified epithelium after airway injury. These findings support a novel paradigm for the development of squamous metaplasia of the airway epithelium and for developing therapeutic strategies for circulating progenitor epithelial cells in airway diseases. PMID:16424223

  1. Midbody remnant licenses primary cilia formation in epithelial cells.

    PubMed

    Ott, Carolyn M

    2016-08-01

    Tethered midbody remnants dancing across apical microvilli, encountering the centrosome, and beckoning forth a cilium-who would have guessed this is how polarized epithelial cells coordinate the end of mitosis and the beginning of ciliogenesis? New evidence from Bernabé-Rubio et al. (2016. J. Cell Biol http://dx.doi.org/10.1083/jcb.201601020) supports this emerging model. PMID:27482049

  2. Review: Corneal epithelial stem cells, their niche and wound healing

    PubMed Central

    2013-01-01

    Stem cells emerged as a concept during the second half of 19th century, first as a theoretical entity, but then became one of the most promising research fields in cell biology. This work describes the most important characteristics of adult stem cells, including the experimental criteria used to identify them, and discusses current knowledge that led to the proposal that stem cells existed in different parts of the eye, such as the retina, lens, conjunctiva, corneal stroma, Descemet’s membrane, and the subject of this review: the corneal epithelium. Evidence includes results that support the presence of corneal epithelial stem cells at the limbus, as well as the major obstacles to isolating them as pure cell populations. Part of this review describes the variation in the basement membrane composition between the limbus and the central cornea, to show the importance of the corneal stem cell niche, its structure, and the participation of extracellular matrix (ECM) components in regulating corneal stem cell compartment. Results obtained by various laboratories suggest that the extracellular matrix plays a central role in regulating stem cell commitment, corneal differentiation, and participation in corneal wound healing, in addition to other environmental signals such as cytokines and growth factors. The niche could define cell division patterns in corneal stem cell populations, establishing whether stem cells divide asymmetrically or symmetrically. Characterization and understanding of the factors that regulate corneal epithelial stem cells should open up new paths for developing new therapies and strategies for accelerating and improving corneal wound healing. PMID:23901244

  3. MUCOSAL IMMUNOLOGY. The microbiota regulates type 2 immunity through RORγt⁺ T cells.

    PubMed

    Ohnmacht, Caspar; Park, Joo-Hong; Cording, Sascha; Wing, James B; Atarashi, Koji; Obata, Yuuki; Gaboriau-Routhiau, Valérie; Marques, Rute; Dulauroy, Sophie; Fedoseeva, Maria; Busslinger, Meinrad; Cerf-Bensussan, Nadine; Boneca, Ivo G; Voehringer, David; Hase, Koji; Honda, Kenya; Sakaguchi, Shimon; Eberl, Gérard

    2015-08-28

    Changes to the symbiotic microbiota early in life, or the absence of it, can lead to exacerbated type 2 immunity and allergic inflammations. Although it is unclear how the microbiota regulates type 2 immunity, it is a strong inducer of proinflammatory T helper 17 (T(H)17) cells and regulatory T cells (T(regs)) in the intestine. Here, we report that microbiota-induced T(regs) express the nuclear hormone receptor RORγt and differentiate along a pathway that also leads to T(H)17 cells. In the absence of RORγt(+) T(regs), T(H)2-driven defense against helminths is more efficient, whereas T(H)2-associated pathology is exacerbated. Thus, the microbiota regulates type 2 responses through the induction of type 3 RORγt(+) T(regs) and T(H)17 cells and acts as a key factor in balancing immune responses at mucosal surfaces. PMID:26160380

  4. Vangl2 Regulates E-Cadherin in Epithelial Cells

    PubMed Central

    Nagaoka, Tadahiro; Inutsuka, Ayumu; Begum, Khadiza; hafiz, Khandakar musabbir bin; Kishi, Masashi

    2014-01-01

    E-cadherin belongs to the classic cadherin subfamily of calcium-dependent cell adhesion molecules and is crucial for the formation and function of epithelial adherens junctions. In this study, we demonstrate that Vangl2, a vertebrate regulator of planar cell polarity (PCP), controls E-cadherin in epithelial cells. E-cadherin co-immunoprecipitates with Vangl2 from embryonic kidney extracts, and this association is also observed in transfected fibroblasts. Vangl2 enhances the internalization of E-cadherin when overexpressed. Conversely, the quantitative ratio of E-cadherin exposed to the cell surface is increased in cultured renal epithelial cells derived from Vangl2Lpt/+ mutant mice. Interestingly, Vangl2 is also internalized through protein traffic involving Rab5- and Dynamin-dependent endocytosis. Taken together with recent reports regarding the transport of Frizzled3, MMP14 and nephrin, these results suggest that one of the molecular functions of Vangl2 is to enhance the internalization of specific plasma membrane proteins with broad selectivity. This function may be involved in the control of intercellular PCP signalling or in the PCP-related rearrangement of cell adhesions. PMID:25373475

  5. Sef Regulates Epithelial-Mesenchymal Transition in Breast Cancer Cells.

    PubMed

    He, Qing; Gong, Yan; Gower, Lindsey; Yang, Xuehui; Friesel, Robert E

    2016-10-01

    Sef (similar expression to fgf), also know as IL17RD, is a transmembrane protein shown to inhibit fibroblast growth factor signaling in developmental and cancer contexts; however, its role as a tumor suppressor remains to be fully elucidated. Here, we show that Sef regulates epithelial-mesenchymal transition (EMT) in breast cancer cell lines. Sef expression was highest in the normal breast epithelial cell line MCF10A, intermediate expression in MCF-7 cells and lowest in MDA-MB-231 cells. Knockdown of Sef increased the expression of genes associated with EMT, and promoted cell migration, invasion, and a fibroblastic morphology of MCF-7 cells. Overexpression of Sef inhibited the expression of EMT marker genes and inhibited cell migration and invasion in MCF-7 cells. Induction of EMT in MCF10A cells by TGF-β and TNF-α resulted in downregulation of Sef expression concomitant with upregulation of EMT gene expression and loss of epithelial morphology. Overexpression of Sef in MCF10A cells partially blocked cytokine-induced EMT. Sef was shown to block β-catenin mediated luciferase reporter activity and to cause a decrease in the nuclear localization of active β-catenin. Furthermore, Sef was shown to co-immunoprecipitate with β-catenin. In a mouse orthotopic xenograft model, Sef overexpression in MDA-MB-231 cells slowed tumor growth and reduced expression of EMT marker genes. Together, these data indicate that Sef plays a role in the negative regulation of EMT in a β-catenin dependent manner and that reduced expression of Sef in breast tumor cells may be permissive for EMT and the acquisition of a more metastatic phenotype. J. Cell. Biochem. 117: 2346-2356, 2016. © 2016 Wiley Periodicals, Inc. PMID:26950413

  6. Lipidome and Transcriptome Profiling of Pneumolysin Intoxication Identifies Networks Involved in Statin-Conferred Protection of Airway Epithelial Cells

    PubMed Central

    Statt, Sarah; Ruan, Jhen-Wei; Huang, Chih-Ting; Wu, Reen; Kao, Cheng-Yuan

    2015-01-01

    Pneumonia remains one of the leading causes of death in both adults and children worldwide. Despite the adoption of a wide variety of therapeutics, the mortality from community-acquired pneumonia has remained relatively constant. Although viral and fungal acute airway infections can result in pneumonia, bacteria are the most common cause of community-acquired pneumonia, with Streptococcus pneumoniae isolated in nearly 50% of cases. Pneumolysin is a cholesterol-dependent cytolysin or pore-forming toxin produced by Streptococcus pneumonia and has been shown to play a critical role in bacterial pathogenesis. Airway epithelium is the initial site of many bacterial contacts and its barrier and mucosal immunity functions are central to infectious lung diseases. In our studies, we have shown that the prior exposure to statins confers significant resistance of airway epithelial cells to the cytotoxicity of pneumolysin. We decided to take this study one step further, assessing changes in both the transcriptome and lipidome of human airway epithelial cells exposed to toxin, statin or both. Our current work provides the first global view in human airway epithelial cells of both the transcriptome and the lipid interactions that result in cellular protection from pneumolysin. PMID:26023727

  7. Exposure to zearalenone mycotoxin alters in vitro porcine intestinal epithelial cells by differential gene expression.

    PubMed

    Taranu, Ionelia; Braicu, Cornelia; Marin, Daniela Eliza; Pistol, Gina Cecilia; Motiu, Monica; Balacescu, Loredana; Beridan Neagoe, Ioana; Burlacu, Radu

    2015-01-01

    The gut represents the main route of intoxication with mycotoxins. To evaluate the effect and the underlying molecular changes that occurred when the intestine is exposed to zearalenone, a Fusarium sp mycotoxin, porcine epithelial cells (IPEC-1) were treated with 10μM of ZEA for 24h and analysed by microarray using Gene Spring GX v.11.5. Our results showed that 10μM of ZEA did not affect cell viability, but can increase the expression of toll like receptors (TLR1-10) and of certain cytokines involved in inflammation (TNF-α, IL-1β, IL-6, IL-8, MCP-1, IL-12p40, CCL20) or responsible for the recruitment of immune cells (IL-10, IL-18). Microarray results identified 190 genes significantly and differentially expressed, of which 70% were up-regulated. ZEA determined the over expression of ITGB5 gene, essential against the attachment and adhesion of ETEC to porcine jejunal cells and of TFF2 implicated in mucosal protection. An up-regulation of glutathione peroxidase enzymes (GPx6, GPx2, GPx1) was also observed. Upon ZEA challenge, genes like GTF3C4 responsible for the recruitment of polymerase III and initiation of tRNA transcription in eukaryotes and STAT5B were significantly higher induced. The up-regulation of CD97 gene and the down-regulation of tumour suppressor genes (DKK-1, PCDH11X and TC531386) demonstrates the carcinogenic potential of ZEA. PMID:25455459

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

    PubMed Central

    Fehrenbach, Heinz

    2001-01-01

    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. PMID:11686863

  9. Generation of Stratified Squamous Epithelial Progenitor Cells from Mouse Induced Pluripotent Stem Cells

    PubMed Central

    Yoshida, Satoru; Yasuda, Miyuki; Miyashita, Hideyuki; Ogawa, Yoko; Yoshida, Tetsu; Matsuzaki, Yumi; Tsubota, Kazuo; Okano, Hideyuki; Shimmura, Shigeto

    2011-01-01

    Background Application of induced pluripotent stem (iPS) cells in regenerative medicine will bypass ethical issues associated with use of embryonic stem cells. In addition, patient-specific IPS cells can be useful to elucidate the pathophysiology of genetic disorders, drug screening, and tailor-made medicine. However, in order to apply iPS cells to mitotic tissue, induction of tissue stem cells that give rise to progeny of the target organ is required. Methodology/Principal Findings We induced stratified epithelial cells from mouse iPS cells by co-culture with PA6 feeder cells (SDIA-method) with use of BMP4. Clusters of cells positive for the differentiation markers KRT1 or KRT12 were observed in KRT14-positive colonies. We successfully cloned KRT14 and p63 double-positive stratified epithelial progenitor cells from iPS-derived epithelial cells, which formed stratified epithelial sheets consisting of five- to six-polarized epithelial cells in vitro. When these clonal cells were cultured on denuded mouse corneas, a robust stratified epithelial layer was observed with physiological cell polarity including high levels of E-cadherin, p63 and K15 expression in the basal layer and ZO-1 in the superficial layer, recapitulating the apico-basal polarity of the epithelium in vivo. Conclusions/Significance These results suggest that KRT14 and p63 double-positive epithelial progenitor cells can be cloned from iPS cells in order to produce polarized multilayer epithelial cell sheets. PMID:22174914

  10. STARVATION INDUCED PROXIMAL GUT MUCOSAL ATROPHY DIMINISHED WITH AGING

    PubMed Central

    Song, Juquan; Wolf, Steven E.; Wu, Xiao-Wu; Finnerty, Celeste C.; Gauglitz, Gerd G.; Herndon, David N.; Jeschke, Marc G.

    2013-01-01

    Background Starvation induces small bowel atrophy with increased intestinal epithelial apoptosis and decreased proliferation. Here, we examined these parameters after starvation in aged animals. Methods Sixty-four 6 week-old and 26 month-old C57BL/6 mice were randomly assigned to either an ad libitum fed or fasted group. The small bowel was harvested at 12, 48, and 72 hours following starvation. Proximal gut mucosal height was measured and epithelial cells counted. Apoptosis was identified by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. Proliferation was determined by immunohistochemical staining for proliferating cell nuclear antigen (PCNA). Comparison of fed vs. fasted and adult vs. old groups was done by one-way ANOVA with Tukey’s test and unpaired t-test. Significance was accepted at p<0.05. Results Aged mice had higher proximal gut weights, mucosal heights and cell numbers at baseline compared with the adult group (p<0.05). The rate of apoptosis was lower in the aged (p<0.05) while proliferation was not different between groups before starvation. After starvation, proximal gut wet weight decreased only in adult mice (p<0.05); Gut mucosal height and mucosal cell number decreased greater in adult than in aged mice (p<0.05). This was related to decreased proliferation only in the adult group (p<0.05). The fold of epithelial apoptosis increased was higher in the aged group than in the adult after starvation (p<0.05). Conclusions Gut mucosal kinetics change with age had lower rates of apoptosis and greater mucosal mass; the character of starvation-induced atrophy is diminished with aging. PMID:19126762

  11. Cytokeratin changes in cell culture systems of epithelial cells isolated from oral mucosa: a short review.

    PubMed

    Gasparoni, Alberto; Squier, Christopher Alan; Fonzi, Luciano

    2005-01-01

    In the past three decades, many studies have analyzed ultrastructural and molecular markers of differentiation in squamous stratified epithelial tissues. In these tissues, epithelial cells migrating from the basal layer to the upper layers undergo drastic changes, which involve membrane-associated proteins, DNA synthesis, phenotypic aspects, lipid composition, and cytoskeletal components. Cytoskeletal components include a large and heterogeneous group, including intermediate filaments, components of the cornified envelope, and of the stratum corneum. When grown in mono- and multilayer cell cultures, epithelial cells isolated from the oral mucosa may reproduce many of the biochemical and morphological aspects of epithelial tissue in vivo. In the present paper, we examine phenotypic changes, development of suprabasal layer, and Involucrin expression occurring in differentiating oral epithelial cells, based on literature review and original data. PMID:16277157

  12. Temporal Monitoring of Differentiated Human Airway Epithelial Cells Using Microfluidics

    PubMed Central

    Blume, Cornelia; Reale, Riccardo; Held, Marie; Millar, Timothy M.; Collins, Jane E.; Davies, Donna E.; Morgan, Hywel; Swindle, Emily J.

    2015-01-01

    The airway epithelium is exposed to a variety of harmful agents during breathing and appropriate cellular responses are essential to maintain tissue homeostasis. Recent evidence has highlighted the contribution of epithelial barrier dysfunction in the development of many chronic respiratory diseases. Despite intense research efforts, the responses of the airway barrier to environmental agents are not fully understood, mainly due to lack of suitable in vitro models that recapitulate the complex in vivo situation accurately. Using an interdisciplinary approach, we describe a novel dynamic 3D in vitro model of the airway epithelium, incorporating fully differentiated primary human airway epithelial cells at the air-liquid interface and a basolateral microfluidic supply of nutrients simulating the interstitial flow observed in vivo. Through combination of the microfluidic culture system with an automated fraction collector the kinetics of cellular responses by the airway epithelium to environmental agents can be analysed at the early phases for the first time and with much higher sensitivity compared to common static in vitro models. Following exposure of primary differentiated epithelial cells to pollen we show that CXCL8/IL–8 release is detectable within the first 2h and peaks at 4–6h under microfluidic conditions, a response which was not observed in conventional static culture conditions. Such a microfluidic culture model is likely to have utility for high resolution temporal profiling of toxicological and pharmacological responses of the airway epithelial barrier, as well as for studies of disease mechanisms. PMID:26436734

  13. Temporal Monitoring of Differentiated Human Airway Epithelial Cells Using Microfluidics.

    PubMed

    Blume, Cornelia; Reale, Riccardo; Held, Marie; Millar, Timothy M; Collins, Jane E; Davies, Donna E; Morgan, Hywel; Swindle, Emily J

    2015-01-01

    The airway epithelium is exposed to a variety of harmful agents during breathing and appropriate cellular responses are essential to maintain tissue homeostasis. Recent evidence has highlighted the contribution of epithelial barrier dysfunction in the development of many chronic respiratory diseases. Despite intense research efforts, the responses of the airway barrier to environmental agents are not fully understood, mainly due to lack of suitable in vitro models that recapitulate the complex in vivo situation accurately. Using an interdisciplinary approach, we describe a novel dynamic 3D in vitro model of the airway epithelium, incorporating fully differentiated primary human airway epithelial cells at the air-liquid interface and a basolateral microfluidic supply of nutrients simulating the interstitial flow observed in vivo. Through combination of the microfluidic culture system with an automated fraction collector the kinetics of cellular responses by the airway epithelium to environmental agents can be analysed at the early phases for the first time and with much higher sensitivity compared to common static in vitro models. Following exposure of primary differentiated epithelial cells to pollen we show that CXCL8/IL-8 release is detectable within the first 2h and peaks at 4-6h under microfluidic conditions, a response which was not observed in conventional static culture conditions. Such a microfluidic culture model is likely to have utility for high resolution temporal profiling of toxicological and pharmacological responses of the airway epithelial barrier, as well as for studies of disease mechanisms. PMID:26436734

  14. Retinoic acid promotes primary fetal alveolar epithelial type II cell proliferation and differentiation to alveolar epithelial type I cells.

    PubMed

    Gao, Rui-wei; Kong, Xiang-yong; Zhu, Xiao-xi; Zhu, Guo-qing; Ma, Jin-shuai; Liu, Xiu-xiang

    2015-05-01

    Retinoic acid (RA) plays an important role in lung development and maturation. Many stimuli can induce alveolar epithelial cell damage which will result in the injury of lung parenchyma. The aim of this study was to observe the effect of RA on the proliferation and differentiation of primary fetal alveolar epithelial type II cells (fAECIIs). Primary fAECIIs were isolated from fetal rats at 19 d of gestation and purified by a differential centrifugation and adhesion method. The cells were randomly divided into control (dimethyl sulfoxide, DMSO) and RA groups. Cell proliferation, viability, apoptosis, cycle, and expression of target protein were examined at 24, 48, and 72 h. We found that the proliferation and viability of cells in the RA-exposed group significantly increased compared with the DMSO control group. The proportion (%) of cells in the G2 and S phases in the RA group was significantly higher than that in control group cells. The proportion (%) of both early apoptotic cells and late apoptotic cells decreased significantly in cells exposed to RA compared with cells exposed to DMSO. RA significantly enhanced the expression of aquaporin 5 (AQP5). The expression level of pulmonary surfactant C (SPC) was elevated after cells were exposed to RA for 24 and 72 h but was inhibited when cells were exposed to RA for 48 h. These results suggest that RA promotes fAECII proliferation by improving cell viability, promoting S phase entry and inhibiting apoptosis and RA promotes fAECIIs differentiation to alveolar epithelial type I cells (AECIs). PMID:25515249

  15. Oral mucosal manifestations of autoimmune skin diseases.

    PubMed

    Mustafa, Mayson B; Porter, Stephen R; Smoller, Bruce R; Sitaru, Cassian

    2015-10-01

    A group of autoimmune diseases is characterised by autoantibodies against epithelial adhesion structures and/or tissue-tropic lymphocytes driving inflammatory processes resulting in specific pathology at the mucosal surfaces and the skin. The most frequent site of mucosal involvement in autoimmune diseases is the oral cavity. Broadly, these diseases include conditions affecting the cell-cell adhesion causing intra-epithelial blistering and those where autoantibodies or infiltration lymphocytes cause a loss of cell-matrix adhesion or interface inflammation. Clinically, patients present with blistering, erosions and ulcers that may affect the skin as well as further mucosal surfaces of the eyes, nose and genitalia. While the autoimmune disease may be suspected based on clinical manifestations, demonstration of tissue-bound and circulating autoantibodies, or lymphocytic infiltrates, by various methods including histological examination, direct and indirect immunofluorescence microscopy, immunoblotting and quantitative immunoassay is a prerequisite for definitive diagnosis. Given the frequency of oral involvement and the fact that oral mucosa is the initially affected site in many cases, the informed practitioner should be well acquainted with diagnostic and therapeutic aspects of autoimmune dermatosis with oral involvement. This paper reviews the pathogenesis and clinical presentation of these conditions in the oral cavity with a specific emphasis on their differential diagnosis and current management approaches. PMID:26117595

  16. Human airway xenograft models of epithelial cell regeneration.

    PubMed

    Puchelle, E; Peault, B

    2000-01-01

    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. PMID:11667974

  17. Epithelial Cell Proliferation Contributes to Airway Remodeling in Severe Asthma

    PubMed Central

    Cohen, Lance; E, Xueping; Tarsi, Jaime; Ramkumar, Thiruvamoor; Horiuchi, Todd K.; Cochran, Rebecca; DeMartino, Steve; Schechtman, Kenneth B.; Hussain, Iftikhar; Holtzman, Michael J.; Castro, Mario

    2007-01-01

    Rationale: Despite long-term therapy with corticosteroids, patients with severe asthma develop irreversible airway obstruction. Objectives: To evaluate if there are structural and functional differences in the airway epithelium in severe asthma associated with airway remodeling. Methods: In bronchial biopsies from 21 normal subjects, 11 subjects with chronic bronchitis, 9 subjects with mild asthma, and 31 subjects with severe asthma, we evaluated epithelial cell morphology: epithelial thickness, lamina reticularis (LR) thickness, and epithelial desquamation. Levels of retinoblastoma protein (Rb), Ki67, and Bcl-2 were measured, reflecting cellular proliferation and death. Terminal deoxynucleotidyl-mediated dUTP nick end labeling (TUNEL) was used to study cellular apoptosis. Measurements and Main Results: Airway epithelial and LR thickness was greater in subjects with severe asthma compared with those with mild asthma, normal subjects, and diseased control subjects (p = 0.009 and 0.033, respectively). There was no significant difference in epithelial desquamation between groups. Active, hypophosphorylated Rb expression was decreased (p = 0.002) and Ki67 was increased (p < 0.01) in the epithelium of subjects with severe asthma as compared with normal subjects, indicating increased cellular proliferation. Bcl-2 expression was decreased (p < 0.001), indicating decreased cell death suppression. There was a greater level of apoptotic activity in the airway biopsy in subjects with severe asthma as compared with the normal subjects using the TUNEL assay (p = 0.002), suggesting increased cell death. Conclusions: In subjects with severe asthma, as compared with subjects with mild asthma, normal subjects, and diseased control subjects, we found novel evidence of increased cellular proliferation in the airway contributing to a thickened epithelium and LR. These changes may contribute to the progressive decline in lung function and airway remodeling in patients with severe

  18. Effects of phenol on barrier function of a human intestinal epithelial cell line correlate with altered tight junction protein localization.

    PubMed

    McCall, Ingrid C; Betanzos, Abigail; Weber, Dominique A; Nava, Porfirio; Miller, Gary W; Parkos, Charles A

    2009-11-15

    Phenol contamination of soil and water has raised concerns among people living near phenol-producing factories and hazardous waste sites containing the chemical. Phenol, particularly in high concentrations, is an irritating and corrosive substance, making mucosal membranes targets of toxicity in humans. However, few data on the effects of phenol after oral exposure exist. We used an in vitro model employing human intestinal epithelial cells (SK-CO15) cultured on permeable supports to examine effects of phenol on epithelial barrier function. We hypothesized that phenol disrupts epithelial barrier by altering tight junction (TJ) protein expression. The dose-response effect of phenol on epithelial barrier function was determined using transepithelial electrical resistance (TER) and FITC-dextran permeability measurements. We studied phenol-induced changes in cell morphology and expression of several tight junction proteins by immunofluorescence and Western blot analysis. Effects on cell viability were assessed by MTT, Trypan blue, propidium iodide and TUNEL staining. Exposure to phenol resulted in decreased TER and increased paracellular flux of FITC-dextran in a dose-dependent manner. Delocalization of claudin-1 and ZO-1 from TJs to cytosol correlated with the observed increase in permeability after phenol treatment. Additionally, the decrease in TER correlated with changes in the distribution of a membrane raft marker, suggesting phenol-mediated effects on membrane fluidity. Such observations were independent of effects of phenol on cell viability as enhanced permeability occurred at doses of phenol that did not cause cell death. Overall, these findings suggest that phenol may affect transiently the lipid bilayer of the cell membrane, thus destabilizing TJ-containing microdomains. PMID:19679145

  19. Effects of phenol on barrier function of a human intestinal epithelial cell line correlate with altered tight junction protein localization

    PubMed Central

    McCall, Ingrid C.; Betanzos, Abigail; Weber, Dominique A.; Nava, Porfirio; Miller, Gary W.; Parkos, Charles A.

    2010-01-01

    Phenol contamination of soil and water has raised concerns among people living near phenol-producing factories and hazardous waste sites containing the chemical. Phenol, particularly in high concentrations, is an irritating and corrosive substance, making mucosal membranes targets of toxicity in humans. However, few data on the effects of phenol after oral exposure exist. We used an in vitro model employing human intestinal epithelial cells (SK-CO15) cultured on permeable supports to examine effects of phenol on epithelial barrier function. We hypothesized that phenol disrupts epithelial barrier by altering tight junction (TJ) protein expression. The dose-response effect of phenol on epithelial barrier function was determined using transepithelial electrical resistance (TER) and FITC-dextran permeability measurements. We studied phenol-induced changes in cell morphology and expression of several tight junction proteins by immunofluorescence and Western blot analysis. Effects on cell viability were assessed by MTT, Trypan blue, propidium iodide and TUNEL staining. Exposure to phenol resulted in decreased TER and increased paracellular flux of FITC-dextran in a dose-dependent manner. Delocalization of claudin-1 and ZO-1 from TJs to cytosol correlated with the observed increase in permeability after phenol treatment. Additionally, the decrease in TER correlated with changes in the distribution of a membrane raft marker, suggesting phenol-mediated effects on membrane fluidity. Such observations were independent of effects of phenol on cell viability as enhanced permeability occurred at doses of phenol that did not cause cell death. Overall, these findings suggest that phenol may affect transiently the lipid bilayer of the cell membrane, thus destabilizing TJ-containing microdomains. PMID:19679145

  20. Effects of phenol on barrier function of a human intestinal epithelial cell line correlate with altered tight junction protein localization

    SciTech Connect

    McCall, Ingrid C.; Betanzos, Abigail; Weber, Dominique A.; Nava, Porfirio; Miller, Gary W.; Parkos, Charles A.

    2009-11-15

    Phenol contamination of soil and water has raised concerns among people living near phenol-producing factories and hazardous waste sites containing the chemical. Phenol, particularly in high concentrations, is an irritating and corrosive substance, making mucosal membranes targets of toxicity in humans. However, few data on the effects of phenol after oral exposure exist. We used an in vitro model employing human intestinal epithelial cells (SK-CO15) cultured on permeable supports to examine effects of phenol on epithelial barrier function. We hypothesized that phenol disrupts epithelial barrier by altering tight junction (TJ) protein expression. The dose-response effect of phenol on epithelial barrier function was determined using transepithelial electrical resistance (TER) and FITC-dextran permeability measurements. We studied phenol-induced changes in cell morphology and expression of several tight junction proteins by immunofluorescence and Western blot analysis. Effects on cell viability were assessed by MTT, Trypan blue, propidium iodide and TUNEL staining. Exposure to phenol resulted in decreased TER and increased paracellular flux of FITC-dextran in a dose-dependent manner. Delocalization of claudin-1 and ZO-1 from TJs to cytosol correlated with the observed increase in permeability after phenol treatment. Additionally, the decrease in TER correlated with changes in the distribution of a membrane raft marker, suggesting phenol-mediated effects on membrane fluidity. Such observations were independent of effects of phenol on cell viability as enhanced permeability occurred at doses of phenol that did not cause cell death. Overall, these findings suggest that phenol may affect transiently the lipid bilayer of the cell membrane, thus destabilizing TJ-containing microdomains.

  1. Transcriptional Regulation of Tlr11 Gene Expression in Epithelial Cells*

    PubMed Central

    Cai, Zhenyu; Shi, Zhongcheng; Sanchez, Amir; Zhang, Tingting; Liu, Mingyao; Yang, Jianghua; Wang, Fen; Zhang, Dekai

    2009-01-01

    As sensors of invading microorganisms, Toll-like receptors (TLRs) are expressed not only on macrophages and dendritic cells (DCs) but also on epithelial cells. In the TLR family, Tlr11 appears to have the unique feature in that it is expressed primarily on epithelial cells, although it is also expressed on DCs and macrophages. Here, we demonstrate that transcription of the Tlr11 gene is regulated through two cis-acting elements, one Ets-binding site and one interferon regulatory factor (IRF)-binding site. The Ets element interacts with the epithelium-specific transcription factors, ESE-1 and ESE-3, and the IRF motif interacts with IRF-8. Thus, Tlr11 expression on epithelial cells is regulated by the transcription factors that are presumably distinct from transcription factors that regulate the expression of TLRs in innate immune cells such as macrophages and DCs. Our results imply that the distinctive transcription regulatory machinery for TLRs on epithelium may represent a promising new avenue for the development of epithelia-specific therapeutic interventions. PMID:19801549

  2. Differentiation capacity of epithelial cells in the sponge Suberites domuncula.

    PubMed

    Schröder, Heinz C; Perović-Ottstadt, Sanja; Wiens, Matthias; Batel, Renato; Müller, Isabel M; Müller, Werner E G

    2004-05-01

    Sponges (phylum Porifera) represent the oldest metazoans. Their characteristic metazoan adhesion molecules and transcription factors enable them to establish a complex "Bauplan"; three major differentiated cell types (epithelial cells, skeletal cells/sclerocytes, and contractile cells) can be distinguished. Since no molecular markers are as yet available to distinguish these somatic cells or the corresponding embryonic cells from which they originate, we have selected the following three genes for their characterization: noggin (a signaling molecule in development), a caspase that encodes an apoptotic molecule, and silicatein. Silicatein is an enzyme that is involved in the synthesis of siliceous spicules and can hence be considered as a marker for scleroblasts. We have used the demosponge Suberites domuncula as a model system. During the hatching of the gemmules (asexual reproduction bodies) of S. domuncula, the expression of both noggin and caspase increases, whereas no transcripts for silicatein can be detected, irrespective of the presence of silicate or ferric iron (Fe3+) in the medium. In contrast, in adult specimens, silicate/Fe3+ cause an increased expression of these genes. In situ analysis has revealed that the first cells that express noggin, caspase, and silicatein lie in the epithelial layer of the pinacoderm. In a later phase, the noggin- and silicatein-positive cells migrate into the mesohyl, where they are found in association with spicules. Thus, the pinacoderm of sponges contains cells that have a differentiating capacity and from which somatic cells, such as skeletal cells/sclerocytes, derive. PMID:15024642

  3. Radiogenic transformation of human mammary epithelial cells in vitro

    NASA Technical Reports Server (NTRS)

    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.

  4. Nucleus Morphometry in Cultured Epithelial Cells Correlates with Phenotype.

    PubMed

    Khan, Ayyad Z; Utheim, Tor P; Jackson, Catherine J; Reppe, Sjur; Lyberg, Torstein; Eidet, Jon R

    2016-06-01

    Phenotype of cultured ocular epithelial transplants has been shown to affect clinical success rates following transplantation to the cornea. The purpose of this study was to evaluate the relationship between cell nucleus morphometry and phenotype in three types of cultured epithelial cells. This study provides knowledge for the development of a non-invasive method of determining the phenotype of cultured epithelium before transplantation. Cultured human conjunctival epithelial cells (HCjE), human epidermal keratinocytes (HEK), and human retinal pigment epithelial cells (HRPE) were analyzed by quantitative immunofluorescence. Assessments of nucleus morphometry and nucleus-to-cytoplasm ratio (N/C ratio) were performed using ImageJ. Spearman's correlation coefficient was employed for statistical analysis. Levels of the proliferation marker PCNA in HCjE, HEK, and HRPE correlated positively with nuclear area. Nuclear area correlated significantly with levels of the undifferentiated cell marker ABCG2 in HCjE. Bmi1 levels, but not p63α levels, correlated significantly with nuclear area in HEK. The N/C ratio did not correlate significantly with any of the immunomarkers in HCjE (ABCG2, CK7, and PCNA) and HRPE (PCNA). In HEK, however, the N/C ratio was negatively correlated with levels of the undifferentiated cell marker CK14 and positively correlated with Bmi1 expression. The size of the nuclear area correlated positively with proliferation markers in all three epithelia. Morphometric indicators of phenotype in cultured epithelia can be identified using ImageJ. Conversely, the N/C ratio did not show a uniform relationship with phenotype in HCjE, HEK, or HRPE. N/C ratio therefore, may not be a useful morphometric marker for in vitro assessment of phenotype in these three epithelia. PMID:27329312

  5. Zinc modulates cytokine-induced lung epithelial cell barrier permeability.

    PubMed

    Bao, Shenying; Knoell, Daren L

    2006-12-01

    Apoptosis plays a causative role in acute lung injury in part due to epithelial cell loss. We recently reported that zinc protects the lung epithelium during inflammatory stress whereas depletion of intracellular zinc enhances extrinsic apoptosis. In this investigation, we evaluated the relationship between zinc, caspase-3, and cell-to-cell contact via proteins that form the adherens junction complex. Cell adhesion proteins are directly responsible for formation of the mechanical barrier of the lung epithelium. We hypothesized that exposure to inflammatory cytokines, in conjunction with zinc deprivation, would induce caspase-3, leading to degradation of junction proteins, loss of cell-to-cell contact, and compromised barrier function. Primary human upper airway and type I/II alveolar epithelial cultures were obtained from multiple donors and exposed to inflammatory stimuli that provoke extrinsic apoptosis in addition to depletion of intracellular zinc. We observed that zinc deprivation combined with tumor necrosis factor-alpha, interferon-gamma, and Fas receptor ligation accelerates caspase-3 activation, proteolysis of E-cadherin and beta-catenin, and cellular apoptosis, leading to increased paracellular leak across monolayers of both upper airway and alveolar lung epithelial cultures. Zinc supplementation inhibited apoptosis and paracellular leak, whereas caspase inhibition was less effective. We conclude that zinc is a vital factor in the lung epithelium that protects against death receptor-mediated apoptosis and barrier dysfunction. Furthermore, our findings suggest that although caspase-3 inhibition reduces lung epithelial apoptosis it does not prevent mechanical dysfunction. These findings facilitate future studies aimed at developing therapeutic strategies to prevent acute lung injury. PMID:16844947

  6. Wound repair: role of immune–epithelial interactions

    PubMed Central

    Leoni, G; Neumann, P-A; Sumagin, R; Denning, TL; Nusrat, A

    2016-01-01

    The epithelium serves as a highly selective barrier at mucosal surfaces. Upon injury, epithelial wound closure is orchestrated by a series of events that emanate from the epithelium itself as well as by the temporal recruitment of immune cells into the wound bed. Epithelial cells adjoining the wound flatten out, migrate, and proliferate to rapidly cover denuded surfaces and re-establish mucosal homeostasis. This process is highly regulated by proteins and lipids, proresolving mediators such as Annexin A1 protein and resolvins released into the epithelial milieu by the epithelium itself and infiltrating innate immune cells including neutrophils and macrophages. Failure to achieve these finely tuned processes is observed in chronic inflammatory diseases that are associated with non-healing wounds. An improved understanding of mechanisms that mediate repair is important in the development of therapeutics aimed to promote mucosal wound repair. PMID:26174765

  7. Borrelia burgdorferi bind to epithelial cell proteoglycans.

    PubMed Central

    Isaacs, R D

    1994-01-01

    Borrelia burgdorferi adhere to mammalian cells in vitro but neither the ligand(s) nor the receptor(s) has (have) been clearly established. Using an in vitro attachment-inhibition assay, a B. burgdorferi attachment mechanism has been identified. Heparin, heparan sulfate, and dermatan sulfate reduced the attachment of virulent B. burgdorferi strain 297 to HeLa cells by approximately 60%. In addition, virulent, but not avirulent, B. burgdorferi strains B31, N40, and HB19 demonstrated heparin attachment-inhibition. Attachment to Chinese hamster ovary cells deficient in heparan sulfate proteoglycans was reduced by 68% compared to attachment to wild-type cells and was identical to attachment at maximum heparin inhibition to the wild-type cells. Pretreatment of HeLa cell monolayers with heparitinase, heparinase, and chondroitinase ABC, but not with chondroitinase AC, reduced borrelial attachment by approximately 50%. A moderately high affinity, low copy number, promiscuous B. burgdorferi glycosaminoglycan receptor was demonstrated by equilibrium binding studies. A 39-kD polypeptide, purified by heparin affinity chromatography from Triton X-100 extracts derived from virulent borrelia, was a candidate for this receptor. These studies indicate that one mode of B. burgdorferi attachment to eukaryotic cells