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Sample records for mucosal epithelial barrier

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

    PubMed

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

    2016-10-01

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

  2. 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

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

    PubMed

    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.

  4. HIV and mucosal barrier interactions: consequences for transmission and pathogenesis.

    PubMed

    Burgener, Adam; McGowan, Ian; Klatt, Nichole R

    2015-10-01

    The mucosal barrier plays an integral function in human health as it is the primary defense against pathogens, and provides a critical transition between the external environment and the human internal body. In the context of HIV infection, the most relevant mucosal surfaces include those of the gastrointestinal (GI) and genital tract compartments. Several components help maintain the effectiveness of this mucosal surface, including the physical anatomy of the barrier, cellular immunity, soluble factors, and interactions between the epithelial barrier and the local microenvironment, including mucus and host microbiota. Any defects in barrier integrity or function can rapidly lead to an increase in acquisition risk, or with established infection may result in increased pathogenesis, morbidities, or mortality. Indeed, a key feature to all aspects of HIV infection from transmission to pathogenesis is disruption and/or dysfunction of mucosal barriers. Herein, we will detail the host-pathogen relationship of HIV and mucosal barriers in both of these scenarios.

  5. 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

  6. The Anti-Inflammatory Activity of Curcumin Protects the Genital Mucosal Epithelial Barrier from Disruption and Blocks Replication of HIV-1 and HSV-2

    PubMed Central

    Ferreira, Victor H.; 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

  7. Cytoskeletal Regulation of Epithelial Barrier Function During Inflammation

    PubMed Central

    Ivanov, Andrei I.; Parkos, Charles A.; Nusrat, Asma

    2010-01-01

    Increased epithelial permeability is a common and important consequence of mucosal inflammation that results in perturbed body homeostasis and enhanced exposure to external pathogens. The integrity and barrier properties of epithelial layers are regulated by specialized adhesive plasma membrane structures known as intercellular junctions. It is generally believed that inflammatory stimuli increase transepithelial permeability by inducing junctional disassembly. This review highlights molecular events that lead to disruption of epithelial junctions during inflammation. We specifically focus on key mechanisms of junctional regulation that are dependent on reorganization of the perijunctional F-actin cytoskeleton. We discuss critical roles of myosin-II–dependent contractility and actin filament turnover in remodeling of the F-actin cytoskeleton that drive disruption of epithelial barriers under different inflammatory conditions. Finally, we highlight signaling pathways induced by inflammatory mediators that regulate reorganization of actin filaments and junctional disassembly in mucosal epithelia. PMID:20581053

  8. Helminthic therapy: improving mucosal barrier function

    PubMed Central

    Wolff, Martin J.; Broadhurst, Mara J.

    2014-01-01

    The epidemiology of autoimmune diseases and helminth infections led to suggestions that helminths could improve inflammatory conditions, which was then tested using animal models. This has translated to clinical investigations aimed at the safe and controlled reintroduction of helminthic exposure to patients suffering from autoimmune diseases (so-called “helminthic therapy”) in an effort to mitigate the inflammatory response. In this review, we will summarize the results of recent clinical trials of helminthic therapy, with particular attention to mechanisms of action. Whereas previous reviews have emphasized immune regulatory mechanisms activated by helminths, we propose that enhancement of mucosal barrier function may have an equally important role in improving conditions of inflammatory bowel diseases. PMID:22464690

  9. Physiology and immunology of mucosal barriers in catfish (Ictalurus spp.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mucosal barriers of catfish (Ictalurus spp.) constitute the first line of defense against pathogen invasion while simultaneously carrying out a diverse array of other critical physiological processes, including nutrient adsorption, osmoregulation, waste excretion, and environmental sensing. Catf...

  10. Epithelial barrier and oral bacterial infection.

    PubMed

    Groeger, Sabine E; Meyle, Joerg

    2015-10-01

    The oral epithelial barrier separates the host from the environment and provides the first line of defense against pathogens, exogenous substances and mechanical stress. It consists of underlying connective tissue and a stratified keratinized epithelium with a basement membrane, whose cells undergo terminal differentiation resulting in the formation of a mechanically resistant surface. Gingival keratinocytes are connected by various transmembrane proteins, such as tight junctions, adherens junctions and gap junctions, each of which has a specialized structure and specific functions. Periodontal pathogens are able to induce inflammatory responses that lead to attachment loss and periodontal destruction. A number of studies have demonstrated that the characteristics of pathogenic oral bacteria influence the expression and structural integrity of different cell-cell junctions. Tissue destruction can be mediated by host cells following stimulation with cytokines and bacterial products. Keratinocytes, the main cell type in gingival epithelial tissues, express a variety of proinflammatory cytokines and chemokines, including interleukin-1alpha, interleukin-1beta, interleukin-6, interleukin-8 and tumor necrosis factor-alpha. Furthermore, the inflammatory mediators that may be secreted by oral keratinocytes are vascular endothelial growth factor, prostaglandin E2 , interleukin-1 receptor antagonist and chemokine (C-C motif) ligand 2. The protein family of matrix metalloproteinases is able to degrade all types of extracellular matrix protein, and can process a number of bioactive molecules. Matrix metalloproteinase activities under inflammatory conditions are mostly deregulated and often increased, and those mainly relevant in periodontal disease are matrix metalloproteinases 1, 2, 3, 8, 9, 13 and 24. Viral infection may also influence the epithelial barrier. Studies show that the expression of HIV proteins in the mucosal epithelium is correlated with the disruption of

  11. [Recent studies on corneal epithelial barrier function].

    PubMed

    Liu, F F; Li, W; Liu, Z G; Chen, W S

    2016-08-01

    Corneal epithelium, the outermost layer of eyeball, is the main route for foreign materials to enter the eye. Under physiological conditions, the corneal epithelial superficial cells form a functionally selective permeability barrier. Integral corneal epithelial barrier function not only ensures the enrolling of nutrients which is required for regular metabolism, but also prevents foreign bodies, or disease-causing microorganism invasion. Recently, a large number of clinical and experimental studies have shown that abnormal corneal epithelial barrier function is the pathological basis for many ocular diseases. In addition, some study found that corneal epithelial barrier constitutes a variety of proteins involved in cell proliferation, differentiation, apoptosis, and a series of physiological and pathological processes. This paper reviewed recent studies specifically on the corneal epithelial barrier, highlights of its structure, function and influence factors. (Chin J Ophthalmol, 2016, 52: 631-635). PMID:27562284

  12. Physiology and immunology of mucosal barriers in catfish (Ictalurus spp.)

    PubMed Central

    Peatman, Eric; Lange, Miles; Zhao, Honggang; Beck, Benjamin H

    2015-01-01

    The mucosal barriers of catfish (Ictalurus spp) constitute the first line of defense against pathogen invasion while simultaneously carrying out a diverse array of other critical physiological processes, including nutrient adsorption, osmoregulation, waste excretion, and environmental sensing. Catfish depend more heavily on mucosal barriers than their terrestrial counterparts as they are continuously interacting with the aquatic microbiota. Our understanding of these barriers, while growing, is still limited relative to that of mammalian model systems. Nevertheless, a combination of molecular and cellular studies in catfish over the last few decades, and particularly within the last few years, has helped to elucidate many of the primary actors and pathways critical to their mucosal health. Here we describe aspects of innate and adaptive immune responses in the primary mucosal tissues (skin, gill, and intestine) of catfish, focusing on mucus-driven responses, pathogen recognition, soluble mediators, and immunoglobulin and T-cell derived immunity. Modulation of mucosal barriers will be critical moving forward for crafting better diets, improving vaccine delivery, enhancing water quality, and ensuring sustainable production practices in catfish. PMID:26716071

  13. Epithelial barrier function: at the frontline of asthma immunology and allergic airway inflammation

    PubMed Central

    Georas, Steve N.; Rezaee, Fariba

    2014-01-01

    Airway epithelial cells form a barrier to the outside world, and are at the frontline of mucosal immunity. Epithelial apical junctional complexes are multi-protein subunits that promote cell-cell adhesion and barrier integrity. Recent studies in the skin and GI tract suggest that disruption of cell-cell junctions is required to initiate epithelial immune responses, but how this applies to mucosal immunity in the lung is not clear. Increasing evidence indicates that defective epithelial barrier function is a feature of airway inflammation in asthma. One challenge in this area is that barrier function and junctional integrity are difficult to study in the intact lung, but innovative approaches should provide new knowledge in this area in the near future. In this article, we review the structure and function of epithelial apical junctional complexes, emphasizing how regulation of the epithelial barrier impacts innate and adaptive immunity. We discuss why defective epithelial barrier function may be linked to Th2 polarization in asthma, and propose a rheostat model of barrier dysfunction that implicates the size of inhaled allergen particles as an important factor influencing adaptive immunity. PMID:25085341

  14. Human immunodeficiency virus-associated disruption of mucosal barriers and its role in HIV transmission and pathogenesis of HIV/AIDS disease.

    PubMed

    Tugizov, Sharof

    2016-01-01

    Oral, intestinal and genital mucosal epithelia have a barrier function to prevent paracellular penetration by viral, bacterial and other pathogens, including human immunodeficiency virus (HIV). HIV can overcome these barriers by disrupting the tight and adherens junctions of mucosal epithelia. HIV-associated disruption of epithelial junctions may also facilitate paracellular penetration and dissemination of other viral pathogens. This review focuses on possible molecular mechanisms of HIV-associated disruption of mucosal epithelial junctions and its role in HIV transmission and pathogenesis of HIV and acquired immune deficiency syndrome (AIDS). PMID:27583187

  15. How do polymeric micelles cross epithelial barriers?

    PubMed

    Pepić, Ivan; Lovrić, Jasmina; Filipović-Grčić, Jelena

    2013-09-27

    Non-parenteral delivery of drugs using nanotechnology-based delivery systems is a promising non-invasive way to achieve effective local or systemic drug delivery. The efficacy of drugs administered non-parenterally is limited by their ability to cross biological barriers, and epithelial tissues particularly present challenges. Polymeric micelles can achieve transepithelial drug delivery because of their ability to be internalized into cells and/or cross epithelial barriers, thereby delivering drugs either locally or systematically following non-parenteral administration. This review discusses the particular characteristics of various epithelial barriers and assesses their potential as non-parenteral routes of delivery. The material characteristics of polymeric micelles (e.g., size, surface charge, and surface decoration) and of unimers dissociated from polymeric micelles determine their interactions (non-specific and/or specific) with mucus and epithelial cells as well as their intracellular fate. This paper outlines the mechanisms governing the major modes of internalization of polymeric micelles into epithelial cells, with an emphasis on specific recent examples of the transport of drug-loaded polymeric micelles across epithelial barriers.

  16. Epithelial IL-18 Equilibrium Controls Barrier Function in Colitis

    PubMed Central

    Nowarski, Roni; Jackson, Ruaidhrí; Gagliani, Nicola; de Zoete, Marcel R.; Palm, Noah W.; Bailis, Will; Low, Jun Siong; Harman, Christian C.D.; Graham, Morven; Elinav, Eran; Flavell, Richard A.

    2016-01-01

    SUMMARY The intestinal mucosal barrier controlling the resident microbiome is dependent on a protective mucus layer generated by goblet cells, impairment of which is a hallmark of the inflammatory bowel disease Ulcerative Colitis. Here we show that IL-18 is critical in driving the pathologic breakdown of barrier integrity in a model of colitis. Deletion of Il18 or its receptor Il18r1 in intestinal epithelial cells (Δ/EC) conferred protection from colitis and mucosal damage in mice. In contrast, deletion of the IL-18 negative regulator Il18bp resulted in severe colitis associated with loss of mature goblet cells. Colitis and goblet cell loss were rescued in Il18bp−/−;Il18rΔ/EC mice, demonstrating that colitis severity is controlled at the level of IL-18 signaling in intestinal epithelial cells. IL-18 inhibited goblet cell maturation by regulating the transcriptional program instructing goblet cell development. These results inform on the mechanism of goblet cell dysfunction which underlies the pathology of Ulcerative Colitis. PMID:26638073

  17. Helicobacter pylori: a pathogenic threat to the gastric mucosal barrier.

    PubMed

    Oluwole, F S

    2015-12-01

    Peptic ulcer disease is a multi-factorial disorder of the gastrointestinal tract with a global prevalence affecting about 4.6 million people annually and having a mortality of one death per 10,000 cases. Helicobacter pylori (H. pylori) plays a profound role in the pathogenesis of chronic gastritis, peptic ulcer, including gastric mucosa-associated lymphoid tissue and carcinoma. Any compromise to the gastric mucosal barrier will greatly affect the integrity of the stomach. H. pylori is an organism which mediates a compromise of the gastric mucosal barrier by stimulating increased gastric acid secretion, causing alteration of certain immune factors, penetration of the mucosal layer and provoking persistent inflammation even without invading the mucus membrane. All the different lines of therapy have not shown maximal efficacy in the eradication/cure of the infection in patients. Consequently, alternative therapies including phytomedicines and probiotics have been introduced both in the quest for better eradication therapies and in addressing the problem of H. pylori relapse. In the light of the increasing antibiotic resistance associated with current therapies, the use of herbal preparations or its concomitant use with current therapy has the potential to contribute additive and synergistic effect in the eradication of the H. pylori infection. This review highlights the anti-H. pylori herbal preparations tested and in current use.

  18. Helicobacter pylori: a pathogenic threat to the gastric mucosal barrier.

    PubMed

    Oluwole, F S

    2015-12-01

    Peptic ulcer disease is a multi-factorial disorder of the gastrointestinal tract with a global prevalence affecting about 4.6 million people annually and having a mortality of one death per 10,000 cases. Helicobacter pylori (H. pylori) plays a profound role in the pathogenesis of chronic gastritis, peptic ulcer, including gastric mucosa-associated lymphoid tissue and carcinoma. Any compromise to the gastric mucosal barrier will greatly affect the integrity of the stomach. H. pylori is an organism which mediates a compromise of the gastric mucosal barrier by stimulating increased gastric acid secretion, causing alteration of certain immune factors, penetration of the mucosal layer and provoking persistent inflammation even without invading the mucus membrane. All the different lines of therapy have not shown maximal efficacy in the eradication/cure of the infection in patients. Consequently, alternative therapies including phytomedicines and probiotics have been introduced both in the quest for better eradication therapies and in addressing the problem of H. pylori relapse. In the light of the increasing antibiotic resistance associated with current therapies, the use of herbal preparations or its concomitant use with current therapy has the potential to contribute additive and synergistic effect in the eradication of the H. pylori infection. This review highlights the anti-H. pylori herbal preparations tested and in current use. PMID:27462690

  19. Mucosal barrier injury, fever and infection in neutropenic patients with cancer: introducing the paradigm febrile mucositis.

    PubMed

    van der Velden, Walter J F M; Herbers, Alexandra H E; Netea, Mihai G; Blijlevens, Nicole M A

    2014-11-01

    Infection remains one of the most prominent complications after cytotoxic treatment for cancer. The connection between neutropenia and both infections and fever has long been designated as 'febrile neutropenia', but treatment with antimicrobial agents and haematopoietic growth factors has failed to significantly reduce its incidence. Moreover, emerging antimicrobial resistance is becoming a concern that necessitates the judicious use of available antimicrobial agents. In addition to neutropenia, patients who receive cytotoxic therapy experience mucosal barrier injury (MBI) or 'mucositis'. MBI creates a port-de-entrée for resident micro-organisms to cause blood stream infections and contributes directly to the occurrence of fever by disrupting the highly regulated host-microbe interactions, which, even in the absence of an infection, can result in strong inflammatory reactions. Indeed, MBI has been shown to be a pivotal factor in the occurrence of inflammatory complications after cytotoxic therapy. Hence, the concept 'febrile neutropenia' alone may no longer suffice and a new concept 'febrile mucositis' should be recognized as the two are at least complementary. This review we summarizes the existing evidence for both paradigms and proposes new therapeutic approaches to tackle the perturbed host-microbe interactions arising from cytotoxic therapy-induced tissue damage in order to reduce fever in neutropenic patients with cancer.

  20. Restoring airway epithelial barrier dysfunction: a new therapeutic challenge in allergic airway disease.

    PubMed

    Steelant, B; Seys, S F; Boeckxstaens, G; Akdis, C A; Ceuppens, J L; Hellings, P W

    2016-09-01

    An intact functional mucosal barrier is considered to be crucial for the maintenance of airway homeostasis as it protects the host immune system from exposure to allergens and noxious environmental triggers. Recent data provided evidence for the contribution of barrier dysfunction to the development of inflammatory diseases in the airways, skin and gut. A defective barrier has been documented in chronic rhinosinusitis, allergic rhinitis, asthma, atopic dermatitis and inflammatory bowel diseases. However, it remains to be elucidated to what extent primary (genetic) versus secondary (inflammatory) mechanisms drive barrier dysfunction. The precise pathogenesis of barrier dysfunction in patients with chronic mucosal inflammation and its implications on tissue inflammation and systemic absorption of exogenous particles are only partly understood. Since epithelial barrier defects are linked with chronicity and severity of airway inflammation, restoring the barrier integrity may become a useful approach in the treatment of allergic diseases. We here provide a state-of-the-art review on epithelial barrier dysfunction in upper and lower airways as well as in the intestine and the skin and on how barrier dysfunction can be restored from a therapeutic perspective.

  1. 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

  2. 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

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

    PubMed

    Hammad, Hamida; Lambrecht, Bart N

    2015-07-21

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

  4. Lactobacillus protects the integrity of intestinal epithelial barrier damaged by pathogenic bacteria

    PubMed Central

    Yu, Qinghua; Yuan, Lixia; Deng, Jun; Yang, Qian

    2015-01-01

    Pathogens invade intestinal mucosal barrier through phagocytosis of antigen presenting cells (dendritic cell, microfold cells), or through the invasion into the intestinal epithelial directly. Some pathogens could damage the cell junction between epithelial cells and use the paracellular pathway as an entrance to invade. Moreover, some Lactobacillus could inhibit the adhesion of the pathogens and protect the integrity of the cell junction and mucosal barrier. This research focused on the potential therapeutic effect of Lactobacillus fructosus (L. fructosus) C2 to attenuate ETEC K88 or S. typhimurium SL1344 induced changes to mucosal barrier. The results demonstrated that treatment of polarized Caco-2 cells with L. fructosus C2 reduced the permeation of dextran, and expression of IL-8, p-ERK, and p-JNK when cells were infected with pathogenic bacteria. The findings indicated that L. fructosus C2 exerted a protective effect against the damage to the integrity of Caco-2 cells by ETEC or S. typhimurium infection. PMID:25859435

  5. Nonmuscle Myosin IIA Regulates Intestinal Epithelial Barrier in vivo and Plays a Protective Role During Experimental Colitis.

    PubMed

    Naydenov, Nayden G; Feygin, Alex; Wang, Dongdong; Kuemmerle, John F; Harris, Gianni; Conti, Mary Anne; Adelstein, Robert S; Ivanov, Andrei I

    2016-01-01

    The actin cytoskeleton is a critical regulator of intestinal mucosal barrier permeability, and the integrity of epithelial adherens junctions (AJ) and tight junctions (TJ). Non muscle myosin II (NM II) is a key cytoskeletal motor that controls actin filament architecture and dynamics. While NM II has been implicated in the regulation of epithelial junctions in vitro, little is known about its roles in the intestinal mucosa in vivo. In this study, we generated a mouse model with an intestinal epithelial-specific knockout of NM IIA heavy chain (NM IIA cKO) and examined the structure and function of normal gut barrier, and the development of experimental colitis in these animals. Unchallenged NM IIA cKO mice showed increased intestinal permeability and altered expression/localization of several AJ/TJ proteins. They did not develop spontaneous colitis, but demonstrated signs of a low-scale mucosal inflammation manifested by prolapses, lymphoid aggregates, increased cytokine expression, and neutrophil infiltration in the gut. NM IIA cKO animals were characterized by a more severe disruption of the gut barrier and exaggerated mucosal injury during experimentally-induced colitis. Our study provides the first evidence that NM IIA plays important roles in establishing normal intestinal barrier, and protection from mucosal inflammation in vivo. PMID:27063635

  6. Nonmuscle Myosin IIA Regulates Intestinal Epithelial Barrier in vivo and Plays a Protective Role During Experimental Colitis

    PubMed Central

    Naydenov, Nayden G.; Feygin, Alex; Wang, Dongdong; Kuemmerle, John F.; Harris, Gianni; Conti, Mary Anne; Adelstein, Robert S.; Ivanov, Andrei I.

    2016-01-01

    The actin cytoskeleton is a critical regulator of intestinal mucosal barrier permeability, and the integrity of epithelial adherens junctions (AJ) and tight junctions (TJ). Non muscle myosin II (NM II) is a key cytoskeletal motor that controls actin filament architecture and dynamics. While NM II has been implicated in the regulation of epithelial junctions in vitro, little is known about its roles in the intestinal mucosa in vivo. In this study, we generated a mouse model with an intestinal epithelial-specific knockout of NM IIA heavy chain (NM IIA cKO) and examined the structure and function of normal gut barrier, and the development of experimental colitis in these animals. Unchallenged NM IIA cKO mice showed increased intestinal permeability and altered expression/localization of several AJ/TJ proteins. They did not develop spontaneous colitis, but demonstrated signs of a low-scale mucosal inflammation manifested by prolapses, lymphoid aggregates, increased cytokine expression, and neutrophil infiltration in the gut. NM IIA cKO animals were characterized by a more severe disruption of the gut barrier and exaggerated mucosal injury during experimentally-induced colitis. Our study provides the first evidence that NM IIA plays important roles in establishing normal intestinal barrier, and protection from mucosal inflammation in vivo. PMID:27063635

  7. Hydrophobicity of mucosal surface and its relationship to gut barrier function.

    PubMed

    Qin, Xiaofa; Caputo, Francis J; Xu, Da-Zhong; Deitch, Edwin A

    2008-03-01

    Loss of the gut barrier has been implicated in the pathogenesis of the multiple organ dysfunction syndrome, and, thus, understanding the intestinal barrier is of potential clinical importance. An important, but relatively neglected, component of the gut barrier is the unstirred mucus layer, which through its hydrophobic and other properties serves as an important barrier to bacterial and other factors within the gut lumen. Thus, the goal of this study was to establish a reproducible method of measuring mucosal hydrophobicity and test the hypothesis that conditions that decrease mucosal hydrophobicity are associated with increased gut permeability. Hydrophobicity was measured in various segments of normal gut by measuring the contact angle of an aqueous droplet placed on the mucosal surface using a commercial goniometer. Second, the effect of the mucolytic agent N-acetyl cysteine on mucosal hydrophobicity and gut permeability was measured, as was the effects of increasing periods of in vivo gut ischemia on these parameters. Gut ischemia was induced by superior mesenteric artery occlusion, and gut permeability was measured by the mucosal-to-serosal passage of fluoresceine isothiocyanate-dextran (4.3 kDa) (FD4) across the everted sacs of ileum. Intestinal mucosal hydrophobicity showed a gradual increase from the duodenum to the end of the ileum and remained at high level in the cecum, colon, and rectum. Both N-acetyl cysteine treatment and ischemia caused a dose-dependent decrease in mucosal hydrophobicity, which significantly correlated increased gut permeability. Mucosal hydrophobicity of the intestine can be reproducibly measured, and decreases in mucosal hydrophobicity closely correlate with increased gut permeability. These results suggest that mucosal hydrophobicity can be a reliable method of measuring the barrier function of the unstirred mucus layer and a useful parameter in evaluating the pathogenesis of gut barrier dysfunction. PMID:17693944

  8. Hydrophobicity of mucosal surface and its relationship to gut barrier function.

    PubMed

    Qin, Xiaofa; Caputo, Francis J; Xu, Da-Zhong; Deitch, Edwin A

    2008-03-01

    Loss of the gut barrier has been implicated in the pathogenesis of the multiple organ dysfunction syndrome, and, thus, understanding the intestinal barrier is of potential clinical importance. An important, but relatively neglected, component of the gut barrier is the unstirred mucus layer, which through its hydrophobic and other properties serves as an important barrier to bacterial and other factors within the gut lumen. Thus, the goal of this study was to establish a reproducible method of measuring mucosal hydrophobicity and test the hypothesis that conditions that decrease mucosal hydrophobicity are associated with increased gut permeability. Hydrophobicity was measured in various segments of normal gut by measuring the contact angle of an aqueous droplet placed on the mucosal surface using a commercial goniometer. Second, the effect of the mucolytic agent N-acetyl cysteine on mucosal hydrophobicity and gut permeability was measured, as was the effects of increasing periods of in vivo gut ischemia on these parameters. Gut ischemia was induced by superior mesenteric artery occlusion, and gut permeability was measured by the mucosal-to-serosal passage of fluoresceine isothiocyanate-dextran (4.3 kDa) (FD4) across the everted sacs of ileum. Intestinal mucosal hydrophobicity showed a gradual increase from the duodenum to the end of the ileum and remained at high level in the cecum, colon, and rectum. Both N-acetyl cysteine treatment and ischemia caused a dose-dependent decrease in mucosal hydrophobicity, which significantly correlated increased gut permeability. Mucosal hydrophobicity of the intestine can be reproducibly measured, and decreases in mucosal hydrophobicity closely correlate with increased gut permeability. These results suggest that mucosal hydrophobicity can be a reliable method of measuring the barrier function of the unstirred mucus layer and a useful parameter in evaluating the pathogenesis of gut barrier dysfunction.

  9. Evaluation of tumorigenic risk of tissue-engineered oral mucosal epithelial cells by using combinational examinations.

    PubMed

    Thépot, A; Morel, A P; Justin, V; Desanlis, A; Thivillier, L; Hoffman, E; Till, M; Accardi, R; Tommasino, M; Breton, P; Hainaut, P; Damour, O

    2010-01-01

    Recently, oral mucosal epithelial cells were proposed as a cell source of the autologous cell transplant therapy for corneal trauma or disease. The question addressed is to know if the biological conditions of grafting could induce certain cellular, molecular, and genetic alterations that might increase the risk of mutations and possibly of cellular transformation. Recent progress in cancer research enables us to depict the generation mechanisms and basic characteristics of human cancer cells from molecular, cytological, and biological aspects. The aim of this study is to evaluate the risk of tumorigenicity of the oral mucosal epithelial culture process in order to mitigate that risk, if any, before clinical application. Oral mucosal epithelial cells from three different human donors were investigated by combinational examinations to detect possible tumorigenic transformation. We investigated (i) clonogenic and karyology types, (ii) the validation of proliferation rate, (iii) the epithelial-mesenchymal transition, (iv) anchorage-independent growth potential, and (v) tumorigenicity on nude mice. Results show that the culture process used in this study presents no risk of tumorigenicity. PMID:20977830

  10. Entamoeba histolytica contains an occludin-like protein that can alter colonic epithelial barrier function.

    PubMed

    Goplen, Michael; Lejeune, Manigandan; Cornick, Steve; Moreau, France; Chadee, Kris

    2013-01-01

    The exact mechanism by which Entamoeba histolytica disrupts the human colonic epithelium and invades the mucosa has yet to be clearly elucidated. E. histolytica produces a diverse array of putative virulent factors such as glycosidase, cysteine proteinases and amebapore that can modulate and/or disrupt epithelial barrier functions. However, it is currently thought that E. histolytica produces numerous other molecules and strategies to disrupt colonic mucosal defenses. In this study, we document a putative mechanism whereby the parasite alters the integrity of human epithelium by expressing a cognate tight junction protein of the host. We detected this protein as "occludin-like" as revealed by immunoblotting and immunoprecipitation studies and visualization by confocal microscopy using antibodies highly specific for human occludin. We propose that E. histolytica occludin-like protein might displace mucosal epithelial occludin-occludin tight junction interactions resulting in epithelial disruption analogous to sub mucosal human dendritic cells sampling luminal contents. These results indicate that E. histolytica occludin is a putative virulent component that can play a role in the pathogenesis of intestinal amebiasis.

  11. Combined probiotic bacteria promotes intestinal epithelial barrier function in interleukin-10-gene-deficient mice

    PubMed Central

    Shi, Chen-Zhang; Chen, Hong-Qi; Liang, Yong; Xia, Yang; Yang, Yong-Zhi; Yang, Jun; Zhang, Jun-Dong; Wang, Shu-Hai; Liu, Jing; Qin, Huan-Long

    2014-01-01

    AIM: To investigate the protective effects of combinations of probiotic (Bifico) on interleukin (IL)-10-gene-deficient (IL-10 KO) mice and Caco-2 cell monolayers. METHODS: IL-10 KO mice were used to assess the benefits of Bifico in vivo. IL-10 KO and control mice received approximately 1.5 × 108 cfu/d of Bifico for 4 wk. Colons were then removed and analyzed for epithelial barrier function by Ussing Chamber, while an ELISA was used to evaluate proinflammatory cytokines. The colon epithelial cell line, Caco-2, was used to test the benefit of Bifico in vitro. Enteroinvasive Escherichia coli (EIEC) and the probiotic mixture Bifico, or single probiotic strains, were applied to cultured Caco-2 monolayers. Barrier function was determined by measuring transepithelial electrical resistance and tight junction protein expression. RESULTS: Treatment of IL-10 KO mice with Bifico partially restored body weight, colon length, and epithelial barrier integrity to wild-type levels. In addition, IL-10 KO mice receiving Bifico treatment had reduced mucosal secretion of tumor necrosis factor-α and interferon-γ, and attenuated colonic disease. Moreover, treatment of Caco-2 monolayers with Bifico or single-strain probiotics in vitro inhibited EIEC invasion and reduced the secretion of proinflammatory cytokines. CONCLUSION: Bifico reduced colon inflammation in IL-10 KO mice, and promoted and improved epithelial-barrier function, enhanced resistance to EIEC invasion, and decreased proinflammatory cytokine secretion. PMID:24782616

  12. 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.

  13. 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

  14. Gut barrier structure, mucosal immunity and intestinal microbiota in the pathogenesis and treatment of HIV infection.

    PubMed

    Tincati, Camilla; Douek, Daniel C; Marchetti, Giulia

    2016-01-01

    Over the past 10 years, extensive work has been carried out in the field of microbial translocation in HIV infection, ranging from studies on its clinical significance to investigations on its pathogenic features. In the present work, we review the most recent findings on this phenomenon, focusing on the predictive role of microbial translocation in HIV-related morbidity and mortality, the mechanisms by which it arises and potential therapeutic approaches. From a clinical perspective, current work has shown that markers of microbial translocation may be useful in predicting clinical events in untreated HIV infection, while conflicting data exist on their role in cART-experienced subjects, possibly due to the inclusion of extremely varied patient populations in cohort studies. Results from studies addressing the pathogenesis of microbial translocation have improved our knowledge of the damage of the gastrointestinal epithelial barrier occurring in HIV infection. However, the extent to which mucosal impairment translates directly to increased gastrointestinal permeability remains an open issue. In this respect, novel work has established a role for IL-17 and IL-22-secreting T cell populations in limiting microbial translocation and systemic T-cell activation/inflammation, thus representing a possible target of immune-therapeutic interventions shown to be promising in the animal model. Further, recent reports have not only confirmed the presence of a dysbiotic intestinal community in the course of HIV infection but have also shown that it may be linked to mucosal damage, microbial translocation and peripheral immune activation. Importantly, technical advances have also shed light on the metabolic activity of gut microbes, highlighting the need for novel therapeutic approaches to correct the function, as well as the composition, of the gastrointestinal microbiota. PMID:27073405

  15. 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.

  16. The mucosal T cell integrin alpha M290 beta 7 recognizes a ligand on mucosal epithelial cell lines.

    PubMed

    Roberts, K; Kilshaw, P J

    1993-07-01

    The integrin alpha M290 beta 7 is expressed at high levels on mucosal T cells, particularly on those within the epithelium of the gut. We now report that a mouse T cell hybridoma, MTC-1, with similar surface expression of this molecule, adhered strongly to cells of the mouse rectal carcinoma line CMT93 and that adhesion was blocked completely by the monoclonal antibody (mAb) M290. Other mAb to the alpha M290 or beta 7 subunits had little or no inhibitory effect. M290 also inhibited adhesion of the hybridoma to cells of the mouse lung carcinomas CTM64/61 and KLN205 but had little or no effect on adhesion to seven other mouse epithelial cell lines or to the human colon carcinoma line, HT29. Intraepithelial lymphocytes (IEL) isolated from the small intestine of BALB/c mice displayed potent T cell receptor-dependent cytotoxic effector function against CMT93 in the presence of low concentrations of Phytolacca americana lectin. This cytotoxic activity also was inhibited by the M290 mAb. Treatment of CMT93 cells with tumor necrosis factor-alpha and interferon-gamma induced expression de novo of ICAM-1 and reduced the inhibitory effect of M290 in tests both for adhesion and cytotoxicity. In further experiments cytotoxic activity of IEL against the mastocytoma P815 was investigated. This target cell was considered not to possess a ligand for the integrin. In this case cytotoxic effector function was triggered by anti-CD3 mAb and, in contrast to results with CMT93, target cell lysis was increased in the presence of M290 and other antibodies to the integrin, suggesting a co-stimulatory effect. These results show that alpha M290 beta 7 recognizes a ligand on the surface of certain epithelial cell lines. Further, they provide the first clear indication that this integrin may play an important role in functional interactions between T cells and the mucosal epithelium.

  17. 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.

  18. Intestinal epithelial apoptosis initiates gut mucosal injury during extracorporeal membrane oxygenation in the newborn piglet.

    PubMed

    MohanKumar, Krishnan; Killingsworth, Cheryl R; McIlwain, R Britt; Timpa, Joseph G; Jagadeeswaran, Ramasamy; Namachivayam, Kopperuncholan; Kurundkar, Ashish R; Kelly, David R; Garzon, Steven A; Maheshwari, Akhil

    2014-02-01

    Neonates and young infants exposed to extracorporeal circulation during extracorporeal membrane oxygenation (ECMO) and cardiopulmonary bypass are at risk of developing a systemic inflammatory response syndrome with multi-organ dysfunction. We used a piglet model of ECMO to investigate the hypothesis that epithelial apoptosis is an early event that precedes villous damage during ECMO-related bowel injury. Healthy 3-week-old piglets were subjected to ECMO for up to 8 h. Epithelial apoptosis was measured in histopathological analysis, nuclear imaging, and terminal deoxynucleotidyl transferase dUTP nick end labeling. Plasma intestinal fatty acid-binding protein (I-FABP) levels were measured by enzyme immunoassay. Intestinal mast cells were isolated by fluorescence-assisted cell sorting. Cleaved caspase-8, caspase-9, phospho-p38 MAPK, and fas ligand expression were investigated by immunohistochemistry, western blots, and reverse transcriptase-quantitative PCR. Piglet ECMO was associated with increased gut epithelial apoptosis. Extensive apoptotic changes were noted on villus tips and in scattered crypt cells after 2 h of ECMO. After 8 h, the villi were denuded and apoptotic changes were evident in a majority of crypt cells. Increased circulating I-FABP levels, a marker of gut epithelial injury, showed that epithelial injury occurred during ECMO. We detected increased cleaved caspase-8, but not cleaved caspase-9, in epithelial cells indicating that the extrinsic apoptotic pathway was active. ECMO was associated with increased fas ligand expression in intestinal mast cells, which was induced through activation of the p38 mitogen-activated protein kinase. We conclude that epithelial apoptosis is an early event that initiates gut mucosal injury in a piglet model of ECMO.

  19. Methionine restriction fundamentally supports health by tightening epithelial barriers.

    PubMed

    Mullin, James M; Skrovanek, Sonja M; Ramalingam, Arivudainambi; DiGuilio, Katherine M; Valenzano, Mary C

    2016-01-01

    Dietary methionine restriction (MR) has been found to affect one of the most primary tissue-level functions of an organism: the efficiency with which the epithelial linings of major organs separate the fluid compartments that they border. This process, epithelial barrier function, is basic for proper function of all organs, including the lung, liver, gastrointestinal tract, reproductive tract, blood-brain barrier, and kidney. Specifically, MR has been found to modify the protein composition of tight junctional complexes surrounding individual epithelial cells in a manner that renders the complexes less leaky. This has been observed in both a renal epithelial cell culture model and in gastrointestinal tissue. In both cases, MR increased the transepithelial electrical resistance across the epithelium, while decreasing passive leak of small nonelectrolytes. However, the specific target protein modifications involved were unique to each case. Overall, this provides an example of the primary level on which MR functions to modify, and improve, an organism.

  20. 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

  1. 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

  2. CaMKII regulates the strength of the epithelial barrier

    PubMed Central

    Shiomi, Ryo; Shigetomi, Kenta; Inai, Tetsuichiro; Sakai, Masami; Ikenouchi, Junichi

    2015-01-01

    Epithelial cells define the boundary between the outside and the inside of our body by constructing the diffusion barrier. Tight junctions (TJs) of epithelial cells function as barriers against invasion of harmful microorganisms into the human body and free diffusion of water or ions from the body. Therefore, formation of TJs has to be strictly controlled in epithelial cells. However, the molecular mechanisms governing this regulation are largely unknown. In this study, we identified Ca2+/calmodulin-dependent protein kinase II (CaMKII) as a regulator of the barrier function of TJs. CaMKII inhibition led to enlargement of TJ-areas and up-regulation of the barrier function. CaMKII inhibition induced excess TJ formation in part by the activation of AMP-activated protein kinase (AMPK) and subsequent phosphorylation of claudin-1. As up-regulation of epithelial barriers is essential for the prevention of chronic inflammatory diseases, the identification of CaMKII as a modulator of TJ function paves the way for the development of new drugs to treat these diseases. PMID:26281891

  3. AMP-18 protects barrier function of colonic epithelial cells: role of tight junction proteins

    PubMed Central

    Walsh-Reitz, Margaret M.; Huang, Erick F.; Musch, Mark W.; Chang, Eugene B.; Martin, Terence E.; Kartha, Sreedharan; Toback, F. Gary

    2005-01-01

    AMP-18, a novel gastric antrum mucosal protein, and a synthetic peptide of amino acids 77-97, have mitogenic and motogenic properties for epithelial cells. The possibility that AMP-18 is also protective was evaluated in the colonic mucosa of mice and monolayer cultures of human colonic epithelial Caco2/bbe (C2) cells. Administration of AMP peptide to mice with dextran sulfate sodium (DSS)-induced colonic injury delayed the onset of bloody diarrhea, and reduced weight loss. Treatment of C2 cells with AMP peptide protected monolayers against decreases in transepithelial electrical resistance (TER) induced by the oxidant monochloramine, indomethacin, or DSS. A molecular mechanism for these barrier-protective effects was sought by asking if AMP peptide acted on specific tight junction (TJ) proteins. Immunoblots of detergent-insoluble fractions of C2 cells treated with AMP peptide exhibited increased accumulation of specific TJ proteins. Occludin immunoreactivity was also increased in detergent-insoluble fractions obtained from colonic mucosal cells of mice injected with AMP peptide. Laser scanning confocal microscopy (CF) supported the capacity of AMP peptide to enhance accumulation of occludin and ZO-1 in TJ domains of C2 cell monolayers, and together with immunoblot analysis showed that the peptide protected against loss of these TJ proteins following oxidant injury. AMP peptide also protected against a fall in TER during disruption of actin filaments by cytochalasin D, and stabilized perijunctional actin during oxidant injury when assessed by CF. These findings suggest that AMP-18 could protect the intestinal mucosal barrier by acting on specific TJ proteins and stabilizing perijunctional actin. PMID:15961882

  4. Mechanism of intestinal mucosal barrier dysfunction in a rat model of chronic obstructive pulmonary disease: An observational study

    PubMed Central

    Xin, Xiaofeng; Dai, Wei; Wu, Jie; Fang, Liping; Zhao, Ming; Zhang, Pengpeng; Chen, Min

    2016-01-01

    The aim of the present study was to investigate intestinal mucosal barrier dysfunction in a rat model of chronic obstructive pulmonary disease (COPD). Male Sprague Dawley rats (n=40) were evenly randomized into control and COPD groups and the COPD model was established by regulated exposure to cigarette smoke for 6 months. Histopathological changes of the lung and intestinal tissues were detected by hematoxylin and eosin staining. Expression of the tight junction proteins occludin and zona occludens-1 (ZO-1) in the intestinal tissues were analyzed by western blotting, serum diamine oxidase (DAO) activity was detected by spectrophotometry, the urinary lactulose to mannitol ratio (L/M) was evaluated by high performance liquid chromatography, and intestinal tissue secretion of tumor necrosis factor (TNF)-α, interferon (IFN)-γ and interleukin (IL)-8 were detected by ELISA. Lung histopathology revealed thinned alveolar walls, ruptured alveolar septa, enlarged and deformed alveoli, and the formation of bullae and emphysema due to alveolar fusion in the COPD group, while intestinal histopathology indicated clearly swollen intestines with darkened and gray mucosa, neutrophil infiltration of the intestinal mucosal and regional epithelial shedding. The occludin and ZO-1 expression levels were significantly lower in the COPD group compared with those in the corresponding control group (P<0.05), while the urinary L/M ratio was significantly higher (P<0.05). Furthermore, the serum DAO activity and secretion of TNF-α, IFN-γ and IL-8 in the intestinal tissues were significantly higher in the COPD group than in the control group (each P<0.05). Dysfunctional and structural changes were observed in the intestinal mucosal barrier in COPD model rats, which may be associated with the increased intestinal inflammatory responses. PMID:27588054

  5. Host-microbial interactions and regulation of intestinal epithelial barrier function: From physiology to pathology

    PubMed Central

    Yu, Linda Chia-Hui; Wang, Jin-Town; Wei, Shu-Chen; Ni, Yen-Hsuan

    2012-01-01

    The gastrointestinal tract is the largest reservoir of commensal bacteria in the human body, providing nutrients and space for the survival of microbes while concurrently operating mucosal barriers to confine the microbial population. The epithelial cells linked by tight junctions not only physically separate the microbiota from the lamina propria, but also secrete proinflammatory cytokines and reactive oxygen species in response to pathogen invasion and metabolic stress and serve as a sentinel to the underlying immune cells. Accumulating evidence indicates that commensal bacteria are involved in various physiological functions in the gut and microbial imbalances (dysbiosis) may cause pathology. Commensal bacteria are involved in the regulation of intestinal epithelial cell turnover, promotion of epithelial restitution and reorganization of tight junctions, all of which are pivotal for fortifying barrier function. Recent studies indicate that aberrant bacterial lipopolysaccharide-mediated signaling in gut mucosa may be involved in the pathogenesis of chronic inflammation and carcinogenesis. Our perception of enteric commensals has now changed from one of opportunistic pathogens to active participants in maintaining intestinal homeostasis. This review attempts to explain the dynamic interaction between the intestinal epithelium and commensal bacteria in disease and health status. PMID:22368784

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

    PubMed

    Utheim, Tor Paaske; Utheim, Øygunn Aass; 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

  7. 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

  8. Fecal microbiota transplantation and bacterial consortium transplantation have comparable effects on the re-establishment of mucosal barrier function in mice with intestinal dysbiosis

    PubMed Central

    Li, Ming; Liang, Pin; Li, Zhenzhen; Wang, Ying; Zhang, Guobin; Gao, Hongwei; Wen, Shu; Tang, Li

    2015-01-01

    Fecal microbiota transplantation (FMT) is a promising therapy, despite some reports of adverse side effects. Bacterial consortia transplantation (BCT) for targeted restoration of the intestinal ecosystem is considered a relatively safe and simple procedure. However, no systematic research has assessed the effects of FMT and BCT on immune responses of intestinal mucosal barrier in patients. We conducted complementary studies in animal models on the effects of FMT and BCT, and provide recommendations for improving the clinical outcomes of these treatments. To establish the dysbiosis model, male BALB/c mice were treated with ceftriaxone intra-gastrically for 7 days. After that, FMT and BCT were performed on ceftriaxone-treated mice for 3 consecutive days to rebuild the intestinal ecosystem. Post-FMT and post-BCT changes of the intestinal microbial community and mucosal barrier functions were investigated and compared. Disruption of intestinal microbial homeostasis impacted the integrity of mucosal epithelial layer, resulting in increased intestinal permeability. These outcomes were accompanied by overexpression of Muc2, significant decrease of SIgA secretion, and overproduction of defensins and inflammatory cytokines. After FMT and BCT, the intestinal microbiota recovered quickly, this was associated with better reconstruction of mucosal barriers and re-establishment of immune networks compared with spontaneous recovery (SR). Although based on a short-term study, our results suggest that FMT and BCT promote the re-establishment of intestinal microbial communities in mice with antibiotic-induced dysbiosis, and contribute to the temporal and spatial interactions between microbiota and mucosal barriers. The effects of BCT are comparable to that of FMT, especially in normalizing the intestinal levels of Muc2, SIgA, and defensins. PMID:26217323

  9. Research Advance in Intestinal Mucosal Barrier and Pathogenesis of Crohn's Disease.

    PubMed

    Wang, Kuan; Wu, Lu-Yi; Dou, Chuan-Zi; Guan, Xin; Wu, Huan-Gan; Liu, Hui-Rong

    2016-01-01

    To date, the etiology and pathogenesis of Crohn's disease (CD) have not been fully elucidated. It is widely accepted that genetic, immune, and environment factors are closely related to the development of CD. As an important defensive line for human body against the environment, intestinal mucosa is able to protect the homeostasis of gut bacteria and alleviate the intestinal inflammatory and immune response. It is evident that the dysfunction of intestinal mucosa barriers plays a crucial role in CD initiation and development. Yet researches are insufficient on intestinal mucosal barrier's action in the prevention of CD onset. This article summarizes the research advances about the correlations between the disorders of intestinal mucosal barriers and CD. PMID:27651792

  10. Research Advance in Intestinal Mucosal Barrier and Pathogenesis of Crohn's Disease

    PubMed Central

    Dou, Chuan-zi; Guan, Xin; Wu, Huan-gan

    2016-01-01

    To date, the etiology and pathogenesis of Crohn's disease (CD) have not been fully elucidated. It is widely accepted that genetic, immune, and environment factors are closely related to the development of CD. As an important defensive line for human body against the environment, intestinal mucosa is able to protect the homeostasis of gut bacteria and alleviate the intestinal inflammatory and immune response. It is evident that the dysfunction of intestinal mucosa barriers plays a crucial role in CD initiation and development. Yet researches are insufficient on intestinal mucosal barrier's action in the prevention of CD onset. This article summarizes the research advances about the correlations between the disorders of intestinal mucosal barriers and CD.

  11. Research Advance in Intestinal Mucosal Barrier and Pathogenesis of Crohn's Disease

    PubMed Central

    Dou, Chuan-zi; Guan, Xin; Wu, Huan-gan

    2016-01-01

    To date, the etiology and pathogenesis of Crohn's disease (CD) have not been fully elucidated. It is widely accepted that genetic, immune, and environment factors are closely related to the development of CD. As an important defensive line for human body against the environment, intestinal mucosa is able to protect the homeostasis of gut bacteria and alleviate the intestinal inflammatory and immune response. It is evident that the dysfunction of intestinal mucosa barriers plays a crucial role in CD initiation and development. Yet researches are insufficient on intestinal mucosal barrier's action in the prevention of CD onset. This article summarizes the research advances about the correlations between the disorders of intestinal mucosal barriers and CD. PMID:27651792

  12. Epithelial microRNAs regulate gut mucosal immunity via epithelium-T cell crosstalk.

    PubMed

    Biton, Moshe; Levin, Avi; Slyper, Michal; Alkalay, Irit; Horwitz, Elad; Mor, Hagar; Kredo-Russo, Sharon; Avnit-Sagi, Tali; Cojocaru, Gady; Zreik, Farid; Bentwich, Zvi; Poy, Matthew N; Artis, David; Walker, Michael D; Hornstein, Eran; Pikarsky, Eli; Ben-Neriah, Yinon

    2011-03-01

    Colonic homeostasis entails epithelium-lymphocyte cooperation, yet many participants in this process are unknown. We show here that epithelial microRNAs mediate the mucosa-immune system crosstalk necessary for mounting protective T helper type 2 (T(H)2) responses. Abolishing the induction of microRNA by gut-specific deletion of Dicer1 (Dicer1(Δgut)), which encodes an enzyme involved in microRNA biogenesis, deprived goblet cells of RELMβ, a key T(H)2 antiparasitic cytokine; this predisposed the host to parasite infection. Infection of Dicer1(Δgut) mice with helminths favored a futile T(H)1 response with hallmarks of inflammatory bowel disease. Interleukin 13 (IL-13) induced the microRNA miR-375, which regulates the expression of TSLP, a T(H)2-facilitating epithelial cytokine; this indicated a T(H)2-amplification loop. We found that miR-375 was required for RELMβ expression in vivo; miR-375-deficient mice had significantly less intestinal RELMβ, which possibly explains the greater susceptibility of Dicer1(Δgut) mice to parasites. Our findings indicate that epithelial microRNAs are key regulators of gut homeostasis and mucosal immunity.

  13. Modulation of distal colonic epithelial barrier function by dietary fibre in normal rats

    PubMed Central

    Mariadason, J; Catto-Smith, A; Gibson, P

    1999-01-01

    BACKGROUND—Dietary fibre influences the turnover and differentiation of the colonic epithelium, but its effects on barrier function are unknown. 
AIMS—To determine whether altering the type and amount of fibre in the diet affects paracellular permeability of intestinal epithelium, and to identify the mechanisms of action. 
METHODS—Rats were fed isoenergetic low fibre diets with or without supplements of wheat bran (10%) or methylcellulose (10%), for four weeks. Paracellular permeability was determined by measurement of conductance and 51Cr-EDTA flux across tissue mounted in Ussing chambers. Faecal short chain fatty acid (SCFA) concentrations were assessed by gas chromatography, epithelial kinetics stathmokinetically, and mucosal brush border hydrolase activities spectrophotometrically. 
RESULTS—Body weight was similar across the dietary groups. Conductance and 51Cr-EDTA flux were approximately 25% higher in animals fed no fibre, compared with those fed wheat bran or methylcellulose in the distal colon, but not in the caecum or jejunum. Histologically, there was no evidence of epithelial injury or erosion associated with any diet. The fibres exerted different spectra of effects on luminal SCFA concentrations and pH, and on mucosal indexes, but both bulked the faeces, were trophic to the epithelium, and stimulated expression of a marker of epithelial differentiation. 
CONCLUSIONS—Both a fermentable and a non-fermentable fibre reduce paracellular permeability specifically in the distal colon, possibly by promoting epithelial cell differentiation. The mechanisms by which the two fibres exert their effects are likely to be different. 

 Keywords: colon; differentiation; epithelium; fibre; paracellular permeability; proliferation PMID:10026327

  14. Pulmonary epithelial barrier function: some new players and mechanisms

    PubMed Central

    Brune, Kieran; Frank, James; Schwingshackl, Andreas; Finigan, James

    2015-01-01

    The pulmonary epithelium serves as a barrier to prevent access of the inspired luminal contents to the subepithelium. In addition, the epithelium dictates the initial responses of the lung to both infectious and noninfectious stimuli. One mechanism by which the epithelium does this is by coordinating transport of diffusible molecules across the epithelial barrier, both through the cell and between cells. In this review, we will discuss a few emerging paradigms of permeability changes through altered ion transport and paracellular regulation by which the epithelium gates its response to potentially detrimental luminal stimuli. This review is a summary of talks presented during a symposium in Experimental Biology geared toward novel and less recognized methods of epithelial barrier regulation. First, we will discuss mechanisms of dynamic regulation of cell-cell contacts in the context of repetitive exposure to inhaled infectious and noninfectious insults. In the second section, we will briefly discuss mechanisms of transcellular ion homeostasis specifically focused on the role of claudins and paracellular ion-channel regulation in chronic barrier dysfunction. In the next section, we will address transcellular ion transport and highlight the role of Trek-1 in epithelial responses to lung injury. In the final section, we will outline the role of epithelial growth receptor in barrier regulation in baseline, acute lung injury, and airway disease. We will then end with a summary of mechanisms of epithelial control as well as discuss emerging paradigms of the epithelium role in shifting between a structural element that maintains tight cell-cell adhesion to a cell that initiates and participates in immune responses. PMID:25637609

  15. Role of EP4 receptor and prostaglandin transporter in prostaglandin E2-induced alteration in colonic epithelial barrier integrity.

    PubMed

    Lejeune, Manigandan; Leung, Pearl; Beck, Paul L; Chadee, Kris

    2010-11-01

    Prostaglandin E(2) (PGE(2)) is a proinflammatory lipid mediator produced in excess in inflammatory bowel disease (IBD). PGE(2) couples to and signals via four different E-prostanoid (EP) receptors, namely EP1, EP2, EP3, and EP4. In this study, we determined a role for PGE(2) and EP4 receptors in altering colonic epithelial barrier integrity. In healthy colonic mucosa, EP4 receptors were localized on apical plasma membrane of epithelial cells at the tip of mucosal folds, whereas, in patients with IBD and in rats with dextran sodium sulfate (DSS)-induced colitis, they were diffusely overexpressed throughout the mucosa. Similarly, expression of EP4 receptor was polarized in T84 colonic epithelial monolayer and mimics the normal epithelium. Apical exposure of T84 monolayer with high levels of PGE(2) decreased barrier integrity, which was abrogated by an EP4 receptor antagonist. To reveal the mechanism of vectorial transport of basally produced PGE(2) toward apical EP4 receptors, we identified prostaglandin transporters (PGT) in human colonic epithelia. PGT were least expressed on epithelial cells at the colonic mucosal folds of control subjects but overexpressed in epithelial cells of patients with IBD or animals with DSS-induced colitis. T84 monolayer also expressed PGT, which increased twofold following stimulation with TNF-α. Importantly, in T84 monolayer stimulated with TNF-α, there was a corresponding increase in the uptake and vectorial transport of (3)H-PGE(2) to the apical surface. Knockdown or pharmacological inhibition of PGT significantly decreased vectorial transport of (3)H-PGE(2). These studies unravel a mechanism whereby EP4 receptor and PGT play a role in PGE(2)-induced alteration of epithelial barrier integrity in colitis.

  16. Bacillus anthracis Lethal Toxin Reduces Human Alveolar Epithelial Barrier Function

    PubMed Central

    Langer, Marybeth; Duggan, Elizabeth Stewart; Booth, John Leland; Patel, Vineet Indrajit; Zander, Ryan A.; Silasi-Mansat, Robert; Ramani, Vijay; Veres, Tibor Zoltan; Prenzler, Frauke; Sewald, Katherina; Williams, Daniel M.; Coggeshall, Kenneth Mark; Awasthi, Shanjana; Lupu, Florea; Burian, Dennis; Ballard, Jimmy Dale; Braun, Armin

    2012-01-01

    The lung is the site of entry for Bacillus anthracis in inhalation anthrax, the deadliest form of the disease. Bacillus anthracis produces virulence toxins required for disease. Alveolar macrophages were considered the primary target of the Bacillus anthracis virulence factor lethal toxin because lethal toxin inhibits mouse macrophages through cleavage of MEK signaling pathway components, but we have reported that human alveolar macrophages are not a target of lethal toxin. Our current results suggest that, unlike human alveolar macrophages, the cells lining the respiratory units of the lung, alveolar epithelial cells, are a target of lethal toxin in humans. Alveolar epithelial cells expressed lethal toxin receptor protein, bound the protective antigen component of lethal toxin, and were subject to lethal-toxin-induced cleavage of multiple MEKs. These findings suggest that human alveolar epithelial cells are a target of Bacillus anthracis lethal toxin. Further, no reduction in alveolar epithelial cell viability was observed, but lethal toxin caused actin rearrangement and impaired desmosome formation, consistent with impaired barrier function as well as reduced surfactant production. Therefore, by compromising epithelial barrier function, lethal toxin may play a role in the pathogenesis of inhalation anthrax by facilitating the dissemination of Bacillus anthracis from the lung in early disease and promoting edema in late stages of the illness. PMID:23027535

  17. Probiotics Prevent Intestinal Barrier Dysfunction in Acute Pancreatitis in Rats via Induction of Ileal Mucosal Glutathione Biosynthesis

    PubMed Central

    Lutgendorff, Femke; Nijmeijer, Rian M.; Sandström, Per A.; Trulsson, Lena M.; Magnusson, Karl-Eric; Timmerman, Harro M.; van Minnen, L. Paul; Rijkers, Ger T.; Gooszen, Hein G.; Akkermans, Louis M. A.; Söderholm, Johan D.

    2009-01-01

    Background During acute pancreatitis (AP), oxidative stress contributes to intestinal barrier failure. We studied actions of multispecies probiotics on barrier dysfunction and oxidative stress in experimental AP. Methodology/Principal Findings Fifty-three male Spraque-Dawley rats were randomly allocated into five groups: 1) controls, non-operated, 2) sham-operated, 3) AP, 4) AP and probiotics and 5) AP and placebo. AP was induced by intraductal glycodeoxycholate infusion and intravenous cerulein (6 h). Daily probiotics or placebo were administered intragastrically, starting five days prior to AP. After cerulein infusion, ileal mucosa was collected for measurements of E. coli K12 and 51Cr-EDTA passage in Ussing chambers. Tight junction proteins were investigated by confocal immunofluorescence imaging. Ileal mucosal apoptosis, lipid peroxidation, and glutathione levels were determined and glutamate-cysteine-ligase activity and expression were quantified. AP-induced barrier dysfunction was characterized by epithelial cell apoptosis and alterations of tight junction proteins (i.e. disruption of occludin and claudin-1 and up-regulation of claudin-2) and correlated with lipid peroxidation (r>0.8). Probiotic pre-treatment diminished the AP-induced increase in E. coli passage (probiotics 57.4±33.5 vs. placebo 223.7±93.7 a.u.; P<0.001), 51Cr-EDTA flux (16.7±10.1 vs. 32.1±10.0 cm/s10−6; P<0.005), apoptosis, lipid peroxidation (0.42±0.13 vs. 1.62±0.53 pmol MDA/mg protein; P<0.001), and prevented tight junction protein disruption. AP-induced decline in glutathione was not only prevented (14.33±1.47 vs. 8.82±1.30 nmol/mg protein, P<0.001), but probiotics even increased mucosal glutathione compared with sham rats (14.33±1.47 vs. 10.70±1.74 nmol/mg protein, P<0.001). Glutamate-cysteine-ligase activity, which is rate-limiting in glutathione biosynthesis, was enhanced in probiotic pre-treated animals (probiotics 2.88±1.21 vs. placebo 1.94±0.55 nmol/min/mg protein; P<0

  18. Concise review: transplantation of cultured oral mucosal epithelial cells for treating limbal stem cell deficiency-current status and future perspectives.

    PubMed

    Utheim, Tor Paaske

    2015-06-01

    A number of diseases and external factors can deplete limbal stem cells, causing pain and visual loss. Ten years have passed since the first transplantation of cultured oral mucosal epithelial cells in humans, representing the first autologous cell-based therapy for severe bilateral limbal stem cell deficiency. Its steady increase in popularity since then can be attributed to the accumulating evidence of its efficacy in reverting limbal stem cell deficiency. In this review, the focus is on clinical, and to a lesser degree laboratory, features of cultured oral mucosal epithelial transplants over the past 10 years. Comparisons with other available technologies are made. Avenues for research to stimulate further improvements in clinical results and allow worldwide distribution of limbal stem cell therapy based on oral mucosal cells are discussed. These include storage and transportation of cultured oral mucosal epithelial sheets and in vivo culture of oral mucosal epithelial cells.

  19. Electroacupuncture at Bilateral Zusanli Points (ST36) Protects Intestinal Mucosal Immune Barrier in Sepsis

    PubMed Central

    Zhu, Mei-fei; Xing, Xi; Lei, Shu; Wu, Jian-nong; Wang, Ling-cong; Huang, Li-quan; Jiang, Rong-lin

    2015-01-01

    Sepsis results in high morbidity and mortality. Immunomodulation strategies could be an adjunctive therapy to treat sepsis. Acupuncture has also been used widely for many years in China to treat sepsis. However, the underlying mechanisms are not well-defined. We demonstrated here that EA preconditioning at ST36 obviously ameliorated CLP-induced intestinal injury and high permeability and reduced the mortality of CLP-induced sepsis rats. Moreover, electroacupuncture (EA) pretreatment exerted protective effects on intestinal mucosal immune barrier by increasing the concentration of sIgA and the percentage of CD3+, γ/δ, and CD4+ T cells and the ratio of CD4+/CD8+ T cells. Although EA at ST36 treatments immediately after closing the abdomen in the CLP procedure with low-frequency or high-frequency could not reduce the mortality of CLP-induced sepsis in rats, these EA treatments could also significantly improve intestinal injury index in rats with sepsis and obviously protected intestinal mucosal immune barrier. In conclusion, our findings demonstrated that EA at ST36 could improve intestinal mucosal immune barrier in sepsis induced by CLP, while the precise mechanism underlying the effects needs to be further elucidated. PMID:26346309

  20. Chronic Kidney Disease Induced Intestinal Mucosal Barrier Damage Associated with Intestinal Oxidative Stress Injury

    PubMed Central

    Yu, Chao; Wang, Qiang; Zhou, Chunyu; Kang, Xin; Zhao, Shuang; Liu, Shuai; Fu, Huijun; Yu, Zhen; Peng, Ai

    2016-01-01

    Background. To investigate whether intestinal mucosal barrier was damaged or not in chronic kidney disease progression and the status of oxidative stress. Methods. Rats were randomized into two groups: a control group and a uremia group. The uremia rat model was induced by 5/6 kidney resection. In postoperative weeks (POW) 4, 6, 8, and 10, eight rats were randomly selected from each group to prepare samples for assessing systemic inflammation, intestinal mucosal barrier changes, and the status of intestinal oxidative stress. Results. The uremia group presented an increase trend over time in the serum tumor necrosis factor-alpha, interleukin-6 (IL-6) and IL-10, serum D-lactate and diamine oxidase, and intestinal permeability, and these biomarkers were significantly higher than those in control group in POW 8 and/or 10. Chiu's scores in uremia group were also increased over time, especially in POW 8 and 10. Furthermore, the intestinal malondialdehyde, superoxide dismutase, and glutathione peroxidase levels were significantly higher in uremia group when compared with those in control group in POW 8 and/or 10. Conclusions. The advanced chronic kidney disease could induce intestinal mucosal barrier damage and further lead to systemic inflammation. The underlying mechanism may be associated with the intestinal oxidative stress injury. PMID:27493661

  1. 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

  2. Metallic oxide nanoparticle translocation across the human bronchial epithelial barrier

    NASA Astrophysics Data System (ADS)

    George, Isabelle; Naudin, Grégoire; Boland, Sonja; Mornet, Stéphane; Contremoulins, Vincent; Beugnon, Karine; Martinon, Laurent; Lambert, Olivier; Baeza-Squiban, Armelle

    2015-02-01

    Inhalation is the most frequent route of unintentional exposure to nanoparticles (NPs). Our aim was to quantify the translocation of different metallic NPs across human bronchial epithelial cells and to determine the factors influencing this translocation. Calu-3 cells forming a tight epithelial barrier when grown on a porous membrane in a two compartment chamber were exposed to fluorescently labelled NPs to quantify the NP translocation. NP translocation and uptake by cells were also studied by confocal and transmission electron microscopy. Translocation was characterized according to NP size (16, 50, or 100 nm), surface charge (negative or positive SiO2), composition (SiO2 or TiO2), presence of proteins or phospholipids and in an inflammatory context. Our results showed that NPs can translocate through the Calu-3 monolayer whatever their composition (SiO2 or TiO2), but this translocation was increased for the smallest and negatively charged NPs. Translocation was not associated with an alteration of the integrity of the epithelial monolayer, suggesting a transcytosis of the internalized NPs. By modifying the NP corona, the ability of NPs to cross the epithelial barrier differed depending on their intrinsic properties, making positively charged NPs more prone to translocate. NP translocation can be amplified by using agents known to open tight junctions and to allow paracellular passage. NP translocation was also modulated when mimicking an inflammatory context frequently found in the lungs, altering the epithelial integrity and inducing transient tight junction opening. This in vitro evaluation of NP translocation could be extended to other inhaled NPs to predict their biodistribution.Inhalation is the most frequent route of unintentional exposure to nanoparticles (NPs). Our aim was to quantify the translocation of different metallic NPs across human bronchial epithelial cells and to determine the factors influencing this translocation. Calu-3 cells forming a

  3. Metallic oxide nanoparticle translocation across the human bronchial epithelial barrier.

    PubMed

    George, Isabelle; Naudin, Grégoire; Boland, Sonja; Mornet, Stéphane; Contremoulins, Vincent; Beugnon, Karine; Martinon, Laurent; Lambert, Olivier; Baeza-Squiban, Armelle

    2015-03-14

    Inhalation is the most frequent route of unintentional exposure to nanoparticles (NPs). Our aim was to quantify the translocation of different metallic NPs across human bronchial epithelial cells and to determine the factors influencing this translocation. Calu-3 cells forming a tight epithelial barrier when grown on a porous membrane in a two compartment chamber were exposed to fluorescently labelled NPs to quantify the NP translocation. NP translocation and uptake by cells were also studied by confocal and transmission electron microscopy. Translocation was characterized according to NP size (16, 50, or 100 nm), surface charge (negative or positive SiO2), composition (SiO2 or TiO2), presence of proteins or phospholipids and in an inflammatory context. Our results showed that NPs can translocate through the Calu-3 monolayer whatever their composition (SiO2 or TiO2), but this translocation was increased for the smallest and negatively charged NPs. Translocation was not associated with an alteration of the integrity of the epithelial monolayer, suggesting a transcytosis of the internalized NPs. By modifying the NP corona, the ability of NPs to cross the epithelial barrier differed depending on their intrinsic properties, making positively charged NPs more prone to translocate. NP translocation can be amplified by using agents known to open tight junctions and to allow paracellular passage. NP translocation was also modulated when mimicking an inflammatory context frequently found in the lungs, altering the epithelial integrity and inducing transient tight junction opening. This in vitro evaluation of NP translocation could be extended to other inhaled NPs to predict their biodistribution.

  4. Ginsenoside Rb1 protects the intestinal mucosal barrier following peritoneal air exposure

    PubMed Central

    Zhou, Feng; Zhang, Peichen; Chen, Xiaoxi; Yan, Jingyi; Yao, Jiangao; Yu, Zhen; Chen, Xiaolei

    2016-01-01

    Ginsenoside Rb1 (GRb1), which is one of the main ingredients derived from Panax ginseng, has been widely used to treat various gastrointestinal disorders. The present study aimed to determine whether GRb1 was able to prevent intestinal mucosal barrier damage in rats following peritoneal air exposure for 3 h. GRb1 (5, 10, and 20 mg/kg) was orally administrated via gavage four times prior to and following surgery. Blood and terminal ileum were sampled 24 h following surgery. Levels of serum D-lactate (D-LA) were detected using an enzyme-linked immunosorbent assay kit. Intestinal permeability was assessed by determining the intestinal clearance of fluorescein isothiocyanate-dextran (FD4). Activity of intestinal myeloperoxidase was measured to assess intestinal inflammation, and intestinal histopathology was assessed by light microscopy. The results showed that GRb1 reduced the level of serum D-LA, intestinal clearance of FD4, and the activity of intestinal myeloperoxidase. Intestinal edema and inflammation were also ameliorated by GRb1, and the Chiu's scores employed for assessing intestinal mucosal damage were also reduced in the GRb1-treated peritoneal air exposure group. In addition, GRb1 induced a significant difference at 10 and 20 mg/kg, indicating a dose-dependent effect. The results of the present study suggest that GRb1 may be able to protect the intestinal mucosal barrier against damage induced by peritoneal air exposure, which may be associated with its anti-inflammatory action. PMID:27703510

  5. Evidence of the survival of ectopically transplanted oral mucosal epithelial stem cells after repeated wounding of cornea.

    PubMed

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

    2014-08-01

    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.

  6. Enteral feeding and its impact on the gut immune system and intestinal mucosal barrier

    PubMed Central

    Kruszewski, Wiesław J.; Buczek, Tomasz

    2015-01-01

    Enteral feeding is the preferred method of nutritional therapy. Mucosal lack of contact with nutrients leads do lymphoid tissue atrophy, immune system functional decline, and intensification in bacterial translocation. Currently, it is assumed that microbiome is one of the body organs that has a significant impact on health. The composition of microbiome is not affected by age, sex, or place of residence, although it changes rapidly after diet modification. The composition of the microbiome is determined by enterotype, which is specific for each organism. It has a significant impact on the risk of diabetes, cancer, atherosclerosis, and other diseases. This review gathers data on interaction between gut-associated lymphoid tissue, mucosa-associated lymphoid tissue, microbiome, and the intestinal mucosal barrier. Usually, the information on the aforementioned is scattered in specialist-subject magazines such as gastroenterology, microbiology, genetics, biochemistry, and others. PMID:26557936

  7. Convective diffusion of nanoparticles from the epithelial barrier toward regional lymph nodes.

    PubMed

    Dukhin, Stanislav S; Labib, Mohamed E

    2013-11-01

    Drug delivery using nanoparticles as drug carriers has recently attracted the attention of many investigators. Targeted delivery of nanoparticles to the lymph nodes is especially important to prevent cancer metastasis or infection, and to diagnose disease stage. However, systemic injection of nanoparticles often results in organ toxicity because they reach and accumulate in all the lymph nodes in the body. An attractive strategy would be to deliver the drug-loaded nanoparticles to a subset of draining lymph nodes corresponding to a specific site or organ to minimize systemic toxicity. In this respect, mucosal delivery of nanoparticles to regional draining lymph nodes of a selected site creates a new opportunity to accomplish this task with minimal toxicity. One example is the delivery of nanoparticles from the vaginal lumen to draining lymph nodes to prevent the transmission of HIV in women. Other known examples include mucosal delivery of vaccines to induce immunity. In all cases, molecular and particle transport by means of diffusion and convective diffusion play a major role. The corresponding transport processes have common inherent regularities and are addressed in this review. Here we use nanoparticle delivery from the vaginal lumen to the lymph nodes as an example to address the many aspects of associated transport processes. In this case, nanoparticles penetrate the epithelial barrier and move through the interstitium (tissue) to the initial lymphatics until they finally reach the lymph nodes. Since the movement of interstitial liquid near the epithelial barrier is retarded, nanoparticle transport was found to take place through special foci present in the epithelium. Immediately after nanoparticles emerge from the foci, they move through the interstitium due to diffusion affected by convection (convective diffusion). Specifically, the convective transport of nanoparticles occurs due to their convection together with interstitial fluid through the

  8. Moving towards a new era in the research of tonsils and mucosal barriers.

    PubMed

    Yamanaka, Noboru

    2011-01-01

    The palatine and nasopharyngeal tonsils (adenoids) are lymphoepithelial tissues located in strategic anatomical areas of the oral pharynx and nasopharynx. These immunocompetent tissues represent the first line of defense against ingested or inhaled foreign proteins such as bacteria, viruses, or food antigens. Accompanying the advances being made in the field of medicine today, the role of the tonsils in immunocompetence is becoming extremely important. Upper respiratory tract infections such as acute otitis media, acute rhinosinusitis and acute pharyngo-tonsillitis are diseases that occur with extremely high frequency, and the antimicrobial agents used to treat these diseases account for a large proportion of health care costs. The increasingly refractory nature of upper respiratory tract infections caused by drug-resistant bacteria has become a major worldwide concern. The elucidation of the immune functions of the tonsils and mucosal membranes of the upper respiratory tract is considered to have important significance. The tonsils are also considered to play an important role as one of the causes of sleep apnea syndrome, and have been reported to be intimately involved in the manifestation of IgA nephropathy and palmoplantar pustulosis, a kind of skin disorder. Interest has continued to grow in this symposium with each session ever since it was first held in Kyoto, Japan in 1987. Since then, the symposium has been held every 3-4 years; in Pavia in 1991, in Sapporo in 1995, in Ghent in 1999, in Wakayama in 2003, and in Siena in 2006. Since the 5th symposium in Wakayama, the topics were extended to mucosal barriers of upper airways including the mucosal immune system, innate immunity, and mucosal vaccine. Recent fine technologies and information on molecular biological approaches for upper airways will continue to advance our understanding of epidemiology, etiology, pathogenesis, diagnosis and management of tonsil-related disorders and various upper respiratory

  9. Pregnane X receptor agonists enhance intestinal epithelial wound healing and repair of the intestinal barrier following the induction of experimental colitis.

    PubMed

    Terc, Joshua; Hansen, Ashleigh; Alston, Laurie; Hirota, Simon A

    2014-05-13

    The intestinal epithelial barrier plays a key role in the maintenance of homeostasis within the gastrointestinal tract. Barrier dysfunction leading to increased epithelial permeability is associated with a number of gastrointestinal disorders including the inflammatory bowel diseases (IBD) - Crohn's disease and ulcerative colitis. It is thought that the increased permeability in patients with IBD may be driven by alterations in the epithelial wound healing response. To this end considerable study has been undertaken to identify signaling pathways that may accelerate intestinal epithelial wound healing and normalize the barrier dysfunction observed in IBD. In the current study we examined the role of the pregnane X receptor (PXR) in modulating the intestinal epithelial wound healing response. Mutations and reduced mucosal expression of the PXR are associated with IBD, and others have reported that PXR agonists can dampen intestinal inflammation. Furthermore, stimulation of the PXR has been associated with increased cell migration and proliferation, two of the key processes involved in wound healing. We hypothesized that PXR agonists would enhance intestinal epithelial repair. Stimulation of Caco-2 intestinal epithelial cells with rifaximin, rifampicin and SR12813, all potent agonists of the PXR, significantly increased wound closure. This effect was driven by p38 MAP kinase-dependent cell migration, and occurred in the absence of cell proliferation. Treating mice with a rodent specific PXR agonist, pregnenolone 16α-carbonitrile (PCN), attenuated the intestinal barrier dysfunction observed in the dextran sulphate sodium (DSS) model of experimental colitis, an effect that occurred independent of the known anti-inflammatory effects of PCN. Taken together our data indicate that the activation of the PXR can enhance intestinal epithelial repair and suggest that targeting the PXR may help to normalize intestinal barrier dysfunction observed in patients with IBD

  10. Epithelial restitution and cellular proliferation after gastric mucosal damage caused by hypertonic NaCl in rats.

    PubMed

    Sørbye, H; Svanes, C; Stangeland, L; Kvinnsland, S; Svanes, K

    1988-01-01

    Hypertonic NaCl enhances gastric cancer in rats induced by N-nitroso compounds. This study was designed to examine the structural changes and alterations in mitotic activity occurring after mucosal exposure to hypertonic NaCl. Wistar rats were given one ml of 4.5 M NaCl by gastric tube and groups of 4-5 animals were sacrificed at different time intervals up to 120 h. An i.p. injection of thymidine was given 1 h before death. Samples of antral and corpus mucosa were prepared for microscopy and autoradiography. Hypertonic NaCl caused uniform destruction of surface mucous cells and pits in the corpus and antrum. Epithelial restitution with the formation of a thin epithelial layer occurred within one h of damage. The mucosa changed towards normal within 24-48 h. The distance between mucosal surface and the replicating cells decreased during the first 2 h. The proliferation zone remained in the middle of the glandular layer throughout the experiment. The proliferative activity increased during the first 24 h after mucosal damage. The number of labelled cells per unit area of mucosa was somewhat larger in the corpus than the antrum, but in the corpus the distance between proliferating cells and mucosal surface was double that of the antrum. Hypertonic NaCl causes a series of changes in the gastric mucosa. The increased mitotic activity can only partly explain the cocarcinogenic effect, since N-nitroso-induced adenocarcinomas occur predominantly in the antrum while the mitotic activity is maximal in the corpus.

  11. Convective diffusion of nanoparticles from the epithelial barrier towards regional lymph nodes

    PubMed Central

    Dukhin, Stanislav S; Labib, Mohamed E.

    2013-01-01

    Drug delivery using nanoparticles as drug carriers has recently attracted the attention of many investigators. Targeted delivery of nanoparticles to lymph nodes is especially important to prevent cancer metastasis or infection, and to diagnose disease stage. However, systemic injection of nanoparticles often results in organ toxicity because they reach and accumulate in all the lymph nodes in the body. An attractive strategy would be to deliver the drug-loaded nanoparticles to a subset of draining lymph nodes corresponding to a specific site or organ to minimize systemic toxicity. In this respect, mucosal delivery of nanoparticles to regional draining lymph nodes of a selected site creates a new opportunity to accomplish this task with minimal toxicity. One example is the delivery of nanoparticles from the vaginal lumen to draining lymph nodes to prevent the transmission of HIV in women. Other known examples include mucosal delivery of vaccines to induce immunity. In all cases, molecular and particle transport by means of diffusion and convective diffusion play a major role. The corresponding transport processes have common inherent regularities and are addressed in this review. Here we use nanoparticles delivery from the vaginal lumen to lymph nodes as an example to address the many aspects of associated transport processes. In this case, nanoparticles penetrate the epithelial barrier and move through the interstitium (tissue) to the initial lymphatics until they finally reach the lymph nodes. Since the movement of interstitial liquid near the epithelial barrier is retarded, nanoparticles transport was found to take place through special foci present in the epithelium. Immediately after nanoparticles emerge from the foci, they move through the interstitium due to diffusion affected by convection (convective diffusion). Specifically, the convective transport of nanoparticles occurs due to their convection together with interstitial fluid through the interstitium

  12. Convective diffusion of nanoparticles from the epithelial barrier toward regional lymph nodes.

    PubMed

    Dukhin, Stanislav S; Labib, Mohamed E

    2013-11-01

    Drug delivery using nanoparticles as drug carriers has recently attracted the attention of many investigators. Targeted delivery of nanoparticles to the lymph nodes is especially important to prevent cancer metastasis or infection, and to diagnose disease stage. However, systemic injection of nanoparticles often results in organ toxicity because they reach and accumulate in all the lymph nodes in the body. An attractive strategy would be to deliver the drug-loaded nanoparticles to a subset of draining lymph nodes corresponding to a specific site or organ to minimize systemic toxicity. In this respect, mucosal delivery of nanoparticles to regional draining lymph nodes of a selected site creates a new opportunity to accomplish this task with minimal toxicity. One example is the delivery of nanoparticles from the vaginal lumen to draining lymph nodes to prevent the transmission of HIV in women. Other known examples include mucosal delivery of vaccines to induce immunity. In all cases, molecular and particle transport by means of diffusion and convective diffusion play a major role. The corresponding transport processes have common inherent regularities and are addressed in this review. Here we use nanoparticle delivery from the vaginal lumen to the lymph nodes as an example to address the many aspects of associated transport processes. In this case, nanoparticles penetrate the epithelial barrier and move through the interstitium (tissue) to the initial lymphatics until they finally reach the lymph nodes. Since the movement of interstitial liquid near the epithelial barrier is retarded, nanoparticle transport was found to take place through special foci present in the epithelium. Immediately after nanoparticles emerge from the foci, they move through the interstitium due to diffusion affected by convection (convective diffusion). Specifically, the convective transport of nanoparticles occurs due to their convection together with interstitial fluid through the

  13. 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

  14. 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.

  15. 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.

  16. Influenza virus damages the alveolar barrier by disrupting epithelial cell tight junctions.

    PubMed

    Short, Kirsty R; Kasper, Jennifer; van der Aa, Stijn; Andeweg, Arno C; Zaaraoui-Boutahar, Fatiha; Goeijenbier, Marco; Richard, Mathilde; Herold, Susanne; Becker, Christin; Scott, Dana P; Limpens, Ronald W A L; Koster, Abraham J; Bárcena, Montserrat; Fouchier, Ron A M; Kirkpatrick, Charles James; Kuiken, Thijs

    2016-03-01

    A major cause of respiratory failure during influenza A virus (IAV) infection is damage to the epithelial-endothelial barrier of the pulmonary alveolus. Damage to this barrier results in flooding of the alveolar lumen with proteinaceous oedema fluid, erythrocytes and inflammatory cells. To date, the exact roles of pulmonary epithelial and endothelial cells in this process remain unclear.Here, we used an in vitro co-culture model to understand how IAV damages the pulmonary epithelial-endothelial barrier. Human epithelial cells were seeded on the upper half of a transwell membrane while human endothelial cells were seeded on the lower half. These cells were then grown in co-culture and IAV was added to the upper chamber.We showed that the addition of IAV (H1N1 and H5N1 subtypes) resulted in significant barrier damage. Interestingly, we found that, while endothelial cells mounted a pro-inflammatory/pro-coagulant response to a viral infection in the adjacent epithelial cells, damage to the alveolar epithelial-endothelial barrier occurred independently of endothelial cells. Rather, barrier damage was associated with disruption of tight junctions amongst epithelial cells, and specifically with loss of tight junction protein claudin-4.Taken together, these data suggest that maintaining epithelial cell integrity is key in reducing pulmonary oedema during IAV infection.

  17. 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

  18. The role of intestinal epithelial barrier function in the development of NEC

    PubMed Central

    Halpern, Melissa D; Denning, Patricia W

    2015-01-01

    The intestinal epithelial barrier plays an important role in maintaining host health. Breakdown of intestinal barrier function is known to play a role in many diseases such as infectious enteritis, idiopathic inflammatory bowel disease, and neonatal inflammatory bowel diseases. Recently, increasing research has demonstrated the importance of understanding how intestinal epithelial barrier function develops in the premature neonate in order to develop strategies to promote its maturation. Optimizing intestinal barrier function is thought to be key to preventing neonatal inflammatory bowel diseases such as necrotizing enterocolitis. In this review, we will first summarize the key components of the intestinal epithelial barrier, what is known about its development, and how this may explain NEC pathogenesis. Finally, we will review what therapeutic strategies may be used to promote optimal development of neonatal intestinal barrier function in order to reduce the incidence and severity of NEC. PMID:25927016

  19. Toward improving mucosal barrier defenses: rhG-CSF plus IgG antibody.

    PubMed

    Simmonds, Aryeh; LaGamma, Edmund F

    2006-11-01

    Epithelial cell functions ultimately define the ability of the extremely low birth weight human fetus to survive outside of the uterus. These specialized epithelial cell capacities manage all human interactions with the ex utero world including: (i) lung mechanics, surface chemistry and gas exchange, (ii) renal tubular balance of fluid and electrolytes, (iii) barrier functions of the intestine and skin for keeping bacteria out and water in, plus enabling intestinal digestion, as well as (iv) maintaining an intact neuroepithelium lining of the ventricles of the brain and retina. In Part I of this two part review, the authors describe why the gut barrier is a clinically relevant model system for studying the complex interplay between innate and adaptive immunity, dendritic &epithelial cell interactions, intraepithelial lymphocytes, M-cells, as well as the gut associated lymphoid tissues where colonization after birth, clinician feeding practices, use of antibiotics as well as exposure to prebiotics, probiotics and maternal vaginal flora all program the neonate for a life-time of immune competence distinguishing "self" from foreign antigens. These barrier defense capacities become destructive during disease processes like necrotizing enterocolitis (NEC) when an otherwise maturationally normal, yet dysregulated and immature, immune defense system is associated with high levels of certain inflammatory mediators like TNFa. In Part II the authors discuss the rationale for why rhG-CSF has theoretical advantages in managing NEC or sepsis by augmenting neonatal neutrophil number, neutrophil expression of Fcg and complement receptors, as well as phagocytic function and oxidative burst. rhG-CSF also has potent anti-TNFa functions that may serve to limit extension of tissue destruction while not impairing bacterial killing capacity. Healthy, non-infected neutropenic and septic neonates differ in their ability to respond to rhG-CSF; however, no neonatal clinical trials to date

  20. Vocal Fold Epithelial Barrier in Health and Injury A Research Review

    PubMed Central

    Levendoski, Elizabeth Erickson; Leydon, Ciara; Thibeault, Susan L.

    2015-01-01

    Purpose Vocal fold epithelium is composed of layers of individual epithelial cells joined by junctional complexes constituting a unique interface with the external environment. This barrier provides structural stability to the vocal folds and protects underlying connective tissue from injury while being nearly continuously exposed to potentially hazardous insults including environmental or systemic-based irritants such as pollutants and reflux, surgical procedures, and vibratory trauma. Small disruptions in the epithelial barrier may have a large impact on susceptibility to injury and overall vocal health. The purpose of this article is to provide a broad-based review of our current knowledge of the vocal fold epithelial barrier. Methods A comprehensive review of the literature was conducted. Details of the structure of the vocal fold epithelial barrier are presented and evaluated in the context of function in injury and pathology. The importance of the epithelial-associated vocal fold mucus barrier is also introduced. Results/Conclusions Information presented in this review is valuable for clinicians and researchers as it highlights the importance of this understudied portion of the vocal folds to overall vocal health and disease. Prevention and treatment of injury to the epithelial barrier is a significant area awaiting further investigation. PMID:24686981

  1. Alix-mediated assembly of the actomyosin–tight junction polarity complex preserves epithelial polarity and epithelial barrier

    PubMed Central

    Campos, Yvan; Qiu, Xiaohui; Gomero, Elida; Wakefield, Randall; Horner, Linda; Brutkowski, Wojciech; Han, Young-Goo; Solecki, David; Frase, Sharon; Bongiovanni, Antonella; d'Azzo, Alessandra

    2016-01-01

    Maintenance of epithelial cell polarity and epithelial barrier relies on the spatial organization of the actin cytoskeleton and proper positioning/assembly of intercellular junctions. However, how these processes are regulated is poorly understood. Here we reveal a key role for the multifunctional protein Alix in both processes. In a knockout mouse model of Alix, we identified overt structural changes in the epithelium of the choroid plexus and in the ependyma, such as asymmetrical cell shape and size, misplacement and abnormal beating of cilia, blebbing of the microvilli. These defects culminate in excessive cell extrusion, enlargement of the lateral ventricles and hydrocephalus. Mechanistically, we find that by interacting with F-actin, the Par complex and ZO-1, Alix ensures the formation and maintenance of the apically restricted actomyosin–tight junction complex. We propose that in this capacity Alix plays a role in the establishment of apical–basal polarity and in the maintenance of the epithelial barrier. PMID:27336173

  2. MicroRNAs regulate tight junction proteins and modulate epithelial/endothelial barrier functions.

    PubMed

    Cichon, Christoph; Sabharwal, Harshana; Rüter, Christian; Schmidt, M Alexander

    2014-01-01

    Tightly controlled epithelial and endothelial barriers are a prerequisite for life as these barriers separate multicellular organisms from their environment and serve as first lines of defense. Barriers between neighboring epithelial cells are formed by multiple intercellular junctions including the 'apical junctional complex-AJC' with tight junctions (TJ), adherens junctions (AJ), and desmosomes. TJ consist of tetraspan transmembrane proteins like occludin, various claudins that directly control paracellular permeability, and the 'Junctional Adhesion Molecules' (JAMs). For establishing tight barriers TJ are essential but at the same time have to allow also selective permeability. For this, TJ need to be tightly regulated and controlled. This is organized by a variety of adaptor molecules, i.e., protein kinases, phosphatases and GTPases, which in turn are regulated and fine-tuned involving microRNAs (miRNAs). In this review we summarize available data on the role and targeting of miRNAs in the maintenance of epithelial and/or endothelial barriers.

  3. Claudin-based barrier differentiation in the colonic epithelial crypt niche involves Hopx/Klf4 and Tcf7l2/Hnf4-α cascades.

    PubMed

    Lili, Loukia N; Farkas, Attila E; Gerner-Smidt, Christian; Overgaard, Christian E; Moreno, Carlos S; Parkos, Charles A; Capaldo, Christopher T; Nusrat, Asma

    2016-01-01

    Colonic enterocytes form a rapidly renewing epithelium and barrier to luminal antigens. During renewal, coordinated expression of the claudin family of genes is vital to maintain the epithelial barrier. Disruption of this process contributes to barrier compromise and mucosal inflammatory diseases. However, little is known about the regulation of this critical aspect of epithelial cell differentiation. In order to identify claudin regulatory factors we utilized high-throughput gene microarrays and correlation analyses. We identified complex expression gradients for the transcription factors Hopx, Hnf4a, Klf4 and Tcf7l2, as well as 12 claudins, during differentiation. In vitro confirmatory methods identified 2 pathways that stimulate claudin expression; Hopx/Klf4 activation of Cldn4, 7 and 15, and Tcf7l2/Hnf4a up-regulation of Cldn23. Chromatin immunoprecipitation confirmed a Tcf7l2/Hnf4a/Claudin23 cascade. Furthermore, Hnf4a conditional knockout mice fail to induce Cldn23 during colonocyte differentiation. In conclusion, we report a comprehensive screen of colonic claudin gene expression and discover spatiotemporal Hopx/Klf4 and Tcf7l2/Hnf4a signaling as stimulators of colonic epithelial barrier differentiation. PMID:27583195

  4. Tight junction disruption: Helicobacter pylori and dysregulation of the gastric mucosal barrier.

    PubMed

    Caron, Tyler J; Scott, Kathleen E; Fox, James G; Hagen, Susan J

    2015-10-28

    Long-term chronic infection with Helicobacter pylori (H. pylori) is a risk factor for gastric cancer development. In the multi-step process that leads to gastric cancer, tight junction dysfunction is thought to occur and serve as a risk factor by permitting the permeation of luminal contents across an otherwise tight mucosa. Mechanisms that regulate tight junction function and structure in the normal stomach, or dysfunction in the infected stomach, however, are largely unknown. Although conventional tight junction components are expressed in gastric epithelial cells, claudins regulate paracellular permeability and are likely the target of inflammation or H. pylori itself. There are 27 different claudin molecules, each with unique properties that render the mucosa an intact barrier that is permselective in a way that is consistent with cell physiology. Understanding the architecture of tight junctions in the normal stomach and then changes that occur during infection is important but challenging, because most of the reports that catalog claudin expression in gastric cancer pathogenesis are contradictory. Furthermore, the role of H. pylori virulence factors, such as cytotoxin-associated gene A and vacoulating cytotoxin, in regulating tight junction dysfunction during infection is inconsistent in different gastric cell lines and in vivo, likely because non-gastric epithelial cell cultures were initially used to unravel the details of their effects on the stomach. Hampering further study, as well, is the relative lack of cultured cell models that have tight junction claudins that are consistent with native tissues. This summary will review the current state of knowledge about gastric tight junctions, normally and in H. pylori infection, and make predictions about the consequences of claudin reorganization during H. pylori infection.

  5. Tight junction disruption: Helicobacter pylori and dysregulation of the gastric mucosal barrier

    PubMed Central

    Caron, Tyler J; Scott, Kathleen E; Fox, James G; Hagen, Susan J

    2015-01-01

    Long-term chronic infection with Helicobacter pylori (H. pylori) is a risk factor for gastric cancer development. In the multi-step process that leads to gastric cancer, tight junction dysfunction is thought to occur and serve as a risk factor by permitting the permeation of luminal contents across an otherwise tight mucosa. Mechanisms that regulate tight junction function and structure in the normal stomach, or dysfunction in the infected stomach, however, are largely unknown. Although conventional tight junction components are expressed in gastric epithelial cells, claudins regulate paracellular permeability and are likely the target of inflammation or H. pylori itself. There are 27 different claudin molecules, each with unique properties that render the mucosa an intact barrier that is permselective in a way that is consistent with cell physiology. Understanding the architecture of tight junctions in the normal stomach and then changes that occur during infection is important but challenging, because most of the reports that catalog claudin expression in gastric cancer pathogenesis are contradictory. Furthermore, the role of H. pylori virulence factors, such as cytotoxin-associated gene A and vacoulating cytotoxin, in regulating tight junction dysfunction during infection is inconsistent in different gastric cell lines and in vivo, likely because non-gastric epithelial cell cultures were initially used to unravel the details of their effects on the stomach. Hampering further study, as well, is the relative lack of cultured cell models that have tight junction claudins that are consistent with native tissues. This summary will review the current state of knowledge about gastric tight junctions, normally and in H. pylori infection, and make predictions about the consequences of claudin reorganization during H. pylori infection. PMID:26523106

  6. The guggulsterone derivative GG-52 inhibits NF-κB signaling in gastric epithelial cells and ameliorates ethanol-induced gastric mucosal lesions in mice.

    PubMed

    Kim, Jung Mogg; Kim, Su Hyun; Ko, Su Hyuk; Jung, Jireh; Chun, Jaeyoung; Kim, Nayoung; Jung, Hyun Chae; Kim, Joo Sung

    2013-01-15

    Gastric mucosal inflammation can develop after challenge with noxious stimuli such as alcohol. Specially, alcohol stimulates the release of inflammatory cytokines but does not increase gastric acid secretion, leading to gastric mucosal damage. The plant sterol guggulsterone and its novel derivative GG-52 have been reported to inhibit nuclear factor-κB (NF-κB) signaling in intestinal epithelial cells and experimental colitis. In the present study, we investigated the anti-inflammatory effects of GG-52 on gastric epithelial cells and on ethanol-induced gastric mucosal inflammation in mice. GG-52 inhibited the expression of interleukin-8 (IL-8) in gastric epithelial AGS and MKN-45 cell lines stimulated with tumor necrosis factor (TNF)-α in a dose-dependent manner. Pretreatment with GG-52 suppressed TNF-α-induced activation of IκB kinase (IKK) and NF-κB signaling in MKN-45 cells. In contrast, the inactive analog GG-46 did not produce significant changes in IL-8 expression or NF-κB activation. In a model of ethanol-induced murine gastritis, administration of GG-52 significantly reduced the severity of gastritis, as assessed by macroscopic and histological evaluation of gastric mucosal damage. In addition, the ethanol-induced upregulation of chemokine KC, a mouse homolog of IL-8, and phosphorylated p65 NF-κB signals were significantly inhibited in murine gastric mucosa pretreated with GG-52. These results indicate that GG-52 suppresses NF-κB activation in gastric epithelial cells and ameliorates ethanol-induced gastric mucosal lesions in mice, suggesting that GG-52 may be a potential gastroprotective agent.

  7. Intestinal mucosal barrier dysfunction participates in the progress of nonalcoholic fatty liver disease.

    PubMed

    Mao, Jing-Wei; Tang, Hai-Ying; Zhao, Ting; Tan, Xiao-Yan; Bi, Jian; Wang, Bing-Yuan; Wang, Ying-De

    2015-01-01

    Intestinal mucosal barrier dysfunction is closely related to liver diseases, which implies impaired gut-liver axis may play a role in the pathogenesis of NAFLD. In our study, rats were divided into three groups: normal chow diet (NCD) group, high-fat diet (HFD) group and TNBS-induced colitis with high-fat diet (C-HFD) group. Liver tissues were obtained for histological observation and TNF-α, IL-6 mRNA determination and blood samples were collected for liver enzymes and LPS analysis. Ultrastructural changes of jejuna epithelium, SIBO and amounts of CD103(+)MHCII(+)DCs and CD4(+)CD25(+)FoxP3(+)T-regs in terms of percentage in mesenteric lymph nodes (MLN) were observed by electron microscope, bacterial cultivation and flow cytometry, respectively. The results demonstrated the pathological characteristics accorded with nonalcoholic simple fatty liver (NAFL) and NASH in HFD group by week 8 and 12, respectively. Besides, the degree of hepatic steatosis and steatohepatitis was more severe in C-HFD group compared with HFD-group at the same time point. NAFLD activity score (NAS), liver enzymes, concentration of LPS and mRNA expressions of TNF-α, IL-6 were higher significantly in C-HFD group compared with HFD and NCD group at week 4, 8 and 12, respectively. In HFD group, epithelium microvilli atrophy, disruptive tight junctions and SIBO were present, and these changes were more severe in NASH compared with NAFL. The percentage of CD103+MHCII+DCs and CD4+CD25+FoxP3+T-regs decreased significantly in NAFL and NASH compared with NCD group. Our conclusion was that gut-liver axis was impaired in NAFLD, which played crucial role in the pathogenesis of NAFLD. PMID:26097546

  8. Effects of simulated weightlessness on the intestinal mucosal barrier of rats

    NASA Astrophysics Data System (ADS)

    Chen, Ying; Yang, Chun-min; Mao, Gao-ping; Liu, Qing-sen; Guo, Ming-zhou

    2011-07-01

    This study employed a rat tail-suspension model to investigate the effects of simulated weightlessness on the intestinal mucosal barrier. Twenty-four Wistar rats were randomly divided into control (CON), 14-day tail-suspension (SUS-14d), and 21-day tail-suspension (SUS-21d) groups ( n = 8 per group). Expression of occludin and zonula occludins-1 (ZO-1), proteins of the tight junction (TJ), in the intestinal mucosa was measured by immunohistochemical analysis, Western blotting, and mRNA fluorescent quantitation PCR. Plasma concentrations of diamine oxidase (DAO) and D-lactate were determined using an enzymatic spectrophotometric assay. Expression of occludin and ZO-1 was reduced in the SUS-14d and SUS-21d groups as compared to the CON group, with lowest expression observed in the SUS-21d group ( P < 0.01). Examination by transmission electron microscopy (TEM) of the jejunal epithelium revealed increased intercellular space, decreased TJ and desmosome densities, and destruction of microvilli in the SUS-14d and SUS-21d groups. Plasma DAO and D-lactate concentrations in the SUS-21d group were higher than those in SUS-14d group and significantly higher than those in the CON group ( P < 0.01). In all three groups, the expression of occludin and ZO-1 was found to correlate negatively with DAO ( P < 0.01) and D-lactate ( P < 0.01) concentrations. It is concluded that simulated weightless results in down-regulation of expression of TJ proteins in the rat intestinal mucosa. Simulated weightlessness is proposed to increase intestinal permeability through damage to the TJ.

  9. NFkB and Nrf2 in esophageal epithelial barrier function

    PubMed Central

    Chen, Hao; Fang, Yu; Li, Wenbo; Orlando, Roy C; Shaheen, Nicholas; Chen, Xiaoxin Luke

    2013-01-01

    The stratified squamous epithelium of the esophagus forms a tight protective barrier. Defects of the barrier function contribute to gastroesophageal reflux disease (GERD), which is manifested as damage to the esophageal epithelium due to exposure to the gastrointestinal refluxate. In this review, we discuss the involvement of NFkB and Nrf2 in esophageal epithelial barrier function. Understanding these molecular pathways in the esophagus may help us develop therapeutic strategies to improve clinical outcomes in patients with GERD. PMID:24790804

  10. Mucosal Vaccination Overcomes the Barrier to Recombinant Vaccinia Immunization Caused by Preexisting Poxvirus Immunity

    NASA Astrophysics Data System (ADS)

    Belyakov, Igor M.; Moss, Bernard; Strober, Warren; Berzofsky, Jay A.

    1999-04-01

    Overcoming preexisting immunity to vaccinia virus in the adult population is a key requirement for development of otherwise potent recombinant vaccinia vaccines. Based on our observation that s.c. immunization with vaccinia induces cellular and antibody immunity to vaccinia only in systemic lymphoid tissue and not in mucosal sites, we hypothesized that the mucosal immune system remains naive to vaccinia and therefore amenable to immunization with recombinant vaccinia vectors despite earlier vaccinia exposure. We show that mucosal immunization of vaccinia-immune BALB/c mice with recombinant vaccinia expressing HIV gp160 induced specific serum antibody and strong HIV-specific cytotoxic T lymphocyte responses. These responses occurred not only in mucosal but also in systemic lymphoid tissue, whereas systemic immunization was ineffective under these circumstances. In this context, intrarectal immunization was more effective than intranasal immunization. Boosting with a second dose of recombinant vaccinia was also more effective via the mucosal route. The systemic HIV-specific cytotoxic T lymphocyte response was enhanced by coadministration of IL-12 at the mucosal site. These results also demonstrate the independent compartmentalization of the mucosal versus systemic immune systems and the asymmetric trafficking of lymphocytes between them. This approach to circumvent previous vaccinia immunity may be useful for induction of protective immunity against infectious diseases and cancer in the sizable populations with preexisting immunity to vaccinia from smallpox vaccination.

  11. Herbal prescription Chang'an II repairs intestinal mucosal barrier in rats with post-inflammation irritable bowel syndrome

    PubMed Central

    Wang, Feng-yun; Su, Min; Zheng, Yong-qiu; Wang, Xiao-ge; Kang, Nan; Chen, Ting; Zhu, En-lin; Bian, Zhao-xiang; Tang, Xu-dong

    2015-01-01

    Aim: The herbal prescription Chang'an II is derived from a classical TCM formula Tong-Xie-Yao-Fang for the treatment of liver-qi stagnation and spleen deficiency syndrome of irritable bowel syndrome (IBS). In this study we investigated the effects of Chang'an II on the intestinal mucosal immune barrier in a rat post-inflammation IBS (PI-IBS) model. Methods: A rat model of PI-IBS was established using a multi-stimulation paradigm including early postnatal sibling deprivation, bondage and intrarectal administration of TNBS. Four weeks after TNBS administration, the rats were treated with Chang'an II (2.85, 5.71 and 11.42 g·kg−1·d−1, ig) for 14 d. Intestinal sensitivity was assessed based on the abdominal withdrawal reflex (AWR) scores and fecal water content. Open field test and two-bottle sucrose intake test were used to evaluate the behavioral changes. CD4+ and CD8+ cells were counted and IL-1β and IL-4 levels were measured in intestinal mucosa. Transmission electron microscopy was used to evaluate ultrastructural changes of the intestinal mucosal barrier. Results: PI-IBS model rats showed significantly increased AWR reactivity and fecal water content, and decreased locomotor activity and sucrose intake. Chang'an II treatment not only reduced AWR reactivity and fecal water content, but also suppressed the anxiety and depressive behaviors. Ultrastructural study revealed that the gut mucosal barrier function was severely damaged in PI-IBS model rats, whereas Chang'an II treatment relieved intestinal mucosal inflammation and repaired the gut mucosal barrier. Furthermore, PI-IBS model rats showed a significantly reduced CD4+/CD8+ cell ratio in lamina propria and submucosa, and increased IL-1β and reduced IL-4 expression in intestinal mucosa, whereas Chang'an II treatment reversed PI-IBS-induced changes in CD4+/CD8+ cell ratio and expression of IL-1β and IL-4. Conclusion: Chang'an II treatment protects the intestinal mucosa against PI-IBS through anti

  12. Constitutive activation of epithelial TLR4 augments inflammatory responses to mucosal injury and drives colitis-associated tumorigenesis

    PubMed Central

    Fukata, Masayuki; Shang, Limin; Santaolalla, Rebeca; Sotolongo, John; Pastorini, Cristhine; España, Cecilia; Ungaro, Ryan; Harpaz, Noam; Cooper, Harry S.; Elson, Greg; Kosco-Vilbois, Marie; Zaias, Julia; Perez, Maria T.; Mayer, Lloyd; Vamadevan, Arunan S.; Lira, Sergio A.; Abreu, Maria T.

    2010-01-01

    Chronic intestinal inflammation culminates in cancer and a link to TLR4 has been suggested by our observation that TLR4 deficiency prevents colitis-associated neoplasia. In the current study, we address the effect of the aberrant activation of epithelial TLR4 on induction of colitis and colitis-associated tumor development. We take a translational approach to address the consequences of increased TLR signaling in the intestinal mucosa. Mice transgenic for a constitutively-active TLR4 under the intestine-specific villin promoter (villin-TLR4 mice) were treated with DSS for acute colitis and azoxymethane-dextran sulfate sodium. TLR4 expression was analyzed by immunohistochemistry in colonic tissue from patients with ulcerative colitis and ulcerative colitis associated cancer. The effect of an antagonist TLR4 Ab was tested in prevention of colitis-associated neoplasia in the AOM-DSS model. Villin-TLR4 mice were highly susceptible to both acute colitis and colitis-associated neoplasia. Villin-TLR4 mice had increased epithelial expression of COX-2 and mucosal PGE2 production at baseline. Increased severity of colitis in villin-TLR4 mice was characterized by enhanced expression of inflammatory mediators and increased neutrophilic infiltration. In human UC samples, TLR4 expression was upregulated in almost all CAC and progressively increases with grade of dysplasia. As a proof of principle, a TLR4/MD-2 antagonist antibody inhibited colitis-associated neoplasia in the mouse model. Our results show that regulation of TLR's can affect the outcome of both acute colitis and its consequences—cancer. Targeting TLR4 and other TLR's may ultimately play a role in prevention or treatment of colitis-associated cancer. PMID:21674704

  13. Benign serrated colorectal fibroblastic polyps/intramucosal perineuriomas are true mixed epithelial-stromal polyps (hybrid hyperplastic polyp/mucosal perineurioma) with frequent BRAF mutations.

    PubMed

    Agaimy, Abbas; Stoehr, Robert; Vieth, Michael; Hartmann, Arndt

    2010-11-01

    Colorectal fibroblastic polyp and intramucosal perineurioma are 2 synonyms for a recently described benign mucosal lesion with a predilection for the rectosigmoid colon. These lesions are characterized by aggregates of bland spindled cells separating and distorting mucosal crypts. The latter frequently showed a serrated architecture. The pathogenesis of fibroblastic polyp/intramucosal perineurioma and the nature of serrated crypts observed in them are poorly understood. We analyzed the clinicopathological features of 29 fibroblastic polyps and investigated them for the first time for mutations known to be involved in serrated colorectal epithelial polyps (BRAF, KRAS, and PIK3CA). Patients were 23 women and 6 men with a mean age of 64 years (range: 47 to 84 y). All lesions represented asymptomatic solitary polyps (mean size 3.5 mm) localized predominantly in the rectosigmoid colon (81%). Hyperplastic polyps, classical adenoma, and sessile serrated adenoma/lesion coexisted in 12 (44%), 12 (44%), and 5 (17%) patients, respectively. All lesions showed irregular aggregates of bland spindled cells separating and distorting mucosal crypts. Serrated (hyperplastic) crypts were observed on the top or contiguous with the lesion in all cases. Immunohistochemistry revealed expression of at least one perineurial cell marker (epithelial membrane antigen, claudin-1, and glucose transporter-1) in 26 out of 27 lesions (96%), but expression of CD34 was less common (8 of 27; 30%). Immunostaining for hMLH1 showed a normal nuclear expression. Molecular analysis in 22 cases showed V600E BRAF mutation in 14 cases (63%) and KRAS mutation in 1 (4%). The remainder were wild-type for all 3 genes. Our results indicate that serrated fibroblastic polyps/intramucosal perineuriomas represent a unique type of mixed epithelial-stromal polyps (hybrid hyperplastic polyp/mucosal perineurioma). The perineurial stromal component might be derived from modified pericryptic fibroblasts as a consequence

  14. NITROTYROSINATION OF A TUBULIN INDUCES EPITHELIAL BARRIER DYSFUNCTION

    EPA Science Inventory

    Nitrotyrosination of a-Tubulin Induces Epithelial Transport Dysfunction. Yuh-Chin Huang, Lisa Dailey, Wen-Li Zhang and Ilona Jaspers. ORD, Environmental Protection Agency and CEMLB, University of North Carolina

    a-Tubulin undergoes a cyclic removal and readdition of tyrosin...

  15. Lipid rafts are disrupted in mildly inflamed intestinal microenvironments without overt disruption of the epithelial barrier.

    PubMed

    Bowie, Rachel V; Donatello, Simona; Lyes, Clíona; Owens, Mark B; Babina, Irina S; Hudson, Lance; Walsh, Shaun V; O'Donoghue, Diarmuid P; Amu, Sylvie; Barry, Sean P; Fallon, Padraic G; Hopkins, Ann M

    2012-04-15

    Intestinal epithelial barrier disruption is a feature of inflammatory bowel disease (IBD), but whether barrier disruption precedes or merely accompanies inflammation remains controversial. Tight junction (TJ) adhesion complexes control epithelial barrier integrity. Since some TJ proteins reside in cholesterol-enriched regions of the cell membrane termed lipid rafts, we sought to elucidate the relationship between rafts and intestinal epithelial barrier function. Lipid rafts were isolated from Caco-2 intestinal epithelial cells primed with the proinflammatory cytokine interferon-γ (IFN-γ) or treated with methyl-β-cyclodextrin as a positive control for raft disruption. Rafts were also isolated from the ilea of mice in which colitis had been induced in conjunction with in vivo intestinal permeability measurements, and lastly from intestinal biopsies of ulcerative colitis (UC) patients with predominantly mild or quiescent disease. Raft distribution was analyzed by measuring activity of the raft-associated enzyme alkaline phosphatase and by performing Western blot analysis for flotillin-1. Epithelial barrier integrity was estimated by measuring transepithelial resistance in cytokine-treated cells or in vivo permeability to fluorescent dextran in colitic mice. Raft and nonraft fractions were analyzed by Western blotting for the TJ proteins occludin and zonula occludens-1 (ZO-1). Our results revealed that lipid rafts were disrupted in IFN-γ-treated cells, in the ilea of mice with subclinical colitis, and in UC patients with quiescent inflammation. This was not associated with a clear pattern of occludin or ZO-1 relocalization from raft to nonraft fractions. Significantly, a time-course study in colitic mice revealed that disruption of lipid rafts preceded the onset of increased intestinal permeability. Our data suggest for the first time that lipid raft disruption occurs early in the inflammatory cascade in murine and human colitis and, we speculate, may contribute to

  16. 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.

  17. 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

  18. Cooperation of Salmonella pathogenicity islands 1 and 4 is required to breach epithelial barriers.

    PubMed

    Gerlach, Roman G; Cláudio, Nuno; Rohde, Manfred; Jäckel, Daniela; Wagner, Carolin; Hensel, Michael

    2008-11-01

    Invasion is an important microbial virulence strategy to overcome the barrier formed by polarized epithelial cells. Salmonella enterica is a food-borne pathogen that deploys a type III secretion system for the manipulation of the actin cytoskeleton and to trigger internalization into epithelial cells. Here we show that this function is not sufficient to enter polarized cells and report that penetration of epithelia from the luminal side requires both the type III secretion system and novel virulence functions conferred by Salmonella pathogenicity island 4. Salmonella pathogenicity island 4 encodes a type I secretion system for the giant non-fimbrial adhesin SiiE that mediates intimate contact of Salmonella to microvilli on the apical membrane. Mutant strains lacking SiiE fail to invade polarized cells, to destroy epithelial barrier functions and to efface the apical brush border. Deletion analyses of repetitive domains in SiiE indicate that the large size of the adhesin is of functional importance. Our observations demonstrate that efficient penetration of epithelial barriers requires the cooperative activity of two Salmonella pathogenicity islands encoding different secretion systems. These findings underline the role of the epithelial brush border and reveal a new mechanism used by bacterial pathogens to overcome this barrier.

  19. Ocular surface reconstruction with a tissue-engineered nasal mucosal epithelial cell sheet for the treatment of severe ocular surface diseases.

    PubMed

    Kobayashi, Masakazu; Nakamura, Takahiro; Yasuda, Makoto; Hata, Yuiko; Okura, Shoki; Iwamoto, Miyu; Nagata, Maho; Fullwood, Nigel J; Koizumi, Noriko; Hisa, Yasuo; Kinoshita, Shigeru

    2015-01-01

    Severe ocular surface diseases (OSDs) with severe dry eye can be devastating and are currently some of the most challenging eye disorders to treat. To investigate the feasibility of using an autologous tissue-engineered cultivated nasal mucosal epithelial cell sheet (CNMES) for ocular surface reconstruction, we developed a novel technique for the culture of nasal mucosal epithelial cells expanded ex vivo from biopsy-derived human nasal mucosal tissues. After the protocol, the CNMESs had 4-5 layers of stratified, well-differentiated cells, and we successfully generated cultured epithelial sheets, including numerous goblet cells. Immunohistochemistry confirmed the presence of keratins 3, 4, and 13; mucins 1, 16, and 5AC; cell junction and basement membrane assembly proteins; and stem/progenitor cell marker p75 in the CNMESs. We then transplanted the CNMESs onto the ocular surfaces of rabbits and confirmed the survival of this tissue, including the goblet cells, up to 2 weeks. The present report describes an attempt to overcome the problems of treating severe OSDs with the most severe dry eye by treating them using tissue-engineered CNMESs to supply functional goblet cells and to stabilize and reconstruct the ocular surface. The present study is a first step toward assessing the use of tissue-engineered goblet-cell transplantation of nonocular surface origin for ocular surface reconstruction.

  20. Side chain variations radically alter the diffusion of poly(2-alkyl-2-oxazoline) functionalised nanoparticles through a mucosal barrier.

    PubMed

    Mansfield, Edward D H; de la Rosa, Victor R; Kowalczyk, Radoslaw M; Grillo, Isabelle; Hoogenboom, Richard; Sillence, Katy; Hole, Patrick; Williams, Adrian C; Khutoryanskiy, Vitaliy V

    2016-08-16

    Functionalised nanomaterials are gaining popularity for use as drug delivery vehicles and, in particular, mucus penetrating nanoparticles may improve drug bioavailability via the oral route. To date, few polymers have been investigated for their muco-penetration, and the effects of systematic structural changes to polymer architectures on the penetration and diffusion of functionalised nanomaterials through mucosal tissue have not been reported. We investigated the influence of poly(2-oxazoline) alkyl side chain length on nanoparticle diffusion; poly(2-methyl-2-oxazoline), poly(2-ethyl-2-oxazoline), and poly(2-n-propyl-2-oxazoline) were grafted onto the surface of thiolated silica nanoparticles and characterised by FT-IR, Raman and NMR spectroscopy, thermogravimetric analysis, and small angle neutron scattering. Diffusion coefficients were determined in water and in a mucin dispersion (using Nanoparticle Tracking Analysis), and penetration through a mucosal barrier was assessed using an ex vivo fluorescence technique. The addition of a single methylene group in the side chain significantly altered the penetration and diffusion of the materials in both mucin dispersions and mucosal tissue. Nanoparticles functionalised with poly(2-methyl-2-oxazoline) were significantly more diffusive than particles with poly(2-ethyl-2-oxazoline) while particles with poly(2-n-propyl-2-oxazoline) showed no significant increase compared to the unfunctionalised particles. These data show that variations in the polymer structure can radically alter their diffusive properties with clear implications for the future design of mucus penetrating systems. PMID:27400181

  1. Bile salts disrupt human esophageal squamous epithelial barrier function by modulating tight junction proteins.

    PubMed

    Chen, Xin; Oshima, Tadayuki; Shan, Jing; Fukui, Hirokazu; Watari, Jiro; Miwa, Hiroto

    2012-07-15

    Reflux of acid and bile acids contributes to epithelial tissue injury in gastro-esophageal reflux disease. However, the influence of refluxed material on human esophageal stratified epithelial barrier function and tight junction (TJ) proteins has not been fully elucidated. Here, we investigated the influence of acid and bile acids on barrier function and TJ protein distribution using a newly developed air-liquid interface (ALI) in vitro culture model of stratified squamous epithelium based on primary human esophageal epithelial cells (HEECs). Under ALI conditions, HEECs formed distinct epithelial layers on Transwell inserts after 7 days of culture. The epithelial layers formed TJ, and the presence of claudin-1, claudin-4, and occludin were detected by immunofluorescent staining. The NP-40-insoluble fraction of these TJ proteins was significantly higher by day 7 of ALI culture. Exposure of HEECs to pH 2, and taurocholic acid (TCA) and glycocholic acid (GCA) at pH 3, but not pH 4, for 1 h decreased transepithelial electrical resistance (TEER) and increased paracellular permeability. Exposure of cell layers to GCA (pH 3) and TCA (pH 3) for 1 h also markedly reduced the insoluble fractions of claudin-1 and -4. We found that deoxycholic acid (pH 7.4 or 6, 1 h) and pepsin (pH 3, 24 h) significantly decreased TEER and increased permeability. Based on these findings, ALI-cultured HEECs represent a new in vitro model of human esophageal stratified epithelium and are suitable for studying esophageal epithelial barrier functions. Using this model, we demonstrated that acid, bile acids, and pepsin disrupt squamous epithelial barrier function partly by modulating TJ proteins. These results provide new insights into understanding the role of TJ proteins in esophagitis.

  2. Measles virus breaks through epithelial cell barriers to achieve transmission

    PubMed Central

    Takeda, Makoto

    2008-01-01

    Measles is a highly contagious disease that causes immunosuppression in patients. Measles virus infection has been thought to begin in the respiratory epithelium and then spread to lymphoid tissue. In this issue of the JCI, Leonard et al. provide data to suggest an alternative model of measles virus pathogenesis (see the related article beginning on page 2448). In human primary epithelial cells and rhesus monkeys in vivo, the authors show that initial infection of respiratory epithelium is not necessary for the virus to enter the host but that viral entry into epithelial cells via interaction of the virus with a receptor located on the basolateral side of the epithelium is required for viral shedding into the airway and subsequent transmission. PMID:18568081

  3. NLRP3 protects alveolar barrier integrity by an inflammasome-independent increase of epithelial cell adherence

    PubMed Central

    Kostadinova, Elena; Chaput, Catherine; Gutbier, Birgitt; Lippmann, Juliane; Sander, Leif E.; Mitchell, Timothy J.; Suttorp, Norbert; Witzenrath, Martin; Opitz, Bastian

    2016-01-01

    Bacterial pneumonia is a major cause of acute lung injury and acute respiratory distress syndrome, characterized by alveolar barrier disruption. NLRP3 is best known for its ability to form inflammasomes and to regulate IL-1β and IL-18 production in myeloid cells. Here we show that NLRP3 protects the integrity of the alveolar barrier in a mouse model of Streptococcus pneumoniae-induced pneumonia, and ex vivo upon treatment of isolated perfused and ventilated lungs with the purified bacterial toxin, pneumolysin. We reveal that the preserving effect of NLRP3 on the lung barrier is independent of inflammasomes, IL-1β and IL-18. NLRP3 improves the integrity of alveolar epithelial cell monolayers by enhancing cellular adherence. Collectively, our study uncovers a novel function of NLRP3 by demonstrating that it protects epithelial barrier function independently of inflammasomes. PMID:27476670

  4. Metabolic regulation of intestinal epithelial barrier during inflammation

    PubMed Central

    Colgan, Sean P; Curtis, Valerie F; Lanis, Jordi M; Glover, Louise E

    2014-01-01

    The gastrointestinal mucosa has proven to be an interesting tissue for which to investigate disease-related metabolism. In this review, we outline some evidence that implicates metabolic signaling as important features of barrier in the healthy and disease. Studies from cultured cell systems, animal models and human patients have revealed that metabolites generated within the inflammatory microenvironment are central to barrier regulation. These studies have revealed a prominent role for hypoxia and hypoxia-inducible factor (HIF) at key steps in adenine nucleotide metabolism and within the creatine kinase pathway. Results from animal models of intestinal inflammation have demonstrated an almost uniformly beneficial influence of HIF stabilization on disease outcomes and barrier function. Studies underway to elucidate the contribution of immune responses will provide additional insight into how metabolic changes contribute to the complexity of the gastrointestinal tract and how such information might be harnessed for therapeutic benefit. PMID:25838978

  5. 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

  6. Hypertonic challenge to porcine vocal folds: Effects on epithelial barrier function

    PubMed Central

    Sivasankar, Mahalakshmi; Erickson, Elizabeth; Rosenblatt, Mark; Branski, Ryan C.

    2010-01-01

    Objective Dehydration challenges can increase the chemical composition of surface fluid overlying vocal fold epithelia (hypertonic surface fluid). The vocal fold epithelium is posited to act as a barrier, shielding the lamina propria from perturbations in the airway lumen. However, the effects of hypertonic surface fluid on the barrier functions of vocal fold epithelia have not been quantified. We, therefore, sought to investigate whether hypertonic surface fluid compromises epithelial barrier function. We examined the effects of hypertonic surface fluid on vocal fold epithelial resistance, paracellular pathway morphology, and tight junction protein integrity. Study Design Ex vivo, between group design. Setting Laboratory. Methods Porcine vocal folds (n = 24) were exposed to hypertonic or isotonic challenge and examined by electrophysiology, transmission electron microscopy, and Western blot analyses. Results Hypertonic, but not isotonic, challenge significantly reduced transepithelial resistance. This decrease in resistance was observed immediately after the challenge and was consistent with the appearance of dilated paracellular pathway morphology. However, hypertonic challenge did not alter protein levels of occludin, zona occludens-1, E-cadherin, or β-catenin. Conclusion Hypertonic surface fluid alters epithelial barrier function in the vocal folds. Specifically, exposure to hypertonic challenges increases epithelial permeability. Given the important role of the vocal fold epithelium in shielding the underlying mucosa from inhaled pathogens and pollutants, our data provide the impetus for future studies on pharmacological treatments aimed at restoring the hydration level and chemical composition of vocal fold surface fluid. PMID:20096227

  7. Vocal Fold Epithelial Barrier in Health and Injury: A Research Review

    ERIC Educational Resources Information Center

    Levendoski, Elizabeth Erickson; Leydon, Ciara; Thibeault, Susan L.

    2014-01-01

    Purpose: Vocal fold epithelium is composed of layers of individual epithelial cells joined by junctional complexes constituting a unique interface with the external environment. This barrier provides structural stability to the vocal folds and protects underlying connective tissue from injury while being nearly continuously exposed to potentially…

  8. Low molecular weight components of pollen alter bronchial epithelial barrier functions.

    PubMed

    Blume, Cornelia; Swindle, Emily J; Gilles, Stefanie; Traidl-Hoffmann, Claudia; Davies, Donna E

    2015-01-01

    The bronchial epithelium plays a key role in providing a protective barrier against many environmental substances of anthropogenic or natural origin which enter the lungs during breathing. Appropriate responses to these agents are critical for regulation of tissue homeostasis, while inappropriate responses may contribute to disease pathogenesis. Here, we compared epithelial barrier responses to different pollen species, characterized the active pollen components and the signaling pathways leading to epithelial activation. Polarized bronchial cells were exposed to extracts of timothy grass (Phleum pratense), ragweed (Ambrosia artemisifolia), mugwort (Artemisia vulgaris), birch (Betula alba) and pine (Pinus sylvestris) pollens. All pollen species caused a decrease in ionic permeability as monitored trans-epithelial electrical resistance (TER) and induced polarized release of mediators analyzed by ELISA, with grass pollen showing the highest activity. Ultrafiltration showed that the responses were due to components <3kDa. However, lipid mediators, including phytoprostane E1, had no effect on TER, and caused only modest induction of mediator release. Reverse-phase chromatography separated 2 active fractions: the most hydrophilic maximally affected cytokine release whereas the other only affected TER. Inhibitor studies revealed that JNK played a more dominant role in regulation of barrier permeability in response to grass pollen exposure, whereas ERK and p38 controlled cytokine release. Adenosine and the flavonoid isorhamnetin present in grass pollen contributed to the overall effect on airway epithelial barrier responses. In conclusion, bronchial epithelial barrier functions are differentially affected by several low molecular weight components released by pollen. Furthermore, ionic permeability and innate cytokine production are differentially regulated. PMID:26451347

  9. The pH-sensing receptor OGR1 improves barrier function of epithelial cells and inhibits migration in an acidic environment.

    PubMed

    de Vallière, Cheryl; Vidal, Solange; Clay, Ieuan; Jurisic, Giorgia; Tcymbarevich, Irina; Lang, Silvia; Ludwig, Marie-Gabrielle; Okoniewski, Michal; Eloranta, Jyrki J; Kullak-Ublick, Gerd A; Wagner, Carsten A; Rogler, Gerhard; Seuwen, Klaus

    2015-09-15

    The pH-sensing receptor ovarian cancer G protein-coupled receptor 1 (OGR1; GPR68) is expressed in the gut. Inflammatory bowel disease is typically associated with a decrease in local pH, which may lead to altered epithelial barrier function and subsequent gastrointestinal repair involving epithelial cell adhesion and migration. As the mechanisms underlying the response to pH changes are not well understood, we have investigated OGR1-mediated, pH-dependent signaling pathways in intestinal epithelial cells. Caco-2 cells stably overexpressing OGR1 were created and validated as tools to study OGR1 signaling. Barrier function, migration, and proliferation were measured using electric cell-substrate impedance-sensing technology. Localization of the tight junction proteins zonula occludens protein 1 and occludin and the rearrangement of cytoskeletal actin were examined by confocal microscopy. Paracellular permeability and protein and gene expression analysis using DNA microarrays were performed on filter-grown Caco-2 monolayers. We report that an acidic pH shift from pH 7.8 to 6.6 improved barrier function and stimulated reorganization of filamentous actin with prominent basal stress fiber formation. Cell migration and proliferation during in vitro wound healing were inhibited. Gene expression analysis revealed significant upregulation of genes related to cytoskeleton remodeling, cell adhesion, and growth factor signaling. We conclude that acidic extracellular pH can have a signaling function and impact the physiology of intestinal epithelial cells. The deconstruction of OGR1-dependent signaling may aid our understanding of mucosal inflammation mechanisms.

  10. MicroRNAs regulate tight junction proteins and modulate epithelial/endothelial barrier functions

    PubMed Central

    Cichon, Christoph; Sabharwal, Harshana; Rüter, Christian; Schmidt, M Alexander

    2014-01-01

    Tightly controlled epithelial and endothelial barriers are a prerequisite for life as these barriers separate multicellular organisms from their environment and serve as first lines of defense. Barriers between neighboring epithelial cells are formed by multiple intercellular junctions including the ‘apical junctional complex—AJC’ with tight junctions (TJ), adherens junctions (AJ), and desmosomes. TJ consist of tetraspan transmembrane proteins like occludin, various claudins that directly control paracellular permeability, and the ‘Junctional Adhesion Molecules’ (JAMs). For establishing tight barriers TJ are essential but at the same time have to allow also selective permeability. For this, TJ need to be tightly regulated and controlled. This is organized by a variety of adaptor molecules, i.e., protein kinases, phosphatases and GTPases, which in turn are regulated and fine-tuned involving microRNAs (miRNAs). In this review we summarize available data on the role and targeting of miRNAs in the maintenance of epithelial and/or endothelial barriers. PMID:25610754

  11. 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.

  12. Dietary Lactobacillus rhamnosus GG Supplementation Improves the Mucosal Barrier Function in the Intestine of Weaned Piglets Challenged by Porcine Rotavirus

    PubMed Central

    Mao, Xiangbing; Gu, Changsong; Hu, Haiyan; Tang, Jun; Chen, Daiwen; Yu, Bing; He, Jun; Yu, Jie; Luo, Junqiu; Tian, Gang

    2016-01-01

    Lactobacillus rhamnosus GG (LGG) has been regarded as a safe probiotic strain. The aim of this study was to investigate whether dietary LGG supplementation could alleviate diarrhea via improving jejunal mucosal barrier function in the weaned piglets challenged by RV, and further analyze the potential roles for apoptosis of jejunal mucosal cells and intestinal microbiota. A total of 24 crossbred barrows weaned at 21 d of age were assigned randomly to 1 of 2 diets: the basal diet and LGG supplementing diet. On day 11, all pigs were orally infused RV or the sterile essential medium. RV infusion increased the diarrhea rate, increased the RV-Ab, NSP4 and IL-2 concentrations and the Bax mRNA levels of jejunal mucosa (P<0.05), decreased the villus height, villus height: crypt depth, the sIgA, IL-4 and mucin 1 concentrations and the ZO-1, occludin and Bcl-2 mRNA levels of jejunal mucosa (P<0.05), and affected the microbiota of ileum and cecum (P<0.05) in the weaned pigs. Dietary LGG supplementation increased the villus height and villus height: crypt depth, the sIgA, IL-4, mucin 1 and mucin 2 concentrations, and the ZO-1, occludin and Bcl-2 mRNA levels of the jejunal mucosa (P<0.05) reduced the Bax mRNA levels of the jejunal mucosa (P<0.05) in weaned pigs. Furthermore, dietary LGG supplementation alleviated the increase of diarrhea rate in the weaned pigs challenged by RV (P<0.05), and relieve the effect of RV infection on the villus height, crypt depth and the villus height: crypt depth of the jejunal mucosa (P<0.05), the NSP4, sIgA, IL-2, IL-4, mucin 1 and mucin 2 concentrations of jejunal mucosa (P<0.05), the ZO-1, occludin, Bax and Bcl-2 mRNA levels of the jejunal mucosa (P<0.05), and the microbiota of ileum and cecum (P<0.05) in the weaned pigs challenged by RV. These results suggest that supplementing LGG in diets alleviated the diarrhea of weaned piglets challenged by RV via inhibiting the virus multiplication and improving the jejunal mucosal barrier function

  13. Modified Pulsatilla decoction attenuates oxazolone-induced colitis in mice through suppression of inflammation and epithelial barrier disruption

    PubMed Central

    Wang, Xuewei; Fan, Fugang; Cao, Qin

    2016-01-01

    Inflammatory bowel diseases (IBDs) are chronic inflammatory gastrointestinal disorders caused by a dysregulated mucosal immune response and epithelial barrier disruption. Conventional treatment of IBD is currently limited to overcoming patient symptoms and is often associated with severe adverse effects from the drugs used. Modified Pulsatilla decoction has been used previously to treat ulcerative colitis (UC) in clinical practice in China, however, the underlying mechanism in the treatment of UC remains to be elucidated. In the present study, the efficiency and mechanisms of modified Pulsatilla decoction in the treatment of oxazolone-induced colitis were investigated. Assessment of clinical colitis and histological examination found that the administration of modified Pulsatilla decoction attenuated the severity of oxazolone-induced colitis in mice. Measurement of cytokine concentration, western blotting and reverse transcription-quantitative polymerase chain reaction demonstrated modified Pulsatilla decoction treatment significantly reduced the secretion of pro-inflammatory cytokines and restored alterations in tight junction proteins in the colon tissues. In addition, modified Pulsatilla decoction suppressed the activation of the nuclear factor-κB signaling pathway. Thus, the findings of the present study demonstrated that modified Pulsatilla decoction offers an effective therapeutic approach for the treatment of IBD and revealed the underlying mechanisms of action offered by modified Pulsatilla decoction. PMID:27278299

  14. Heparan sulfate and syndecan-1 are essential in maintaining murine and human intestinal epithelial barrier function

    PubMed Central

    Bode, Lars; Salvestrini, Camilla; Park, Pyong Woo; Li, Jin-Ping; Esko, Jeffrey D.; Yamaguchi, Yu; Murch, Simon; Freeze, Hudson H.

    2007-01-01

    Patients with protein-losing enteropathy (PLE) fail to maintain intestinal epithelial barrier function and develop an excessive and potentially fatal efflux of plasma proteins. PLE occurs in ostensibly unrelated diseases, but emerging commonalities in clinical observations recently led us to identify key players in PLE pathogenesis. These include elevated IFN-γ, TNF-α, venous hypertension, and the specific loss of heparan sulfate proteoglycans from the basolateral surface of intestinal epithelial cells during PLE episodes. Here we show that heparan sulfate and syndecan-1, the predominant intestinal epithelial heparan sulfate proteoglycan, are essential in maintaining intestinal epithelial barrier function. Heparan sulfate– or syndecan-1–deficient mice and mice with intestinal-specific loss of heparan sulfate had increased basal protein leakage and were far more susceptible to protein loss induced by combinations of IFN-γ, TNF-α, and increased venous pressure. Similarly, knockdown of syndecan-1 in human epithelial cells resulted in increased basal and cytokine-induced protein leakage. Clinical application of heparin has been known to alleviate PLE in some patients but its unknown mechanism and severe side effects due to its anticoagulant activity limit its usefulness. We demonstrate here that non-anticoagulant 2,3-de-O-sulfated heparin could prevent intestinal protein leakage in syndecan-deficient mice, suggesting that this may be a safe and effective therapy for PLE patients. PMID:18064305

  15. Human alveolar epithelial cells expressing tight junctions to model the air-blood barrier.

    PubMed

    Kuehn, Anna; Kletting, Stephanie; de Souza Carvalho-Wodarz, Cristiane; Repnik, Urska; Griffiths, Gareth; Fischer, Ulrike; Meese, Eckart; Huwer, Hanno; Wirth, Dagmar; May, Tobias; Schneider-Daum, Nicole; Lehr, Claus-Michael

    2016-01-01

    This paper describes a new human alveolar epithelial cell line (hAELVi - human Alveolar Epithelial Lentivirus immortalized) with type I-like characteristics and functional tight junctions, suitable to model the air-blood barrier of the peripheral lung. Primary human alveolar epithelial cells were immortalized by a novel regimen, grown as monolayers on permeable filter supports and characterized morphologically, biochemically and biophysically. hAELVi cells maintain the capacity to form tight intercellular junctions, with high trans-epithelial electrical resistance (> 1000 Ω*cm²). The cells could be kept in culture over several days, up to passage 75, under liquid-liquid as well as air-liquid conditions. Ultrastructural analysis and real time PCR revealed type I-like cell properties, such as the presence of caveolae, expression of caveolin-1, and absence of surfactant protein C. Accounting for the barrier properties, inter-digitations sealed with tight junctions and desmosomes were also observed. Low permeability of the hydrophilic marker sodium fluorescein confirmed the suitability of hAELVi cells for in vitro transport studies across the alveolar epithelium. These results suggest that hAELVi cells reflect the essential features of the air-blood barrier, as needed for an alternative to animal testing to study absorption and toxicity of inhaled drugs, chemicals and nanomaterials. PMID:26985677

  16. Arsenic Compromises Conducting Airway Epithelial Barrier Properties in Primary Mouse and Immortalized Human Cell Cultures

    PubMed Central

    Sherwood, Cara L.; Liguori, Andrew E.; Olsen, Colin E.; Lantz, R. Clark; Burgess, Jefferey L.; Boitano, Scott

    2013-01-01

    Arsenic is a lung toxicant that can lead to respiratory illness through inhalation and ingestion, although the most common exposure is through contaminated drinking water. Lung effects reported from arsenic exposure include lung cancer and obstructive lung disease, as well as reductions in lung function and immune response. As part of their role in innate immune function, airway epithelial cells provide a barrier that protects underlying tissue from inhaled particulates, pathogens, and toxicants frequently found in inspired air. We evaluated the effects of a five-day exposure to environmentally relevant levels of arsenic {<4μM [~300 μg/L (ppb)] as NaAsO2} on airway epithelial barrier function and structure. In a primary mouse tracheal epithelial (MTE) cell model we found that both micromolar (3.9 μM) and submicromolar (0.8 μM) arsenic concentrations reduced transepithelial resistance, a measure of barrier function. Immunofluorescent staining of arsenic-treated MTE cells showed altered patterns of localization of the transmembrane tight junction proteins claudin (Cl) Cl-1, Cl-4, Cl-7 and occludin at cell-cell contacts when compared with untreated controls. To better quantify arsenic-induced changes in tight junction transmembrane proteins we conducted arsenic exposure experiments with an immortalized human bronchial epithelial cell line (16HBE14o-). We found that arsenic exposure significantly increased the protein expression of Cl-4 and occludin as well as the mRNA levels of Cl-4 and Cl-7 in these cells. Additionally, arsenic exposure resulted in altered phosphorylation of occludin. In summary, exposure to environmentally relevant levels of arsenic can alter both the function and structure of airway epithelial barrier constituents. These changes likely contribute to the observed arsenic-induced loss in basic innate immune defense and increased infection in the airway. PMID:24349408

  17. Permeabilization of the Blood-Brain Barrier via Mucosal Engrafting: Implications for Drug Delivery to the Brain

    PubMed Central

    Bleier, Benjamin S.; Kohman, Richie E.; Feldman, Rachel E.; Ramanlal, Shreshtha; Han, Xue

    2013-01-01

    Utilization of neuropharmaceuticals for central nervous system(CNS) disease is highly limited due to the blood-brain barrier(BBB) which restricts molecules larger than 500Da from reaching the CNS. The development of a reliable method to bypass the BBB would represent an enormous advance in neuropharmacology enabling the use of many potential disease modifying therapies. Previous attempts such as transcranial catheter implantation have proven to be temporary and associated with multiple complications. Here we describe a novel method of creating a semipermeable window in the BBB using purely autologous tissues to allow for high molecular weight(HMW) drug delivery to the CNS. This approach is inspired by recent advances in human endoscopic transnasal skull base surgical techniques and involves engrafting semipermeable nasal mucosa within a surgical defect in the BBB. The mucosal graft thereby creates a permanent transmucosal conduit for drugs to access the CNS. The main objective of this study was to develop a murine model of this technique and use it to evaluate transmucosal permeability for the purpose of direct drug delivery to the brain. Using this model we demonstrate that mucosal grafts allow for the transport of molecules up to 500 kDa directly to the brain in both a time and molecular weight dependent fashion. Markers up to 40 kDa were found within the striatum suggesting a potential role for this technique in the treatment of Parkinson’s disease. This proof of principle study demonstrates that mucosal engrafting represents the first permanent and stable method of bypassing the BBB thereby providing a pathway for HMW therapeutics directly into the CNS. PMID:23637885

  18. Campylobacter infection in chickens modulates the intestinal epithelial barrier function.

    PubMed

    Awad, Wageha A; Molnár, Andor; Aschenbach, Jörg R; Ghareeb, Khaled; Khayal, Basel; Hess, Claudia; Liebhart, Dieter; Dublecz, Károly; Hess, Michael

    2015-02-01

    Asymptomatic carriage of Campylobacter jejuni is highly prevalent in chicken flocks. Thus, we investigated whether chronic Campylobacter carriage affects chicken intestinal functions despite the absence of clinical symptoms. An experiment was carried out in which commercial chickens were orally infected with C. jejuni (1 × 10(8) CFU/bird) at 14 days of life. Changes in ion transport and barrier function were assessed by short-circuit current (I(sc)) and transepithelial ion conductance (G(t)) in Ussing chambers. G(t) increased in cecum and colon of Campylobacter-infected chicken 7 d post-infection (DPI), whereas G t initially decreased in the jejunum at 7 DPI and increased thereafter at 14 DPI. The net charge transfer across the epithelium was reduced or tended to be reduced in all segments, as evidenced by a decreased I sc. Furthermore, the infection induced intestinal histomorphological changes, most prominently including a decrease in villus height, crypt depth and villus surface area in the jejunum at 7 DPI. Furthermore, body mass gain was decreased by Campylobacter carriage. This study demonstrates, for the first time, changes in the intestinal barrier function in Campylobacter-infected chickens and these changes were associated with a decrease in growth performance in otherwise healthy-appearing birds.

  19. Effect of carbon nanoparticles on renal epithelial cell structure, barrier function, and protein expression.

    PubMed

    Blazer-Yost, Bonnie L; Banga, Amiraj; Amos, Adam; Chernoff, Ellen; Lai, Xianyin; Li, Cheng; Mitra, Somenath; Witzmann, Frank A

    2011-09-01

    To assess effects of carbon nanoparticle (CNP) exposure on renal epithelial cells, fullerenes (C(60)), single-walled carbon nanotubes (SWNT), and multi-walled carbon nanotubes (MWNT) were incubated with a confluent renal epithelial line for 48 h. At low concentrations, CNP-treated cells exhibited significant decreases in transepithelial electrical resistance (TEER) but no changes in hormone-stimulated ion transport or CNP-induced toxicity or stress responses as measured by lactate dehydrogenase or cytokine release. The changes in TEER, manifested as an inverse relationship with CNP concentration, were mirrored by an inverse correlation between dose and changes in protein expression. Lower, more physiologically relevant, concentrations of CNP have the most profound effects on barrier cell function and protein expression. These results indicate an impact of CNPs on renal epithelial cells at concentrations lower than have been previously studied and suggest caution with regard to increasing CNP levels entering the food chain due to increasing environmental pollution. PMID:21067278

  20. Acute Acrolein Exposure Induces Impairment of Vocal Fold Epithelial Barrier Function

    PubMed Central

    Zheng, Wei; Sivasankar, M. Preeti

    2016-01-01

    Acrolein is a ubiquitous pollutant abundant in cigarette smoke, mobile exhaust, and industrial waste. There is limited literature on the effects of acrolein on vocal fold tissue, although there are clinical reports of voice changes after pollutant exposures. Vocal folds are responsible for voice production. The overall objective of this study was to investigate the effects of acrolein exposure on viable, excised vocal fold epithelial tissue and to characterize the mechanism underlying acrolein toxicity. Vocal fold epithelia were studied because they form the outermost layer of the vocal folds and are a primary recipient of inhaled pollutants. Porcine vocal fold epithelia were exposed to 0, 50, 100, 500, 900 or 1300 μM of acrolein for 3 hours; the metabolic activity, epithelial resistance, epithelial permeability, tight junction protein (occludin and claudin 3) expression, cell membrane integrity and lipid peroxidation were investigated. The data demonstrated that acrolein exposure at 500 μM significantly reduced vocal fold epithelial metabolic activity by 27.2% (p≤0.001). Incubation with 100 μM acrolein caused a marked increase in epithelial permeability by 130.5% (p<0.05) and a reduction in transepithelial electrical resistance (TEER) by 180.0% (p<0.001). While the expression of tight junctional protein did not change in acrolein-treated samples, the cell membrane integrity was significantly damaged with a 45.6% increase of lipid peroxidation as compared to controls (p<0.05). Taken together, these data provide evidence that acute acrolein exposure impairs vocal fold epithelial barrier integrity. Lipid peroxidation-induced cell membrane damage may play an important role in reducing the barrier function of the epithelium. PMID:27643990

  1. Acute Acrolein Exposure Induces Impairment of Vocal Fold Epithelial Barrier Function.

    PubMed

    Liu, Xinxin; Zheng, Wei; Sivasankar, M Preeti

    2016-01-01

    Acrolein is a ubiquitous pollutant abundant in cigarette smoke, mobile exhaust, and industrial waste. There is limited literature on the effects of acrolein on vocal fold tissue, although there are clinical reports of voice changes after pollutant exposures. Vocal folds are responsible for voice production. The overall objective of this study was to investigate the effects of acrolein exposure on viable, excised vocal fold epithelial tissue and to characterize the mechanism underlying acrolein toxicity. Vocal fold epithelia were studied because they form the outermost layer of the vocal folds and are a primary recipient of inhaled pollutants. Porcine vocal fold epithelia were exposed to 0, 50, 100, 500, 900 or 1300 μM of acrolein for 3 hours; the metabolic activity, epithelial resistance, epithelial permeability, tight junction protein (occludin and claudin 3) expression, cell membrane integrity and lipid peroxidation were investigated. The data demonstrated that acrolein exposure at 500 μM significantly reduced vocal fold epithelial metabolic activity by 27.2% (p≤0.001). Incubation with 100 μM acrolein caused a marked increase in epithelial permeability by 130.5% (p<0.05) and a reduction in transepithelial electrical resistance (TEER) by 180.0% (p<0.001). While the expression of tight junctional protein did not change in acrolein-treated samples, the cell membrane integrity was significantly damaged with a 45.6% increase of lipid peroxidation as compared to controls (p<0.05). Taken together, these data provide evidence that acute acrolein exposure impairs vocal fold epithelial barrier integrity. Lipid peroxidation-induced cell membrane damage may play an important role in reducing the barrier function of the epithelium. PMID:27643990

  2. A novel dual-flow bioreactor simulates increased fluorescein permeability in epithelial tissue barriers.

    PubMed

    Giusti, Serena; Sbrana, Tommaso; La Marca, Margherita; Di Patria, Valentina; Martinucci, Valentina; Tirella, Annalisa; Domenici, Claudio; Ahluwalia, Arti

    2014-09-01

    Permeability studies across epithelial barriers are of primary importance in drug delivery as well as in toxicology. However, traditional in vitro models do not adequately mimic the dynamic environment of physiological barriers. Here, we describe a novel two-chamber modular bioreactor for dynamic in vitro studies of epithelial cells. The fluid dynamic environment of the bioreactor was characterized using computational fluid dynamic models and measurements of pressure gradients for different combinations of flow rates in the apical and basal chambers. Cell culture experiments were then performed with fully differentiated Caco-2 cells as a model of the intestinal epithelium, comparing the effect of media flow applied in the bioreactor with traditional static transwells. The flow increases barrier integrity and tight junction expression of Caco-2 cells with respect to the static controls. Fluorescein permeability increased threefold in the dynamic system, indicating that the stimulus induced by flow increases transport across the barrier, closely mimicking the in vivo situation. The results are of interest for studying the influence of mechanical stimuli on cells, and underline the importance of developing more physiologically relevant in vitro tissue models. The bioreactor can be used to study drug delivery, chemical, or nanomaterial toxicity and to engineer barrier tissues.

  3. Culture of choroid plexus epithelial cells and in vitro model of blood-CSF barrier.

    PubMed

    Monnot, Andrew D; Zheng, Wei

    2013-01-01

    Chemical homeostasis in the extracellular fluid of the central nervous system (CNS) is maintained by two brain barrier systems, i.e., the blood-brain barrier (BBB) that separates the blood circulation from brain interstitial fluid and the blood-cerebrospinal fluid barrier (BCB) that separates the blood from the cerebrospinal fluid (CSF). The choroid plexus, where the BCB is located, is a polarized tissue, with the basolateral side of the choroidal epithelium facing the blood and the apical microvilli in direct contact with the CSF. The tissue plays a wide range of roles in brain development, aging, nutrient transport, endocrine regulation, and pathogenesis of certain neurodegenerative disorders. This chapter describes two in vitro cultures that have been well established to allow for study of the BCB structure and function. The primary choroidal epithelial cell culture can be established from rat choroid plexus tissue, and a similar immortalized murine choroidal epithelial cell culture known as Z310 cells has also been established. Both cultures display a dominant polygonal morphology, and immunochemical studies demonstrate the presence of transthyretin, a thyroxine transport protein known to be exclusively produced by the choroidal epithelia in the CNS. These cultures have been adapted for use on freely permeable Transwell(®) membranes sandwiched between two culture chambers, facilitating transport studies of various compounds across this barrier in vitro. These choroidal epithelia cultures with the Transwell system will perceivably assist blood-CSF barrier research. PMID:23097098

  4. Culture of choroid plexus epithelial cells and in vitro model of blood-CSF barrier.

    PubMed

    Monnot, Andrew D; Zheng, Wei

    2013-01-01

    Chemical homeostasis in the extracellular fluid of the central nervous system (CNS) is maintained by two brain barrier systems, i.e., the blood-brain barrier (BBB) that separates the blood circulation from brain interstitial fluid and the blood-cerebrospinal fluid barrier (BCB) that separates the blood from the cerebrospinal fluid (CSF). The choroid plexus, where the BCB is located, is a polarized tissue, with the basolateral side of the choroidal epithelium facing the blood and the apical microvilli in direct contact with the CSF. The tissue plays a wide range of roles in brain development, aging, nutrient transport, endocrine regulation, and pathogenesis of certain neurodegenerative disorders. This chapter describes two in vitro cultures that have been well established to allow for study of the BCB structure and function. The primary choroidal epithelial cell culture can be established from rat choroid plexus tissue, and a similar immortalized murine choroidal epithelial cell culture known as Z310 cells has also been established. Both cultures display a dominant polygonal morphology, and immunochemical studies demonstrate the presence of transthyretin, a thyroxine transport protein known to be exclusively produced by the choroidal epithelia in the CNS. These cultures have been adapted for use on freely permeable Transwell(®) membranes sandwiched between two culture chambers, facilitating transport studies of various compounds across this barrier in vitro. These choroidal epithelia cultures with the Transwell system will perceivably assist blood-CSF barrier research.

  5. Involvement of PI3K/AKT and MAPK Pathways for TNF-α Production in SiHa Cervical Mucosal Epithelial Cells Infected with Trichomonas vaginalis.

    PubMed

    Yang, Jung-Bo; Quan, Juan-Hua; Kim, Ye-Eun; Rhee, Yun-Ee; Kang, Byung-Hyun; Choi, In-Wook; Cha, Guang-Ho; Yuk, Jae-Min; Lee, Young-Ha

    2015-08-01

    Trichomonas vaginalis; induces proinflammation in cervicovaginal mucosal epithelium. To investigate the signaling pathways in TNF-α production in cervical mucosal epithelium after T. vaginalis infection, the phosphorylation of PI3K/AKT and MAPK pathways were evaluated in T. vaginalis-infected SiHa cells in the presence and absence of specific inhibitors. T. vaginalis increased TNF-α production in SiHa cells, in a parasite burden-dependent and incubation time-dependent manner. In T. vaginalis-infected SiHa cells, AKT, ERK1/2, p38 MAPK, and JNK were phosphorylated from 1 hr after infection; however, the phosphorylation patterns were different from each other. After pretreatment with inhibitors of the PI3K/AKT and MAPK pathways, TNF-α production was significantly decreased compared to the control; however, TNF-α reduction patterns were different depending on the type of PI3K/MAPK inhibitors. TNF-α production was reduced in a dose-dependent manner by treatment with wortmannin and PD98059, whereas it was increased by SP600125. These data suggested that PI3K/AKT and MAPK signaling pathways are important in regulation of TNF-α production in cervical mucosal epithelial SiHa cells. However, activation patterns of each pathway were different from the types of PI3K/MAPK pathways.

  6. Pregnane X Receptor Activation Attenuates Inflammation-Associated Intestinal Epithelial Barrier Dysfunction by Inhibiting Cytokine-Induced Myosin Light-Chain Kinase Expression and c-Jun N-Terminal Kinase 1/2 Activation.

    PubMed

    Garg, Aditya; Zhao, Angela; Erickson, Sarah L; Mukherjee, Subhajit; Lau, Aik Jiang; Alston, Laurie; Chang, Thomas K H; Mani, Sridhar; Hirota, Simon A

    2016-10-01

    The inflammatory bowel diseases (IBDs) are chronic inflammatory disorders with a complex etiology. IBD is thought to arise in genetically susceptible individuals in the context of aberrant interactions with the intestinal microbiota and other environmental risk factors. Recently, the pregnane X receptor (PXR) was identified as a sensor for microbial metabolites, whose activation can regulate the intestinal epithelial barrier. Mutations in NR1I2, the gene that encodes the PXR, have been linked to IBD, and in animal models, PXR deletion leads to barrier dysfunction. In the current study, we sought to assess the mechanism(s) through which the PXR regulates barrier function during inflammation. In Caco-2 intestinal epithelial cell monolayers, tumor necrosis factor-α/interferon-γ exposure disrupted the barrier and triggered zonula occludens-1 relocalization, increased expression of myosin light-chain kinase (MLCK), and activation of c-Jun N-terminal kinase 1/2 (JNK1/2). Activation of the PXR [rifaximin and [[3,5-Bis(1,1-dimethylethyl)-4-hydroxyphenyl]ethenylidene]bis-phosphonic acid tetraethyl ester (SR12813); 10 μM] protected the barrier, an effect that was associated with attenuated MLCK expression and JNK1/2 activation. In vivo, activation of the PXR [pregnenolone 16α-carbonitrile (PCN)] attenuated barrier disruption induced by toll-like receptor 4 activation in wild-type, but not Pxr-/-, mice. Furthermore, PCN treatment protected the barrier in the dextran-sulfate sodium model of experimental colitis, an effect that was associated with reduced expression of mucosal MLCK and phosphorylated JNK1/2. Together, our data suggest that the PXR regulates the intestinal epithelial barrier during inflammation by modulating cytokine-induced MLCK expression and JNK1/2 activation. Thus, targeting the PXR may prove beneficial for the treatment of inflammation-associated barrier disruption in the context of IBD. PMID:27440420

  7. Pregnane X Receptor Activation Attenuates Inflammation-Associated Intestinal Epithelial Barrier Dysfunction by Inhibiting Cytokine-Induced Myosin Light-Chain Kinase Expression and c-Jun N-Terminal Kinase 1/2 Activation.

    PubMed

    Garg, Aditya; Zhao, Angela; Erickson, Sarah L; Mukherjee, Subhajit; Lau, Aik Jiang; Alston, Laurie; Chang, Thomas K H; Mani, Sridhar; Hirota, Simon A

    2016-10-01

    The inflammatory bowel diseases (IBDs) are chronic inflammatory disorders with a complex etiology. IBD is thought to arise in genetically susceptible individuals in the context of aberrant interactions with the intestinal microbiota and other environmental risk factors. Recently, the pregnane X receptor (PXR) was identified as a sensor for microbial metabolites, whose activation can regulate the intestinal epithelial barrier. Mutations in NR1I2, the gene that encodes the PXR, have been linked to IBD, and in animal models, PXR deletion leads to barrier dysfunction. In the current study, we sought to assess the mechanism(s) through which the PXR regulates barrier function during inflammation. In Caco-2 intestinal epithelial cell monolayers, tumor necrosis factor-α/interferon-γ exposure disrupted the barrier and triggered zonula occludens-1 relocalization, increased expression of myosin light-chain kinase (MLCK), and activation of c-Jun N-terminal kinase 1/2 (JNK1/2). Activation of the PXR [rifaximin and [[3,5-Bis(1,1-dimethylethyl)-4-hydroxyphenyl]ethenylidene]bis-phosphonic acid tetraethyl ester (SR12813); 10 μM] protected the barrier, an effect that was associated with attenuated MLCK expression and JNK1/2 activation. In vivo, activation of the PXR [pregnenolone 16α-carbonitrile (PCN)] attenuated barrier disruption induced by toll-like receptor 4 activation in wild-type, but not Pxr-/-, mice. Furthermore, PCN treatment protected the barrier in the dextran-sulfate sodium model of experimental colitis, an effect that was associated with reduced expression of mucosal MLCK and phosphorylated JNK1/2. Together, our data suggest that the PXR regulates the intestinal epithelial barrier during inflammation by modulating cytokine-induced MLCK expression and JNK1/2 activation. Thus, targeting the PXR may prove beneficial for the treatment of inflammation-associated barrier disruption in the context of IBD.

  8. Modulation of bronchial epithelial cell barrier function by in vitro ozone exposure

    SciTech Connect

    Yu, Xiao-Ying; Takahashi, Nobuyoshi; Croxton, T.L.; Spannhake, E.W.

    1994-12-01

    The epithelial cells lining the small, peripheral airways function as important targets for the action of inspired ozone. Loss of epithelial barrier integrity in these regions is a common element in ozone-induced airway inflammation. To investigate the direct effect of ozone on epithelial barrier function, canine bronchial epithelial (CBE) cells grown with an air interface were exposed for 3hr to 0.2, 0.5, or 0.8 ppm ozone or to air. Mannitol flux, used as an index of paracellular permeability, increased above air controls by 461%, 774%, and 1172% at the three ozone concentrations, respectively. Transcellular electrical resistance exhibited a dose-related decrease. The immediate effect of 0.8 ppm ozone on permeability was significantly inhibited by preincubation for 48 hr in the presence of 1 ng/ml vitamin E (33%) or 1 {mu}M vitamin A (34%). Responses to 0.5 ppm or 0.8 ppm were inhibited by pretreatment of the cells with 0.1 {mu}M of the actin polymerizing agent phalloidin (34% and 25% inhibition, respectively). The increases in permeability induced by 0.2 and 0.5 ppm ozone were attenuated by 54% and 22%, respectively, at 18 hr after exposure, whereas that to 0.8 ppm was further enhanced by 42% at this time. The effects of ozone are modulated by the availability of antioxidants to the cells and appear to be associated with cytoskeletal dysfunction in CBE cells. The data are consistent with a loss of barrier function linked to a direct oxidative effect of ozone on individual CBE cells and indicate that the reversible or progressive nature of this effect is dose dependent. 31 refs., 5 figs.

  9. Modulation of bronchial epithelial cell barrier function by in vitro jet propulsion fuel 8 exposure.

    PubMed

    Robledo, R F; Barber, D S; Witten, M L

    1999-09-01

    The loss of epithelial barrier integrity in bronchial and bronchiolar airways may be an initiating factor in the observed onset of toxicant-induced lung injuries. Acute 1-h inhalation exposures to aerosolized jet propulsion fuel 8 (JP-8) have been shown to induce cellular and morphological indications of pulmonary toxicity that was associated with increased respiratory permeability to 99mTc-DTPA. To address the hypothesis that JP-8 jet fuel-induced lung injury is initiated through a disruption in the airway epithelial barrier function, paracellular mannitol flux of BEAS-2B human bronchial epithelial cells was measured. Incubation of confluent cell cultures with non-cytotoxic concentrations of JP-8 or n-tetradecane (C14), a primary constituent of JP-8, for a 1-h exposure period resulted in dose-dependent increases of paracellular flux. Following exposures of 0.17, 0.33, 0.50, or 0.67 mg/ml, mannitol flux increased above vehicle controls by 10, 14, 29, and 52%, respectively, during a 2-h incubation period immediately after each JP-8 exposure. C14 caused greater mannitol flux increases of 37, 42, 63, and 78%, respectively, following identical exposure conditions. The effect on transepithelial mannitol flux reached a maximum at 12 h and spontaneously reversed to control values over a 48-h recovery period, for both JP-8 and C14 exposure. These data indicate that non-cytotoxic exposures to JP-8 or C14 exert a noxious effect on bronchial epithelial barrier function that may preclude pathological lung injury.

  10. Bifidobacteria Prevent Tunicamycin-Induced Endoplasmic Reticulum Stress and Subsequent Barrier Disruption in Human Intestinal Epithelial Caco-2 Monolayers.

    PubMed

    Akiyama, Takuya; Oishi, Kenji; Wullaert, Andy

    2016-01-01

    Endoplasmic reticulum (ER) stress is caused by accumulation of unfolded and misfolded proteins in the ER, thereby compromising its vital cellular functions in protein production and secretion. Genome wide association studies in humans as well as experimental animal models linked ER stress in intestinal epithelial cells (IECs) with intestinal disorders including inflammatory bowel diseases. However, the mechanisms linking the outcomes of ER stress in IECs to intestinal disease have not been clarified. In this study, we investigated the impact of ER stress on intestinal epithelial barrier function using human colon carcinoma-derived Caco-2 monolayers. Tunicamycin-induced ER stress decreased the trans-epithelial electrical resistance of Caco-2 monolayers, concomitant with loss of cellular plasma membrane integrity. Epithelial barrier disruption in Caco-2 cells after ER stress was not caused by caspase- or RIPK1-dependent cell death but was accompanied by lysosomal rupture and up-regulation of the ER stress markers Grp78, sXBP1 and Chop. Interestingly, several bifidobacteria species inhibited tunicamycin-induced ER stress and thereby diminished barrier disruption in Caco-2 monolayers. Together, these results showed that ER stress compromises the epithelial barrier function of Caco-2 monolayers and demonstrate beneficial impacts of bifidobacteria on ER stress in IECs. Our results identify epithelial barrier loss as a potential link between ER stress and intestinal disease development, and suggest that bifidobacteria could exert beneficial effects on this phenomenon. PMID:27611782

  11. Bifidobacteria Prevent Tunicamycin-Induced Endoplasmic Reticulum Stress and Subsequent Barrier Disruption in Human Intestinal Epithelial Caco-2 Monolayers

    PubMed Central

    Akiyama, Takuya; Oishi, Kenji

    2016-01-01

    Endoplasmic reticulum (ER) stress is caused by accumulation of unfolded and misfolded proteins in the ER, thereby compromising its vital cellular functions in protein production and secretion. Genome wide association studies in humans as well as experimental animal models linked ER stress in intestinal epithelial cells (IECs) with intestinal disorders including inflammatory bowel diseases. However, the mechanisms linking the outcomes of ER stress in IECs to intestinal disease have not been clarified. In this study, we investigated the impact of ER stress on intestinal epithelial barrier function using human colon carcinoma-derived Caco-2 monolayers. Tunicamycin-induced ER stress decreased the trans-epithelial electrical resistance of Caco-2 monolayers, concomitant with loss of cellular plasma membrane integrity. Epithelial barrier disruption in Caco-2 cells after ER stress was not caused by caspase- or RIPK1-dependent cell death but was accompanied by lysosomal rupture and up-regulation of the ER stress markers Grp78, sXBP1 and Chop. Interestingly, several bifidobacteria species inhibited tunicamycin-induced ER stress and thereby diminished barrier disruption in Caco-2 monolayers. Together, these results showed that ER stress compromises the epithelial barrier function of Caco-2 monolayers and demonstrate beneficial impacts of bifidobacteria on ER stress in IECs. Our results identify epithelial barrier loss as a potential link between ER stress and intestinal disease development, and suggest that bifidobacteria could exert beneficial effects on this phenomenon. PMID:27611782

  12. β-1,3/1,6-Glucan alleviated intestinal mucosal barrier impairment of broiler chickens challenged with Salmonella enterica serovar Typhimurium.

    PubMed

    Shao, Yujing; Guo, Yuming; Wang, Zhong

    2013-07-01

    This study investigated the protective effect of β-1,3/1,6-glucan on gut morphology, intestinal epithelial tight junctions, and bacterial translocation of broiler chickens challenged with Salmonella enterica serovar Typhimurium. Ninety Salmonella-free Arbor Acre male broiler chickens were randomly divided into 3 groups: negative control group (NC), Salmonella Typhimurium-infected positive group (PC), and the Salmonella Typhimurium-infected group with dietary 100 mg/kg of β-1,3/1,6-glucan supplementation (T) to determine the effect of β-1,3/1,6-glucan on intestinal barrier function. Salmonella Typhimurium challenge alone significantly decreased villus height (P < 0.001), villus height/crypt depth ratio (P < 0.05), and the number of goblet cells (P < 0.001) in the jejunum at 14 d postinfection (dpi), but significantly increased the number of intestinal secretory IgA (sIgA)-expressing cells at 14 dpi (P < 0.01) and total sIgA levels in the jejunum at 7 (P < 0.05) and 14 dpi (P < 0.01) compared with the unchallenged birds (NC). Dietary β-1,3/1,6-glucan supplementation not only significantly increased villus height, villus height/crypt depth ratio, and the number of goblet cells (P < 0.01), but also increased the number of sIgA-expressing cells (P < 0.05) and sIgA content in the jejunum at 14 dpi (P < 0.01) in birds challenged with Salmonella Typhimurium in comparison with Salmonella Typhimurium challenge alone. β-1,3/1,6-Glucan addition had significant inhibitory effects (P < 0.05) on cecal Salmonella colonization levels and liver Salmonella invasion of the Salmonella Typhimurium-infected birds compared with the PC group. Intestinal tight junction proteins claudin-1, claudin-4, and occludin mRNA expression in the jejunum at 14 dpi was significantly decreased by Salmonella Typhimurium challenge alone (P < 0.01) compared with that of the NC group, whereas β-1,3/1,6-glucan supplementation significantly increased claudin-1 and occludin mRNA expression (P < 0.01) at

  13. Establishment of a novel in vitro model of stratified epithelial wound healing with barrier function

    PubMed Central

    Gonzalez-Andrades, Miguel; Alonso-Pastor, Luis; Mauris, Jérôme; Cruzat, Andrea; Dohlman, Claes H.; Argüeso, Pablo

    2016-01-01

    The repair of wounds through collective movement of epithelial cells is a fundamental process in multicellular organisms. In stratified epithelia such as the cornea and skin, healing occurs in three steps that include a latent, migratory, and reconstruction phases. Several simple and inexpensive assays have been developed to study the biology of cell migration in vitro. However, these assays are mostly based on monolayer systems that fail to reproduce the differentiation processes associated to multilayered systems. Here, we describe a straightforward in vitro wound assay to evaluate the healing and restoration of barrier function in stratified human corneal epithelial cells. In this assay, circular punch injuries lead to the collective migration of the epithelium as coherent sheets. The closure of the wound was associated with the restoration of the transcellular barrier and the re-establishment of apical intercellular junctions. Altogether, this new model of wound healing provides an important research tool to study the mechanisms leading to barrier function in stratified epithelia and may facilitate the development of future therapeutic applications. PMID:26759072

  14. Autophagy enhances intestinal epithelial tight junction barrier function by targeting claudin-2 protein degradation.

    PubMed

    Nighot, Prashant K; Hu, Chien-An Andy; Ma, Thomas Y

    2015-03-13

    Autophagy is an intracellular degradation pathway and is considered to be an essential cell survival mechanism. Defects in autophagy are implicated in many pathological processes, including inflammatory bowel disease. Among the innate defense mechanisms of intestinal mucosa, a defective tight junction (TJ) barrier has been postulated as a key pathogenic factor in the causation and progression of inflammatory bowel disease by allowing increased antigenic permeation. The cross-talk between autophagy and the TJ barrier has not yet been described. In this study, we present the novel finding that autophagy enhances TJ barrier function in Caco-2 intestinal epithelial cells. Nutrient starvation-induced autophagy significantly increased transepithelial electrical resistance and reduced the ratio of sodium/chloride paracellular permeability. Nutrient starvation reduced the paracellular permeability of small-sized urea but not larger molecules. The role of autophagy in the modulation of paracellular permeability was confirmed by pharmacological induction as well as pharmacological and genetic inhibition of autophagy. Consistent with the autophagy-induced reduction in paracellular permeability, a marked decrease in the level of the cation-selective, pore-forming TJ protein claudin-2 was observed after cell starvation. Starvation reduced the membrane presence of claudin-2 and increased its cytoplasmic, lysosomal localization. Therefore, our data show that autophagy selectively reduces epithelial TJ permeability of ions and small molecules by lysosomal degradation of the TJ protein claudin-2.

  15. Autophagy Enhances Intestinal Epithelial Tight Junction Barrier Function by Targeting Claudin-2 Protein Degradation*

    PubMed Central

    Nighot, Prashant K.; Hu, Chien-An Andy; Ma, Thomas Y.

    2015-01-01

    Autophagy is an intracellular degradation pathway and is considered to be an essential cell survival mechanism. Defects in autophagy are implicated in many pathological processes, including inflammatory bowel disease. Among the innate defense mechanisms of intestinal mucosa, a defective tight junction (TJ) barrier has been postulated as a key pathogenic factor in the causation and progression of inflammatory bowel disease by allowing increased antigenic permeation. The cross-talk between autophagy and the TJ barrier has not yet been described. In this study, we present the novel finding that autophagy enhances TJ barrier function in Caco-2 intestinal epithelial cells. Nutrient starvation-induced autophagy significantly increased transepithelial electrical resistance and reduced the ratio of sodium/chloride paracellular permeability. Nutrient starvation reduced the paracellular permeability of small-sized urea but not larger molecules. The role of autophagy in the modulation of paracellular permeability was confirmed by pharmacological induction as well as pharmacological and genetic inhibition of autophagy. Consistent with the autophagy-induced reduction in paracellular permeability, a marked decrease in the level of the cation-selective, pore-forming TJ protein claudin-2 was observed after cell starvation. Starvation reduced the membrane presence of claudin-2 and increased its cytoplasmic, lysosomal localization. Therefore, our data show that autophagy selectively reduces epithelial TJ permeability of ions and small molecules by lysosomal degradation of the TJ protein claudin-2. PMID:25616664

  16. Checkpoint Kinase 1 Activation Enhances Intestinal Epithelial Barrier Function via Regulation of Claudin-5 Expression

    PubMed Central

    Watari, Akihiro; Hasegawa, Maki; Yagi, Kiyohito; Kondoh, Masuo

    2016-01-01

    Several stressors are known to influence epithelial tight junction (TJ) integrity, but the association between DNA damage and TJ integrity remains unclear. Here we examined the effects of daunorubicin and rebeccamycin, two anti-tumor chemicals that induce DNA damage, on TJ integrity in human intestinal epithelial cells. Daunorubicin and rebeccamycin dose-dependently enhanced transepithelial electrical resistance (TER) and decreased flux of the 4 kDa FITC-dextran in Caco-2 cell monolayer. Daunorubicin- or rebeccamycin-induced enhancement of the TJ barrier function partly rescued attenuation of the barrier function by the inflammatory cytokines TNF-α and IFN-γ. Daunorubicin and rebeccamycin increased claudin-5 expression and the product was distributed in the actin cytoskeleton fraction, which was enriched with TJ proteins. Caffeine, which is an inhibitor of ataxia telangiectasia mutated protein (ATM) and ataxia telangiectasia mutated and Rad3-related protein (ATR), and the Chk1 inhibitor inhibited the TER increases induced by daunorubicin and rebeccamycin, whereas a Chk2 inhibitor did not. Treatment with Chk1 siRNA also significantly inhibited the TER increases. Induction of claudin-5 expression was inhibited by Chk1 inhibitor and by siRNA treatment. Our results suggest that Chk1 activation by daunorubicin and rebeccamycin induced claudin-5 expression and enhanced TJ barrier function in Caco-2 cell monolayer, which suggests a link between DNA damage and TJ integrity in the human intestine. PMID:26727128

  17. Moxibustion regulates inflammatory mediators and colonic mucosal barrier in ulcerative colitis rats

    PubMed Central

    Ma, Tie-Ming; Xu, Na; Ma, Xian-De; Bai, Zeng-Hua; Tao, Xing; Yan, Hong-Chi

    2016-01-01

    occludin and ZO-1 in colonic tissue represent a potential mechanism for improved intestinal mucosal tissue repair with grain-sized moxibustion. PMID:26937144

  18. Protective effects of nonionic tri-block copolymers on bile acid-mediated epithelial barrier disruption.

    SciTech Connect

    Edelstein, A.; Fink, D.; Musch, M.; Valuckaite, V.; Zabornia, O.; Grubjesic, S.; Firestone, M. A.; Matthews, J. B.; Alverdy, J. C.

    2011-11-01

    Translocation of bacteria and other luminal factors from the intestine following surgical injury can be a major driver of critical illness. Bile acids have been shown to play a key role in the loss of intestinal epithelial barrier function during states of host stress. Experiments to study the ability of nonionic block copolymers to abrogate barrier failure in response to bile acid exposure are described. In vitro experiments were performed with the bile salt sodium deoxycholate on Caco-2 enterocyte monolayers using transepithelial electrical resistance to assay barrier function. A bisphenol A coupled triblock polyethylene glycol (PEG), PEG 15-20, was shown to prevent sodium deoxycholate-induced barrier failure. Enzyme-linked immunosorbent assay, lactate dehydrogenase, and caspase 3-based cell death detection assays demonstrated that bile acid-induced apoptosis and necrosis were prevented with PEG 15-20. Immunofluorescence microscopic visualization of the tight junctional protein zonula occludens 1 (ZO-1) demonstrated that PEG 15-20 prevented significant changes in tight junction organization induced by bile acid exposure. Preliminary transepithelial electrical resistance-based studies examining structure-function correlates of polymer protection against bile acid damage were performed with a small library of PEG-based copolymers. Polymer properties associated with optimal protection against bile acid-induced barrier disruption were PEG-based compounds with a molecular weight greater than 10 kd and amphiphilicity. The data demonstrate that PEG-based copolymer architecture is an important determinant that confers protection against bile acid injury of intestinal epithelia.

  19. Unique Transcompartmental Bridge: Antigen-Presenting Cells Sampling across Endothelial and Mucosal Barriers.

    PubMed

    Allen, Frederick; Tong, Alexander A; Huang, Alex Y

    2016-01-01

    Potentially harmful pathogens can gain access to tissues and organ systems through body sites that are in direct contact with the outside environment, such as the skin, the gut, and the airway mucosa. Antigen-presenting cells (APCs) represent a bridge between the innate and adaptive immunity, and their capacity for constant immune surveillance and rapid sampling of incoming pathogens and other potentially harmful antigens is central for mounting an effective and robust protective host response. The classical view is that APCs perform this task efficiently within the tissue to sense invading agents intra-compartmentally. However, recent data based on high resolution imaging support an additional transcompartmental surveillance behavior by APC by reaching across intact physical barriers. In this review, we summarize intravital microscopic evidences of APC to sample antigens transcompartmentally at the gut mucosa and other body sites. PMID:27375624

  20. Unique Transcompartmental Bridge: Antigen-Presenting Cells Sampling across Endothelial and Mucosal Barriers

    PubMed Central

    Allen, Frederick; Tong, Alexander A.; Huang, Alex Y.

    2016-01-01

    Potentially harmful pathogens can gain access to tissues and organ systems through body sites that are in direct contact with the outside environment, such as the skin, the gut, and the airway mucosa. Antigen-presenting cells (APCs) represent a bridge between the innate and adaptive immunity, and their capacity for constant immune surveillance and rapid sampling of incoming pathogens and other potentially harmful antigens is central for mounting an effective and robust protective host response. The classical view is that APCs perform this task efficiently within the tissue to sense invading agents intra-compartmentally. However, recent data based on high resolution imaging support an additional transcompartmental surveillance behavior by APC by reaching across intact physical barriers. In this review, we summarize intravital microscopic evidences of APC to sample antigens transcompartmentally at the gut mucosa and other body sites. PMID:27375624

  1. Protective Effect of Huoxiang Zhengqi Oral Liquid on Intestinal Mucosal Mechanical Barrier of Rats with Postinfectious Irritable Bowel Syndrome Induced by Acetic Acid

    PubMed Central

    Liu, Yao; Liu, Wei; Peng, Qiu-Xian; Peng, Jiang-Li; Yu, Lin-Zhong; Hu, Jian-Lan

    2014-01-01

    In this study, a rat model with acetic acid-induced PI-IBS was used to study the role of HXZQ oral liquid in repairing the colonic epithelial barrier and reducing intestinal permeability. Pathomorphism of colonic tissue, epithelial ultrastructure, DAO activity in serum, and the protein expression of ZO-1 and occludin were examined to investigate protective effect mechanisms of HXZQ on intestinal mucosa barrier and then present experimental support for its use for prevention and cure of PI-IBS. PMID:25254052

  2. Severity of pancreatitis-associated intestinal mucosal barrier injury is reduced following treatment with the NADPH oxidase inhibitor apocynin

    PubMed Central

    Deng, Wenhong; Abliz, Ablikim; Xu, Sheng; Sun, Rongze; Guo, Wenyi; Shi, Qiao; Yu, Jia; Wang, Weixing

    2016-01-01

    Recent studies demonstrated that apocynin, a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) inhibitor, significantly decreased acute pancreatitis-associated inflammatory and oxidative stress parameters. In addition, apocynin was able to reduce ischemic reperfusion injury-associated damage; however, the exact effects of apocynin on acute pancreatitis-associated intestinal mucosal injury have yet to be fully clarified. The present study aimed to investigate the protective effects of apocynin on intestinal mucosal injury in a rat model of severe acute pancreatitis (SAP). A total of 60 male Sprague Dawley rats were randomly divided into four groups (n=15/group): Sham operation group (SO), SAP group, apocynin treatment (APO) group and drug control (APO-CON) group. SAP was induced by retrograde injection of 5% sodium taurocholate into the biliopancreatic duct. Apocynin was administered 30 min prior to SAP induction in the APO group. All rats were sacrificed 12 h after SAP induction. Intestinal integrity was assessed by measuring diamine oxidase (DAO) levels. Morphological alterations to intestinal tissue were determined under light and transmission electron microscopy. NOX2, p38 mitogen-activated protein kinases (MAPK) and nuclear factor (NF)-κB expression levels were detected in the intestine by immunohistochemical staining. Oxidative stress was detected by measuring intestinal malondialdehyde (MDA) and superoxide dismutase content. In addition, blood inflammatory cytokines, and amylase (AMY) and lipase (LIP) levels were evaluated. The results demonstrated that apocynin attenuated the following: i) Serum AMY, LIP and DAO levels; ii) pancreatic and intestinal pathological injury; iii) intestinal MDA content; iv) intestinal ultrastructural alterations; v) serum interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α levels; and vi) NOX2, p38 MAPK and NF-κB expression in intestinal tissues. These results suggested that apocynin may attenuate

  3. Severity of pancreatitis‑associated intestinal mucosal barrier injury is reduced following treatment with the NADPH oxidase inhibitor apocynin.

    PubMed

    Deng, Wenhong; Abliz, Ablikim; Xu, Sheng; Sun, Rongze; Guo, Wenyi; Shi, Qiao; Yu, Jia; Wang, Weixing

    2016-10-01

    Recent studies demonstrated that apocynin, a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) inhibitor, significantly decreased acute pancreatitis‑associated inflammatory and oxidative stress parameters. In addition, apocynin was able to reduce ischemic reperfusion injury‑associated damage; however, the exact effects of apocynin on acute pancreatitis‑associated intestinal mucosal injury have yet to be fully clarified. The present study aimed to investigate the protective effects of apocynin on intestinal mucosal injury in a rat model of severe acute pancreatitis (SAP). A total of 60 male Sprague Dawley rats were randomly divided into four groups (n=15/group): Sham operation group (SO), SAP group, apocynin treatment (APO) group and drug control (APO‑CON) group. SAP was induced by retrograde injection of 5% sodium taurocholate into the biliopancreatic duct. Apocynin was administered 30 min prior to SAP induction in the APO group. All rats were sacrificed 12 h after SAP induction. Intestinal integrity was assessed by measuring diamine oxidase (DAO) levels. Morphological alterations to intestinal tissue were determined under light and transmission electron microscopy. NOX2, p38 mitogen‑activated protein kinases (MAPK) and nuclear factor (NF)‑κB expression levels were detected in the intestine by immunohistochemical staining. Oxidative stress was detected by measuring intestinal malondialdehyde (MDA) and superoxide dismutase content. In addition, blood inflammatory cytokines, and amylase (AMY) and lipase (LIP) levels were evaluated. The results demonstrated that apocynin attenuated the following: i) Serum AMY, LIP and DAO levels; ii) pancreatic and intestinal pathological injury; iii) intestinal MDA content; iv) intestinal ultrastructural alterations; v) serum interleukin (IL)‑1β, IL‑6 and tumor necrosis factor (TNF)‑α levels; and vi) NOX2, p38 MAPK and NF‑κB expression in intestinal tissues. These results suggested

  4. Why mucosal health?

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aquaculture species depend more heavily on mucosal barriers than their terrestrial agricultural counterparts as they are continuously interacting with the aquatic microbiota. Unlike classical immune centers, such as the spleen and kidney, the accessibility of mucosal surfaces through immersion/dip t...

  5. The relative balance of GM-CSF and TGF-β1 regulates lung epithelial barrier function.

    PubMed

    Overgaard, Christian E; Schlingmann, Barbara; Dorsainvil White, StevenClaude; Ward, Christina; Fan, Xian; Swarnakar, Snehasikta; Brown, Lou Ann S; Guidot, David M; Koval, Michael

    2015-06-15

    Lung barrier dysfunction is a cardinal feature of the acute respiratory distress syndrome (ARDS). Alcohol abuse, which increases the risk of ARDS two- to fourfold, induces transforming growth factor (TGF)-β1, which increases epithelial permeability and impairs granulocyte/macrophage colony-stimulating factor (GM-CSF)-dependent barrier integrity in experimental models. We hypothesized that the relative balance of GM-CSF and TGF-β1 signaling regulates lung epithelial barrier function. GM-CSF and TGF-β1 were tested separately and simultaneously for their effects on lung epithelial cell barrier function in vitro. TGF-β1 alone caused an ∼ 25% decrease in transepithelial resistance (TER), increased paracellular flux, and was associated with projections perpendicular to tight junctions ("spikes") containing claudin-18 that colocalized with F-actin. In contrast, GM-CSF treatment induced an ∼ 20% increase in TER, decreased paracellular flux, and showed decreased colocalization of spike-associated claudin-18 with F-actin. When simultaneously administered to lung epithelial cells, GM-CSF antagonized the effects of TGF-β1 on epithelial barrier function in cultured cells. Given this, GM-CSF and TGF-β1 levels were measured in bronchoalveolar lavage (BAL) fluid from patients with ventilator-associated pneumonia and correlated with markers for pulmonary edema and patient outcome. In patient BAL fluid, protein markers of lung barrier dysfunction, serum α2-macroglobulin, and IgM levels were increased at lower ratios of GM-CSF/TGF-β1. Critically, patients who survived had significantly higher GM-CSF/TGF-β1 ratios than nonsurviving patients. This study provides experimental and clinical evidence that the relative balance between GM-CSF and TGF-β1 signaling is a key regulator of lung epithelial barrier function. The GM-CSF/TGF-β1 ratio in BAL fluid may provide a concentration-independent biomarker that can predict patient outcomes in ARDS.

  6. Breakdown of Epithelial Barrier Integrity and Overdrive Activation of Alveolar Epithelial Cells in the Pathogenesis of Acute Respiratory Distress Syndrome and Lung Fibrosis

    PubMed Central

    Yanagi, Shigehisa; Tsubouchi, Hironobu; Miura, Ayako; Matsumoto, Nobuhiro; Nakazato, Masamitsu

    2015-01-01

    Individual alveolar epithelial cells (AECs) collaboratively form a tight barrier between atmosphere and fluid-filled tissue to enable normal gas exchange. The tight junctions of AECs provide intercellular sealing and are integral to the maintenance of the AEC barrier integrity. Disruption and failure of reconstitution of AEC barrier result in catastrophic consequences, leading to alveolar flooding and subsequent devastating fibrotic scarring. Recent evidences reveal that many of the fibrotic lung diseases involve AECs both as a frequent target of injury and as a driver of ongoing pathological processes. Aberrantly activated AECs express most of the growth factors and chemokines responsible for the proliferation, migration, and activation of fibroblasts. Current evidences suggest that AECs may acquire overdrive activation in the initial step of fibrosis by several mechanisms, including abnormal recapitulation of the developmental pathway, defects of the molecules essential for epithelial integrity, and acceleration of aging-related properties. Among these initial triggering events, epithelial Pten, a multiple phosphatase that negatively regulates the PI3K/Akt pathway and is crucial for lung development, is essential for the prevention of alveolar flooding and lung fibrosis through the regulation of AEC barrier integrity after injury. Reestablishment of AEC barrier integrity also involves the deployment of specialized stem/progenitor cells. PMID:26523279

  7. Development of an in vitro model of human bronchial epithelial barrier to study nanoparticle translocation.

    PubMed

    George, Isabelle; Vranic, Sandra; Boland, Sonja; Courtois, Arnaud; Baeza-Squiban, Armelle

    2015-02-01

    Inhalation is the most frequent route of unintentional exposure to nanoparticles (NPs). Our aim was to compare different in vitro models of human lung epithelial monolayers for their suitability to assess the translocation of 50 nm fluorescently labelled silica NPs (50 nm-SiO(2)-FITC-NPs). Human bronchial epithelial cell lines NCI-H292 and Calu-3 as well as human alveolar cell line A549 were seeded onto Transwell filters (TF) separating the well into an apical and a basal compartment. Measurements of the transepithelial electric resistance and monitoring the paracellular transport of a fluorescent marker (Lucifer Yellow) have shown that only Calu-3 cells formed a tight epithelium. In the absence of cells 4% of the initially applied NP concentration was found to cross the TF but the majority remained trapped inside the filter. After 24 h of treatment, 50 nm-SiO(2)-FITC-NPs were taken up by all cell types but their translocation was inversely correlated to the efficiency to prevent LY passage: translocation represented 3% of the initially apically applied NP concentration for Calu-3 cells, 9% for NCI-H292 cells and 35% for A549 cells. In conclusion, 50 nm-SiO(2)-FITC-NPs can cross different bronchial epithelial barriers, but the Calu-3 cell line appears to be the most relevant model for studying NP translocation. PMID:25197033

  8. Innate Immunity and the Role of Epithelial Barrier During Aspergillus fumigatus Infection

    PubMed Central

    Svirshchevskaya, Elena; Zubkov, Dmitrii; Mouyna, Isabelle; Berkova, Nadia

    2012-01-01

    Fungi are the most important eukaryotic infective agents in Europe which largely overpass parasite infections. Total number of people dying of fungal infection is increasing and this trend is likely to continue due to the increase in immunosuppressive treatments. The opportunistic pathogen Aspergillus fumigatus (Af) is a saprophytic filamentous fungus that can cause invasive pulmonary diseases in immuno-compromised hosts. In veterinary medicine aspergillosis is also a recurrent problem since it infects various species, birds are particularly susceptible. It propagates through airborne conidia (spores), which are inhaled into the small airways where they may germinate and initiate an infection. The host epithelium has permanent contact with the environment and a multitude of diverse microorganisms, resulting in a network of the host’s defense mechanisms. Pathogens use various strategies to invade epithelial barriers, to exploit eukaryotic host function to their own benefit and disseminate throughout the host using the epithelium as a reservoir. The current revue will discuss the ways how epithelial and innate immunity cells can contlol Af infection. We will focus on Af strategies for the host’s invasion, antifungal innate immune response and antimicrobial activities of the respiratory epithelial cells. PMID:23255875

  9. Crossing of the epithelial barriers by Bacillus anthracis: the Known and the Unknown

    PubMed Central

    Goossens, Pierre L.; Tournier, Jean-Nicolas

    2015-01-01

    Anthrax, caused by Bacillus anthracis, a Gram-positive spore-forming bacterium, is initiated by the entry of spores into the host body. There are three types of human infection: cutaneous, inhalational, and gastrointestinal. For each form, B. anthracis spores need to cross the cutaneous, respiratory or digestive epithelial barriers, respectively, as a first obligate step to establish infection. Anthrax is a toxi-infection: an association of toxemia and rapidly spreading infection progressing to septicemia. The pathogenicity of Bacillus anthracis mainly depends on two toxins and a capsule. The capsule protects bacilli from the immune system, thus promoting systemic dissemination. The toxins alter host cell signaling, thereby paralyzing the immune response of the host and perturbing the endocrine and endothelial systems. In this review, we will mainly focus on the events and mechanisms leading to crossing of the respiratory epithelial barrier, as the majority of studies have addressed inhalational infection. We will discuss the critical gaps of knowledge that need to be addressed to gain a comprehensive view of the initial steps of inhalational anthrax. We will then discuss the few data available on B. anthracis crossing the cutaneous and digestive epithelia. PMID:26500645

  10. Junctional Adhesion Molecule A Promotes Epithelial Tight Junction Assembly to Augment Lung Barrier Function

    PubMed Central

    Mitchell, Leslie A.; Ward, Christina; Kwon, Mike; Mitchell, Patrick O.; Quintero, David A.; Nusrat, Asma; Parkos, Charles A.; Koval, Michael

    2016-01-01

    Epithelial barrier function is maintained by tight junction proteins that control paracellular fluid flux. Among these proteins is junctional adhesion molecule A (JAM-A), an Ig fold transmembrane protein. To assess JAM-A function in the lung, we depleted JAM-A in primary alveolar epithelial cells using shRNA. In cultured cells, loss of JAM-A caused an approximately 30% decrease in transepithelial resistance, decreased expression of the tight junction scaffold protein zonula occludens 1, and disrupted junctional localization of the structural transmembrane protein claudin-18. Consistent with findings in other organs, loss of JAM-A decreased β1 integrin expression and impaired filamentous actin formation. Using a model of mild systemic endoxotemia induced by i.p. injection of lipopolysaccharide, we report that JAM-A−/− mice showed increased susceptibility to pulmonary edema. On injury, the enhanced susceptibility of JAM-A−/− mice to edema correlated with increased, transient disruption of claudin-18, zonula occludens 1, and zonula occludens 2 localization to lung tight junctions in situ along with a delay in up-regulation of claudin-4. In contrast, wild-type mice showed no change in lung tight junction morphologic features in response to mild systemic endotoxemia. These findings support a key role of JAM-A in promoting tight junction homeostasis and lung barrier function by coordinating interactions among claudins, the tight junction scaffold, and the cytoskeleton. PMID:25438062

  11. TNFα/IFNγ Mediated Intestinal Epithelial Barrier Dysfunction Is Attenuated by MicroRNA-93 Downregulation of PTK6 in Mouse Colonic Epithelial Cells

    PubMed Central

    Beard, Richard S.; Eitner, Rebecca A.; Chen, Liwei; Wu, Mack H.

    2016-01-01

    Since inflammatory bowel diseases (IBD) represent significant morbidity and mortality in the US, the need for defining novel drug targets and inflammatory mechanisms would be of considerable benefit. Although protein tyrosine kinase 6 (PTK6, also known as breast tumor kinase BRK) has been primarily studied in an oncogenic context, it was noted that PTK6 null mice exhibited significantly enhanced colonic epithelial barrier function. Considering that the inflammatory functions of PTK6 have not yet been explored, we hypothesized that cytokines responsible for mediating IBD, such as TNFα/IFNγ, may solicit the action of PTK6 to alter barrier function. After first assessing critical mediators of TNFα/IFNγ driven epithelial barrier dysfunction, we further explored the possibility of PTK6 in this inflammatory context. In this report, we showed that PTK6 siRNA and PTK6 null young adult mouse colonic epithelial cells (YAMC) exhibited significant attenuation of TNFα/IFNγ induced barrier dysfunction as measured by electric cell-substrate impedance sensing (ECIS) assay and permeability assays. In addition, PTK6 null cells transfected with PTK6 cDNA displayed restored barrier dysfunction in response to TNFα/IFNγ, while the cells transfected with vector alone showed similar attenuation of barrier dysfunction. Furthermore, using subcellular fractionation and immunocytochemistry experiments, we found that PTK6 plays a role in FoxO1 nuclear accumulation leading to down-regulation of claudin-3, a tight junction protein. Moreover, we searched for relevant miRNA candidates putative for targeting PTK6 in order to identify and assess the impact of microRNA that target PTK6 with respect to TNFα/IFNγ induced barrier dysfunction. Subsequently, we assayed likely targets and determined their effectiveness in attenuating PTK6 expression as well as cytokine induced barrier dysfunction. Results showed that miR-93 reduced PTK6 expression and attenuated TNFα/IFNγ imposed decrease in

  12. Protective effect and mechanism of hydrogen treatment on lung epithelial barrier dysfunction in rats with sepsis.

    PubMed

    Liu, L-D; Wu, X-Y; Tao, B-D; Wang, N; Zhang, J

    2016-01-01

    This study aimed to explore the protective effect of hydrogen and to investigate the underlying mechanism of its preliminary effect on the alveolar epithelial barrier function in septic rats. Forty-five male Sprague-Dawley rats were divided randomly into three groups (N = 15): control [saline injection (intraperitoneal, ip), air drawing; SA], acute lung injury group [lipopolysaccharide (LPS) injection (ip, 15 mg/kg), air drawing; LA], and acute lung injury combined with hydrogen drawing group [LPS injection (ip, 15 mg/kg), 2% hydrogen drawing; LH]. The rats were euthanized after 6 h of treatment, and the extravascular lung water (EVLW), pulmonary alveolar-arterial oxygen pressure (A-aDO2), and respiratory index (RI) of each group were measured. The aquaporin-1 (AQP-1) protein expression in the lung tissues was detected using immunohistochemistry and western blotting, and the correlation between the EVLW and AQP-1 was analyzed. The lung morphology was observed with light and electron microscopy. In the LA group, EVLW (0.87 ± 0.17), A-aDO2 (113.21 ± 13.92), RI (0.65 ± 0.26), and AQP-1 expression increased. Additionally, thickened alveolar walls, significant invasion of inflammatory cells around the vessels, capillary ectasia, hyperemia/hemorrhage in the alveolar space, significantly swollen mitochondria, and increased vacuolar degeneration were observed. A significant negative correlation between AQP-1 expression and EVLW was observed (R2 = 0.8806). Compared with the LA group, EVLW (0.71 ± 0.19), A-aDO2 (132.42 ± 17.39), RI (0.75 ± 0.24), and inflammatory reaction decreased and AQP-1 expression increased in the LH group. The damage to pulmonary epithelial cells improved after hydrogen treatment in rats with sepsis; hydrogen could protect the pulmonary epithelial barrier function by acting on AQP-1. PMID:26909920

  13. Roles of cellular polyamines in mucosal healing in the gastrointestinal tract.

    PubMed

    Gao, J-H; Guo, L-J; Huang, Z-Y; Rao, J N; Tang, C-W

    2013-12-01

    The mammalian gastrointestinal (GI) mucosa is a rapidly self-renewing tissue in the body, and its integrity is preserved through the strict regulation of epithelial cell proliferation, growth arrest, and apoptosis. Polyamines are shown to play an important role in the regulation of gastrointestinal mucosal growth and healing after injury under physiological and various pathological conditions. In this review, we highlight the importance of cellular polyamines in the control GI mucosal proliferation, migration, apoptosis, angiogenesis and GI barrier function during mucosal repair after injury.

  14. Interferon-gamma increased epithelial barrier function via upregulating claudin-7 expression in human submandibular gland duct epithelium.

    PubMed

    Abe, Ayumi; Takano, Kenichi; Kojima, Takashi; Nomura, Kazuaki; Kakuki, Takuya; Kaneko, Yakuto; Yamamoto, Motohisa; Takahashi, Hiroki; Himi, Tetsuo

    2016-06-01

    Tight junctions (TJs) are necessary for salivary gland function and may serve as indicators of salivary gland epithelial dysfunction. IgG4-related disease (IgG4-RD) is a newly recognized fibro-inflammatory condition which disrupts the TJ associated epithelial barrier. The salivary glands are one of the most frequently involved organs in IgG4-RD, however, changes of the TJ associated epithelial barrier in salivary gland duct epithelium is poorly understood. Here, we investigated the regulation and function of TJs in human submandibular gland ductal epithelial cells (HSDECs) in normal and IgG4-RD. We examined submandibular gland (SMG) tissue from eight control individuals and 22 patients with IgG4-RD and established an HSDEC culture system. Immunohistochemistry, immunocytochemistry, western blotting, and measurement of transepithelial electrical resistance (TER) were performed. Claudin-4, claudin-7, occludin, and JAM-A were expressed at the apical side of the duct epithelium in submandibular gland (SMG) tissue and at the cell borders in HSDECs of normal and IgG4-RD. The expression and distribution of TJs in SMG tissue were not different in control individuals and patients with IgG4-RD in vivo and in vitro. Although interferon-gamma (IFNγ) generally disrupts the integrity and function of TJs, as manifested by decreased epithelial barrier function, IFNγ markedly increased the epithelial barrier function of HSDECs via upregulation of claudin-7 expression in HSDECs from patients with IgG4-RD. This is the first report showing an IFNγ-dependent increase in epithelial barrier function in the salivary gland duct epithelium. Our results provide insights into the functional significance of TJs in salivary gland duct epithelium in physiological and pathological conditions, including IgG4-RD.

  15. Interferon-gamma increased epithelial barrier function via upregulating claudin-7 expression in human submandibular gland duct epithelium.

    PubMed

    Abe, Ayumi; Takano, Kenichi; Kojima, Takashi; Nomura, Kazuaki; Kakuki, Takuya; Kaneko, Yakuto; Yamamoto, Motohisa; Takahashi, Hiroki; Himi, Tetsuo

    2016-06-01

    Tight junctions (TJs) are necessary for salivary gland function and may serve as indicators of salivary gland epithelial dysfunction. IgG4-related disease (IgG4-RD) is a newly recognized fibro-inflammatory condition which disrupts the TJ associated epithelial barrier. The salivary glands are one of the most frequently involved organs in IgG4-RD, however, changes of the TJ associated epithelial barrier in salivary gland duct epithelium is poorly understood. Here, we investigated the regulation and function of TJs in human submandibular gland ductal epithelial cells (HSDECs) in normal and IgG4-RD. We examined submandibular gland (SMG) tissue from eight control individuals and 22 patients with IgG4-RD and established an HSDEC culture system. Immunohistochemistry, immunocytochemistry, western blotting, and measurement of transepithelial electrical resistance (TER) were performed. Claudin-4, claudin-7, occludin, and JAM-A were expressed at the apical side of the duct epithelium in submandibular gland (SMG) tissue and at the cell borders in HSDECs of normal and IgG4-RD. The expression and distribution of TJs in SMG tissue were not different in control individuals and patients with IgG4-RD in vivo and in vitro. Although interferon-gamma (IFNγ) generally disrupts the integrity and function of TJs, as manifested by decreased epithelial barrier function, IFNγ markedly increased the epithelial barrier function of HSDECs via upregulation of claudin-7 expression in HSDECs from patients with IgG4-RD. This is the first report showing an IFNγ-dependent increase in epithelial barrier function in the salivary gland duct epithelium. Our results provide insights into the functional significance of TJs in salivary gland duct epithelium in physiological and pathological conditions, including IgG4-RD. PMID:26956365

  16. Intestinal epithelial cell apoptosis and loss of barrier function in the setting of altered microbiota with enteral nutrient deprivation

    PubMed Central

    Demehri, Farokh R.; Barrett, Meredith; Ralls, Matthew W.; Miyasaka, Eiichi A.; Feng, Yongjia; Teitelbaum, Daniel H.

    2013-01-01

    Total parenteral nutrition (TPN), a commonly used treatment for patients who cannot receive enteral nutrition, is associated with significant septic complications due in part to a loss of epithelial barrier function (EBF). While the underlying mechanisms of TPN-related epithelial changes are poorly understood, a mouse model of TPN-dependence has helped identify several contributing factors. Enteral deprivation leads to a shift in intestinal microbiota to predominantly Gram-negative Proteobacteria. This is associated with an increase in expression of proinflammatory cytokines within the mucosa, including interferon-γ and tumor necrosis factor-α. A concomitant loss of epithelial growth factors leads to a decrease in epithelial cell proliferation and increased apoptosis. The resulting loss of epithelial tight junction proteins contributes to EBF dysfunction. These mechanisms identify potential strategies of protecting against TPN-related complications, such as modification of luminal bacteria, blockade of proinflammatory cytokines, or growth factor replacement. PMID:24392360

  17. Boswellia serrata Preserves Intestinal Epithelial Barrier from Oxidative and Inflammatory Damage

    PubMed Central

    Catanzaro, Daniela; Rancan, Serena; Orso, Genny; Dall’Acqua, Stefano; Brun, Paola; Giron, Maria Cecilia; Carrara, Maria; Castagliuolo, Ignazio; Ragazzi, Eugenio; Caparrotta, Laura; Montopoli, Monica

    2015-01-01

    Aminosalicylates, corticosteroids and immunosuppressants are currently the therapeutic choices in inflammatory bowel diseases (IBD), however, with limited remission and often serious side effects. Meanwhile complementary and alternative medicine (CAM) use is increasing, particularly herbal medicine. Boswellia serrata is a traditional Ayurvedic remedy with anti-inflammatory properties, of interest for its usefulness in IBDs. The mechanism of this pharmacological potential of Boswellia serrata was investigated in colonic epithelial cell monolayers exposed to H2O2 or INF-γ+TNF-α, chosen as in vitro experimental model of intestinal inflammation. The barrier function was evaluated by the transepithelial electrical resistance (TEER) and paracellular permeability assay, and by the tight junction proteins (zonula occludens-1, ZO-1 and occludin) immunofluorescence. The expression of phosphorylated NF-κB and reactive oxygen species (ROS) generation were determined by immunoblot and cytofluorimetric assay, respectively. Boswellia serrata oleo-gum extract (BSE) and its pure derivative acetyl-11-keto-β-boswellic acid (AKBA), were tested at 0.1-10 μg/ml and 0.027μg/ml, respectively. BSE and AKBA safety was demonstrated by no alteration of intestinal cell viability and barrier function and integrity biomarkers. H2O2 or INF-γ+TNF-α treatment of Caco-2 cell monolayers significantly reduced TEER, increased paracellular permeability and caused the disassembly of tight junction proteins occludin and ZO-1. BSE and AKBA pretreatment significantly prevented functional and morphological alterations and also the NF-κB phosphorylation induced by the inflammatory stimuli. At the same concentrations BSE and AKBA counteracted the increase of ROS caused by H2O2 exposure. Data showed the positive correlation of the antioxidant activity with the mechanism involved in the physiologic maintenance of the integrity and function of the intestinal epithelium. This study elucidates the

  18. Boswellia serrata Preserves Intestinal Epithelial Barrier from Oxidative and Inflammatory Damage.

    PubMed

    Catanzaro, Daniela; Rancan, Serena; Orso, Genny; Dall'Acqua, Stefano; Brun, Paola; Giron, Maria Cecilia; Carrara, Maria; Castagliuolo, Ignazio; Ragazzi, Eugenio; Caparrotta, Laura; Montopoli, Monica

    2015-01-01

    Aminosalicylates, corticosteroids and immunosuppressants are currently the therapeutic choices in inflammatory bowel diseases (IBD), however, with limited remission and often serious side effects. Meanwhile complementary and alternative medicine (CAM) use is increasing, particularly herbal medicine. Boswellia serrata is a traditional Ayurvedic remedy with anti-inflammatory properties, of interest for its usefulness in IBDs. The mechanism of this pharmacological potential of Boswellia serrata was investigated in colonic epithelial cell monolayers exposed to H2O2 or INF-γ+TNF-α, chosen as in vitro experimental model of intestinal inflammation. The barrier function was evaluated by the transepithelial electrical resistance (TEER) and paracellular permeability assay, and by the tight junction proteins (zonula occludens-1, ZO-1 and occludin) immunofluorescence. The expression of phosphorylated NF-κB and reactive oxygen species (ROS) generation were determined by immunoblot and cytofluorimetric assay, respectively. Boswellia serrata oleo-gum extract (BSE) and its pure derivative acetyl-11-keto-β-boswellic acid (AKBA), were tested at 0.1-10 μg/ml and 0.027 μg/ml, respectively. BSE and AKBA safety was demonstrated by no alteration of intestinal cell viability and barrier function and integrity biomarkers. H2O2 or INF-γ+TNF-α treatment of Caco-2 cell monolayers significantly reduced TEER, increased paracellular permeability and caused the disassembly of tight junction proteins occludin and ZO-1. BSE and AKBA pretreatment significantly prevented functional and morphological alterations and also the NF-κB phosphorylation induced by the inflammatory stimuli. At the same concentrations BSE and AKBA counteracted the increase of ROS caused by H2O2 exposure. Data showed the positive correlation of the antioxidant activity with the mechanism involved in the physiologic maintenance of the integrity and function of the intestinal epithelium. This study elucidates the

  19. Lactobacillus plantarum inhibits intestinal epithelial barrier dysfunction induced by unconjugated bilirubin.

    PubMed

    Zhou, Yukun; Qin, Huanlong; Zhang, Ming; Shen, Tongyi; Chen, Hongqi; Ma, Yanlei; Chu, Zhaoxin; Zhang, Peng; Liu, Zhihua

    2010-08-01

    Although a large number of in vitro and in vivo tests have confirmed that taking probiotics can improve the intestinal barrier, few studies have focused on the relationship between probiotics and the intestinal epithelial barrier in hyperbilirubinaemia. To investigate the effects of and mechanisms associated with probiotic bacteria (Lactobacillus plantarum; LP) and unconjugated bilirubin (UCB) on the intestinal epithelial barrier, we measured the viability, apoptotic ratio and protein kinase C (PKC) activity of Caco-2 cells. We also determined the distribution and expression of tight junction proteins such as occludin, zonula occludens (ZO)-1, claudin-1, claudin-4, junctional adhesion molecule (JAM)-1 and F-actin using confocal laser scanning microscopy, immunohistochemistry, Western blotting and real-time quantitative PCR. The present study demonstrated that high concentrations of UCB caused obvious cytotoxicity and decreased the transepithelial electrical resistance (TER) of the Caco-2 cell monolayer. Low concentrations of UCB inhibited the expression of tight junction proteins and PKC but could induce UDP-glucuronosyltransferases 1 family-polypeptide A1 (UGT1A1) expression. UCB alone caused decreased PKC activity, serine phosphorylated occludin and ZO-1 levels. After treatment with LP, the effects of UCB on TER and apoptosis were mitigated; LP also prevented aberrant expression and rearrangement of tight junction proteins. Moreover, PKC activity and serine phosphorylated tight junction protein levels were partially restored after treatment with LP, LP exerted a protective effect against UCB damage to Caco-2 monolayer cells, and it restored the structure and distribution of tight junction proteins by activating the PKC pathway. In addition, UGT1A1 expression induced by UCB in Caco-2 cells could ameliorate the cytotoxicity of UCB. PMID:20412608

  20. Gut microbiota facilitates dietary heme-induced epithelial hyperproliferation by opening the mucus barrier in colon

    PubMed Central

    Ijssennagger, Noortje; Belzer, Clara; Hooiveld, Guido J.; Dekker, Jan; van Mil, Saskia W. C.; Müller, Michael; Kleerebezem, Michiel; van der Meer, Roelof

    2015-01-01

    Colorectal cancer risk is associated with diets high in red meat. Heme, the pigment of red meat, induces cytotoxicity of colonic contents and elicits epithelial damage and compensatory hyperproliferation, leading to hyperplasia. Here we explore the possible causal role of the gut microbiota in heme-induced hyperproliferation. To this end, mice were fed a purified control or heme diet (0.5 μmol/g heme) with or without broad-spectrum antibiotics for 14 d. Heme-induced hyperproliferation was shown to depend on the presence of the gut microbiota, because hyperproliferation was completely eliminated by antibiotics, although heme-induced luminal cytotoxicity was sustained in these mice. Colon mucosa transcriptomics revealed that antibiotics block heme-induced differential expression of oncogenes, tumor suppressors, and cell turnover genes, implying that antibiotic treatment prevented the heme-dependent cytotoxic micelles to reach the epithelium. Our results indicate that this occurs because antibiotics reinforce the mucus barrier by eliminating sulfide-producing bacteria and mucin-degrading bacteria (e.g., Akkermansia). Sulfide potently reduces disulfide bonds and can drive mucin denaturation and microbial access to the mucus layer. This reduction results in formation of trisulfides that can be detected in vitro and in vivo. Therefore, trisulfides can serve as a novel marker of colonic mucolysis and thus as a proxy for mucus barrier reduction. In feces, antibiotics drastically decreased trisulfides but increased mucin polymers that can be lysed by sulfide. We conclude that the gut microbiota is required for heme-induced epithelial hyperproliferation and hyperplasia because of the capacity to reduce mucus barrier function. PMID:26216954

  1. Candidalysin is a fungal peptide toxin critical for mucosal infection

    PubMed Central

    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.

    2016-01-01

    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. Until now human pathogenic fungi were not known to possess such toxins. Here we identify the first fungal cytolytic peptide toxin in the opportunistic pathogen Candida albicans. This secreted toxin directly damages epithelial membranes, triggers a danger response signaling pathway and activates epithelial immunity. Toxin-mediated membrane permeabilization is enhanced by a positively charged C-terminus and 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. TNFα Induces Choroid Plexus Epithelial Cell Barrier Alterations by Apoptotic and Nonapoptotic Mechanisms

    PubMed Central

    Schwerk, Christian; Rybarczyk, Kasia; Essmann, Frank; Seibt, Annette; Mölleken, Marie-Louise; Zeni, Patrick; Schroten, Horst; Tenenbaum, Tobias

    2010-01-01

    The choroid plexus epithelium constitutes the structural basis of the blood-cerebrospinal fluid barrier. Since the cytokine TNFα is markedly increased during inflammatory diseases in the blood and the central nervous system, we investigated by which mechanisms TNFα induces barrier alteration in porcine choroid plexus epithelial cells. We found a dose-dependent decrease of transepithelial electrical resistance, increase of paracellular inulin-flux, and induction of histone-associated DNA fragmentation and caspase-3 activation after TNFα stimulation. This response was strongly aggravated by the addition of cycloheximide and could partially be inhibited by the NF-κB inhibitor CAPE, but most effectively by the pan-caspase-inhibitor zVAD-fmk and not by the JNK inhibitor SP600125. Partial loss of cell viability could also be attenuated by CAPE. Immunostaining showed cell condensation and nuclear binding of high-mobility group box 1 protein as a sign of apoptosis after TNFα stimulation. Taken together our findings indicate that TNFα compromises PCPEC barrier function by caspase and NF-κB dependent mechanisms. PMID:20369072

  3. Live Faecalibacterium prausnitzii in an apical anaerobic model of the intestinal epithelial barrier.

    PubMed

    Ulluwishewa, Dulantha; Anderson, Rachel C; Young, Wayne; McNabb, Warren C; van Baarlen, Peter; Moughan, Paul J; Wells, Jerry M; Roy, Nicole C

    2015-02-01

    Faecalibacterium prausnitzii, an abundant member of the human commensal microbiota, has been proposed to have a protective role in the intestine. However, it is an obligate anaerobe, difficult to co-culture in viable form with oxygen-requiring intestinal cells. To overcome this limitation, a unique apical anaerobic model of the intestinal barrier, which enabled co-culture of live obligate anaerobes with the human intestinal cell line Caco-2, was developed. Caco-2 cells remained viable and maintained an intact barrier for at least 12 h, consistent with gene expression data, which suggested Caco-2 cells had adapted to survive in an oxygen-reduced atmosphere. Live F. prausnitzii cells, but not ultraviolet (UV)-killed F. prausnitzii, increased the permeability of mannitol across the epithelial barrier. Gene expression analysis showed inflammatory mediators to be expressed at lower amounts in Caco-2 cells exposed to live F. prausnitzii than UV-killed F. prausnitzii, This, consistent with previous reports, implies that live F. prausnitzii produces an anti-inflammatory compound in the culture supernatant, demonstrating the value of a physiologically relevant co-culture system that allows obligate anaerobic bacteria to remain viable.

  4. DIGESTIVE PHYSIOLOGY OF THE PIG SYMPOSIUM: Involvement of gut chemosensing in the regulation of mucosal barrier function and defense mechanisms1,2

    PubMed Central

    Kaji, I.; Akiba, Y.; Kaunitz, J. D.

    2016-01-01

    Meal ingestion is followed by release of numerous hormones from enteroendocrine cells interspersed among the epithelial cells lining the intestine. Recently, the de-orphanization of G protein-coupled receptor (GPCR)-type nutrient receptors, expressed on the apical membranes of enteroendocrine cells, has suggested a plausible mechanism whereby luminal nutrients trigger the release of gut hormones. Activation of nutrient receptors triggers intracellular signaling mechanisms that promote exocytosis of hormone-containing granules into the submucosal space. Hormones released by foregut enteroendocrine cells include the glucagon-like peptides (GLP) affecting glycemic control (GLP-1) and releasing pro-proliferative, hypertrophy-inducing growth factors (GLP-2). The foregut mucosa, being exposed to pulses of concentrated HCl, is protected by a system of defense mechanisms, which includes epithelial bicarbonate and mucus secretion and augmentation of mucosal blood flow. We have reported that luminal co-perfusion of AA with nucleotides in anesthetized rats releases GLP-2 into the portal vein, associated with increased bicarbonate and mucus secretion and mucosal blood flow. The GLP-2 increases bicarbonate secretion via release of vasoactive intestinal peptide (VIP) from myenteric nerves. Luminal bile acids also release gut hormones due to activation of the bile-acid receptor known as G Protein-Coupled Receptor (GPR) 131, G Protein Bile Acid Receptor (GPBAR) 1, or Takeda G Protein-Coupled Receptor (TGR) 5, also expressed on enteroendocrine cells. The GLP are metabolized by dipeptidyl peptidase IV (DPPIV), an enzyme of particular interest to pharmaceutical, because its inhibition increases plasma concentrations of GLP-1 to treat diabetes. We have also reported that DPPIV inhibition enhances the secretory effects of nutrient-evoked GLP-2. Understanding the release mechanism and the metabolic pathways of gut hormones is of potential utility to the formulation of feedstuff

  5. High-mobility group box 1 impairs airway epithelial barrier function through the activation of the RAGE/ERK pathway

    PubMed Central

    HUANG, WUFENG; ZHAO, HAIJIN; DONG, HANGMING; WU, YUE; YAO, LIHONG; ZOU, FEI; CAI, SHAOXI

    2016-01-01

    Recent studies have indicated that high-mobility group box 1 protein (HMGB1) and the receptor for advanced glycation end-products (RAGE) contribute to the pathogenesis of asthma. However, whether the activation of the HMGB1/RAGE axis mediates airway epithelial barrier dysfunction remains unknown. Thus, the aim of this study was to examine the effects of HMGB1 and its synergistic action with interleukin (IL)-1β on airway epithelial barrier properties. We evaluated the effects of recombinant human HMGB1 alone or in combination with IL-1β on ionic and macromolecular barrier permeability, by culturing air-liquid interface 16HBE cells with HMGB1 to mimic the differentiated epithelium. Western blot analysis and immunofluorescence staining were utilized to examine the level and structure of major junction proteins, namely E-cadherin, β-catenin, occludin and claudin-1. Furthermore, we examined the effects of RAGE neutralizing antibodies and mitogen-activated protein kinase (MAPK) inhibitors on epithelial barrier properties in order to elucidate the mechanisms involved. HMGB1 increased FITC-dextran permeability, but suppressed epithelial resistance in a dose-and time-dependent manner. HMGB1-mediated barrier hyperpermeability was accompanied by a disruption of cell-cell contacts, the selective downregulation of occludin and claudin-1, and the redistribution of E-cadherin and β-catenin. HMGB1 in synergy with IL-1β induced a similar, but greater barrier hyperpermeability and induced the disruption of junction proteins. Furthermore, HMGB1 elicited the activation of the RAGE/extracellular signal-related kinase (ERK)1/2 signaling pathway, which correlated with barrier dysfunction in the 16HBE cells. Anti-RAGE antibody and the ERK1/2 inhibitor, U0126, attenuated the HMGB1-mediated changes in barrier permeability, restored the expression levels of occludin and claudin-1 and pevented the redistribution of E-cadherin and β-catenin. Taken together, the findings of our study

  6. Glial cell line-derived neurotrophic factor promotes barrier maturation and wound healing in intestinal epithelial cells in vitro.

    PubMed

    Meir, Michael; Flemming, Sven; Burkard, Natalie; Bergauer, Lisa; Metzger, Marco; Germer, Christoph-Thomas; Schlegel, Nicolas

    2015-10-15

    Recent data suggest that neurotrophic factors from the enteric nervous system are involved in intestinal epithelial barrier regulation. In this context the glial cell line-derived neurotrophic factor (GDNF) was shown to affect gut barrier properties in vivo directly or indirectly by largely undefined processes in a model of inflammatory bowel disease (IBD). We further investigated the potential role and mechanisms of GDNF in the regulation of intestinal barrier functions. Immunostaining of human gut specimen showed positive GDNF staining in enteric neuronal plexus and in enterocytes. In Western blots of the intestinal epithelial cell lines Caco2 and HT29B6, significant amounts of GDNF were detected, suggesting that enterocytes represent an additional source of GDNF. Application of recombinant GDNF on Caco2 and HT29B6 cells for 24 h resulted in significant epithelial barrier stabilization in monolayers with immature barrier functions. Wound-healing assays showed a significantly faster closure of the wounded areas after GDNF application. GDNF augmented cAMP levels and led to significant inactivation of p38 MAPK in immature cells. Activation of p38 MAPK signaling by SB-202190 mimicked GDNF-induced barrier maturation, whereas the p38 MAPK activator anisomycin blocked GDNF-induced effects. Increasing cAMP levels had adverse effects on barrier maturation, as revealed by permeability measurements. However, increased cAMP augmented the proliferation rate in Caco2 cells, and GDNF-induced proliferation of epithelial cells was abrogated by the PKA inhibitor H89. Our data show that enterocytes represent an additional source of GDNF synthesis. GDNF contributes to wound healing in a cAMP/PKA-dependent manner and promotes barrier maturation in immature enterocytes cells by inactivation of p38 MAPK signaling.

  7. 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

  8. Simulation of in-vivo-equivalent epithelial barriers using a micro fluidic device.

    PubMed

    Greß, C; Jeziorski, M; Saumer, M; Schäfer, K-H

    2014-04-01

    In biomedical approaches cell culture models do often not fully represent their biological counterparts. Often the methods used do not completely mimic the in-vivo situation, either by using only single-cell-type culture approaches, or by using inadequate culture conditions. We therefore developed a variable system based on individual modules to simulate in vitro equivalent cell-barriers (e.g. for mucous layers). This system allows the growth of different communicating cell types in micro channels. Hot embossing is used to fabricate the micro structured polymer sheets. The stamp for hot embossing is fabricated by UV-lithography/electroforming or by micro milling. The system consists of a container with micro fluidic modules and a pump-system for a continuous medium-supply. An individual module is made of two micro-structured polycarbonate-sheets separated by a transmissible polycarbonate membrane. The two sheets are arranged orthogonally to induce a cross flow. The system is highly variable by channel-geometry (height and width), capacity (number of micro fluidic modules), and pore sizes of the transmissible membranes. In a first approach we simulated the intestinal mucosa. Epithelial cells and primary neurons of the enteric nervous system were cultured on both sides of the transmissible membrane within the two different compartments. So the cells could be supplied with two different media. We kept a mono-culture of primary neurons or epithelial cells for 5 days and a co-culture between these two cell-types was established for 4 days. The proposed system delivers a sophisticated model for the simulation of various epithelial layers which takes the specific biological properties into account.

  9. DHA protects against experimental colitis in IL-10-deficient mice associated with the modulation of intestinal epithelial barrier function.

    PubMed

    Zhao, Jie; Shi, Peiliang; Sun, Ye; Sun, Jing; Dong, Jian-Ning; Wang, Hong-Gang; Zuo, Lu-Gen; Gong, Jian-Feng; Li, Yi; Gu, Li-Li; Li, Ning; Li, Jie-Shou; Zhu, Wei-Ming

    2015-07-01

    A defect in the intestinal barrier is one of the characteristics of Crohn's disease (CD). The tight junction (TJ) changes and death of epithelial cells caused by intestinal inflammation play an important role in the development of CD. DHA, a long-chain PUFA, has been shown to be helpful in treating inflammatory bowel disease in experimental models by inhibiting the NF-κB pathway. The present study aimed at investigating the specific effect of DHA on the intestinal barrier function in IL-10-deficient mice. IL-10-deficient mice (IL-10(-/-)) at 16 weeks of age with established colitis were treated with DHA (i.g. 35.5 mg/kg per d) for 2 weeks. The severity of their colitis, levels of pro-inflammatory cytokines, epithelial gene expression, the distributions of TJ proteins (occludin and zona occludens (ZO)-1), and epithelial apoptosis in the proximal colon were measured at the end of the experiment. DHA treatment attenuated the established colitis and was associated with reduced infiltration of inflammatory cells in the colonic mucosa, lower mean histological scores and decreased levels of pro-inflammatory cytokines (IL-17, TNF-α and interferon-γ). Moreover, enhanced barrier function was observed in the DHA-treated mice that resulted from attenuated colonic permeability, rescued expression and corrected distributions of occludin and ZO-1. The results of the present study indicate that DHA therapy may ameliorate experimental colitis in IL-10(-/-) mice by improving the intestinal epithelial barrier function.

  10. Age-related differences in mucosal barrier function and morphology of the small intestine in low and normal birth weight piglets.

    PubMed

    Huygelen, V; De Vos, M; Willemen, S; Fransen, E; Casteleyn, C; Van Cruchten, S; Van Ginneken, C

    2014-08-01

    To test the hypothesis that the mucosal maturation of the small intestine is altered in low birth weight piglets, pairs of naturally suckled low birth weight (LBW, n = 20) and normal birth weight (NBW, n = 20) littermate piglets were selected and sampled after 0, 3, 10, and 28 d of suckling. In vivo intestinal permeability was evaluated via a lactulose-mannitol absorption test. Other indirect measurements for mucosal barrier functioning included sampling for histology and immunohistochemistry (intestinal trefoil factor [ITF]), measuring intestinal alkaline phosphatase (IAP) activity, and immunoblotting for occludin, caspase-3, and proliferating cell nuclear antigen (PCNA). The lactulose-mannitol ratio did not differ between NBW and LBW piglets, but a significant increase in this ratio was observed in 28-d-old piglets (P = 0.001). Small intestinal villus height did not differ with age (P = 0.02) or birth weight (P = 0.20). In contrast, villus width (P = 0.02) and crypt depth (P < 0.05) increased gradually with age, but no birth-weight-related differences were observed. LBW piglets had significantly (P = 0.03) more ITF immunoreactive positive cells per villus area compared to NBW piglets, whereas no age (P = 0.82) or region-related (P = 0.13) differences could be observed. The activity of IAP in the small intestine was higher in newborn piglets compared to the older piglets. No significant differences in cell proliferation in the small intestine was observed (P = 0.47) between NBW and LBW piglets; the highest proliferation was seen in piglets of 28 d of age (P = 0.01). Newborn piglets had significantly fewer apoptotic cells, whereas more apoptotic cells were seen in piglets of 10 d of age (P < 0.01). In conclusion, birth weight did not affect the parameters related to intestinal barrier function investigated in this study, suggesting that the mucosal barrier function is not altered in LBW piglets. Nevertheless, these results confirm that the mucosal barrier function

  11. The mechanisms of gastric mucosal injury: focus on microvascular endothelium as a key target.

    PubMed

    Tarnawski, A S; Ahluwalia, A; Jones, M K

    2012-01-01

    This paper reviews and updates current views on gastric mucosal injury with a focus on the microvascular endothelium as the key target and the role of the anti-apoptosis protein survivin. Under normal conditions, mucosal integrity is maintained by well structured and mutually amplifying defense mechanisms, which include pre-epithelial "barrier"--the first line of defense; and, an epithelial "barrier". Other important defense mechanisms of gastric mucosa include: continuous epithelial cell renewal, blood flow through mucosal microvessels (providing oxygen and nutrients), an endothelial microvascular "barrier," sensory innervation, and generation of PGs, nitric oxide and hydrogen sulfide. The microvascular endothelium lining gastric mucosal blood microvessels severs not only as a barrier but is a biologically active tissue involved in many synthetic and metabolic functions. It allows transport of oxygen and nutrients, and produces prostaglandins and leukotriens, procoagulant factors, nitric oxide, endothelin, ghrelin, HSP, growth factors such VEGF, bFGF, angiopoietin 2 and others, specific types of collagen, plasminogen activator, and can also actively contract. Accumulating evidence indicates that the gastric microvascular endothelium is a critical target for injury by ethanol, NSAIDs, free radicals, ischemia-reperfusion and other damaging factors. The injury--microvessel rupture, plasma and erythrocyte extravasation, platelet aggregation and fibrin deposition caused by these damaging factors--occurs early (1-5 min), precedes glandular epithelial cell injury and results in cessation of blood flow, ischemia, hypoxia and impaired oxygen and nutrient transport. As a consequence, mucosal necrosis develops. One of the main reasons for the increased susceptibility of gastric microvascular endothelial (vs. epithelial) cells to injury is reduced expression levels of survivin, an anti-apoptosis protein, which is a regulator of both proliferation and cell survival. PMID

  12. Meprin A impairs epithelial barrier function, enhances monocyte migration, and cleaves the tight junction protein occludin

    PubMed Central

    Bao, Jialing; Yura, Renee E.; Matters, Gail L.; Bradley, S. Gaylen; Shi, Pan; Tian, Fang

    2013-01-01

    Meprin metalloproteases are highly expressed at the luminal interface of the intestine and kidney and in certain leukocytes. Meprins cleave a variety of substrates in vitro, including extracellular matrix proteins, adherens junction proteins, and cytokines, and have been implicated in a number of inflammatory diseases. The linkage between results in vitro and pathogenesis, however, has not been elucidated. The present study aimed to determine whether meprins are determinative factors in disrupting the barrier function of the epithelium. Active meprin A or meprin B applied to Madin-Darby canine kidney (MDCK) cell monolayers increased permeability to fluorescein isothiocyanate-dextran and disrupted immunostaining of the tight junction protein occludin but not claudin-4. Meprin A, but not meprin B, cleaved occludin in MDCK monolayers. Experiments with recombinant occludin demonstrated that meprin A cleaves the protein between Gly100 and Ser101 on the first extracellular loop. In vivo experiments demonstrated that meprin A infused into the mouse bladder increased the epithelium permeability to sodium fluorescein. Furthermore, monocytes from meprin knockout mice on a C57BL/6 background were less able to migrate through an MDCK monolayer than monocytes from their wild-type counterparts. These results demonstrate the capability of meprin A to disrupt epithelial barriers and implicate occludin as one of the important targets of meprin A that may modulate inflammation. PMID:23804454

  13. Deoxynivalenol affects in vitro intestinal epithelial cell barrier integrity through inhibition of protein synthesis

    SciTech Connect

    Van De Walle, Jacqueline; Sergent, Therese; Piront, Neil; Toussaint, Olivier; Schneider, Yves-Jacques; Larondelle, Yvan

    2010-06-15

    Deoxynivalenol (DON), one of the most common mycotoxin contaminants of raw and processed cereal food, adversely affects the gastrointestinal tract. Since DON acts as a protein synthesis inhibitor, the constantly renewing intestinal epithelium could be particularly sensitive to DON. We analyzed the toxicological effects of DON on intestinal epithelial protein synthesis and barrier integrity. Differentiated Caco-2 cells, as a widely used model of the human intestinal barrier, were exposed to realistic intestinal concentrations of DON (50, 500 and 5000 ng/ml) during 24 h. DON caused a concentration-dependent decrease in total protein content associated with a reduction in the incorporation of [{sup 3}H]-leucine, demonstrating its inhibitory effect on protein synthesis. DON simultaneously increased the paracellular permeability of the monolayer as reflected through a decreased transepithelial electrical resistance associated with an increased paracellular flux of the tracer [{sup 3}H]-mannitol. A concentration-dependent reduction in the expression level of the tight junction constituent claudin-4 was demonstrated by Western blot, which was not due to diminished transcription, increased degradation, or NF-{kappa}B, ERK or JNK activation, and was also observed for a tight junction independent protein, i.e. intestinal alkaline phosphatase. These results demonstrate a dual toxicological effect of DON on differentiated Caco-2 cells consisting in an inhibition of protein synthesis as well as an increase in monolayer permeability, and moreover suggest a possible link between them through diminished synthesis of the tight junction constituent claudin-4.

  14. Expression of claudins -2 and -4 and cingulin is coordinated with the start of stratification and differentiation in corneal epithelial cells: retinoic acid reversibly disrupts epithelial barrier

    PubMed Central

    Ortiz-Melo, María Teresa; Sánchez-Guzmán, Erika; González-Robles, Arturo; Valdés, Jesús; Gómez-Flores, Eber; Castro-Muñozledo, Federico

    2013-01-01

    Summary Although tight junctions (TJ) have been extensively studied in simple epithelial cells, it is still unknown whether their organization is coupled to cell differentiation in stratified epithelia. We studied the expression of TJ in RCE1(5T5) cells, an in vitro model which mimics the sequential steps of rabbit corneal epithelial differentiation. RCE1(5T5) cells expressed TJ components which were assembled once cells constituted differentiated epithelia, as suggested by the increase of transepithelial electrical resistance (TER) which followed a similar kinetic to the expression of the early differentiation marker Pax-6. TJ were functional as indicated by the establishment of an epithelial barrier nonpermeable to ruthenium red or a biotin tracer. In immunostaining experiments, TJ were located at the superficial cells from the suprabasal layers; Western blot and RT-PCR suggested that TJ were composed of claudins (cldn) -1, -2, -4, cingulin (cgn), occludin (ocln) and ZO-1. Semi-quantitative RT-PCR and TER measurements showed that TJ became organized when cells began to form a 3–5 layers stratified epithelium; TER increased once cells reached confluence, with a time course comparable to the raise in the expression of cgn, cldn-2 and -4. Nevertheless, cldn-1, -2, ZO-1 and ocln were present in the cells from the beginning of cultivation, suggesting that TER increases mainly depend on TJ assembly. While EGF increased epithelial barrier strength, retinoic acid disrupted it, increasing paracellular flux about 2-fold; this effect was concentration dependent and completely reversible. Our results suggest that TJ assembly is tightly linked to the expression of corneal epithelial terminal phenotype. PMID:23429425

  15. Characterization and mucosal responses of interleukin 17 family ligand and receptor genes in channel catfish Ictalurus punctatus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Interleukin (IL) 17 family cytokines are important mediators of mucosal immune responses, tightly regulated by signals from the complex milieu of pathogenic and commensal microbes, epithelial cells and innate and adaptive leukocytes found at tissue barriers. In mammals, IL17 ligand expression has be...

  16. Activation of muscarinic cholinoceptor ameliorates tumor necrosis factor-α-induced barrier dysfunction in intestinal epithelial cells.

    PubMed

    Khan, Md Rafiqul Islam; Uwada, Junsuke; Yazawa, Takashi; Islam, Md Tariqul; Krug, Susanne M; Fromm, Michael; Karaki, Shin-ichiro; Suzuki, Yuichi; Kuwahara, Atsukazu; Yoshiki, Hatsumi; Sada, Kiyonao; Muramatsu, Ikunobu; Anisuzzaman, Abu Syed Md; Taniguchi, Takanobu

    2015-11-30

    Impaired intestinal barrier function is one of the critical issues in inflammatory bowel diseases. The aim of this study is to investigate muscarinic cholinoceptor (mAChR)-mediated signaling for the amelioration of cytokine-induced barrier dysfunction in intestinal epithelium. Rat colon challenged with TNF-α and interferon γ reduced transepithelial electrical resistance (TER). This barrier injury was attenuated by muscarinic stimulation. In HT-29/B6 intestinal epithelial cells, muscarinic stimulation suppressed TNF-α-induced activation of NF-κB signaling and barrier disruption. Finally, muscarinic stimulation promoted the shedding of TNFR1, which would be a mechanism for the attenuation of TNF-α/NF-κB signaling and barrier disruption via mAChR.

  17. Glutamine Prevents Total Parenteral Nutrition-Associated Changes to Intraepithelial Lymphocyte Phenotype and Function: A Potential Mechanism for the Preservation of Epithelial Barrier Function

    PubMed Central

    2010-01-01

    Total parenteral nutrition (TPN) results in a number of derangements to the intestinal epithelium, including a loss of epithelial barrier function (EBF). As TPN supplemented with glutamine has been thought to prevent this loss, this article further defined the impact of glutamine on EBF, and investigated potential mechanisms that contributed to the preservation of EBF. C57BL/6J male mice were randomized to enteral nutrition (control), TPN, or TPN supplemented with glutamine (TPN+GLN). Changes in intraepithelial lymphocyte (IEL)-derived cytokine expression were measured, and EBF was assessed with electrophysiologic methods and assessment of junctional protein expression. TPN resulted in a significant decline in EBF, and this loss of EBF was significantly prevented in the TPN+GLN group. Coincident with these changes was a loss of intraepithelial lymphocyte (IEL, mucosal lymphocyte)-derived IL-10 and increase in interferon-γ (IFN-γ) expression, and a decline in IEL numbers in the TPN group. A prevention in the increase in IFN-γ and decline in IL-10 expression was seen in the TPN+GLN group. To determine the mechanism responsible for these glutamine-associated cytokine changes, we tested whether blockade of the IL-7 signaling pathway between epithelial cells (EC) and IEL would prevent these changes; however, blockade failed to influence IEL-derived cytokine changes. Glutamine-supplemented TPN leads to a specific IEL-derived cytokine profile, which may account for the preservation of EBF; and such action may be due to a direct action of glutamine on the IEL. PMID:20028208

  18. Decreased epithelial barrier function evoked by exposure to metabolic stress and nonpathogenic E. coli is enhanced by TNF-alpha.

    PubMed

    Lewis, Kimberley; Caldwell, Jackie; Phan, Van; Prescott, David; Nazli, Aisha; Wang, Arthur; Soderhölm, Johan D; Perdue, Mary H; Sherman, Philip M; McKay, Derek M

    2008-03-01

    A defect in mitochondrial activity contributes to many diseases. We have shown that monolayers of the human colonic T84 epithelial cell line exposed to dinitrophenol (DNP, uncouples oxidative phosphorylation) and nonpathogenic Escherichia coli (E. coli) (strain HB101) display decreased barrier function. Here the impact of DNP on macrophage activity and the effect of TNF-alpha, DNP, and E. coli on epithelial permeability were assessed. DNP treatment of the human THP-1 macrophage cell line resulted in reduced ATP synthesis, and, although hyporesponsive to LPS, the metabolically stressed macrophages produced IL-1beta, IL-6, and TNF-alpha. Given the role of TNF-alpha in inflammatory bowel disease (IBD) and the association between increased permeability and IBD, recombinant TNF-alpha (10 ng/ml) was added to the DNP (0.1 mM) + E. coli (10(6) colony-forming units), and this resulted in a significantly greater loss of T84 epithelial barrier function than that elicited by DNP + E. coli. This increased epithelial permeability was not due to epithelial death, and the enhanced E. coli translocation was reduced by pharmacological inhibitors of NF-kappabeta signaling (pyrrolidine dithiocarbamate, NF-kappabeta essential modifier-binding peptide, BAY 11-7082, and the proteosome inhibitor, MG132). In contrast, the drop in transepithelial electrical resistance was unaffected by the inhibitors of NF-kappabeta. Thus, as an integrative model system, our findings support the induction of a positive feedback loop that can severely impair epithelial barrier function and, as such, could contribute to existing inflammation or trigger relapses in IBD. Thus metabolically stressed epithelia display increased permeability in the presence of viable nonpathogenic E. coli that is exaggerated by TNF-alpha released by activated immune cells, such as macrophages, that retain this ability even if they themselves are experiencing a degree of metabolic stress.

  19. An in vitro triple cell co-culture model with primary cells mimicking the human alveolar epithelial barrier.

    PubMed

    Lehmann, Andrea D; Daum, Nicole; Bur, Michael; Lehr, Claus-Michael; Gehr, Peter; Rothen-Rutishauser, Barbara M

    2011-04-01

    A triple cell co-culture model was recently established by the authors, consisting of either A549 or 16HBE14o- epithelial cells, human blood monocyte-derived macrophages and dendritic cells, which offers the possibility to study the interaction of xenobiotics with those cells. The 16HBE14o- containing co-culture model mimics the airway epithelial barrier, whereas the A549 co-cultures mimic the alveolar type II-like epithelial barrier. The goal of the present work was to establish a new triple cell co-culture model composed of primary alveolar type I-like cells isolated from human lung biopsies (hAEpC) representing a more realistic alveolar epithelial barrier wall, since type I epithelial cells cover >93% of the alveolar surface. Monocultures of A549 and 16HBE14o- were morphologically and functionally compared with the hAEpC using laser scanning microscopy, as well as transmission electron microscopy, and by determining the epithelial integrity. The triple cell co-cultures were characterized using the same methods. It could be shown that the epithelial integrity of hAEpC (mean ± SD, 1180 ± 188 Ω cm(2)) was higher than in A549 (172 ± 59 Ω cm(2)) but similar to 16HBE14o- cells (1469 ± 156 Ω cm(2)). The triple cell co-culture model with hAEpC (1113 ± 30 Ω cm(2)) showed the highest integrity compared to the ones with A549 (93 ± 14 Ω cm(2)) and 16HBE14o- (558 ± 267 Ω cm(2)). The tight junction protein zonula occludens-1 in hAEpC and 16HBE14o- were more regularly expressed but not in A549. The epithelial alveolar model with hAEpC combined with two immune cells (i.e. macrophages and dendritic cells) will offer a novel and more realistic cell co-culture system to study possible cell interactions of inhaled xenobiotics and their toxic potential on the human alveolar type I epithelial wall.

  20. Remodeling of Tight Junctions and Enhancement of Barrier Integrity of the CACO-2 Intestinal Epithelial Cell Layer by Micronutrients.

    PubMed

    Valenzano, Mary Carmen; DiGuilio, Katherine; Mercado, Joanna; Teter, Mimi; To, Julie; Ferraro, Brendan; Mixson, Brittany; Manley, Isabel; Baker, Valerissa; Moore, Beverley A; Wertheimer, Joshua; Mullin, James M

    2015-01-01

    The micronutrients zinc, quercetin, butyrate, indole and berberine were evaluated for their ability to induce remodeling of epithelial tight junctions (TJs) and enhance barrier integrity in the CACO-2 gastrointestinal epithelial cell culture model. All five of these chemically very diverse micronutrients increased transepithelial electrical resistance (Rt) significantly, but only berberine also improved barrier integrity to the non-electrolyte D-mannitol. Increases of Rt as much as 200% of untreated controls were observed. Each of the five micronutrients also induced unique, signature-like changes in TJ protein composition, suggesting multiple pathways (and TJ arrangements) by which TJ barrier function can be enhanced. Decreases in abundance by as much as 90% were observed for claudin-2, and increases of over 300% could be seen for claudins -5 and -7. The exact effects of the micronutrients on barrier integrity and TJ protein composition were found to be highly dependent on the degree of differentiation of the cell layer at the time it was exposed to the micronutrient. The substratum to which the epithelial layer adheres was also found to regulate the response of the cell layer to the micronutrient. The implications of these findings for therapeutically decreasing morbidity in Inflammatory Bowel Disease are discussed.

  1. Remodeling of Tight Junctions and Enhancement of Barrier Integrity of the CACO-2 Intestinal Epithelial Cell Layer by Micronutrients

    PubMed Central

    Valenzano, Mary Carmen; DiGuilio, Katherine; Mercado, Joanna; Teter, Mimi; To, Julie; Ferraro, Brendan; Mixson, Brittany; Manley, Isabel; Baker, Valerissa; Moore, Beverley A.; Wertheimer, Joshua; Mullin, James M.

    2015-01-01

    The micronutrients zinc, quercetin, butyrate, indole and berberine were evaluated for their ability to induce remodeling of epithelial tight junctions (TJs) and enhance barrier integrity in the CACO-2 gastrointestinal epithelial cell culture model. All five of these chemically very diverse micronutrients increased transepithelial electrical resistance (Rt) significantly, but only berberine also improved barrier integrity to the non-electrolyte D-mannitol. Increases of Rt as much as 200% of untreated controls were observed. Each of the five micronutrients also induced unique, signature-like changes in TJ protein composition, suggesting multiple pathways (and TJ arrangements) by which TJ barrier function can be enhanced. Decreases in abundance by as much as 90% were observed for claudin-2, and increases of over 300% could be seen for claudins -5 and -7. The exact effects of the micronutrients on barrier integrity and TJ protein composition were found to be highly dependent on the degree of differentiation of the cell layer at the time it was exposed to the micronutrient. The substratum to which the epithelial layer adheres was also found to regulate the response of the cell layer to the micronutrient. The implications of these findings for therapeutically decreasing morbidity in Inflammatory Bowel Disease are discussed. PMID:26226276

  2. Cystic fibrosis transmembrane conductance regulator trafficking modulates the barrier function of airway epithelial cell monolayers.

    PubMed

    LeSimple, Pierre; Liao, Jie; Robert, Renaud; Gruenert, Dieter C; Hanrahan, John W

    2010-04-15

    The cystic fibrosis transmembrane conductance regulator (CFTR) is an integral membrane glycoprotein which functions as an anion channel and influences diverse cellular processes. We studied its role in the development of epithelial tightness by expressing wild-type (WT-CFTR) or mutant (Delta F508-CFTR) CFTR in human airway epithelial cell monolayers cultured at the air-liquid interface. Green fluorescent protein (GFP)-tagged WT or Delta F508 constructs were expressed in the CF bronchial cell line CFBE41o(-) using adenoviruses, and the results were compared with those obtained using CFBE41o(-) lines stably complemented with wild-type or mutant CFTR. As predicted, GFP-Delta WT-CFTR reached the apical membrane whereas GFP-F508-CFTR was only detected intracellularly. Although CFTR expression would be expected to reduce transepithelial resistance (TER), expressing GFP-CFTR significantly increased the TER of CFBE41o(-) monolayers whilst GFP-Delta F508-CFTR had no effect. Similar results were obtained with cell lines stably overexpressing Delta F508-CFTR or WT-CFTR. Preincubating Delta F508-CFTR monolayers at 29 degrees C reduced mannitol permeability and restored TER, and the effect on TER was reversible during temperature oscillations. Expression of GFP-Delta F508-CFTR or GFP-WT-CFTR in a cell line already containing endogenous WT-CFTR (Calu-3) did not alter TER. The CFTR- and temperature-dependence of TER were not affected by the CFTR inhibitor CFTR(inh)172 or low-chloride medium; therefore the effect of CFTR on barrier function was unrelated to its ion channel activity. Modulation of TER was blunted but not eliminated by genistein, implying the involvement of tyrosine phosphorylation and other mechanisms. Modulation of CFTR trafficking was correlated with an increase in tight junction depth. The results suggest that CFTR trafficking is required for the normal organisation and function of tight junctions. A reduction in barrier function caused by endoplasmic reticulum

  3. Immunoglobulin Responses at the Mucosal Interface

    PubMed Central

    Chen, Kang; Chorny, Alejo

    2011-01-01

    Mucosal surfaces are colonized by large communities of commensal bacteria and represent the primary site of entry for pathogenic agents. To prevent microbial intrusion, mucosal B cells release large amounts of immunoglobulin (Ig) molecules through multiple follicular and extrafollicular pathways. IgA is the most abundant antibody isotype in mucosal secretions and owes its success in frontline immunity to its ability to undergo transcytosis across epithelial cells. In addition to translocating IgA onto the mucosal surface, epithelial cells educate the mucosal immune system as to the composition of the local microbiota and instruct B cells to initiate IgA responses that generate immune protection while preserving immune homeostasis. Here we review recent advances in our understanding of the cellular interactions and signaling pathways governing IgA production at mucosal surfaces and discuss new findings on the regulation and function of mucosal IgD, the most enigmatic isotype of our mucosal antibody repertoire. PMID:21219173

  4. Desmoglein-1 regulates esophageal epithelial barrier function and immune responses in eosinophilic esophagitis

    PubMed Central

    Sherrill, J D; KC, K; Wu, D; Djukic, Z; Caldwell, J M; Stucke, E M; Kemme, K A; Costello, M S; Mingler, M K; Blanchard, C; Collins, M H; Abonia, J P; Putnam, P E; Dellon, E S; Orlando, R C; Hogan, S P; Rothenb, M E

    2014-01-01

    The desmosomal cadherin desmoglein-1 (DSG1) is an essential intercellular adhesion molecule that is altered in various human cutaneous disorders; however, its regulation and function in allergic disease remains unexplored. Herein, we demonstrate a specific reduction in DSG1 in esophageal biopsies from patients with eosinophilic esophagitis (EoE), an emerging allergic disorder characterized by chronic inflammation within the esophageal mucosa. Further, we show that DSG1 gene silencing weakens esophageal epithelial integrity, and induces cell separation and impaired barrier function (IBF) despite high levels of desmoglein-3 (DSG3). Moreover, DSG1 deficiency induces transcriptional changes that partially overlap with the transcriptome of inflamed esophageal mucosa; notably, periostin, a multipotent pro-inflammatory extracellular matrix molecule, is the top induced overlapping gene. We further demonstrate that IBF is a pathological feature in EoE, which can be partially induced through the downregulation of DSG1 by interleukin-13 (IL-13). Taken together, these data identify a functional role for DSG1 and its dysregulation by IL-13 in the pathophysiology of EoE and suggest that the loss of DSG1 may potentiate allergic inflammation through the induction of pro-inflammatory mediators such as periostin. PMID:24220297

  5. Desmoglein-1 regulates esophageal epithelial barrier function and immune responses in eosinophilic esophagitis.

    PubMed

    Sherrill, J D; Kc, K; Wu, D; Djukic, Z; Caldwell, J M; Stucke, E M; Kemme, K A; Costello, M S; Mingler, M K; Blanchard, C; Collins, M H; Abonia, J P; Putnam, P E; Dellon, E S; Orlando, R C; Hogan, S P; Rothenberg, M E

    2014-05-01

    The desmosomal cadherin desmoglein-1 (DSG1) is an essential intercellular adhesion molecule that is altered in various human cutaneous disorders; however, its regulation and function in allergic disease remains unexplored. Herein, we demonstrate a specific reduction in DSG1 in esophageal biopsies from patients with eosinophilic esophagitis (EoE), an emerging allergic disorder characterized by chronic inflammation within the esophageal mucosa. Further, we show that DSG1 gene silencing weakens esophageal epithelial integrity, and induces cell separation and impaired barrier function (IBF) despite high levels of desmoglein-3. Moreover, DSG1 deficiency induces transcriptional changes that partially overlap with the transcriptome of inflamed esophageal mucosa; notably, periostin (POSTN), a multipotent pro-inflammatory extracellular matrix molecule, is the top induced overlapping gene. We further demonstrate that IBF is a pathological feature in EoE, which can be partially induced through the downregulation of DSG1 by interleukin-13 (IL-13). Taken together, these data identify a functional role for DSG1 and its dysregulation by IL-13 in the pathophysiology of EoE and suggest that the loss of DSG1 may potentiate allergic inflammation through the induction of pro-inflammatory mediators such as POSTN. PMID:24220297

  6. Acid modulates the squamous epithelial barrier function by modulating the localization of claudins in the superficial layers.

    PubMed

    Oshima, Tadayuki; Koseki, Junichi; Chen, Xin; Matsumoto, Takayuki; Miwa, Hiroto

    2012-01-01

    Acid is a major cause of gastro-esophageal reflux disease. However, the influence of acid on the esophageal stratified epithelial barrier function and tight junction (TJ) proteins is not fully understood. Here, we explore the influence of acid on barrier function and TJ proteins using a newly developed model of the esophageal-like squamous epithelial cell layers that employs an air-liquid interface (ALI) system. Barrier function was determined by measuring trans-epithelial electrical resistance (TEER) and diffusion of paracellular tracers. TJ-related protein (claudin-1, claudin-4, occludin and ZO-1) expression and localization was examined by immunofluorescent staining, and by western blotting of 1% NP-40 soluble and insoluble fractions. We also examined the influence of acid (pH 2-4) on the barrier created by these cells. The in vitro ALI culture system showed a tight barrier (1500-2500 Ω·cm(2)) with the expression of claudin-1, claudin-4, occludin and ZO-1 in the superficial layers. Claudin-1, claudin-4, occludin and ZO-1 were detected as dots and whisker-like lines in the superficial layers, and as a broad line in the suprabasal layers. These localization patterns are similar to those in the human esophagus. On day 7 under ALI culture, TJ proteins were detected in the superficial layers with functional properties, including decreased permeability and increased TEER. Dilated intercellular spaces were detected at the suprabasal cell layers even under the control conditions of ALI cells. pH 2 acid on the apical side significantly reduced the TEER in ALI-cultured cells. This decrease in TEER by the acid was in parallel with the decreased amount of detergent-insoluble claudin-4. Claudin-4 delocalization was confirmed by immunofluorescent staining. In conclusion, TJs are located in the superficial layers of the esophagus, and acid stimulation disrupts barrier function, at least in part by modulating the amount and localization of claudin-4 in the superficial layers.

  7. Lactobacillus rhamnosus strain GG prevents enterohemorrhagic Escherichia coli O157:H7-induced changes in epithelial barrier function.

    PubMed

    Johnson-Henry, K C; Donato, K A; Shen-Tu, G; Gordanpour, M; Sherman, P M

    2008-04-01

    Enterohemorrhagic Escherichia coli (EHEC) O157:H7 intimately attaches to intestinal epithelial monolayers and produces attaching and effacing (A/E) lesions. In addition, EHEC infection causes disruptions of intercellular tight junctions, leading to clinical sequelae that include acute diarrhea, hemorrhagic colitis, and the hemolytic-uremic syndrome. Current therapy remains supportive since antibiotic therapy increases the risk of systemic complications. This study focused on the potential therapeutic effect of an alternative form of therapy, probiotic Lactobacillus rhamnosus strain GG, to attenuate EHEC-induced changes in paracellular permeability in polarized MDCK-I and T84 epithelial cell monolayers. Changes in epithelial cell morphology, electrical resistance, dextran permeability, and distribution and expression of claudin-1 and ZO-1 were assessed using phase-contrast, immunofluorescence, and transmission electron microscopy and macromolecular flux. This study demonstrated that pretreatment of polarized MDCK-I and T84 cells with the probiotic L. rhamnosus GG reduced morphological changes and diminished the number of A/E lesions induced in response to EHEC O157:H7 infection. With probiotic pretreatment there was corresponding attenuation of the EHEC-induced drop in electrical resistance and the increase in barrier permeability assays. In addition, L. rhamnosus GG protected epithelial monolayers against EHEC-induced redistribution of the claudin-1 and ZO-1 tight junction proteins. In contrast to the effects seen with the live probiotic, heat-inactivated L. rhamnosus GG had no effect on EHEC binding and A/E lesion formation or on disruption of the barrier function. Collectively, these findings provide in vitro evidence that treatment with the probiotic L. rhamnosus strain GG could prove to be an effective management treatment for preventing injury of the epithelial cell barrier induced by A/E bacterial enteropathogens.

  8. Glycogen Synthase Kinase 3 (GSK-3) influences epithelial barrier function by regulating Occludin, Claudin-1 and E-cadherin expression

    SciTech Connect

    Severson, Eric A.; Kwon, Mike; Hilgarth, Roland S.; Parkos, Charles A.; Nusrat, Asma

    2010-07-02

    The Apical Junctional Complex (AJC) encompassing the tight junction (TJ) and adherens junction (AJ) plays a pivotal role in regulating epithelial barrier function and epithelial cell proliferative processes through signaling events that remain poorly characterized. A potential regulator of AJC protein expression is Glycogen Synthase Kinase-3 (GSK-3). GSK-3 is a constitutively active kinase that is repressed during epithelial-mesenchymal transition (EMT). In the present study, we report that GSK-3 activity regulates the structure and function of the AJC in polarized model intestinal (SK-CO15) and kidney (Madin-Darby Canine Kidney (MDCK)) epithelial cells. Reduction of GSK-3 activity, either by small molecule inhibitors or siRNA targeting GSK-3 alpha and beta mRNA, resulted in increased permeability to both ions and bulk solutes. Immunofluorescence labeling and immunoblot analyses revealed that the barrier defects correlated with decreased protein expression of AJC transmembrane proteins Occludin, Claudin-1 and E-cadherin without influencing other TJ proteins, Zonula Occludens-1 (ZO-1) and Junctional Adhesion Molecule A (JAM-A). The decrease in Occludin and E-cadherin protein expression correlated with downregulation of the corresponding mRNA levels for these respective proteins following GSK-3 inhibition. These observations implicate an important role of GSK-3 in the regulation of the structure and function of the AJC that is mediated by differential modulation of mRNA transcription of key AJC proteins, Occludin, Claudin-1 and E-cadherin.

  9. A new role for reticulon-4B/NOGO-B in the intestinal epithelial barrier function and inflammatory bowel disease.

    PubMed

    Rodríguez-Feo, Juan Antonio; Puerto, Marta; Fernández-Mena, Carolina; Verdejo, Cristina; Lara, José Manuel; Díaz-Sánchez, María; Álvarez, Emilio; Vaquero, Javier; Marín-Jiménez, Ignacio; Bañares, Rafael; Menchén, Luis

    2015-06-15

    Inflammatory bowel disease (IBD) is characterized by an impaired intestinal barrier function. We aimed to investigate the role of reticulon-4B (RTN-4B/NOGO-B), a structural protein of the endoplasmic reticulum, in intestinal barrier function and IBD. We used immunohistochemistry, confocal microscopy, real-time PCR, and Western blotting to study tissue distribution and expression levels of RTN-4B/NOGO-B in control and IBD samples from mouse and humans. We also targeted RTN-4B/NOGO-B using siRNAs in cultured human intestinal epithelial cell (IECs). Epithelial barrier permeability was assessed by transepithelial electrical resistance (TEER) measurement. RTN-4B/NOGO-B is expressed in the intestine mainly by IECs. Confocal microscopy revealed a colocalization of RTN-4B, E-cadherin, and polymerized actin fibers in tissue and cultured IECs. RTN-4B mRNA and protein expression were lower in the colon of IL-10(-/-) compared with wild-type mice. Colocalization of RTN-4B/E-cadherin/actin was reduced in the colon of IL-10(-/-) mice. Analysis of endoscopic biopsies from IBD patients showed a significant reduction of RTN-4B/NOGO-B expression in inflamed mucosa compared with control. Treatment of IECs with H2O2 reduced TEER values and triggered phosphorylation of RTN-4B in serine 107 residues as well as downregulation of RTN-4B expression. Acute RTN-4B/NOGO-B knockdown by siRNAs resulted in a decreased TEER values and reduction of E-cadherin and α-catenin expression and in the amount of F-actin-rich filaments in IECs. Epithelial RTN-4B/NOGO-B was downregulated in human and experimental IBD. RTN-4B participates in the intestinal epithelial barrier function, most likely via its involvement in E-cadherin, α-catenin expression, and actin cytoskeleton organization at sites of cell-to-cell contacts.

  10. Rhubarb extract partially improves mucosal integrity in chemotherapy-induced intestinal mucositis

    PubMed Central

    Bajic, Juliana E; Eden, Georgina L; Lampton, Lorrinne S; Cheah, Ker Y; Lymn, Kerry A; Pei, Jinxin V; Yool, Andrea J; Howarth, Gordon S

    2016-01-01

    AIM To investigate the effects of orally gavaged aqueous rhubarb extract (RE) on 5-fluorouracil (5-FU)-induced intestinal mucositis in rats. METHODS Female Dark Agouti rats (n = 8/group) were gavaged daily (1 mL) with water, high-dose RE (HDR; 200 mg/kg) or low-dose RE (LDR; 20mg/kg) for eight days. Intestinal mucositis was induced (day 5) with 5-FU (150 mg/kg) via intraperitoneal injection. Intestinal tissue samples were collected for myeloperoxidase (MPO) activity and histological examination. Xenopus oocytes expressing aquaporin 4 water channels were prepared to examine the effect of aqueous RE on cell volume, indicating a potential mechanism responsible for modulating net fluid absorption and secretion in the gastrointestinal tract. Statistical significance was assumed at P < 0.05 by one-way ANOVA. RESULTS Bodyweight was significantly reduced in rats administered 5-FU compared to healthy controls (P < 0.01). Rats administered 5-FU significantly increased intestinal MPO levels (≥ 307%; P < 0.001), compared to healthy controls. However, LDR attenuated this effect in 5-FU treated rats, significantly decreasing ileal MPO activity (by 45%; P < 0.05), as compared to 5-FU controls. 5-FU significantly reduced intestinal mucosal thickness (by ≥ 29% P < 0.001) as compared to healthy controls. LDR significantly increased ileal mucosal thickness in 5-FU treated rats (19%; P < 0.05) relative to 5-FU controls. In xenopus oocytes expressing AQP4 water channels, RE selectively blocked water influx into the cell, induced by a decrease in external osmotic pressure. As water efflux was unaltered by the presence of extracellular RE, the directional flow of water across the epithelial barrier, in the presence of extracellular RE, indicated that RE may alleviate water loss across the epithelial barrier and promote intestinal health in chemotherapy-induced intestinal mucositis. CONCLUSION In summary, low dose RE improves selected parameters of mucosal integrity and reduces ileal

  11. Modulation of the epithelial barrier by dexamethasone and prolactin in cultured Madin-Darby canine kidney (MDCK) cells.

    PubMed

    Peixoto, E B M I; Collares-Buzato, C B

    2006-02-01

    Glucocorticoids and prolactin (PRL) have a direct effect on the formation and maintenance of tight junctions (TJs) in cultured endothelial and mammary gland epithelial cells. In this work, we investigated the effect of a synthetic glucocorticoid dexamethasone (DEX) and PRL on the paracellular barrier function in MDCK renal epithelial cells. DEX (4 microM)+PRL (2 microg/ml) and DEX alone increased significantly the transepithelial electrical resistance after chronic treatment (4 days) of confluent MDCK monolayers or after 24 h treatment of subconfluent monolayers. Immunoblotting and immunocytochemistry revealed no changes in the expression and distribution of TJ-associated proteins occludin, ZO-1 and claudin-1 in confluent monolayers after hormone addition. However, a marked increase in junctional content for occludin and ZO-1 with no changes in their total expression was observed in subconfluent MDCK monolayers 24 h exposed to DEX or DEX+PRL. No change in cell proliferation/growth was detected at subconfluent conditions following hormone treatment. An increase in the total number of viable cells was observed only in confluent MDCK monolayers after exposure to DEX+PRL suggesting that the main effect of these hormones on already established barrier may be associated with the inhibition of cell death. In conclusion, our data suggest that these hormones (specially dexamethasone) have an effect on TJ structure and function only during the formation of MDCK epithelial barrier by probably modulating the localization, stability or assembly of TJ proteins to membrane sites of intercellular contact.

  12. Gut epithelial barrier dysfunction in human immunodeficiency virus-hepatitis C virus coinfected patients: Influence on innate and acquired immunity

    PubMed Central

    Márquez, Mercedes; Fernández Gutiérrez del Álamo, Clotilde; Girón-González, José Antonio

    2016-01-01

    Even in cases where viral replication has been controlled by antiretroviral therapy for long periods of time, human immunodeficiency virus (HIV)-infected patients have several non-acquired immunodeficiency syndrome (AIDS) related co-morbidities, including liver disease, cardiovascular disease and neurocognitive decline, which have a clear impact on survival. It has been considered that persistent innate and acquired immune activation contributes to the pathogenesis of these non-AIDS related diseases. Immune activation has been related with several conditions, remarkably with the bacterial translocation related with the intestinal barrier damage by the HIV or by hepatitis C virus (HCV)-related liver cirrhosis. Consequently, increased morbidity and mortality must be expected in HIV-HCV coinfected patients. Disrupted gut barrier lead to an increased passage of microbial products and to an activation of the mucosal immune system and secretion of inflammatory mediators, which in turn might increase barrier dysfunction. In the present review, the intestinal barrier structure, measures of intestinal barrier dysfunction and the modifications of them in HIV monoinfection and in HIV-HCV coinfection will be considered. Both pathogenesis and the consequences for the progression of liver disease secondary to gut microbial fragment leakage and immune activation will be assessed. PMID:26819512

  13. Dual Function of Novel Pollen Coat (Surface) Proteins: IgE-binding Capacity and Proteolytic Activity Disrupting the Airway Epithelial Barrier

    PubMed Central

    Bashir, Mohamed Elfatih H.; Ward, Jason M.; Cummings, Matthew; Karrar, Eltayeb E.; Root, Michael; Mohamed, Abu Bekr A.; Naclerio, Robert M.; Preuss, Daphne

    2013-01-01

    Background The pollen coat is the first structure of the pollen to encounter the mucosal immune system upon inhalation. Prior characterizations of pollen allergens have focused on water-soluble, cytoplasmic proteins, but have overlooked much of the extracellular pollen coat. Due to washing with organic solvents when prepared, these pollen coat proteins are typically absent from commercial standardized allergenic extracts (i.e., “de-fatted”), and, as a result, their involvement in allergy has not been explored. Methodology/Principal Findings Using a unique approach to search for pollen allergenic proteins residing in the pollen coat, we employed transmission electron microscopy (TEM) to assess the impact of organic solvents on the structural integrity of the pollen coat. TEM results indicated that de-fatting of Cynodon dactylon (Bermuda grass) pollen (BGP) by use of organic solvents altered the structural integrity of the pollen coat. The novel IgE-binding proteins of the BGP coat include a cysteine protease (CP) and endoxylanase (EXY). The full-length cDNA that encodes the novel IgE-reactive CP was cloned from floral RNA. The EXY and CP were purified to homogeneity and tested for IgE reactivity. The CP from the BGP coat increased the permeability of human airway epithelial cells, caused a clear concentration-dependent detachment of cells, and damaged their barrier integrity. Conclusions/Significance Using an immunoproteomics approach, novel allergenic proteins of the BGP coat were identified. These proteins represent a class of novel dual-function proteins residing on the coat of the pollen grain that have IgE-binding capacity and proteolytic activity, which disrupts the integrity of the airway epithelial barrier. The identification of pollen coat allergens might explain the IgE-negative response to available skin-prick-testing proteins in patients who have positive symptoms. Further study of the role of these pollen coat proteins in allergic responses is

  14. H19 Long Noncoding RNA Regulates Intestinal Epithelial Barrier Function via MicroRNA 675 by Interacting with RNA-Binding Protein HuR.

    PubMed

    Zou, Tongtong; Jaladanki, Suraj K; Liu, Lan; Xiao, Lan; Chung, Hee Kyoung; Wang, Jun-Yao; Xu, Yan; Gorospe, Myriam; Wang, Jian-Ying

    2016-05-01

    The disruption of the intestinal epithelial barrier function occurs commonly in various pathologies, but the exact mechanisms responsible are unclear. The H19 long noncoding RNA (lncRNA) regulates the expression of different genes and has been implicated in human genetic disorders and cancer. Here, we report that H19 plays an important role in controlling the intestinal epithelial barrier function by serving as a precursor for microRNA 675 (miR-675). H19 overexpression increased the cellular abundance of miR-675, which in turn destabilized and repressed the translation of mRNAs encoding tight junction protein ZO-1 and adherens junction E-cadherin, resulting in the dysfunction of the epithelial barrier. Increasing the level of the RNA-binding protein HuR in cells overexpressing H19 prevented the stimulation of miR-675 processing from H19, promoted ZO-1 and E-cadherin expression, and restored the epithelial barrier function to a nearly normal level. In contrast, the targeted deletion of HuR in intestinal epithelial cells enhanced miR-675 production in the mucosa and delayed the recovery of the gut barrier function after exposure to mesenteric ischemia/reperfusion. These results indicate that H19 interacts with HuR and regulates the intestinal epithelial barrier function via the H19-encoded miR-675 by altering ZO-1 and E-cadherin expression posttranscriptionally.

  15. H19 Long Noncoding RNA Regulates Intestinal Epithelial Barrier Function via MicroRNA 675 by Interacting with RNA-Binding Protein HuR

    PubMed Central

    Zou, Tongtong; Jaladanki, Suraj K.; Liu, Lan; Xiao, Lan; Chung, Hee Kyoung; Wang, Jun-Yao; Xu, Yan; Gorospe, Myriam

    2016-01-01

    The disruption of the intestinal epithelial barrier function occurs commonly in various pathologies, but the exact mechanisms responsible are unclear. The H19 long noncoding RNA (lncRNA) regulates the expression of different genes and has been implicated in human genetic disorders and cancer. Here, we report that H19 plays an important role in controlling the intestinal epithelial barrier function by serving as a precursor for microRNA 675 (miR-675). H19 overexpression increased the cellular abundance of miR-675, which in turn destabilized and repressed the translation of mRNAs encoding tight junction protein ZO-1 and adherens junction E-cadherin, resulting in the dysfunction of the epithelial barrier. Increasing the level of the RNA-binding protein HuR in cells overexpressing H19 prevented the stimulation of miR-675 processing from H19, promoted ZO-1 and E-cadherin expression, and restored the epithelial barrier function to a nearly normal level. In contrast, the targeted deletion of HuR in intestinal epithelial cells enhanced miR-675 production in the mucosa and delayed the recovery of the gut barrier function after exposure to mesenteric ischemia/reperfusion. These results indicate that H19 interacts with HuR and regulates the intestinal epithelial barrier function via the H19-encoded miR-675 by altering ZO-1 and E-cadherin expression posttranscriptionally. PMID:26884465

  16. Non-animal models of epithelial barriers (skin, intestine and lung) in research, industrial applications and regulatory toxicology.

    PubMed

    Gordon, Sarah; Daneshian, Mardas; Bouwstra, Joke; Caloni, Francesca; Constant, Samuel; Davies, Donna E; Dandekar, Gudrun; Guzman, Carlos A; Fabian, Eric; Haltner, Eleonore; Hartung, Thomas; Hasiwa, Nina; Hayden, Patrick; Kandarova, Helena; Khare, Sangeeta; Krug, Harald F; Kneuer, Carsten; Leist, Marcel; Lian, Guoping; Marx, Uwe; Metzger, Marco; Ott, Katharina; Prieto, Pilar; Roberts, Michael S; Roggen, Erwin L; Tralau, Tewes; van den Braak, Claudia; Walles, Heike; Lehr, Claus-Michael

    2015-01-01

    Models of the outer epithelia of the human body - namely the skin, the intestine and the lung - have found valid applications in both research and industrial settings as attractive alternatives to animal testing. A variety of approaches to model these barriers are currently employed in such fields, ranging from the utilization of ex vivo tissue to reconstructed in vitro models, and further to chip-based technologies, synthetic membrane systems and, of increasing current interest, in silico modeling approaches. An international group of experts in the field of epithelial barriers was convened from academia, industry and regulatory bodies to present both the current state of the art of non-animal models of the skin, intestinal and pulmonary barriers in their various fields of application, and to discuss research-based, industry-driven and regulatory-relevant future directions for both the development of new models and the refinement of existing test methods. Issues of model relevance and preference, validation and standardization, acceptance, and the need for simplicity versus complexity were focal themes of the discussions. The outcomes of workshop presentations and discussions, in relation to both current status and future directions in the utilization and development of epithelial barrier models, are presented by the attending experts in the current report.

  17. High therapeutic efficacy of Cathelicidin-WA against postweaning diarrhea via inhibiting inflammation and enhancing epithelial barrier in the intestine

    PubMed Central

    Yi, Hongbo; Zhang, Lin; Gan, Zhenshun; Xiong, Haitao; Yu, Caihua; Du, Huahua; Wang, Yizhen

    2016-01-01

    Diarrhea is a leading cause of death among young mammals, especially during weaning. Here, we investigated the effects of Cathelicidin-WA (CWA) on diarrhea, intestinal morphology, inflammatory responses, epithelial barrier and microbiota in the intestine of young mammals during weaning. Piglets with clinical diarrhea were selected and treated with saline (control), CWA or enrofloxacin (Enro) for 4 days. Both CWA and Enro effectively attenuated diarrhea. Compared with the control, CWA decreased IL-6, IL-8 and IL-22 levels and reduced neutrophil infiltration into the jejunum. CWA inhibited inflammation by down-regulating the TLR4-, MyD88- and NF-κB-dependent pathways. Additionally, CWA improved intestinal morphology by increasing villus and microvillus heights and enhancing intestinal barrier function by increasing tight junction (TJ) protein expression and augmenting wound-healing ability in intestinal epithelial cells. CWA also improved microbiota composition and increased short-chain fatty acid (SCFA) levels in feces. By contrast, Enro not only disrupted the intestinal barrier but also negatively affected microbiota composition and SCFA levels in the intestine. In conclusion, CWA effectively attenuated inflammation, enhanced intestinal barrier function, and improved microbiota composition in the intestines of weaned piglets. These results suggest that CWA could be an effective and safe therapy for diarrhea or other intestinal diseases in young mammals. PMID:27181680

  18. Non-animal models of epithelial barriers (skin, intestine and lung) in research, industrial applications and regulatory toxicology.

    PubMed

    Gordon, Sarah; Daneshian, Mardas; Bouwstra, Joke; Caloni, Francesca; Constant, Samuel; Davies, Donna E; Dandekar, Gudrun; Guzman, Carlos A; Fabian, Eric; Haltner, Eleonore; Hartung, Thomas; Hasiwa, Nina; Hayden, Patrick; Kandarova, Helena; Khare, Sangeeta; Krug, Harald F; Kneuer, Carsten; Leist, Marcel; Lian, Guoping; Marx, Uwe; Metzger, Marco; Ott, Katharina; Prieto, Pilar; Roberts, Michael S; Roggen, Erwin L; Tralau, Tewes; van den Braak, Claudia; Walles, Heike; Lehr, Claus-Michael

    2015-01-01

    Models of the outer epithelia of the human body - namely the skin, the intestine and the lung - have found valid applications in both research and industrial settings as attractive alternatives to animal testing. A variety of approaches to model these barriers are currently employed in such fields, ranging from the utilization of ex vivo tissue to reconstructed in vitro models, and further to chip-based technologies, synthetic membrane systems and, of increasing current interest, in silico modeling approaches. An international group of experts in the field of epithelial barriers was convened from academia, industry and regulatory bodies to present both the current state of the art of non-animal models of the skin, intestinal and pulmonary barriers in their various fields of application, and to discuss research-based, industry-driven and regulatory-relevant future directions for both the development of new models and the refinement of existing test methods. Issues of model relevance and preference, validation and standardization, acceptance, and the need for simplicity versus complexity were focal themes of the discussions. The outcomes of workshop presentations and discussions, in relation to both current status and future directions in the utilization and development of epithelial barrier models, are presented by the attending experts in the current report. PMID:26536291

  19. Protective Effects of Ferulic Acid against Heat Stress-Induced Intestinal Epithelial Barrier Dysfunction In Vitro and In Vivo

    PubMed Central

    He, Shasha; Liu, Fenghua; Xu, Lei; Yin, Peng; Li, Deyin; Mei, Chen; Jiang, Linshu; Ma, Yunfei; Xu, Jianqin

    2016-01-01

    Heat stress is important in the pathogenesis of intestinal epithelial barrier dysfunction. Ferulic acid (FA), a phenolic acid widely found in fruits and vegetables, can scavenge free radicals and activate cell stress responses. This study is aimed at investigating protective effects of FA on heat stress-induced dysfunction of the intestinal epithelial barrier in vitro and in vivo. Intestinal epithelial (IEC-6) cells were pretreated with FA for 4 h and then exposed to heat stress. Heat stress caused decreased transepithelial electrical resistance (TER) and increased permeability to 4-kDa fluorescein isothiocyanate (FITC)-dextran (FD4). Both effects were inhibited by FA in a dose-dependent manner. FA significantly attenuated the decrease in occludin, ZO-1 and E-cadherin expression observed with heat stress. The distortion and redistribution of occludin, ZO-1 and E-cadherin proteins were also effectively prevented by FA pretreatment. Moreover, heat stress diminished electron-dense material detected in tight junctions (TJs), an effect also alleviated by FA in a dose-dependent manner. In an in vivo heat stress model, FA (50 mg/kg) was administered to male Sprague–Dawley rats for 7 consecutive days prior to exposure to heat stress. FA pretreatment significantly attenuated the effects of heat stress on the small intestine, including the increased FD4 permeability, disrupted tight junctions and microvilli structure, and reduced occludin, ZO-1 and E-cadherin expression. Taken together, our results demonstrate that FA pretreatment is potentially protective against heat stress-induced intestinal epithelial barrier dysfunction. PMID:26894689

  20. Protective Effects of Ferulic Acid against Heat Stress-Induced Intestinal Epithelial Barrier Dysfunction In Vitro and In Vivo.

    PubMed

    He, Shasha; Liu, Fenghua; Xu, Lei; Yin, Peng; Li, Deyin; Mei, Chen; Jiang, Linshu; Ma, Yunfei; Xu, Jianqin

    2016-01-01

    Heat stress is important in the pathogenesis of intestinal epithelial barrier dysfunction. Ferulic acid (FA), a phenolic acid widely found in fruits and vegetables, can scavenge free radicals and activate cell stress responses. This study is aimed at investigating protective effects of FA on heat stress-induced dysfunction of the intestinal epithelial barrier in vitro and in vivo. Intestinal epithelial (IEC-6) cells were pretreated with FA for 4 h and then exposed to heat stress. Heat stress caused decreased transepithelial electrical resistance (TER) and increased permeability to 4-kDa fluorescein isothiocyanate (FITC)-dextran (FD4). Both effects were inhibited by FA in a dose-dependent manner. FA significantly attenuated the decrease in occludin, ZO-1 and E-cadherin expression observed with heat stress. The distortion and redistribution of occludin, ZO-1 and E-cadherin proteins were also effectively prevented by FA pretreatment. Moreover, heat stress diminished electron-dense material detected in tight junctions (TJs), an effect also alleviated by FA in a dose-dependent manner. In an in vivo heat stress model, FA (50 mg/kg) was administered to male Sprague-Dawley rats for 7 consecutive days prior to exposure to heat stress. FA pretreatment significantly attenuated the effects of heat stress on the small intestine, including the increased FD4 permeability, disrupted tight junctions and microvilli structure, and reduced occludin, ZO-1 and E-cadherin expression. Taken together, our results demonstrate that FA pretreatment is potentially protective against heat stress-induced intestinal epithelial barrier dysfunction. PMID:26894689

  1. Infectious salmon anaemia virus (ISAV) mucosal infection in Atlantic salmon.

    PubMed

    Aamelfot, Maria; McBeath, Alastair; Christiansen, Debes H; Matejusova, Iveta; Falk, Knut

    2015-01-01

    All viruses infecting fish must cross the surface mucosal barrier to successfully enter a host. Infectious salmon anaemia virus (ISAV), the causative agent of the economically important infectious salmon anaemia (ISA) in Atlantic salmon, Salmo salar L., has been shown to use the gills as its entry point. However, other entry ports have not been investigated despite the expression of virus receptors on the surface of epithelial cells in the skin, the gastrointestinal (GI) tract and the conjunctiva. Here we investigate the ISAV mucosal infection in Atlantic salmon after experimental immersion (bath) challenge and in farmed fish collected from a confirmed outbreak of ISA in Norway. We show for the first time evidence of early replication in several mucosal surfaces in addition to the gills, including the pectoral fin, skin and GI tract suggesting several potential entry points for the virus. Initially, the infection is localized and primarily infecting epithelial cells, however at later stages it becomes systemic, infecting the endothelial cells lining the circulatory system. Viruses of low and high virulence used in the challenge revealed possible variation in virus progression during infection at the mucosal surfaces. PMID:26490835

  2. Effects of early enteral nutrition on the gastrointestinal motility and intestinal mucosal barrier of patients with burn-induced invasive fungal infection

    PubMed Central

    Zhang, Yu; Gu, Fang; Wang, Fengxian; Zhang, Yuanda

    2016-01-01

    Objective: To evaluate the effects of early enteral nutrition on the gastrointestinal motility and intestinal mucosal barrier of patients with burn-induced invasive fungal infection. Methods: A total of 120 patients with burn-induced invasive fungal infection were randomly divided into an early enteral nutrition (EN) group and a parenteral nutrition (PN) group (n=60). The patients were given nutritional support intervention for 14 days, and the expression levels of serum transferrin, albumin, total protein, endotoxin, D-lactic acid and inflammatory cytokines were detected on the 1st, 7th and 14th days respectively. Results: As the treatment progressed, the levels of serum transferrin, albumin and total protein of the EN group were significantly higher than those of the PN group (P<0.05), while the levels of serum endotoxin and D-lactic acid of the form group were significantly lower (P<0.05). After treatment, the expression levels of IL-6 and TNF-α were decreased in the EN group, which were significantly different from those of the PN group (P<0.05). During treatment, the incidence rates of complications such as abdominal distension, diarrhea, sepsis, nausea, vomiting and gastric retention were similar. The mean healing time of wound surface was 9.34±0.78 days in the EN group and 12.46±2.19 days in the PN group, i.e. such time of the former was significantly shorter than that of the latter (P<0.05). Conclusion: Treating patients having burn-induced invasive fungal infection by early enteral nutrition support with arginine can safely alleviate malnutrition and stress reaction, strengthen cellular immune function and promote wound healing, thereby facilitating the recovery of gastrointestinal motility and the function of intestinal mucosal barrier. PMID:27375697

  3. Role of Oral Mucosal Fluid and Electrolyte Absorption and Secretion in Dry Mouth.

    PubMed

    Zhang, Guo H; Castro, Robert

    2015-09-01

    Dry mouth is induced by dehydration of the oral mucosa, resulting from an imbalance of fluid supply and clearance within the oral cavity. Saliva is the major source of oral mucosal fluid, whereas oral fluid clearance includes evaporation and swallowing. Oral mucosal fluid absorption has been suggested to play a critical role in oral fluid clearance; over-absorption of water and ions across the oral mucosa under certain conditions may be a major component for oral fluid imbalance, leading to mucosal dehydration. While numerous studies have confirmed that the oral mucosa absorbs fluid and electrolytes, the pathways and mechanisms mediating the absorption remain undefined. The transcellular pathway regulating oral mucosal epithelial absorption includes aquaporins, epithelial Na+ channel and/or Na+/H+ exchanger, whereas the paracellular transport is likely to be mediated by tight junctions. The regulatory mechanisms of these pathways require further elucidation. It remains unclear whether the oral mucosa also secretes fluid and ions into the oral cavity. Although intercellular lipids secreted by epithelial cells form the major barrier to paracellular water and ion transport, the role and regulation of these lipids in oral mucosal hydration in physiological and pathological conditions need further investigation. Delineation of these mechanisms will be conducive to the development of preventive and therapeutic interventions for dry mouth. PMID:26485506

  4. Retinoic acid and hydrocortisone strengthen the barrier function of human RPMI 2650 cells, a model for nasal epithelial permeability.

    PubMed

    Kürti, Levente; Veszelka, Szilvia; Bocsik, Alexandra; Ozsvári, Béla; Puskás, László G; Kittel, Agnes; Szabó-Révész, Piroska; Deli, Mária A

    2013-05-01

    The nasal pathway represents an alternative route for non-invasive systemic administration of drugs. The main advantages of nasal drug delivery are the rapid onset of action, the avoidance of the first-pass metabolism in the liver and the easy applicability. In vitro cell culture systems offer an opportunity to model biological barriers. Our aim was to develop and characterize an in vitro model based on confluent layers of the human RPMI 2650 cell line. Retinoic acid, hydrocortisone and cyclic adenosine monophosphate, which influence cell attachment, growth and differentiation have been investigated on the barrier formation and function of the nasal epithelial cell layers. Real-time cell microelectronic sensing, a novel label-free technique was used for dynamic monitoring of cell growth and barrier properties of RPMI 2650 cells. Treatments enhanced the formation of adherens and tight intercellular junctions visualized by electron microscopy, the presence and localization of junctional proteins ZO-1 and β-catenin demonstrated by fluorescent immunohistochemistry, and the barrier function of nasal epithelial cell layers. The transepithelial resistance of the RPMI 2650 cell model reached 50 to 200 Ω × cm(2), the permeability coefficient for 4.4 kDa FITC-dextran was 9.3 to 17 × 10(-6) cm/s, in agreement with values measured on nasal mucosa from in vivo and ex vivo experiments. Based on these results human RPMI 2650 cells seem to be a suitable nasal epithelial model to test different pharmaceutical excipients and various novel formulations, such as nanoparticles for toxicity and permeability.

  5. Nanomaterial interactions with and trafficking across the lung alveolar epithelial barrier: implications for health effects of air-pollution particles

    PubMed Central

    Yacobi, Nazanin R.; Fazllolahi, Farnoosh; Kim, Yong Ho; Sipos, Arnold; Borok, Zea; Kim, Kwang-Jin

    2014-01-01

    Studies on the health effects of air-pollution particles suggest that injury may result from inhalation of airborne ultrafine particles (<100 nm in diameter). Engineered nanomaterials (<100 nm in at least one dimension) may also be harmful if inhaled. Nanomaterials deposited on the respiratory epithelial tract are thought to cross the air-blood barrier, especially via the expansive alveolar region, into the systemic circulation to reach end organs (e.g., myocardium, liver, pancreas, kidney, and spleen). Since ambient ultrafine particles are difficult to track, studies of defined engineered nanomaterials have been used to obtain valuable information on how nanomaterials interact with and traffic across the air-blood barrier of mammalian lungs. Since specific mechanistic information on how nanomaterials interact with the lung is difficult to obtain using in vivo or ex vivo lungs due to their complex anatomy, in vitro alveolar epithelial models have been of considerable value in determining nanomaterial-lung interactions. In this review, we provide information on mechanisms underlying lung alveolar epithelial injury caused by various nanomaterials and on nanomaterial trafficking across alveolar epithelium that may lead to end-organ injury. PMID:25568662

  6. 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

  7. Stepwise DNA Methylation Changes Are Linked to Escape from Defined Proliferation Barriers and Mammary Epithelial Cell Immortalization

    SciTech Connect

    Novak, Petr; Jensen, Taylor J.; Garbe, James C.; Stampfer, Martha R.; Futscher, Bernard W.

    2009-04-20

    The timing and progression of DNA methylation changes during carcinogenesis are not completely understood. To develop a timeline of aberrant DNA methylation events during malignant transformation, we analyzed genome-wide DNA methylation patterns in an isogenic human mammary epithelial cell (HMEC) culture model of transformation. To acquire immortality and malignancy, the cultured finite lifespan HMEC must overcome two distinct proliferation barriers. The first barrier, stasis, is mediated by the retinoblastoma protein and can be overcome by loss of p16(INK4A) expression. HMEC that escape stasis and continue to proliferate become genomically unstable before encountering a second more stringent proliferation barrier, telomere dysfunction due to telomere attrition. Rare cells that acquire telomerase expression may escape this barrier, become immortal, and develop further malignant properties. Our analysis of HMEC transitioning from finite lifespan to malignantly transformed showed that aberrant DNA methylation changes occur in a stepwise fashion early in the transformation process. The first aberrant DNA methylation step coincides with overcoming stasis, and results in few to hundreds of changes, depending on how stasis was overcome. A second step coincides with immortalization and results in hundreds of additional DNA methylation changes regardless of the immortalization pathway. A majority of these DNA methylation changes are also found in malignant breast cancer cells. These results show that large-scale epigenetic remodeling occurs in the earliest steps of mammary carcinogenesis, temporally links DNA methylation changes and overcoming cellular proliferation barriers, and provides a bank of potential epigenetic biomarkers that mayprove useful in breast cancer risk assessment.

  8. Effects of Soybean Agglutinin on Mechanical Barrier Function and Tight Junction Protein Expression in Intestinal Epithelial Cells from Piglets

    PubMed Central

    Pan, Li; Qin, Guixin; Zhao, Yuan; Wang, Jun; Liu, Feifei; Che, Dongsheng

    2013-01-01

    In this study, we sought to investigate the role of soybean agglutinin (SBA) in mediating membrane permeability and the mechanical barrier function of intestinal epithelial cells. The IPEC-J2 cells were cultured and treated with 0, 0.5, 1.0, 1.5, 2.0, 2.5, or 3.0 mg/mL SBA. Transepithelial electrical resistance (TEER) and alkaline phosphatase (AP) activity were measured to evaluate membrane permeability. The results showed a significant decrease in TEER values (p < 0.05) in a time- and dose-dependent manner, and a pronounced increase in AP activity (p < 0.05). Cell growth and cell morphology were used to evaluate the cell viability. A significant cell growth inhibition (p < 0.05) and alteration of morphology were observed when the concentration of SBA was increased. The results of western blotting showed that the expression levels of occludin and claudin-3 were decreased by 31% and 64% compared to those of the control, respectively (p < 0.05). In addition, immunofluorescence labeling indicated an obvious decrease in staining of these targets and changes in their localizations. In conclusion, SBA increased the membrane permeability, inhibited the cell viability and reduced the levels of tight junction proteins (occludin and claudin-3), leading to a decrease in mechanical barrier function in intestinal epithelial cells. PMID:24189218

  9. Entosis allows timely elimination of the luminal epithelial barrier for embryo implantation.

    PubMed

    Li, Yingju; Sun, Xiaofei; Dey, Sudhansu K

    2015-04-21

    During implantation, uterine luminal epithelial (LE) cells first interact with the blastocyst trophectoderm. Within 30 hr after the initiation of attachment, LE cells surrounding the blastocyst in the implantation chamber (crypt) disappear, allowing trophoblast cells to make direct physical contact with the underneath stroma for successful implantation. The mechanism for the extraction of LE cells was thought to be mediated by apoptosis. Here, we show that LE cells in direct contact with the blastocyst are endocytosed by trophoblast cells by adopting the nonapoptotic cell-in-cell invasion process (entosis) in the absence of caspase 3 activation. Our in vivo observations were reinforced by the results of co-culture experiments with primary uterine epithelial cells with trophoblast stem cells or blastocysts showing internalization of epithelial cells by trophoblasts. We have identified entosis as a mechanism to remove LE cells by trophoblast cells in implantation, conferring a role for entosis in an important physiological process. PMID:25865893

  10. AMP-18 Targets p21 to Maintain Epithelial Homeostasis.

    PubMed

    Chen, Peili; Li, Yan Chun; Toback, F Gary

    2015-01-01

    Dysregulated homeostasis of epithelial cells resulting in disruption of mucosal barrier function is an important pathogenic mechanism in inflammatory bowel diseases (IBD). We have characterized a novel gastric protein, Antrum Mucosal Protein (AMP)-18, that has pleiotropic properties; it is mitogenic, anti-apoptotic and can stimulate formation of tight junctions. A 21-mer synthetic peptide derived from AMP-18 exhibits the same biological functions as the full-length protein and is an effective therapeutic agent in mouse models of IBD. In this study we set out to characterize therapeutic mechanisms and identify molecular targets by which AMP-18 maintains and restores disrupted epithelial homeostasis in cultured intestinal epithelial cells and a mouse model of IBD. Tumor necrosis factor (TNF)-α, a pro-inflammatory cytokine known to mediate gastrointestinal (GI) mucosal injury in IBD, was used to induce intestinal epithelial cell injury, and study the effects of AMP-18 on apoptosis and the cell cycle. An apoptosis array used to search for targets of AMP-18 in cells exposed to TNF-α identified the cyclin-dependent kinase inhibitor p21 WAF1/CIP1. Treatment with AMP-18 blunted increases in p21 expression and apoptosis, while reversing disturbed cell cycle kinetics induced by TNF-α. AMP-18 appears to act through PI3K/AKT pathways to increase p21 phosphorylation, thereby reducing its nuclear accumulation to overcome the antiproliferative effects of TNF-α. In vitamin D receptor-deficient mice with TNBS-induced IBD, the observed increase in p21 expression in colonic epithelial cells was suppressed by treatment with AMP peptide. The results indicate that AMP-18 can maintain and/or restore the homeostatic balance between proliferation and apoptosis in intestinal epithelial cells to protect and repair mucosal barrier homeostasis and function, suggesting a therapeutic role in IBD.

  11. Differential expression of gastric MUC5AC in colonic epithelial cells: TFF3-wired IL1 β/Akt crosstalk-induced mucosal immune response against Shigella dysenteriae infection.

    PubMed

    Raja, Subramaniya Bharathi; Murali, Malliga Raman; Devaraj, Halagowder; Devaraj, Sivasithamparam Niranjali

    2012-02-01

    An understanding of the signaling mechanism(s) that regulate the differential expression of gastric mucin MUC5AC in colonic epithelial cells would contribute significantly to investigations of its role in colonic mucosa infected with the bacterial pathogen Shigella dysenteriae. Here we show that S. dysenteriae-Sinduced expression of interleukin-1β upregulates MUC2 expression and the differential expression of MUC5AC. Differential expression of MUC5AC involves crosstalk between interleukin-1β and Akt, whereby the trefoil factor family peptide TFF3 activates Akt by phosphorylation of EGFR. TFF3 also downregulates E-cadherin expression, causing accumulation of β-catenin in the cytosol. Phosphorylation of GSK-3β (inactivated) by activated Akt inhibits ubiquitylation of β-catenin, leading to its nuclear translocation, which then induces the expression of MUC5AC and cyclin D1. Accumulation of cyclin D1 alters the cell cycle, promoting cell survival and proliferation. Human colon HT29MTX cells, which overexpress MUC5AC, were resistant to adherence and invasion of S. dysenteriae when compared with other mucin-secreting HT29 cell types. Thus, during infection with S. dysenteriae, crosstalk between interleukin-1β and Akt wired by TFF3 induces expression of MUC5AC in colonic epithelial cells. Differentially expressed gastric MUC5AC aids in mucosal clearance of S. dysenteriae, inhibiting adherence and invasion of the pathogen to colonic epithelial cells, which protects the host.

  12. Delivery of berberine using chitosan/fucoidan-taurine conjugate nanoparticles for treatment of defective intestinal epithelial tight junction barrier.

    PubMed

    Wu, Shao-Jung; Don, Trong-Ming; Lin, Cheng-Wei; Mi, Fwu-Long

    2014-11-01

    Bacterial-derived lipopolysaccharides (LPS) can cause defective intestinal barrier function and play an important role in the development of inflammatory bowel disease. In this study, a nanocarrier based on chitosan and fucoidan was developed for oral delivery of berberine (Ber). A sulfonated fucoidan, fucoidan-taurine (FD-Tau) conjugate, was synthesized and characterized by Fourier transform infrared (FTIR) spectroscopy. The FD-Tau conjugate was self-assembled with berberine and chitosan (CS) to form Ber-loaded CS/FD-Tau complex nanoparticles with high drug loading efficiency. Berberine release from the nanoparticles had fast release in simulated intestinal fluid (SIF, pH 7.4), while the release was slow in simulated gastric fluid (SGF, pH 2.0). The effect of the berberine-loaded nanoparticles in protecting intestinal tight-junction barrier function against nitric oxide and inflammatory cytokines released from LPS-stimulated macrophage was evaluated by determining the transepithelial electrical resistance (TEER) and paracellular permeability of a model macromolecule fluorescein isothiocyanate-dextran (FITC-dextran) in a Caco-2 cells/RAW264.7 cells co-culture system. Inhibition of redistribution of tight junction ZO-1 protein by the nanoparticles was visualized using confocal laser scanning microscopy (CLSM). The results suggest that the nanoparticles may be useful for local delivery of berberine to ameliorate LPS-induced intestinal epithelia tight junction disruption, and that the released berberine can restore barrier function in inflammatory and injured intestinal epithelial. PMID:25421323

  13. Neisseria meningitidis subverts the polarized organization and intracellular trafficking of host cells to cross the epithelial barrier.

    PubMed

    Barrile, Riccardo; Kasendra, Magdalena; Rossi-Paccani, Silvia; Merola, Marcello; Pizza, Mariagrazia; Baldari, Cosima; Soriani, Marco; Aricò, Beatrice

    2015-09-01

    Translocation of the nasopharyngeal barrier by Neisseria meningitidis occurs via an intracellular microtubule-dependent pathway and represents a crucial step in its pathogenesis. Despite this fact, the interaction of invasive meningococci with host subcellular compartments and the resulting impact on their organization and function have not been investigated. The influence of serogroup B strain MC58 on host cell polarity and intracellular trafficking system was assessed by confocal microscopy visualization of different plasma membrane-associated components (such as E-cadherin, ZO-1 and transferrin receptor) and evaluation of the transferrin uptake and recycling in infected Calu-3 monolayers. Additionally, the association of N. meningitidis with different endosomal compartments was evaluated through the concomitant staining of bacteria and markers specific for Rab11, Rab22a, Rab25 and Rab3 followed by confocal microscopy imaging. Subversion of the host cell architecture and intracellular trafficking system, denoted by mis-targeting of cell plasma membrane components and perturbations of transferrin transport, was shown to occur in response to N. meningitidis infection. Notably, the appearance of all of these events seems to positively correlate with the efficiency of N. meningitidis to cross the epithelial barrier. Our data reveal for the first time that N. meningitidis is able to modulate the host cell architecture and function, which might serve as a strategy of this pathogen for overcoming the nasopharyngeal barrier without affecting the monolayer integrity.

  14. The roles and functional mechanisms of interleukin-17 family cytokines in mucosal immunity

    PubMed Central

    Song, Xinyang; He, Xiao; Li, Xiaoxia; Qian, Youcun

    2016-01-01

    The mucosal immune system serves as our front-line defense against pathogens. It also tightly maintains immune tolerance to self-symbiotic bacteria, which are usually called commensals. Sensing both types of microorganisms is modulated by signalling primarily through various pattern-recognition receptors (PRRs) on barrier epithelial cells or immune cells. After sensing, proinflammatory molecules such as cytokines are released by these cells to mediate either defensive or tolerant responses. The interleukin-17 (IL-17) family members belong to a newly characterized cytokine subset that is critical for the maintenance of mucosal homeostasis. In this review, we will summarize recent progress on the diverse functions and signals of this family of cytokines at different mucosal edges. PMID:27018218

  15. Duodenal Chemosensing and Mucosal Defenses

    PubMed Central

    Akiba, Yasutada; Kaunitz, Jonathan D.

    2011-01-01

    The duodenal mucosa is exposed to endogenous and exogenous chemicals, including acid, CO2, bile acids and nutrients. Mucosal chemical sensors are necessary to exert physiological responses such as secretion, digestion, absorption, and motility. We propose a mucosal chemosensing system by which luminal chemicals are sensed via mucosal acid sensors and G-protein-coupled receptors. Luminal acid/CO2 sensing consists of ecto- and cytosolic carbonic anhydrases, epithelial ion transporters, and acid sensors expressed on the afferent nerves in the duodenum. Furthermore, a luminal L-glutamate signal is mediated via mucosal L-glutamate receptors, including metabotropic glutamate receptors and taste receptor 1 family heterodimers, with activation of afferent nerves and cyclooxygenase, whereas luminal Ca2+ is differently sensed via the calcium-sensing receptor in the duodenum. Recent studies also show the involvement of enteroendocrine G-protein-coupled receptors in bile acid and fatty acid sensing in the duodenum. These luminal chemosensors help activate mucosal defense mechanisms in or- der to maintain the mucosal integrity and physiological responses. Stimulation of luminal chemosensing in the duodenal mucosa may prevent mucosal injury, affect nutrient metabolism, and modulate sensory nerve activity. PMID:21389725

  16. Exocyst Sec10 protects epithelial barrier integrity and enhances recovery following oxidative stress, by activation of the MAPK pathway.

    PubMed

    Park, Kwon Moo; Fogelgren, Ben; Zuo, Xiaofeng; Kim, Jinu; Chung, Daniel C; Lipschutz, Joshua H

    2010-03-01

    Cell-cell contacts are essential for epithelial cell function, and disruption is associated with pathological conditions including ischemic kidney injury. We hypothesize that the exocyst, a highly-conserved eight-protein complex that targets secretory vesicles carrying membrane proteins, is involved in maintaining renal epithelial barrier integrity. Accordingly, increasing exocyst expression in renal tubule cells may protect barrier function from oxidative stress resulting from ischemia and reperfusion (I/R) injury. When cultured on plastic, Madin-Darby canine kidney (MDCK) cells overexpressing Sec10, a central exocyst component, formed domes showing increased resistance to hydrogen peroxide (H2O2). Transepithelial electric resistance (TER) of Sec10-overexpressing MDCK cells grown on Transwell filters was higher than in control MDCK cells, and the rate of TER decrease following H2O2 treatment was less in Sec10-overexpressing MDCK cells compared with control MDCK cells. After removal of H2O2, TER returned to normal more rapidly in Sec10-overexpressing compared with control MDCK cells. In collagen culture MDCK cells form cysts, and H2O2 treatment damaged Sec10-overexpressing MDCK cell cysts less than control MDCK cell cysts. The MAPK pathway has been shown to protect animals from I/R injury. Levels of active ERK, the final MAPK pathway step, were higher in Sec10-overexpressing compared with control MDCK cells. U0126 inhibited ERK activation, exacerbated the H2O2-induced decrease in TER and cyst disruption, and delayed recovery of TER following H2O2 removal. Finally, in mice with renal I/R injury, exocyst expression decreased early and returned to normal concomitant with functional recovery, suggesting that the exocyst may be involved in the recovery following I/R injury.

  17. Intra-epithelial vaccination with COPV L1 DNA by particle-mediated DNA delivery protects against mucosal challenge with infectious COPV in beagle dogs.

    PubMed

    Stanley, M A; Moore, R A; Nicholls, P K; Santos, E B; Thomsen, L; Parry, N; Walcott, S; Gough, G

    2001-04-01

    Protection against viral challenge with canine oral papillomavirus (COPV) was achieved by immunisation via particle-mediated DNA delivery (PMDD) of a plasmid encoding the COPV L1 gene to cutaneous and oral mucosal sites in beagle dogs. The initial dose of approximately 9 microg of DNA was followed by two booster doses at 6 week intervals. A similar approach was used to vaccinate a control group of animals with plasmid DNA encoding the Hepatitis B virus S gene. Following challenge at the oral mucosa with COPV all animals vaccinated with the COPV L1 gene were protected against disease. However five of six animals in the control group developed COPV induced papillomas at the oral mucosa. Both cell-mediated lymphoproliferative and humoral antibody responses to the DNA vaccine were observed. Our data indicate that PMDD of plasmid DNA can protect against mucosal challenge with papillomavirus. PMID:11282188

  18. Clinical Characteristics Associated with Post-Operative Intestinal Epithelial Barrier Dysfunction in Children with Congenital Heart Disease

    PubMed Central

    Typpo, Katri V; Larmonier, Claire B.; Deschenes, Jendar; Redford, Daniel; Kiela, Pawel R.; Ghishan, Fayez K.

    2014-01-01

    Objective Children with congenital heart disease (CHD) have loss of intestinal epithelial barrier function (EBF), which increases their risk for post-operative sepsis and organ dysfunction. We do not understand how post-operative cardiopulmonary support or the inflammatory response to cardiopulmonary bypass (CPB) might alter intestinal EBF. We examined variation in a panel of plasma biomarkers to reflect intestinal EBF (cellular and paracellular structure and function) after CPB and in response to routine ICU care. Design Prospective cohort Setting University medical center cardiac intensive care unit Patients Twenty children aged newborn to 18 years undergoing repair or palliation of CHD with CPB. Interventions We measured baseline and repeated plasma FABP2, citrulline, claudin 3, and dual sugar permeability test (DSPT) to reflect intestinal epithelial integrity, epithelial function, paracellular integrity, and paracellular function, respectively. We measured baseline and repeated plasma pro-inflammatory (IL-6, TNF-α, IFN-γ) and anti-inflammatory (IL4, IL10) cytokines, known to modulate intestinal EBF in murine models of CPB. Measurements and Main Results All patients had abnormal baseline FABP2 concentrations (mean 3815.5 pg/mL), (normal 41–336 pg/mL). Cytokine response to CPB was associated with early, but not late changes in plasma concentrations of FABP2 and citrulline. Variation in biomarker concentrations over time were associated with aspects of ICU care indicating greater severity of illness: claudin 3, FABP2, and DSPT ratio were associated with symptoms of feeding intolerance (p<0.05) while FABP2 was positively associated with vasoactive-inotrope score (VIS) (p=0.04). Citrulline was associated with larger arteriovenous O2 saturation difference (p=0.04) and had a complex relationship with VIS. Conclusions Children undergoing CPB for repair or palliation of CHD are at risk for intestinal injury and often present with evidence for loss of intestinal

  19. Polarity Protein Complex Scribble/Lgl/Dlg and Epithelial Cell Barriers

    PubMed Central

    Su, Wen-Hui; Mruk, Dolores D.; Wong, Elissa W.P.; Lui, Wing-Yee; Cheng, C. Yan

    2014-01-01

    The Scribble polarity complex or module is one of the three polarity modules that regulate cell polarity in multiple epithelia including blood-tissue barriers. This protein complex is composed of Scribble, Lethal giant larvae (Lgl) and Discs large (Dlg), which are well conserved across species from fruitflies and worms to mammals. Originally identified in Drosophila and C. elegans where the Scribble complex was found to work with the Par-based and Crumbs-based polarity modules to regulate apicobasal polarity and asymmetry in cells and tissues during embryogenesis, their mammalian homologs have all been identified in recent years. Components of the Scribble complex are known to regulate multiple cellular functions besides cell polarity, which include cell proliferation, assembly and maintenance of adherens junction (AJ) and tight junction (TJ), and they are also tumor suppressors. Herein, we provide an update on the Scribble polarity complex and how this protein complex modulates cell adhesion with some emphasis on its role in Sertoli cell blood-testis barrier (BTB) function. It should be noted that this is a rapidly developing field, in particular the role of this protein module in blood-tissue barriers, and this short chapter attempts to provide the information necessary for investigators studying reproductive biology and blood-tissue barriers to design future studies. We also include results of recent studies from flies and worms since this information will be helpful in planning experiments for future functional studies in the testis to understand how Scribble-based proteins regulate BTB dynamics and spermatogenesis. PMID:23397623

  20. Effects of nitrogen dioxide on alveolar epithelial barrier properties. Research report, September 1983-January 1987

    SciTech Connect

    Crandall, E.D.; Cheek, J.M.; Shaw, M.E.; Postlethwait, E.M.

    1987-10-01

    This study used primary cultures of rat Type II pneumocytes to investigate if, and how, nitrogen dioxide (NO/sub 2/) alters the permeability of the alveolar epithelia. Susceptibility of cells to NO/sub 2/ appears to depend on the nutrient media of the cells, with a 10-fold difference observed between minimum essential medium (MEM) and MEM supplemented with Ham's F12 media. No protective effect was observed when the antioxidant, vitamin E, was added to the media. Bioelectric studies were conducted on cells cultured on a porous substrate. Exposure to 20 ppm NO/sub 2/ caused significant reductions in tissue resistance but not in short-circuit current. It was concluded that NO/sub 2/ primarily affects epithelial active and passive transport via altered tight junctional pathways that result from damage to paracellular tight junctions or, possibly, to transcellular transport membrane pumps. The authors suggest that free-radical formation causes the damage observed.

  1. Cellular zinc is required for intestinal epithelial barrier maintenance via the regulation of claudin-3 and occludin expression.

    PubMed

    Miyoshi, Yuka; Tanabe, Soichi; Suzuki, Takuya

    2016-07-01

    Intracellular zinc is required for a variety of cell functions, but its precise roles in the maintenance of the intestinal tight junction (TJ) barrier remain unclear. The present study investigated the essential roles of intracellular zinc in the preservation of intestinal TJ integrity and the underlying molecular mechanisms. Depletion of intracellular zinc in both intestinal Caco-2 cells and mouse colons through the application of a cell-permeable zinc chelator N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) induced a disruption of the TJ barrier, as indicated by increased FITC-labeled dextran flux and decreased transepithelial electrical resistance. The TPEN-induced TJ disruption is associated with downregulation of two TJ proteins, occludin and claudin-3. Biotinylation of cell surface proteins revealed that the zinc depletion induced the proteolysis of occludin but not claudin-3. Occludin proteolysis was sensitive to the inhibition of calpain activity, and increased calpain activity was observed in the zinc-depleted cells. Although quantitative PCR analysis and promoter reporter assay have demonstrated that the zinc depletion-induced claudin-3 downregulation occurred at transcriptional levels, a site-directed mutation in the egr1 binding site in the claudin-3 promoter sequence induced loss of both the basal promoter activity and the TPEN-induced decreases. Reduced egr1 expression by a specific siRNA also inhibited claudin-3 expression and transepithelial electrical resistance maintenance in cells. This study shows that intracellular zinc has an essential role in the maintenance of the intestinal epithelial TJ barrier through regulation of occludin proteolysis and claudin-3 transcription. PMID:27151944

  2. Lung epithelial barrier function and wound healing are decreased by IL-4 and IL-13 and enhanced by IFN-gamma.

    PubMed

    Ahdieh, M; Vandenbos, T; Youakim, A

    2001-12-01

    To understand the effects of cytokines on epithelial cells in asthma, we have investigated the effects of interleukin (IL)-4, IL-13, and interferon (IFN)-gamma on barrier function and wound healing in Calu-3 human lung epithelial cells. IL-4 and IL-13 treatment of Calu-3 cells grown on Transwell filters resulted in a 70-75% decrease in barrier function as assessed by electrophysiological and [(14)C]mannitol flux measurements. In contrast, IFN-gamma enhanced barrier function threefold using these same parameters. Cells treated concurrently with IFN-gamma and IL-4 or IL-13 showed an initial decline in barrier function that was reversed within 2 days, resulting in barrier levels comparable to control cells. Analysis of the tight junction-associated proteins ZO-1 and occludin showed that IL-4 and IL-13 significantly reduced ZO-1 expression and modestly decreased occludin expression compared with controls. IFN-gamma, quite unexpectedly given its enhancing effect on barrier function, reduced expression of ZO-1 and occludin to almost undetectable levels compared with controls. In wound-healing assays of cells grown on collagen I, IL-4 and IL-13 decreased migration, whereas IFN-gamma treatment enhanced migration, compared with control cells. Addition of IFN-gamma, in combination with IL-4 or IL-13, restored migration of cells to control levels. Migration differences observed between the various cytokine treatments was correlated with expression of the collagen I-binding alpha(2)beta(1)-integrin at the leading edge of cells at the wound front; alpha(2)beta(1)-integrin expression was decreased in IFN-gamma-treated cells compared with controls, whereas it was highest in IL-4- and IL-13-treated cells. These results demonstrate that IL-4 and IL-13 diminish the capacity of Calu-3 cells to maintain barrier function and repair wounds, whereas IFN-gamma promotes epithelial restitution by enhancing barrier function and wound healing. PMID:11698262

  3. Protective effects of ψ taraxasterol 3-O-myristate and arnidiol 3-O-myristate isolated from Calendula officinalis on epithelial intestinal barrier.

    PubMed

    Dall'Acqua, Stefano; Catanzaro, Daniela; Cocetta, Veronica; Igl, Nadine; Ragazzi, Eugenio; Giron, Maria Cecilia; Cecconello, Laura; Montopoli, Monica

    2016-03-01

    The triterpene esters ᴪ taraxasterol-3-O-myristate (1) and arnidiol-3-O-myristate (2) were tested for their ability to protect epithelial intestinal barrier in an in vitro model. Their effects on ROS production and on trans-epithelial resistance were investigated on CaCo-2 cell monolayers both in basal and stress-induced conditions. Both compounds were able to modulate the stress damage induced by H2O2 and INFγ+TNFα, showing a potential use as model compounds for the study of new therapeutic agents for intestinal inflammations. PMID:26791917

  4. Protective effects of ψ taraxasterol 3-O-myristate and arnidiol 3-O-myristate isolated from Calendula officinalis on epithelial intestinal barrier.

    PubMed

    Dall'Acqua, Stefano; Catanzaro, Daniela; Cocetta, Veronica; Igl, Nadine; Ragazzi, Eugenio; Giron, Maria Cecilia; Cecconello, Laura; Montopoli, Monica

    2016-03-01

    The triterpene esters ᴪ taraxasterol-3-O-myristate (1) and arnidiol-3-O-myristate (2) were tested for their ability to protect epithelial intestinal barrier in an in vitro model. Their effects on ROS production and on trans-epithelial resistance were investigated on CaCo-2 cell monolayers both in basal and stress-induced conditions. Both compounds were able to modulate the stress damage induced by H2O2 and INFγ+TNFα, showing a potential use as model compounds for the study of new therapeutic agents for intestinal inflammations.

  5. PKCι interacts with Rab14 and modulates epithelial barrier function through regulation of claudin-2 levels

    PubMed Central

    Lu, Ruifeng; Dalgalan, Dogukan; Mandell, Edward K.; Parker, Sara S.; Ghosh, Sourav; Wilson, Jean M.

    2015-01-01

    PKCι is essential for the establishment of epithelial polarity and the normal assembly of tight junctions. We find that PKCι knockdown does not compromise the steady-state distribution of most tight junction proteins but results in increased transepithelial resistance (TER) and decreased paracellular permeability. Analysis of the levels of tight junction components demonstrates that claudin-2 protein levels are decreased. However, other tight junction proteins, such as claudin-1, ZO-1, and occludin, are unchanged. Incubation with an aPKC pseudosubstrate recapitulates the phenotype of PKCι knockdown, including increased TER and decreased levels of claudin-2. In addition, overexpression of PKCι results in increased claudin-2 levels. ELISA and coimmunoprecipitation show that the TGN/endosomal small GTPase Rab14 and PKCι interact directly. Immunolabeling shows that PKCι and Rab14 colocalize in both intracellular puncta and at the plasma membrane and that Rab14 expression is required for normal PKCι distribution in cysts in 3D culture. We showed previously that knockdown of Rab14 results in increased TER and decreased claudin-2. Our results suggest that Rab14 and aPKC interact to regulate trafficking of claudin-2 out of the lysosome-directed pathway. PMID:25694446

  6. A Deafness Associated Mutant Human Connexin 26 Improves The Epithelial Barrier In Vitro

    PubMed Central

    Man, Y.K. Stella; Trolove, Caroline; Tattersall, Daniel; Thomas, Anna C.; Papakonstantinopoulou, Annie; Patel, Drashnika; Scott, Claire; Chong, Jiehan; Jagger, Daniel J.; O’Toole, Edel A.; Navsaria, Harshad; Curtis, Michael A.; Kelsell, David P.

    2010-01-01

    A large proportion of recessive non-syndromic hearing loss is due to mutations in the GJB2 gene encoding connexin 26 (Cx26), a component of a gap junction. Within different ethnic groups there are specific common recessive mutations each with a relatively high carrier frequency suggesting a possibility of heterozygous advantage. Carriers of the R143W GJB2 allele, the most prevalent in the African population, present with a thicker epidermis than non-carriers. In this study, we show (R143W)Cx26 expressing keratinocytes form a significantly thicker epidermis in an organotypic co-culture skin model. In addition, we show increased migration of cells expressing (R143W)Cx26 compared to (WT)Cx26 overexpressing cells. We also demonstrate that cells expressing (R143W)Cx26 are significantly less susceptible to cellular invasion by the enteric pathogen Shigella flexneri than (WT)Cx26 expressing cells. These in vitro studies suggest an advantageous effect of (R143W)Cx26 in epithelial cells. PMID:17581693

  7. Breaking the epithelial polarity barrier in cancer: the strange case of LKB1/PAR-4

    PubMed Central

    Partanen, Johanna I.; Tervonen, Topi A.; Klefström, Juha

    2013-01-01

    The PAR clan of polarity regulating genes was initially discovered in a genetic screen searching for genes involved in asymmetric cell divisions in the Caenorhabditis elegans embryo. Today, investigations in worms, flies and mammals have established PAR proteins as conserved and fundamental regulators of animal cell polarization in a broad range of biological phenomena requiring cellular asymmetries. The human homologue of invertebrate PAR-4, a serine–threonine kinase LKB1/STK11, has caught attention as a gene behind Peutz–Jeghers polyposis syndrome and as a bona fide tumour suppressor gene commonly mutated in sporadic cancer. LKB1 functions as a master regulator of AMP-activated protein kinase (AMPK) and 12 other kinases referred to as the AMPK-related kinases, including four human homologues of PAR-1. The role of LKB1 as part of the energy sensing LKB1-AMPK module has been intensively studied, whereas the polarity function of LKB1, in the context of homoeostasis or cancer, has gained less attention. Here, we focus on the PAR-4 identity of LKB1, discussing the weight of evidence indicating a role for LKB1 in regulation of cell polarity and epithelial integrity across species and highlight recent investigations providing new insight into the old question: does the PAR-4 identity of LKB1 matter in cancer? PMID:24062587

  8. Toll-like receptor 2 regulates the barrier function of human bronchial epithelial monolayers through atypical protein kinase C zeta, and an increase in expression of claudin-1

    PubMed Central

    Ragupathy, Sakthikumar; Esmaeili, Farnaz; Paschoud, Serge; Sublet, Emmanuelle; Citi, Sandra; Borchard, Gerrit

    2014-01-01

    We investigated the role of Toll-like receptor (TLR) 2 in maintaining the integrity of the airway epithelial barrier using the human bronchial epithelial cell line Calu-3. Activation of TLR2 by its ligands, Pam3CysSK4 and Peptidoglycan showed a concentration dependent increase in epithelial barrier function, as measured by transepithelial electrical resistance (TEER). This was confirmed by a decrease in paracellular flux of fluorescein sodium. This TLR2 induced increase in TEER was significantly reduced by pretreatment with polyclonal anti-human TLR2-neutralizing antibody. TLR2 stimulation in Calu-3 cell monolayers resulted in an increased expression of the tight junction proteins claudin-1 and ZO-1, and a decreased expression of occludin, at both the mRNA and protein levels. A pseudosubstrate inhibitor to PKCζ significantly prevented the TLR2 mediated increase in barrier function. It also prevented the increase in claudin-1 in a concentration dependent manner up to 1 µM. TLR2 stimulation led to an increase in phosphorylation of atypical PKC ζ, which was prevented by the pseudosubstrate inhibitor in a concentration dependent manner. Taken together, our observations support a model whereby increased tight junction barrier function induced by activation of TLR2 occurs through increased expression of claudin-1, and through modulation of PKC ζ activity. PMID:25101232

  9. Medroxyprogesterone acetate and levonorgestrel increase genital mucosal permeability and enhance susceptibility to genital herpes simplex virus type 2 infection

    PubMed Central

    Calla, Nirk E Quispe; Miguel, Rodolfo D Vicetti; Boyaka, Prosper N; Hall-Stoodley, Luanne; Kaur, Balveen; Trout, Wayne; Pavelko, Stephen D; Cherpes, Thomas L

    2016-01-01

    Depot-medroxyprogesterone acetate (DMPA) is a hormonal contraceptive especially popular in areas with high prevalence of HIV and other sexually transmitted infections (STI). While observational studies identify DMPA as an important STI risk factor, mechanisms underlying this connection are undefined. Levonorgestrel (LNG) is another progestin used for hormonal contraception, but its effect on STI susceptibility is much less explored. Using a mouse model of genital HSV-2 infection, we herein found DMPA and LNG similarly reduced genital expression of the desmosomal cadherin desmoglein-1α (DSG1α), enhanced access of inflammatory cells to genital tissue by increasing mucosal epithelial permeability, and increased susceptibility to viral infection. Additional studies with uninfected mice revealed DMPA-mediated increases in mucosal permeability promoted tissue inflammation by facilitating endogenous vaginal microbiota invasion. Conversely, concomitant treatment of mice with DMPA and intravaginal estrogen restored mucosal barrier function and prevented HSV-2 infection. Evaluating ectocervical biopsy tissue from women before and 1 month after initiating DMPA remarkably revealed inflammation and barrier protection were altered by treatment identically to changes seen in progestin-treated mice. Together, our work reveals DMPA and LNG diminish the genital mucosal barrier; a first-line defense against all STI, but may offer foundation for new contraceptive strategies less compromising of barrier protection. PMID:27007679

  10. Sieving characteristics of cytokine- and peroxide-induced epithelial barrier leak: Inhibition by berberine

    PubMed Central

    DiGuilio, Katherine M; Mercogliano, Christina M; Born, Jillian; Ferraro, Brendan; To, Julie; Mixson, Brittany; Smith, Allison; Valenzano, Mary Carmen; Mullin, James M

    2016-01-01

    AIM: To study whether the inflammatory bowel disease (IBD) colon which exhibits varying severity and cytokine levels across its mucosa create varying types of transepithelial leak. METHODS: We examined the effects of tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-1-β (IL1β) and hydrogen peroxide (H2O2) - singly and in combinations - on barrier function of CACO-2 cell layers. Our focus was on the type (not simply the magnitude) of transepithelial leak generated by these agents as measured by transepithelial electrical resistance (TER) and transepithelial flux of 14C-D-mannitol, 3H-Lactulose and 14C-Polyethylene glycol as radiolabeled probe molecules. The isoquinoline alkaloid, berberine, was then examined for its ability to reduce specific types of transepithelial leak. RESULTS: Exposure to TNF-α alone (200 ng/mL; 48 h) induced a 50% decrease in TER, i.e., increased leak of Na+ and Cl- - with only a marginal but statistically significant increase in transepithelial leak of 14C-mannitol (Jm). Exposure to TNF-α + IFN-γ (200 ng/mL; 48 h) + IL1β (50 ng/mL; 48 h) did not increase the TER change (from TNF-α alone), but there was now a 100% increase in Jm. There however was no increase in transepithelial leak of two larger probe molecules, 3H-lactulose and 14C-polyethylene glycol (PEG). However, exposure to TNF-α + IFN-γ + IL1β followed by a 5 h exposure to 2 mmol/L H2O2 resulted in a 500% increase in 14C-PEG leak as well as leak to the luminal mitogen, epidermal growth factor. CONCLUSION: This model of graded transepithelial leak is useful in evaluating therapeutic agents reducing IBD morbidity by reducing barrier leak to various luminal substances. PMID:27190695

  11. Effects of Phonation Time and Magnitude Dose on Vocal Fold Epithelial Genes, Barrier Integrity, and Function

    PubMed Central

    Kojima, Tsuyoshi; Valenzuela, Carla V.; Novaleski, Carolyn K.; Van Deusen, Mark; Mitchell, Joshua R.; Garrett, C. Gaelyn; Sivasankar, M. Preeti; Rousseau, Bernard

    2014-01-01

    Objective To investigate the effects of increasing time and magnitude doses of vibration exposure on transcription of the vocal fold's junctional proteins, structural alterations, and functional tissue outcomes. Study Design Animal study. Methods 100 New Zealand White breeder rabbits were studied. Dependent variables were measured in response to increasing time doses (30, 60, or 120 minutes) and magnitude doses (control, modal intensity, and raised intensity) of vibration exposure. Messenger RNA expression of occludin, zonula occluden-1 (ZO-1), E-cadherin, β-catenin, interleukin 1β (IL-1β), cyclooxygenase-2 (COX-2), transforming growth factor β-1 (TGFβ1), and fibronectin were measured. Tissue structural alterations were assessed using transmission electron microscopy (TEM). Transepithelial resistance was used to measure functional tissue outcomes. Results Occludin gene expression was downregulated in vocal folds exposed to 120 minute time doses of raised intensity phonation, relative to control, and modal intensity phonation. ZO-1 gene expression was upregulated following a 120 minute time dose of modal intensity phonation, compared to control, and downregulated after a 120 minute time dose of raised intensity phonation, compared to modal intensity phonation. E-cadherin gene expression was downregulated after a120 minute time dose of raised intensity phonation, compared to control and modal intensity phonation. TEM revealed extensive desquamation of the stratified squamous epithelial cells with increasing time and magnitude doses of vibration exposure. A general observation of lower transepithelial resistance measures was made in tissues exposed to raised intensity phonation, compared to all other groups. Conclusions This study provides evidence of vocal fold tissue responses to varying time and magnitude doses of vibration exposure. Level of Evidence N/A PMID:25073715

  12. Anti-mouse CD52 monoclonal antibody ameliorates intestinal epithelial barrier function in interleukin-10 knockout mice with spontaneous chronic colitis.

    PubMed

    Wang, Honggang; Dong, Jianning; Shi, Peiliang; Liu, Jianhui; Zuo, Lugen; Li, Yi; Gong, Jianfeng; Gu, Lili; Zhao, Jie; Zhang, Liang; Zhang, Wei; Zhu, Weiming; Li, Ning; Li, Jieshou

    2015-02-01

    Intestinal inflammation causes tight junction changes and death of epithelial cells, and plays an important role in the development of Crohn's disease (CD). CD52 monoclonal antibody (CD52 mAb) directly targets the cell surface CD52 and is effective in depleting mature lymphocytes by cytolytic effects in vivo, leading to long-lasting changes in adaptive immunity. The aim of this study was to investigate the therapeutic effect of CD52 mAb on epithelial barrier function in animal models of IBD. Interleukin-10 knockout mice (IL-10(-/-) ) of 16 weeks with established colitis were treated with CD52 mAb once a week for 2 weeks. Severity of colitis, CD4(+) lymphocytes and cytokines in the lamina propria, epithelial expression of tight junction proteins, morphology of tight junctions, tumour necrosis factor-α (TNF-α)/TNF receptor 2 (TNFR2) mRNA expression, myosin light chain kinase (MLCK) expression and activity, as well as epithelial apoptosis in proximal colon were measured at the end of the experiment. CD52 mAb treatment effectively attenuated colitis associated with decreased lamina propria CD4(+) lymphocytes and interferon-γ/IL-17 responses in colonic mucosa in IL-10(-/-) mice. After CD52 mAb treatment, attenuation of colonic permeability, increased epithelial expression and correct localization of tight junction proteins (occludin and zona occludens protein-1), as well as ameliorated tight junction morphology were observed in IL-10(-/-) mice. CD52 mAb treatment also effectively suppressed the epithelial apoptosis, mucosa TNF-α mRNA expression, epithelial expression of long MLCK, TNFR2 and phosphorylation of MLC. Our results indicated that anti-CD52 therapy may inhibit TNF-α/TNFR2-mediated epithelial apoptosis and MLCK-dependent tight junction permeability by depleting activated T cells in the gut mucosa.

  13. Nutritional Keys for Intestinal Barrier Modulation

    PubMed Central

    De Santis, Stefania; Cavalcanti, Elisabetta; Mastronardi, Mauro; Jirillo, Emilio; Chieppa, Marcello

    2015-01-01

    The intestinal tract represents the largest interface between the external environment and the human body. Nutrient uptake mostly happens in the intestinal tract, where the epithelial surface is constantly exposed to dietary antigens. Since inflammatory response toward these antigens may be deleterious for the host, a plethora of protective mechanisms take place to avoid or attenuate local damage. For instance, the intestinal barrier is able to elicit a dynamic response that either promotes or impairs luminal antigens adhesion and crossing. Regulation of intestinal barrier is crucial to control intestinal permeability whose increase is associated with chronic inflammatory conditions. The cross talk among bacteria, immune, and dietary factors is able to modulate the mucosal barrier function, as well as the intestinal permeability. Several nutritional products have recently been proposed as regulators of the epithelial barrier, even if their effects are in part contradictory. At the same time, the metabolic function of the microbiota generates new products with different effects based on the dietary content. Besides conventional treatments, novel therapies based on complementary nutrients are now growing. Fecal therapy has been recently used for the clinical treatment of refractory Clostridium difficile infection instead of the classical antibiotic therapy. In the present review, we will outline the epithelial response to nutritional components derived from dietary intake and microbial fermentation focusing on the consequent effects on the integrity of the epithelial barrier. PMID:26697008

  14. Mucosal immunology

    PubMed Central

    Bienenstock, J.; Befus, A. D.

    1980-01-01

    In this review, we shall highlight some recent advances in mucosal immunology and also those concepts which seem to us to merit more attention than they normally receive. Since we cannot hope to be all inclusive, we recommend the following articles and books to the reader (Tomasi & Bienenstock, 1968; Tomasi & Grey, 1972; Bienenstock, 1974; Heremans, 1974; Mestecky & Lawton, 1974; Lamm, 1976; Tomasi, 1976; Waksman & Ozer, 1976; Porter & Knight, 1977; McGhee, Mestecky & Babb, 1978; Ogra & Dayton, 1979; Befus & Bienenstock, 1980). PMID:7002769

  15. FRET-based dual-emission and pH-responsive nanocarriers for enhanced delivery of protein across intestinal epithelial cell barrier.

    PubMed

    Lu, Kun-Ying; Lin, Cheng-Wei; Hsu, Chun-Hua; Ho, Yi-Cheng; Chuang, Er-Yuan; Sung, Hsing-Wen; Mi, Fwu-Long

    2014-10-22

    The oral route is a convenient and commonly employed way for drug delivery. However, therapeutic proteins have poor bioavailability upon oral administration due to the impermeable barrier from intestinal epithelial tight junction (TJ). Moreover, the pH of the small intestine varies among different regions of the intestinal tract where digestion and absorption occur at different levels. In this study, a tunable dual-emitting and pH-responsive nanocarrier that can alter the fluorescent color and emission intensity in response to pH changes and can trigger the opening of intestinal epithelial TJ at different levels were developed from chitosan-N-arginine and poly(γ-glutamic acid)-taurine conjugates. As pH increased from 6.0 to 8.0, the binding affinity of the oppositely charged polyions decreased, whereas the ratio of the intensity of the donor-to-acceptor emission intensity (ID/IA) increased by 27-fold. The fluorescent and pH-responsive nanocarrier was able to monitor the pH change of intestinal environment and to control the release of an anti-angiogenic protein in response to the pH gradient. The nanocarrier triggered the opening of intestinal epithelial TJ and consequently enhanced the permeation of the released protein through the intestinal epithelial barrier model (Caco-2 cell monolayer) to inhibit tube formation of human umbilical vein endothelial cells. PMID:25260022

  16. Intestinal water absorption through aquaporin 1 expressed in the apical membrane of mucosal epithelial cells in seawater-adapted Japanese eel.

    PubMed

    Aoki, Mayumi; Kaneko, Toyoji; Katoh, Fumi; Hasegawa, Sanae; Tsutsui, Naoaki; Aida, Katsumi

    2003-10-01

    To elucidate the mechanisms associated with water absorption in the intestine, we compared drinking and intestinal water absorption in freshwater- and seawater-adapted Japanese eels, and investigated a possible involvement of aquaporin (AQP) in the absorption of water in the intestine. Seawater eels ingested more water than freshwater eels, the drinking rate being 0.02 ml kg(-1) h(-1) in fresh water and 0.82 ml kg(-1) h(-1) in sea water. In intestinal sacs prepared from freshwater and seawater eels, water absorption increased in time- and hydrostatic pressure-dependent manners. The water absorption rates were greater in seawater sacs than in freshwater sacs, and also greater in the posterior intestine than in the anterior. In view of the enhanced water permeability in the intestine of seawater eel, we cloned two cDNAs encoding AQP from the seawater eel intestine, and identified two eel homologues (S-AQP and L-AQP) of mammalian AQP1. S-AQP and L-AQP possessed the same amino acid sequence, except that one amino acid was lacking in S-AQP and two amino acids were substituted. Eel AQP1 was expressed predominantly in the intestine, and the expression levels were higher in seawater eel than in freshwater eel. Immunocytochemical studies revealed intense AQP1 immunoreaction in the apical surface of columnar epithelial cells in seawater eel, in which the immunoreaction was stronger in the posterior intestine than in the anterior. In contrast, the immunoreaction was faint in the freshwater eel intestine. Preferential localization of AQP1 in the apical membrane of epithelial cells in the posterior intestine of seawater eel indicates that this region of the intestine is responsible for water absorption, and that AQP1 may act as a water entry site in the epithelial cells.

  17. Nerve growth factor injected into the gastric ulcer base incorporates into endothelial, neuronal, glial and epithelial cells: implications for angiogenesis, mucosal regeneration and ulcer healing.

    PubMed

    Tanigawa, T; Ahluwalia, A; Watanabe, T; Arakawa, T; Tarnawski, A S

    2015-08-01

    A previous study has demonstrated that locally administered growth factors such as epidermal growth factor, basic fibroblast growth factor and hepatocyte growth factor can accelerate healing of experimental gastric ulcers in rats. That study indicates that locally administered growth factors can exert potent biological effects resulting in enhanced gastric ulcers healing. However, the fate of injected growth factors, their retention and localization to specific cellular compartments have not been examined. In our preliminary study, we demonstrated that local injection of nerve growth factor to the base of experimental gastric ulcers dramatically accelerates ulcer healing, increases angiogenesis - new blood vessel formation, and improves the quality of vascular and epithelial regeneration. Before embarking on larger, definitive and time sequence studies, we wished to determine whether locally injected nerve growth factor is retained in gastric ulcer's tissues and taken up by specific cells during gastric ulcer healing. Gastric ulcers were induced in anesthetized rats by local application of acetic acid using standard methods; and, 60 min later fluorescein isothiocyanate-labeled nerve growth factor was injected locally to the ulcer base. Rats were euthanized 2, 5 and 10 days later. Gastric specimens were obtained and processed for histology. Unstained paraffin sections were examined under a fluorescence microscope, and the incorporation of fluorescein isothiocyanate-labeled nerve growth factor into various gastric tissue cells was determined and quantified. In addition, we performed immunostaining for S100β protein that is expressed in neural components. Five and ten days after ulcer induction labeled nerve growth factor (injected to the gastric ulcer base) was incorporated into endothelial cells of blood vessels, neuronal, glial and epithelial cells, myofibroblasts and muscle cells. This study demonstrates for the first time that during gastric ulcer healing

  18. Mucosal transmission of HIV-1: first stop dendritic cells.

    PubMed

    Wilkinson, John; Cunningham, Anthony L

    2006-12-01

    Worldwide the heterosexual route is the prevalent mode of transmission of HIV, increasing the demand for measures that block the sexual spread of HIV infection. Vaccines designed to prevent mucosal transmission of HIV should be considered a component of vaccine strategies against HIV (in addition to cytotoxic T cells required for clearance and to prevent viral dissemination) and include antibodies, which are capable of blocking HIV entry at mucosal epithelial barriers, and prevent initial infection of target cells in the mucosa. However, in the interim and in the absence of an effective vaccine, the development of microbicides, topical preparations that block the early steps of HIV infection and transmission, may represent a more viable alternative to condom use in many HIV infected regions of the world especially by empowering women. To date there has been some success with antiviral antibodies applied as a microbicide capable of preventing SIV infection in macaques and reports of vaccines capable of preventing intravaginal and intrarectal inoculated SIV. However, for such success in humans a much greater understanding of the mechanisms involved in the very early stages of mucosal transmission in HIV infection are required. These may lead to additional strategies to inactivate or inhibit viral uptake and replication before a potentially life threatening acute infection develops. Such measures will lead to the development of effective microbicides and vaccines that will diminish the global spread of HIV. PMID:17168831

  19. Dual effect of nitrogen dioxide on barrier properties of guinea pig tracheobronchial epithelial monolayers cultured in an air interface

    SciTech Connect

    Robison, T.W.; Kim, K.J.

    1995-08-01

    Nitrogen dioxide (NO{sub 2}) is an oxidant gas that may injure the airway epithelial lining, leading to decrements in barrier and active ion transport properties. The present studies examined alterations of bioelectric properties and solute flux by guinea pig tracheobronchial epithelial (GPTE) monolayers exposed in vitro to NO{sub 2}. Confluent GPTE monolayers were exposed to NO{sub 2} levels between 0.5 and 5 ppm, while controls were exposed to air. Following exposure, monolayers were mounted in Ussing chambers for measurement of transepithelial resistance (R{sub te}) and short-circuit current (SCC). A 1-h exposure to 1 ppm NO{sub 2} significantly increased SCC to 131.3 {+-} 8.7% of air controls, while R{sub te} with a value of 109.3 {+-} 13.8% was unchanged. In contrast, a 1-h exposure to 2 or 5 ppm NO{sub 2} significantly decreased R{sub te} to 39.0 {+-} 1.6 or 33.5 {+-} 7.3% of air controls, respectively, while SCC values of 140.3 {+-} 10.4 or 153.3 {+-} 8.6%, respectively, were also significantly elevated. A 1-h exposure to 2 or 5 ppm NO{sub 2} significantly increased sucrose permeability across GPTE monolayers to 446.8 {+-} 117 or 313.3 {+-} 39.5% of air controls, respectively, while glycerol permeability was unchanged. In contrast, a 1-h exposure to 1 ppm NO{sub 2} produced no alterations of sucrose or glycerol flux. The SCC of control GPTE monolayers (1-h air exposure) consisted of 50% bumetanide-sensitive and 40% amiloride-sensitive current; exposure for 1 h to 2 ppm NO{sub 2} led to no changes in the corresponding SCC components. Active ion transport (i.e., SCC) across the airway epithelium was significantly increased after exposure to NO{sub 2} levels {le}1 ppm with no change of paracellular pathways for diffusion, suggesting that this reactive gas alters cell membrane function. The increased SCC may lead to impairment of fluid balance and mucociliary clearance. 34 refs., 4 figs.

  20. JAK-STAT and intestinal mucosal immunology

    PubMed Central

    Heneghan, Aaron F; Pierre, Joseph F; Kudsk, Kenneth A

    2013-01-01

    The intestinal mucosal immune system is challenged with bacteria, viruses, and parasites, in addition to food and environmental antigens, that require dynamic immune responsiveness for homeostasis. One central signaling pathway is JAK-STAT, which regulates the adaptive and innate immune arms of mucosal immunity as well as epithelial repair and regeneration. Adaptive immunity includes lymphocyte mediated secretion of specific antibodies, while innate immune respones include secretion of non-antigen specific compounds. This review examines effects of specialized nutrition support on JAK-STAT in innate immune function and in lymphocyte modulation and epithelial antibody transport in gut-associated lymphoid tissue. PMID:24416649

  1. The serine protease-mediated increase in intestinal epithelial barrier function is dependent on occludin and requires an intact tight junction.

    PubMed

    Ronaghan, Natalie J; Shang, Judie; Iablokov, Vadim; Zaheer, Raza; Colarusso, Pina; Turner, Jerrold R; MacNaughton, Wallace K

    2016-09-01

    Barrier dysfunction is a characteristic of the inflammatory bowel diseases (IBD), Crohn's disease and ulcerative colitis. Understanding how the tight junction is modified to maintain barrier function may provide avenues for treatment of IBD. We have previously shown that the apical addition of serine proteases to intestinal epithelial cell lines causes a rapid and sustained increase in transepithelial electrical resistance (TER), but the mechanisms are unknown. We hypothesized that serine proteases increase barrier function through trafficking and insertion of tight junction proteins into the membrane, and this could enhance recovery of a disrupted monolayer after calcium switch or cytokine treatment. In the canine epithelial cell line, SCBN, we showed that matriptase, an endogenous serine protease, could potently increase TER. Using detergent solubility-based cell fractionation, we found that neither trypsin nor matriptase treatment changed levels of tight junction proteins at the membrane. In a fast calcium switch assay, serine proteases did not enhance the rate of recovery of the junction. In addition, serine proteases could not reverse barrier disruption induced by IFNγ and TNFα. We knocked down occludin in our cells using siRNA and found this prevented the serine protease-induced increase in TER. Using fluorescence recovery after photobleaching (FRAP), we found serine proteases induce a greater mobile fraction of occludin in the membrane. These data suggest that a functional tight junction is needed for serine proteases to have an effect on TER, and that occludin is a crucial tight junction protein in this mechanism. PMID:27492333

  2. [Kinetics of immunoglobulin G transport through the multi-layer epithelial-hematic barrier of the respiratory tract].

    PubMed

    Ermakov, N V; Krekhnov, B V; Chenchikova, E Iu; Cherchenko, N G; Ishkov, A G; Remizova, E N; Sveshnikov, P G; Miroshnichenko, E V; Nazarov, Iu V; Morozov, A P

    1991-05-01

    A new class of drugs is now utilized for in vivo diagnoses and therapy of many widespread diseases. In these pharmacological and diagnostic preparations the active substance is conjugated with a vector which transports the drug to specific biological targets. Monoclonal antibodies are the most commonly used vectors: estimation of their permeability through multilayer and unilayer biomembranes is an important step in the analysis of efficiency of vector drugs. Experiments with Sprague-Dawley rats (mature females weighing 500 to 160 g) have demonstrated the ability of immunoglobulins G to penetrate through the respiratory epithelial-hematic barrier. Using solid phase ELISA, it was found that 5-25% of the total amount of mouse antiinsulin immunoglobulins G1 injected into the trachea under hexenal anesthesia can penetrate into the blood plasma. Accumulation of antibodies in the blood begins 4 hours and ceases 32 hours after the drug application in a dose of 400 micrograms. The kinetics of transmembrane transport is described by an S-like saturation function: C(t) = Cmax/(1+e-(at-b]. Penetration of monoclonal antibodies into the blood is accompanied by their distribution in the organs and tissues as well as by their clearance from the blood plasma. The clearance of monoclonal antibodies is characterized by a 24 hour half-life and is described by an exponential equation: C(t) = C0 x exp-kt. An algorithm for the interaction of these processes which should be taken into account during measurements of the transport of monoclonal antibodies and their complexes through biomembranes is proposed.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1747414

  3. 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

  4. A role for IL-22 in the relationship between intestinal helminths, gut microbiota and mucosal immunity.

    PubMed

    Leung, Jacqueline M; Loke, P'ng

    2013-03-01

    The intestinal tract is home to nematodes as well as commensal bacteria (microbiota), which have coevolved with the mammalian host. The mucosal immune system must balance between an appropriate response to dangerous pathogens and an inappropriate response to commensal microbiota that may breach the epithelial barrier, in order to maintain intestinal homeostasis. IL-22 has been shown to play a critical role in maintaining barrier homeostasis against intestinal pathogens and commensal bacteria. Here we review the advances in our understanding of the role of IL-22 in helminth infections, as well as in response to commensal and pathogenic bacteria of the intestinal tract. We then consider the relationship between intestinal helminths and gut microbiota and hypothesize that this relationship may explain how helminths may improve symptoms of inflammatory bowel diseases. We propose that by inducing an immune response that includes IL-22, intestinal helminths may enhance the mucosal barrier function of the intestinal epithelium. This may restore the mucosal microbiota populations from dysbiosis associated with colitis and improve intestinal homeostasis.

  5. 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

  6. Vitamin D/VDR signaling attenuates lipopolysaccharide-induced acute lung injury by maintaining the integrity of the pulmonary epithelial barrier

    PubMed Central

    SHI, YONG-YAN; LIU, TIAN-JING; FU, JIAN-HUA; XU, WEI; WU, LIN-LIN; HOU, A-NA; XUE, XIN-DONG

    2016-01-01

    Vitamin D and its receptor have a protective effect on epithelial barriers in various tissues. Low levels of vitamin D are associated with numerous pulmonary diseases, including acute lung injury (ALI) and acute respiratory distress syndrome. The present study investigated whether the vitamin D/vitamin D receptor (VDR) pathway may ameliorate lipopolysaccharide (LPS)-induced ALI through maintaining the integrity of the alveolar epithelial barrier. This was investigated by exposing wild-type (WT) and VDR knockout C57BL/6J mice to LPS, then comparing the healthy and LPS-treated mice lungs and bronchoalveolar lavage fluid (BALF). More specifically, lung histology, mRNA levels of proinflammatory cytokines and chemokines, and protein expression levels of tight junction proteins were determined. In addition, a vitamin D analog (paricalcitol) was administered to WT mice in order to investigate the effect of vitamin D on the alveolar epithelial barrier following exposure to LPS. VDR knockout mice exhibited severe lung injuries (P<0.001), increased alveolar permeability [demonstrated by a higher wet-dry ratio of lung weight (P<0.05), greater expression levels of BALF protein (P<0.001) and fluorescein isothiocyanate-conjugated 4 kDa dextran (P<0.001) leakage into the alveolar space], elevated proinflammatory cytokine and chemokine mRNA levels, as demonstrated by reverse transcription-quantitative polymerase chain reaction (P<0.05), and decreased protein and mRNA expression levels of occludin (P<0.01) and zonula occludens-1 (ZO-1; P<0.01) compared with WT mice. Paricalcitol treatment partially inhibited these pathological changes in WT mice by maintaining the mRNA and protein expression levels of occludin (P<0.01) and ZO-1 (P<0.05). A lack of VDRs in the pulmonary epithelial barrier appeared to compromise its defense, leading to more severe LPS-induced lung injury. Furthermore, vitamin D treatment alleviated LPS-induced lung injury and preserved alveolar barrier function

  7. Vitamin D/VDR signaling attenuates lipopolysaccharide‑induced acute lung injury by maintaining the integrity of the pulmonary epithelial barrier.

    PubMed

    Shi, Yong-Yan; Liu, Tian-Jing; Fu, Jian-Hua; Xu, Wei; Wu, Lin-Lin; Hou, A-Na; Xue, Xin-Dong

    2016-02-01

    Vitamin D and its receptor have a protective effect on epithelial barriers in various tissues. Low levels of vitamin D are associated with numerous pulmonary diseases, including acute lung injury (ALI) and acute respiratory distress syndrome. The present study investigated whether the vitamin D/vitamin D receptor (VDR) pathway may ameliorate lipopolysaccharide (LPS)‑induced ALI through maintaining the integrity of the alveolar epithelial barrier. This was investigated by exposing wild‑type (WT) and VDR knockout C57BL/6J mice to LPS, then comparing the healthy and LPS‑treated mice lungs and bronchoalveolar lavage fluid (BALF). More specifically, lung histology, mRNA levels of proinflammatory cytokines and chemokines, and protein expression levels of tight junction proteins were determined. In addition, a vitamin D analog (paricalcitol) was administered to WT mice in order to investigate the effect of vitamin D on the alveolar epithelial barrier following exposure to LPS. VDR knockout mice exhibited severe lung injuries (P<0.001), increased alveolar permeability [demonstrated by a higher wet‑dry ratio of lung weight (P<0.05), greater expression levels of BALF protein (P<0.001) and fluorescein isothiocyanate‑conjugated 4 kDa dextran (P<0.001) leakage into the alveolar space], elevated proinflammatory cytokine and chemokine mRNA levels, as demonstrated by reverse transcription‑quantitative polymerase chain reaction (P<0.05), and decreased protein and mRNA expression levels of occludin (P<0.01) and zonula occludens‑1 (ZO‑1; P<0.01) compared with WT mice. Paricalcitol treatment partially inhibited these pathological changes in WT mice by maintaining the mRNA and protein expression levels of occludin (P<0.01) and ZO‑1 (P<0.05). A lack of VDRs in the pulmonary epithelial barrier appeared to compromise its defense, leading to more severe LPS‑induced lung injury. Furthermore, vitamin D treatment alleviated LPS‑induced lung injury and preserved alveolar

  8. Effects of feminine hygiene products on the vaginal mucosal biome

    PubMed Central

    Fashemi, Bisiayo; Delaney, Mary L.; Onderdonk, Andrew B.; Fichorova, Raina N.

    2013-01-01

    Background Over-the-counter (OTC) feminine hygiene products come with little warning about possible side effects. This study evaluates in-vitro their effects on Lactobacillus crispatus, which is dominant in the normal vaginal microbiota and helps maintain a healthy mucosal barrier essential for normal reproductive function and prevention of sexually transmitted infections and gynecologic cancer. Methods A feminine moisturizer (Vagisil), personal lubricant, and douche were purchased OTC. A topical spermicide (nonoxynol-9) known to alter the vaginal immune barrier was used as a control. L. crispatus was incubated with each product for 2 and 24h and then seeded on agar for colony forming units (CFU). Human vaginal epithelial cells were exposed to products in the presence or absence of L. crispatus for 24h, followed by epithelium-associated CFU enumeration. Interleukin-8 was immunoassayed and ANOVA was used for statistical evaluation. Results Nonoxynol-9 and Vagisil suppressed Lactobacillus growth at 2h and killed all bacteria at 24h. The lubricant decreased bacterial growth insignificantly at 2h but killed all at 24h. The douche did not have a significant effect. At full strength, all products suppressed epithelial viability and all, except the douche, suppressed epithelial-associated CFU. When applied at non-toxic dose in the absence of bacteria, the douche and moisturizer induced an increase of IL-8, suggesting a potential to initiate inflammatory reaction. In the presence of L. crispatus, the proinflammatory effects of the douche and moisturizer were countered, and IL-8 production was inhibited in the presence of the other products. Conclusion Some OTC vaginal products may be harmful to L. crispatus and alter the vaginal immune environment. Illustrated through these results, L. crispatus is essential in the preservation of the function of vaginal epithelial cells in the presence of some feminine hygiene products. More research should be invested toward these

  9. Airway and lung pathology due to mucosal surface dehydration in β-Epithelial Na+ Channel-overexpressing mice: role of TNFα and IL-4Rα signaling, influence of neonatal development, and limited efficacy of glucocorticoid treatment

    PubMed Central

    Livraghi, Alessandra; Grubb, Barbara R.; Hudson, Elizabeth J.; Wilkinson, Kristen J.; Sheehan, John K.; Mall, Marcus A.; O'Neal, Wanda K.; Boucher, Richard C.; Randell, Scott H.

    2009-01-01

    Overexpression of the epithelial Na+ channel β subunit (Scnn1b gene, βENaC protein) in transgenic (Tg) mouse airways dehydrates mucosal surfaces, producing mucus obstruction, inflammation, and neonatal mortality. Airway inflammation includes macrophage activation, neutrophil and eosinophil recruitment, and elevated KC, TNFα and chitinase levels. These changes recapitulate aspects of complex human obstructive airway diseases, but their molecular mechanisms are poorly understood. We used genetic and pharmacologic approaches to identify pathways relevant to the development of Scnn1b-Tg mouse lung pathology. Genetic deletion of tumor necrosis factor alpha (TNFα) or its receptor, TNFR1, had no measurable effect on the phenotype. Deletion of the interleukin-4 receptor alpha subunit (IL-4Rα) abolished transient mucous secretory cell (MuSC) abundance and eosinophilia normally observed in neonatal wild-type (WT) mice. Similarly, IL-4Rα deficiency decreased MuSC and eosinophils in neonatal Scnn1b-Tg mice, which correlated with improved neonatal survival. However, chronic lung pathology in adult Scnn1b-Tg mice was not affected by IL-4Rα status. Prednisolone treatment ablated eosinophilia and MuSC in adult Scnn1b-Tg mice, but did not decrease mucus plugging or neutrophilia. These studies demonstrate that: 1) normal neonatal mouse airway development entails an IL-4Rα-dependent, transient abundance of MuSC and eosinophils; 2) absence of IL-4Rα improved neonatal survival of Scnn1b-Tg mice, likely reflecting decreased formation of asphyxiating mucus plugs; and 3) in Scnn1b-Tg mice, neutrophilia, mucus obstruction, and airspace enlargement are IL-4Rα- and TNFα-independent, and only MuSC and eosinophilia are sensitive to glucocorticoids. Thus, manipulation of multiple pathways will likely be required to treat the complex pathogenesis caused by airway surface dehydration. PMID:19299736

  10. Characterization of the Probiotic Yeast Saccharomyces boulardii in the Healthy Mucosal Immune System.

    PubMed

    Hudson, Lauren E; McDermott, Courtney D; Stewart, Taryn P; Hudson, William H; Rios, Daniel; Fasken, Milo B; Corbett, Anita H; Lamb, Tracey J

    2016-01-01

    The probiotic yeast Saccharomyces boulardii has been shown to ameliorate disease severity in the context of many infectious and inflammatory conditions. However, use of S. boulardii as a prophylactic agent or therapeutic delivery vector would require delivery of S. boulardii to a healthy, uninflamed intestine. In contrast to inflamed mucosal tissue, the diverse microbiota, intact epithelial barrier, and fewer inflammatory immune cells within the healthy intestine may all limit the degree to which S. boulardii contacts and influences the host mucosal immune system. Understanding the nature of these interactions is crucial for application of S. boulardii as a prophylactic agent or therapeutic delivery vehicle. In this study, we explore both intrinsic and immunomodulatory properties of S. boulardii in the healthy mucosal immune system. Genomic sequencing and morphological analysis of S. boulardii reveals changes in cell wall components compared to non-probiotic S. cerevisiae that may partially account for probiotic functions of S. boulardii. Flow cytometry and immunohistochemistry demonstrate limited S. boulardii association with murine Peyer's patches. We also show that although S. boulardii induces a systemic humoral immune response, this response is small in magnitude and not directed against S. boulardii itself. RNA-seq of the draining mesenteric lymph nodes indicates that even repeated administration of S. boulardii induces few transcriptional changes in the healthy intestine. Together these data strongly suggest that interaction between S. boulardii and the mucosal immune system in the healthy intestine is limited, with important implications for future work examining S. boulardii as a prophylactic agent and therapeutic delivery vehicle.

  11. Multi-Faceted Functions of Secretory IgA at Mucosal Surfaces

    PubMed Central

    Corthésy, Blaise

    2013-01-01

    Secretory IgA (SIgA) plays an important role in the protection and homeostatic regulation of intestinal, respiratory, and urogenital mucosal epithelia separating the outside environment from the inside of the body. This primary function of SIgA is referred to as immune exclusion, a process that limits the access of numerous microorganisms and mucosal antigens to these thin and vulnerable mucosal barriers. SIgA has been shown to be involved in avoiding opportunistic pathogens to enter and disseminate in the systemic compartment, as well as tightly controlling the necessary symbiotic relationship existing between commensals and the host. Clearance by peristalsis appears thus as one of the numerous mechanisms whereby SIgA fulfills its function at mucosal surfaces. Sampling of antigen-SIgA complexes by microfold (M) cells, intimate contact occurring with Peyer’s patch dendritic cells (DC), down-regulation of inflammatory processes, modulation of epithelial, and DC responsiveness are some of the recently identified processes to which the contribution of SIgA has been underscored. This review aims at presenting, with emphasis at the biochemical level, how the molecular complexity of SIgA can serve these multiple and non-redundant modes of action. PMID:23874333

  12. Factor XIII Transglutaminase Supports the Resolution of Mucosal Damage in Experimental Colitis

    PubMed Central

    Andersson, Christina; Kvist, Peter H.; McElhinney, Kathryn; Baylis, Richard; Gram, Luise K.; Pelzer, Hermann; Lauritzen, Brian; Holm, Thomas L.; Hogan, Simon; Wu, David; Turpin, Brian; Miller, Whitney; Palumbo, Joseph S.

    2015-01-01

    The thrombin-activated transglutaminase factor XIII (FXIII) that covalently crosslinks and stablizes provisional fibrin matrices is also thought to support endothelial and epithelial barrier function and to control inflammatory processes. Here, gene-targeted mice lacking the FXIII catalytic A subunit were employed to directly test the hypothesis that FXIII limits colonic pathologies associated with experimental colitis. Wildtype (WT) and FXIII-/- mice were found to be comparable in their initial development of mucosal damage following exposure to dextran sulfate sodium (DSS) challenge. However, unlike FXIII-sufficient mice, FXIII-deficient cohorts failed to efficiently resolve colonic inflammatory pathologies and mucosal damage following withdrawal of DSS. Consistent with prior evidence of ongoing coagulation factor activation and consumption in individuals with active colitis, plasma FXIII levels were markedly decreased in colitis-challenged WT mice. Treatment of colitis-challenged mice with recombinant human FXIII-A zymogen significantly mitigated weight loss, intestinal bleeding, and diarrhea, regardless of whether cohorts were FXIII-sufficient or were genetically devoid of FXIII. Similarly, both qualitative and quantitative microscopic analyses of colonic tissues revealed that exogenous FXIII improved the resolution of multiple colitis disease parameters in both FXIII-/- and WT mice. The most striking differences were seen in the resolution of mucosal ulceration, the most severe histopathological manifestation of DSS-induced colitis. These findings directly demonstrate that FXIII is a significant determinant of mucosal healing and clinical outcome following inflammatory colitis induced mucosal injury and provide a proof-of-principle that clinical interventions supporting FXIII activity may be a means to limit colitis pathology and improve resolution of mucosal damage. PMID:26098308

  13. The effects of an epithelial barrier protective cationic aerosol on allergen-induced airway inflammation in asthma: a randomized, placebo-controlled clinical trial.

    PubMed

    Nair, P; Denis, S; Cancelliere, L; Radford, K; Efthimiadis, A; Rosano, M; Hanrahan, J

    2014-09-01

    Inhaled cationic airway lining modulator (iCALM) is a cationic aerosol therapy comprised of 1.29% calcium chloride dissolved in 0.9% isotonic saline that enhances the biophysical barrier function of the airway lining fluid and primes the host defense response. It's ability to attenuate bronchitis caused by inhaled particles was investigated using an allergen-inhalation model in a proof-of-concept study. In a randomized, double-blind, placebo-controlled cross-over trial of 6 mild atopic steroid-naïve asthmatic subjects, 3 doses of iCALM were well tolerated and they attenuated allergen-induced increase in sputum eosinophils, and levels of IL-5, MCP-1 and eotaxin. This study provides an opportunity to investigate the role of enhancing epithelial barrier to decrease airway inflammation provoked by inhaled particles in a variety of airway diseases.

  14. Immortalization of normal human mammary epithelial cells in two steps by direct targeting of senescence barriers does not require gross genomic alterations

    SciTech Connect

    Garbe, James C.; Vrba, Lukas; Sputova, Klara; Fuchs, Laura; Novak, Petr; Brothman, Arthur R.; Jackson, Mark; Chin, Koei; LaBarge, Mark A.; Watts, George; Futscher, Bernard W.; Stampfer, Martha R.

    2014-10-29

    Telomerase reactivation and immortalization are critical for human carcinoma progression. However, little is known about the mechanisms controlling this crucial step, due in part to the paucity of experimentally tractable model systems that can examine human epithelial cell immortalization as it might occur in vivo. We achieved efficient non-clonal immortalization of normal human mammary epithelial cells (HMEC) by directly targeting the 2 main senescence barriers encountered by cultured HMEC. The stress-associated stasis barrier was bypassed using shRNA to p16INK4; replicative senescence due to critically shortened telomeres was bypassed in post-stasis HMEC by c-MYC transduction. Thus, 2 pathologically relevant oncogenic agents are sufficient to immortally transform normal HMEC. The resultant non-clonal immortalized lines exhibited normal karyotypes. Most human carcinomas contain genomically unstable cells, with widespread instability first observed in vivo in pre-malignant stages; in vitro, instability is seen as finite cells with critically shortened telomeres approach replicative senescence. Our results support our hypotheses that: (1) telomere-dysfunction induced genomic instability in pre-malignant finite cells may generate the errors required for telomerase reactivation and immortalization, as well as many additional “passenger” errors carried forward into resulting carcinomas; (2) genomic instability during cancer progression is needed to generate errors that overcome tumor suppressive barriers, but not required per se; bypassing the senescence barriers by direct targeting eliminated a need for genomic errors to generate immortalization. Achieving efficient HMEC immortalization, in the absence of “passenger” genomic errors, should facilitate examination of telomerase regulation during human carcinoma progression, and exploration of agents that could prevent immortalization.

  15. Immortalization of normal human mammary epithelial cells in two steps by direct targeting of senescence barriers does not require gross genomic alterations

    DOE PAGES

    Garbe, James C.; Vrba, Lukas; Sputova, Klara; Fuchs, Laura; Novak, Petr; Brothman, Arthur R.; Jackson, Mark; Chin, Koei; LaBarge, Mark A.; Watts, George; et al

    2014-10-29

    Telomerase reactivation and immortalization are critical for human carcinoma progression. However, little is known about the mechanisms controlling this crucial step, due in part to the paucity of experimentally tractable model systems that can examine human epithelial cell immortalization as it might occur in vivo. We achieved efficient non-clonal immortalization of normal human mammary epithelial cells (HMEC) by directly targeting the 2 main senescence barriers encountered by cultured HMEC. The stress-associated stasis barrier was bypassed using shRNA to p16INK4; replicative senescence due to critically shortened telomeres was bypassed in post-stasis HMEC by c-MYC transduction. Thus, 2 pathologically relevant oncogenic agentsmore » are sufficient to immortally transform normal HMEC. The resultant non-clonal immortalized lines exhibited normal karyotypes. Most human carcinomas contain genomically unstable cells, with widespread instability first observed in vivo in pre-malignant stages; in vitro, instability is seen as finite cells with critically shortened telomeres approach replicative senescence. Our results support our hypotheses that: (1) telomere-dysfunction induced genomic instability in pre-malignant finite cells may generate the errors required for telomerase reactivation and immortalization, as well as many additional “passenger” errors carried forward into resulting carcinomas; (2) genomic instability during cancer progression is needed to generate errors that overcome tumor suppressive barriers, but not required per se; bypassing the senescence barriers by direct targeting eliminated a need for genomic errors to generate immortalization. Achieving efficient HMEC immortalization, in the absence of “passenger” genomic errors, should facilitate examination of telomerase regulation during human carcinoma progression, and exploration of agents that could prevent immortalization.« less

  16. Pseudomonas aeruginosa Outer Membrane Vesicles Triggered by Human Mucosal Fluid and Lysozyme Can Prime Host Tissue Surfaces for Bacterial Adhesion

    PubMed Central

    Metruccio, Matteo M. E.; Evans, David J.; Gabriel, Manal M.; Kadurugamuwa, Jagath L.; Fleiszig, Suzanne M. J.

    2016-01-01

    Pseudomonas aeruginosa is a leading cause of human morbidity and mortality that often targets epithelial surfaces. Host immunocompromise, or the presence of indwelling medical devices, including contact lenses, can predispose to infection. While medical devices are known to accumulate bacterial biofilms, it is not well understood why resistant epithelial surfaces become susceptible to P. aeruginosa. Many bacteria, including P. aeruginosa, release outer membrane vesicles (OMVs) in response to stress that can fuse with host cells to alter their function. Here, we tested the hypothesis that mucosal fluid can trigger OMV release to compromise an epithelial barrier. This was tested using tear fluid and corneal epithelial cells in vitro and in vivo. After 1 h both human tear fluid, and the tear component lysozyme, greatly enhanced OMV release from P. aeruginosa strain PAO1 compared to phosphate buffered saline (PBS) controls (∼100-fold). Transmission electron microscopy (TEM) and SDS-PAGE showed tear fluid and lysozyme-induced OMVs were similar in size and protein composition, but differed from biofilm-harvested OMVs, the latter smaller with fewer proteins. Lysozyme-induced OMVs were cytotoxic to human corneal epithelial cells in vitro and murine corneal epithelium in vivo. OMV exposure in vivo enhanced Ly6G/C expression at the corneal surface, suggesting myeloid cell recruitment, and primed the cornea for bacterial adhesion (∼4-fold, P < 0.01). Sonication disrupted OMVs retained cytotoxic activity, but did not promote adhesion, suggesting the latter required OMV-mediated events beyond cell killing. These data suggest that mucosal fluid induced P. aeruginosa OMVs could contribute to loss of epithelial barrier function during medical device-related infections. PMID:27375592

  17. TGF-β1 improves mucosal IgA dysfunction and dysbiosis following intestinal ischaemia-reperfusion in mice.

    PubMed

    Zhang, Xu-Yu; Liu, Zi-Meng; Zhang, Hu-Fei; Li, Yun-Sheng; Wen, Shi-Hong; Shen, Jian-Tong; Huang, Wen-Qi; Liu, Ke-Xuan

    2016-06-01

    Intestinal ischaemia/reperfusion (I/R) severely disrupts gut barriers and leads to high mortality in the critical care setting. Transforming growth factor (TGF)-β1 plays a pivotal role in intestinal cellular and immune regulation. However, the effects of TGF-β1 on intestinal I/R injury remain unclear. Thus, we aimed to investigate the effects of TGF-β1 on gut barriers after intestinal I/R and the molecular mechanisms. Intestinal I/R model was produced in mice by clamping the superior mesenteric artery for 1 hr followed by reperfusion. Recombinant TGF-β1 was intravenously infused at 15 min. before ischaemia. The results showed that within 2 hrs after reperfusion, intestinal I/R disturbed intestinal immunoglobulin A class switch recombination (IgA CSR), the key process of mucosal IgA synthesis, and resulted in IgA dysfunction, as evidenced by decreased production and bacteria-binding capacity of IgA. Meanwhile, the disruptions of intestinal microflora and mucosal structure were exhibited. Transforming growth factor-β1 activated IgA CSR as evidenced by the increased activation molecules and IgA precursors. Strikingly, TGF-β1 improved intestinal mucosal IgA dysfunction, dysbiosis and epithelial damage at the early stage after reperfusion. In addition, SB-431542, a specific inhibitor of activating mothers against decapentaplegic homologue (SMAD) 2/3, totally blocked the inductive effect of TGF-β1 on IgA CSR and almost abrogated the above protective effects on intestinal barriers. Taken together, our study demonstrates that TGF-β1 protects intestinal mucosal IgA immunity, microbiota and epithelial integrity against I/R injury mainly through TGF-β receptor 1/SMAD 2/3 pathway. Induction of IgA CSR may be involved in the protection conferred by TGF-β1.

  18. TcpC protein from E. coli Nissle improves epithelial barrier function involving PKCζ and ERK1/2 signaling in HT-29/B6 cells.

    PubMed

    Hering, N A; Richter, J F; Fromm, A; Wieser, A; Hartmann, S; Günzel, D; Bücker, R; Fromm, M; Schulzke, J D; Troeger, H

    2014-03-01

    The probiotic Escherichia coli Nissle 1917 (EcN) is widely used to maintain remission in ulcerative colitis. This is thought to be mediated by various immunomodulatory and barrier-stabilizing effects in the intestine. In this study, the mechanisms of barrier modulation by EcN were studied in the human epithelial HT-29/B6 cell culture model.EcN supernatant increased transepithelial resistance (TER) and reduced permeability to mannitol because of sealing of the paracellular passage pathway as revealed by two-path impedance spectroscopy. This increase in TER was attributed to the TcpC protein of EcN. TcpC induced protein kinase C-ζ (PKCζ) and extracellular-signal-regulated kinase 1/2 (ERK1/2) phosphorylation, which in turn resulted in upregulation of the barrier-forming tight junction protein claudin-14. By specific silencing of protein expression by small interfering RNA (siRNA), the sealing function of claudin-14 was confirmed. In conclusion, the TcpC protein of EcN affects innate immunity by improving intestinal barrier function through upregulation of claudin-14 via PKCζ and ERK1/2 signaling. PMID:23900194

  19. 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

  20. Structure and barrier properties of human embryonic stem cell-derived retinal pigment epithelial cells are affected by extracellular matrix protein coating.

    PubMed

    Sorkio, Anni; Hongisto, Heidi; Kaarniranta, Kai; Uusitalo, Hannu; Juuti-Uusitalo, Kati; Skottman, Heli

    2014-02-01

    Extracellular matrix (ECM) interactions play a vital role in cell morphology, migration, proliferation, and differentiation of cells. We investigated the role of ECM proteins on the structure and function of human embryonic stem cell-derived retinal pigment epithelial (hESC-RPE) cells during their differentiation and maturation from hESCs into RPE cells in adherent differentiation cultures on several human ECM proteins found in native human Bruch's membrane, namely, collagen I, collagen IV, laminin, fibronectin, and vitronectin, as well as on commercial substrates of xeno-free CELLstart™ and Matrigel™. Cell pigmentation, expression of RPE-specific proteins, fine structure, as well as the production of basal lamina by hESC-RPE on different protein coatings were evaluated after 140 days of differentiation. The integrity of hESC-RPE epithelium and barrier properties on different coatings were investigated by measuring transepithelial resistance. All coatings supported the differentiation of hESC-RPE cells as demonstrated by early onset of cell pigmentation and further maturation to RPE monolayers after enrichment. Mature RPE phenotype was verified by RPE-specific gene and protein expression, correct epithelial polarization, and phagocytic activity. Significant differences were found in the degree of RPE cell pigmentation and tightness of epithelial barrier between different coatings. Further, the thickness of self-assembled basal lamina and secretion of the key ECM proteins found in the basement membrane of the native RPE varied between hESC-RPE cultured on compared protein coatings. In conclusion, this study shows that the cell culture substrate has a major effect on the structure and basal lamina production during the differentiation and maturation of hESC-RPE potentially influencing the success of cell integrations and survival after cell transplantation.

  1. Intra-subtype variation in enteroadhesion accounts for differences in epithelial barrier disruption and is associated with metronidazole resistance in Blastocystis subtype-7.

    PubMed

    Wu, Zhaona; Mirza, Haris; Tan, Kevin Shyong Wei

    2014-05-01

    Blastocystis is an extracellular, enteric pathogen that induces intestinal disorders in a range of hosts including humans. Recent studies have identified potential parasite virulence factors in and host responses to this parasite; however, little is known about Blastocystis-host attachment, which is crucial for colonization and virulence of luminal stages. By utilizing 7 different strains of the parasite belonging to two clinically relevant subtypes ST-4 and ST-7, we investigated Blastocystis-enterocyte adhesion and its association with parasite-induced epithelial barrier disruption. We also suggest that drug resistance in ST-7 strains might result in fitness cost that manifested as impairment of parasite adhesion and, consequently, virulence. ST-7 parasites were generally highly adhesive to Caco-2 cells and preferred binding to intercellular junctions. These strains also induced disruption of ZO-1 and occludin tight junction proteins as well as increased dextran-FITC flux across epithelial monolayers. Interestingly, their adhesion was correlated with metronidazole (Mz) susceptibility. Mz resistant (Mzr) strains were found to be less pathogenic, owing to compromised adhesion. Moreover, tolerance of nitrosative stress was also reduced in the Mzr strains. In conclusion, the findings indicate that Blastocystis attaches to intestinal epithelium and leads to epithelial barrier dysfunction and that drug resistance might entail a fitness cost in parasite virulence by limiting entero-adhesiveness. This is the first study of the cellular basis for strain-to-strain variation in parasite pathogenicity. Intra- and inter-subtype variability in cytopathogenicity provides a possible explanation for the diverse clinical outcomes of Blastocystis infections.

  2. Immunology of Gut Mucosal Vaccines

    PubMed Central

    Pasetti, Marcela F.; Simon, Jakub K.; Sztein, Marcelo B.; Levine, Myron M.

    2011-01-01

    Summary Understanding the mechanisms underlying the induction of immunity in the gastrointestinal mucosa following oral immunization and the cross-talk between mucosal and systemic immunity should expedite the development of vaccines to diminish the global burden caused by enteric pathogens. Identifying an immunological correlate of protection in the course of field trials of efficacy, animal models (when available), or human challenge studies is also invaluable. In industrialized country populations, live attenuated vaccines (e.g. polio, typhoid, and rotavirus) mimic natural infection and generate robust protective immune responses. In contrast, a major challenge is to understand and overcome the barriers responsible for the diminished immunogenicity and efficacy of the same enteric vaccines in underprivileged populations in developing countries. Success in developing vaccines against some enteric pathogens has heretofore been elusive (e.g. Shigella). Different types of oral vaccines can selectively or inclusively elicit mucosal secretory immunoglobulin A and serum immunoglobulin G antibodies and a variety of cell-mediated immune responses. Areas of research that require acceleration include interaction between the gut innate immune system and the stimulation of adaptive immunity, development of safe yet effective mucosal adjuvants, better understanding of homing to the mucosa of immunologically relevant cells, and elicitation of mucosal immunologic memory. This review dissects the immune responses elicited in humans by enteric vaccines. PMID:21198669

  3. The role of immunomodulators on intestinal barrier homeostasis in experimental models.

    PubMed

    Andrade, Maria Emília Rabelo; Araújo, Raquel Silva; de Barros, Patrícia Aparecida Vieira; Soares, Anne Danieli Nascimento; Abrantes, Fernanda Alves; Generoso, Simone de Vasconcelos; Fernandes, Simone Odília Antunes; Cardoso, Valbert Nascimento

    2015-12-01

    The intestinal epithelium is composed of specialized epithelial cells that form a physical and biochemical barrier to commensal and pathogenic microorganisms. However, dysregulation of the epithelial barrier function can lead to increased intestinal permeability and bacterial translocation across the intestinal mucosa, which contributes to local and systemic immune activation. The increase in these parameters is associated with inflammatory bowel disease, physical exercise under heat stress, intestinal obstruction, ischemia, and mucositis, among other conditions. Lately, there has been growing interest in immunomodulatory nutrients and probiotics that can regulate host immune and inflammatory responses and possibly restore the intestinal barrier. Immunomodulators such as amino acids (glutamine, arginine, tryptophan, and citrulline), fatty acids (short-chain and omega-3 fatty acids and conjugated linoleic acids), and probiotics (Bifidobacterium, Saccharomyces, and Lactobacillus) have been reported in the literature. Here, we review the critical roles of immunomodulatory nutrients in supporting gut barrier integrity and function. PMID:25660317

  4. Evidence for a plasma membrane-mediated permeability barrier to Tat basic domain in well-differentiated epithelial cells: lack of correlation with heparan sulfate.

    PubMed

    Violini, Stefania; Sharma, Vijay; Prior, Julie L; Dyszlewski, Mary; Piwnica-Worms, David

    2002-10-22

    Membrane permeation peptides, such as Tat basic domain, have emerged as useful membrane transduction agents with potential utility in therapeutic delivery and diagnostic imaging. While generally thought to universally permeate all cells by a nonselective process, the mechanism of membrane transduction remains poorly characterized. To examine vectorial transport properties of Tat basic domain in well-differentiated epithelial cells possessing tight junctions, L and D stereoisomers of Tat(48-57) peptide conjugates labeled with (99m)Tc were quantitatively analyzed in confluent monolayers of MDCK renal epithelial and CaCo-2 colonic carcinoma cells grown in transwell configurations. In both cell lines, vectorial transepithelial apparent permeability coefficients (P(app)) for L- and D-[(99m)Tc]Tat-peptides ranged from 30 to 70 nm/s, comparable to values for the macromolecular impermeant marker inulin in both apical-to-basolateral and basolateral-to-apical directions, but 100-fold less than the P(app) values for propranolol, a highly permeable control compound. Upon direct instillation of [(99m)Tc]Tat-peptide into the urinary bladder of living rats in vivo, no transepithelial permeation into other tissues was identified. Furthermore, MDCK and CaCo-2 cells showed a complete lack of intracellular accumulation of fluorescein conjugated Tat-peptide. However, translocation into cells was induced by treatment with plasma membrane permeabilizing agents such as digitonin and acetone/methanol, while cholesterol depletion with beta-methyl-cyclodextrin and metabolic inhibition with CCCP or 4 degrees C showed no effect. By contrast, in Hela and KB 3-1 cells, epithelial lines that do not form tight junctions in monolayer culture, baseline cytoplasmic and nucleolar accumulation was readily observed. Because all four cell lines expressed heparan sulfate proteoglycans, putative receptors for Tat basic peptides, we found no correlation between heparan sulfate and the permeation barrier

  5. MicroRNA-346 Mediates Tumor Necrosis Factor-α-induced Down-Regulation of Gut Epithelial Vitamin D Receptor in Inflammatory Bowel Diseases

    PubMed Central

    Chen, Yunzi; Du, Jie; Zhang, Zhongyi; Liu, Tianjing; Shi, Yongyan; Ge, Xin; Li, Yan Chun

    2015-01-01

    Background We recently reported that the gut epithelial vitamin D receptor (VDR) signaling inhibits colitis through inhibition of intestinal epithelial cell apoptosis, and the level of colonic epithelial VDR is markedly reduced in patients with inflammatory bowel diseases (IBD). VDR down-regulation promotes colitis, but the mechanism underlying VDR down-regulation in IBD is unknown. Methods VDR expression was analyzed in colon cancer cells under pro-inflammatory cytokine treatment. VDR as a target of miR-346 was confirmed using colon cancer cell culture. The relationship among inflammation, miR-346 and VDR was assessed in human IBD biopsies and experimental colitis models. Results We showed that TNF-α suppresses VDR expression while simultaneously up-regulating miR-346 in human colon cancer cells. Further studies demonstrated that miR-346 inhibits VDR by a specific target sequence in the 3′ untranslated region of the human VDR transcript, and blockade of miR-346 with a hairpin inhibitor abrogates the ability of TNF-α to inhibit VDR, confirming that TNF-α down-regulates VDR by inducing miR-346. Consistently, in human IBD biopsies the reduction of epithelial VDR is associated with increased immune cell infiltration and elevation of TNF-α and miR-346. In an experimental model of colitis mucosal VDR expression is reduced over time with the progression of colitis, inversely correlated with the induction of TNF-α and miR-346 in the mucosa. Conclusion These data suggest that during mucosal inflammation TNF-α induces miR-346, which down-regulates epithelial VDR. Mucosal VDR reduction in turn compromises the integrity of the mucosal epithelial barrier, further driving mucosal inflammation and colitis development. PMID:25192497

  6. 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

  7. The inner foreskin of healthy males at risk of HIV infection harbors epithelial CD4+ CCR5+ cells and has features of an inflamed epidermal barrier.

    PubMed

    Lemos, Maria P; Lama, Javier R; Karuna, Shelly T; Fong, Youyi; Montano, Silvia M; Ganoza, Carmela; Gottardo, Raphael; Sanchez, Jorge; McElrath, M Juliana

    2014-01-01

    Male circumcision provides partial protection against multiple sexually transmitted infections (STIs), including HIV, but the mechanisms are not fully understood. To examine potential vulnerabilities in foreskin epithelial structure, we used Wilcoxon paired tests adjusted using the false discovery rate method to compare inner and outer foreskin samples from 20 healthy, sexually active Peruvian males who have sex with males or transgender females, ages 21-29, at elevated risk of HIV infection. No evidence of epithelial microtrauma was identified, as assessed by keratinocyte activation, fibronectin deposition, or parakeratosis. However, multiple suprabasal tight junction differences were identified: 1) inner foreskin stratum corneum was thinner than outer (p = 0.035); 2) claudin 1 had extended membrane-bound localization throughout inner epidermis stratum spinosum (p = 0.035); 3) membrane-bound claudin 4 was absent from inner foreskin stratum granulosum (p = 0.035); and 4) occludin had increased membrane deposition in inner foreskin stratum granulosum (p = 0.042) versus outer. Together, this suggests subclinical inflammation and paracellular transport modifications to the inner foreskin. A setting of inflammation was further supported by inner foreskin epithelial explant cultures secreting higher levels of GM-CSF (p = 0.029), IP-10 (p = 0.035) and RANTES (p = 0.022) than outer foreskin, and also containing an increased density of CCR5+ and CD4+ CCR5+ cells (p = 0.022). Inner foreskin dermis also secreted more RANTES than outer (p = 0.036), and had increased density of CCR5+ cells (p = 0.022). In conclusion, subclinical changes to the inner foreskin of sexually active males may support an inflammatory state, with availability of target cells for HIV infection and modifications to epidermal barriers, potentially explaining the benefits of circumcision for STI prevention. PMID:25268493

  8. The Inner Foreskin of Healthy Males at Risk of HIV Infection Harbors Epithelial CD4+ CCR5+ Cells and Has Features of an Inflamed Epidermal Barrier

    PubMed Central

    Lemos, Maria P.; Lama, Javier R.; Karuna, Shelly T.; Fong, Youyi; Montano, Silvia M.; Ganoza, Carmela; Gottardo, Raphael; Sanchez, Jorge; McElrath, M. Juliana

    2014-01-01

    Male circumcision provides partial protection against multiple sexually transmitted infections (STIs), including HIV, but the mechanisms are not fully understood. To examine potential vulnerabilities in foreskin epithelial structure, we used Wilcoxon paired tests adjusted using the false discovery rate method to compare inner and outer foreskin samples from 20 healthy, sexually active Peruvian males who have sex with males or transgender females, ages 21–29, at elevated risk of HIV infection. No evidence of epithelial microtrauma was identified, as assessed by keratinocyte activation, fibronectin deposition, or parakeratosis. However, multiple suprabasal tight junction differences were identified: 1) inner foreskin stratum corneum was thinner than outer (p = 0.035); 2) claudin 1 had extended membrane-bound localization throughout inner epidermis stratum spinosum (p = 0.035); 3) membrane-bound claudin 4 was absent from inner foreskin stratum granulosum (p = 0.035); and 4) occludin had increased membrane deposition in inner foreskin stratum granulosum (p = 0.042) versus outer. Together, this suggests subclinical inflammation and paracellular transport modifications to the inner foreskin. A setting of inflammation was further supported by inner foreskin epithelial explant cultures secreting higher levels of GM-CSF (p = 0.029), IP-10 (p = 0.035) and RANTES (p = 0.022) than outer foreskin, and also containing an increased density of CCR5+ and CD4+ CCR5+ cells (p = 0.022). Inner foreskin dermis also secreted more RANTES than outer (p = 0.036), and had increased density of CCR5+ cells (p = 0.022). In conclusion, subclinical changes to the inner foreskin of sexually active males may support an inflammatory state, with availability of target cells for HIV infection and modifications to epidermal barriers, potentially explaining the benefits of circumcision for STI prevention. PMID:25268493

  9. The inner foreskin of healthy males at risk of HIV infection harbors epithelial CD4+ CCR5+ cells and has features of an inflamed epidermal barrier.

    PubMed

    Lemos, Maria P; Lama, Javier R; Karuna, Shelly T; Fong, Youyi; Montano, Silvia M; Ganoza, Carmela; Gottardo, Raphael; Sanchez, Jorge; McElrath, M Juliana

    2014-01-01

    Male circumcision provides partial protection against multiple sexually transmitted infections (STIs), including HIV, but the mechanisms are not fully understood. To examine potential vulnerabilities in foreskin epithelial structure, we used Wilcoxon paired tests adjusted using the false discovery rate method to compare inner and outer foreskin samples from 20 healthy, sexually active Peruvian males who have sex with males or transgender females, ages 21-29, at elevated risk of HIV infection. No evidence of epithelial microtrauma was identified, as assessed by keratinocyte activation, fibronectin deposition, or parakeratosis. However, multiple suprabasal tight junction differences were identified: 1) inner foreskin stratum corneum was thinner than outer (p = 0.035); 2) claudin 1 had extended membrane-bound localization throughout inner epidermis stratum spinosum (p = 0.035); 3) membrane-bound claudin 4 was absent from inner foreskin stratum granulosum (p = 0.035); and 4) occludin had increased membrane deposition in inner foreskin stratum granulosum (p = 0.042) versus outer. Together, this suggests subclinical inflammation and paracellular transport modifications to the inner foreskin. A setting of inflammation was further supported by inner foreskin epithelial explant cultures secreting higher levels of GM-CSF (p = 0.029), IP-10 (p = 0.035) and RANTES (p = 0.022) than outer foreskin, and also containing an increased density of CCR5+ and CD4+ CCR5+ cells (p = 0.022). Inner foreskin dermis also secreted more RANTES than outer (p = 0.036), and had increased density of CCR5+ cells (p = 0.022). In conclusion, subclinical changes to the inner foreskin of sexually active males may support an inflammatory state, with availability of target cells for HIV infection and modifications to epidermal barriers, potentially explaining the benefits of circumcision for STI prevention.

  10. Effect of nonylphenol ethoxylates (NPEs) on barrier functions of epithelial cell membranes: opening of tight junctions and competitive inhibition of P-gp-mediated efflux.

    PubMed

    Doo, Min-Ho; Li, Hong; Jang, Hye-In; Song, Im-Sook; Chung, Suk-Jae; Shim, Chang-Koo

    2005-09-30

    The effect of nonylphenol ethoxylates (NPEs) on selected barrier functions of biological membranes, such as tight junction and P-gp efflux pump of epithelial membranes, against the transport of xenobiotics was examined. The Caco-2 cell line was used to evaluate the transport of mannitol and daunomycin across the cell monolayer as well as the cellular uptake of daunomycin. In the presence of NPEs, the transport of mannitol was increased, with NP-9 showing a maximal effect, and the transepithelial electrical resistance (TEER) was reduced. The onset of this effect of NP-9 was fairly rapid and reversible for a short term (e.g., 2 h) treatment, while irreversible for a long term (e.g., 72 h) treatment. In the presence of NP-9, the apical uptake of daunomycin was increased, suggesting competitive inhibition between NP-9 and daunomycin in the efflux via the P-gp system. However, a 72 h pretreatment of the cells with NP-9 (up to 1000 nM) did not affect the apparent cellular uptake of daunomycin, suggesting no significant effect of NPEs on the expression of P-gp. In conclusion, NPEs appear to rapidly open the tight junction of epithelial cell membranes and to competitively inhibit the efflux of P-gp substrates, thereby reducing the self-protection ability of the organism against xenobiotics or hazardous environmental compounds that are transported via the paracellular pathway (i.e., uptake) or the P-gp system (i.e., efflux).

  11. Collective epithelial cell invasion overcomes mechanical barriers of collagenous extracellular matrix by a narrow tube-like geometry and MMP14-dependent local softening†

    PubMed Central

    Alcaraz, Jordi; Mori, Hidetoshi; Ghajar, Cyrus M.; Brownfield, Doug; Galgoczy, Roland; Bissell, Mina J.

    2013-01-01

    Collective cell invasion (CCI) through interstitial collagenous extracellular matrix (ECM) is crucial to the initial stages of branching morphogenesis, and a hallmark of tissue repair and dissemination of certain tumors. The collagenous ECM acts as a mechanical barrier against CCI. However, the physical nature of this barrier and how it is overcome by cells remains incompletely understood. To address these questions, we performed theoretical and experimental analysis of mammary epithelial branching morphogenesis in 3D type I collagen (collagen-I) gels. We found that the mechanical resistance of collagen-I is largely due to its elastic rather than its viscous properties. We also identified two strategies utilized by mammary epithelial cells that can independently minimize ECM mechanical resistance during CCI. First, cells adopt a narrow tube-like geometry during invasion, which minimizes the elastic opposition from the ECM as revealed by theoretical modeling of the most frequent invasive shapes and sizes. Second, the stiffness of the collagenous ECM is reduced at invasive fronts due to its degradation by matrix metalloproteinases (MMPs), as indicated by direct measurements of collagen-I microelasticity by atomic force microscopy. Molecular techniques further specified that the membrane-bound MMP14 mediates degradation of collagen-I at invasive fronts. Thus, our findings reveal that MMP14 is necessary to efficiently reduce the physical restraints imposed by collagen-I during branching morphogenesis, and help our overall understanding of how forces are balanced between cells and their surrounding ECM to maintain collective geometry and mechanical stability during CCI. PMID:21993836

  12. Mucosal permeability and immune activation as potential therapeutic targets of probiotics in irritable bowel syndrome.

    PubMed

    Barbara, Giovanni; Zecchi, Lisa; Barbaro, Raffaella; Cremon, Cesare; Bellacosa, Lara; Marcellini, Marco; De Giorgio, Roberto; Corinaldesi, Roberto; Stanghellini, Vincenzo

    2012-10-01

    There is increasingly convincing evidence supporting the participation of the gut microenvironment in the pathophysiology of irritable bowel syndrome (IBS). Studies particularly suggest an interplay between luminal factors (eg, foods and bacteria residing in the intestine), the epithelial barrier, and the mucosal immune system. Decreased expression and structural rearrangement of tight junction proteins in the small bowel and colon leading to increased intestinal permeability have been observed, particularly in postinfectious IBS and in IBS with diarrhea. These abnormalities are thought to contribute to the outflow of antigens through the leaky epithelium, causing overstimulation of the mucosal immune system. Accordingly, subsets of patients with IBS show higher numbers and an increased activation of mucosal immunocytes, particularly mast cells. Immune factors, released by these cells, including proteases, histamine, and prostanoids, participate in the perpetuation of the permeability dysfunction and contribute to the activation of abnormal neural responses involved in abdominal pain perception and changes in bowel habits. All these mechanisms represent new targets for therapeutic approaches in IBS. Probiotics are an attractive therapeutic option in IBS given their recognized safety and by virtue of positive biological effects they can exert on the host. Of importance for the IBS pathophysiology is that preclinical studies have shown that selective probiotic strains exhibit potentially useful properties including anti-inflammatory effects, improvement of mucosal barrier homeostasis, beneficial effects on intestinal microbiota, and a reduction of visceral hypersensitivity. The effect of probiotics on IBS is positive in most randomized, controlled studies, although the gain over the placebo is small. Identifying tailored probiotic approaches for subgroups of IBS patients represents a challenge for the future.

  13. Glycoprotein A33 deficiency: a new mouse model of impaired intestinal epithelial barrier function and inflammatory disease.

    PubMed

    Williams, Benjamin B; Tebbutt, Niall C; Buchert, Michael; Putoczki, Tracy L; Doggett, Karen; Bao, Shisan; Johnstone, Cameron N; Masson, Frederick; Hollande, Frederic; Burgess, Antony W; Scott, Andrew M; Ernst, Matthias; Heath, Joan K

    2015-08-01

    The cells of the intestinal epithelium provide a selectively permeable barrier between the external environment and internal tissues. The integrity of this barrier is maintained by tight junctions, specialised cell-cell contacts that permit the absorption of water and nutrients while excluding microbes, toxins and dietary antigens. Impairment of intestinal barrier function contributes to multiple gastrointestinal disorders, including food hypersensitivity, inflammatory bowel disease (IBD) and colitis-associated cancer (CAC). Glycoprotein A33 (GPA33) is an intestinal epithelium-specific cell surface marker and member of the CTX group of transmembrane proteins. Roles in cell-cell adhesion have been demonstrated for multiple CTX family members, suggesting a similar function for GPA33 within the gastrointestinal tract. To test a potential requirement for GPA33 in intestinal barrier function, we generated Gpa33(-/-) mice and subjected them to experimental regimens designed to produce food hypersensitivity, colitis and CAC. Gpa33(-/-) mice exhibited impaired intestinal barrier function. This was shown by elevated steady-state immunosurveillance in the colonic mucosa and leakiness to oral TRITC-labelled dextran after short-term exposure to dextran sodium sulphate (DSS) to injure the intestinal epithelium. Gpa33(-/-) mice also exhibited rapid onset and reduced resolution of DSS-induced colitis, and a striking increase in the number of colitis-associated tumours produced by treatment with the colon-specific mutagen azoxymethane (AOM) followed by two cycles of DSS. In contrast, Gpa33(-/-) mice treated with AOM alone showed no increase in sporadic tumour formation, indicating that their increased tumour susceptibility is dependent on inflammatory stimuli. Finally, Gpa33(-/-) mice displayed hypersensitivity to food allergens, a common co-morbidity in humans with IBD. We propose that Gpa33(-/-) mice provide a valuable model to study the mechanisms linking intestinal

  14. Intestinal epithelial vitamin D receptor signaling inhibits experimental colitis.

    PubMed

    Liu, Weicheng; Chen, Yunzi; Golan, Maya Aharoni; Annunziata, Maria L; Du, Jie; Dougherty, Urszula; Kong, Juan; Musch, Mark; Huang, Yong; Pekow, Joel; Zheng, Changqing; Bissonnette, Marc; Hanauer, Stephen B; Li, Yan Chun

    2013-09-01

    The inhibitory effects of vitamin D on colitis have been previously documented. Global vitamin D receptor (VDR) deletion exaggerates colitis, but the relative anticolitic contribution of epithelial and nonepithelial VDR signaling is unknown. Here, we showed that colonic epithelial VDR expression was substantially reduced in patients with Crohn's disease or ulcerative colitis. Moreover, targeted expression of human VDR (hVDR) in intestinal epithelial cells (IECs) protected mice from developing colitis. In experimental colitis models induced by 2,4,6-trinitrobenzenesulfonic acid, dextran sulfate sodium, or CD4(+)CD45RB(hi) T cell transfer, transgenic mice expressing hVDR in IECs were highly resistant to colitis, as manifested by marked reductions in clinical colitis scores, colonic histological damage, and colonic inflammation compared with WT mice. Reconstitution of Vdr-deficient IECs with the hVDR transgene completely rescued Vdr-null mice from severe colitis and death, even though the mice still maintained a hyperresponsive Vdr-deficient immune system. Mechanistically, VDR signaling attenuated PUMA induction in IECs by blocking NF-κB activation, leading to a reduction in IEC apoptosis. Together, these results demonstrate that gut epithelial VDR signaling inhibits colitis by protecting the mucosal epithelial barrier, and this anticolitic activity is independent of nonepithelial immune VDR actions.

  15. Multiple roles for keratin intermediate filaments in the regulation of epithelial barrier function and apico-basal polarity

    PubMed Central

    Salas, Pedro J.; Forteza, Radia; Mashukova, Anastasia

    2016-01-01

    abstract As multicellular organisms evolved a family of cytoskeletal proteins, the keratins (types I and II) expressed in epithelial cells diversified in more than 20 genes in vertebrates. There is no question that keratin filaments confer mechanical stiffness to cells. However, such a number of genes can hardly be explained by evolutionary advantages in mechanical features. The use of transgenic mouse models has revealed unexpected functional relationships between keratin intermediate filaments and intracellular signaling. Accordingly, loss of keratins or mutations in keratins that cause or predispose to human diseases, result in increased sensitivity to apoptosis, regulation of innate immunity, permeabilization of tight junctions, and mistargeting of apical proteins in different epithelia. Precise mechanistic explanations for these phenomena are still lacking. However, immobilization of membrane or cytoplasmic proteins, including chaperones, on intermediate filaments (“scaffolding”) appear as common molecular mechanisms and may explain the need for so many different keratin genes in vertebrates. PMID:27583190

  16. Multiple roles for keratin intermediate filaments in the regulation of epithelial barrier function and apico-basal polarity.

    PubMed

    Salas, Pedro J; Forteza, Radia; Mashukova, Anastasia

    2016-01-01

    As multicellular organisms evolved a family of cytoskeletal proteins, the keratins (types I and II) expressed in epithelial cells diversified in more than 20 genes in vertebrates. There is no question that keratin filaments confer mechanical stiffness to cells. However, such a number of genes can hardly be explained by evolutionary advantages in mechanical features. The use of transgenic mouse models has revealed unexpected functional relationships between keratin intermediate filaments and intracellular signaling. Accordingly, loss of keratins or mutations in keratins that cause or predispose to human diseases, result in increased sensitivity to apoptosis, regulation of innate immunity, permeabilization of tight junctions, and mistargeting of apical proteins in different epithelia. Precise mechanistic explanations for these phenomena are still lacking. However, immobilization of membrane or cytoplasmic proteins, including chaperones, on intermediate filaments ("scaffolding") appear as common molecular mechanisms and may explain the need for so many different keratin genes in vertebrates. PMID:27583190

  17. Reconstituting the barrier properties of a water-tight epithelial membrane by design of leaflet-specific liposomes.

    PubMed

    Hill, W G; Zeidel, M L

    2000-09-29

    To define aspects of lipid composition and bilayer asymmetry critical to barrier function, we examined the permeabilities of liposomes that model individual leaflets of the apical membrane of a barrier epithelium, Madin-Darby canine kidney type 1 cells. Using published lipid compositions we prepared exofacial liposomes containing phosphatidylcholine, sphingomyelin, glycosphingolipids, and cholesterol; and cytoplasmic liposomes containing phosphatidylethanolamine, phosphatidylserine, and cholesterol. The osmotic permeability of cytoplasmic liposomes to water (P(f)), solutes, and NH(3) was 18-90-fold higher than for the exofacial liposomes (P(f(ex)) = 2.4 +/- 0.4 x 10(-4) cm/s, P(f(cy)) = 4.4 +/- 0.3 x 10(-3) cm/s; P(glycerol(ex)) = 2.5 +/- 0.3 x 10(-8) cm/s, P(glycerol(cy)) = 2.2 +/- 0.02 x 10(-6) cm/s; P(NH3(ex)) = 0. 13 +/- 0.4 x 10(-4) cm/s, P(NH3(cy)) = 7.9 +/- 1.0 x 10(-3) cm/s). By contrast, the apparent proton permeability of exofacial liposomes was 4-fold higher than cytoplasmic liposomes (P(H+(ex)) = 1.1 +/- 0. 1 x 10(-2) cm/s, P(H+(cy)) = 2.7 +/- 0.6 x 10(-3) cm/s). By adding single leaflet permeabilities, we calculated a theoretical P(f) for a Madin-Darby canine kidney apical membrane of 4.6 x 10(-4) cm/s, which compares favorably with experimentally determined values. In exofacial liposomes lacking glycosphingolipids or sphingomyelin, permeabilities were 2-7-fold higher, indicating that both species play a role in barrier function. Removal of cholesterol resulted in 40-280-fold increases in permeability. We conclude: 1) that we have reconstituted the biophysical properties of a barrier membrane, 2) that the barrier resides in the exofacial leaflet, 3) that both sphingomyelin and glycosphingolipids play a role in reducing membrane permeability but that there is an absolute requirement for cholesterol to mediate this effect, 4) that these results further validate the hypothesis that each leaflet offers an independent resistance to permeation, and 5) that

  18. Impact of exogenous lipase supplementation on growth, intestinal function, mucosal immune and physical barrier, and related signaling molecules mRNA expression of young grass carp (Ctenopharyngodon idella).

    PubMed

    Liu, Sen; Feng, Lin; Jiang, Wei-Dan; Liu, Yang; Jiang, Jun; Wu, Pei; Zeng, Yun-Yun; Xu, Shu-De; Kuang, Sheng-Yao; Tang, Ling; Tang, Wu-Neng; Zhang, Yong-An; Zhou, Xiao-Qiu

    2016-08-01

    This study investigated the effects of exogenous lipase supplementation on the growth performance, intestinal growth and function, immune response and physical barrier function, and related signaling molecules mRNA expression of young grass carp (Ctenopharyngodon idella). A total of 450 grass carp (255.02 ± 0.34 g) were fed five diets for 60 days. There were 5 dietary treatments that included a normal protein and lipid diet containing 30% crude protein (CP) with 5% ether extract (EE), and the low-protein and high-lipid diets (28% CP, 6% EE) supplemented with graded levels of exogenous lipase supplementation activity at 0, 1193, 2560 and 3730 U/kg diet. The results indicated that compared with a normal protein and lipid diet (30% CP, 5% EE), a low-protein and high-lipid diet (28% CP, 6% EE) (un-supplemented lipase) improved lysozyme activities and complement component 3 contents in the distal intestine (DI), interleukin 10 mRNA expression in the proximal intestine (PI), and glutathione S-transferases activity and glutathione content in the intestine of young grass carp. In addition, in low-protein and high-lipid diets, optimal exogenous lipase supplementation significantly increased acid phosphatase (ACP) activities and complement component 3 (C3) contents (P < 0.05), up-regulated the relative mRNA levels of antimicrobial peptides (liver expressed antimicrobial peptide 2 and hepcidin) and anti-inflammatory cytokines (interleukin 10 and transforming growth factor β1) and signaling molecules inhibitor protein-κBα (IκBα) and target of rapamycin (TOR) (P < 0.05), down-regulated the mRNA levels of pro-inflammatory cytokines (tumor necrosis factor α, interleukin 8, interferon γ2, and interleukin 1β), and signaling molecules (nuclear factor kappa B p65, IκB kinase β, IκB kinase γ) (P < 0.05) in the intestine of young grass carp. Moreover, optimal exogenous lipase supplementation significantly decreased reactive oxygen species (ROS), malondialdehyde

  19. Lymphocytes accelerate epithelial tight junction assembly: role of AMP-activated protein kinase (AMPK).

    PubMed

    Tang, Xiao Xiao; Chen, Hao; Yu, Sidney; Zhang, Li; Caplan, Michael J; Chan, Hsiao Chang

    2010-01-01

    The tight junctions (TJs), characteristically located at the apicolateral borders of adjacent epithelial cells, are required for the proper formation of epithelial cell polarity as well as for sustaining the mucosal barrier to the external environment. The observation that lymphocytes are recruited by epithelial cells to the sites of infection [1] suggests that they may play a role in the modulation of epithelial barrier function and thus contribute to host defense. To test the ability of lymphocytes to modulate tight junction assembly in epithelial cells, we set up a lymphocyte-epithelial cell co-culture system, in which Madin-Darby canine kidney (MDCK) cells, a well-established model cell line for studying epithelial TJ assembly [2], were co-cultured with mouse lymphocytes to mimic an infection state. In a typical calcium switch experiment, the TJ assembly in co-culture was found to be accelerated compared to that in MDCK cells alone. This accelaration was found to be mediated by AMP-activated protein kinase (AMPK). AMPK activation was independent of changes in cellular ATP levels but it was found to be activated by the pro-inflammatory cytokine TNF-alpha. Forced suppression of AMPK, either with a chemical inhibitor or by knockdown, abrogated the accelerating effect of lymphocytes on TJ formation. Similar results were also observed in a co-culture with lymphocytes and Calu-3 human airway epithelial cells, suggesting that the activation of AMPK may be a general mechanism underlying lymphocyte-accelerated TJ assembly in different epithelia. These results suggest that signals from lymphocytes, such as cytokines, facilitate TJ assembly in epithelial cells via the activation of AMPK. PMID:20808811

  20. Helicobacter pylori antigens, acetylsalicylic acid, LDL and 7-ketocholesterol - their potential role in destabilizing the gastric epithelial cell barrier. An in vitro model of Kato III cells.

    PubMed

    Gajewski, Adrian; Mnich, Eliza; Szymański, Karol; Hinc, Krzysztof; Obuchowski, Michał; Moran, Anthony P; Chmiela, Magdalena

    2016-01-01

    Colonization of gastric tissue in humans by H. pylori Gram-negative bacteria initiates gastric and duodenal ulcers and even gastric cancers. Infections promote inflammation and damage to gastric epithelium which might be followed by the impairment of its barrier function. The role of H. pylori components in these processes has not been specified. H. pylori cytotoxicity may potentially increase in the milieu of anti-inflammatory drugs including acetylsalicylic acid (ASA). The lipid transport-associated molecule such as low density lipoprotein (LDL), which is a classic risk factor of coronary heart disease (CHD) and 7-ketocholesterol (7-kCh) a product of cholesterol oxidation, which may occur during the oxidative stress in LDL could also be considered as pro-inflammatory. The aim of this study was to evaluate the cytotoxicity of H. pylori antigens, ASA, LDL and 7-kCh towards Kato III gastric epithelial cells, on the basis of the cell ability to reduce tetrazolium salt (MTT) and morphology of cell nuclei assessed by 4',6-diamidino-2-phenylindole (DAPI) staining. Kato III cells were stimulated for 24 h, at 37°C and 5% CO2, with H. pylori antigens: cytotoxin associated gene A (CagA) protein, the urease A subunit (UreA), lipopolysaccharide (LPS) and ASA, LDL or 7-kCh. H. pylori LPS, ASA, LDL and 7-kCh, but not H. pylori glycine acid extract (GE), demonstrated cytotoxicity against Kato III cells, which was related to a diminished percentage of MTT reducing cells and to an increased cell population with the signs of DNA damage. The results suggest that damage to gastric epithelial cells can be induced independently by H. pylori antigens, ASA and endogenous lipid transport-associated molecules. During H. pylori infection in vivo, especially in CHD patients, synergistic or antagonistic interactions between these factors might possibly influence the disease course. Further study is necessary to explain these potential effects. PMID:26619253

  1. Pathogen Resistance Mediated by IL-22 Signaling at the Epithelial-Microbiota Interface.

    PubMed

    Schreiber, Fernanda; Arasteh, Julia Maryam; Lawley, Trevor D

    2015-11-20

    Intestinal colonization resistance to bacterial pathogens is generally associated, among other factors, with mucosal homeostasis that preserves the integrity of the intestinal barrier. Mucosal homeostasis depends on physical and molecular interactions between three components: the resident microbiota, the epithelial layer and the local immune system. The cytokine IL-22 helps to orchestrate this three-way interaction. IL-22 is produced by immune cells present beneath the epithelium and is induced by bacteria present in the intestine. IL-22 stimulates the epithelial cells via the IL-22RA1-IL-10R2 receptor complex inducing changes in the expression of genes involved in the maintenance of epithelial barrier integrity, with a variety of functions in pathogen resistance such as mucus layer modifications and hydration, tight junction fortification and the production of a broad range of bactericidal compounds. These mechanisms of pathogen resistance, in turn, affect the microbiota composition and create an environment that excludes pathogens. Here we highlight the role of IL-22 as key mediator in the give-and-take relationship between the microbiota and the host that impacts pathogen resistance.

  2. The role of oral flora in the development of chemotherapy-induced oral mucositis.

    PubMed

    Stringer, Andrea M; Logan, Richard M

    2015-02-01

    Chemotherapy-induced mucositis is considered to be a major oncological problem, caused by the cytotoxic effects of cancer chemotherapy. In the last 10 years, there have been significant advances in the understanding of mucositis pathobiology. At the basic level, it is now well-understood that it is not just an epithelial process, but rather a complex interaction between epithelial and connective tissue compartments. There is also potential interaction between the oral microenvironment and the development of mucositis. Changes occur in the resident oral flora (commensal) throughout cancer treatment, and it is conceivable that these organisms and changes that occur may have an influence on the development of mucosal toxicity associated with cancer treatment. The aim of this review was to examine the potential contributions of oral microflora in the pathobiology of mucositis and identify pathways and interactions that could be targeted for therapeutic management of mucositis.

  3. A coculture model mimicking the intestinal mucosa reveals a regulatory role for myofibroblasts in immune-mediated barrier disruption.

    PubMed

    Willemsen, L E M; Schreurs, C C H M; Kroes, H; Spillenaar Bilgen, E J; Van Deventer, S J H; Van Tol, E A F

    2002-10-01

    The pathogenesis of Crohn's disease involves a mucosal inflammatory response affecting the barrier function of the gut. Myofibroblasts directly underlining the intestinal epithelium may have a regulatory role in immune-mediated barrier disruption. A coculture system of T84 epithelial and CCD-18Co myofibroblasts was established in order to mimic the in situ spatial interactions between these cell types and to evaluate their role in barrier: integrity. Lamina propria mononuclear cells (LPMC) were introduced in co- and monocultures. Effects of immune cells on barrier integrity was determined by measuring resistance and permeability for macromolecules. Introduction of LPMC in both culture systems caused a time-dependent decrease in barrier integrity. This was found to be less pronounced in cocultures indicating a regulatory role for mesenchymal cells. The effects were also found to depend on the route of LPMC stimulation. Additional analyses suggested that the regulatory role of myofibroblasts in barrier integrity involves production of growth factors. PMID:12395905

  4. Oral mucositis - self-care

    MedlinePlus

    Cancer treatment - mucositis; Cancer treatment - mouth pain; Cancer treatment - mouth sores; Chemotherapy - mucositis; Chemotherapy - mouth pain; Chemotherapy - mouth sores; Radiation therapy - mucositis; Radiation therapy - mouth pain; Radiation therapy - mouth ...

  5. Regulation of the Intestinal Barrier Function by Host Defense Peptides.

    PubMed

    Robinson, Kelsy; Deng, Zhuo; Hou, Yongqing; Zhang, Guolong

    2015-01-01

    Intestinal barrier function is achieved primarily through regulating the synthesis of mucins and tight junction (TJ) proteins, which are critical for maintaining optimal gut health and animal performance. An aberrant expression of TJ proteins results in increased paracellular permeability, leading to intestinal and systemic disorders. As an essential component of innate immunity, host defense peptides (HDPs) play a critical role in mucosal defense. Besides broad-spectrum antimicrobial activities, HDPs promotes inflammation resolution, endotoxin neutralization, wound healing, and the development of adaptive immune response. Accumulating evidence has also indicated an emerging role of HDPs in barrier function and intestinal homeostasis. HDP deficiency in the intestinal tract is associated with barrier dysfunction and dysbiosis. Several HDPs were recently shown to enhance mucosal barrier function by directly inducing the expression of multiple mucins and TJ proteins. Consistently, dietary supplementation of HDPs often leads to an improvement in intestinal morphology, production performance, and feed efficiency in livestock animals. This review summarizes current advances on the regulation of epithelial integrity and homeostasis by HDPs. Major signaling pathways mediating HDP-induced mucin and TJ protein synthesis are also discussed. As an alternative strategy to antibiotics, supplementation of exogenous HDPs or modulation of endogenous HDP synthesis may have potential to improve intestinal barrier function and animal health and productivity. PMID:26664984

  6. Regulation of the Intestinal Barrier Function by Host Defense Peptides

    PubMed Central

    Robinson, Kelsy; Deng, Zhuo; Hou, Yongqing; Zhang, Guolong

    2015-01-01

    Intestinal barrier function is achieved primarily through regulating the synthesis of mucins and tight junction (TJ) proteins, which are critical for maintaining optimal gut health and animal performance. An aberrant expression of TJ proteins results in increased paracellular permeability, leading to intestinal and systemic disorders. As an essential component of innate immunity, host defense peptides (HDPs) play a critical role in mucosal defense. Besides broad-spectrum antimicrobial activities, HDPs promotes inflammation resolution, endotoxin neutralization, wound healing, and the development of adaptive immune response. Accumulating evidence has also indicated an emerging role of HDPs in barrier function and intestinal homeostasis. HDP deficiency in the intestinal tract is associated with barrier dysfunction and dysbiosis. Several HDPs were recently shown to enhance mucosal barrier function by directly inducing the expression of multiple mucins and TJ proteins. Consistently, dietary supplementation of HDPs often leads to an improvement in intestinal morphology, production performance, and feed efficiency in livestock animals. This review summarizes current advances on the regulation of epithelial integrity and homeostasis by HDPs. Major signaling pathways mediating HDP-induced mucin and TJ protein synthesis are also discussed. As an alternative strategy to antibiotics, supplementation of exogenous HDPs or modulation of endogenous HDP synthesis may have potential to improve intestinal barrier function and animal health and productivity. PMID:26664984

  7. Mucosal immunization and adjuvants.

    PubMed

    Hasegawa, Hideki; van Reit, Elly; Kida, Hiroshi

    2015-01-01

    The goal of the influenza vaccine is to prevent influenza virus infection and control the yearly seasonal epidemic and pandemic. However, the presently available parenteral influenza vaccine induces only systemic humoral immunity, which does not prevent influenza virus infection on the mucosal surface. Secretary IGA antibodies play an important role in preventing natural infection. Moreover, the IgA antibody response mediates cross-protection against variant viruses in animal models. Thus, a mucosal influenza vaccine that induces mucosal immunity would be a powerful tool to protect individuals from the influenza virus. Although the function of the mucosal immune system, especially in the respiratory tract, is not completely understood, there are several studies underway to develop mucosal influenza vaccines. Here, we will review current knowledge concerning the induction of IgA, the role of B-cell production of influenza virus specific IgA antibodies in anti-influenza immunity, and the role of humoral memory responses induced upon vaccination.

  8. 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

  9. A Gut Microbial Metabolite of Linoleic Acid, 10-Hydroxy-cis-12-octadecenoic Acid, Ameliorates Intestinal Epithelial Barrier Impairment Partially via GPR40-MEK-ERK Pathway*

    PubMed Central

    Miyamoto, Junki; Mizukure, Taichi; Park, Si-Bum; Kishino, Shigenobu; Kimura, Ikuo; Hirano, Kanako; Bergamo, Paolo; Rossi, Mauro; Suzuki, Takuya; Arita, Makoto; Ogawa, Jun; Tanabe, Soichi

    2015-01-01

    Gut microbial metabolites of polyunsaturated fatty acids have attracted much attention because of their various physiological properties. Dysfunction of tight junction (TJ) in the intestine contributes to the pathogenesis of many disorders such as inflammatory bowel disease. We evaluated the effects of five novel gut microbial metabolites on tumor necrosis factor (TNF)-α-induced barrier impairment in Caco-2 cells and dextran sulfate sodium-induced colitis in mice. 10-Hydroxy-cis-12-octadecenoic acid (HYA), a gut microbial metabolite of linoleic acid, suppressed TNF-α and dextran sulfate sodium-induced changes in the expression of TJ-related molecules, occludin, zonula occludens-1, and myosin light chain kinase. HYA also suppressed the expression of TNF receptor 2 (TNFR2) mRNA and protein expression in Caco-2 cells and colonic tissue. In addition, HYA suppressed the protein expression of TNFR2 in murine intestinal epithelial cells. Furthermore, HYA significantly up-regulated G protein-coupled receptor (GPR) 40 expression in Caco-2 cells. It also induced [Ca2+]i responses in HEK293 cells expressing human GPR40 with higher sensitivity than linoleic acid, its metabolic precursor. The barrier-recovering effects of HYA were abrogated by a GPR40 antagonist and MEK inhibitor in Caco-2 cells. Conversely, 10-hydroxyoctadacanoic acid, which is a gut microbial metabolite of oleic acid and lacks a carbon-carbon double bond at Δ12 position, did not show these TJ-restoring activities and down-regulated GPR40 expression. Therefore, HYA modulates TNFR2 expression, at least partially, via the GPR40-MEK-ERK pathway and may be useful in the treatment of TJ-related disorders such as inflammatory bowel disease. PMID:25505251

  10. Maturation of the Intestinal Epithelial Barrier in Neonatal Rats Coincides with Decreased FcRn Expression, Replacement of Vacuolated Enterocytes and Changed Blimp-1 Expression

    PubMed Central

    Arévalo Sureda, Ester; Weström, Björn; Pierzynowski, Stefan G.; Prykhodko, Olena

    2016-01-01

    Background The intestinal barrier is immature in newborn mammals allowing for transfer of bioactive macromolecules, e.g. protecting antibodies, from mother’s milk to the blood circulation and in neonatal rodents lasts until weaning. This passage involves the neonatal-Fc-receptor (FcRn) binding IgG in the proximal and highly endocytic vacuolated enterocytes in the distal immature small intestine (SI). Recent studies have suggested an involvement of the transcription factor B-lymphocyte-induced maturation-protein-1 (Blimp-1) in the regulation of SI maturation in mice. Hence, the objective of the present study was to monitor the development of the intestinal barrier function, in relation to Blimp-1 expression during both natural and precociously induced intestinal maturation in rats. Results During the suckling period IgG plasma levels increased, while after gut closure it temporarily decreased. This corresponded to a high expression of FcRn in the proximal SI epithelium and the presence of vacuolated enterocytes in the distal SI. The immature foetal-type epithelium was replaced after weaning or induced precocious maturation, by an adult-type epithelium with FcRnneg cells in the proximal and by non-vacuolated enterocytes in the distal SI. In parallel to this epithelial shift, Blimp-1 expression decreased in the distal SI. Conclusion The switch from foetal- to adult-type epithelium, with decreased proximal expression of FcRn and distal replacement of vacuolated enterocytes, was concurrent in the two SI regions and could be used for monitoring SI maturation in the rat. The changes in expression of Blimp-1 in the distal SI epithelium followed the maturation pattern. PMID:27736989

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

    PubMed

    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.

  12. Characterization of the Probiotic Yeast Saccharomyces boulardii in the Healthy Mucosal Immune System.

    PubMed

    Hudson, Lauren E; McDermott, Courtney D; Stewart, Taryn P; Hudson, William H; Rios, Daniel; Fasken, Milo B; Corbett, Anita H; Lamb, Tracey J

    2016-01-01

    The probiotic yeast Saccharomyces boulardii has been shown to ameliorate disease severity in the context of many infectious and inflammatory conditions. However, use of S. boulardii as a prophylactic agent or therapeutic delivery vector would require delivery of S. boulardii to a healthy, uninflamed intestine. In contrast to inflamed mucosal tissue, the diverse microbiota, intact epithelial barrier, and fewer inflammatory immune cells within the healthy intestine may all limit the degree to which S. boulardii contacts and influences the host mucosal immune system. Understanding the nature of these interactions is crucial for application of S. boulardii as a prophylactic agent or therapeutic delivery vehicle. In this study, we explore both intrinsic and immunomodulatory properties of S. boulardii in the healthy mucosal immune system. Genomic sequencing and morphological analysis of S. boulardii reveals changes in cell wall components compared to non-probiotic S. cerevisiae that may partially account for probiotic functions of S. boulardii. Flow cytometry and immunohistochemistry demonstrate limited S. boulardii association with murine Peyer's patches. We also show that although S. boulardii induces a systemic humoral immune response, this response is small in magnitude and not directed against S. boulardii itself. RNA-seq of the draining mesenteric lymph nodes indicates that even repeated administration of S. boulardii induces few transcriptional changes in the healthy intestine. Together these data strongly suggest that interaction between S. boulardii and the mucosal immune system in the healthy intestine is limited, with important implications for future work examining S. boulardii as a prophylactic agent and therapeutic delivery vehicle. PMID:27064405

  13. Characterization of the Probiotic Yeast Saccharomyces boulardii in the Healthy Mucosal Immune System

    PubMed Central

    Hudson, Lauren E.; McDermott, Courtney D.; Stewart, Taryn P.; Hudson, William H.; Rios, Daniel; Fasken, Milo B.; Corbett, Anita H.; Lamb, Tracey J.

    2016-01-01

    The probiotic yeast Saccharomyces boulardii has been shown to ameliorate disease severity in the context of many infectious and inflammatory conditions. However, use of S. boulardii as a prophylactic agent or therapeutic delivery vector would require delivery of S. boulardii to a healthy, uninflamed intestine. In contrast to inflamed mucosal tissue, the diverse microbiota, intact epithelial barrier, and fewer inflammatory immune cells within the healthy intestine may all limit the degree to which S. boulardii contacts and influences the host mucosal immune system. Understanding the nature of these interactions is crucial for application of S. boulardii as a prophylactic agent or therapeutic delivery vehicle. In this study, we explore both intrinsic and immunomodulatory properties of S. boulardii in the healthy mucosal immune system. Genomic sequencing and morphological analysis of S. boulardii reveals changes in cell wall components compared to non-probiotic S. cerevisiae that may partially account for probiotic functions of S. boulardii. Flow cytometry and immunohistochemistry demonstrate limited S. boulardii association with murine Peyer’s patches. We also show that although S. boulardii induces a systemic humoral immune response, this response is small in magnitude and not directed against S. boulardii itself. RNA-seq of the draining mesenteric lymph nodes indicates that even repeated administration of S. boulardii induces few transcriptional changes in the healthy intestine. Together these data strongly suggest that interaction between S. boulardii and the mucosal immune system in the healthy intestine is limited, with important implications for future work examining S. boulardii as a prophylactic agent and therapeutic delivery vehicle. PMID:27064405

  14. 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

  15. Helicobacter pylori impedes acid-induced tightening of gastric epithelial junctions

    PubMed Central

    Marcus, Elizabeth A.; Vagin, Olga; Tokhtaeva, Elmira; Sachs, George

    2013-01-01

    Gastric infection by Helicobacter pylori is the most common cause of ulcer disease and gastric cancer. The mechanism of progression from gastritis and inflammation to ulcers and cancer in a fraction of those infected is not definitively known. Significant acidity is unique to the gastric environment and is required for ulcer development. The interplay between gastric acidity and H. pylori pathogenesis is important in progression to advanced disease. The aim of this study was to characterize the impact of acid on gastric epithelial integrity and cytokine release and how H. pylori infection alters these responses. Human gastric epithelial (HGE-20) cells were grown on porous inserts, and survival, barrier function, and cytokine release were studied at various apical pH levels in the presence and absence of H. pylori. With apical acidity, gastric epithelial cells demonstrate increased barrier function, as evidenced by increased transepithelial electrical resistance (TEER) and decreased paracellular permeability. This effect is reduced in the presence of wild-type, but not urease knockout, H. pylori. The epithelial inflammatory response is also modulated by acidity and H. pylori infection. Without H. pylori, epithelial IL-8 release decreases in acid, while IL-6 release increases. In the presence of H. pylori, acidic pH diminishes the magnitude of the previously reported increase in IL-8 and IL-6 release. H. pylori interferes with the gastric epithelial response to acid, contributing to altered barrier function and inflammatory response. H. pylori diminishes acid-induced tightening of cell junctions in a urease-dependent manner, suggesting that local pH elevation promotes barrier compromise and progression to mucosal damage. PMID:23989011

  16. CUGBP1 and HuR regulate E-cadherin translation by altering recruitment of E-cadherin mRNA to processing bodies and modulate epithelial barrier function.

    PubMed

    Yu, Ting-Xi; Gu, Bei-Lin; Yan, Jun-Kai; Zhu, Jie; Yan, Wei-Hui; Chen, Jie; Qian, Lin-Xi; Cai, Wei

    2016-01-01

    The effectiveness and stability of epithelial barrier depend on apical junctional complexes, which consist of tight junctions (TJs) and adherens junctions (AJs). E-cadherin is the primary component of AJs, and it is essential for maintenance of cell-to-cell interactions and regulates the epithelial barrier. However, the exact mechanism underlying E-cadherin expression, particularly at the posttranscriptional level, remains largely unknown. RNA-binding proteins CUG-binding protein 1 (CUGBP1) and HU antigen R (HuR) are highly expressed in the intestinal epithelial tissues and modulate the stability and translation of target mRNAs. Here, we present evidence that CUGBP1 and HuR interact directly with the 3'-untranslated region of E-cadherin mRNA and regulate E-cadherin translation. CUGBP1 overexpression in Caco-2 cells inhibited E-cadherin translation by increasing the recruitment of E-cadherin mRNA to processing bodies (PBs), thus resulting in an increase in paracellular permeability. Overexpression of HuR exhibited an opposite effect on E-cadherin expression by preventing the translocation of E-cadherin mRNA to PBs and therefore prevented CUGBP1-induced repression of E-cadherin expression. Elevation of HuR also abolished the CUGBP1-induced epithelial barrier dysfunction. These findings indicate that CUGBP1 and HuR negate each other's effects in regulating E-cadherin translation by altering the recruitment of E-cadherin mRNA to PBs and play an important role in the regulation of intestinal barrier integrity under various pathophysiological conditions.

  17. Nasal mucosal biopsy

    MedlinePlus

    Biopsy - nasal mucosa; Nose biopsy ... to fast for a few hours before the biopsy. ... Nasal mucosal biopsy is usually done when abnormal tissue is seen during examination of the nose. It may also be done ...

  18. Epithelial Transport in Inflammatory Bowel Diseases

    PubMed Central

    Ghishan, Fayez K.; Kiela, Pawel R.

    2014-01-01

    The epithelium of the gastrointestinal tract is one of the most versatile tissues in the organism, responsible for providing a tight barrier between dietary and bacterial antigens and the mucosal and systemic immune system, while maintaining efficient digestive and absorptive processes to ensure adequate nutrient and energy supply. Inflammatory Bowel Diseases (IBD; Crohn’s disease and ulcerative colitis) are associated with a breakdown of both functions, which in some cases are clearly interrelated. In this updated literature review, we focus on the effects of intestinal inflammation and the associated immune mediators on selected aspects of the transepithelial transport of macro- and micronutrients. The mechanisms responsible for nutritional deficiencies are not always clear and could be related to decreased intake, malabsorption and excess losses. We summarize the known causes of nutrient deficiencies and the mechanism of IBD-associated diarrhea. We also overview the consequences of impaired epithelial transport, which infrequently transcend its primary purpose to affect the gut microbial ecology and epithelial integrity. While some of those regulatory mechanisms are relatively well established, more work needs to be done to determine how inflammatory cytokines can alter the transport process of nutrients across the gastrointestinal and renal epithelia. PMID:24691115

  19. Use of Transparent Film Dressing for Dermoscopy of Mucosal Lesions.

    PubMed

    Sorrell, Jennifer; Lauren, Christine T

    2016-01-01

    Genital and mucosal nevi are not uncommonly encountered in children. These type of nevi highlight challenges in performing a thorough dermoscopic examination. Contact dermoscopy can provide additional information about a nevus that non-contact dermoscopy cannot. We propose applying a sterile transparent film dressing over the dermatoscope to act as a waterproof barrier that is also impermeable to bacteria and viruses. This provides a sanitary way to evaluate nevi in genital skin as well as on other mucosal surfaces. PMID:26250593

  20. 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

  1. Symbiotic Bacterial Metabolites Regulate Gastrointestinal Barrier Function via the Xenobiotic Sensor PXR and Toll-like Receptor 4

    PubMed Central

    Venkatesh, Madhukumar; Mukherjee, Subhajit; Wang, Hongwei; Li, Hao; Sun, Katherine; Benechet, Alaxandre P.; Qiu, Zhijuan; Maher, Leigh; Redinbo, Matthew R.; Phillips, Robert S.; Fleet, James C.; Kortagere, Sandhya; Mukherjee, Paromita; Fasano, Alessio; Le Ven, Jessica; Nicholson, Jeremy K.; Dumas, Marc E.; Khanna, Kamal M.; Mani, Sridhar

    2014-01-01

    SUMMARY Intestinal microbial metabolites are conjectured to affect mucosal integrity through an incompletely characterized mechanism. Here we showed microbial-specific indoles regulated intestinal barrier function through the xenobiotic sensor, pregnane X receptor (PXR). Indole 3-propionic acid (IPA), in the context of indole, is as a ligand for PXR in vivo, and IPA down-regulated enterocyte TNF–α while up-regulated junctional protein-coding mRNAs. PXR-deficient (Nr1i2−/−) mice showed a distinctly “leaky” gut physiology coupled with up-regulation of the Toll-like receptor (TLR) signaling pathway. These defects in the epithelial barrier were corrected in Nr1i2−/−Tlr4−/− mice. Our results demonstrate that a direct chemical communication between the intestinal symbionts and PXR regulates mucosal integrity through a pathway which involves luminal sensing and signaling by TLR4. PMID:25065623

  2. 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

  3. NOD-Like Receptors in Intestinal Homeostasis and Epithelial Tissue Repair

    PubMed Central

    Parlato, Marianna; Yeretssian, Garabet

    2014-01-01

    The intestinal epithelium constitutes a dynamic physical barrier segregating the luminal content from the underlying mucosal tissue. Following injury, the epithelial integrity is restored by rapid migration of intestinal epithelial cells (IECs) across the denuded area in a process known as wound healing. Hence, through a sequence of events involving restitution, proliferation and differentiation of IECs the gap is resealed and homeostasis reestablished. Relapsing damage followed by healing of the inflamed mucosa is a hallmark of several intestinal disorders including inflammatory bowel diseases (IBD). While several regulatory peptides, growth factors and cytokines stimulate restitution of the epithelial layer after injury, recent evidence in the field underscores the contribution of innate immunity in controlling this process. In particular, nucleotide-binding and oligomerization domain-like receptors (NLRs) play critical roles in sensing the commensal microbiota, maintaining homeostasis, and regulating intestinal inflammation. Here, we review the process of intestinal epithelial tissue repair and we specifically focus on the impact of NLR-mediated signaling mechanisms involved in governing epithelial wound healing during disease. PMID:24886810

  4. Role of mucus in ischemia/reperfusion-induced gastric mucosal injury in rats.

    PubMed

    Mojzis, J; Hegedüsová, R; Mirossay, L

    2000-01-01

    Gastric mucus plays an important role in gastric mucosal protection. Apart from its "barrier" function, it has been demonstrated that mucus protects gastric epithelial cells against toxic oxygen metabolites derived from the xanthine/ xanthine oxidase system. In this study, we investigated the effect of malotilate and sucralfate (mucus production stimulators) and N-acetylcysteine (mucolytic agent) on ischemia/reperfusion-induced gastric mucosal injury. Gastric ischemia was induced by 30 min clamping of the coeliac artery followed by 30 min of reperfusion. The mucus content was determined by the Alcian blue method. Sucralfate (100 mg/kg), malotilate (100 mg/kg), and N-acetylcysteine (100 mg/kg) were given orally 30 min before surgery. Both sucralfate and malotilate increased the mucus production in control rats. On the other hand, N-acetyloysteine significantly decreased mucus content in control (sham) group. A significant decrease of mucus content was found in the control and the N-acetylcysteine pretreated group during the period of ischemia. On the other hand, sucralfate and malotilate prevented the decrease the content of mucus during ischemia. A similar result can be seen after ischemia/reperfusion. In the control group and N-acetylcysteine pretreated group a significant decrease of adherent mucus content was found. However, sucralfate and malotilate increased mucus production (sucralfate significantly). Sucralfate and malotilate also significantly protected the gastric mucosa against ischemia/reperfusion-induced injury. However, N-acetylcysteine significantly increased gastric mucosal injury after ischemia/reperfusion. These results suggest that gastric mucus may be involved in the protection of gastric mucosa after ischemia/reperfusion.

  5. Mucosal healing in inflammatory bowel diseases: a systematic review.

    PubMed

    Neurath, Markus F; Travis, Simon P L

    2012-11-01

    Recent studies have identified mucosal healing on endoscopy as a key prognostic parameter in the management of inflammatory bowel diseases (IBD), thus highlighting the role of endoscopy for monitoring of disease activity in IBD. In fact, mucosal healing has emerged as a key treatment goal in IBD that predicts sustained clinical remission and resection-free survival of patients. The structural basis of mucosal healing is an intact barrier function of the gut epithelium that prevents translocation of commensal bacteria into the mucosa and submucosa with subsequent immune cell activation. Thus, mucosal healing should be considered as an initial event in the suppression of inflammation of deeper layers of the bowel wall, rather than as a sign of complete healing of gut inflammation. In this systematic review, the clinical studies on mucosal healing are summarised and the effects of anti-inflammatory or immunosuppressive drugs such as 5-aminosalicylates, corticosteroids, azathioprine, ciclosporin and anti-TNF antibodies (adalimumab, certolizumab pegol, infliximab) on mucosal healing are discussed. Finally, the implications of mucosal healing for subsequent clinical management in patients with IBD are highlighted. PMID:22842618

  6. Aluminum enhances inflammation and decreases mucosal healing in experimental colitis in mice

    PubMed Central

    Pineton de Chambrun, G; Body-Malapel, M; Frey-Wagner, I; Djouina, M; Deknuydt, F; Atrott, K; Esquerre, N; Altare, F; Neut, C; Arrieta, M C; Kanneganti, T-D; Rogler, G; Colombel, J-F; Cortot, A; Desreumaux, P; Vignal, C

    2014-01-01

    The increasing incidence of inflammatory bowel diseases (IBDs) in developing countries has highlighted the critical role of environmental pollutants as causative factors in their pathophysiology. Despite its ubiquity and immune toxicity, the impact of aluminum in the gut is not known. This study aimed to evaluate the effects of environmentally relevant intoxication with aluminum in murine models of colitis and to explore the underlying mechanisms. Oral administration of aluminum worsened intestinal inflammation in mice with 2,4,6-trinitrobenzene sulfonic acid- and dextran sodium sulfate-induced colitis and chronic colitis in interleukin 10-negative (IL10−/−) mice. Aluminum increased the intensity and duration of macroscopic and histologic inflammation, colonic myeloperoxidase activity, inflammatory cytokines expression, and decreased the epithelial cell renewal compared with control animals. Under basal conditions, aluminum impaired intestinal barrier function. In vitro, aluminum induced granuloma formation and synergized with lipopolysaccharide to stimulate inflammatory cytokines expression by epithelial cells. Deleterious effects of aluminum on intestinal inflammation and mucosal repair strongly suggest that aluminum might be an environmental IBD risk factor. PMID:24129165

  7. The Role of E-Cadherin in Maintaining the Barrier Function of Corneal Epithelium after Treatment with Cultured Autologous Oral Mucosa Epithelial Cell Sheet Grafts for Limbal Stem Deficiency

    PubMed Central

    Hoft, Richard H.; Wood, Andrew; Oliva, Joan; Niihara, Hope; Makalinao, Andrew; Thropay, Jacquelyn; Pan, Derek; Tiger, Kumar; Garcia, Julio; Laporte, Amanda; French, Samuel W.; Niihara, Yutaka

    2016-01-01

    The role of E-cadherin in epithelial barrier function of cultured autologous oral mucosa epithelial cell sheet (CAOMECS) grafts was examined. CAOMECS were cultured on a temperature-responsive surface and grafted onto rabbit corneas with Limbal Stem Cell Deficiency (LSCD). E-cadherin levels were significantly higher in CAOMECS compared to normal and LSCD epithelium. Beta-catenin colocalized with E-cadherin in CAOMECS cell membranes while phosphorylated beta-catenin was significantly increased. ZO-1, occludin, and Cnx43 were also strongly expressed in CAOMECS. E-cadherin and beta-catenin localization at the cell membrane was reduced in LSCD corneas, while CAOMECS-grafted corneas showed a restoration of E-cadherin and beta-catenin expression. LSCD corneas did not show continuous staining for ZO-1 or for Cnx43, while CAOMECS-grafted corneas showed a positive expression of ZO-1 and Cnx43. Cascade Blue® hydrazide did not pass through CAOMECS. Because E-cadherin interactions are calcium-dependent, EGTA was used to chelate calcium and disrupt cell adhesion. EGTA-treated CAOMECS completely detached from cell culture surface, and E-cadherin levels were significantly decreased. In conclusion, E cadherin high expression contributed to CAOMECS tight and gap junction protein recruitment at the cell membrane, thus promoting cellular adhesion and a functional barrier to protect the ocular surface. PMID:27777792

  8. Interleukin-18 facilitates neutrophil transmigration via myosin light chain kinase-dependent disruption of occludin, without altering epithelial permeability.

    PubMed

    Lapointe, Tamia K; Buret, Andre G

    2012-02-01

    Compromised epithelial barrier function and tight junction alterations are hallmarks of a number of gastrointestinal disorders, including inflammatory bowel disease (IBD). Increased levels of IL-18 have been observed in mucosal samples from Crohn's disease and ulcerative colitis patients. Remarkably, several reports have demonstrated that immunological or genetic blockage of IL-18 ameliorates the severity of colitis in multiple in vivo models of IBD. Nevertheless, the effects of IL-18 on intestinal epithelial barrier function remain unclear. We hypothesized that IL-18 could disrupt intestinal epithelial barrier structure and function, thus contributing to tissue damage in the context of IBD. The aims of the present study were to determine the effects of IL-18 on epithelial barrier structure and function and to characterize the mechanisms involved in these modulatory properties. Human colonic epithelial Caco-2 monolayers were coincubated with IL-18 for 24 h and processed for immunocytochemistry, immunoblotting, quantitative PCR, and permeability measurements (transepithelial resistance, FITC-dextran fluxes, and bacterial translocation). Our findings indicate that IL-18 selectively disrupts tight junctional occludin, without affecting the distribution pattern of claudin-4, claudin-5, zonula occludens-1, or E-cadherin. This effect coincided with a significant increase in myosin light chain kinase (MLCK) protein levels and activity. Pharmacological inhibition of MLCK and NF-κB prevented IL-18-induced loss of occludin. Although too subtle to alter paracellular permeability, these fine changes correlated with an MLCK-dependent increase in neutrophil transepithelial migration. In conclusion, our data suggest that IL-18 may potentiate inflammation in the context of IBD by facilitating neutrophil transepithelial migration via MLCK-dependent disruption of tight junctional occludin.

  9. Mucosal immunology and probiotics.

    PubMed

    Dongarrà, Maria Luisa; Rizzello, Valeria; Muccio, Letizia; Fries, Walter; Cascio, Antonio; Bonaccorsi, Irene; Ferlazzo, Guido

    2013-02-01

    The cross-talk between the mucosa-associated immune system and microbiota is critical in mucosal tissue homeostasis as well as in protection against infectious and inflammatory diseases occurring at mucosal sites. This recent evidence has paved the way to therapeutic approaches aimed at modulating the mucosa-associated immune system using probiotics. Different strains of probiotics possess the ability to finely regulate dendritic cell (DC) activation, polarizing the subsequent T cell activity toward Th1 (e.g. Lactobacillus (Lb) acidophilus), Th2 (Lb.reuteri and Bifidobacterium bifidum) or, as more recently demonstrated, Th17 responses induced by specific strains such as Lb.rhamnosus GG and Lac23a, the latter isolated in our laboratory. Here, we review some recent advances in our understanding of probiotics effects on mucosal immunology, particularly on cells of the innate immunity such as DCs. We also highlight our own experiences in modulating DC functions by commensal bacteria and discuss the relevance of probiotics administration in the treatment of human immunopathologies.

  10. 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

  11. 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

  12. The development and function of mucosal lymphoid tissues: a balancing act with micro-organisms.

    PubMed

    Randall, T D; Mebius, R E

    2014-05-01

    Mucosal surfaces are constantly exposed to environmental antigens, colonized by commensal organisms and used by pathogens as points of entry. As a result, the immune system has devoted the bulk of its resources to mucosal sites to maintain symbiosis with commensal organisms, prevent pathogen entry, and avoid unnecessary inflammatory responses to innocuous antigens. These functions are facilitated by a variety of mucosal lymphoid organs that develop during embryogenesis in the absence of microbial stimulation as well as ectopic lymphoid tissues that develop in adults following microbial exposure or inflammation. Each of these lymphoid organs samples antigens from different mucosal sites and contributes to immune homeostasis, commensal containment, and immunity to pathogens. Here we discuss the mechanisms, mostly based on mouse studies, that control the development of mucosal lymphoid organs and how the various lymphoid tissues cooperate to maintain the integrity of the mucosal barrier. PMID:24569801

  13. The development and function of mucosal lymphoid tissues: a balancing act with micro-organisms.

    PubMed

    Randall, T D; Mebius, R E

    2014-05-01

    Mucosal surfaces are constantly exposed to environmental antigens, colonized by commensal organisms and used by pathogens as points of entry. As a result, the immune system has devoted the bulk of its resources to mucosal sites to maintain symbiosis with commensal organisms, prevent pathogen entry, and avoid unnecessary inflammatory responses to innocuous antigens. These functions are facilitated by a variety of mucosal lymphoid organs that develop during embryogenesis in the absence of microbial stimulation as well as ectopic lymphoid tissues that develop in adults following microbial exposure or inflammation. Each of these lymphoid organs samples antigens from different mucosal sites and contributes to immune homeostasis, commensal containment, and immunity to pathogens. Here we discuss the mechanisms, mostly based on mouse studies, that control the development of mucosal lymphoid organs and how the various lymphoid tissues cooperate to maintain the integrity of the mucosal barrier.

  14. Rho-A prenylation and signaling link epithelial homeostasis to intestinal inflammation

    PubMed Central

    López-Posadas, Rocío; Becker, Christoph; Günther, Claudia; Tenzer, Stefan; Amann, Kerstin; Billmeier, Ulrike; Atreya, Raja; Fiorino, Gionata; Vetrano, Stefania; Danese, Silvio; Ekici, Arif B.; Wirtz, Stefan; Thonn, Veronika; Watson, Alastair J.M.; Brakebusch, Cord; Bergö, Martin; Neurath, Markus F.; Atreya, Imke

    2016-01-01

    Although defects in intestinal barrier function are a key pathogenic factor in patients with inflammatory bowel diseases (IBDs), the molecular pathways driving disease-specific alterations of intestinal epithelial cells (IECs) are largely unknown. Here, we addressed this issue by characterizing the transcriptome of IECs from IBD patients using a genome-wide approach. We observed disease-specific alterations in IECs with markedly impaired Rho-A signaling in active IBD patients. Localization of epithelial Rho-A was shifted to the cytosol in IBDs, and inflammation was associated with suppressed Rho-A activation due to reduced expression of the Rho-A prenylation enzyme geranylgeranyltransferase-I (GGTase-I). Functionally, we found that mice with conditional loss of Rhoa or the gene encoding GGTase-I, Pggt1b, in IECs exhibit spontaneous chronic intestinal inflammation with accumulation of granulocytes and CD4+ T cells. This phenotype was associated with cytoskeleton rearrangement and aberrant cell shedding, ultimately leading to loss of epithelial integrity and subsequent inflammation. These findings uncover deficient prenylation of Rho-A as a key player in the pathogenesis of IBDs. As therapeutic triggering of Rho-A signaling suppressed intestinal inflammation in mice with GGTase-I–deficient IECs, our findings suggest new avenues for treatment of epithelial injury and mucosal inflammation in IBD patients. PMID:26752649

  15. 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

  16. The uptake of soluble and particulate antigens by epithelial cells in the mouse small intestine.

    PubMed

    Howe, Savannah E; Lickteig, Duane J; Plunkett, Kyle N; Ryerse, Jan S; Konjufca, Vjollca

    2014-01-01

    Intestinal epithelial cells (IECs) overlying the villi play a prominent role in absorption of digested nutrients and establish a barrier that separates the internal milieu from potentially harmful microbial antigens. Several mechanisms by which antigens of dietary and microbial origin enter the body have been identified; however whether IECs play a role in antigen uptake is not known. Using in vivo imaging of the mouse small intestine, we investigated whether epithelial cells (enterocytes) play an active role in the uptake (sampling) of lumen antigens. We found that small molecular weight antigens such as chicken ovalbumin, dextran, and bacterial LPS enter the lamina propria, the loose connective tissue which lies beneath the epithelium via goblet cell associated passageways. However, epithelial cells overlying the villi can internalize particulate antigens such as bacterial cell debris and inert nanoparticles (NPs), which are then found co-localizing with the CD11c+ dendritic cells in the lamina propria. The extent of NP uptake by IECs depends on their size: 20-40 nm NPs are taken up readily, while NPs larger than 100 nm are taken up mainly by the epithelial cells overlying Peyer's patches. Blocking NPs with small proteins or conjugating them with ovalbumin does not inhibit their uptake. However, the uptake of 40 nm NPs can be inhibited when they are administered with an endocytosis inhibitor (chlorpromazine). Delineating the mechanisms of antigen uptake in the gut is essential for understanding how tolerance and immunity to lumen antigens are generated, and for the development of mucosal vaccines and therapies.

  17. Mucosal adjuvants: Opportunities and challenges.

    PubMed

    Zeng, Lingbin

    2016-09-01

    Most pathogens access the body via mucosal surfaces. Mucosal vaccination is a highly effective and recommended method to prevent mucosally transmitted infections. Compared with immunization via intramuscular injection, mucosal immunization offers remarkable advantages, including non-invasiveness, low costs and reduced risk of transmission of blood-borne diseases, which make it more acceptable to human beings, especially to young children. However, only few mucosal vaccines are licensed for human, which is mainly due to the deficiency of safe and effective mucosal adjuvants. Adjuvants, as important components of most vaccines, are essential to enhance immunity and induce immune memory. The development of mucosal adjuvants, unfortunately, has been severely hampered by research strategies based on empiric trials and non-comprehensive methods for safety evaluation. Therefore, changing the research and development strategies of mucosal adjuvant field from empiricism based discovery to rational design based invention is highly demanded. The change of strategies mainly depends upon clarification of mechanism of mucosal adjuvant activity though a combination of life science, information science and materials science. PMID:27159278

  18. Epithelial IL-23R Signaling Licenses Protective IL-22 Responses in Intestinal Inflammation.

    PubMed

    Aden, Konrad; Rehman, Ateequr; Falk-Paulsen, Maren; Secher, Thomas; Kuiper, Jan; Tran, Florian; Pfeuffer, Steffen; Sheibani-Tezerji, Raheleh; Breuer, Alexandra; Luzius, Anne; Jentzsch, Marlene; Häsler, Robert; Billmann-Born, Susanne; Will, Olga; Lipinski, Simone; Bharti, Richa; Adolph, Timon; Iovanna, Juan L; Kempster, Sarah L; Blumberg, Richard S; Schreiber, Stefan; Becher, Burkhard; Chamaillard, Mathias; Kaser, Arthur; Rosenstiel, Philip

    2016-08-23

    A plethora of functional and genetic studies have suggested a key role for the IL-23 pathway in chronic intestinal inflammation. Currently, pathogenic actions of IL-23 have been ascribed to specific effects on immune cells. Herein, we unveil a protective role of IL-23R signaling. Mice deficient in IL-23R expression in intestinal epithelial cells (Il23R(ΔIEC)) have reduced Reg3b expression, show a disturbed colonic microflora with an expansion of flagellated bacteria, and succumb to DSS colitis. Surprisingly, Il23R(ΔIEC) mice show impaired mucosal IL-22 induction in response to IL-23. αThy-1 treatment significantly deteriorates colitis in Il23R(ΔIEC) animals, which can be rescued by IL-22 application. Importantly, exogenous Reg3b administration rescues DSS-treated Il23R(ΔIEC) mice by recruiting neutrophils as IL-22-producing cells, thereby restoring mucosal IL-22 levels. The study identifies a critical barrier-protective immune pathway that originates from, and is orchestrated by, IL-23R signaling in intestinal epithelial cells. PMID:27524624

  19. Mucus barrier-triggered disassembly of siRNA nanocarriers

    NASA Astrophysics Data System (ADS)

    Thomsen, Troels B.; Li, Leon; Howard, Kenneth A.

    2014-10-01

    The mucus overlying mucosal epithelial surfaces presents not only a biological barrier to the penetration of potential pathogens, but also therapeutic modalities including RNAi-based nanocarriers. Movement of nanomedicines across the mucus barriers of the gastrointestinal mucosa is modulated by interactions of the nanomedicine carriers with mucin glycoproteins inside the mucus, potentiated by the large surface area of the nanocarrier. We have developed a fluorescence activation-based reporter system showing that the interaction between polyanionic mucins and the cationic chitosan/small interfering RNA (siRNA) nanocarriers (polyplexes) results in the disassembly and consequent triggered release of fluorescent siRNA. The quantity of release was found to be dependent on the molar ratio between chitosan amino groups and siRNA phosphate groups (NP ratio) of the polyplexes with a maximal estimated 48.6% release of siRNA over 30 min at NP 60. Furthermore, a microfluidic in vitro model of the gastrointestinal mucus barrier was used to visualize the dynamic interaction between chitosan/siRNA nanocarriers and native purified porcine stomach mucins. We observed strong interactions and aggregations at the mucin-liquid interface, followed by an NP ratio dependent release and consequent diffusion of siRNA across the mucin barrier. This work describes a new model of interaction at the nanocarrier-mucin interface and has important implications for the design and development of nucleic acid-based nanocarrier therapeutics for mucosal disease treatments and also provides insights into nanoscale pathogenic processes.The mucus overlying mucosal epithelial surfaces presents not only a biological barrier to the penetration of potential pathogens, but also therapeutic modalities including RNAi-based nanocarriers. Movement of nanomedicines across the mucus barriers of the gastrointestinal mucosa is modulated by interactions of the nanomedicine carriers with mucin glycoproteins inside the

  20. Proteomic and Bioinformatic Profile of Primary Human Oral Epithelial Cells

    PubMed Central

    Ghosh, Santosh K.; Yohannes, Elizabeth; Bebek, Gurkan; Weinberg, Aaron; Jiang, Bin; Willard, Belinda; Chance, Mark R.; Kinter, Michael T.; McCormick, Thomas S.

    2012-01-01

    Wounding of the oral mucosa occurs frequently in a highly septic environment. Remarkably, these wounds heal quickly and the oral cavity, for the most part, remains healthy. Deciphering the normal human oral epithelial cell (NHOEC) proteome is critical for understanding the mechanism(s) of protection elicited when the mucosal barrier is intact, as well as when it is breached. Combining 2D gel electrophoresis with shotgun proteomics resulted in identification of 1662 NHOEC proteins. Proteome annotations were performed based on protein classes, molecular functions, disease association and membership in canonical and metabolic signaling pathways. Comparing the NHOEC proteome with a database of innate immunity-relevant interactions (InnateDB) identified 64 common proteins associated with innate immunity. Comparison with published salivary proteomes revealed that 738/1662 NHOEC proteins were common, suggesting that significant numbers of salivary proteins are of epithelial origin. Gene ontology analysis showed similarities in the distributions of NHOEC and saliva proteomes with regard to biological processes, and molecular functions. We also assessed the inter-individual variability of the NHOEC proteome and observed it to be comparable with other primary cells. The baseline proteome described in this study should serve as a resource for proteome studies of the oral mucosa, especially in relation to disease processes. PMID:23035736

  1. Milk fermentation products of L. helveticus R389 activate calcineurin as a signal to promote gut mucosal immunity

    PubMed Central

    Vinderola, Gabriel; Matar, Chantal; Perdigón, Gabriela

    2007-01-01

    Background Fermented milks containing probiotic bacteria are a way of delivering bioactive constituents to targets in the gastrointestinal tract. We reported previously that the fermentation of milk at constant pH 6 by L. helveticus R389 increased its content of peptide fractions, and the oral administration of the non-bacterial fraction (FMSpH6) to mice increased total secretory IgA in the intestinal lumen and enhanced the number of IgA and various cytokines producing cells as well as the secretion of IL-6 by small intestine epithelial cells. We also demonstrated that this FMSpH6 was effective for the prevention of Salmonella typhimurium infection in mice. In this work, we studied in mice the impact of the oral administration of the supernatant of milk fermented by L. helveticus R389 on the gut physiology by measuring parameters such as calcium channels and E-cadherin expression, the activation of the biological signal calcineurin and mast and goblet cells, as a way to determine some mechanisms involved in the immunomodulating effects of the milk fermentation products, observed in previous studies. We analyzed the impact of the supernatant of milk fermented by L. helveticus R389 at pH6-controlled on the expression of calcineurin and on the reinforcement of the ephitelial barrier, measuring parameters such as calcium channels and E-cadherin expression and in the reinforcement of the non-specific immunity determining mast cells and goblet cells associated to the gut. Results We observed an enhanced expression of TRPV6 channels in the duodenum, indicating an improved capacity for dietary Ca2+ uptake. We demonstrated an enhanced expression of calcineurin in the small intestine, able to upregulate immune parameters such as IL-2 and TNF production, with an increase in the number of these cytokines secreting cells. We determined an increase in the number of mucosal mast cells and goblet cells, which would mean an improved state of mucosal surveillance at sites of

  2. Mice lacking myosin IXb, an inflammatory bowel disease susceptibility gene, have impaired intestinal barrier function and superficial ulceration in the ileum.

    PubMed

    Hegan, Peter S; Chandhoke, Surjit K; Barone, Christina; Egan, Marie; Bähler, Martin; Mooseker, Mark S

    2016-04-01

    Genetic studies have implicated MYO9B, which encodes myosin IXb (Myo9b), a motor protein with a Rho GTPase activating domain (RhoGAP), as a susceptibility gene for inflammatory bowel disease (IBD). Moreover, we have recently shown that knockdown of Myo9b in an intestinal epithelial cell line impairs wound healing and barrier function. Here, we investigated whether mice lacking Myo9b have impaired intestinal barrier function and features of IBD. Myo9b knock out (KO) mice exhibit impaired weight gain and fecal occult blood (indicator of gastrointestinal bleeding), and increased intestinal epithelial cell apoptosis could be detected along the entire intestinal axis. Histologic analysis revealed intestinal mucosal damage, most consistently observed in the ileum, which included superficial ulceration and neutrophil infiltration. Focal lesions contained neutrophils and ultrastructural examination confirmed epithelial discontinuity and the deposition of extracellular matrix. We also observed impaired mucosal barrier function in KO mice. Transepithelial electrical resistance of KO ileum is >3 fold less than WT ileum. The intestinal mucosa is also permeable to high molecular weight dextran, presumably due to the presence of mucosal surface ulcerations. There is loss of tight junction-associated ZO-1, decreased lateral membrane associated E-cadherin, and loss of terminal web associated cytokeratin filaments. Consistent with increased Rho activity in the KO, there is increased subapical expression of activated myosin II (Myo2) based on localization of phosphorylated Myo2 regulatory light chain. Except for a delay in disease onset in the KO, no difference in dextran sulfate sodium-induced colitis and lethality was observed between wild-type and Myo9b KO mice.

  3. Epithelial Cells Augment Barrier Function via Activation of the Toll-Like Receptor 2/Phosphatidylinositol 3-Kinase Pathway upon Recognition of Salmonella enterica Serovar Typhimurium Curli Fibrils in the Gut

    PubMed Central

    Oppong, Gertrude O.; Rapsinski, Glenn J.; Newman, Tiffanny N.; Nishimori, Jessalyn H.; Biesecker, Steven G.

    2013-01-01

    Curli fibrils, the best-characterized functional bacterial amyloids, are an important component of enterobacterial biofilms. We have previously shown that curli fibrils are recognized by the Toll-like receptor 2 (TLR2)/TLR1 heterodimer complex. Utilizing polarized T-84 cells, an intestinal epithelial cell line derived from colon carcinoma grown on semipermeable tissue culture inserts, we determined that infection with a Salmonella enterica serovar Typhimurium csgBA mutant, which does not express curli, resulted in an increase in intestinal barrier permeability and an increase in bacterial translocation compared to infection with curliated wild-type S. Typhimurium. When the TLR2 downstream signaling molecule phosphatidylinositol 3-kinase (PI3K) was blocked using wortmannin or LY294002, the difference in disruption of the intestinal epithelium and bacterial translocation was no longer observed. Additionally, disruption of polarized T-84 cells treated basolaterally with the TLR5 ligand flagellin was prevented when the polarized cells were simultaneously treated with the synthetic TLR2/TLR1 ligand Pam3CSK4 or with purified curli fibrils in the apical compartment. Similar to in vitro observations, C57BL/6 mice infected with the csgBA mutant suffered increased disruption of the intestinal epithelium and therefore greater dissemination of the bacteria to the mesenteric lymph nodes than mice infected with wild-type S. Typhimurium. The differences in disruption of the intestinal epithelium and bacterial dissemination in the mice infected with csgBA mutant or wild-type S. Typhimurium were not apparent in TLR2-deficient mice. Overall, these studies report for the first time that activation of the TLR2/PI3K pathway by microbial amyloids plays a critical role in regulating the intestinal epithelial barrier as well as monitoring bacterial translocation during infection. PMID:23208603

  4. Characterization of the Molecular Interplay between Moraxella catarrhalis and Human Respiratory Tract Epithelial Cells

    PubMed Central

    de Vries, Stefan P. W.; Eleveld, Marc J.; Hermans, Peter W. M.; Bootsma, Hester J.

    2013-01-01

    Moraxella catarrhalis is a mucosal pathogen that causes childhood otitis media and exacerbations of chronic obstructive pulmonary disease in adults. During the course of infection, M. catarrhalis needs to adhere to epithelial cells of different host niches such as the nasopharynx and lungs, and consequently, efficient adhesion to epithelial cells is considered an important virulence trait of M. catarrhalis. By using Tn-seq, a genome-wide negative selection screenings technology, we identified 15 genes potentially required for adherence of M. catarrhalis BBH18 to pharyngeal epithelial Detroit 562 and lung epithelial A549 cells. Validation with directed deletion mutants confirmed the importance of aroA (3-phosphoshikimate 1-carboxyvinyl-transferase), ecnAB (entericidin EcnAB), lgt1 (glucosyltransferase), and MCR_1483 (outer membrane lipoprotein) for cellular adherence, with ΔMCR_1483 being most severely attenuated in adherence to both cell lines. Expression profiling of M. catarrhalis BBH18 during adherence to Detroit 562 cells showed increased expression of 34 genes in cell-attached versus planktonic bacteria, among which ABC transporters for molybdate and sulfate, while reduced expression of 16 genes was observed. Notably, neither the newly identified genes affecting adhesion nor known adhesion genes were differentially expressed during adhesion, but appeared to be constitutively expressed at a high level. Profiling of the transcriptional response of Detroit 562 cells upon adherence of M. catarrhalis BBH18 showed induction of a panel of pro-inflammatory genes as well as genes involved in the prevention of damage of the epithelial barrier. In conclusion, this study provides new insight into the molecular interplay between M. catarrhalis and host epithelial cells during the process of adherence. PMID:23936538

  5. MicroRNAs and Epithelial Immunity

    PubMed Central

    Liu, Jun; Drescher, Kristen M.; Chen, Xian-Ming

    2009-01-01

    MicroRNAs are required for development and maintenance of the epithelial barrier. It is hypothesized that microRNAs are involved in regulating epithelial anti-microbial defenses by targeting key epithelial effector molecules and/or influencing intracellular signaling pathways. Additionally, aberrant microRNA expression has been implicated in the pathogenesis of various diseases at the skin and mucosa. Increased understanding of the role of microRNAs in epithelial immunoregulation and identification of microRNAs of pathogenetic significance will enhance our understanding of epithelial immunobiology and immunopathology. PMID:19811319

  6. Alteration in Intrapulmonary Pharmacokinetics of Aerosolized Model Compounds Due to Disruption of the Alveolar Epithelial Barriers Following Bleomycin-Induced Pulmonary Fibrosis in Rats.

    PubMed

    Togami, Kohei; Chono, Sumio; Tada, Hitoshi

    2016-03-01

    Idiopathic pulmonary fibrosis is a lethal lung disease that is characterized by the accumulation of extracellular matrix and a change in lung structure. In this study, intrapulmonary pharmacokinetics of aerosolized model compounds were evaluated using rats with bleomycin-induced pulmonary fibrosis. Aerosol formulations of indocyanine green, 6-carboxyfluorescein (6-CF), and fluorescein isothiocyanate dextrans (FD; 4.4, 10, 70, and 250 kDa) were administered to rat lungs using a MicroSprayer. Indocyanine green fluorescence signals were significantly weaker in fibrotic lungs than in control lungs and 6-CF and FD concentrations in the plasma of pulmonary fibrotic animals were markedly higher than in the plasma of control animals. Moreover, disrupted epithelial tight junctions, including claudins-1, -3, and -5, were observed in pulmonary fibrotic lesions using immunofluorescence microscopy. In addition, destruction of tight junctions on model alveolar epithelial cells (NCI-H441) by transforming growth factor-β1 treatment enhanced the permeability of 6-CF and FDs through NCI-H441 cell monolayers. These results indicate that aerosolized drugs are easily distributed into the plasma after leakage through damaged tight junctions of alveolar epithelium. Therefore, the development of delivery systems for anti-fibrotic agents to improve intrapulmonary pharmacokinetics may be necessary for effective idiopathic pulmonary fibrosis therapy.

  7. 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

  8. Mucosal effects of tenofovir 1% gel

    PubMed Central

    Hladik, Florian; Burgener, Adam; Ballweber, Lamar; Gottardo, Raphael; Vojtech, Lucia; Fourati, Slim; Dai, James Y; Cameron, Mark J; Strobl, Johanna; Hughes, Sean M; Hoesley, Craig; Andrew, Philip; Johnson, Sherri; Piper, Jeanna; Friend, David R; Ball, T Blake; Cranston, Ross D; Mayer, Kenneth H; McElrath, M Juliana; McGowan, Ian

    2015-01-01

    Tenofovir gel is being evaluated for vaginal and rectal pre-exposure prophylaxis against HIV transmission. Because this is a new prevention strategy, we broadly assessed its effects on the mucosa. In MTN-007, a phase-1, randomized, double-blinded rectal microbicide trial, we used systems genomics/proteomics to determine the effect of tenofovir 1% gel, nonoxynol-9 2% gel, placebo gel or no treatment on rectal biopsies (15 subjects/arm). We also treated primary vaginal epithelial cells from four healthy women with tenofovir in vitro. After seven days of administration, tenofovir 1% gel had broad-ranging effects on the rectal mucosa, which were more pronounced than, but different from, those of the detergent nonoxynol-9. Tenofovir suppressed anti-inflammatory mediators, increased T cell densities, caused mitochondrial dysfunction, altered regulatory pathways of cell differentiation and survival, and stimulated epithelial cell proliferation. The breadth of mucosal changes induced by tenofovir indicates that its safety over longer-term topical use should be carefully monitored. Clinical trial registration: NCT01232803. DOI: http://dx.doi.org/10.7554/eLife.04525.001 PMID:25647729

  9. Mucosal effects of tenofovir 1% gel.

    PubMed

    Hladik, Florian; Burgener, Adam; Ballweber, Lamar; Gottardo, Raphael; Vojtech, Lucia; Fourati, Slim; Dai, James Y; Cameron, Mark J; Strobl, Johanna; Hughes, Sean M; Hoesley, Craig; Andrew, Philip; Johnson, Sherri; Piper, Jeanna; Friend, David R; Ball, T Blake; Cranston, Ross D; Mayer, Kenneth H; McElrath, M Juliana; McGowan, Ian

    2015-01-01

    Tenofovir gel is being evaluated for vaginal and rectal pre-exposure prophylaxis against HIV transmission. Because this is a new prevention strategy, we broadly assessed its effects on the mucosa. In MTN-007, a phase-1, randomized, double-blinded rectal microbicide trial, we used systems genomics/proteomics to determine the effect of tenofovir 1% gel, nonoxynol-9 2% gel, placebo gel or no treatment on rectal biopsies (15 subjects/arm). We also treated primary vaginal epithelial cells from four healthy women with tenofovir in vitro. After seven days of administration, tenofovir 1% gel had broad-ranging effects on the rectal mucosa, which were more pronounced than, but different from, those of the detergent nonoxynol-9. Tenofovir suppressed anti-inflammatory mediators, increased T cell densities, caused mitochondrial dysfunction, altered regulatory pathways of cell differentiation and survival, and stimulated epithelial cell proliferation. The breadth of mucosal changes induced by tenofovir indicates that its safety over longer-term topical use should be carefully monitored. PMID:25647729

  10. Pharmacological Protection From Radiation {+-} Cisplatin-Induced Oral Mucositis

    SciTech Connect

    Cotrim, Ana P.; Yoshikawa, Masanobu; Sunshine, Abraham N.; Zheng Changyu; Sowers, Anastasia L.; Thetford, Angela D.; Cook, John A.; Mitchell, James B.; Baum, Bruce J.

    2012-07-15

    Purpose: To evaluate if two pharmacological agents, Tempol and D-methionine (D-met), are able to prevent oral mucositis in mice after exposure to ionizing radiation {+-} cisplatin. Methods and Materials: Female C3H mice, {approx}8 weeks old, were irradiated with five fractionated doses {+-} cisplatin to induce oral mucositis (lingual ulcers). Just before irradiation and chemotherapy, mice were treated, either alone or in combination, with different doses of Tempol (by intraperitoneal [ip] injection or topically, as an oral gel) and D-met (by gavage). Thereafter, mice were sacrificed and tongues were harvested and stained with a solution of Toluidine Blue. Ulcer size and tongue epithelial thickness were measured. Results: Significant lingual ulcers resulted from 5 Multiplication-Sign 8 Gy radiation fractions, which were enhanced with cisplatin treatment. D-met provided stereospecific partial protection from lingual ulceration after radiation. Tempol, via both routes of administration, provided nearly complete protection from lingual ulceration. D-met plus a suboptimal ip dose of Tempol also provided complete protection. Conclusions: Two fairly simple pharmacological treatments were able to markedly reduce chemoradiation-induced oral mucositis in mice. This proof of concept study suggests that Tempol, alone or in combination with D-met, may be a useful and convenient way to prevent the severe oral mucositis that results from head-and-neck cancer therapy.

  11. Growth inhibition of Candida by human oral epithelial cells.

    PubMed

    Steele, C; Leigh, J; Swoboda, R; Fidel, P L

    2000-11-01

    Oropharyngeal candidiasis (OPC) caused by Candida albicans is a significant problem in human immunodeficiency virus (HIV)-infected persons. Recognizing the paucity of information on innate and/or adaptive mucosal host defenses against C. albicans, we recently reported that human and nonhuman primate and mouse vaginal epithelial cells inhibit the growth of C. albicans in vitro. In the present study, oral epithelial cells collected from saliva of healthy volunteers and a purified oral epithelial cell line were found to inhibit blastoconidia and/or hyphal growth of several Candida species. Cell contact was a strict requirement for the epithelial cell anti-Candida activity; neither saliva nor culture supernatants alone inhibited Candida growth, and addition of saliva to the coculture did not modulate the epithelial cell activity. Finally, epithelial cell anti-Candida activity was significantly lower in HIV-infected persons with OPC. Together, these results suggest that oral epithelial cells may play a role in innate resistance against OPC.

  12. Increased iNOS activity is essential for intestinal epithelial tight junction dysfunction in endotoxemic mice.

    PubMed

    Han, Xiaonan; Fink, Mitchell P; Yang, Runkuan; Delude, Russell L

    2004-03-01

    We tested the hypothesis that increased production of nitric oxide (NO.) associated with lipopolysaccharide (LPS)-induced systemic inflammation leads to functionally significant alterations in the expression and/or targeting of key tight junction (TJ) proteins in ileal and colonic epithelium. Wild-type or inducible NO. synthase (iNOS) knockout male C57B1/6J mice were injected intraperitoneally with 2 mg/kg Escherichia coli O111:B4 LPS. iNOS was inhibited using intraperitoneal L-N(6)-(1-iminoethyl)lysine (L-NIL; 5 mg/kg). Immunoblotting of total protein and NP-40 insoluble proteins revealed decreased expression and decreased TJ localization, respectively, of the TJ proteins, zonula occludens (ZO)-1, ZO-2, ZO-3, and/or occludin in ileal mucosa and colonic mucosa (total protein only) after injection of C57B1/6J mice with LPS. Immunohistochemistry showed deranged distribution of ZO-1 and occludin in both tissues from endotoxemic mice. Endotoxemia was associated with evidence of gut epithelial barrier dysfunction evidenced by increased ileal mucosal permeability to fluorescein isothiocyanate-dextran (Mr=4 kDa) and increased bacterial translocation to mesenteric lymph nodes. Pharmacologic inhibition of iNOS activity using L-NIL or genetic ablation of the iNOS gene ameliorated LPS-induced changes in TJ protein expression and gut mucosal barrier function. These results support the view that at least one mechanism contributing to the pathogenesis of gastrointestinal epithelial dysfunction secondary to systemic inflammation is increased iNOS-dependent NO. production leading to altered expression and localization of key TJ proteins.

  13. 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

  14. 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

  15. 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.

  16. Langerhans cells and their role in oral mucosal diseases.

    PubMed

    Upadhyay, Juhi; Upadhyay, Ram B; Agrawal, Pankaj; Jaitley, Shweta; Shekhar, Rhitu

    2013-09-01

    Dendritic cells are arguably the most potent antigen-presenting cells and may be the only cells capable of initiating the adaptive immune response. The epithelial residents of dendritic cells are Langerhans cells, which serve as the "sentinels" of the mucosa, altering the immune system not only to pathogen entry but also of tolerance to self antigen and commensal microbes. Oral mucosal Langerhans cells are capable of engaging and internalizing a wide variety of pathogens and have been found responsive to nickel in patients with nickel allergies, oral Candida species, oral lichen planus, lichenoid drug eruptions, graft versus host diseases, periodontal diseases median rhomboid glossitis, human immunodeficiency virus infection, hairy leukoplakia of the tongue, and oral squamous cell carcinoma. Review focuses on the role of antigen-presenting cells in particular Langerhans cells to better understand the mechanisms underlying immune responses. In this review, comprehensive detail about mucosal diseases has been compiled using the PubMed database and through textbooks. PMID:24251267

  17. A novel method for imaging sites of paracellular passage of macromolecules in epithelial sheets.

    PubMed

    Richter, Jan F; Schmauder, Ralf; Krug, Susanne M; Gebert, Andreas; Schumann, Michael

    2016-05-10

    Understanding the dynamics of intestinal barrier function is key to elucidating oral delivery routes of therapeutics as well as to understanding various diseases that involve the mucosal immune system. Passage of macromolecules across barrier-forming epithelia is classically analyzed by means of various tracer flux measurements. This approach averages over contributions from many cells and lacks labeling of passage-sites. Thus, abundance and nature of involved cells have remained unidentified. We present a novel method that allowed for optical analysis of passage of various macromolecules on large-scale and single-cell level. To achieve tracking of passage loci in epithelia at submicrometer resolution we used biotinylated and fluorescent macromolecules that bind to basolateral membranes pre-labeled with cell-adherent avidin. We applied this method to epithelial cell lines and isolated mucosae in order to 3-dimensionally determine barrier leak properties over time. Tracer passage was found in all epithelia examined. However, it was infrequent, strikingly inhomogeneous, depended on culture duration and tightness of the monolayer. Stimulating passage with barrier-perturbing agents increased the number of leaks exposition time-dependently in cell lines and explanted mucosae. After stepwise opening of the paracellular passage pathway, integrated tracer-signal measured by our assay strictly correlated to simultaneously performed standard fluxes. Thus, our assay allows for the study of transepithelial macromolecule passage in various physiological and pathological conditions. PMID:26995760

  18. A novel method for imaging sites of paracellular passage of macromolecules in epithelial sheets.

    PubMed

    Richter, Jan F; Schmauder, Ralf; Krug, Susanne M; Gebert, Andreas; Schumann, Michael

    2016-05-10

    Understanding the dynamics of intestinal barrier function is key to elucidating oral delivery routes of therapeutics as well as to understanding various diseases that involve the mucosal immune system. Passage of macromolecules across barrier-forming epithelia is classically analyzed by means of various tracer flux measurements. This approach averages over contributions from many cells and lacks labeling of passage-sites. Thus, abundance and nature of involved cells have remained unidentified. We present a novel method that allowed for optical analysis of passage of various macromolecules on large-scale and single-cell level. To achieve tracking of passage loci in epithelia at submicrometer resolution we used biotinylated and fluorescent macromolecules that bind to basolateral membranes pre-labeled with cell-adherent avidin. We applied this method to epithelial cell lines and isolated mucosae in order to 3-dimensionally determine barrier leak properties over time. Tracer passage was found in all epithelia examined. However, it was infrequent, strikingly inhomogeneous, depended on culture duration and tightness of the monolayer. Stimulating passage with barrier-perturbing agents increased the number of leaks exposition time-dependently in cell lines and explanted mucosae. After stepwise opening of the paracellular passage pathway, integrated tracer-signal measured by our assay strictly correlated to simultaneously performed standard fluxes. Thus, our assay allows for the study of transepithelial macromolecule passage in various physiological and pathological conditions.

  19. Regulation of the gut microbiota by the mucosal immune system in mice

    PubMed Central

    Hasegawa, Mizuho

    2014-01-01

    The benefits of commensal bacteria to the health of the host have been well documented, such as providing stimulation to potentiate host immune responses, generation of useful metabolites, and direct competition with pathogens. However, the ability of the host immune system to control the microbiota remains less well understood. Recent microbiota analyses in mouse models have revealed detailed structures and diversities of microbiota at different sites of the digestive tract in mouse populations. The contradictory findings of previous studies on the role of host immune responses in overall microbiota composition are likely attributable to the high β-diversity in mouse populations as well as technical limitations of the methods to analyze microbiota. The host employs multiple systems to strictly regulate their interactions with the microbiota. A spatial segregation between the host and microbiota is achieved with the mucosal epithelium, which is further fortified with a mucus layer on the luminal side and Paneth cells that produce antimicrobial peptides. When commensal bacteria or pathogens breach the epithelial barrier and translocate to peripheral tissues, the host immune system is activated to eliminate them. Defective segregation and tissue elimination of commensals result in exaggerated inflammatory responses and possibly death of the host. In this review, we discuss the current understanding of mouse microbiota, its common features with human microbiota, the technologies utilized to analyze microbiota, and finally the challenges faced to delineate the role of host immune responses in the composition of the luminal microbiota. PMID:24792038

  20. Esophageal mucosal protection--why do we need a special approach?

    PubMed

    Simon, L

    1992-01-01

    The epidemiology and natural history of reflux induced peptic esophageal diseases remain incompletely understood. That is why it is easy to explain that the traditional therapeutic efforts were mostly restricted to the use of acid-reducing or neutralizing drogs. The author tries to survey--mainly on theoretical bases--a new approach of the maintenance treatment of peptic esophagitis and consequential columnar metaplasia. The mechanism of the esophageal antireflux barrier is composed by the (a) lower esophageal sphincter tone, (b) upper esophageal sphincter tone, (c) esophageal acid clearance and (d) esophageal epithelial resistance. The data of a 100-patient-group of gastroesophageal reflux disease cases were retrospectively evaluated principally considering the efficacy of antisecretory treatment relating to the accompanying diseases, recurrence of symptoms and prevention the development of Barrett's columnar lined esophagus and Barrett's ulceration. The decrease of exposure by damaging factors is an essential criterion of antisecretory therapy, having several disadvantages. Based only to logically well established arguments the author believes that gastroesophageal reflux disease and consecutive conditions might be an ideal model for studying and introducing esophageal cyto (-mucosal, -tissue) protection, considering that in the esophagus--in contradiction to the stomach--the cell and tissue injury, induced by several pathogenic agents, does not develop rapidly, and when the organ damage develops gradually, interventions may be possible to protect esophageal cell and the mucosa directly.

  1. Systemic Staphylococcus aureus infection mediated by Candida albicans hyphal invasion of mucosal tissue

    PubMed Central

    Schlecht, Lisa Marie; Peters, Brian M.; Krom, Bastiaan P.; Freiberg, Jeffrey A.; Hänsch, Gertrud M.; Filler, Scott G.

    2015-01-01

    Candida albicans and Staphylococcus aureus are often co-isolated in cases of biofilm-associated infections. C. albicans can cause systemic disease through morphological switch from the rounded yeast to the invasive hyphal form. Alternatively, systemic S. aureus infections arise from seeding through breaks in host epithelial layers although many patients have no documented portal of entry. We describe a novel strategy by which S. aureus is able to invade host tissue and disseminate via adherence to the invasive hyphal elements of Candida albicans. In vitro and ex vivo findings demonstrate a specific binding of the staphylococci to the candida hyphal elements. The C. albicans cell wall adhesin Als3p binds to multiple staphylococcal adhesins. Furthermore, Als3p is required for C. albicans to transport S. aureus into the tissue and cause a disseminated infection in an oral co-colonization model. These findings suggest that C. albicans can facilitate the invasion of S. aureus across mucosal barriers, leading to systemic infection in co-colonized patients. PMID:25332378

  2. What interactions drive the salivary mucosal pellicle formation?

    PubMed Central

    Gibbins, Hannah L.; Yakubov, Gleb E.; Proctor, Gordon B.; Wilson, Stephen; Carpenter, Guy H.

    2014-01-01

    The bound salivary pellicle is essential for protection of both the enamel and mucosa in the oral cavity. The enamel pellicle formation is well characterised, however the mucosal pellicle proteins have only recently been clarified and what drives their formation is still unclear. The aim of this study was to examine the salivary pellicle on particles with different surface properties (hydrophobic or hydrophilic with a positive or negative charge), to determine a suitable model to mimic the mucosal pellicle. A secondary aim was to use the model to test how transglutaminase may alter pellicle formation. Particles were incubated with resting whole mouth saliva, parotid saliva and submandibular/sublingual saliva. Following incubation and two PBS and water washes bound salivary proteins were eluted with two concentrations of SDS, which were later analysed using SDS-PAGE and Western blotting. Experiments were repeated with purified transglutaminase to determine how this epithelial-derived enzyme may alter the bound pellicle. Protein pellicles varied according to the starting salivary composition and the particle chemistry. Amylase, the single most abundant protein in saliva, did not bind to any particle indicating specific protein binding. Most proteins bound through hydrophobic interactions and a few according to their charges. The hydrophobic surface most closely matched the known salivary mucosal pellicle by containing mucins, cystatin and statherin but an absence of amylase and proline-rich proteins. This surface was further used to examine the effect of added transglutaminase. At the concentrations used only statherin showed any evidence of crosslinking with itself or another saliva protein. In conclusion, the formation of the salivary mucosal pellicle is probably mediated, at least in part, by hydrophobic interactions to the epithelial cell surface. PMID:24921197

  3. Molecular Signatures of Immune Activation and Epithelial Barrier Remodeling Are Enhanced during the Luteal Phase of the Menstrual Cycle: Implications for HIV Susceptibility

    PubMed Central

    Arnold, Kelly B.; Novak, Richard M.; McCorrister, Stuart; Shaw, Souradet; Westmacott, Garrett R.; Ball, Terry B.; Lauffenburger, Douglas A.; Burgener, Adam

    2015-01-01

    ABSTRACT The variable infectivity and transmissibility of HIV/SHIV has been recently associated with the menstrual cycle, with particular susceptibility observed during the luteal phase in nonhuman primate models and ex vivo human explant cultures, but the mechanism is poorly understood. Here, we performed an unbiased, mass spectrometry-based proteomic analysis to better understand the mucosal immunological processes underpinning this observed susceptibility to HIV infection. Cervicovaginal lavage samples (n = 19) were collected, characterized as follicular or luteal phase using days since last menstrual period, and analyzed by tandem mass spectrometry. Biological insights from these data were gained using a spectrum of computational methods, including hierarchical clustering, pathway analysis, gene set enrichment analysis, and partial least-squares discriminant analysis with LASSO feature selection. Of the 384 proteins identified, 43 were differentially abundant between phases (P < 0.05, ≥2-fold change). Cell-cell adhesion proteins and antiproteases were reduced, and leukocyte recruitment (interleukin-8 pathway, P = 1.41E–5) and extravasation proteins (P = 5.62E–4) were elevated during the luteal phase. LASSO/PLSDA identified a minimal profile of 18 proteins that best distinguished the luteal phase. This profile included cytoskeletal elements and proteases known to be involved in cellular movement. Gene set enrichment analysis associated CD4+ T cell and neutrophil gene set signatures with the luteal phase (P < 0.05). Taken together, our findings indicate a strong association between proteins involved in tissue remodeling and leukocyte infiltration with the luteal phase, which may represent potential hormone-associated mechanisms of increased susceptibility to HIV. IMPORTANCE Recent studies have discovered an enhanced susceptibility to HIV infection during the progesterone-dominant luteal phase of the menstrual cycle. However, the mechanism responsible for

  4. Effect of Eriobotrya japonica seed extract on 5-fluorouracil-induced mucositis in hamsters.

    PubMed

    Takuma, Daisuke; Guangchen, Sun; Yokota, Junko; Hamada, Atsuhide; Onogawa, Masahide; Yoshioka, Saburo; Kusunose, Masahiko; Miyamura, Mitsuhiko; Kyotani, Syojiro; Nishioka, Yutaka

    2008-02-01

    In this study, we investigated the effects of an Eriobotrya japonica seed extract (ESE) on mucositis using a 5-fluorouracil (5-FU)-induced mucositis hamster model. This model was prepared by intraperitoneally administering 90 mg/kg of 5-FU to hamsters on Day 1, scratching 1 cm(2) of the left cheek pouch of hamsters with a wire brush on Days 2, 3, and 4, and intraperitoneally administering 60 mg/kg of 5-FU on Day 5. Mucositis was evaluated based on the mucositis score at the mucositis site, left cheek pouch thickness, histological findings on HE staining, and plasma lipid peroxide levels. On Day 10, the mucositis score and left cheek pouch thickness in the ESE group were significantly lower than those in the tap water group. Histologically, the two groups showed a defect of the cheek pouch epithelium on Day 6. On Day 10, epithelial injury and bacterial infection were noted in the tap water group. However, in the ESE group, similar findings were not observed. On Day 6, the plasma lipid peroxide level in the tap water group was significantly higher than that in the normal group. In the ESE group, the plasma lipid peroxide level was significantly lower than that in the tap water group. These results suggest that ESE is useful for treating chemotherapy-induced mucositis.

  5. Mucosal B cells: phenotypic characteristics, transcriptional regulation, and homing properties.

    PubMed

    Brandtzaeg, Per; Johansen, Finn-Eirik

    2005-08-01

    Mucosal antibody defense depends on a complex cooperation between local B cells and secretory epithelia. Mucosa-associated lymphoid tissue gives rise to B cells with striking J-chain expression that are seeded to secretory effector sites. Such preferential homing constitutes the biological basis for local production of polymeric immunoglobulin A (pIgA) and pentameric IgM with high affinity to the epithelial pIg receptor that readily can export these antibodies to the mucosal surface. This ultimate functional goal of mucosal B-cell differentiation appears to explain why the J chain is also expressed by IgG- and IgD-producing plasma cells (PCs) occurring at secretory tissue sites; these immunocytes may be considered as 'spin-offs' from early effector clones that through class switch are on their way to pIgA production. Abundant evidence supports the notion that intestinal PCs are largely derived from B cells initially activated in gut-associated lymphoid tissue (GALT). Nevertheless, insufficient knowledge exists concerning the relative importance of M cells, major histocompatibility complex class II-expressing epithelial cells, and professional antigen-presenting cells for the uptake, processing, and presentation of luminal antigens in GALT to accomplish the extensive and sustained priming and expansion of mucosal B cells. Likewise, it is unclear how the germinal center reaction in GALT so strikingly can promote class switch to IgA and expression of J chain. Although B-cell migration from GALT to the intestinal lamina propria is guided by rather well-defined adhesion molecules and chemokines/chemokine receptors, the cues directing preferential homing to different segments of the gut require better definition. This is even more so for the molecules involved in homing of mucosal B cells to secretory effector sites beyond the gut, and in this respect, the role of Waldever's ring (including the palatine tonsils and adenoids) as a regional inductive tissue needs further

  6. LRRC31 is induced by IL-13 and regulates kallikrein expression and barrier function in the esophageal epithelium

    PubMed Central

    D’Mello, RJ; Caldwell, JM; Azouz, NP; Wen, T; Sherrill, JD; Hogan, SP; Rothenberg, ME

    2015-01-01

    Eosinophilic esophagitis (EoE) is an allergic inflammatory disease of the esophagus featuring increased esophageal interleukin 13 (IL-13) levels and impaired barrier function. Herein, we investigated leucine-rich repeat–containing protein 31 (LRRC31) in human EoE esophageal tissue and IL-13–treated esophageal epithelial cells. LRRC31 had basal mRNA expression in colonic and airway mucosal epithelium. Esophageal LRRC31 mRNA and protein increased in active EoE and strongly correlated with esophageal eosinophilia and IL13 and CCL26 mRNA expression. IL-13 treatment increased LRRC31 mRNA and protein in air-liquid interface–differentiated esophageal epithelial cells (EPC2s). At baseline, differentiated LRRC31-overexpressing EPC2s had increased barrier function (1.9-fold increase in transepithelial electrical resistance [P < 0.05] and 2.8-fold decrease in paracellular flux [P < 0.05]). RNA sequencing analysis of differentiated LRRC31-overexpressing EPC2s identified 38 dysregulated genes (P < 0.05), including 5 kallikrein (KLK) serine proteases. Notably, differentiated LRRC31-overexpressing EPC2s had decreased KLK expression and activity, whereas IL-13–treated, differentiated LRRC31 gene-silenced EPC2s had increased KLK expression and suprabasal epithelial detachment. We identified similarly dysregulated KLK expression in the esophagus of patients with active EoE and in IL-13–treated esophageal epithelial cells. We propose that LRRC31 is induced by IL-13 and modulates epithelial barrier function, potentially through KLK regulation. PMID:26462420

  7. Denatured globular protein and bile salt-coated nanoparticles for poorly water-soluble drugs: Penetration across the intestinal epithelial barrier into the circulation system and enhanced oral bioavailability.

    PubMed

    He, Wei; Yang, Ke; Fan, Lifang; Lv, Yaqi; Jin, Zhu; Zhu, Shumin; Qin, Chao; Wang, Yiao; Yin, Lifang

    2015-11-10

    Oral drug delivery is the most preferred route for patients; however, the low solubility of drugs and the resultant poor absorption compromise the benefits of oral administration. On the other hand, for years, the overwhelmingly accepted mechanism for enhanced oral absorption using lipid nanocarriers was based on the process of lipid digestion and drug solubilization in the small intestine. Few reports indicated that other bypass pathways are involved in drug absorption in the gastrointestinal tract (GIT) for oral delivery of nanocarriers. Herein, we report a new nanoemulsion system with a denatured globular protein with a diameter of 30 nm, soybean protein isolates (SPI), and bile salt as emulsifiers, aiming to enhance the absorption of insoluble drugs and explore other pathways for absorption. A BCS class II drug, fenofibrate (FB), was used as the model drug. The SPI and bile salt-coated Ns with a diameter of approximately 150 nm were prepared via a high-pressure homogenizing procedure. Interestingly, the present Ns could be converted to solid dosage form using fluid-bed coating technology, maintaining a nanoscale size. Most importantly, in a model of in situ rat intestinal perfusion, Ns could penetrate across the intestinal epithelial barrier into the systemic circulation and then obtain biodistribution into other tissues. In addition, Ns significantly improved FB oral absorption, exhibited as a greater than 2- and 2.5-fold increase in Cmax and AUC0-t, respectively, compared to the suspension formulation. Overall, the present Ns are promising nanocarriers for the oral delivery of insoluble drugs, and the penetration of intact Ns across the GIT barrier into systemic circulation may be a new strategy for improved drug absorption with the use of nanocarriers.

  8. Ontogeny of Intestinal Epithelial Innate Immune Responses

    PubMed Central

    Hornef, Mathias W.; Fulde, Marcus

    2014-01-01

    Emerging evidence indicates that processes during postnatal development might significantly influence the establishment of mucosal host-microbial homeostasis. Developmental and adaptive immunological processes but also environmental and microbial exposure early after birth might thus affect disease susceptibility and health during adult life. The present review aims at summarizing the current understanding of the intestinal epithelial innate immune system and its developmental and adaptive changes after birth. PMID:25346729

  9. Interleukin-10 Enhances the Intestinal Epithelial Barrier in the Presence of Corticosteroids through p38 MAPK Activity in Caco-2 Monolayers: A Possible Mechanism for Steroid Responsiveness in Ulcerative Colitis

    PubMed Central

    Lorén, Violeta; Cabré, Eduard; Ojanguren, Isabel; Domènech, Eugeni; Pedrosa, Elisabet; García-Jaraquemada, Arce; Mañosa, Miriam; Manyé, Josep

    2015-01-01

    Glucocorticosteroids are the first line therapy for moderate-severe flare-ups of ulcerative colitis. Despite that, up to 60% of patients do not respond adequately to steroid treatment. Previously, we reported that low IL-10 mRNA levels in intestine are associated with a poor response to glucocorticoids in active Crohn’s disease. Here, we test whether IL-10 can favour the response to glucocorticoids by improving the TNFα-induced intestinal barrier damage (assessed by transepithelial electrical resistance) in Caco-2 monolayers, and their possible implications on glucocorticoid responsiveness in active ulcerative colitis. We show that the association of IL-10 and glucocorticoids improves the integrity of TNFα-treated Caco-2 cells and that p38 MAPK plays a key role. In vitro, IL-10 facilitates the nuclear translocation of p38 MAPK-phosphorylated thereby modulating glucocorticoids-receptor-α, IL-10-receptor-α and desmoglein-2 expression. In glucocorticoids-refractory patients, p38 MAPK phosphorylation and membrane desmoglein-2 expression are reduced in colonic epithelial cells. These results suggest that p38 MAPK-mediated synergism between IL-10 and glucocorticoids improves desmosome straightness contributing to the recovery of intestinal epithelium and reducing luminal antigens contact with lamina propria in ulcerative colitis. This study highlights the link between the intestinal epithelium in glucocorticoids-response in ulcerative colitis. PMID:26090671

  10. Claudin-4 Overexpression in Epithelial Ovarian Cancer Is Associated with Hypomethylation and Is a Potential Target for Modulation of Tight Junction Barrier Function Using a C-Terminal Fragment of Clostridium perfringens Enterotoxin1

    PubMed Central

    Litkouhi, Babak; Kwong, Joseph; Lo, Chun-Min; Smedley, James G; McClane, Bruce A; Aponte, Margarita; Gao, Zhijian; Sarno, Jennifer L; Hinners, Jennifer; Welch, William R; Berkowitz, Ross S; Mok, Samuel C; Garner, Elizabeth I O

    2007-01-01

    Background Claudin-4, a tight junction (TJ) protein and receptor for the C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE), is overexpressed in epithelial ovarian cancer (EOC). Previous research suggests DNA methylation is a mechanism for claudin-4 overexpression in cancer and that C-CPE acts as an absorption-enhancing agent in claudin-4-expressing cells. We sought to correlate claudin-4 overexpression in EOC with clinical outcomes and TJ barrier function, investigate DNA methylation as a mechanism for overexpression, and evaluate the effect of C-CPE on the TJ. Methods Claudin-4 expression in EOC was quantified and correlated with clinical outcomes. Claudin-4 methylation status was determined, and claudin-4-negative cell lines were treated with a demethylating agent. Electric cell-substrate impedance sensing was used to calculate junctional (paracellular) resistance (Rb) in EOC cells after claudin-4 silencing and after C-CPE treatment. Results Claudin-4 overexpression in EOC does not correlate with survival or other clinical endpoints and is associated with hypomethylation. Claudin-4 overexpression correlates with Rb and C-CPE treatment of EOC cells significantly decreased Rb in a dose- and claudin-4-dependent noncytotoxic manner. Conclusions C-CPE treatment of EOC cells leads to altered TJ function. Further research is needed to determine the potential clinical applications of C-CPE in EOC drug delivery strategies. PMID:17460774

  11. The Anti-Inflammatory Effect of Spray-Dried Plasma Is Mediated by a Reduction in Mucosal Lymphocyte Activation and Infiltration in a Mouse Model of Intestinal Inflammation

    PubMed Central

    Pérez-Bosque, Anna; Miró, Lluïsa; Amat, Concepció; Polo, Javier; Moretó, Miquel

    2016-01-01

    Spray-dried preparations from porcine and bovine plasma can alleviate mucosal inflammation in experimental models and improve symptoms in patients with enteropathy. In rodents, dietary supplementation with porcine spray-dried plasma (SDP) attenuates intestinal inflammation and improves the epithelial barrier function during intestinal inflammation induced by Staphylococcus aureus enterotoxin B (SEB). The aim of this study was to discern the molecular mechanisms involved in the anti-inflammatory effects of SDP. Male C57BL/6 mice were fed with 8% SDP or control diet (based on milk proteins) for two weeks, from weaning until day 33. On day 32, the mice were given a SEB dose (i.p., 25 µg/mouse) or vehicle. SEB administration increased cell recruitment to mesenteric lymph nodes and the percentage of activated Th lymphocytes and SDP prevented these effects). SDP supplementation increased the expression of interleukin 10 (IL-10) or transforming growth factor- β (TGF-β) compared to the SEB group. The SEB challenge increased six-fold the expression of mucosal addressin cell adhesion molecule 1 (MAdCAM-1) and intercellular adhesion molecule 1 (ICAM-1); and these effects were attenuated by SDP supplementation. SEB also augmented NF-κB phosphorylation, an effect that was prevented by dietary SDP. Our results indicate that the anti-inflammatory effects of SDP involve the regulation of transcription factors and adhesion molecules that reduce intestinal cell infiltration and the degree of the inflammatory response. PMID:27782068

  12. Claudin-4 forms a paracellular barrier, revealing the interdependence of claudin expression in the loose epithelial cell culture model opossum kidney cells.

    PubMed

    Borovac, Jelena; Barker, Reid S; Rievaj, Juraj; Rasmussen, Andrew; Pan, Wanling; Wevrick, Rachel; Alexander, R Todd

    2012-12-15

    The effect of claudins on paracellular fluxes has been predominantly studied in either Madin-Darby canine kidney (MDCK) or LLCPK cells. Neither model system has a very low transepithelial resistance (TER) as observed in leaky epithelia. Moreover, results from one model system are not always consistent with another. Opossum kidney (OK) cells form tight junctions yet have a very low TER. We therefore set out to characterize the paracellular transport properties of this cell culture model. Ussing chamber dilution potential measurements revealed that OK cells exhibit a very low TER (11.7 ± 1.4 Ω·cm(2)), slight cation selectivity (P(Na)/P(Cl) = 1.10 ± 0.01), and the Eisenman permeability sequence IV; the permeability of monovalent cations ranking K(+) > Cs(+) > Rb(+) > Na(+) > Li(+). Quantitative real-time PCR studies found that OK cells endogenously express claudin-4 > -1 > -6 > -20 > -9 > -12 > -11 > -15. Overexpression of claudin-4 significantly increased TER, decreased Na(+) and Cl(-) permeability, and increased levels of claudin-1, -6, and -9 mRNA. Knockdown of claudin-4 in the overexpressing cells significantly decreased TER without altering claudin expression; thus claudin-4 forms a barrier in OK cells. Knockdown of endogenous claudin-4 decreased claudin-1, -9, and -12 expression without altering TER. Claudin-2 overexpression decreased TER, significantly increased Na(+) and Cl(-) permeability, and decreased claudin-12 and -6 expression. Together these results demonstrate that claudin expression is tightly coupled in OK cells.

  13. Chemo-radiotherapy induced oral mucositis during IMRT for head and neck cancer - An assessment

    PubMed Central

    2015-01-01

    Background This study is conducted mainly to evaluate the changes in quality and quantity of oral epithelial cells during the course of IMRT. Material and Methods 30 Patients undergoing chemo-radiotherapy were followed through course of treatment. They were compared with a group of age- and sex-matched healthy individuals. The procedure involved WHO clinical scoring, collection of oral washings and preparation of buccal smears from both study group and control group. The changes occurred were recorded as a way of assessing the severity of oral mucositis. Results Revealed a significant occurrence of oral mucositis in almost all patients during weekly follow up. There was a significant increase in percentage of viable buccal epithelial cells in study group when compared to normal controls (P<0.005) during and at the end of chemo-radiotherapy. Conclusions Quantification of oral mucositis can be done at cellular level by determining the oral mucosal cell viability and their maturation during IMRT. Key words:Oral mucositis, in vitro assay, quantification, radiotherapy, chemotherapy, viable cells. PMID:25662542

  14. A Single Dose of Oral BCG Moreau Fails to Boost Systemic IFN-γ Responses to Tuberculin in Children in the Rural Tropics: Evidence for a Barrier to Mucosal Immunization

    PubMed Central

    Vaca, Maritza; Moncayo, Ana-Lucia; Cosgrove, Catherine A.; Chico, Martha E.; Castello-Branco, Luiz R.; Lewis, David J.; Cooper, Philip J.

    2012-01-01

    Immune responses to oral vaccines are impaired in populations living in conditions of poverty in developing countries, and there is evidence that concurrent geohelminth infections may contribute to this effect. We vaccinated 48 children living in rural communities in Ecuador with a single oral dose of 100 mg of BCG Moreau RDJ and measured the frequencies of tuberculin-stimulated peripheral blood mononuclear cells expressing IFN-γ before and after vaccination. Vaccinated children had active ascariasis (n = 20) or had been infected but received short- (n = 13) or long-term (n = 15) repeated treatments with albendazole prior to vaccination to treat ascariasis. All children had a BCG scar from neonatal vaccination. There was no evidence of a boosting of postvaccination IFN-γ responses in any of the 3 study groups. Our data provide support for the presence of a barrier to oral vaccination among children from the rural tropics that appeared to be independent of concurrent ascariasis. PMID:22287972

  15. Other causes of GI mucosal injury: upper intestinal content.

    PubMed

    Ritchie, W P

    1987-05-01

    Factors in upper intestinal content that can produce acute injury to the gastric mucosa include lysolecithin and the bile acids. Both damage the gastric mucosal barrier by increasing mucosal permeability. The secondary and deconjugated bile acids are more toxic in this regard than are the primary or conjugated ones. The extent of injury is highly pH-dependent. Although the bile acids do not affect the gel properties of gastric mucus, they do produce significant inhibition of carbonic anhydrase activity and gastric bicarbonate secretion. In concert with other topical damaging agents, bile acids increase mucosal blood flow. However, gross mucosal lesions are rarely observed under these circumstances. Chronic exposure of acid-peptic-secreting mucosa to upper intestinal content results in the development of a severe atrophic gastritis within 6 months. The ability of atrophic mucosa to maintain an intraluminal pH gradient is impaired, and it ulcerates with great regularity when exposed to a highly acid environment. Clinically, excessive enterogastric reflux has been implicated in the pathogenesis of both benign gastric ulcer and the post-gastrectomy syndrome of alkaline reflux gastritis. The evidence to support this view for both disease entities is reviewed. PMID:3113802

  16. Bifidobacterium longum CCM 7952 Promotes Epithelial Barrier Function and Prevents Acute DSS-Induced Colitis in Strictly Strain-Specific Manner

    PubMed Central

    Srutkova, Dagmar; Schwarzer, Martin; Hudcovic, Tomas; Zakostelska, Zuzana; Drab, Vladimir; Spanova, Alena; Rittich, Bohuslav; Kozakova, Hana; Schabussova, Irma

    2015-01-01

    Background Reduced microbial diversity has been associated with inflammatory bowel disease (IBD) and probiotic bacteria have been proposed for its prevention and/or treatment. Nevertheless, comparative studies of strains of the same subspecies for specific health benefits are scarce. Here we compared two Bifidobacterium longum ssp. longum strains for their capacity to prevent experimental colitis. Methods Immunomodulatory properties of nine probiotic bifidobacteria were assessed by stimulation of murine splenocytes. The immune responses to B. longum ssp. longum CCM 7952 (Bl 7952) and CCDM 372 (Bl 372) were further characterized by stimulation of bone marrow-derived dendritic cell, HEK293/TLR2 or HEK293/NOD2 cells. A mouse model of dextran sulphate sodium (DSS)-induced colitis was used to compare their beneficial effects in vivo. Results The nine bifidobacteria exhibited strain-specific abilities to induce cytokine production. Bl 372 induced higher levels of both pro- and anti-inflammatory cytokines in spleen and dendritic cell cultures compared to Bl 7952. Both strains engaged TLR2 and contain ligands for NOD2. In a mouse model of DSS-induced colitis, Bl 7952, but not Bl 372, reduced clinical symptoms and preserved expression of tight junction proteins. Importantly, Bl 7952 improved intestinal barrier function as demonstrated by reduced FITC-dextran levels in serum. Conclusions We have shown that Bl 7952, but not Bl 372, protected mice from the development of experimental colitis. Our data suggest that although some immunomodulatory properties might be widespread among the genus Bifidobacterium, others may be rare and characteristic only for a specific strain. Therefore, careful selection might be crucial in providing beneficial outcome in clinical trials with probiotics in IBD. PMID:26218526

  17. Adhesins and ligands involved in the interaction of Candida spp. with epithelial and endothelial surfaces.

    PubMed Central

    Hostetter, M K

    1994-01-01

    Adhesion of candidal species to the epithelium of the gastrointestinal or genitourinary tract stands as a critical first step in the pathogenesis of candidal infection. After colonization and replication at mucosal surfaces, Candida albicans and other pathogenic species may penetrate the mucosal barrier, enter the vascular tree, and disseminate hematogenously. The consequences of this pathogenic cascade evoke considerable morbidity and mortality, especially among immunocompromised patients. Thus, interactions of C. albicans and other candidal species with epithelium and endothelium may lead to serious consequences for the human host. This review evaluates candidate candidal adhesions for epithelial and endothelial surfaces, with emphasis on the specificity of the interaction, the inhibitors that have been employed, and the ligands that have been identified on mammalian cells or matrices. Three types of interactions are described: protein-protein interactions, lectin-like interactions, and incompletely defined interactions in which the adhesive ligand is as yet unidentified. Special attention is given to the roles of integrin-like proteins. Differences in the mechanisms of candidal attachment to epithelium and endothelium are delineated. Last, on the basis of the available literature, avenues of potentially fruitful investigation are proposed. Images PMID:8118789

  18. Brain barriers: Crosstalk between complex tight junctions and adherens junctions

    PubMed Central

    Tietz, Silvia

    2015-01-01

    Unique intercellular junctional complexes between the central nervous system (CNS) microvascular endothelial cells and the choroid plexus epithelial cells form the endothelial blood–brain barrier (BBB) and the epithelial blood–cerebrospinal fluid barrier (BCSFB), respectively. These barriers inhibit paracellular diffusion, thereby protecting the CNS from fluctuations in the blood. Studies of brain barrier integrity during development, normal physiology, and disease have focused on BBB and BCSFB tight junctions but not the corresponding endothelial and epithelial adherens junctions. The crosstalk between adherens junctions and tight junctions in maintaining barrier integrity is an understudied area that may represent a promising target for influencing brain barrier function. PMID:26008742

  19. Pathogenesis of human papillomavirus-associated mucosal disease.

    PubMed

    Groves, Ian J; Coleman, Nicholas

    2015-03-01

    Human papillomaviruses (HPVs) are a necessary cause of carcinoma of the cervix and other mucosal epithelia. Key events in high-risk HPV (HRHPV)-associated neoplastic progression include persistent infection, deregulated expression of virus early genes in basal epithelial cells and genomic instability causing secondary host genomic imbalances. There are multiple mechanisms by which deregulated virus early gene expression may be achieved. Integration of virus DNA into host chromosomes is observed in the majority of cervical squamous cell carcinomas (SCCs), although in ∼15% of cases the virus remains extrachromosomal (episomal). Interestingly, not all integration events provide a growth advantage to basal cervical epithelial cells or lead to increased levels of the virus oncogenes E6 and E7, when compared with episome-containing basal cells. The factors that provide a competitive advantage to some integrants, but not others, are complex and include virus and host contributions. Gene expression from integrated and episomal HRHPV is regulated through host epigenetic mechanisms affecting the virus long control region (LCR), which appear to be of functional importance. New approaches to treating HRHPV-associated mucosal neoplasia include knockout of integrated HRHPV DNA, depletion of virus transcripts and inhibition of virus early gene transcription through targeting or use of epigenetic modifiers. Copyright © 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  20. SIgA Binding to Mucosal Surfaces Is Mediated by Mucin-Mucin Interactions

    PubMed Central

    Gibbins, Hannah L.; Proctor, Gordon B.; Yakubov, Gleb E.; Wilson, Stephen; Carpenter, Guy H.

    2015-01-01

    The oral mucosal pellicle is a layer of absorbed salivary proteins, including secretory IgA (SIgA), bound onto the surface of oral epithelial cells and is a useful model for all mucosal surfaces. The mechanism by which SIgA concentrates on mucosal surfaces is examined here using a tissue culture model with real saliva. Salivary mucins may initiate the formation of the mucosal pellicle through interactions with membrane-bound mucins on cells. Further protein interactions with mucins may then trigger binding of other pellicle proteins. HT29 colon cell lines, which when treated with methotrexate (HT29-MTX) produce a gel-forming mucin, were used to determine the importance of these mucin-mucin interactions. Binding of SIgA to cells was then compared using whole mouth saliva, parotid (mucin-free) saliva and a source of purified SIgA. Greatest SIgA binding occurred when WMS was incubated with HT29-MTX expressing mucus. Since salivary MUC5B was only able to bind to cells which produced mucus and purified SIgA showed little binding to the same cells we conclude that most SIgA binding to mucosal cells occurs because SIgA forms complexes with salivary mucins which then bind to cells expressing membrane-bound mucins. This work highlights the importance of mucin interactions in the development of the mucosal pellicle. PMID:25793390

  1. Identification and mucosal expression analysis of cathepsin B in channel catfish (Ictalurus punctatus) following bacterial challenge.

    PubMed

    Li, Chao; Song, Lin; Tan, Fenghua; Su, Baofeng; Zhang, Dongdong; Zhao, Honggang; Peatman, Eric

    2015-12-01

    The mucosal surfaces of fish (skin, gill and intestine) constitute the primary line of defense against pathogen invasion. Although the importance of fish mucosal surfaces as the first barriers against pathogens cannot be overstated, the knowledge of teleost mucosal immunity are still limited. Cathepsin B, a lysosomal cysteine protease, is involved in multiple levels of physiological and biological processes, and playing crucial roles for host immune defense against pathogen infection. In this regard, we identified the cathepsin B (ctsba) of channel catfish and investigated the expression patterns of the ctsba in mucosal tissues following Edwardsiella ictaluri and Flavobacterium columnare challenge. Here, catfish ctsba gene was widely expressed in all examined tissues with the lowest expression level in muscle, and the highest expression level in trunk kidney, followed by spleen, gill, head kidney, intestine, liver and skin. In addition, the phylogenetic analysis showed the catfish ctsba had the strongest relationship to zebrafish. Moreover, the ctsba showed a general trend of up-regulated in mucosal tissues following both Gram-negative bacterial challenge. Taken together, the increased expression of ctsba in mucosal surfaces indicated the protective function of ctsba against bacterial infection, and the requirement for effective clearance of invading bacteria. Further studies are needed, indeed, to expand functional characterization and examine whether ctsba may play additional physiological and biological roles in catfish mucosal tissues. PMID:26497091

  2. The mucosal expression signatures of g-type lysozyme in turbot (Scophthalmus maximus) following bacterial challenge.

    PubMed

    Gao, Chengbin; Fu, Qiang; Zhou, Shun; Song, Lin; Ren, Yichao; Dong, Xiaoyu; Su, Baofeng; Li, Chao

    2016-07-01

    The mucosal surfaces constitute the first line of host defense against infection, and also serve as the dynamic interfaces that simultaneously mediate a diverse array of critical physiological processes, while in constantly contact with a wide range of pathogens. The lysozymes are considered as key components for innate immune response to pathogen infection with their strong antibacterial activities. But their activities in mucosal immune responses were always overlooked, especially for g-type lysozymes, whose expression patterns in mucosal tissues following bacterial challenge are still limited. Towards to this end, here, we characterized the g-type lysozymes, Lyg1 and Lyg2 in turbot, and determined their expression patterns in mucosal barriers following different bacterial infection. The phylogenetic analysis revealed the turbot g-type lysozyme genes showed the closest relationship to Cynoglossus semilaevis. The two lysozyme genes showed different expression patterns following challenge. Lyg2 was significantly up-regulated in mucosal tissues following Vibrio anguillarum and Streptococcus iniae challenge, while Lyg1 showed a general trend of down-regulation. The significant mucosal expression signatures of g-type lysozyme genes indicated their key roles to prevent pathogen attachment and entry in the first line of host defense system. Further functional studies should be carried out to better characterize the availability of utilization of g-type lysozyme to increase the disease resistance in the mucosal surfaces and facilitate the disease resistant breeding selection. PMID:27189917

  3. Mucus-penetrating nanoparticles for drug and gene delivery to mucosal tissues

    PubMed Central

    Lai, Samuel K.; Wang, Ying-Ying; Hanes, Justin

    2009-01-01

    Mucus is a viscoelastic and adhesive gel that protects the lung airways, gastrointestinal (GI) tract, vagina, eye and other mucosal surfaces. Most foreign particulates, including conventional particle-based drug delivery systems, are efficiently trapped in human mucus layers by steric obstruction and/or adhesion. Trapped particles are typically removed from the mucosal tissue within seconds to a few hours depending on anatomical location, thereby strongly limiting the duration of sustained drug delivery locally. A number of debilitating diseases could be treated more effectively and with fewer side effects if drugs and genes could be more efficiently delivered to the underlying mucosal tissues in a controlled manner. This review first describes the tenacious mucus barrier properties that have precluded the efficient penetration of therapeutic particles. It then reviews the design and development of new mucus-penetrating particles that may avoid rapid mucus clearance mechanisms, and thereby provide targeted or sustained drug delivery for localized therapies in mucosal tissues. PMID:19133304

  4. Role of topical drugs in treatment of oral mucosal diseases. A literature review.

    PubMed

    Sheikh, Soheyl; Gupta, Deepak; Pallagatti, Shambulingappa; Singla, Isha; Gupta, Rajesh; Goel, Varun

    2013-11-01

    Few topical formulations have been designed specifically to treat oral mucosal diseases. Local drug delivery may provide a more targeted and efficient option than systemic delivery for diseases of the oral mucosa. The permeability to the topical drugs differs according to the thickness of the epithelium and the extent of keratinization. The loss of the permeability barrier in the oral mucosa, due to ulceration or erosion, leads to rapid diffusion of the drug into tissues as compared to the intact areas of the mucosa. Oral mucosal delivery has the potential to treat many different conditions and diseases, such as oral cancer, mucositis, lichen planus, herpes simplex, candidiasis, recurrent aphthous stomatitis, vesiculo-bullous diseases, neuropathic pain and salivary dysfunction. Each therapy requires distinct penetration and drug retention profiles in order to optimize treatment and minimize side effects. In this paper, topical medications are discussed, as these are advantageous for the treatment of oral mucosal lesions with fewer side effects.

  5. 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

  6. Reinforcement of epithelial cell adhesion to basement membrane by a bacterial pathogen as a new infectious stratagem.

    PubMed

    Kim, Minsoo; Ogawa, Michinaga; Mimuro, Hitomi; Sasakawa, Chihiro

    2010-01-01

    The intestinal epithelium undergoes a rapid turnover in addition to rapid exfoliation in response to bacterial infection, thus acting as an intrinsic defense against microbial intruders. It has long been questioned how mucosal pathogens can circumvent the intestinal defense systems. Our recent discovery of a bacterial ploy used by Shigella provided us with fresh insight. Shigella delivers OspE via the type III secretion system during multiplication within epithelial cells. This effector protein reinforces epithelial adherence to the basement membrane by interacting with integrin-linked kinase (ILK), a unique intracellular Ser/Thr kinase that links the cell-adhesion receptors, integrin, and growth factors to the actin cytoskeleton. The interaction between OspE and ILK increased formation of focal adhesions (FAs) and surface levels of b1-integrin, while suppressing phosphorylation of FAK and paxillin, thus suppressing rapid turnover of FAs, reducing cell motility and promoting cell adhesion to extracellular matrix. The impact of this OspE-ILK interplay was demonstrated both in vitro and in vivo by infecting polarized epithelial cell monolayers and guinea pig colons with Shigella possessing or lacking the ospE gene. The findings thus establish a new class of virulence-associated factors, and provide new insight into the functioning of the intestinal barrier and bacterial strategies for circumventing it. PMID:21178415

  7. The junction zone of human epithelium and foreign stroma. Ultrastructural observations in human oral mucosal transplants in nude mice.

    PubMed

    Holmstrup, P; Andersen, L; Harder, F

    1984-07-01

    This study describes ultrastructural features of epithelial-stromal junction of human buccal and palatal mucosal transplants and their outgrowths in nude mice. The investigation included transplant epithelium overlying human connective tissue in 27 cases, epithelial outgrowth formed over murine connective tissue in 33 cases, and over Millipore filter in 12 cases. The epithelial-stromal junction of the transplants differed from the normal state only in the presence of lamina densa loops projecting into the connective tissue and in lamina densa interruptions and duplications. In contrast the epithelial outgrowths demonstrated flattening of epithelial basal cells, lack of proximal epithelial cell projections, lack of complete hemidesmosome complexes, lack of distinct lamina densa, and lack of anchoring fibrils. It is suggested that these changes may be due to lack of necessary interaction between the human epithelium and the foreign stroma.

  8. Primary mucosal melanomas: a comprehensive review

    PubMed Central

    Mihajlovic, Marija; Vlajkovic, Slobodan; Jovanovic, Predrag; Stefanovic, Vladisav

    2012-01-01

    Primary mucosal melanomas arise from melanocytes located in mucosal membranes lining respiratory, gastrointestinal and urogenital tract. Although a majority of mucosal melanomas originate from the mucosa of the nasal cavity and accessory sinuses, oral cavity, anorectum, vulva and vagina, they can arise in almost any part of mucosal membranes. Most of mucosal melanomas occur in occult sites, which together with the lack of early and specific signs contribute to late diagnosis, and poor prognosis. Because of their rareness the knowledge about their pathogenesis and risk factors is insufficient, and also there are not well established protocols for staging and treatment of mucosal melanomas. Surgery is the mainstay of treatment, with trends toward more conservative treatment since radical surgery did not show an advantage for survival. Radiotherapy can provide better local control in some locations, but did not show improvement in survival. There is no effective systemic therapy for these aggressive tumors. Compared with cutaneous and ocular melanoma, mucosal melanomas have lowest percent of five-year survival. Recently revealed molecular changes underlying mucosal melanomas offer new hope for development of more effective systemic therapy for mucosal melanomas. Herein we presented a comprehensive review of various locations of primary melanoma along mucosal membranes, their epidemiological and clinical features, and treatment options. We also gave a short comparison of some characteristics of cutaneous and mucosal melanomas. PMID:23071856

  9. The Mucosal Immune System of Teleost Fish

    PubMed Central

    Salinas, Irene

    2015-01-01

    Teleost fish possess an adaptive immune system associated with each of their mucosal body surfaces. Evidence obtained from mucosal vaccination and mucosal infection studies reveal that adaptive immune responses take place at the different mucosal surfaces of teleost. The main mucosa-associated lymphoid tissues (MALT) of teleosts are the gut-associated lymphoid tissue (GALT), skin-associated lymphoid tissue (SALT), the gill-associated lymphoid tissue (GIALT) and the recently discovered nasopharynx-associated lymphoid tissue (NALT). Teleost MALT includes diffuse B cells and T cells with specific phenotypes different from their systemic counterparts that have co-evolved to defend the microbe-rich mucosal environment. Both B and T cells respond to mucosal infection or vaccination. Specific antibody responses can be measured in the gills, gut and skin mucosal secretions of teleost fish following mucosal infection or vaccination. Rainbow trout studies have shown that IgT antibodies and IgT+ B cells are the predominant B cell subset in all MALT and respond in a compartmentalized manner to mucosal infection. Our current knowledge on adaptive immunity in teleosts is limited compared to the mammalian literature. New research tools and in vivo models are currently being developed in order to help reveal the great intricacy of teleost mucosal adaptive immunity and help improve mucosal vaccination protocols for use in aquaculture. PMID:26274978

  10. 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

  11. 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

  12. Role of HMGB1 in propofol protection of rat intestinal epithelial cells injured by heat shock.

    PubMed

    Tang, Jing; Deng, Peng; Jiang, Yu; Tang, Youqing; Chen, Bin; Su, Lei; Liu, Zhifeng

    2013-03-01

    Gut-derived endotoxin and pathogenic bacteria may be important causative factors of morbidity and death during heat stroke. However, as the key component of intestinal mucosal barrier, the molecular mechanism of how intestinal epithelial cells are injured by heat shock is remains unclear. After rat intestinal epithelial cells (IEC-6) had been exposed to heat shock, their viability was measured. Propofol, which plays an important role in anti-inflammation and organ protection, was investigated to see how it affected viability under this stress. Changes of high mobility group box 1 (HMGB1) in IEC-6 cells were measured with RT-PCR and Western blot assay at transcription and translational levels, respectively. Ethyl pyruvate (EP), a specific inhibitor of HMGB1 that can inhibit the release of HMGB1 without affecting its intracellular synthesis, was also investigated. Heat shock significantly reduced the intracellular level of HMGB1, and propofol inhibit its reduction. Propofol protected the heat shock-injured cells, at least partly through inhibiting the release of intracellular HMGB1 to reduce the direct or indirect cell damage caused by HMGB1. Pretreatment with high concentrations of EP also attenuated heat-shock injury.

  13. Cryopreservation of Human Mucosal Leukocytes

    PubMed Central

    Shu, Zhiquan; Levy, Claire N.; Ferre, April L.; Hartig, Heather; Fang, Cifeng; Lentz, Gretchen; Fialkow, Michael; Kirby, Anna C.; Adams Waldorf, Kristina M.; Veazey, Ronald S.; Germann, Anja; von Briesen, Hagen; McElrath, M. Juliana; Dezzutti, Charlene S.; Sinclair, Elizabeth; Baker, Chris A. R.; Shacklett, Barbara L.; Gao, Dayong; Hladik, Florian

    2016-01-01

    Background Understanding how leukocytes in the cervicovaginal and colorectal mucosae respond to pathogens, and how medical interventions affect these responses, is important for developing better tools to prevent HIV and other sexually transmitted infections. An effective cryopreservation protocol for these cells following their isolation will make studying them more feasible. Methods and Findings To find an optimal cryopreservation protocol for mucosal mononuclear leukocytes, we compared cryopreservation media and procedures using human vaginal leukocytes and confirmed our results with endocervical and colorectal leukocytes. Specifically, we measured the recovery of viable vaginal T cells and macrophages after cryopreservation with different cryopreservation media and handling procedures. We found several cryopreservation media that led to recoveries above 75%. Limiting the number and volume of washes increased the fraction of cells recovered by 10–15%, possibly due to the small cell numbers in mucosal samples. We confirmed that our cryopreservation protocol also works well for both endocervical and colorectal leukocytes. Cryopreserved leukocytes had slightly increased cytokine responses to antigenic stimulation relative to the same cells tested fresh. Additionally, we tested whether it is better to cryopreserve endocervical cells on the cytobrush or in suspension. Conclusions Leukocytes from cervicovaginal and colorectal tissues can be cryopreserved with good recovery of functional, viable cells using several different cryopreservation media. The number and volume of washes has an experimentally meaningful effect on the percentage of cells recovered. We provide a detailed, step-by-step protocol with best practices for cryopreservation of mucosal leukocytes. PMID:27232996

  14. Intestinal trefoil factor activates the PI3K/Akt signaling pathway to protect gastric mucosal epithelium from damage.

    PubMed

    Sun, Zhaorui; Liu, Hongmei; Yang, Zhizhou; Shao, Danbing; Zhang, Wei; Ren, Yi; Sun, Baodi; Lin, Jinfeng; Xu, Min; Nie, Shinan

    2014-09-01

    Intestinal trefoil factor (ITF, also named as trefoil factor 3, TFF3) is a member of the TFF-domain peptide family, which plays an essential role in the regulation of cell survival, cell migration and maintains mucosal epithelial integrity in the gastrointestinal tract. However, the underlying mechanisms and associated molecules remain unclear. The aim of this study was to explore the protective effects of ITF on gastric mucosal epithelium injury and its possible molecular mechanisms of action. In the present study, we show that ITF was able to promote the proliferation and migration of GES-1 cells via a mechanism that involves the PI3K/Akt signaling pathway. Western blot results indicated that ITF induced a dose- and time-dependent increase in the Akt signaling pathway. ITF also plays an essential role in the restitution of GES-1 cell damage induced by lipopolysaccharide (LPS). LPS induced the apoptosis of GES-1 cells, decreased cell viability significantly (P<0.01) and led to epithelial tight junction damage, which is attenuated via ITF treatment. The protective effect of ITF on the integrity of GES-1 was abrogated by inhibition of the PI3K/Akt pathway. Taken together, our results demonstrate that ITF promotes the proliferation and migration of gastric mucosal epithelial cells and preserves gastric mucosal epithelial integrity after damage is mediated by activation of the PI3K/Akt signaling pathway. This study suggested that the PI3K/Akt pathway could act as a key intracellular pathway in the gastric mucosal epithelium that may serve as a therapeutic target to preserve epithelial integrity during injury.

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

    PubMed

    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.

  16. 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

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

    PubMed

    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

  18. Advances and challenges in mucosal adjuvant technology.

    PubMed

    Newsted, Daniel; Fallahi, Firouzeh; Golshani, Ashkan; Azizi, Ali

    2015-05-15

    Adjuvants play attractive roles in enhancement of immune response during vaccination; however, due to several challenges, only a limited number of adjuvants are licensed by health authorities. The lack of an effective mucosal adjuvant is even more significant as none of the licensed adjuvants revealed a strong enhancement in immune system after mucosal administration. Over the past two decades, several mucosal adjuvants have been developed to deliver antigens to the target cells in the mucosal immune system and increase specific immune responses. However, the safety and efficacy of these adjuvants for testing in human trials is still an important issue, requiring further study. In this article, we briefly review the challenges associated with most common mucosal adjuvants and discuss potential strategies for targeting the mucosal immune system.

  19. Assessment of mucosal immunity to HIV-1.

    PubMed

    Jespers, Vicky; Harandi, Ali M; Hinkula, Jorma; Medaglini, Donata; Le Grand, Roger; Stahl-Hennig, Christiane; Bogers, Willy; El Habib, Raphaelle; Wegmann, Frank; Fraser, Carol; Cranage, Martin; Shattock, Robin J; Spetz, Anna-Lena

    2010-04-01

    A key gap in the development and evaluation of HIV-1 vaccines is insufficient knowledge with regard to sampling techniques and assessment of mucosal immune responses required for early prevention and inhibition of viral dissemination. In an attempt to start bridging this gap, the EUROPRISE network of scientists working on HIV-1 vaccine and microbicide research organized a workshop with the aim to review the types of mucosal responses/biomarkers currently measured in mucosal immunology and to define how the mucosal responses/biomarkers are measured and/or the assays and sampling methods used. The Workshop addressed two critical questions: first whether, with current knowledge, it would be possible to define a consensus set of mucosal sampling methods to facilitate cross-species comparisons and ensure standardized implementation in clinical trials; second to determine the remaining challenges (technical and logistical) and their possible solutions for assessing mucosal responses to HIV-1 vaccines. PMID:20370549

  20. Focal epithelial hyperplasia - an update.

    PubMed

    Said, Ahmed K; Leao, Jair C; Fedele, Stefano; Porter, Stephen R

    2013-07-01

    Focal epithelial hyperplasia (FEH) is an asymptomatic benign mucosal disease, which is mostly observed in specific groups in certain geographical regions. FEH is usually a disease of childhood and adolescence and is generally associated with people who live in poverty and of low socioeconomic status. Clinically, FEH is typically characterized by multiple, painless, soft, sessile papules, plaques or nodules, which may coalesce to give rise to larger lesions. Human papillomavirus (HPV), especially genotypes 13 and 32, have been associated and detected in the majority of FEH lesions. The clinical examination and social history often allow diagnosis, but histopathological examination of lesional tissue is usually required to confirm the exact diagnosis. FEH sometimes resolves spontaneously however, treatment is often indicated as a consequence of aesthetic effects or any interference with occlusion. There remains no specific therapy for FEH, although surgical removal, laser excision or possibly topical antiviral agents may be of benefit. There remains no evidence that FEH is potentially malignant.

  1. Chemotherapy or radiation-induced oral mucositis.

    PubMed

    Lalla, Rajesh V; Saunders, Deborah P; Peterson, Douglas E

    2014-04-01

    Oral mucositis is a significant toxicity of systemic chemotherapy and of radiation therapy to the head and neck region. The morbidity of oral mucositis can include pain, nutritional compromise, impact on quality of life, alteration in cancer therapy, risk for infection, and economic costs. Management includes general symptomatic support and targeted therapeutic interventions for the prevention or treatment of oral mucositis. Evidence-based clinical practice guidelines are available to guide clinicians in the selection of effective management strategies.

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

    PubMed

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

    2001-04-01

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

  3. 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

  4. Lactose maldigestion during methotrexate-induced gastrointestinal mucositis in a rat model.

    PubMed

    Fijlstra, M; Rings, E H H M; Verkade, H J; van Dijk, T H; Kamps, W A; Tissing, W J E

    2011-02-01

    Patients with chemotherapy-induced gastrointestinal mucositis suffer from anorexia, diarrhea, and stomach pain, often causing weight loss and malnutrition. When the intestinal function during mucositis would be known, a rational feeding strategy might improve the nutritional state, accelerate recuperation, and increase survival of mucositis patients. We developed a methotrexate (MTX)-induced mucositis rat model to study nutrient digestion and absorption. To determine lactose digestion and absorption of its derivative glucose during mucositis, we injected Wistar rats intravenously with MTX (60 mg/kg) or 0.9% NaCl (controls). Four days later, we orally administered trace amounts of [1-(13)C]lactose and [U-(13)C]glucose and quantified the appearance of labeled glucose in the blood for 3 h. Finally, we determined plasma citrulline level and harvested the small intestine to assess histology, myeloperoxidase level, glycohydrolase activity, immunohistochemical protein, and mRNA expression. MTX-treated rats showed profound villus atrophy and epithelial damage. During the experimental period, the absorption of lactose-derived [1-(13)C]glucose was 4.2-fold decreased in MTX-treated rats compared with controls (P < 0.01). Lactose-derived [1-(13)C]glucose absorption correlated strongly with villus length (rho = 0.86, P < 0.001) and with plasma citrulline level (rho = 0.81, P < 0.001). MTX treatment decreased jejunal lactase activity (19.5-fold, P < 0.01) and immunohistochemical protein and mRNA expression (39.7-fold, P < 0.01) compared with controls. Interestingly, MTX treatment did not affect the absorption of [U-(13)C]glucose during the experimental period. We conclude that lactose digestion is severely decreased during mucositis while glucose absorption is still intact, when supplied in trace amounts. Plasma citrulline level might be a useful objective, noninvasive marker for lactose maldigestion during mucositis in clinic.

  5. ALDH2 Deficiency Promotes Ethanol-Induced Gut Barrier Dysfunction and Fatty Liver in Mice

    PubMed Central

    Chaudhry, Kamljit K.; Samak, Geetha; Shukla, Pradeep K.; Mir, Hina; Gangwar, Ruchika; Manda, Bhargavi; Isse, Toyohi; Kawamoto, Toshihiro; Salaspuro, Mikko; Kaihovaara, Pertti; Dietrich, Paula; Dragatsis, Ioannis; Nagy, Laura E.; Rao, RadhaKrishna

    2015-01-01

    Background Acetaldehyde, the toxic ethanol metabolite, disrupts intestinal epithelial barrier function. Aldehyde dehydrogenase (ALDH) detoxifies acetaldehyde into acetate. Sub populations of Asians and Native Americans show polymorphism with loss of function mutations in ALDH2. We evaluated the effect of ALDH2 deficiency on ethanol-induced disruption of intestinal epithelial tight junctions and adherens junctions, gut barrier dysfunction and liver injury. Methods Wild type and ALDH2 deficient mice were fed (1–6%) in Lieber-DeCarli diet for 4 weeks. Gut permeability in vivo measured by plasma-to-luminal flux of FITC-inulin, tight junction and adherens junction integrity analyzed by confocal microscopy and liver injury was assessed by analysis of plasma transaminase activity, histopathology and liver triglyceride. Results Ethanol feeding elevated colonic mucosal acetaldehyde, which was significantly greater in ALDH2 deficient mice. ALDH2−/− mice showed a drastic reduction in the ethanol diet intake. Therefore, this study was continued only in wild type and ALDH2+/− mice. Ethanol feeding elevated mucosal inulin permeability in distal colon, but not in proximal colon, ileum or jejunum of wild type mice. In ALDH2+/− mice, ethanol-induced inulin permeability in distal colon was not only higher than that in wild type mice, but inulin permeability was also elevated in the proximal colon, ileum and jejunum. Greater inulin permeability in distal colon of ALDH2+/− mice was associated with a more severe redistribution of tight junction and adherens junction proteins from the intercellular junctions. In ALDH2+/− mice, but not in wild type mice, ethanol feeding caused a loss of junctional distribution of tight junction and adherens junction proteins in the ileum. Histopathology, plasma transaminases and liver triglyceride analyses showed that ethanol-induced liver damage was significantly greater in ALDH2+/− mice compared to wild type mice. Conclusion These data

  6. Voice Disorders in Mucosal Leishmaniasis

    PubMed Central

    Ruas, Ana Cristina Nunes; Lucena, Márcia Mendonça; da Costa, Ananda Dutra; Vieira, Jéssica Rafael; de Araújo-Melo, Maria Helena; Terceiro, Benivaldo Ramos Ferreira; de Sousa Torraca, Tania Salgado; de Oliveira Schubach, Armando; Valete-Rosalino, Claudia Maria

    2014-01-01

    Introduction Leishmaniasis is considered as one of the six most important infectious diseases because of its high detection coefficient and ability to produce deformities. In most cases, mucosal leishmaniasis (ML) occurs as a consequence of cutaneous leishmaniasis. If left untreated, mucosal lesions can leave sequelae, interfering in the swallowing, breathing, voice and speech processes and requiring rehabilitation. Objective To describe the anatomical characteristics and voice quality of ML patients. Materials and Methods A descriptive transversal study was conducted in a cohort of ML patients treated at the Laboratory for Leishmaniasis Surveillance of the Evandro Chagas National Institute of Infectious Diseases - Fiocruz, between 2010 and 2013. The patients were submitted to otorhinolaryngologic clinical examination by endoscopy of the upper airways and digestive tract and to speech-language assessment through directed anamnesis, auditory perception, phonation times and vocal acoustic analysis. The variables of interest were epidemiologic (sex and age) and clinic (lesion location, associated symptoms and voice quality. Results 26 patients under ML treatment and monitored by speech therapists were studied. 21 (81%) were male and five (19%) female, with ages ranging from 15 to 78 years (54.5+15.0 years). The lesions were distributed in the following structures 88.5% nasal, 38.5% oral, 34.6% pharyngeal and 19.2% laryngeal, with some patients presenting lesions in more than one anatomic site. The main complaint was nasal obstruction (73.1%), followed by dysphonia (38.5%), odynophagia (30.8%) and dysphagia (26.9%). 23 patients (84.6%) presented voice quality perturbations. Dysphonia was significantly associated to lesions in the larynx, pharynx and oral cavity. Conclusion We observed that vocal quality perturbations are frequent in patients with mucosal leishmaniasis, even without laryngeal lesions; they are probably associated to disorders of some resonance

  7. Free fucose is a danger signal to human intestinal epithelial cells.

    PubMed

    Chow, Wai Ling; Lee, Yuan Kun

    2008-03-01

    Fucose is present in foods, and it is a major component of human mucin glycoproteins and glycolipids. l-Fucose can also be found at the terminal position of many cell-surface oligosaccharide ligands that mediate cell-recognition and adhesion-signalling pathways. Mucin fucose can be released through the hydrolytic activity of pathogens and indigenous bacteria, leading to the release of free fucose into the intestinal lumen. The immunomodulating effects of free fucose on intestinal epithelial cells (enterocyte-like Caco-2) were investigated. It was found that the presence of l-fucose up regulated genes and secretion of their encoded proteins that are involved in both the innate and adaptive immune responses, possibly via the toll-like receptor-2 signalling pathway. These include TNFSF5, TNFSF7, TNF-alpha, IL12, IL17 and IL18. Besides modulating immune reactions in differentiated Caco-2 cells, fucose induced a set of cytokine genes that are involved in the development and proliferation of immune cells. These include the bone morphogenetic proteins (BMP) BMP2, BMP4, IL5, thrombopoietin and erythropoietin. In addition, the up regulated gene expression of fibroblast growth factor-2 may help to promote epithelial cell restitution in conjunction with the enhanced expression of transforming growth factor-beta mRNA. Since the exogenous fucose was not metabolised by the differentiated Caco-2 cells as a carbon source, the reactions elicited were suggested to be a result of the direct interaction of fucose and differentiated Caco-2 cells. The presence of free fucose may signal the invasion of mucin-hydrolysing microbial cells and breakage of the mucosal barrier. The intestinal epithelial cells respond by up regulation and secretion of cytokines, pre-empting the actual invasion of pathogens.

  8. Interleukin 17A evoked mucosal damage is attenuated by cannabidiol and anandamide in a human colonic explant model.

    PubMed

    Harvey, B S; Sia, T C; Wattchow, D A; Smid, S D

    2014-02-01

    Interleukin 17A (IL-17A) is a cytokine linked to inflammatory bowel disease. We investigated IL-17A expression in human colonic mucosa, whether IL-17A can elicit colonic mucosal damage in a human explant model and modulate gastrointestinal epithelial permeability in cell culture. We also tested if select cannabinoid ligands, shown to be protective in colitis models could attenuate damage caused by IL-17A. In addition, the ability of pro-inflammatory cytokines TNF-α and IL-1β to modulate levels of IL-17A in the explant colitis model was also explored. IL-17A incubation caused significant mucosal epithelial and crypt damage which were attenuated following hydrocortisone treatment, and also reduced following anandamide or cannabidiol incubation. IL-17A-evoked mucosal damage was also associated with an increase in matrix metalloprotease activity. However, IL-17A did not induce any significant changes in epithelial permeability in confluent Caco-2 cell monolayers over a 48h incubation period. IL-17A was located predominantly in human mucosal epithelium together with IL-17C, but both IL-17A and IL-17C were also expressed in the lamina propria and submucosa. Incubation of human colonic mucosal tissue or Caco-2 cells with pro-inflammatory cytokines TNF-α and IL-1β however did not alter IL-17A expression. These results indicate IL-17A has a widespread distribution in the human colon and the capacity to elicit mucosal damage which can be attenuated by cannabinoid ligands.

  9. Innate Immune Responses after Airway Epithelial Stimulation with Mycobacterium bovis Bacille-Calmette Guérin

    PubMed Central

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

    2016-01-01

    Mycobacterium bovis bacilli Calmette-Guerin (BCG) is used as a benchmark to compare the immunogenicity of new vaccines against tuberculosis. This live vaccine is administered intradermal, but several new studies show that changing the route to mucosal immunisation represents an improved strategy. We analysed the immunomodulatory functions of BCG on human neutrophils and primary airway epithelial cells (AECs), as the early events of mucosal immune activation are unclear. Neutrophils and the primary epithelial cells were found to express the IL-17A receptor subunit IL-17RA, while the expression of IL-17RE was only observed on epithelial cells. BCG stimulation specifically reduced neutrophil IL-17RA and epithelial IL-17RE expression. BCG induced neutrophil extracellular traps (NETs), but did not have an effect on apoptosis as measured by transcription factor forkhead box O3 (FOXO3). BCG stimulation of AECs induced CXCL8 secretion and neutrophil endothelial passage towards infected epithelia. Infected epithelial cells and neutrophils were not found to be a source of IL-17 cytokines or the interstitial collagenase MMP-1. However, the addition of IFNγ or IL-17A to BCG stimulated primary epithelial cells increased epithelial IL-6 secretion, while the presence of IFNγ reduced neutrophil recruitment. Using our model of mucosal infection we revealed that BCG induces selective mucosal innate immune responses that could lead to induction of vaccine-mediated protection of the lung. PMID:27723804

  10. Airway cooling and mucosal injury during cold weather exercise.

    PubMed

    Davis, M S; Lockard, A J; Marlin, D J; Freed, A N

    2002-09-01

    In human subjects that exercise strenuously in cold weather, there is evidence that hyperventilation with cold air leads to peripheral airway cooling, desiccation and mucosal injury. Our hypothesis was that hyperventilation with cold air can result in penetration of unconditioned air (air that is not completely warmed and humidified) into the peripheral airways of exercising horses, resulting in peripheral airway mucosal injury. To test this hypothesis, a thermister-tipped catheter was inserted through the midcervical trachea and advanced into a sublobar bronchus in three horses that cantered on a treadmill at 6.6 m/s while breathing cold (5 degrees C) air. The mean (+/- s.e.) intra-airway temperature during cantering was 33.3 +/- 0.4 degrees C, a value comparable to the bronchial lumen temperatures measured in man during maximal exercise while breathing subfreezing dry air. In a second experiment, 6 fit Thoroughbred racehorses with satisfactory performance were used to determine whether strenuous exercise in cold conditions can produce airway injury. Horses were assigned to Exercise (E) or Control (C) groups in a random crossover design. Samples of bronchoalveolar lavage fluid (BALF) in the E treatment were recovered within 30 min of galloping exercise in 4 degrees C, 100% relative humidity (E), while in C BALF samples were obtained when the horses had not performed any exercise for at least 48 h prior. Ciliated epithelial cells in BALF were higher in E than in the C treatment. Similar results have been found in human athletes and laboratory animal models of cold weather exercise. These results support the hypothesis that, similar to man, horses that exercise in cold weather experience peripheral airway mucosal injury due to the penetration of unconditioned air. Furthermore, these results suggest that airway cooling and desiccation may be a factor in airway inflammation commonly found in equine athletes.

  11. Effect of wild-type Shigella species and attenuated Shigella vaccine candidates on small intestinal barrier function, antigen trafficking, and cytokine release.

    PubMed

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

    2014-01-01

    Bacterial dysentery due to Shigella species is a major cause of morbidity and mortality worldwide. The pathogenesis of Shigella is based on the bacteria's ability to invade and replicate within the colonic epithelium, resulting in severe intestinal inflammatory response and epithelial destruction. Although the mechanisms of pathogenesis of Shigella in the colon have been extensively studied, little is known on the effect of wild-type Shigella on the small intestine and the role of the host response in the development of the disease. Moreover, to the best of our knowledge no studies have described the effects of apically administered Shigella flexneri 2a and S. dysenteriae 1 vaccine strains on human small intestinal enterocytes. The aim of this study was to assess the coordinated functional and immunological human epithelial responses evoked by strains of Shigella and candidate vaccines on small intestinal enterocytes. To model the interactions of Shigella with the intestinal mucosa, we apically exposed monolayers of human intestinal Caco2 cells to increasing bacterial inocula. We monitored changes in paracellular permeability, examined the organization of tight-junctions and the pro-inflammatory response of epithelial cells. Shigella infection of Caco2 monolayers caused severe mucosal damage, apparent as a drastic increase in paracellular permeability and disruption of tight junctions at the cell-cell boundary. Secretion of pro-inflammatory IL-8 was independent of epithelial barrier dysfunction. Shigella vaccine strains elicited a pro-inflammatory response without affecting the intestinal barrier integrity. Our data show that wild-type Shigella infection causes a severe alteration of the barrier function of a small intestinal cell monolayer (a proxy for mucosa) and might contribute (along with enterotoxins) to the induction of watery diarrhea. Diarrhea may be a mechanism by which the host attempts to eliminate harmful bacteria and transport them from the small to

  12. Mucosal Wave Measurement and Visualization Techniques

    PubMed Central

    Krausert, Christopher R.; Olszewski, Aleksandra E.; Taylor, Lindsay N.; McMurray, James S.; Dailey, Seth H.; Jiang, Jack J.

    2010-01-01

    Organized vibration of the vocal folds is critical to high quality voice production. When the vocal folds oscillate, the superficial tissue of the vocal fold is displaced in a wave-like fashion, creating the so called “mucosal wave”. Because the mucosal wave is dependent on vocal fold structure, physical alterations of that structure cause mucosal wave abnormalities. Visualization and quantification of mucosal wave properties have become useful parameters in diagnosing and managing vocal fold pathology. Mucosal wave measurement provides information about vocal fold characteristics that cannot be determined with other assessment techniques. Here, we discuss the benefits, disadvantages, and clinical applicability of the different mucosal wave measurement techniques, such as electroglottography (EGG), photoglottography (PGG), and ultrasound and visualization techniques that include videokymography (VKG), stroboscopy, and high-speed digital imaging (HSDI). The various techniques and their specific uses are reviewed with the intention of helping researchers and clinicians choose a method for a given situation and understand its limitations as well as its potential applications. Recent applications of these techniques for quantitative assessment demonstrate that additional research must be conducted to realize the full potential of these tools. Evaluations of existing research and recommendations for future research are given to promote both the quantitative study of the mucosal wave through accurate and standardized measurement of mucosal wave parameters and the development of reliable methods with which physicians can diagnose vocal disorders. PMID:20471798

  13. Vagal nerve and the gastric mucosal defense.

    PubMed

    Mózsik, G; Karádi, O; Király, A; Matus, Z; Sütö, G; Tóth, G; Vincze, A

    1993-01-01

    An essential role for an intact vagal nerve has been proven in the development of gastric mucosal cyto- and general protection. On the other hand, chemically-induced (ethanol, HCl, indomethacin) gastric mucosal damage is enhanced after acute surgical vagotomy. The aims of this paper were to study the possible mechanisms of the vagal nerve in the development of gastric mucosal defense. The following questions were addressed: 1) effect of surgical vagotomy on the development of ethanol- (ETOH), HCl-, and indomethacin (IND)-induced gastric mucosal damage: 2) changes in the gastric mucosal defense by scavengers, prostacyclin and other compounds (small doses of atropine and cimetidine; 3) changes in the gastric mucosal vascular permeability due to chemicals; 4) effect of indomethacin in the ETOH and HCl models with and without surgical vagotomy; 5) changes in the gastric mucosal content of prostacyclin and PGE2 in the ETOH and HCl models after surgical vagotomy; and 6) changes in the role of SH-groups in gastric mucosal defense after surgical vagotomy.(ABSTRACT TRUNCATED AT 250 WORDS)

  14. Biology and Mucosal Immunity to Myxozoans

    PubMed Central

    Gómez, Daniela; Bartholomew, Jerri; Sunyer, J. Oriol

    2014-01-01

    Myxozoans are among the most abundant parasites in nature. Their life cycles involve two hosts: an invertebrate, usually an annelid, and a vertebrate, usually a fish. They affect fish species in their natural habitats but also constitute a menace for fish aquaculture. Using different strategies they are able to parasitize and cause damage in multiple organs, including mucosal tissues, which they use also as portals of entry. In fish, the main mucosal sites include the intestine, skin and gills. Recently the finding of a specific mucosal immunoglobulin in teleost (IgT), analogous to mammalian IgA, and the capacity of fish to develop a specific mucosal immune response against different pathogens, has highlighted the importance of studying immune responses at mucosal sites. In this review, we describe the major biological characteristics of myxozoan parasites and present the data available regarding immune responses for species that infect mucosal sites. As models for mucosal immunity we review the responses to Enteromyxum spp. and Ceratomyxa shasta, both of which parasitize the intestine. The immune response at the skin and gills is also described, as these mucosal tissues are used by myxozoans as attaching surfaces and portal of entry, and some species also parasitize these sites. Finally, the development of immunoprophylactic strategies is discussed. PMID:23994774

  15. Mucosal vaccines: a paradigm shift in the development of mucosal adjuvants and delivery vehicles.

    PubMed

    Srivastava, Atul; Gowda, Devegowda Vishakante; Madhunapantula, SubbaRao V; Shinde, Chetan G; Iyer, Meenakshi

    2015-04-01

    Mucosal immune responses are the first-line defensive mechanisms against a variety of infections. Therefore, immunizations of mucosal surfaces from which majority of infectious agents make their entry, helps to protect the body against infections. Hence, vaccinization of mucosal surfaces by using mucosal vaccines provides the basis for generating protective immunity both in the mucosal and systemic immune compartments. Mucosal vaccines offer several advantages over parenteral immunization. For example, (i) ease of administration; (ii) non-invasiveness; (iii) high-patient compliance; and (iv) suitability for mass vaccination. Despite these benefits, to date, only very few mucosal vaccines have been developed using whole microorganisms and approved for use in humans. This is due to various challenges associated with the development of an effective mucosal vaccine that can work against a variety of infections, and various problems concerned with the safe delivery of developed vaccine. For instance, protein antigen alone is not just sufficient enough for the optimal delivery of antigen(s) mucosally. Hence, efforts have been made to develop better prophylactic and therapeutic vaccines for improved mucosal Th1 and Th2 immune responses using an efficient and safe immunostimulatory molecule and novel delivery carriers. Therefore, in this review, we have made an attempt to cover the recent advancements in the development of adjuvants and delivery carriers for safe and effective mucosal vaccine production.

  16. Mucosal vaccines: novel strategies and applications for the control of pathogens and tumors at mucosal sites.

    PubMed

    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.

  17. Distinct defensin profiles in Neisseria gonorrhoeae and Chlamydia trachomatis urethritis reveal novel epithelial cell-neutrophil interactions.

    PubMed

    Porter, Edith; Yang, Huixia; Yavagal, Sujata; Preza, Gloria C; Murillo, Omar; Lima, Heriberto; Greene, Sheila; Mahoozi, Laily; Klein-Patel, Marcia; Diamond, Gill; Gulati, Sunita; Ganz, Tomas; Rice, Peter A; Quayle, Alison J

    2005-08-01

    Defensins are key participants in mucosal innate defense. The varied antimicrobial activity and differential distribution of defensins at mucosal sites indicate that peptide repertoires are tailored to site-specific innate defense requirements. Nonetheless, few studies have investigated changes in peptide profiles and function after in vivo pathogen challenge. Here, we determined defensin profiles in urethral secretions of healthy men and men with Chlamydia trachomatis- and Neisseria gonorrhoeae-mediated urethritis by immunoblotting for the epithelial defensins HBD1, HBD2, and HD5 and the neutrophil defensins HNP1 to -3 (HNP1-3). HBD1 was not detectable in secretions, and HBD2 was only induced in a small proportion of the urethritis patients; however, HD5 and HNP1-3 were increased in C. trachomatis infection and significantly elevated in N. gonorrhoeae infection. When HNP1-3 levels were low, HD5 appeared mostly as the propeptide; however, when HNP1-3 levels were >10 microg/ml, HD5 was proteolytically processed, suggesting neutrophil proteases might contribute to HD5 processing. HD5 and HNP1-3 were bactericidal against C. trachomatis and N. gonorrhoeae, but HD5 activity was dependent upon N-terminal processing of the peptide. In vitro proteolysis of proHD5 by neutrophil proteases and analysis of urethral secretions by surface-enhanced laser desorption ionization substantiated that neutrophils contribute the key convertases for proHD5 in the urethra during these infections. This contrasts with the small intestine, where Paneth cells secrete both proHD5 and its processing enzyme, trypsin. In conclusion, we describe a unique defensin expression repertoire in response to inflammatory sexually transmitted infections and a novel host defense mechanism wherein epithelial cells collaborate with neutrophils to establish an antimicrobial barrier during infection. PMID:16040996

  18. Impact of Helicobacter pylori on the healing process of the gastric barrier

    PubMed Central

    Mnich, Eliza; Kowalewicz-Kulbat, Magdalena; Sicińska, Paulina; Hinc, Krzysztof; Obuchowski, Michał; Gajewski, Adrian; Moran, Anthony P; Chmiela, Magdalena

    2016-01-01

    AIM To determine the impact of selected well defined Helicobacter pylori (H. pylori) antigens on gastric barrier cell turnover. METHODS In this study, using two cellular models of gastric epithelial cells and fibroblasts, we have focused on exploring the effects of well defined H. pylori soluble components such as glycine acid extract antigenic complex (GE), subunit A of urease (UreA), cytotoxin associated gene A protein (CagA) and lipopolysaccharide (LPS) on cell turnover by comparing the wound healing capacity of the cells in terms of their proliferative and metabolic activity as well as cell cycle distribution. Toxic effects of H. pylori components have been assessed in an association with damage to cell nuclei and inhibition of signal transducer and activator of transcription 3 (STAT3) phosphorylation. RESULTS We showed that H. pylori GE, CagA and UreA promoted regeneration of epithelial cells and fibroblasts, which is necessary for effective tissue healing. However, in vivo increased proliferative activity of these cells may constitute an increased risk of gastric neoplasia. In contrast, H. pylori LPS showed a dose-dependent influence on the process of wound healing. At a low concentration (1 ng/mL) H. pylori LPS accelerated of healing epithelial cells, which was linked to significantly enhanced cell proliferation and MTT reduction as well as lack of alterations in cell cycle and downregulation of epidermal growth factor (EGF) production as well as cell nuclei destruction. By comparison, H. pylori LPS at a high concentration (25 ng/mL) inhibited the process of wound repair, which was related to diminished proliferative activity of the cells, cell cycle arrest, destruction of cell nuclei and downregulation of the EGF/STAT3 signalling pathway. CONCLUSION In vivo H. pylori LPS driven effects might lead to the maintenance of chronic inflammatory response and pathological disorders on the level of the gastric mucosal barrier.

  19. Impact of Helicobacter pylori on the healing process of the gastric barrier

    PubMed Central

    Mnich, Eliza; Kowalewicz-Kulbat, Magdalena; Sicińska, Paulina; Hinc, Krzysztof; Obuchowski, Michał; Gajewski, Adrian; Moran, Anthony P; Chmiela, Magdalena

    2016-01-01

    AIM To determine the impact of selected well defined Helicobacter pylori (H. pylori) antigens on gastric barrier cell turnover. METHODS In this study, using two cellular models of gastric epithelial cells and fibroblasts, we have focused on exploring the effects of well defined H. pylori soluble components such as glycine acid extract antigenic complex (GE), subunit A of urease (UreA), cytotoxin associated gene A protein (CagA) and lipopolysaccharide (LPS) on cell turnover by comparing the wound healing capacity of the cells in terms of their proliferative and metabolic activity as well as cell cycle distribution. Toxic effects of H. pylori components have been assessed in an association with damage to cell nuclei and inhibition of signal transducer and activator of transcription 3 (STAT3) phosphorylation. RESULTS We showed that H. pylori GE, CagA and UreA promoted regeneration of epithelial cells and fibroblasts, which is necessary for effective tissue healing. However, in vivo increased proliferative activity of these cells may constitute an increased risk of gastric neoplasia. In contrast, H. pylori LPS showed a dose-dependent influence on the process of wound healing. At a low concentration (1 ng/mL) H. pylori LPS accelerated of healing epithelial cells, which was linked to significantly enhanced cell proliferation and MTT reduction as well as lack of alterations in cell cycle and downregulation of epidermal growth factor (EGF) production as well as cell nuclei destruction. By comparison, H. pylori LPS at a high concentration (25 ng/mL) inhibited the process of wound repair, which was related to diminished proliferative activity of the cells, cell cycle arrest, destruction of cell nuclei and downregulation of the EGF/STAT3 signalling pathway. CONCLUSION In vivo H. pylori LPS driven effects might lead to the maintenance of chronic inflammatory response and pathological disorders on the level of the gastric mucosal barrier. PMID:27672275

  20. 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.

  1. Balancing acts: the role of TGF-β in the mucosal immune system

    PubMed Central

    Konkel, Joanne E.; Chen, Wanjun

    2011-01-01

    The gastrointestinal mucosal immune system faces unique challenges in dealing not only with fed antigens but also both commensal and pathogenic bacteria. It is tasked with digesting, transporting and using nutritional antigens whilst protecting the host from pathogenic organisms. As such, mechanisms that mediate effective immunity and immune tolerance are active within the gut environment. To accomplish this, the mucosal immune system has evolved sophisticated mechanisms that safeguard the integrity of the mucosal barrier. Transforming growth factor-β (TGF-β) emerges as a key mediator, balancing the tolerogenic and immunogenic forces at play in the gut. In this review we discuss the role of TGF-β in the generation and functioning of gut lymphocyte populations. We highlight recent findings, summarize controversies, outline remaining questions and provide our personal perspectives. PMID:21890412

  2. A single mild episode of subacute ruminal acidosis does not affect ruminal barrier function in the short term.

    PubMed

    Penner, G B; Oba, M; Gäbel, G; Aschenbach, J R

    2010-10-01

    Twenty-four German Merino sheep (72.3±10.1 kg of body weight) were fed an all-hay diet and assigned to either the subacute ruminal acidosis (SARA) treatment (n=17) or sham treatment (n=7). The SARA sheep were orally dosed with a 2.2 M glucose solution to supply 5 g of glucose/kg of body weight, whereas sham sheep received an equal volume of water. Ruminal pH was measured for 48 h before and 3 h after the oral dose. Sheep were then killed and ruminal epithelia from the ventral sac were mounted in Ussing chambers. The serosal-to-mucosal flux rate of partially (3)H-labeled mannitol (J(mannitol-SM)), an indicator of barrier function, was measured while epithelia were exposed to 3 sequential in vitro measurement periods lasting 1 h each. The measurement periods consisted of baseline, challenge, and recovery periods and were interspersed by 30-min periods for treatment equilibration. Baseline conditions were pH 6.1 (mucosal solution) and pH 7.4 (serosal solution) with a bilateral osmolarity of 293 mOsm/L. During the challenge period, the mucosal side of the epithelia was exposed to either an acidotic challenge (pH 5.2, osmolarity 293 mOsm/L) or an osmotic challenge (pH 6.1, osmolarity 450 mOsm/L); a third group served as control (pH 6.1, osmolarity 293 mOsm/L). The mucosal buffer solution was replaced for the recovery period. In vivo, sheep on the SARA treatment had lower mean (5.77 vs. 6.67) and nadir (5.48 vs. 6.47) ruminal pH for the 3h following the oral drench compared with sham sheep, indicating the successful induction of SARA with the oral glucose dose. Despite the marked reduction in pH in vivo, induction of SARA had no detectable effects on the baseline measurements of J(mannitol-SM), tissue conductance (G(t)), and short-circuit current (I(sc)) in vitro. However, reducing mucosal pH to 5.2 in vitro had negative effects on epithelial barrier function in the recovery period, including increased J(mannitol-SM), increased G(t), and decreased I(sc). The osmotic

  3. Generation of Mouse Small Intestinal Epithelial Cell Lines That Allow the Analysis of Specific Innate Immune Functions

    PubMed Central

    Schwerk, Johannes; Köster, Mario; Hauser, Hansjörg; Rohde, Manfred; Fulde, Marcus; Hornef, Mathias W.; May, Tobias

    2013-01-01

    Cell lines derived from the small intestine that reflect authentic properties of the originating intestinal epithelium are of high value for studies on mucosal immunology and host microbial homeostasis. A novel immortalization procedure was applied to generate continuously proliferating cell lines from murine E19 embryonic small intestinal tissue. The obtained cell lines form a tight and polarized epithelial cell layer, display characteristic tight junction, microvilli and surface protein expression and generate increasing transepithelial electrical resistance during in vitro culture. Significant up-regulation of Cxcl2 and Cxcl5 chemokine expression upon exposure to defined microbial innate immune stimuli and endogenous cytokines is observed. Cell lines were also generated from a transgenic interferon reporter (Mx2-Luciferase) mouse, allowing reporter technology-based quantification of the cellular response to type I and III interferon. Thus, the newly created cell lines mimic properties of the natural epithelium and can be used for diverse studies including testing of the absorption of drug candidates. The reproducibility of the method to create such cell lines from wild type and transgenic mice provides a new tool to study molecular and cellular processes of the epithelial barrier. PMID:23940817

  4. Mucosal acid causes gastric mucosal microcirculatory disturbance in nonsteroidal anti-inflammatory drug-treated rats.

    PubMed

    Funatsu, Toshiyuki; Chono, Koji; Hirata, Takuya; Keto, Yoshihiro; Kimoto, Aishi; Sasamata, Masao

    2007-01-01

    The mechanism by which nonsteroidal anti-inflammatory drugs (NSAIDs) suppress gastric mucosal blood flow is not fully understood, although the depletion of mucosal prostaglandin E2 has been proposed as one possible explanation. We investigated the role of gastric acid on gastric mucosal blood flow in NSAID-treated rats. A rat stomach was mounted in an ex vivo chamber, and gastric mucosal blood flow was measured sequentially in a 5-mm2 area of the gastric corpus using a scanning laser Doppler perfusion image system. Results showed that diclofenac (5 mg/kg s.c.) and indomethacin (10 mg/kg s.c.) did not affect gastric mucosal blood flow, although both strongly decreased mucosal prostaglandin E2 when saline was instilled into the gastric chamber. On replacement of the saline in the chamber with 100 mM hydrochloric acid, these drugs caused a decrease in gastric mucosal blood flow levels within 30 min. The specific cyclooxygenase (COX)-2 inhibitors celecoxib (50 mg/kg s.c.) and rofecoxib (25 mg/kg s.c.) did not affect mucosal prostaglandin E2 level, nor did they decrease gastric mucosal blood flow, even when hydrochloric acid was added to the chamber. Furthermore, measurement of vasoconstrictive factors present in the mucosa showed that endothelin-1 levels increased after administration of diclofenac s.c. in the presence of intragastric hydrochloric acid. This indicates that the presence of mucosal hydrochloric acid plays an important role in the NSAID-induced decrease in gastric mucosal blood flow, while the COX-1-derived basal prostaglandin E2, which is unlikely to control gastric mucosal blood flow itself, protects microcirculatory systems from mucosal hydrochloric acid.

  5. Treatment of oral mucositis due to chemotherapy

    PubMed Central

    Bagán-Sebastián, José V

    2016-01-01

    Introduction The management of oral mucositis is a challenge, due to its complex biological nature. Over the last 10 years, different strategies have been developed for the management of oral mucositis caused by chemotherapy in cancer patients. Material and Methods An exhaustive search was made of the PubMed-Medline, Cochrane Library and Scopus databases, crossing the key words “oral mucositis”, “prevention” and “treatment” with the terms “chemotherapy” and “radiotherapy” by means of the boolean operators “AND” and “NOT”. A total of 268 articles were obtained, of which 96 met the inclusion criteria. Results Several interventions for the prevention of oral mucositis, such as oral hygiene protocols, amifostine, benzidamine, calcium phosphate, cryotherapy and iseganan, among others, were found to yield only limited benefits. Other studies have reported a decrease in the appearance and severity of mucositis with the use of cytoprotectors (sucralfate, oral glutamine, hyaluronic acid), growth factors, topical polyvinylpyrrolidone, and low power laser irradiation. Conclusions Very few interventions of confirmed efficacy are available for the management of oral mucositis due to chemotherapy. However, according to the reviewed literature, the use of palifermin, cryotherapy and low power laser offers benefits, reducing the incidence and severity of oral mucositis – though further studies are needed to confirm the results obtained. Key words:Chemotherapy-Induced Oral Mucositis Treatment. PMID:27034762

  6. Chemotherapy-induced and/or radiation therapy-induced oral mucositis--complicating the treatment of cancer.

    PubMed

    Naidu, Maddireddy Umameshwar Rao; Ramana, Gogula Venkat; Rani, Pingali Usha; Mohan, Iyyapu Krishna; Suman, Avula; Roy, Priyadarshni

    2004-01-01

    The term mucositis is coined to describe the adverse effects of radiation and chemotherapy treatments. Mucositis is one of the most common adverse reactions encountered in radiation therapy for head and neck cancers, as well as in chemotherapy, in particular with drugs affecting DNA synthesis (S-phase-specific agents such as fluorouracil, methotrexate, and cytarabine). Mucositis may limit the patient's ability to tolerate chemotherapy or radiation therapy, and nutritional status is compromised. It may drastically affect cancer treatment as well as the patient's quality of life. The incidence and severity of mucositis will vary from patient to patient. It will also vary from treatment to treatment. It is estimated that there is 40% incidence of mucositis in patients treated with standard chemotherapy and this will not only increase with the number of treatment cycles but also with previous episodes. Similarly, patients who undergo bone marrow transplantation and who receive high doses of chemotherapy have a 76% chance of getting mucositis. Patients receiving radiation, in particular to head and neck cancers, have a 30% to 60% chance. The exact pathophysiology of development is not known, but it is thought to be divided into direct and indirect mucositis. Chemotherapy and/or radiation therapy will interfere with the normal turnover of epithelial, cells leading to mucosal injury; subsequently, it can also occur due to indirect invasion of Gram-negative bacteria and fungal species because most of the cancer drugs will cause changes in blood counts. With the advancement in cytology, a more precise mechanism has been established. With this understanding, we can select and target particular mediators responsible for the mucositis. Risk factors such as age, nutritional status, type of malignancy, and oral care during treatment will play important roles in the development of mucositis. Many treatment options are available to prevent and treat this condition, but none of

  7. Mucosal adjuvants and delivery systems for protein-, DNA- and RNA-based vaccines.

    PubMed

    Vajdy, Michael; Srivastava, Indresh; Polo, John; Donnelly, John; O'Hagan, Derek; Singh, Manmohan

    2004-12-01

    Almost all vaccinations today are delivered through parenteral routes. Mucosal vaccination offers several benefits over parenteral routes of vaccination, including ease of administration, the possibility of self-administration, elimination of the chance of injection with infected needles, and induction of mucosal as well as systemic immunity. However, mucosal vaccines have to overcome several formidable barriers in the form of significant dilution and dispersion; competition with a myriad of various live replicating bacteria, viruses, inert food and dust particles; enzymatic degradation; and low pH before reaching the target immune cells. It has long been known that vaccination through mucosal membranes requires potent adjuvants to enhance immunogenicity, as well as delivery systems to decrease the rate of dilution and degradation and to target the vaccine to the site of immune function. This review is a summary of current approaches to mucosal vaccination, and it primarily focuses on adjuvants as immunopotentiators and vaccine delivery systems for mucosal vaccines based on protein, DNA or RNA. In this context, we define adjuvants as protein or oligonucleotides with immunopotentiating properties co-administered with pathogen-derived antigens, and vaccine delivery systems as chemical formulations that are more inert and have less immunomodulatory effects than adjuvants, and that protect and deliver the vaccine through the site of administration. Although vaccines can be quite diverse in their composition, including inactivated virus, virus-like particles and inactivated bacteria (which are inert), protein-like vaccines, and non-replicating viral vectors such as poxvirus and adenovirus (which can serve as DNA delivery systems), this review will focus primarily on recombinant protein antigens, plasmid DNA, and alphavirus-based replicon RNA vaccines and delivery systems. This review is not an exhaustive list of all available protein, DNA and RNA vaccines, with related

  8. CXCR4 Antagonist AMD3100 Modulates Claudin Expression and Intestinal Barrier Function in Experimental Colitis

    PubMed Central

    Xia, Xian-Ming; Wang, Fang-Yu; Zhou, Ju; Hu, Kai-Feng; Li, Su-Wen; Zou, Bing-Bing

    2011-01-01

    Ulcerative colitis is a gastrointestinal disorder characterized by local inflammation and impaired epithelial barrier. Previous studies demonstrated that CXC chemokine receptor 4 (CXCR4) antagonists could reduce colonic inflammation and mucosal damage in dextran sulfate sodium (DSS)-induced colitis. Whether CXCR4 antagonist has action on intestinal barrier and the possible mechanism, is largely undefined. In the present study, the experimental colitis was induced by administration of 5% DSS for 7 days, and CXCR4 antagonist AMD3100 was administered intraperitoneally once daily during the study period. For in vitro study, HT-29/B6 colonic cells were treated with cytokines or AMD3100 for 24 h until assay. DSS-induced colitis was characterized by morphologic changes in mice. In AMD3100-treated mice, epithelial destruction, inflammatory infiltration, and submucosal edema were markedly reduced, and the disease activity index was also significantly decreased. Increased intestinal permeability in DSS-induced colitis was also significantly reduced by AMD3100. The expressions of colonic claudin-1, claudin-3, claudin-5, claudin-7 and claudin-8 were markedly decreased after DSS administration, whereas colonic claudin-2 expression was significantly decreased. Treatment with AMD3100 prevented all these changes. However, AMD3100 had no influence on claudin-3, claudin-5, claudin-7 and claudin-8 expression in HT-29/B6 cells. Cytokines as TNF-α, IL-6, and IFN-γ increased apoptosis and monolayer permeability, inhibited the wound-healing and the claudin-3, claudin-7 and claudin-8 expression in HT-29/B6 cells. We suggest that AMD3100 acted on colonic claudin expression and intestinal barrier function, at least partly, in a cytokine-dependent pathway. PMID:22073304

  9. Duodenal mucosal protection by bicarbonate secretion and its mechanisms.

    PubMed

    Konturek, S J; Konturek, P C; Pawlik, T; Sliwowski, Z; Ochmański, W; Hahn, E G

    2004-07-01

    Proximal portion of duodenum is exposed to intermittent pulses of gastric H(+) discharged by the stomach. This review summarizes the mechanisms of duodenal mucosal integrity, mainly the role of mucus-alkaline secretion and the mucous barrier protecting surface epithelium against gastric H(+). The mucous barrier protects the leaky duodenal epithelium against each pulse of gastric H(+), which penetrates this barrier and diffuses into duodenocytes, but fails to damage them due to; a) an enhanced expression of cyclooxygenase-1 (COX-1), with release of protective prostaglandins (PG) and of nitric oxide (NO) synthase (NOS) with, however, production of NO, stimulating duodenal HCO(3)(-) secretion and b) the release of several neurotransmitters also stimulating HCO(3)(-) secretion such as vasoactive intestinal peptide (VIP), pituitary adenylate-cyclase activating polypeptide (PACAP), acetylcholine, melatonin, leptin and ghrelin released by enteric nerves and mucosal cells. At the apical duodenocyte membrane at least two HCO(3)(-)/Cl(-) anion exchangers operate in response to luminal H(+) to provide adequate extrusion of HCO(3)(-) into duodenal lumen. In the basolateral portion of duodenocyte membrane, both non-electrogenic (NBC) and electrogenic (NBC(n)) Na(+) HCO(3)(-) cotransporters are activated by the exposure to duodenal acidification, causing inward movement of HCO(3)(-) from extracellular fluid to duodenocytes. There are also at least three Na(+)/H(+) (NHE1-3) amiloride-sensitive exchangers, eliminating H(+)which diffused into these cells. The Helicobacter pylori (Hp) infection and gastric metaplasia in the duodenum with bacterium inoculating metaplastic mucosa and inhibiting HCO(3)(-) secretion by its endogenous inhibitor, asymmetric dimethyl arginine (ADMA), may result in duodenal ulcerogenesis. PMID:15608357

  10. Brain neuropeptides in gastric mucosal protection.

    PubMed

    Gyires, Klára; Zádori, Zoltán S

    2014-12-01

    The centrally induced gastroprotective effect of neuropeptides has been intensively studied. Besides many similarities, however, differences can also be observed in their gastroprotective actions. The gastroprotective dose-response curve proved to be either sigmoid, or bell-shaped. Additional gastrointestinal effects of neuropeptides can contribute to their mucosal protective effect. Part of the neuropeptides induces gastroprotection by peripheral administration as well. Besides vagal nerve the sympathetic nervous system may also be involved in conveying the central effect to the periphery. Better understanding of the complex mechanism of the maintenance of gastric mucosal integrity may result in the development of new strategy to enhance gastric mucosal resistance against injury.

  11. 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.

  12. The matricellular protein CCN1 promotes mucosal healing in murine colitis through IL-6.

    PubMed

    Choi, J S; Kim, K-H; Lau, L F

    2015-11-01

    The matricellular protein CCN1 (CYR61) is known to function in wound healing and is upregulated in colons of patients with Crohn's disease and ulcerative colitis, yet its specific role in colitis is unknown. Here we have used Ccn1(dm/dm) knockin mice expressing a CCN1 mutant unable to bind integrins α6β1 and αMβ2 as a model to probe CCN1 function in dextran sodium sulfate (DSS)-induced colitis. Ccn1(dm/dm) mice exhibited high mortality, impaired mucosal healing, and diminished interleukin-6 (IL-6) expression during the repair phase of DSS-induced colitis compared with wild-type mice, despite having comparable severity of initial inflammation and tissue injury. CCN1-induced IL-6 expression in macrophages through integrin αMβ2 and in fibroblasts through α6β1, and IL-6 promoted intestinal epithelial cell (IEC) proliferation. Administration of purified CCN1 protein fully rescued Ccn1(dm/dm) mice from DSS-induced mortality, restored IEC proliferation and enhanced mucosal healing, whereas delivery of IL-6 partially rectified these defects. CCN1 therapy accelerated mucosal healing and recovery from DSS-induced colitis even in wild-type mice. These findings reveal a critical role for CCN1 in restoring mucosal homeostasis after intestinal injury in part through integrin-mediated induction of IL-6 expression, and suggest a therapeutic potential for activating the CCN1/IL-6 axis for treating inflammatory bowel disease.

  13. Microbiota and Mucosal Immunity in Amphibians

    PubMed Central

    Colombo, Bruno M.; Scalvenzi, Thibault; Benlamara, Sarah; Pollet, Nicolas

    2015-01-01

    We know that animals live in a world dominated by bacteria. In the last 20 years, we have learned that microbes are essential regulators of mucosal immunity. Bacteria, archeas, and viruses influence different aspects of mucosal development and function. Yet, the literature mainly covers findings obtained in mammals. In this review, we focus on two major themes that emerge from the comparative analysis of mammals and amphibians. These themes concern: (i) the structure and functions of lymphoid organs and immune cells in amphibians, with a focus on the gut mucosal immune system; and (ii) the characteristics of the amphibian microbiota and its influence on mucosal immunity. Lastly, we propose to use Xenopus tadpoles as an alternative small-animal model to improve the fundamental knowledge on immunological functions of gut microbiota. PMID:25821449

  14. Targeting Mucosal Healing in Crohn's Disease

    PubMed Central

    Kakkar, Aarti; Wasan, Sharmeel K.

    2011-01-01

    The goal of medical treatment for Crohn's disease includes improving patients' quality of life while reducing the need for hospitalization and surgery. The current medical armamentarium includes 5-aminosalicylates, corticosteroids, immunomodulators, and biologic agents. In the past, response to treatment was measured by clinical improvement in symptoms; however, with the advent of disease-modifying medications, mucosal healing has emerged as an increasingly important goal of therapy. Mucosal healing, or endoscopic remission, is associated with increased rates of clinical remission, fewer hospitalizations, and fewer abdominal surgeries. Both the immunomodulator and biologic classes of medications are effective at inducing mucosal healing. Despite several limitations, mucosal healing has become a desirable and valid measure of disease activity. PMID:21869869

  15. Microbiota and mucosal immunity in amphibians.

    PubMed

    Colombo, Bruno M; Scalvenzi, Thibault; Benlamara, Sarah; Pollet, Nicolas

    2015-01-01

    We know that animals live in a world dominated by bacteria. In the last 20 years, we have learned that microbes are essential regulators of mucosal immunity. Bacteria, archeas, and viruses influence different aspects of mucosal development and function. Yet, the literature mainly covers findings obtained in mammals. In this review, we focus on two major themes that emerge from the comparative analysis of mammals and amphibians. These themes concern: (i) the structure and functions of lymphoid organs and immune cells in amphibians, with a focus on the gut mucosal immune system; and (ii) the characteristics of the amphibian microbiota and its influence on mucosal immunity. Lastly, we propose to use Xenopus tadpoles as an alternative small-animal model to improve the fundamental knowledge on immunological functions of gut microbiota. PMID:25821449

  16. Transgenic killer commensal bacteria as mucosal protectants.

    PubMed

    Polonelli, L

    2001-05-01

    As first line of defense against the majority of infections and primary site for their transmission, mucosal surfaces of the oral cavity and genitourinary, gastrointestinal, and respiratory tracts represent the most suitable sites to deliver protective agents for the prevention of infectious diseases. Mucosal protection is important not only for life threatening diseases but also for opportunistic infections which currently represent a serious burden in terms of morbidity, mortality, and cost of cures. Candida albicans is among the most prevalent causes of mucosal infections not only in immuno-compromised patients, such as HIV-infected subjects who are frequently affected by oral and esophageal candidiasis, but also in otherwise healthy individuals, as in the case of acute vaginitis. Unfortunately, current strategies for mucosal protection against candidiasis are severely limited by the lack of effective vaccines and the relative paucity and toxicity of commercially available antifungal drugs.

  17. 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

  18. Colonic ornithine decarboxylase in inflammatory bowel disease: ileorectal activity gradient, guanosine triphosphate stimulation, and association with epithelial regeneration but not the degree of inflammation and clinical features.

    PubMed

    Allgayer, Hubert; Roisch, Ulla; Zehnter, Elmar; Ziegenhagen, Dieter J; Dienes, Hans P; Kruis, Wolfgang

    2007-01-01

    The role of colonic mucosal ornithine decarboxylase (ODC) in inflammatory bowel disease (IBD) remains controversial. This study assessed mucosal ODC activity in IBD patients segment by segment with regard to patient characteristics, disease activity/duration, medication, degree of mucosal inflammation, and presence/absence of epithelial regeneration and guanosine triphosphate (GTP) stimulation. Mucosal ODC activity was determined in biopsy specimens from the terminal ileum, cecum/ascending, transverse, and descending colon, and the sigmoid/rectum of 35 patients with IBD (18 with Crohn's disease, 17 with ulcerative colitis) and 29 controls, using the amount of 14CO2 liberated from (carboxyl-14C)ornithine hydrochloride. GTP-stimulatable activity was expressed as the ratio of ODC activity in the presence and absence of GTP (70 micromol/L). Mucosal inflammation was assessed endoscopically/microscopically with previously described criteria. Presence/absence of mucosal regeneration also was determined by predefined criteria. Mucosal ODC-activity did not significantly differ in IBD patients and controls. There was a 4.4-fold activity gradient from the ileum to the rectum. Mucosal ODC activity was significantly higher in areas with epithelial regeneration compared to those without regeneration, and was stimulated by GTP by a factor of 1.42 in Crohn's disease and 1.19 in ulcerative colitis patients compared to controls (p < 0.004). On the other hand, there was no significant association/relationship of mucosal ODC activity with disease activity/duration and the endoscopic/histologic degree of mucosal inflammation. The observation of unchanged mucosal ODC activity in patients with IBD and the absence of a significant relationship with clinical and endoscopic/histologic disease characteristics speaks against a major role of ODC in IBD as a major disease marker. The role of the ileorectal gradient, the enhanced activity in areas with epithelial regeneration, and the GTP

  19. A Novel Peptide for Simultaneously Enhanced Treatment of Head and Neck Cancer and Mitigation of Oral Mucositis

    PubMed Central

    Chen, Peili; Mancini, Maria; Sonis, Stephen T.; Fernandez-Martinez, Juan; Liu, Jing; Cohen, Ezra E. W.; Toback, F. Gary

    2016-01-01

    We have characterized a novel 21 amino acid-peptide derived from Antrum Mucosal Protein (AMP)-18 that mediates growth promotion of cultured normal epithelial cells and mitigates radiation-induced oral mucositis in animal models, while suppressing in vitro function of cancer cells. The objective of this study was to evaluate these dual potential therapeutic effects of AMP peptide in a clinically relevant animal model of head and neck cancer (HNC) by simultaneously assessing its effect on tumor growth and radiation-induced oral mucositis in an orthotopic model of HNC. Bioluminescent SCC-25 HNC cells were injected into the anterior tongue and tumors that formed were then subjected to focal radiation treatment. Tumor size was assessed using an in vivo imaging system, and the extent of oral mucositis was compared between animals treated with AMP peptide or vehicle (controls). Synergism between AMP peptide and radiation therapy was suggested by the finding that tumors in the AMP peptide/radiation therapy cohort demonstrated inhibited growth vs. radiation therapy-only treated tumors, while AMP peptide-treatment delayed the onset and reduced the severity of radiation therapy-induced oral mucositis. A differential effect on apoptosis appears to be one mechanism by which AMP-18 can stimulate growth and repair of injured mucosal epithelial cells while inhibiting proliferation of HNC cells. RNA microarray analysis identified pathways that are differentially targeted by AMP-18 in HNC vs. nontransformed cells. These observations confirm the notion that normal cells and tumor cells may respond differently to common biological stimuli, and that leveraging this finding in the case of AMP-18 may provide a clinically relevant opportunity. PMID:27049860

  20. 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

  1. Occurrence and resolution of focal epithelial hyperplasia in two siblings with leukocyte adhesion deficiency.

    PubMed

    Mealey, B L; Hallmon, W W; Waldrop, T C

    1993-02-01

    Focal epithelial hyperplasia is an uncommon disorder characterized by formation of multiple asymptomatic oral mucosal papular and/or nodular lesions. This article relates the occurrence and clinical course of FEH in 2 adolescent siblings with leukocyte adhesion deficiency. Histological findings are described and insights into potential causes are discussed.

  2. Epithelial Anion Transport as Modulator of Chemokine Signaling.

    PubMed

    Schnúr, Andrea; Hegyi, Péter; Rousseau, Simon; Lukacs, Gergely L; Veit, Guido

    2016-01-01

    The pivotal role of epithelial cells is to secrete and absorb ions and water in order to allow the formation of a luminal fluid compartment that is fundamental for the epithelial function as a barrier against environmental factors. Importantly, epithelial cells also take part in the innate immune system. As a first line of defense they detect pathogens and react by secreting and responding to chemokines and cytokines, thus aggravating immune responses or resolving inflammatory states. Loss of epithelial anion transport is well documented in a variety of diseases including cystic fibrosis, chronic obstructive pulmonary disease, asthma, pancreatitis, and cholestatic liver disease. Here we review the effect of aberrant anion secretion with focus on the release of inflammatory mediators by epithelial cells and discuss putative mechanisms linking these transport defects to the augmented epithelial release of chemokines and cytokines. These mechanisms may contribute to the excessive and persistent inflammation in many respiratory and gastrointestinal diseases. PMID:27382190

  3. Epithelial Anion Transport as Modulator of Chemokine Signaling

    PubMed Central

    Schnúr, Andrea; Hegyi, Péter; Rousseau, Simon; Lukacs, Gergely L.; Veit, Guido

    2016-01-01

    The pivotal role of epithelial cells is to secrete and absorb ions and water in order to allow the formation of a luminal fluid compartment that is fundamental for the epithelial function as a barrier against environmental factors. Importantly, epithelial cells also take part in the innate immune system. As a first line of defense they detect pathogens and react by secreting and responding to chemokines and cytokines, thus aggravating immune responses or resolving inflammatory states. Loss of epithelial anion transport is well documented in a variety of diseases including cystic fibrosis, chronic obstructive pulmonary disease, asthma, pancreatitis, and cholestatic liver disease. Here we review the effect of aberrant anion secretion with focus on the release of inflammatory mediators by epithelial cells and discuss putative mechanisms linking these transport defects to the augmented epithelial release of chemokines and cytokines. These mechanisms may contribute to the excessive and persistent inflammation in many respiratory and gastrointestinal diseases. PMID:27382190

  4. Mucosal toxicity studies of a gel formulation of native pokeweed antiviral protein.

    PubMed

    D'Cruz, Osmond J; Waurzyniak, Barbara; Uckun, Fatih M

    2004-01-01

    Pokeweed antiviral protein (PAP), a 29-kDa plant-derived protein isolated from Phytolacca americana, is a promising nonspermicidal broad-spectrum antiviral microbicide. This study evaluated the mucosal toxicity potential of native PAP in the in vivo rabbit vaginal irritation model as well as the in vitro reconstituted human vaginal epithelial tissue model. Twenty-two New Zealand white rabbits in 4 subgroups were exposed intravaginally to a gel with and without 0.01, 0.1, or 1.0% native PAP for 10 consecutive days. The dose of PAP used represented nearly 200- to 20,000 times its in vitro anti-HIV IC50 value. Animals were euthanized on day 11 and vaginal tissues were evaluated for histologic and immunohistochemical evidence of mucosal toxicity, cellular inflammation, and hyperplasia. Blood was analyzed for changes in hematology and clinical chemistry profiles. Reconstituted human vaginal epithelial tissue grown on membrane filters was exposed to 0.01, 0.1, or 1.0% native PAP in medium or topically via a gel for 24 hours and tissue damage was evaluated by histological assessment. In the in vivo rabbit vaginal irritation model, half of all PAP-treated rabbits (8/16) exhibited an acceptable range of vaginal mucosal irritation (total score <8 out of a possible 16), whereas nearly a third of PAP-treated rabbits (5/16) developed moderate to marked vaginal mucosal irritation (total score >11). However, no treatment-related adverse effects were seen in hematological or clinical chemistry measurements. Furthermore, in vitro exposure of a 3-dimensional human vaginal tissue grown on polycarbonate membrane filters to identical concentrations of PAP either added to culture medium or applied topically via gel formulation did not result in direct toxicity as determined by histologic evaluation. These findings indicate careful monitoring of vaginal irritation will be required in the clinical development of PAP as a nonspermicidal microbicide.

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

    PubMed

    Feng, Yongjia; Tsai, Yu-Hwai; Xiao, Weidong; Ralls, Matthew W; Stoeck, Alex; Wilson, Carole L; Raines, Elaine W; Teitelbaum, Daniel H; Dempsey, Peter J

    2015-11-01

    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.

  6. 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

  7. The interbranchial lymphoid tissue of Atlantic Salmon (Salmo salar L) extends as a diffuse mucosal lymphoid tissue throughout the trailing edge of the gill filament.

    PubMed

    Dalum, Alf S; Austbø, Lars; Bjørgen, Håvard; Skjødt, Karsten; Hordvik, Ivar; Hansen, Tom; Fjelldal, Per G; Press, Charles McL; Griffiths, David J; Koppang, Erling O

    2015-09-01

    The teleost gill forms an extensive, semipermeable barrier that must tolerate intimate contact with the surrounding environment and be able to protect the body from external pathogens. The recent discovery of the interbranchial lymphoid tissue (ILT) has initiated an anatomical and functional investigation of the lymphoid tissue of the salmonid gill. In this article, sectioning of gill arches in all three primary planes revealed an elongation of the ILT outward along the trailing edge of the primary filament to the very distal end, a finding not previously described. This newly found lymphoid tissue was investigated using a range of morphological and transcriptional tools. Avoiding potential salinity-related effects, the study focused on two fresh-water life stages-smoltifying juveniles and mature adults. Aggregates of T-cells continuous with the ILT were found within the thick epithelial lining of the trailing edge of the filament in considerably larger numbers than seen in the epithelium of the leading edge and of the interlamellar area. Only a few of these cells were identified as CD8α(+) -cells, and there was a significantly (P < 0.05) higher relative expression of CD4- than of CD8- related genes in all gill segments investigated. Numerous major histocompatibility complex class II(+) -cells were distributed uniformly throughout the filament epithelial tissue. Few Ig(+) -cells were detected. Overall, the morphological features and comparable immune gene expression of the previously described ILT and the filament trailing edge lymphoid tissue suggest a close functional and anatomical relationship. We propose that the anatomical definition of the ILT must be broadened to include both the previously described ILT (to be renamed proximal ILT) and the trailing edge lymphoid tissue (to be named distal ILT). This extended anatomical localisation identifies the ILT as a widely distributed mucosal lymphoid tissue in the gill of Atlantic salmon.

  8. A protective role for keratinocyte growth factor in a murine model of chemotherapy and radiotherapy-induced mucositis

    SciTech Connect

    Borges, Luis . E-mail: borgesl@amgen.com; Rex, Karen L.; Chen, Jennifer N.; Wei, Ping; Kaufman, Stephen; Scully, Sheila; Pretorius, James K.; Farrell, Catherine L.

    2006-09-01

    Purpose: To evaluate the activity of palifermin (rHuKGF) in a murine model of mucosal damage induced by a radiotherapy/chemotherapy (RT/CT) regimen mimicking treatment protocols used in head-and-neck cancer patients. Methods and Materials: A model of mucosal damage induced by RT/CT was established by injecting female BDF1 mice with cisplatin (10 mg/kg) on Day 1; 5-fluorouracil (40 mg/kg/day) on Days 1-4, and irradiation (5 Gy/day) to the head and neck on Days 1-5. Palifermin was administered subcutaneously on Days -2 to 0 (5 mg/kg/day) and on Day 5 (5 mg/kg). Evaluations included body weight, organ weight, keratinocyte growth factor receptor expression, epithelial thickness, and cellular proliferation. Results: Initiation of the radiochemotherapeutic regimen resulted in a reduction in body weight in control animals. Palifermin administration suppressed weight loss and resulted in increased organ weight (salivary glands and small intestine), epithelial thickness (esophagus and tongue), and cellular proliferation (tongue and salivary glands). Conclusions: Administration of palifermin before RT/CT promotes cell proliferation and increases in epithelial thickness in the oral mucosa, salivary glands, and digestive tract. Palifermin administration before and after RT/CT mitigates weight loss and a trophic effect on the intestinal mucosa and salivary glands, suggesting that palifermin use should be investigated further in the RT/CT settings, in which intestinal mucositis and salivary gland dysfunction are predominant side effects of cytotoxic therapy.

  9. [Methotrexate as inducer of proinflammatory cytokines by epithelial cells].

    PubMed

    Morón-Medina, Alejandra; Viera, Ninoska; de Morales, Thaís Rojas; Alcocer, Sirley; Bohorquez, Dinorath

    2014-03-01

    Methotrexate (MTX), a drug commonly used in childhood cancer, has also been indicated as a cytotoxic agent of the oral mucosa, which can trigger the inflammatory process and increase the vascularity of epithelial tissues during the early stages of oral mucositis. The aim of this study was to determine the production of proinflammatory cytokines IL-1beta, IL-6 y TNF-alpha in epithelial cell cultures treated with MTX. Epithelial cells of human larynx, obtained from the cell line Hep-2, were cultured with different doses of MTX during different incubation times. The drug cytotoxicity was analyzed by means of the colorimetric test, which is based on the metabolic reduction of the bromide of 3-(4, 5-dimetiltiazol-2-ilo)-2,5-difeniltetrazol (MTT); and the proinflammatory cytokines production by the test enzyme-linked immunosorbent assay (ELISA). Cultures of HEp