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Sample records for mucosal epithelial barrier

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

    PubMed

    Ross, Karen F; Herzberg, Mark C

    2016-06-01

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

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

    PubMed

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

    2008-07-01

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

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

    PubMed Central

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

    2014-01-01

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

  4. 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

  5. Intestinal inflammation and mucosal barrier function.

    PubMed

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

    2014-12-01

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

  6. Gastrointestinal mucosal barrier function and diseases.

    PubMed

    Oshima, Tadayuki; Miwa, Hiroto

    2016-08-01

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

  7. 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

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

    PubMed Central

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

    2015-01-01

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

  9. Biliary Mucosal Barrier and Microbiome

    PubMed Central

    Verdier, Julien; Luedde, Tom; Sellge, Gernot

    2015-01-01

    Background The biliary system is in continuous contact with the complex microbiota of the intestine. Microbial products have recently been proposed as potential triggers for biliary diseases. Methods The aim of this review is to provide a summary of the current knowledge regarding the role of the biliary and intestinal microbiome in biliary inflammatory diseases. Results Previously, it was suggested that the healthy biliary system is a sterile organ, while acute cholangitis and cholecystitis may occur from ascending infections. Although non-inflammatory biliary colonization by certain bacteria such as Salmonella spp. has been already recognized since several decades, human and animal studies indicated only very recently that the gallbladder harbors a complex microbiota also under non-pathologic conditions. Novel findings suggested that – similar to the situation in the intestine – the biliary mucosa features a chemical, mechanical, and immunological barrier, ensuring immunological tolerance against commensals. However, microbial triggers might influence acute and chronic inflammatory disease of the biliary system and the whole liver. Conclusion Although yet undefined, dysbiosis of the biliary or intestinal microbiota rather than a single microorganism may influence disease progression. PMID:26468308

  10. Effects of intraepithelial lymphocyte-derived cytokines on intestinal mucosal barrier function.

    PubMed

    Qiu, Yuan; Yang, Hua

    2013-10-01

    The mucosal surface of the gastrointestinal tract directly interacts with the mucosal lumen, which is continuously exposed to foreign antigens. Specialized intraepithelial lymphocytes (IELs), located between the basolateral surfaces of the epithelial cells, are important as the first line of defense against microbes as well as for their role in the maintenance of epithelial barrier homeostasis. Although IELs are mainly composed of T cells, they are phenotypically and functionally distinct from T cells in peripheral blood or the spleen. Not only are IELs stimulated by the antigens of the intestinal lumen but are they also stimulated by regulatory immune cells. The integrity of the intestinal mucosal barrier is closely tied to the IEL function. Cytokines produced by IELs modulate the cellular functions that trigger the downstream signaling pathways and mediate the barrier homeostasis. In this review, we will address the broad spectrum of cytokines that are derived from IELs and the functional regulation of these cytokines on the intestinal barrier. PMID:23692551

  11. Mucosal barrier, bacteria and inflammatory bowel disease: possibilities for therapy.

    PubMed

    Merga, Yvette; Campbell, Barry J; Rhodes, Jonathan M

    2014-01-01

    The mucosal barrier has three major components, the mucus layer, the epithelial glycocalyx and the surface epithelium itself, whose integrity largely depends on tight junction function. In health, there is relatively little direct interaction between the luminal microbiota and the epithelium - the continuous mucus layer in the colon keeps the surface epithelium out of contact with bacteria and the ileo-caecal valve ensures that the distal small intestine is relatively microbe free. Most interaction takes place at the Peyer's patches in the distal ileum and their smaller colonic equivalents, the lymphoid follicles. Peyer's patches are overlain by a 'dome' epithelium, 5% of whose cells are specialised M (microfold) epithelial cells, which act as the major portal of entry for bacteria. There are no goblet cells in the dome epithelium and M cells have a very sparse glycocalyx allowing easy microbial interaction. It is intriguing that the typical age range for the onset of Crohn's disease (CD) is similar to the age at which the number of Peyer's patches is greatest. Peyer's patches are commonly the sites of the initial lesions in CD and the 'anti-pancreatic' antibody associated with CD has been shown to have as its epitope the glycoprotein 2 that is the receptor for type-1 bacterial fimbrial protein (fimH) on M cells. There are many reasons to believe that the mucosal barrier is critically important in the pathogenesis of inflammatory bowel disease (IBD). These include (i) associations between both CD and ulcerative colitis (UC) with genes that are relevant to the mucosal barrier; (ii) increased intestinal permeability in unaffected relatives of CD patients; (iii) increased immune reactivity against bacterial antigens, and (iv) animal models in which altered mucosal barrier, e.g. denudation of the mucus layer associated with oral dextran sulphate in rodents, induces colitis. Whilst some IBD patients may have genetic factors leading to weakening of the mucosal barrier

  12. IgG transport across mucosal barriers by neonatal Fc receptor for IgG and mucosal immunity.

    PubMed

    Yoshida, Masaru; Masuda, Atsuhiro; Kuo, Timothy T; Kobayashi, Kanna; Claypool, Steven M; Takagawa, Tetsuya; Kutsumi, Hiromu; Azuma, Takeshi; Lencer, Wayne I; Blumberg, Richard S

    2006-12-01

    Mucosal secretions of the human gastrointestinal, respiratory, and genital tracts contain significant quantities of IgG. The neonatal Fc receptor for IgG (FcRn) plays a major role in regulating host IgG levels and transporting IgG and associated antigens across polarized epithelial barriers. The FcRn can then recycle the IgG/antigen complex back across the intestinal barrier into the lamina propria for processing by dendritic cells and presentation to CD4(+) T cells in regional organized lymphoid structures. FcRn, through its ability to secrete and absorb IgG, thus integrates luminal antigen encounters with systemic immune compartments and, as such, provides essential host defense and immunoregulatory functions at the mucosal surfaces. PMID:17051393

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

    PubMed Central

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

    2015-01-01

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

  14. 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...

  15. 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

  16. [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

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

    PubMed

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

    2008-01-01

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

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

    PubMed

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

    2015-03-01

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

  19. 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

  20. 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

  1. 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

  2. Epithelial Microvilli Establish an Electrostatic Barrier to Microbial Adhesion

    PubMed Central

    Bennett, Kaila M.; Walker, Sharon L.

    2014-01-01

    Microvilli are membrane extensions on the apical surface of polarized epithelia, such as intestinal enterocytes and tubule and duct epithelia. One notable exception in mucosal epithelia is M cells, which are specialized for capturing luminal microbial particles; M cells display a unique apical membrane lacking microvilli. Based on studies of M cell uptake under different ionic conditions, we hypothesized that microvilli may augment the mucosal barrier by providing an increased surface charge density from the increased membrane surface and associated glycoproteins. Thus, electrostatic charges may repel microbes from epithelial cells bearing microvilli, while M cells are more susceptible to microbial adhesion. To test the role of microvilli in bacterial adhesion and uptake, we developed polarized intestinal epithelial cells with reduced microvilli (“microvillus-minus,” or MVM) but retaining normal tight junctions. When tested for interactions with microbial particles in suspension, MVM cells showed greatly enhanced adhesion and uptake of particles compared to microvillus-positive cells. This preference showed a linear relationship to bacterial surface charge, suggesting that microvilli resist binding of microbes by using electrostatic repulsion. Moreover, this predicts that pathogen modification of electrostatic forces may contribute directly to virulence. Accordingly, the effacement effector protein Tir from enterohemorrhagic Escherichia coli O157:H7 expressed in epithelial cells induced a loss of microvilli with consequent enhanced microbial binding. These results provide a new context for microvillus function in the host-pathogen relationship, based on electrostatic interactions. PMID:24778113

  3. 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

  4. Epithelial Cell Shedding and Barrier Function

    PubMed Central

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

    2015-01-01

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

  5. Probiotic bacteria and intestinal epithelial barrier function.

    PubMed

    Ohland, Christina L; Macnaughton, Wallace K

    2010-06-01

    The intestinal tract is a diverse microenvironment where more than 500 species of bacteria thrive. A single layer of epithelium is all that separates these commensal microorganisms and pathogens from the underlying immune cells, and thus epithelial barrier function is a key component in the arsenal of defense mechanisms required to prevent infection and inflammation. The epithelial barrier consists of a dense mucous layer containing secretory IgA and antimicrobial peptides as well as dynamic junctional complexes that regulate permeability between cells. Probiotics are live microorganisms that confer benefit to the host and that have been suggested to ameliorate or prevent diseases including antibiotic-associated diarrhea, irritable bowel syndrome, and inflammatory bowel disease. Probiotics likely function through enhancement of barrier function, immunomodulation, and competitive adherence to the mucus and epithelium. This review summarizes the evidence about effects of the many available probiotics with an emphasis on intestinal barrier function and the mechanisms affected by probiotics. PMID:20299599

  6. The Effect of Peritoneal Air Exposure on Intestinal Mucosal Barrier

    PubMed Central

    Bao, Jun; Tan, Shanjun; Yu, Wenkui; Lin, Zhiliang; Dong, Yi; Chen, Qiyi; Shi, Jialiang; Duan, Kaipeng; Bai, Xiaowu; Xu, Lin; Li, Jieshou

    2014-01-01

    Background. Damage of the intestinal mucosa barrier may result in intestinal bacterial and endotoxin translocation, leading to local and systemic inflammation. The present study was designed to investigate whether peritoneal air exposure induces damage of intestinal mucosal barrier. Methods. Sprague-Dawley rats (weighing 210 to 230 g) were randomized into five groups (6/group): a control group, a sham group, and three exposure groups with peritoneal air exposure for 1, 2, and 3 h, respectively. At 24 h after surgery, blood and terminal ileum were sampled. The serum D-lactate levels were determined using an ELISA kit. The intestinal permeability was determined by measuring the intestinal clearance of FITC-dextran (FD4). The histopathological changes in terminal ileum were also assessed. Results. Compared with the controls, peritoneal air exposure caused an increase in both serum D-lactate level and intestinal FD4 clearance, which were proportional to the length of peritoneal air exposure and correlated to Chiu's scores, indices for intestinal mucosal injury. Edema and inflammatory cells were also observed in mucosa and submucosa of ileum in three exposure groups. Conclusions. Peritoneal air exposure could induce damage to the intestinal mucosal barrier, which is proportional to the time length of peritoneal air exposure. PMID:25210511

  7. The epithelial barrier is maintained by in vivo tight junction expansion during pathologic intestinal epithelial shedding

    PubMed Central

    Marchiando, Amanda M.; Shen, Le; Graham, W. Vallen; Edelblum, Karen L.; Duckworth, Carrie A.; Guan, Yanfang; Montrose, Marshall H.; Turner, Jerrold R.; Watson, Alastair J.M.

    2011-01-01

    BACKGROUND & AIMS Tumor necrosis factor (TNF) increases intestinal epithelial cell shedding and apoptosis, potentially challenging the barrier between the gastrointestinal lumen and internal tissues. We investigated the mechanism of tight junction remodeling and barrier maintenance, as well as the roles of cytoskeletal regulatory molecules during TNF-induced shedding. METHODS We studied wild-type and transgenic mice that express the fluorescent-tagged proteins enhanced green fluorescent protein–occludin or monomeric red fluorescent protein1–ZO-1. After injection of high doses of TNF (7.5µg, i.p.), laparotomies were performed and segments of small intestine were opened to visualize the mucosa by video confocal microscopy. Pharmacologic inhibitors and knockout mice were used to determine the roles of caspase activation, actomyosin, and microtubule remodeling and membrane trafficking in epithelial shedding. RESULTS Changes detected included redistribution of the tight junction proteins ZO-1 and occluding to lateral membranes of shedding cells. These proteins ultimately formed a funnel around the shedding cell that defined the site of barrier preservation. Claudins, E-cadherin, F-actin, myosin II, Rho-associated kinase (ROCK), and myosin light chain kinase (MLCK) were also recruited to lateral membranes. Caspase activity, myosin motor activity, and microtubules were required to initiate shedding, whereas completion of the process required microfilament remodeling and ROCK, MLCK, and dynamin II activities. CONCLUSIONS Maintenance of the epithelial barrier during TNF-induced cell shedding is a complex process that involves integration of microtubules, microfilaments, and membrane traffic to remove apoptotic cells. This process is accompanied by redistribution of apical junctional complex proteins to form intercellular barriers between lateral membranes and maintain mucosal function. PMID:21237166

  8. Epithelial crosstalk at the microbiota-mucosal interface.

    PubMed

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

    2011-03-15

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

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

    PubMed

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

    2015-05-01

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

  10. Trophic and cytoprotective nutrition for intestinal adaptation, mucosal repair, and barrier function.

    PubMed

    Ziegler, Thomas R; Evans, Mary E; Fernández-Estívariz, Concepción; Jones, Dean P

    2003-01-01

    Intestinal epithelial cell turnover (proliferation, migration, differentiation, and apoptosis) and gut barrier functions are dynamic processes that are markedly affected by nutritional status, the route of feeding, and the adequacy of specific nutrients in the diet. Emerging studies are defining potential therapeutic roles for specific nutrients and diet-derived compounds (including arginine, glutamate, glutamine, glutathione, glycine, vitamin A, zinc, and specific lipids) in gut mucosal turnover, repair, adaptation after massive bowel resection, and barrier function. The role and regulation of endogenous bowel flora in generating short-chain fatty acids from diet-derived fiber and other diet-derived compounds and the effects of these agents on gut function are increasingly being elucidated. Results of these investigations should define new nutritional methods for trophic and cytoprotective effects on the intestine in conditions such as inflammatory bowel disease, malnutrition, and short bowel syndrome. PMID:12626687

  11. Changes in Epithelial Barrier Function in Response to Parasitic Infection: Implications for IBD Pathogenesis

    PubMed Central

    Fernández-Blanco, Joan Antoni; Estévez, Javier; Shea-Donohue, Terez; Vergara, Patri

    2015-01-01

    Background and aims: Mast cells [MCs] are implicated in epithelial barrier alterations that characterize inflammatory and functional bowel disorders. In this study, we describe mast cell proteinases [chymases and tryptases] and tight junction [TJ] proteins kinetics in a rat model of postinfectious gut dysfunction. Methods: Jejunal tissues of control and -infected rats were used. Inflammation-related changes in MCs and the expression of TJ-related proteins were evaluated by immunostaining and reverse transcription-quantitative polymerase chain reaction. Epithelial barrier function was assessed in vitro (Ussing chambers) and in vivo. Results: After infection, intestinal inflammation was associated with a generalized overexpression of MC chymases, peaking between Days 6 and 14. Thereafter, a mucosal MC hyperplasia and a late increase in connective tissue MC counts were observed. From Day 2 post-infection, TJ proteins occludin and claudin-3 expression was down-regulated whereas the pore-forming protein claudin-2 was overexpressed. The expression of proglucagon, precursor of the barrier-enhancing factor glucagon-like peptide-2, was reduced. These changes were associated with an increase in epithelial permeability, both in vitro and in vivo. Conclusions: Proteinases expression and location of mucosal and connective tissue MCs indicate a time-related pattern in the maturation of intestinal MCs following infection. Altered expression of TJ-related proteins is consistent with a loss of epithelial tightness, and provides a molecular mechanism for the enhanced epithelial permeability observed in inflammatory conditions of the gut. PMID:25820018

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

    PubMed

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

    2016-09-01

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

  13. 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

  14. Comparison of the Transmembrane Mucins MUC1 and MUC16 in Epithelial Barrier Function

    PubMed Central

    Gipson, Ilene K.; Spurr-Michaud, Sandra; Tisdale, Ann; Menon, Balaraj B.

    2014-01-01

    Membrane-anchored mucins are present in the apical surface glycocalyx of mucosal epithelial cells, each mucosal epithelium having at least two of the mucins. The mucins have been ascribed barrier functions, but direct comparisons of their functions within the same epithelium have not been done. In an epithelial cell line that expresses the membrane-anchored mucins, MUC1 and MUC16, the mucins were independently and stably knocked down using shRNA. Barrier functions tested included dye penetrance, bacterial adherence and invasion, transepithelial resistance, tight junction formation, and apical surface size. Knockdown of MUC16 decreased all barrier functions tested, causing increased dye penetrance and bacterial invasion, decreased transepithelial resistance, surprisingly, disruption of tight junctions, and greater apical surface cell area. Knockdown of MUC1 did not decrease barrier function, in fact, barrier to dye penetrance and bacterial invasion increased significantly. These data suggest that barrier functions of membrane-anchored mucins vary in the context of other membrane mucins, and MUC16 provides a major barrier when present. PMID:24968021

  15. Gastric mucosal barrier: hydrophobicity of stretched stomach lining.

    PubMed

    Hills, B A; Lichtenberger, L M

    1985-06-01

    Surface hydrophobicity of the luminal lining of the canine stomach has been studied as a very convenient means of following the adsorbed monolayer of surfactant believed to provide the gastric mucosal barrier. Hydrophobicity has been measured as the contact angle (theta) produced when a drop of saline is placed upon the surface. theta was found to decrease from 82 to 62 degrees upon 50% linear extension of samples of oxyntic mucosa from 10 dogs. When the phospholipid believed to cause the hydrophobicity was absorbed to glass slides, the contact angle was found to decrease with lower surface concentration. Thinning or "crazing" of the absorbed surfactant monolayer imparting the very hydrophobic nature of the luminal lining is discussed as a possible reason why ulcers tend to form at the crests of the folds, i.e., at points where the surface has been stretched and the monolayer disrupted. PMID:4003546

  16. 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

  17. 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

  18. 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

  19. TLR2 mediates gap junctional intercellular communication through connexin-43 in intestinal epithelial barrier injury.

    PubMed

    Ey, Birgit; Eyking, Annette; Gerken, Guido; Podolsky, Daniel K; Cario, Elke

    2009-08-14

    Gap junctional intercellular communication (GJIC) coordinates cellular functions essential for sustaining tissue homeostasis; yet its regulation in the intestine is not well understood. Here, we identify a novel physiological link between Toll-like receptor (TLR) 2 and GJIC through modulation of Connexin-43 (Cx43) during acute and chronic inflammatory injury of the intestinal epithelial cell (IEC) barrier. Data from in vitro studies reveal that TLR2 activation modulates Cx43 synthesis and increases GJIC via Cx43 during IEC injury. The ulcerative colitis-associated TLR2-R753Q mutant targets Cx43 for increased proteasomal degradation, impairing TLR2-mediated GJIC during intestinal epithelial wounding. In vivo studies using mucosal RNA interference show that TLR2-mediated mucosal healing depends functionally on intestinal epithelial Cx43 during acute inflammatory stress-induced damage. Mice deficient in TLR2 exhibit IEC-specific alterations in Cx43, whereas administration of a TLR2 agonist protects GJIC by blocking accumulation of Cx43 and its hyperphosphorylation at Ser368 to prevent spontaneous chronic colitis in MDR1alpha-deficient mice. Finally, adding the TLR2 agonist to three-dimensional intestinal mucosa-like cultures of human biopsies preserves intestinal epithelial Cx43 integrity and polarization ex vivo. In conclusion, Cx43 plays an important role in innate immune control of commensal-mediated intestinal epithelial wound repair. PMID:19528242

  20. 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

  1. Functional changes of intestinal mucosal barrier in surgically critical patients

    PubMed Central

    Guo, Yuan-yuan; Liu, Mu-lin; He, Xian-di; Jiang, Cong-qiao; Liu, Rui-lin

    2010-01-01

    BACKGROUND: The gut is capable of inducing multiple organ dysfunction syndrome (MODS). In the diagnosis and treatment of critical ill patients, doctors should pay particular attention to the protection or recovery of intestinal barrier function. However, no reliable diagnostic criteria are available clinically. This study aimed to assess the changes of intestinal mucosal barrier function in surgically critical ill patients as well as their significance. METHODS: Thirty-eight surgically critical ill patients were enrolled as a study group (APACHE II>8 scores), and 15 non-critical ill patients without intestinal dysfunction were selected as a control group (APACHE II<6). General information, symptoms, physical signs, and APACHE II scores of the patients were recorded. The patients in the study group were subdivided into an intestinal dysfunction group (n=26) and a non-intestinal dysfunction group (n=12). Three ml venous blood was collected from the control group on admission and the same volume of plasma was collected from the study group both on admission and in the period of recovery. The plasma concentrations of endotoxin, diamine oxidase (DAO), D-lactate, and intestinal fatty-acid binding protein (iFABP) were detected respectively. The data collected were analyzed by the SPSS 17.0 software for Windows. RESULTS: The levels of variables were significantly higher in the study group than in the control group (P<0.01). They were higher in the intestinal dysfunction group than in the non-intestinal dysfunction group (DAO P<0.05, endotoxin, D-lactate, iFABP P<0.01). In the non-intestinal dysfunction group compared with the control group, the level of endotoxin was not significant (P>0.05), but the levels of DAO, D-lactate and iFABP were statistically significant (P<0.05). The levels of variables in acute stage were higher than those in recovery stage (P<0.01). The death group showed higher levels of variables than the survival group (endotoxin and D-lactate P<0.01, DAO

  2. 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.

  3. Effect of fecal water on an in vitro model of colonic mucosal barrier function.

    PubMed

    Gill, Chris I R; Heavey, Patricia; McConville, Eileen; Bradbury, Ian; Fässler, Caroline; Mueller, Susanne; Cresci, Alberto; Dore, Joel; Norin, Elisabeth; Rowland, Ian

    2007-01-01

    Fecal water (FW) has been shown to exert, in cultured cells, cytotoxic and genotoxic effects that have implications for colorectal cancer (CRC) risk. We have investigated a further biological activity of FW, namely, the ability to affect gap junctions in CACO2 cell monolayers as an index of mucosal barrier function, which is known to be disrupted in cancer. FW samples from healthy, free-living, European subjects that were divided into two broad age groups, adult (40+/-9.7 yr; n=53) and elderly (76+/-7.5 yr; n=55) were tested for effects on gap junction using the transepithelial resistance (TER) assay. Overall, treatment of CACO2 cells with FW samples from adults increased TER (+4%), whereas FW from elderly subjects decreased TER (-5%); the difference between the two groups was significant (P<0.05). We also measured several components of FW potentially associated with modulation of TER, namely, short-chain fatty acid (SCFA) and ammonia. SCFAs (propionic, acetic, and n-butyric) were significantly lower in the elderly population (-30%, -35%, and -21%, respectively, all P A pound 0.01). We consider that FW modulation of in vitro epithelial barrier function is a potentially useful noninvasive biomarker, but it requires further validation to establish its relationship to CRC risk. PMID:17516863

  4. Berberine Prevents Intestinal Mucosal Barrier Damage During Early Phase of Sepsis in Rat through the Toll-Like Receptors Signaling Pathway

    PubMed Central

    Li, Guo-xun; Wang, Xi-mo; Jiang, Tao; Gong, Jian-feng; Niu, Ling-ying

    2015-01-01

    Our previous study has shown berberine prevents damage to the intestinal mucosal barrier during early phase of sepsis in rat through mechanisms independent of the NOD-like receptors signaling pathway. In this study, we explored the regulatory effects of berberine on Toll-like receptors during the intestinal mucosal damaging process in rats. Male Sprague-Dawlay (SD) rats were treated with berberine for 5 d before undergoing cecal ligation and puncture (CLP) to induce polymicrobial sepsis. The expression of Toll-like receptor 2 (TLR 2), TLR 4, TLR 9, the activity of nuclear factor-kappa B (NF-κB), the levels of selected cytokines and chemokines, percentage of cell death in intestinal epithelial cells, and mucosal permeability were investigated at 0, 2, 6, 12 and 24 h after CLP. Results showed that the tumor necrosis factor-α (TNF-α ) and interleukin-6 (IL-6) level were significantly lower in berberine-treated rats compared to the control animals. Conversely, the expression level of tight junction proteins, percentage of cell death in intestinal epithelial cells and the mucosal permeability were significantly higher in berberine-treated rats. The mRNA expression of TLR 2, TLR 4, and TLR 9 were significantly affected by berberine treatment. Our results indicate that pretreatment with berberine attenuates tissue injury and protects the intestinal mucosal barrier in early phase of sepsis and this may possibly have been mediated through the TLRs pathway. PMID:25605990

  5. Trek1 contributes to maintaining nasal epithelial barrier integrity.

    PubMed

    Jiang, Jing; Liu, Jiang-Qi; Li, Jing; Li, Meng; Chen, Hong-Bin; Yan, Hao; Mo, Li-Hua; Qiu, Shu-Qi; Liu, Zhi-Gang; Yang, Ping-Chang

    2015-01-01

    Epithelial barrier integrity is critical to maintain the homeostasis in the body. The regulatory mechanism of the epithelial barrier function has not been fully understood. This study aims to elucidate the role of the TWIK-related potassium channel-1 (Trek1) in the regulation of the epithelial barrier function of the nasal mucosa. In this study, the levels of Trek1 were assessed by real time RT-PCR and Western blotting. The epithelial barrier function of the rat nasal epithelia was evaluated by the Ussing chamber system. The results showed that Trek1 was detected in the human and rat nasal epithelia, which were significantly lower in patients and rats with allergic rhinitis than that in healthy controls. Exposure to the signature T helper 2 cytokine, interleukin (IL)-4, markedly suppressed the expression of Trek1 in the nasal mucosa via up regulating the expression of the histone deacetylase (HDAC)1. The IL-4-induced rat nasal epithelial barrier dysfunction could be blocked by HDAC1 inhibitor (Trichostatin A), or sodium butyrate, or administration of Clostridium Butyricum. We conclude that Trek1 is critical to maintain the nasal epithelial barrier function. PMID:25778785

  6. 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

  7. 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

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

    PubMed

    Wu, Yunpeng; Zhu, Cui; Chen, Zhuang; Chen, Zhongjian; Zhang, Weina; Ma, Xianyong; Wang, Li; Yang, Xuefen; Jiang, Zongyong

    2016-04-01

    Tight junctions (TJs) play an important role in maintaining the mucosal barrier function and gastrointestinal health of animals. Lactobacillus plantarum (L. plantarum) was reported to protect the intestinal barrier function of early-weaned piglets against enterotoxigenic Escherichia coli (ETEC) K88 challenge; however, the underlying cellular mechanism of this protection was unclear. Here, an established intestinal porcine epithelia cell (IPEC-J2) model was used to investigate the protective effects and related mechanisms of L. plantarum on epithelial barrier damages induced by ETEC K88. Epithelial permeability, expression of inflammatory cytokines, and abundance of TJ proteins, were determined. Pre-treatment with L. plantarum for 6h prevented the reduction in transepithelial electrical resistance (TEER) (P<0.05), inhibited the increased transcript abundances of interleukin-8 (IL-8) and tumor necrosis factor (TNF-α) (P<0.05), decreased expression of claudin-1, occludin and zonula occludens (ZO-1) (P<0.05) and protein expression of occludin (P<0.05) of IPEC-J2 cells caused by ETEC K88. Moreover, the mRNA expression of negative regulators of toll-like receptors (TLRs) [single Ig Il-1-related receptor (SIGIRR), B-cell CLL/lymphoma 3 (Bcl3), and mitogen-activated protein kinase phosphatase-1 (MKP-1)] in IPEC-J2 cells pre-treated with L. plantarum were higher (P<0.05) compared with those in cells just exposed to K88. Furthermore, L. plantarum was shown to regulate proteins of nuclear factor κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways. These results indicated that L. plantarum may improve epithelial barrier function by maintenance of TEER, inhibiting the reduction of TJ proteins, and reducing the expression of proinflammatory cytokines induced by ETEC K88, possibly through modulation of TLRs, NF-κB and MAPK pathways. PMID:27032504

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

    PubMed

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

    2016-06-01

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

  10. Nitric Oxide and Airway Epithelial Barrier Function: Regulation of Tight Junction Proteins and Epithelial Permeability

    PubMed Central

    Olson, Nels; Greul, Anne-Katrin; Hristova, Milena; Bove, Peter F.; Kasahara, David I.; van der Vliet, Albert

    2008-01-01

    Acute airway inflammation is associated with enhanced production of nitric oxide (NO•) and altered airway epithelial barrier function, suggesting a role of NO• or its metabolites in epithelial permeability. While high concentrations of S-nitrosothiols disrupted transepithelial resistance (TER) and increased permeability in 16HBE14o- cells, no significant barrier disruption was observed by NONOates, in spite of altered distribution and expression of some TJ proteins. Barrier disruption of mouse tracheal epithelial (MTE) cell monolayers in response to inflammatory cytokines was independent of NOS2, based on similar effects in MTE cells from NOS2-/- mice and a lack of effect of the NOS2-inhibitor 1400W. Cell pre-incubation with LPS protected MTE cells from TER loss and increased permeability by H2O2, which was independent of NOS2. However, NOS2 was found to contribute to epithelial wound repair and TER recovery after mechanical injury. Overall, our results demonstrate that epithelial NOS2 is not responsible for epithelial barrier dysfunction during inflammation, but may contribute to restoration of epithelial integrity. PMID:19100237

  11. 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.

  12. HVEM signalling at mucosal barriers provides host defence against pathogenic bacteria.

    PubMed

    Shui, Jr-Wen; Larange, Alexandre; Kim, Gisen; Vela, Jose Luis; Zahner, Sonja; Cheroutre, Hilde; Kronenberg, Mitchell

    2012-08-01

    The herpes virus entry mediator (HVEM), a member of the tumour-necrosis factor receptor family, has diverse functions, augmenting or inhibiting the immune response. HVEM was recently reported as a colitis risk locus in patients, and in a mouse model of colitis we demonstrated an anti-inflammatory role for HVEM, but its mechanism of action in the mucosal immune system was unknown. Here we report an important role for epithelial HVEM in innate mucosal defence against pathogenic bacteria. HVEM enhances immune responses by NF-κB-inducing kinase-dependent Stat3 activation, which promotes the epithelial expression of genes important for immunity. During intestinal Citrobacter rodentium infection, a mouse model for enteropathogenic Escherichia coli infection, Hvem−/− mice showed decreased Stat3 activation, impaired responses in the colon, higher bacterial burdens and increased mortality. We identified the immunoglobulin superfamily molecule CD160 (refs 7 and 8), expressed predominantly by innate-like intraepithelial lymphocytes, as the ligand engaging epithelial HVEM for host protection. Likewise, in pulmonary Streptococcus pneumoniae infection, HVEM is also required for host defence. Our results pinpoint HVEM as an important orchestrator of mucosal immunity, integrating signals from innate lymphocytes to induce optimal epithelial Stat3 activation, which indicates that targeting HVEM with agonists could improve host defence. PMID:22801499

  13. Differential Targeting of the E-Cadherin/β-Catenin Complex by Gram-Positive Probiotic Lactobacilli Improves Epithelial Barrier Function

    PubMed Central

    Hummel, Stephanie; Veltman, Katharina; Cichon, Christoph; Sonnenborn, Ulrich

    2012-01-01

    The intestinal ecosystem is balanced by dynamic interactions between resident and incoming microbes, the gastrointestinal barrier, and the mucosal immune system. However, in the context of inflammatory bowel diseases (IBD), where the integrity of the gastrointestinal barrier is compromised, resident microbes contribute to the development and perpetuation of inflammation and disease. Probiotic bacteria have been shown to exert beneficial effects, e.g., enhancing epithelial barrier integrity. However, the mechanisms underlying these beneficial effects are only poorly understood. Here, we comparatively investigated the effects of four probiotic lactobacilli, namely, Lactobacillus acidophilus, L. fermentum, L. gasseri, and L. rhamnosus, in a T84 cell epithelial barrier model. Results of DNA microarray experiments indicating that lactobacilli modulate the regulation of genes encoding in particular adherence junction proteins such as E-cadherin and β-catenin were confirmed by quantitative reverse transcription-PCR (qRT-PCR). Furthermore, we show that epithelial barrier function is modulated by Gram-positive probiotic lactobacilli via their effect on adherence junction protein expression and complex formation. In addition, incubation with lactobacilli differentially influences the phosphorylation of adherence junction proteins and the abundance of protein kinase C (PKC) isoforms such as PKCδ that thereby positively modulates epithelial barrier function. Further insight into the underlying molecular mechanisms triggered by these probiotics might also foster the development of novel strategies for the treatment of gastrointestinal diseases (e.g., IBD). PMID:22179242

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

    PubMed

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

    2014-05-22

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

  15. 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

  16. 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

  17. CaMKII regulates the strength of the epithelial barrier.

    PubMed

    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 Ca(2+)/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

  18. 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

  19. Intracellular mediators of JAM-A–dependent epithelial barrier function

    PubMed Central

    Monteiro, Ana C.; Parkos, Charles A.

    2012-01-01

    JAM-A is a critical signaling component of the apical junctional complex, a structure composed of several transmembrane and scaffold molecules that controls the passage of nutrients and solutes across epithelial surfaces. Observations from JAM-A–deficient epithelial cells and JAM-A knockout animals indicate that JAM-A is an important regulator of epithelial paracellular permeability, however the mechanism(s) linking JAM-A to barrier function are not understood. This review highlights recent findings relevant to JAM-A–mediated regulation of epithelial permeability, focusing on the role of upstream and downstream signaling candidates. We draw on what is known about proteins reported to associate with JAM-A in other pathways and on known modulators of barrier function to propose candidate effectors that may mediate JAM-A regulation of epithelial paracellular permeability. Further investigation of pathways highlighted in this review may provide ideas for novel therapeutics that target debilitating conditions associated with barrier dysfunction, such as inflammatory bowel disease. PMID:22671597

  20. 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

  1. 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

  2. Simulated Reflux Decreases Vocal Fold Epithelial Barrier Resistance

    PubMed Central

    Erickson, Elizabeth; Sivasankar, Mahalakshmi

    2010-01-01

    Objectives/Hypothesis The vocal fold epithelium provides a barrier to the entry of inhaled and systemic challenges. However, the location of the epithelium makes it vulnerable to damage. Past research suggests, but does not directly demonstrate, that exposure to gastric reflux adversely affects the function of the epithelial barrier. Understanding the nature of reflux-induced epithelial barrier dysfunction is necessary to better recognize the mechanisms for vocal fold susceptibility to this disease. Therefore, we examined the effects of physiologically relevant reflux challenges on vocal fold transepithelial resistance and gross epithelial and subepithelial appearance. Study Design Ex vivo, mixed design with between-group and repeated-measures analyses. Methods Healthy, native porcine vocal folds (N = 52) were exposed to physiologically relevant acidic pepsin, acid-only, or pepsin-only challenges and examined with electrophysiology and light microscopy. For all challenges, vocal folds exposed to a neutral pH served as control. Results Acidic pepsin and acid-only challenges, but not pepsin-only or control challenges significantly reduced transepithelial resistance within 30 minutes. Reductions in transepithelial resistance were irreversible. Challenge exposure produced minimal gross changes in vocal fold epithelial or subepithelial appearance as evidenced by light microscopy. Conclusions These findings demonstrate that acidic environments characteristic of gastric reflux compromise epithelial barrier function without gross structural changes. In healthy, native vocal folds, reductions in transepithelial resistance could reflect reflux-related epithelial disruption. These results might guide the development of pharmacologic and therapeutic recommendations for patients with reflux, such as continued acid-suppression therapy and patient antireflux behavioral education. PMID:20564752

  3. 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

  4. 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

  5. Glucagon-like peptide-2 protects impaired intestinal mucosal barriers in obstructive jaundice rats

    PubMed Central

    Chen, Jun; Dong, Jia-Tian; Li, Xiao-Jing; Gu, Ye; Cheng, Zhi-Jian; Cai, Yuan-Kun

    2015-01-01

    AIM: To observe the protective effect of glucagon-like peptide-2 (GLP-2) on the intestinal barrier of rats with obstructive jaundice and determine the possible mechanisms of action involved in the protective effect. METHODS: Thirty-six Sprague-Dawley rats were randomly divided into a sham operation group, an obstructive jaundice group, and a GLP-2 group; each group consisted of 12 rats. The GLP-2 group was treated with GLP-2 after the day of surgery, whereas the other two groups were treated with the same concentration of normal saline. Alanine aminotransferase (ALT), total bilirubin, and endotoxin levels were recorded at 1, 3, 7, 10 and 14 d. Furthermore, on the 14th day, body weight, the wet weight of the small intestine, pathological changes of the small intestine and the immunoglobulin A (IgA) expressed by plasma cells located in the small intestinal lamina propria were recorded for each group. RESULTS: In the rat model, jaundice was obvious, and the rats’ activity decreased 4-6 d post bile duct ligation. Compared with the sham operation group, the obstructive jaundice group displayed increased yellow staining of abdominal visceral serosa, decreased small intestine wet weight, thinning of the intestinal muscle layer and villi, villous atrophy, uneven height, fusion, partial villous epithelial cell shedding, substantial inflammatory cell infiltration and significantly reduced IgA expression. However, no significant gross changes were noted between the GLP-2 and sham groups. With time, the levels of ALT, endotoxin and bilirubin in the GLP-2 group were significantly increased compared with the sham group (P < 0.01). The increasing levels of the aforementioned markers were more significant in the obstructive jaundice group than in the GLP-2 group (P < 0.01). CONCLUSION: GLP-2 reduces intestinal mucosal injuries in obstructive jaundice rats, which might be attributed to increased intestinal IgA and reduced bilirubin and endotoxin. PMID:25593463

  6. Traversal of a polarized epithelium by pathogenic Neisseriae: facilitation by type IV pili and maintenance of epithelial barrier function.

    PubMed Central

    Merz, A. J.; Rifenbery, D. B.; Arvidson, C. G.; So, M.

    1996-01-01

    BACKGROUND: Gonococci (GC) and meningococci (MC) are gram-negative bacterial pathogens that infect human mucosal epithelia. We would like to understand the functions of specific bacterial components at each stage of mucosal colonization: adhesion, cell invasion, and traversal into subepithelial tissues. As no animal model of mucosal colonization by GC or MC is available, increasingly sophisticated in vitro approaches have been used to address these issues. MATERIALS AND METHODS: We adapted the polarized T84 human epithelial cell system to study GC and MC colonization. Epithelial barrier function was monitored by permeability to soluble tracers and with electrical resistance measurements. Polarized cells were used to assay bacterial traversal of the monolayers, and cells grown on plastic were used to assay adhesion and cell invasion. RESULTS: All pathogenic Neisseriae examined traversed the monolayers. The traversal times were species specific and identical to times established previously in organ culture studies. In contrast to experiments with some enteric pathogens, transmigration by GC and MC was not accompanied by disruption of the epithelial barrier. GC mutants lacking type IV pili were compromised in adhesion, invasion, and traversal of T84 cells. CONCLUSIONS: Experiments with polarized T84 cells mimic key features of organ culture infections and reveal additional aspects of neisserial infection. Epithelial barrier function can be retained during bacterial traversal. Experiments with a nonpiliated GC mutant and its wild-type parent indicated an unexpected role for pili in cell invasion. Our results are consistent with the hypothesis that bacterial adhesion, invasion, or both are rate-limiting for traversal across the epithelium. Images FIG. 1 FIG. 3 PMID:8972489

  7. 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

  8. 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

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

    PubMed Central

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

    2014-01-01

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

  10. 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

  11. Compromised intestinal epithelial barrier induces adaptive immune compensation that protects from colitis

    PubMed Central

    Khounlotham, Manirath; Kim, Wooki; Peatman, Eric; Nava, Porfirio; Medina-Contreras, Oscar; Addis, Caroline; Koch, Stefan; Fournier, Benedicte; Nusrat, Asma; Denning, Timothy L.; Parkos, Charles A.

    2012-01-01

    SUMMARY Mice lacking Junctional Adhesion Molecule A (JAM-A, encoded by F11r) exhibit enhanced intestinal epithelial permeability, bacterial translocation, and elevated colonic lymphocyte numbers, yet do not develop colitis. To investigate the contribution of adaptive immune compensation in response to increased intestinal epithelial permeability, we examined the susceptibility of F11r-/-Rag1-/- mice to acute colitis. Although negligible contributions of adaptive immunity in F11r-/-Rag1-/- mice were observed, F11r-/-Rag1-/- mice exhibited increased microflora-dependent colitis. Elimination of T cell subsets and cytokine analyses revealed a protective role for TGF-β-producing CD4+ T cells in F11r-/- mice. Additionally, loss of JAM-A resulted in elevated mucosal and serum IgA that was dependent upon CD4+ T cells and TGF-β. Absence of IgA in F11r+/+Igha-/- mice did not affect disease whereas F11r-/-Igha-/- mice displayed markedly increased susceptibility to acute injury induced colitis. These data establish a role for adaptive immune mediated protection from acute colitis under conditions of intestinal epithelial barrier compromise. PMID:22981539

  12. 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

  13. Early weaning stress impairs development of mucosal barrier function in the porcine intestine

    PubMed Central

    Smith, Feli; Clark, Jessica E.; Overman, Beth L.; Tozel, Christena C.; Huang, Jennifer H.; Rivier, Jean E. F.; Blisklager, Anthony T.

    2010-01-01

    Early life stress is a predisposing factor for the development of chronic intestinal disorders in adult life. Here, we show that stress associated with early weaning in pigs leads to impaired mucosal barrier function. Early weaning (15- to 21-day weaning age) resulted in sustained impairment in intestinal barrier function, as indicated by reductions in jejunal transepithelial electrical resistance and elevations in mucosal-to-serosal flux of paracellular probes [3H]mannitol and [14C]inulin measured at 5 and 9 wk of age, compared with that shown in late-weaned pigs (23- to 28-day weaning age). Elevated baseline short-circuit current was observed in jejunum from early-weaned pigs and was shown to be mediated via enhanced Cl− secretion. Jejunal barrier dysfunction in early-weaned pigs coincided with increased lamina propria immune cell density particularly mucosal mast cells. The mast cell stabilizer drug sodium cromoglycolate ameliorated barrier dysfunction and hypersecretion in early-weaned pigs, demonstrating an important role of mast cells. Furthermore, activation of mast cells ex vivo with c48/80 and corticotrophin-releasing factor (CRF) in pig jejunum mounted in Ussing chambers induced barrier dysfunction and elevations in short-circuit current that were inhibited with mast cell protease inhibitors. Experiments in which selective CRF receptor antagonists were administered to early-weaned pigs revealed that CRF receptor 1 (CRFr1) activation mediates barrier dysfunction and hypersecretion, whereas CRFr2 activation may be responsible for novel protective properties in the porcine intestine in response to early life stress. PMID:19926814

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

    PubMed Central

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

    2005-01-01

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

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

    PubMed

    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

  16. Cervicovaginal microbiome dysbiosis is associated with proteome changes related to alterations of the cervicovaginal mucosal barrier.

    PubMed

    Borgdorff, H; Gautam, R; Armstrong, S D; Xia, D; Ndayisaba, G F; van Teijlingen, N H; Geijtenbeek, T B H; Wastling, J M; van de Wijgert, J H H M

    2016-05-01

    Vaginal microbiome (VMB) dysbiosis is associated with increased acquisition of HIV. Cervicovaginal inflammation and other changes to the mucosal barrier are thought to have important roles but human data are scarce. We compared the human cervicovaginal proteome by mass spectrometry of 50 Rwandan female sex workers who had previously been clustered into four VMB groups using a 16S phylogenetic microarray; in order of increasing bacterial diversity: Lactobacillus crispatus-dominated VMB (group 1), Lactobacillus iners-dominated VMB (group 2), moderate dysbiosis (group 3), and severe dysbiosis (group 4). We compared relative protein abundances among these VMB groups using targeted (abundance of pre-defined mucosal barrier proteins) and untargeted (differentially abundant proteins among all human proteins identified) approaches. With increasing bacterial diversity, we found: mucus alterations (increasing mucin 5B and 5AC), cytoskeleton alterations (increasing actin-organizing proteins; decreasing keratins and cornified envelope proteins), increasing lactate dehydrogenase A/B as markers of cell death, increasing proteolytic activity (increasing proteasome core complex proteins/proteases; decreasing antiproteases), altered antimicrobial peptide balance (increasing psoriasin, calprotectin, and histones; decreasing lysozyme and ubiquitin), increasing pro-inflammatory cytokines, and decreasing immunoglobulins immunoglobulin G1/2. Although temporal relationships cannot be derived, our findings support the hypothesis that dysbiosis causes cervicovaginal inflammation and other detrimental changes to the mucosal barrier. PMID:26349657

  17. 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

  18. Zinc modulates cytokine-induced lung epithelial cell barrier permeability.

    PubMed

    Bao, Shenying; Knoell, Daren L

    2006-12-01

    Apoptosis plays a causative role in acute lung injury in part due to epithelial cell loss. We recently reported that zinc protects the lung epithelium during inflammatory stress whereas depletion of intracellular zinc enhances extrinsic apoptosis. In this investigation, we evaluated the relationship between zinc, caspase-3, and cell-to-cell contact via proteins that form the adherens junction complex. Cell adhesion proteins are directly responsible for formation of the mechanical barrier of the lung epithelium. We hypothesized that exposure to inflammatory cytokines, in conjunction with zinc deprivation, would induce caspase-3, leading to degradation of junction proteins, loss of cell-to-cell contact, and compromised barrier function. Primary human upper airway and type I/II alveolar epithelial cultures were obtained from multiple donors and exposed to inflammatory stimuli that provoke extrinsic apoptosis in addition to depletion of intracellular zinc. We observed that zinc deprivation combined with tumor necrosis factor-alpha, interferon-gamma, and Fas receptor ligation accelerates caspase-3 activation, proteolysis of E-cadherin and beta-catenin, and cellular apoptosis, leading to increased paracellular leak across monolayers of both upper airway and alveolar lung epithelial cultures. Zinc supplementation inhibited apoptosis and paracellular leak, whereas caspase inhibition was less effective. We conclude that zinc is a vital factor in the lung epithelium that protects against death receptor-mediated apoptosis and barrier dysfunction. Furthermore, our findings suggest that although caspase-3 inhibition reduces lung epithelial apoptosis it does not prevent mechanical dysfunction. These findings facilitate future studies aimed at developing therapeutic strategies to prevent acute lung injury. PMID:16844947

  19. 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

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

    PubMed

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

    2016-04-01

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

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

    PubMed Central

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

    2014-01-01

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

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

    PubMed

    Szefel, Jarosław; 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

  3. 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

  4. Activation of toll like receptor-3 induces corneal epithelial barrier dysfunction.

    PubMed

    Wei, Jie; Jiang, Hua; Gao, Hongrui; Wang, Guangjie

    2015-06-01

    The epithelial barrier is critical in the maintenance of the homeostasis of the cornea. A number of eye disorders are associated with the corneal epithelial barrier dysfunction. Viral infection is one common eye disease type. This study aims to elucidate the mechanism by which the activation of toll like receptor 3 (TLR3) in the disruption of the corneal epithelial barrier. In this study, HCE cells (a human corneal epithelial cell line) were cultured into epithelial layers using as an in vitro model of the corneal epithelial barrier. PolyI:C was used as a ligand of TLR3. The transepithelial electric resistance (TER) and permeability of the HCE epithelial layer were assessed using as the parameters to evaluate the corneal epithelial barrier integrity. The results showed that exposure to PolyI:C markedly decreased the TER and increased the permeability of the HCE epithelial layers; the levels of cell junction protein, E-cadherin, were repressed by PolyI:C via increasing histone deacetylase-1 (HDAC1), the latter binding to the promoter of E-cadherin and repressed the transcription of E-cadherin. The addition of butyrate (an inhibitor of HDAC1) to the culture blocked the corneal epithelial barrier dysfunction caused by PolyI:C. In conclusion, activation of TLR3 can disrupt the corneal epithelial barrier, which can be blocked by the inhibitor of HDAC1. PMID:25912142

  5. Epithelial barrier modulation by a channel forming peptide.

    PubMed

    Somasekharan, Suma; Brandt, Robert; Iwamoto, Takeo; Tomich, John M; Schultz, Bruce D

    2008-03-01

    NC-1059 is a synthetic channel-forming peptide that provides for ion transport across, and transiently reduces the barrier integrity of, cultured epithelial monolayers derived from canine kidney (MDCK cells). Experiments were conducted to determine whether epithelial cells derived from other sources were similarly affected. Epithelial cells derived from human intestine (T-84), airway (Calu-3), porcine intestine (IPEC-J2) and reproductive duct (PVD9902) were grown on permeable supports. Basal short circuit current (Isc) was <3 microA cm(-2) for T-84, IPEC-J2 and PVD9902 cell monolayers and <8 microA cm(-2) for Calu-3 cells. Apical NC-1059 exposure caused, in all cell types, an increase in Isc to >15 microA cm(-2), indicative of net anion secretion or cation absorption, which was followed by an increase in transepithelial conductance (in mS cm(-2): T-84, 1.6 to 62; PVD9902, 0.2 to 51; IPEC-J2, 0.3 to 26; Calu-3, 2.3 to 13). These results are consistent with the peptide affecting transcellular ion movement, with a likely effect also on the paracellular route. NC-1059 exposure increased dextran permeation when compared to basal permeation, which documents an effect on the paracellular pathway. In order to evaluate membrane ion channels, experiments were conducted to study the dose dependence and stability of the NC-1059-induced membrane conductance in Xenopus laevis oocytes. NC-1059 induced a dose-dependent increase in oocyte membrane conductance that remained stable for greater than 2 h. The results demonstrate that NC-1059 increases transcellular conductance and paracellular permeation in a wide range of epithelia. These effects might be exploited to promote drug delivery across barrier epithelia. PMID:18418541

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

    PubMed

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

    2013-01-01

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

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

    PubMed

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

    2016-05-01

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

  8. Coordinated epithelial NHE3 inhibition and barrier dysfunction are required for TNF-mediated diarrhea in vivo

    PubMed Central

    Clayburgh, Daniel R.; Musch, Mark W.; Leitges, Michael; Fu, Yang-Xin; Turner, Jerrold R.

    2006-01-01

    Acute T cell–mediated diarrhea is associated with increased mucosal expression of proinflammatory cytokines, including the TNF superfamily members TNF and LIGHT. While we have previously shown that epithelial barrier dysfunction induced by myosin light chain kinase (MLCK) is required for the development of diarrhea, MLCK inhibition does not completely restore water absorption. In contrast, although TNF-neutralizing antibodies completely restore water absorption after systemic T cell activation, barrier function is only partially corrected. This suggests that, while barrier dysfunction is critical, other processes must be involved in T cell–mediated diarrhea. To define these processes in vivo, we asked whether individual cytokines might regulate different events in T cell–mediated diarrhea. Both TNF and LIGHT caused MLCK-dependent barrier dysfunction. However, while TNF caused diarrhea, LIGHT enhanced intestinal water absorption. Moreover, TNF, but not LIGHT, inhibited Na+ absorption due to TNF-induced internalization of the brush border Na+/H+ exchanger NHE3. LIGHT did not cause NHE3 internalization. PKCα activation by TNF was responsible for NHE3 internalization, and pharmacological or genetic PKCα inhibition prevented NHE3 internalization, Na+ malabsorption, and diarrhea despite continued barrier dysfunction. These data demonstrate the necessity of coordinated Na+ malabsorption and barrier dysfunction in TNF-induced diarrhea and provide insight into mechanisms of intestinal water transport. PMID:17016558

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

    PubMed Central

    2009-01-01

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

  10. PLEKHA7 modulates epithelial tight junction barrier function

    PubMed Central

    Paschoud, Serge; Jond, Lionel; Guerrera, Diego; Citi, Sandra

    2014-01-01

    PLEKHA7 is a recently identified protein of the epithelial zonula adhaerens (ZA), and is part of a protein complex that stabilizes the ZA, by linking it to microtubules. Since the ZA is important in the assembly and disassembly of tight junctions (TJ), we asked whether PLEKHA7 is involved in modulating epithelial TJ barrier function. We generated clonal MDCK cell lines in which one of four different constructs of PLEKHA7 was inducibly expressed. All constructs were localized at junctions, but constructs lacking the C-terminal region were also distributed diffusely in the cytoplasm. Inducible expression of PLEKHA7 constructs did not affect the expression and localization of TJ proteins, the steady-state value of transepithelial resistance (TER), the development of TER during the calcium switch, and the flux of large molecules across confluent monolayers. In contrast, expression of three out of four constructs resulted both in enhanced recruitment of E-cadherin and associated proteins at the apical ZA and at lateral puncta adherentia (PA), a decreased TER at 18 h after assembly at normal calcium, and an attenuation in the fall in TER after extracellular calcium removal. This latter effect was inhibited when cells were treated with nocodazole. Immunoprecipitation analysis showed that PLEKHA7 forms a complex with the cytoplasmic TJ proteins ZO-1 and cingulin, and this association does not depend on the integrity of microtubules. These results suggest that PLEKHA7 modulates the dynamics of assembly and disassembly of the TJ barrier, through E-cadherin protein complex- and microtubule-dependent mechanisms. PMID:24843844

  11. Central role of the gut epithelial barrier in the pathogenesis of chronic intestinal inflammation: lessons learned from animal models and human genetics.

    PubMed

    Pastorelli, Luca; De Salvo, Carlo; Mercado, Joseph R; Vecchi, Maurizio; Pizarro, Theresa T

    2013-01-01

    The gut mucosa is constantly challenged by a bombardment of foreign antigens and environmental microorganisms. As such, the precise regulation of the intestinal barrier allows the maintenance of mucosal immune homeostasis and prevents the onset of uncontrolled inflammation. In support of this concept, emerging evidence points to defects in components of the epithelial barrier as etiologic factors in the pathogenesis of inflammatory bowel diseases (IBDs). In fact, the integrity of the intestinal barrier relies on different elements, including robust innate immune responses, epithelial paracellular permeability, epithelial cell integrity, as well as the production of mucus. The purpose of this review is to systematically evaluate how alterations in the aforementioned epithelial components can lead to the disruption of intestinal immune homeostasis, and subsequent inflammation. In this regard, the wealth of data from mouse models of intestinal inflammation and human genetics are pivotal in understanding pathogenic pathways, for example, that are initiated from the specific loss of function of a single protein leading to the onset of intestinal disease. On the other hand, several recently proposed therapeutic approaches to treat human IBD are targeted at enhancing different elements of gut barrier function, further supporting a primary role of the epithelium in the pathogenesis of chronic intestinal inflammation and emphasizing the importance of maintaining a healthy and effective intestinal barrier. PMID:24062746

  12. Effects of phenol on barrier function of a human intestinal epithelial cell line correlate with altered tight junction protein localization.

    PubMed

    McCall, Ingrid C; Betanzos, Abigail; Weber, Dominique A; Nava, Porfirio; Miller, Gary W; Parkos, Charles A

    2009-11-15

    Phenol contamination of soil and water has raised concerns among people living near phenol-producing factories and hazardous waste sites containing the chemical. Phenol, particularly in high concentrations, is an irritating and corrosive substance, making mucosal membranes targets of toxicity in humans. However, few data on the effects of phenol after oral exposure exist. We used an in vitro model employing human intestinal epithelial cells (SK-CO15) cultured on permeable supports to examine effects of phenol on epithelial barrier function. We hypothesized that phenol disrupts epithelial barrier by altering tight junction (TJ) protein expression. The dose-response effect of phenol on epithelial barrier function was determined using transepithelial electrical resistance (TER) and FITC-dextran permeability measurements. We studied phenol-induced changes in cell morphology and expression of several tight junction proteins by immunofluorescence and Western blot analysis. Effects on cell viability were assessed by MTT, Trypan blue, propidium iodide and TUNEL staining. Exposure to phenol resulted in decreased TER and increased paracellular flux of FITC-dextran in a dose-dependent manner. Delocalization of claudin-1 and ZO-1 from TJs to cytosol correlated with the observed increase in permeability after phenol treatment. Additionally, the decrease in TER correlated with changes in the distribution of a membrane raft marker, suggesting phenol-mediated effects on membrane fluidity. Such observations were independent of effects of phenol on cell viability as enhanced permeability occurred at doses of phenol that did not cause cell death. Overall, these findings suggest that phenol may affect transiently the lipid bilayer of the cell membrane, thus destabilizing TJ-containing microdomains. PMID:19679145

  13. 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.

  14. 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

  15. Enhanced intranasal delivery of mRNA vaccine by overcoming the nasal epithelial barrier via intra- and paracellular pathways.

    PubMed

    Li, Man; Zhao, Mengnan; Fu, Yao; Li, You; Gong, Tao; Zhang, Zhirong; Sun, Xun

    2016-04-28

    Facing the threat of highly variable virus infection, versatile vaccination systems are urgently needed. Intranasal mRNA vaccination provides a flexible and convenient approach. However, the nasal epithelium remains a major biological barrier to deliver antigens to nasal associated lymphoid tissue (NALT). To address this issue, a potent polymer-based intranasal mRNA vaccination system for HIV-1 treatment was synthesized using cationic cyclodextrin-polyethylenimine 2k conjugate (CP 2k) complexed with anionic mRNA encoding HIV gp120. The delivery vehicle containing CP 2k and mRNA overcame the epithelial barrier by reversibly opening the tight junctions, enhanced the paracellular delivery of mRNA and consequently minimized absorption of toxins in the nasal cavity. Together with the excellent intracellular delivery and prolonged nasal residence time, strong system and mucosal anti-HIV immune responses as well as cytokine productions were achieved with a balanced Th1/Th2/Th17 type. Our study provided the first proof of evidence that cationic polymers can be used as safe and potent intranasal mRNA vaccine carriers to overcome the nasal epithelial barrier. The safe and versatile polymeric delivery system represents a promising vaccination platform for infectious diseases. PMID:26941035

  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. PMID:26743480

  17. 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

  18. Eosinophilic esophagitis–linked calpain 14 is an IL-13–induced protease that mediates esophageal epithelial barrier impairment

    PubMed Central

    Davis, Benjamin P.; Stucke, Emily M.; Khorki, M. Eyad; Litosh, Vladislav A.; Rymer, Jeffrey K.; Rochman, Mark; Travers, Jared; Kottyan, Leah C.; Rothenberg, Marc E.

    2016-01-01

    We recently identified a genome-wide genetic association of eosinophilic esophagitis (EoE) at 2p23 spanning the calpain 14 (CAPN14) gene, yet the causal mechanism has not been elucidated. We now show that recombinant CAPN14 cleaves a calpain-specific substrate and is inhibited by 4 classical calpain inhibitors: MDL-28170, acetyl-calpastatin, E-64, and PD151746. CAPN14 is specifically induced (>100-fold) in esophageal epithelium after IL-13 treatment. Epithelial cells overexpressing CAPN14 display impaired epithelial architecture, characterized by acantholysis, epidermal clefting, and epidermolysis. CAPN14 overexpression impairs epithelial barrier function, as demonstrated by decreased transepithelial resistance (2.1-fold) and increased FITC-dextran flux (2.6-fold). Epithelium with gene-silenced CAPN14 demonstrates increased dilated intercellular spaces (5.5-fold) and less organized basal cell layering (1.5-fold) following IL-13 treatment. Finally, CAPN14 overexpression results in loss of desmoglein 1 (DSG1) expression, whereas the IL-13–induced loss of DSG1 is normalized by CAPN14 gene silencing. Importantly, these findings were specific to CAPN14, as they were not observed with modulation of CAPN1 expression. These results, along with the potent induction of CAPN14 by IL-13 and genetic linkage of EoE to the CAPN14 gene locus, demonstrate a molecular and cellular pathway that contributes to T helper type 2 responses in mucosal epithelium. PMID:27158675

  19. 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

  20. High glucose induces dysfunction of airway epithelial barrier through down-regulation of connexin 43.

    PubMed

    Yu, Hongmei; Yang, Juan; Zhou, Xiangdong; Xiao, Qian; Lü, Yang; Xia, Li

    2016-03-01

    The airway epithelium is a barrier to the inhaled antigens and pathogens. Connexin 43 (Cx43) has been found to play critical role in maintaining the function of airway epithelial barrier and be involved in the pathogenesis of the diabetic retinal vasculature, diabetes nephropathy and diabetes skin. Hyperglycemia has been shown to be an independent risk factor for respiratory infections. We hypothesize that the down-regulation of Cx43 induced by HG alters the expression of tight junctions (zonula occludens-1 (ZO-1) and occludin) and contributes to dysfunction of airway epithelial barrier, and Cx43 plays a critical role in the process in human airway epithelial cells (16 HBE). We show that high glucose (HG) decreased the expression of ZO-1 and occludin, disassociated interaction between Cx43 and tight junctions, and then increased airway epithelial transepithelial electrical resistance (TER) and permeability by down-regulation of Cx43 in human airway epithelial cells. These observations demonstrate an important role for Cx43 in regulating HG-induced dysfunction of airway epithelial barrier. These findings may bring new insights into the molecular pathogenesis of pulmonary infection related to diabetes mellitus and lead to novel therapeutic intervention for the dysfunction of airway epithelial barrier in chronic inflammatory airway diseases. PMID:26902399

  1. 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

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

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

  3. 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

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

    PubMed

    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

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

    PubMed

    Murphy, W A; Lumsden, R D

    1984-08-01

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

  6. Interleukin-13 promotes expression of Alix to compromise renal tubular epithelial barrier function.

    PubMed

    Xu, Chen; Sun, Guangdong; Yang, Jie; Sun, Qianmei; Tong, Zhaohui

    2015-05-01

    The epithelial barrier dysfunction plays a critical role in a number of kidney diseases. The mechanism is unclear. Alix is a protein involving in protein degradation in epithelial cells. This study aims to investigate that interleukin (IL)-13 inhibits Alix to compromise the kidney epithelial barrier function. In this study, the murine collecting duct cell line (M-1) was cultured in Transwell inserts to investigate the significance of Alix in compromising the epithelial barrier functions. T cell (Teff cells) proliferation assay was employed to assess the antigenicity of ovalbumin (OVA) that was transported across the M-1 monolayer barrier. The results showed that M-1 cells express Alix. Exposure to interleukin (IL)-13 markedly decreased the expression of Alix in M-1 cells, which compromised the M-1 monolayer barrier functions by showing the increases in the permeability to OVA. Over-expression of Alix abolished the IL-13-induced M-1 monolayer barrier dysfunction. Knockdown of Alix significantly increased M-1 monolayer permeability. The OVA collected from the Transwell basal chambers induced the OVA-specific T cell proliferation. We conclude that IL-13 compromises M-1 epithelial barrier functions via inhibiting Alix expression. PMID:25597757

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

    PubMed

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

    2015-07-10

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

  8. 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

  9. Interaction of polyacrylates with porcine pepsin and the gastric mucus barrier: a mechanism for mucosal protection.

    PubMed

    Foster, S N; Pearson, J P; Hutton, D A; Allen, A; Dettmar, P W

    1994-12-01

    1. The mechanism of interaction of the polyacrylates, carbopols with the mucus barrier in vivo has been investigated in vitro. 2. Carbopol caused a dramatic increase in the viscosity of porcine gastric mucin solutions that was up to 19-fold greater than that of the sum of the individual polymers. 3. The mucin-carbopol interaction was stable after an initial 30 min period for up to 36 h at 25 degrees C or 37 degrees C. It was reduced by increasing the temperature from 20 degrees C to 45 degrees C, was unaffected by pH and ionic strength, but was enhanced by Ca2+. 4. The magnitude of the interaction between mucin and carbopol depended on the polymeric structure of the mucin and the molecular size and level of cross-linking of the carbopol. 5. The interactions were reversible and increased with increasing carbopol and mucin concentration. The dramatic increase in viscosity can be explained in terms of space filling by the mucin molecules leading to predominantly carbopol-carbopol interactions. 6. Carbopol 934P inhibits pepsin hydrolysis and therefore has potential as a mucosal protective agent in vivo. PMID:7874865

  10. CRF Induces Intestinal Epithelial Barrier Injury via the Release of Mast Cell Proteases and TNF-α

    PubMed Central

    Overman, Elizabeth L.; Rivier, Jean E.; Moeser, Adam J.

    2012-01-01

    Background and Aims Psychological stress is a predisposing factor in the onset and exacerbation of important gastrointestinal diseases including irritable bowel syndrome (IBS) and the inflammatory bowel diseases (IBD). The pathophysiology of stress-induced intestinal disturbances is known to be mediated by corticotropin releasing factor (CRF) but the precise signaling pathways remain poorly understood. Utilizing a porcine ex vivo intestinal model, the aim of this study was to investigate the mechanisms by which CRF mediates intestinal epithelial barrier disturbances. Methodology Ileum was harvested from 6–8 week-old pigs, mounted on Ussing Chambers, and exposed to CRF in the presence or absence of various pharmacologic inhibitors of CRF-mediated signaling pathways. Mucosal-to-serosal flux of 4 kDa-FITC dextran (FD4) and transepithelial electrical resistance (TER) were recorded as indices of intestinal epithelial barrier function. Results Exposure of porcine ileum to 0.05–0.5 µM CRF increased (p<0.05) paracellular flux compared with vehicle controls. CRF treatment had no deleterious effects on ileal TER. The effects of CRF on FD4 flux were inhibited with pre-treatment of tissue with the non-selective CRF1/2 receptor antagonist Astressin B and the mast cell stabilizer sodium cromolyn (10−4 M). Furthermore, anti-TNF-α neutralizing antibody (p<0.01), protease inhibitors (p<0.01) and the neural blocker tetrodotoxin (TTX) inhibited CRF-mediated intestinal barrier dysfunction. Conclusion These data demonstrate that CRF triggers increases in intestinal paracellular permeability via mast cell dependent release of TNF-α and proteases. Furthermore, CRF-mast cell signaling pathways and increases in intestinal permeability require critical input from the enteric nervous system. Therefore, blocking the deleterious effects of CRF may address the enteric signaling of mast cell degranulation, TNFα release, and protease secretion, hallmarks of IBS and IBD. PMID:22768175

  11. 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

  12. Effect of Polysaccharides from Acanthopanax senticosus on Intestinal Mucosal Barrier of Escherichia coli Lipopolysaccharide Challenged Mice.

    PubMed

    Han, Jie; Xu, Yunhe; Yang, Di; Yu, Ning; Bai, Zishan; Bian, Lianquan

    2016-01-01

    To investigate the role of polysaccharide from Acanthopanax senticosus (ASPS) in preventing lipopolysaccharide (LPS)-induced intestinal injury, 18 mice (at 5 wk of age) were assigned to three groups with 6 replicates of one mouse each. Mice were administrated by oral gavage with or without ASPS (300 mg/kg body weight) for 14 days and were injected with saline or LPS at 15 days. Intestinal samples were collected at 4 h post-challenge. The results showed that ASPS ameliorated LPS-induced deterioration of digestive ability of LPS-challenged mice, indicated by an increase in intestinal lactase activity (45%, p<0.05), and the intestinal morphology, as proved by improved villus height (20.84%, p<0.05) and villus height:crypt depth ratio (42%, p<0.05), and lower crypt depth in jejunum (15.55%, p<0.05), as well as enhanced intestinal tight junction proteins expression involving occludin-1 (71.43%, p<0.05). ASPS also prevented intestinal inflammation response, supported by decrease in intestinal inflammatory mediators including tumor necrosis factor α (22.28%, p<0.05) and heat shock protein (HSP70) (77.42%, p<0.05). In addition, intestinal mucus layers were also improved by ASPS, as indicated by the increase in number of goblet cells (24.89%, p<0.05) and intestinal trefoil peptide (17.75%, p<0.05). Finally, ASPS facilitated mRNA expression of epidermal growth factor (100%, p<0.05) and its receptor (200%, p<0.05) gene. These results indicate that ASPS can prevent intestinal mucosal barrier injury under inflammatory conditions, which may be associated with up-regulating gene mRNA expression of epidermal growth factor and its receptor. PMID:26732337

  13. 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

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

    PubMed Central

    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

  15. Effect of Polysaccharides from Acanthopanax senticosus on Intestinal Mucosal Barrier of Escherichia coli Lipopolysaccharide Challenged Mice

    PubMed Central

    Han, Jie; Xu, Yunhe; Yang, Di; Yu, Ning; Bai, Zishan; Bian, Lianquan

    2016-01-01

    To investigate the role of polysaccharide from Acanthopanax senticosus (ASPS) in preventing lipopolysaccharide (LPS)-induced intestinal injury, 18 mice (at 5 wk of age) were assigned to three groups with 6 replicates of one mouse each. Mice were administrated by oral gavage with or without ASPS (300 mg/kg body weight) for 14 days and were injected with saline or LPS at 15 days. Intestinal samples were collected at 4 h post-challenge. The results showed that ASPS ameliorated LPS-induced deterioration of digestive ability of LPS-challenged mice, indicated by an increase in intestinal lactase activity (45%, p<0.05), and the intestinal morphology, as proved by improved villus height (20.84%, p<0.05) and villus height:crypt depth ratio (42%, p<0.05), and lower crypt depth in jejunum (15.55%, p<0.05), as well as enhanced intestinal tight junction proteins expression involving occludin-1 (71.43%, p<0.05). ASPS also prevented intestinal inflammation response, supported by decrease in intestinal inflammatory mediators including tumor necrosis factor α (22.28%, p<0.05) and heat shock protein (HSP70) (77.42%, p<0.05). In addition, intestinal mucus layers were also improved by ASPS, as indicated by the increase in number of goblet cells (24.89%, p<0.05) and intestinal trefoil peptide (17.75%, p<0.05). Finally, ASPS facilitated mRNA expression of epidermal growth factor (100%, p<0.05) and its receptor (200%, p<0.05) gene. These results indicate that ASPS can prevent intestinal mucosal barrier injury under inflammatory conditions, which may be associated with up-regulating gene mRNA expression of epidermal growth factor and its receptor. PMID:26732337

  16. 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.

  17. 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.

  18. 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

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

    PubMed Central

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

    2014-01-01

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

  20. Interleukin-4 and interleukin-13 cause barrier dysfunction in human airway epithelial cells.

    PubMed

    Saatian, Bahman; Rezaee, Fariba; Desando, Samantha; Emo, Jason; Chapman, Tim; Knowlden, Sara; Georas, Steve N

    2013-04-01

    Emerging evidence indicates that airway epithelial barrier function is compromised in asthma, a disease characterized by Th2-skewed immune response against inhaled allergens, but the mechanisms involved are not well understood. The purpose of this study was to investigate the effects of Th2-type cytokines on airway epithelial barrier function. 16HBE14o- human bronchial epithelial cells monolayers were grown on collagen coated Transwell inserts. The basolateral or apical surfaces of airway epithelia were exposed to human interleukin-4 (IL-4), IL-13, IL-25, IL-33, thymic stromal lymphopoietin (TSLP) alone or in combination at various concentrations and time points. We analyzed epithelial apical junctional complex (AJC) function by measuring transepithelial electrical resistance (TEER) and permeability to FITC-conjugated dextran over time. We analyzed AJC structure using immunofluorescence with antibodies directed against key junctional components including occludin, ZO-1, β-catenin and E-cadherin. Transepithelial resistance was significantly decreased after both basolateral and apical exposure to IL-4. Permeability to 3 kDa dextran was also increased in IL-4-exposed cells. Similar results were obtained with IL-13, but none of the innate type 2 cytokines examined (TSLP, IL-25 or IL-33) significantly affected barrier function. IL-4 and IL-13-induced barrier dysfunction was accompanied by reduced expression of membrane AJC components but not by induction of claudin- 2. Enhanced permeability caused by IL-4 was not affected by wortmannin, an inhibitor of PI3 kinase signaling, but was attenuated by a broad spectrum inhibitor of janus associated kinases. Our study indicates that IL-4 and IL-13 have disruptive effect on airway epithelial barrier function. Th2-cytokine induced epithelial barrier dysfunction may contribute to airway inflammation in allergic asthma. PMID:24665390

  1. A Key Claudin Extracellular Loop Domain is Critical for Epithelial Barrier Integrity

    PubMed Central

    Mrsny, Randall J.; Brown, G. Thomas; Gerner-Smidt, Kirsten; Buret, Andre G.; Meddings, Jon B.; Quan, Clifford; Koval, Michael; Nusrat, Asma

    2008-01-01

    Intercellular tight junctions (TJs) regulate epithelial barrier properties. Claudins are major structural constituents of TJs and belong to a large family of tetra-spanning membrane proteins that have two predicted extracellular loops (ELs). Given that claudin-1 is widely expressed in epithelia, we further defined the role of its EL domains in determining TJ function. The effects of several claudin-1 EL mimetic peptides on epithelial barrier structure and function were examined. Incubation of model human intestinal epithelial cells with a 27-amino acid peptide corresponding to a portion of the first EL domain (Cldn-153–80) reversibly interfered with epithelial barrier function by inducing the rearrangement of key TJ proteins: occludin, claudin-1, junctional adhesion molecule-A, and zonula occludens-1. Cldn-153–80 associated with both claudin-1 and occludin, suggesting both the direct interference with the ability of these proteins to assemble into functional TJs and their close interaction under physiological conditions. These effects were specific for Cldn-153–80, because peptides corresponding to other claudin-1 EL domains failed to influence TJ function. Furthermore, the oral administration of Cldn-153–80 to rats increased paracellular gastric permeability. Thus, the identification of a critical claudin-1 EL motif, Cldn-153–80, capable of regulating TJ structure and function, offers a useful adjunct to treatments that require drug delivery across an epithelial barrier. PMID:18349130

  2. 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...

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

    PubMed

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

    2014-01-01

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

  4. 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

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

    PubMed Central

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

    2000-01-01

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

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

    PubMed Central

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

    2015-01-01

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

  7. 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

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

    PubMed

    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

  9. 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

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

    PubMed Central

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

    2010-01-01

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

  11. Targeting Mitochondria-Derived Reactive Oxygen Species to Reduce Epithelial Barrier Dysfunction and Colitis

    PubMed Central

    Wang, Arthur; Keita, Åsa V.; Phan, Van; McKay, Catherine M.; Schoultz, Ida; Lee, Joshua; Murphy, Michael P.; Fernando, Maria; Ronaghan, Natalie; Balce, Dale; Yates, Robin; Dicay, Michael; Beck, Paul L.; MacNaughton, Wallace K.; Söderholm, Johan D.; McKay, Derek M.

    2015-01-01

    Epithelial permeability is often increased in inflammatory bowel diseases. We hypothesized that perturbed mitochondrial function would cause barrier dysfunction and hence epithelial mitochondria could be targeted to treat intestinal inflammation. Mitochondrial dysfunction was induced in human colon-derived epithelial cell lines or colonic biopsy specimens using dinitrophenol, and barrier function was assessed by transepithelial flux of Escherichia coli with or without mitochondria-targeted antioxidant (MTA) cotreatment. The impact of mitochondria-targeted antioxidants on gut permeability and dextran sodium sulfate (DSS)–induced colitis in mice was tested. Mitochondrial superoxide evoked by dinitrophenol elicited significant internalization and translocation of E. coli across epithelia and control colonic biopsy specimens, which was more striking in Crohn’s disease biopsy specimens; the mitochondria-targeted antioxidant, MitoTEMPO, inhibited these barrier defects. Increased gut permeability and reduced epithelial mitochondrial voltage-dependent anion channel expression were observed 3 days after DSS. These changes and the severity of DSS-colitis were reduced by MitoTEMPO treatment. In vitro DSS-stimulated IL-8 production by epithelia was reduced by MitoTEMPO. Metabolic stress evokes significant penetration of commensal bacteria across the epithelium, which is mediated by mitochondria-derived superoxide acting as a signaling, not a cytotoxic, molecule. MitoTEMPO inhibited this barrier dysfunction and suppressed colitis in DSS-colitis, likely via enhancing barrier function and inhibiting proinflammatory cytokine production. These novel findings support consideration of MTAs in the maintenance of epithelial barrier function and the management of inflammatory bowel diseases. PMID:25034594

  12. Hsa-miRNA-31 regulates epithelial cell barrier function by inhibiting TNFSF15 expression.

    PubMed

    Nan, X; Qin, S; Yuan, Z; Li, Y; Zhang, J; Li, C; Tan, X; Yan, Y

    2016-01-01

    Ulcerative colitis (UC) is characterized by epithelial barrier disruption and alterations in immune regulation but with the etiology unknown. MicroRNA-31 is the most consistent differentially expressed miRNA in ulcerative colitis tissue. The aim of this project is to study the important roles of miRNA-31 in regulation of intestinal epithelial barrier function. We found that expression of miRNA-31 is proportional to the proliferation of Caco2-BBE cells and overexpression of miRNA-31 can increase its trans-epithelial resistance (TER) by decreasing the transepithelial permeability. miRNA-31 can directly bind to the 3-UTR of TNFSF15, thereafter negatively regulating its expression in Caco2-BBE cells. BrdU and TUNEL analysis demonstrated that transfection of miRNA-31 stimulates proliferation or apoptosis-resistance. Taken together, these results revealed a novel mecha-nism underlying the regulation of epithelial barrier function by miRNA-31 during its regulation on proliferation of epithelial cells. PMID:27188743

  13. 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

  14. 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…

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

    PubMed Central

    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

  16. 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. PMID:26206859

  17. 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

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

    PubMed

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

    2015-05-15

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

  19. Barrier responses of human bronchial epithelial cells to grass pollen exposure.

    PubMed

    Blume, Cornelia; Swindle, Emily J; Dennison, Patrick; Jayasekera, Nivenka P; Dudley, Sarah; Monk, Phillip; Behrendt, Heidrun; Schmidt-Weber, Carsten B; Holgate, Stephen T; Howarth, Peter H; Traidl-Hoffmann, Claudia; Davies, Donna E

    2013-07-01

    The airway epithelium forms a physical, chemical and immunological barrier against inhaled environmental substances. In asthma, these barrier properties are thought to be abnormal. In this study, we analysed the effect of grass pollen on the physical and immunological barrier properties of differentiated human primary bronchial epithelial cells. Following exposure to Timothy grass (Phleum pratense) pollen extract, the integrity of the physical barrier was not impaired as monitored by measuring the transepithelial resistance and immunofluorescence staining of tight junction proteins. In contrast, pollen exposure affected the immunological barrier properties by modulating vectorial mediator release. CXC chemokine ligand (CXCL)8/interleukin (IL)-8 showed the greatest increase in response to pollen exposure with preferential release to the apical compartment. Inhibition of the extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase pathways selectively blocked apical CXCL8/IL-8 release via a post-transcriptional mechanism. Apical release of CC chemokine ligand (CCL)20/macrophage inflammatory protein-3α, CCL22/monocyte-derived chemokine and tumour necrosis factor-α was significantly increased only in severe asthma cultures, while CCL11/eotaxin-1 and CXCL10/interferon-γ-induced protein-10 were reduced in nonasthmatic cultures. The bronchial epithelial barrier modulates polarised release of mediators in response to pollen without direct effects on its physical barrier properties. The differential response of cells from normal and asthmatic donors suggests the potential for the bronchial epithelium to promote immune dysfunction in asthma. PMID:23143548

  20. 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

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

    PubMed

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

    2015-02-01

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

  2. The Na+/H+ exchange inhibitor HOE642 prevents stress-induced epithelial barrier dysfunction.

    PubMed

    Nowak, Peter; Blaheta, Roman; Schuller, Alina; Cinatl, Jindrich; Wimmer-Greinecker, Gerhard; Moritz, Anton; Scholz, Martin

    2004-08-01

    Recently, evidence has been obtained that the Na+/H+ exchange (NHE) inhibitor HOE642 may stabilize endothelial and epithelial barrier function in vivo. However, the underlying mechanisms are not known. Therefore, we studied the influence of HOE642 on the barrier function of the epithelial cell line CaCo2. The phorbolester phorbol 12-myristate 13-acetate (PMA) was used to induce hyperpermeability of the epithelial layer which was indirectly determined by measuring the transepithelial electrical resistance (TER). Confocal laser scan microscopy (LSM) served to analyze the intracellular localization of adherens and tight junction molecules. In five independent experiments we found that HOE642 increased TER in non-treated CaCo2 cells (control: 350 +/- 28 Omega/cm2; HOE642: 444 +/- 53 Omega/cm2) and prevented PMA-induced barrier dysfunction (PMA: 33 +/- 12 Omega/cm2; PMA plus HOE642: 496 +/- 47 Omega/cm2). LSM showed that HOE642 prevented the PMA-induced disassociation of the zonula adherens molecule beta-catenin from the cell membrane and the decreased expression of the zonula occludens molecule ZO-1. From our data we conclude that HOE642 may prevent stress-induced epithelial dysfunction by stabilization of cell membrane-associated junction molecules. PMID:15254761

  3. Fluticasone Induces Epithelial Injury and Alters Barrier Function in Normal Subjects

    PubMed Central

    MacRedmond, Ruth E.; Singhera, Gurpreet K.; Wadsworth, Samuel J.; Attridge, Susan; Bahzad, Mohammed; Williams, Kristy; Coxson, Harvey O.; White, Steven R.; Dorscheid, Delbert R.

    2014-01-01

    Objective The airway epithelium has a number of roles pivotal to the pathogenesis of asthma, including provision of a physical and immune barrier to the inhaled environment. Dysregulated injury and repair responses in asthma result in loss of airway epithelial integrity. Inhaled corticosteroids are a corner stone of asthma treatment. While effective in controlling asthma symptoms, they fail to prevent airway remodeling. Direct cytopathic effects on the airway epithelium may contribute to this. Methods This study examined the effects of a 4-week treatment regimen of inhaled fluticasone 500 μg twice daily in healthy human subjects. Induced sputum was collected for cell counts and markers of inflammation. Barrier function was examined by diethylenetriaminepentacetic acid (DTPA) clearance measured by nuclear scintillation scan, and albumin concentration in induced sputum. Results Steroid exposure resulted in epithelial injury as measured by a significant increase in the number of airway epithelial cells in induced sputum. There was no change in airway inflammation by induced sputum inflammatory cell counts or cytokine levels. Epithelial shedding was associated with an increase in barrier function, as measured by both a decrease in DTPA clearance and decreased albumin in induced sputum. This likely reflects the normal repair response. Conclusion Inhaled corticosteroids cause injury to normal airway epithelium. These effects warrant further evaluation in asthma, where the dysregulated repair response may contribute to airway remodeling. PMID:25324978

  4. 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

  5. 1,25-Dihydroxyvitamin D3 prevents toluene diisocyanate-induced airway epithelial barrier disruption.

    PubMed

    Li, Wenjia; Dong, Hangming; Zhao, Haijin; Song, Jiafu; Tang, Haixiong; Yao, Lihong; Liu, Laiyu; Tong, Wancheng; Zou, Mengchen; Zou, Fei; Cai, Shaoxi

    2015-07-01

    The loss of airway epithelial integrity contributes significantly to asthma pathogenesis. Evidence suggests that vitamin D plays an important role in the prevention and treatment of asthma. However, its role in airway epithelial barrier function remains uncertain. We have previously demonstrated impaired epithelial junctions in a model of toluene diisocyanate (TDI)-induced asthma. In the present study, we hypothesized that 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] may prevent TDI-induced epithelial barrier disruption. Male BALB/c mice were dermally sensitized and then challenged with TDI. The mice were then administered 1,25(OH)2D3 intraperitoneally prior to challenge with TDI. For in vitro experiments, 16HBE bronchial epithelial cells were cultured and stimulated with TDI-human serum albumin (HSA). The results revealed that the mice treated with 1,25(OH)2D3 displayed decreased airway hyperresponsiveness (AHR), suppressed neutrophil and eosinophil infiltration into the airways, as well as an increased E-cadherin and zonula occludens-1 (ZO-1) expression at the cell-cell contact sites. In vitro, exposure of the cells to TDI-HSA induced a rapid decline in transepithelial electrical resistance (TER) and an increase in cell permeability, followed by a decrease in occludin expression and the redistribution of E-cadherin, accompanied by a significant upregulation in the levels of phosphorylated extracellular signal-regulated kinase (ERK)1/2. These effects were all partly reversed by treatment with either 1,25(OH)2D3 or an ERK1/2 inhibitor. In conclusion, the findings of our study demonstrate that 1,25(OH)2D3 prevents TDI-induced epithelial barrier disruption, and that the ERK1/2 pathway may play a role in this process. PMID:25998793

  6. 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

  7. Epithelial Cell Regulation of Allergic Diseases.

    PubMed

    Gour, Naina; Lajoie, Stephane

    2016-09-01

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

  8. 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

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

    PubMed

    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

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

    PubMed Central

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

    2014-01-01

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

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

    PubMed

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

    2014-01-01

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

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

    PubMed Central

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

    2011-01-01

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

  13. Rapid disruption of intestinal epithelial tight junction and barrier dysfunction by ionizing radiation in mouse colon in vivo: protection by N-acetyl-l-cysteine.

    PubMed

    Shukla, Pradeep K; Gangwar, Ruchika; Manda, Bhargavi; Meena, Avtar S; Yadav, Nikki; Szabo, Erzsebet; Balogh, Andrea; Lee, Sue Chin; Tigyi, Gabor; Rao, RadhaKrishna

    2016-05-01

    The goals of this study were to evaluate the effects of ionizing radiation on apical junctions in colonic epithelium and mucosal barrier function in mice in vivo. Adult mice were subjected to total body irradiation (4 Gy) with or without N-acetyl-l-cysteine (NAC) feeding for 5 days before irradiation. At 2-24 h postirradiation, the integrity of colonic epithelial tight junctions (TJ), adherens junctions (AJ), and the actin cytoskeleton was assessed by immunofluorescence microscopy and immunoblot analysis of detergent-insoluble fractions for TJ and AJ proteins. The barrier function was evaluated by measuring vascular-to-luminal flux of fluorescein isothiocyanate (FITC)-inulin in vivo and luminal-to-mucosal flux in vitro. Oxidative stress was evaluated by measuring protein thiol oxidation. Confocal microscopy showed that radiation caused redistribution of occludin, zona occludens-1, claudin-3, E-cadherin, and β-catenin, as well as the actin cytoskeleton as early as 2 h postirradiation, and this effect was sustained for at least 24 h. Feeding NAC before irradiation blocked radiation-induced disruption of TJ, AJ, and the actin cytoskeleton. Radiation increased mucosal permeability to inulin in colon, which was blocked by NAC feeding. The level of reduced-protein thiols in colon was depleted by radiation with a concomitant increase in the level of oxidized-protein thiol. NAC feeding blocked the radiation-induced protein thiol oxidation. These data demonstrate that radiation rapidly disrupts TJ, AJ, and the actin cytoskeleton by an oxidative stress-dependent mechanism that can be prevented by NAC feeding. PMID:26822914

  14. Abrogation of IFN-γ mediated epithelial barrier disruption by serine protease inhibition

    PubMed Central

    Willemsen, LEM; Hoetjes, JP; Van Deventer, SJH; Van Tol, EAF

    2005-01-01

    The intestinal barrier function is often impaired in a variety of diseases including chronic inflammatory bowel disease. Increased intestinal permeability during episodes of active disease correlates with destruction or rearrangement of the tight junction protein complex. IFN-γ has been widely studied for its effect on barrier function and tight junction structures but its mode of action remains unclear. Since the claudin family of tight junction proteins is proposed to be involved in barrier maintenance we studied the effect of IFN-γ on claudin expression in relation to epithelial barrier function. Cycloheximide and protease inhibitors were used to study mechanisms of IFN-γ mediated barrier disruption. Intestinal epithelial cells were exposed to IFN-γ and permeability was evaluated by horse radish peroxidase (HRP) and 4 kD FITC-dextran fluxes. Occludin and claudin-1, -2, -3, and -4 tight junction protein expression was determined by Western blotting. Occludin and claudin-2 protein expression was dramatically reduced after IFN-γ exposure, which correlated with increased permeability for HRP and FITC-dextran. Interestingly, cleavage of claudin-2 was observed after incubation with IFN-γ. Serine protease inhibitor AEBSF completely abrogated IFN-γ mediated barrier disruption which was associated with preservation of claudin-2 expression. Moreover, IFN-γ induced loss of barrier integrity was found to affect claudin-2 and occludin expression through different mechanisms. Since inhibition of serine protease activity abrogates IFN-γ mediated barrier disruption this may be an important target for therapeutic intervention. PMID:16232214

  15. The Bacteroides fragilis toxin fragilysin disrupts the paracellular barrier of epithelial cells.

    PubMed Central

    Obiso, R J; Azghani, A O; Wilkins, T D

    1997-01-01

    Bacteroides fragilis is a member of the normal colonic microflora of most mammals and is the most commonly isolated anaerobe from human clinical specimens. Some strains produce a toxin (fragilysin, a zinc-metalloproteinase) implicated as a cause of diarrheal disease in farm animals and humans. Studies in our laboratory confirm that the proteolytic activity of this toxin is responsible for the fluid secretion and tissue damage observed in vivo. In this study, we investigated the effects of fragilysin on the paracellular barrier of epithelial cells. Researchers suggest that, since the toxin rapidly intoxicates HT-29 cells, it may be internalized. However, we could not prevent cell rounding by using inhibitors of receptor-mediated endocytosis, which indicates that the toxin may act outside the cell. Based on these observations, we studied the effects of the highly purified B. fragilis fragilysin on the barrier function of cultured epithelial cells. Fragilysin rapidly increased the permeability of the paracellular barrier of epithelial cells to ions (decrease in electrical resistance across monolayers) and to larger molecules (increase in mannitol flux across monolayers). We tested a human colon cell line and cell lines from the lung and the kidney; the human colon cell line was most sensitive, but all three were affected in the same manner. Our studies show that B. fragilis fragilysin alters the barrier function of the epithelial lining, possibly by degrading the tight junction proteins, such as ZO-1. The proteolytic activity is required to cause this effect. The toxin's action has been assumed to be limited to the intestine; however, our studies show that fragilysin could also contribute to the pathogenesis of B. fragilis in extraintestinal infections. PMID:9119484

  16. Mucosal immunoglobulins.

    PubMed

    Woof, Jenny M; Mestecky, Jiri

    2005-08-01

    Due to their vast surface area, the mucosal surfaces of the body represent a major site of potential attack by invading pathogens. The secretions that bathe mucosal surfaces contain significant levels of immunoglobulins (Igs), which play key roles in immune defense of these surfaces. IgA is the predominant antibody class in many external secretions and has many functional attributes, both direct and indirect, that serve to prevent infective agents such as bacteria and viruses from breaching the mucosal barrier. This review details current understanding of the structural and functional characteristics of IgA, including interaction with specific receptors (such as Fc(alpha)RI, Fc(alpha)/microR, and CD71) and presents examples of the means by which certain pathogens circumvent the protective properties of this important Ig. PMID:16048542

  17. 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

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

    PubMed

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

    2014-01-01

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

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

    PubMed Central

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

    2014-01-01

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

  20. The role of anthrolysin O in gut epithelial barrier disruption during Bacillus anthracis infection.

    PubMed

    Bishop, Brian L; Lodolce, James P; Kolodziej, Lauren E; Boone, David L; Tang, Wei Jen

    2010-04-01

    Gastrointestinal (GI) anthrax, caused by the bacterial infection of Bacillus anthracis, posts a significant bioterrorism threat by its relatively high mortality rate in humans. Different from inhalational anthrax by the route of infection, accumulating evidence indicates the bypass of vegetative bacteria across GI epithelium is required to initiate GI anthrax. Previously, we reported that purified anthrolysin O (ALO), instead of tripartite anthrax edema and lethal toxins, is capable of disrupting gut epithelial tight junctions and barrier function in cultured cells. Here, we show that ALO can disrupt intestinal tissue barrier function in an ex vivo mouse model. To explore the effects of ALO in a cell culture model of B. anthracis infection, we showed that anthrax bacteria can effectively reduce the monolayer integrity of human Caco-2 brush-border expressor (C2BBE) cells based on the reduced transepithelial resistance and the increased leakage of fluorescent dye. This disruption is likely caused by tight junction dysfunction observed by the reorganization of the tight junction protein occludin. Consequently, we observe significant passage of vegetative anthrax bacteria across C2BBE cells. This barrier disruption and bacterial crossover requires ALO since ALO-deficient B. anthracis strains fail to induce monolayer dysfunction and allow the passage of anthrax bacteria. Together these findings point to a pivotal role for ALO within the establishment of GI anthrax infection and the initial bypass of the epithelial barrier. PMID:20188700

  1. The Pseudomonas aeruginosa Type III Translocon Is Required for Biofilm Formation at the Epithelial Barrier

    PubMed Central

    Tran, Cindy S.; Rangel, Stephanie M.; Almblad, Henrik; Kierbel, Arlinet; Givskov, Michael; Tolker-Nielsen, Tim; Hauser, Alan R.; Engel, Joanne N.

    2014-01-01

    Clinical infections by Pseudomonas aeruginosa, a deadly Gram-negative, opportunistic pathogen of immunocompromised hosts, often involve the formation of antibiotic-resistant biofilms. Although biofilm formation has been extensively studied in vitro on glass or plastic surfaces, much less is known about biofilm formation at the epithelial barrier. We have previously shown that when added to the apical surface of polarized epithelial cells, P. aeruginosa rapidly forms cell-associated aggregates within 60 minutes of infection. By confocal microscopy we now show that cell-associated aggregates exhibit key characteristics of biofilms, including the presence of extracellular matrix and increased resistance to antibiotics compared to planktonic bacteria. Using isogenic mutants in the type III secretion system, we found that the translocon, but not the effectors themselves, were required for cell-associated aggregation on the surface of polarized epithelial cells and at early time points in a murine model of acute pneumonia. In contrast, the translocon was not required for aggregation on abiotic surfaces, suggesting a novel function for the type III secretion system during cell-associated aggregation. Supernatants from epithelial cells infected with wild-type bacteria or from cells treated with the pore-forming toxin streptolysin O could rescue aggregate formation in a type III secretion mutant, indicating that cell-associated aggregation requires one or more host cell factors. Our results suggest a previously unappreciated function for the type III translocon in the formation of P. aeruginosa biofilms at the epithelial barrier and demonstrate that biofilms may form at early time points of infection. PMID:25375398

  2. The Pseudomonas aeruginosa type III translocon is required for biofilm formation at the epithelial barrier.

    PubMed

    Tran, Cindy S; Rangel, Stephanie M; Almblad, Henrik; Kierbel, Arlinet; Givskov, Michael; Tolker-Nielsen, Tim; Hauser, Alan R; Engel, Joanne N

    2014-11-01

    Clinical infections by Pseudomonas aeruginosa, a deadly Gram-negative, opportunistic pathogen of immunocompromised hosts, often involve the formation of antibiotic-resistant biofilms. Although biofilm formation has been extensively studied in vitro on glass or plastic surfaces, much less is known about biofilm formation at the epithelial barrier. We have previously shown that when added to the apical surface of polarized epithelial cells, P. aeruginosa rapidly forms cell-associated aggregates within 60 minutes of infection. By confocal microscopy we now show that cell-associated aggregates exhibit key characteristics of biofilms, including the presence of extracellular matrix and increased resistance to antibiotics compared to planktonic bacteria. Using isogenic mutants in the type III secretion system, we found that the translocon, but not the effectors themselves, were required for cell-associated aggregation on the surface of polarized epithelial cells and at early time points in a murine model of acute pneumonia. In contrast, the translocon was not required for aggregation on abiotic surfaces, suggesting a novel function for the type III secretion system during cell-associated aggregation. Supernatants from epithelial cells infected with wild-type bacteria or from cells treated with the pore-forming toxin streptolysin O could rescue aggregate formation in a type III secretion mutant, indicating that cell-associated aggregation requires one or more host cell factors. Our results suggest a previously unappreciated function for the type III translocon in the formation of P. aeruginosa biofilms at the epithelial barrier and demonstrate that biofilms may form at early time points of infection. PMID:25375398

  3. 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. PMID:10496683

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

    PubMed Central

    Yu, X Y; Takahashi, N; Croxton, T L; Spannhake, E W

    1994-01-01

    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 3 hr 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 microM vitamin A (34%). Responses to 0.5 ppm or 0.8 ppm were inhibited by pretreatment of the cells with 0.1 microM 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. Images Figure 1. Figure 2. Figure 3. Figure 4. Figure 5. PMID:7713019

  5. [Protection of the mucosal barrier by nutritional strategies. What are the therapeutic options?].

    PubMed

    Lübke, H J

    2000-05-01

    The dysfunction of intestinal barrier allows the translocation of both endotoxin and whole bacterial organisms. It plays an important role in the development of multiple organ failure (MOF). The mucosa ia one component of this barrier. Trauma, atrophy and the "systemic inflammatory response syndrome" increase gastrointestinal permeability. These abnormalities may contribute to the pathophysiology of sepsis. Malnutrition per se compromises the gut's barrier function. Maintenance of gastrointestinal blood flow may be facilitated by (glutamine-enriched?) enteral diets. The most important conclusions of the majority of controlled trials support the concept of the very early enteral nutrition (within 24 hours after trauma): the outcome of seriously ill patients is improved, the rate of complications and infections is reduced. Gastrointestinal motility disorders may interfere with the initiation and tolerance of early enteral nutrition. They may be managed by prokinetic agents (cisapride, erythromycin) or by bypassing the stomach with a nasoenteric tube. PMID:10883361

  6. 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

  7. 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

  8. Ferritin polarization and iron transport across monolayer epithelial barriers in mammals

    PubMed Central

    Meyron-Holtz, Esther G.; Cohen, Lyora A.; Fahoum, Lulu; Haimovich, Yael; Lifshitz, Lena; Magid-Gold, Inbar; Stuemler, Tanja; Truman-Rosentsvit, Marianna

    2014-01-01

    Epithelial barriers are found in many tissues such as the intestine, kidney and brain where they separate the external environment from the body or a specific compartment from its periphery. Due to the tight junctions that connect epithelial barrier-cells (EBCs), the transport of compounds takes place nearly exclusively across the apical or basolateral membrane, the cell-body and the opposite membrane of the polarized EBC, and is regulated on numerous levels including barrier-specific adapted trafficking-machineries. Iron is an essential element but toxic at excess. Therefore, all iron-requiring organisms tightly regulate iron concentrations on systemic and cellular levels. In contrast to most cell types that control just their own iron homeostasis, EBCs also regulate homeostasis of the compartment they enclose or the body as a whole. Iron is transported across EBCs by specialized transporters such as the transferrin receptor and ferroportin. Recently, the iron storage protein ferritin was also attributed a role in the regulation of systemic iron homeostasis and we gathered evidence from the literature and original data that ferritin is polarized in EBC, suggesting also a role for ferritin in iron trafficking across EBCs. PMID:25202274

  9. 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

  10. 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

  11. 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.

  12. Cyclic stretch induces alveolar epithelial barrier dysfunction via calpain-mediated degradation of p120-catenin.

    PubMed

    Wang, Yuelan; Minshall, Richard D; Schwartz, David E; Hu, Guochang

    2011-08-01

    Lung hyperinflation is known to be an important contributing factor in the pathogenesis of ventilator-induced lung injury. Mechanical stretch causes epithelial barrier dysfunction and an increase in alveolar permeability, although the precise mechanisms have not been completely elucidated. p120-catenin is an adherens junction-associated protein that regulates cell-cell adhesion. In this study, we determined the role of p120-catenin in cyclic stretch-induced alveolar epithelial barrier dysfunction. Cultured alveolar epithelial cells (MLE-12) were subjected to uniform cyclic (0.5 Hz) biaxial stretch from 0 to 8 or 20% change in surface area for 0, 1, 2, or 4 h. At the end of the experiments, cells were lysed to determine p120-catenin expression by Western blot analysis. Immunofluorescence staining of p120-catenin and F-actin was performed to assess the integrity of monolayers and interepithelial gap formation. Compared with unstretched control cells, 20% stretch caused a significant loss in p120-catenin expression, which was coupled to interepithelial gap formation. p120-Catenin knockdown with small interfering RNA (siRNA) dose dependently increased stretch-induced gap formation, whereas overexpression of p120-catenin abolished stretch-induced gap formation. Furthermore, pharmacological calpain inhibition or depletion of calpain-1 with a specific siRNA prevented p120-catenin loss and subsequent stretch-induced gap formation. Our findings demonstrate that p120-catenin plays a critical protective role in cyclic stretch-induced alveolar barrier dysfunction, and, thus, maintenance of p120-catenin expression may be a novel therapeutic strategy for the prevention and treatment of ventilator-induced lung injury. PMID:21571907

  13. 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

  14. Induction of mucosal immunity through systemic immunization: Phantom or reality?

    PubMed

    Su, Fei; Patel, Girishchandra B; Hu, Songhua; Chen, Wangxue

    2016-04-01

    Generation of protective immunity at mucosal surfaces can greatly assist the host defense against pathogens which either cause disease at the mucosal epithelial barriers or enter the host through these surfaces. Although mucosal routes of immunization, such as intranasal and oral, are being intensely explored and appear promising for eliciting protective mucosal immunity in mammals, their application in clinical practice has been limited due to technical and safety related challenges. Most of the currently approved human vaccines are administered via systemic (such as intramuscular and subcutaneous) routes. Whereas these routes are acknowledged as being capable to elicit antigen-specific systemic humoral and cell-mediated immune responses, they are generally perceived as incapable of generating IgA responses or protective mucosal immunity. Nevertheless, currently licensed systemic vaccines do provide effective protection against mucosal pathogens such as influenza viruses and Streptococcus pneumoniae. However, whether systemic immunization induces protective mucosal immunity remains a controversial topic. Here we reviewed the current literature and discussed the potential of systemic routes of immunization for the induction of mucosal immunity. PMID:26752023

  15. 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

  16. 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. PMID:25888574

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

    PubMed Central

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

    2015-01-01

    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. PMID:25888574

  18. 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

  19. 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

  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 salvianolic acid B on NASH rat liver through restoring intestinal mucosal barrier function

    PubMed Central

    Wang, Ying-Chun; Jin, Qing-Mei; Kong, Wei-Zong; Chen, Juan

    2015-01-01

    Aim: To investigate the effect of Salvianolic acid B (Sal B) on the disease progress of NASH and change of intestinal barrier function. Methods: Sixty Sprague-Dawley (SD) rats were randomly divided into control group, model group and treated group, with the former given normal diet and the latter 2 groups rats fed high-fat diet. In treated group, rats were infused through the stomach with 1 mg/ml Sal B every day at a dose of 20 mL/kg body weight. All animals were killed at the 24th week and plasma levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), endotoxin (ET) and diamine oxdase (DAO) were analyzed using the blood samples. The histopathology of liver was observed by H&E staining. The expression changes of tight junction protein occludin and ZO-1 were analyzed by immunocytochemistry. Ultrastructural morphology of small intestinal tissues was investigated by transmission electron microscopy. Results: Plasma levels of ALT, AST, TG, TC, ET and DAO were significantly higher in model group than those in both control group and group treated with Sal B. In model group, vacuolated swelling of the cytoplasm with aggregates of chronic inflammatory cells was observed in the liver tissue but not in Sal B-treated group. NAFLD Activity Score in the treated group was significantly lower than that in model group. Immunohistochemical staining showed that Sal B administration recovered the expression of occludin and ZO-1, which was downregulated in the model group. Transmission electron microscopy analysis demonstrated that cell surface microvilli and major intercellular junctional complex including tight junction, gap junction and adherens junction were restored in Sal B-treated group. Conclusion: Sal B exerted protective function against high-fat diet-induced liver damage by restoring healthy barrier function of intestine in NASH rat model. PMID:26191218

  2. Physicochemical Factors that Affect Metal and Metal Oxide Nanoparticle Passage Across Epithelial Barriers

    PubMed Central

    Elder, Alison; Vidyasagar, Sadasivan; DeLouise, Lisa

    2014-01-01

    The diversity of nanomaterials in terms of size, shape, and surface chemistry poses a challenge to those who are trying to characterize the human health and environmental risks associated with incidental and unintentional exposures. There are numerous products that are already commercially available that contain solid metal and metal oxide nanoparticles, either embedded in a matrix or in solution. Exposure assessments for these products are often incomplete or difficult due to technological challenges associated with detection and quantitation of nanoparticles in gaseous or liquid carriers. The main focus of recent research has been on hazard identification. However, risk is a product of hazard and exposure, and one significant knowledge gap is that of the target organ dose following in vivo exposures. In order to reach target organs, nanoparticles must first breech the protective barriers of the respiratory tract, gastrointestinal tract, or skin. The fate of those nanoparticles that reach physiological barriers is in large part determined by the properties of the particles and the barriers themselves. This article reviews the physiological properties of the lung, gut, and skin epithelia, the physicochemical properties of metal and metal oxide nanoparticles that are likely to affect their ability to breech epithelial barriers, and what is known about their fate following in vivo exposures. PMID:20049809

  3. Postnatal requirement of the epithelial sodium channel for maintenance of epidermal barrier function.

    PubMed

    Charles, Roch-Philippe; Guitard, Marjorie; Leyvraz, Céline; Breiden, Bernadette; Haftek, Marek; Haftek-Terreau, Zofia; Stehle, Jean-Christophe; Sandhoff, Konrad; Hummler, Edith

    2008-02-01

    In skin, the physiological consequence of an epithelial sodium channel (ENaC) deficiency is not obvious directly at birth. Nevertheless, within hours after birth, mice deficient for the alpha-subunit of the highly amiloride-sensitive epithelial sodium channel (alphaENaC/Scnn1a) suffer from a significant increased dehydration. This is characterized by a loss of body weight (by 6% in 6 h) and an increased transepidermal water loss, which is accompanied by a higher skin surface pH in 1-day-old pups. Although early and late differentiation markers, as well as tight junction protein distribution and function, seem unaffected, deficiency of alphaENaC severely disturbs the stratum corneum lipid composition with decreased ceramide and cholesterol levels, and increased pro-barrier lipids, whereas covalently bound lipids are drastically reduced. Ultrastructural analysis revealed morphological changes in the formation of intercellular lamellar lipids and the lamellar body secretion. Extracellular formation of the lamellar lipids proved to be abnormal in the knockouts. In conclusion, ENaC deficiency results in progressive dehydration and, consequently, weight loss due to severe impairment of lipid formation and secretion. Our data demonstrate that ENaC expression is required for the postnatal maintenance of the epidermal barrier function but not for its generation. PMID:18039670

  4. 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

  5. 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

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

    PubMed

    Haines, Ricci J; 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

  7. 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

  8. Neutrophil α-Defensins Cause Lung Injury by Disrupting the Capillary–Epithelial Barrier

    PubMed Central

    Bdeir, Khalil; Higazi, Abd Al-Roof; Kulikovskaya, Irina; Christofidou-Solomidou, Melpo; Vinogradov, Sergei A.; Allen, Timothy C.; Idell, Steven; Linzmeier, Rose; Ganz, Tomas; Cines, Douglas B.

    2010-01-01

    Rationale: The involvement of neutrophil activation in the sentinel, potentially reversible, events in the pathogenesis of acute lung injury (ALI) is only partially understood. α-Defensins are the most abundant proteins secreted by activated human neutrophils, but their contribution to ALI in mouse models is hindered by their absence from murine neutrophils and the inability to study their effects in isolation in other species. Objectives: To study the role of α-defensins in the pathogenesis of ALI in a clinically relevant setting using mice transgenic for polymorphonuclear leukocyte expression of α-defensins. Methods: Transgenic mice expressing polymorphonuclear leukocyte α-defensins were generated. ALI was induced by acid aspiration. Pulmonary vascular permeability was studied in vivo using labeled dextran and fibrin deposition. The role of the low-density lipoprotein–related receptor (LRP) in permeability was examined. Measurements and Main Results: Acid aspiration induced neutrophil migration and release of α-defensins into lung parenchyma and airways. ALI was more severe in α-defensin–expressing mice than in wild-type mice, as determined by inspection, influx of neutrophils into the interstitial space and airways, histological evidence of epithelial injury, interstitial edema, extravascular fibrin deposition, impaired oxygenation, and reduced survival. Within 4 hours of insult, α-defensin–expressing mice showed greater disruption of capillary–epithelial barrier function and ALI that was attenuated by systemic or intratracheal administration of specific inhibitors of the LRP. Conclusions: α-Defensins mediate ALI through LRP-mediated loss of capillary–epithelial barrier function, suggesting a potential new approach to intervention. PMID:20093642

  9. A bovine mammary endothelial/epithelial cell culture model of the blood/milk barrier.

    PubMed Central

    Guidry, A J; O'Brien, C N; Douglass, L W

    1998-01-01

    The complex nature of the mammary gland has hampered in-depth studies of the relationship of the circulatory system to cells lining the teat ducts and alveoli of the gland. This study reports an in vitro model of endothelial and epithelial cells separated by a subcellular matrix that simulates the blood milk barrier of the bovine mammary gland. Dual chamber culture dishes with a porous membrane separating the upper and lower chamber were used. Endothelial and epithelial cells were cultured on opposite sides of the porous membrane. A collagen and fibroblast subcellular matrix, separating the 2 cell layers, simulated the in vivo interstitial tissue. Changes in surface binding of anti-bodies to polymorphonuclear neutrophils (PMN) following their migration from the upper to the lower chamber simulated the passage of PMN from blood to milk. Changes in the binding of antibodies to PMN agreed with results observed following the migration of PMN from blood to milk in vivo. This gives credence to the model's potential value for studies where more direct observation of the blood/milk barrier is required. The model will be further tested for its usefulness as an assay for determining: 1) antibiotic diffusion from milk to blood and from blood to milk, 2) cytotoxicity of prophylactic and therapeutic mammary infusion products, 3) factors affecting bacterial adhesion and penetration of mammary epithelial tissue, 4) effectiveness of antibodies present in lacteal secretions in preventing bacterial adhesion, and 5) the feasibility of gene constructs to induce synthesis and secretion of mastitis-preventing compounds and prophylactic and therapeutic compounds for treatment of human disorders. PMID:9553710

  10. Barrier protection via Toll-like receptor 2 signaling in porcine intestinal epithelial cells damaged by deoxynivalnol.

    PubMed

    Gu, Min Jeong; Song, Sun Kwang; Lee, In Kyu; Ko, Seongyeol; Han, Seung Eun; Bae, Suhan; Ji, Sang Yun; Park, Byung-Chul; Song, Ki-Duk; Lee, Hak-Kyo; Han, Seung Hyun; Yun, Cheol-Heui

    2016-01-01

    Intestinal barrier is the first line of defense inside the body and comprises intercellular tight junction (TJ) proteins that regulate paracellular permeability. Deoxynivalenol (DON), a fungal metabolite often found in the contaminated food of domestic animals, is known to impair intestinal barrier function and may be involved in intestinal inflammation. Unlike in humans and mice, the importance of Toll-like receptor (TLR) 2 expressed in porcine intestinal epithelial cells is largely unclear. Therefore, the aim of the present study was to investigate whether TLR2 stimulation enhances intestinal barrier function and protects against DON exposure. We found that the cells treated with TLR2 ligands decreased the epithelial barrier permeability and enhanced TJ protein expression in intestinal porcine epithelial cells (IPEC-J2). In addition, pretreatment with TLR2 ligand, including Pam3CSK4 (PCSK) and lipoteichoic acid from Bacillus subtilis, prevented DON-induced barrier dysfunction by increasing the expression of TJ proteins via the PI3K-Akt-dependent pathway. It is likely that the DON-disrupted intestinal barrier caused biological changes of immune cells in the lamina propria. Thus, we conducted co-culture of differentiated IPEC-J2 cells in the upper well together with peripheral blood mononuclear cells in the bottom well and found that apical TLR2 stimulation of IPEC-J2 cells could alleviate the reduction in cell survival and proliferation of immune cells. Conclusively, TLR2 signaling on intestinal epithelial cells may enhance intestinal barrier function and prevent DON-induced barrier dysfunction of epithelial cells. PMID:26857454

  11. 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

  12. 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...

  13. Gut Epithelial Barrier Dysfunction and Innate Immune Activation Predict Mortality in Treated HIV Infection

    PubMed Central

    Hunt, Peter W.; Sinclair, Elizabeth; Rodriguez, Benigno; Shive, Carey; Clagett, Brian; Funderburg, Nicholas; Robinson, Janet; Huang, Yong; Epling, Lorrie; Martin, Jeffrey N.; Deeks, Steven G.; Meinert, Curtis L.; Van Natta, Mark L.; Jabs, Douglas A.; Lederman, Michael M.

    2014-01-01

    Background. While inflammation predicts mortality in treated human immunodeficiency virus (HIV) infection, the prognostic significance of gut barrier dysfunction and phenotypic T-cell markers remains unclear. Methods. We assessed immunologic predictors of mortality in a case-control study within the Longitudinal Study of the Ocular Complications of AIDS (LSOCA), using conditional logistic regression. Sixty-four case patients who died within 12 months of treatment-mediated viral suppression were each matched to 2 control individuals (total number of controls, 128) by duration of antiretroviral therapy–mediated viral suppression, nadir CD4+ T-cell count, age, sex, and prior cytomegalovirus (CMV) retinitis. A similar secondary analysis was conducted in the SCOPE cohort, which had participants with less advanced immunodeficiency. Results. Plasma gut epithelial barrier integrity markers (intestinal fatty acid binding protein and zonulin-1 levels), soluble CD14 level, kynurenine/tryptophan ratio, soluble tumor necrosis factor receptor 1 level, high-sensitivity C-reactive protein level, and D-dimer level all strongly predicted mortality, even after adjustment for proximal CD4+ T-cell count (all P ≤ .001). A higher percentage of CD38+HLA-DR+ cells in the CD8+ T-cell population was a predictor of mortality before (P = .031) but not after (P = .10) adjustment for proximal CD4+ T-cell count. Frequencies of senescent (defined as CD28−CD57+ cells), exhausted (defined as PD1+ cells), naive, and CMV-specific T cells did not predict mortality. Conclusions. Gut epithelial barrier dysfunction, innate immune activation, inflammation, and coagulation—but not T-cell activation, senescence, and exhaustion—independently predict mortality in individuals with treated HIV infection with a history of AIDS and are viable targets for interventions. PMID:24755434

  14. 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

  15. 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

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

    PubMed

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

    2015-08-11

    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

  17. Critical role for IL-1β in DNA damage-induced mucositis

    PubMed Central

    Kanarek, Naama; Grivennikov, Sergei I.; Leshets, Michael; Lasry, Audrey; Alkalay, Irit; Horwitz, Elad; Shaul, Yoav D.; Stachler, Matthew; Voronov, Elena; Apte, Ron N.; Pagano, Michele; Pikarsky, Eli; Karin, Michael; Ghosh, Sankar; Ben-Neriah, Yinon

    2014-01-01

    β-TrCP, the substrate recognition subunit of SCF-type ubiquitin ligases, is ubiquitously expressed from two distinct paralogs, targeting for degradation many regulatory proteins, among which is the NF-κB inhibitor IκB. To appreciate tissue-specific roles of β-TrCP, we studied the consequences of inducible ablation of three or all four alleles of the E3 in the mouse gut. The ablation resulted in mucositis, a destructive gut mucosal inflammation, which is a common complication of different cancer therapies and represents a major obstacle to successful chemoradiation therapy. We identified epithelial-derived IL-1β as the culprit of mucositis onset, inducing mucosal barrier breach. Surprisingly, epithelial IL-1β is induced by DNA damage via an NF-κB–independent mechanism. Tissue damage caused by gut barrier disruption is exacerbated in the absence of NF-κB, with failure to express the endogenous IL-1β receptor antagonist IL-1Ra upon four-allele loss. Antibody neutralization of IL-1β prevents epithelial tight junction dysfunction and alleviates mucositis in β-TrCP–deficient mice. IL-1β antagonists should thus be considered for prevention and treatment of severe morbidity associated with mucositis. PMID:24469832

  18. Short communication: Differential loss of bovine mammary epithelial barrier integrity in response to lipopolysaccharide and lipoteichoic acid.

    PubMed

    Wellnitz, Olga; Zbinden, Christina; Huang, Xiao; Bruckmaier, Rupert M

    2016-06-01

    In the mammary gland, the blood-milk barrier prevents an uncontrolled intermixture of blood and milk constituents and hence maintains the osmotic gradient to draw water into the mammary secretion. During mastitis, the permeability of the blood-milk barrier is increased, which is reflected by the transfer of blood constituents into milk and vice versa. In this study, we aimed to investigate changes in the barrier function of mammary epithelial cells in vitro as induced by cell wall components of different pathogens. Primary bovine mammary epithelial cells from 3 different cows were grown separately on Transwell (Corning Inc., Corning, NY) inserts. The formation of tight junctions between adjacent epithelial cells was shown by transmission electron microscopy and by immunofluorescence staining of the tight junction protein zona occludens-1. The integrity of the epithelial barrier was assayed by means of transepithelial electrical resistance, as well as by diffusion of the fluorophore Lucifer yellow across the cell layer. The release of lactate dehydrogenase (LDH) was used as an indicator for cytotoxic effects. In response to a 24-h challenge with bacterial endotoxin, barrier integrity was reduced after 3 or 7h, respectively, in response to 0.5mg/mL lipopolysaccharide (LPS) from Escherichia coli or 20mg/mL lipoteichoic acid (LTA) from Staphylococcus aureus. No paracellular leakage was observed in response to 0.2mg/mL LPS or 2mg/mL LTA. Although LPS and LTA affected barrier permeability, most likely by opening the tight junctions, only LPS caused cell damage, reflected by increased LDH concentrations in cell culture medium. These results prove a pathogen-specific loss of blood-milk barrier integrity during mastitis, which is characterized by tight junction opening by both LPS and LTA and by additional epithelial cell destruction through LPS. PMID:27060811

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

    PubMed

    Moyes, David L; Wilson, Duncan; Richardson, Jonathan P; Mogavero, Selene; Tang, Shirley X; Wernecke, Julia; Höfs, Sarah; Gratacap, Remi L; Robbins, Jon; Runglall, Manohursingh; Murciano, Celia; Blagojevic, Mariana; Thavaraj, Selvam; Förster, Toni M; Hebecker, Betty; Kasper, Lydia; Vizcay, Gema; Iancu, Simona I; Kichik, Nessim; Häder, Antje; Kurzai, Oliver; Luo, Ting; Krüger, Thomas; Kniemeyer, Olaf; Cota, Ernesto; Bader, Oliver; Wheeler, Robert T; Gutsmann, Thomas; Hube, Bernhard; Naglik, Julian R

    2016-04-01

    Cytolytic proteins and peptide toxins are classical virulence factors of several bacterial pathogens which disrupt epithelial barrier function, damage cells and activate or modulate host immune responses. Such toxins have not been identified previously in human pathogenic fungi. Here we identify the first, to our knowledge, fungal cytolytic peptide toxin in the opportunistic pathogen Candida albicans. This secreted toxin directly damages epithelial membranes, triggers a danger response signalling pathway and activates epithelial immunity. Membrane permeabilization is enhanced by a positive charge at the carboxy terminus of the peptide, which triggers an inward current concomitant with calcium influx. C. albicans strains lacking this toxin do not activate or damage epithelial cells and are avirulent in animal models of mucosal infection. We propose the name 'Candidalysin' for this cytolytic peptide toxin; a newly identified, critical molecular determinant of epithelial damage and host recognition of the clinically important fungus, C. albicans. PMID:27027296

  20. Local Burn Injury Impairs Epithelial Permeability and Antimicrobial Peptide Barrier Function in Distal Unburned Skin*

    PubMed Central

    Plichta, Jennifer K.; Droho, Steve; Curtis, Brenda J.; Patel, Parita; Gamelli, Richard L.; Radek, Katherine A.

    2014-01-01

    Objectives Our objective was to characterize the mechanisms by which local burn injury compromises epithelial barrier function in burn margin, containing the elements necessary for healing of the burn site, and in distal unburned skin, which serves as potential donor tissue. Design Experimental mouse scald burn injury. Setting University Research Laboratory. Subjects C57/Bl6 Male mice, 8–12 weeks old. Interventions To confirm that dehydration was not contributing to our observed barrier defects, in some experiments mice received 1 mL of saline fluid immediately after burn, while a subgroup received an additional 0.5 mL at 4 hours and 1 mL at 24 hours following burn. We then assessed skin pH and transepidermal water loss every 12 hours on the burn wounds for 72 hours postburn. Measurements and Main Results Burn margin exhibited increased epidermal barrier permeability indicated by higher pH, greater transepidermal water loss, and reduced lipid synthesis enzyme expression and structural protein production up to 96 hours postburn. By contrast, antimicrobial peptide production and protease activity were elevated in burn margin. Skin extracts from burn margin did not exhibit changes in the ability to inhibit bacterial growth. However, distal unburned skin from burned mice also demonstrated an impaired response to barrier disruption, indicated by elevated transepidermal water loss and reduced lipid synthesis enzyme and structural protein expression up to 96 hours postburn. Furthermore, skin extracts from distal unburned skin exhibited greater protease activity and a reduced capacity to inhibit bacterial growth of several skin pathogens. Finally, we established that antimicrobial peptide levels were also altered in the lung and bladder, which are common sites of secondary infection in burn-injured patients. Conclusions These findings reveal several undefined deficiencies in epithelial barrier function at the burn margin, potential donor skin sites, and organs

  1. Emodin enhances alveolar epithelial barrier function in rats with experimental acute pancreatitis

    PubMed Central

    Xia, Xian-Ming; Wang, Fang-Yu; Wang, Zhen-Kai; Wan, Hai-Jun; Xu, Wen-An; Lu, Heng

    2010-01-01

    AIM: To investigate the effect of emodin on expression of claudin-4, claudin-5 and occludin, as well as the alveolar epithelial barrier in rats with pancreatitis induced by sodium taurocholate. METHODS: Experimental pancreatitis was induced by retrograde injection of 5% sodium taurocholate into the biliopancreatic duct. Emodin was injected via the external jugular vein 3 h after induction of acute pancreatitis. Rats from sham operation group and acute pancreatitis group were injected with normal saline (an equivalent volume as emodin) at the same time point. Samples of lung and serum were obtained 6 h after drug administration. Pulmonary morphology was examined with HE staining. Pulmonary edema was estimated by measuring water content in lung tissue samples. Tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) level were measured by enzyme-linked immunospecific assay. Serum amylase and pulmonary myeloperoxidase (MPO) activity were detected by spectrophotometry. Alveolar epithelial barrier was assessed by pulmonary dye extravasation. Expression of claudin-4, claudin-5 and occludin in lung tissue samples was examined by immunohistology, quantitative real-time reverse transcription polymerase chain reaction and Western blotting analysis, respectively. RESULTS: Pancreatitis-associated lung injury was characterized by pulmonary edema, leukocyte infiltration, alveolar collapse, and elevated serum amylase level. The pulmonary damage, pulmonary pathological scores, serum amylase and MPO activity, TNF-α and IL-6 levels, and wet/dry ratio were decreased in rats after treatment with emodin. Immunostaining of claudin-4, claudin-5 and occludin was detected in lung tissue samples from rats in sham operation group, which was distributed in alveolar epithelium, vascular endothelium, and bronchial epithelium, respectively. The mRNA and protein expression levels of claudin-4, claudin-5 and occludin in lung tissue samples were markedly decreased, the expression level of

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

    PubMed

    Huang, Wufeng; Zhao, Haijin; Dong, Hangming; Wu, Yue; Yao, Lihong; Zou, Fei; Cai, Shaoxi

    2016-05-01

    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

  3. 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

  4. 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. PMID:26294673

  5. Nitric oxide attenuates hydrogen peroxide-induced barrier disruption and protein tyrosine phosphorylation in monolayers of intestinal epithelial cell.

    PubMed

    Katsube, Takanori; Tsuji, Hideo; Onoda, Makoto

    2007-06-01

    The intestinal epithelium provides a barrier to the transport of harmful luminal molecules into the systemic circulation. A dysfunctional epithelial barrier is closely associated with the pathogenesis of a variety of intestinal and systemic disorders. We investigated here the effects of nitric oxide (NO) and hydrogen peroxide (H(2)O(2)) on the barrier function of a human intestinal epithelial cell line, Caco-2. When treated with H(2)O(2), Caco-2 cell monolayers grown on permeable supports exhibited several remarkable features of barrier dysfunction as follows: a decrease in transepithelial electrical resistance, an increase in paracellular permeability to dextran, and a disruption of the intercellular junctional localization of the scaffolding protein ZO-1. In addition, an induction of tyrosine phosphorylation of numerous cellular proteins including ZO-1, E-cadherin, and beta-catenin, components of tight and adherens junctions, was observed. On the other hand, combined treatment of Caco-2 monolayers with H(2)O(2) and an NO donor (NOC5 or NOC12) relieved the damage to the barrier function and suppressed the protein tyrosine phosphorylation induced by H(2)O(2) alone. These results suggest that NO protects the barrier function of intestinal epithelia from oxidative stress by modulating some intracellular signaling pathways of protein tyrosine phosphorylation in epithelial cells. PMID:17451824

  6. Epithelial adhesion molecules and the regulation of intestinal homeostasis during neutrophil transepithelial migration.

    PubMed

    Sumagin, Ronen; Parkos, Charles A

    2015-01-01

    Epithelial adhesion molecules play essential roles in regulating cellular function and maintaining mucosal tissue homeostasis. Some form epithelial junctional complexes to provide structural support for epithelial monolayers and act as a selectively permeable barrier separating luminal contents from the surrounding tissue. Others serve as docking structures for invading viruses and bacteria, while also regulating the immune response. They can either obstruct or serve as footholds for the immune cells recruited to mucosal surfaces. Currently, it is well appreciated that adhesion molecules collectively serve as environmental cue sensors and trigger signaling events to regulate epithelial function through their association with the cell cytoskeleton and various intracellular adapter proteins. Immune cells, particularly neutrophils (PMN) during transepithelial migration (TEM), can modulate adhesion molecule expression, conformation, and distribution, significantly impacting epithelial function and tissue homeostasis. This review discusses the roles of key intestinal epithelial adhesion molecules in regulating PMN trafficking and outlines the potential consequences on epithelial function. PMID:25838976

  7. Epithelial adhesion molecules and the regulation of intestinal homeostasis during neutrophil transepithelial migration

    PubMed Central

    Sumagin, Ronen; Parkos, Charles A

    2014-01-01

    Epithelial adhesion molecules play essential roles in regulating cellular function and maintaining mucosal tissue homeostasis. Some form epithelial junctional complexes to provide structural support for epithelial monolayers and act as a selectively permeable barrier separating luminal contents from the surrounding tissue. Others serve as docking structures for invading viruses and bacteria, while also regulating the immune response. They can either obstruct or serve as footholds for the immune cells recruited to mucosal surfaces. Currently, it is well appreciated that adhesion molecules collectively serve as environmental cue sensors and trigger signaling events to regulate epithelial function through their association with the cell cytoskeleton and various intracellular adapter proteins. Immune cells, particularly neutrophils (PMN) during transepithelial migration (TEM), can modulate adhesion molecule expression, conformation, and distribution, significantly impacting epithelial function and tissue homeostasis. This review discusses the roles of key intestinal epithelial adhesion molecules in regulating PMN trafficking and outlines the potential consequences on epithelial function. PMID:25838976

  8. Recovery of mucosal barrier function in ischemic porcine ileum and colon is stimulated by a novel agonist of the ClC-2 chloride channel, lubiprostone.

    PubMed

    Moeser, Adam J; Nighot, Prashant K; Engelke, Kory J; Ueno, Ryuji; Blikslager, Anthony T

    2007-02-01

    Previous studies utilizing an ex vivo porcine model of intestinal ischemic injury demonstrated that prostaglandin (PG)E(2) stimulates repair of mucosal barrier function via a mechanism involving Cl(-) secretion and reductions in paracellular permeability. Further experiments revealed that the signaling mechanism for PGE(2)-induced mucosal recovery was mediated via type-2 Cl(-) channels (ClC-2). Therefore, the objective of the present study was to directly investigate the role of ClC-2 in mucosal repair by evaluating mucosal recovery in ischemia-injured intestinal mucosa treated with the selective ClC-2 agonist lubiprostone. Ischemia-injured porcine ileal mucosa was mounted in Ussing chambers, and short-circuit current (I(sc)) and transepithelial electrical resistance (TER) were measured in response to lubiprostone. Application of 0.01-1 microM lubiprostone to ischemia-injured mucosa induced concentration-dependent increases in TER, with 1 microM lubiprostone stimulating a twofold increase in TER (DeltaTER = 26 Omega.cm(2); P < 0.01). However, lubiprostone (1 microM) stimulated higher elevations in TER despite lower I(sc) responses compared with the nonselective secretory agonist PGE(2) (1 microM). Furthermore, lubiprostone significantly (P < 0.05) reduced mucosal-to-serosal fluxes of (3)H-labeled mannitol to levels comparable to those of normal control tissues and restored occludin localization to tight junctions. Activation of ClC-2 with the selective agonist lubiprostone stimulated elevations in TER and reductions in mannitol flux in ischemia-injured intestine associated with structural changes in tight junctions. Prostones such as lubiprostone may provide a selective and novel pharmacological mechanism of accelerating recovery of acutely injured intestine compared with the nonselective action of prostaglandins such as PGE(2). PMID:17053162

  9. 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

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

    PubMed

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

    2016-03-01

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

  11. PRIMARY CULTURE OF CHOROIDAL EPITHELIAL CELLS: CHARACTERIZATION OF AN IN VITRO MODEL OF BLOOD-CSF BARRIER

    PubMed Central

    ZHENG, WEI; ZHAO, QIUQU; GRAZIANO, JOSEPH H.

    2016-01-01

    Summary A primary rat choroidal epithelial cell culture system was developed to investigate mechanisms of heavy metal toxicity on the blood-cerebrospinal fluid (CSF) barrier. Epithelial cells were dissociated from choroidal tissue by pronase digestion and cultured in standard DMEM culture media supplemented with 10% fetal bovine serum and 10 ng epithelial growth factor per ml. The procedure yielded 2–5 × 104 cells from pooled plexuses of three to four rats, and a viability of 77–85%. The cultures displayed a dominant polygonal type of epithelial cells, with a population doubling time of 2–3 d. The cultures were of distinct choroidal epithelial origins. For example, immunocytochemical studies using monospecific rabbit anti-rat TTR polyclonal antibody revealed a strong positive stain of transthyretin (TTR), a thyroxine transport protein exclusively produced by the choroidal epithelia. Also, reverse-transcriptase polymerase chain reaction (PCR) confirmed the presence of specific TTR mRNA in the cultures. The cultures were further adapted to grow on a freely permeable membrane sandwiched between two culture chambers. The formation of an impermeable confluent monolayer occurred within 5 d after seeding and was verified by the presence of a steady electrical resistance across the membrane (80 ± 10 ohm per cm2). The epithelial barriers appeared to actively transport [125I]-thyroxine from the basal to apical chamber. These results suggest that this primary cell culture system possesses typical choroidal epithelial characteristics and appears to be a suitable model for in vitro mechanistic investigations of blood–CSF barrier. PMID:9542634

  12. Loss of EP2 receptor subtype in colonic cells compromise epithelial barrier integrity by altering claudin-4.

    PubMed

    Lejeune, Manigandan; Moreau, France; Chadee, Kris

    2014-01-01

    Prostaglandin E2 (PGE2) is a bioactive lipid mediator that exerts its biological function through interaction with four different subtypes of E-Prostanoid receptor namely EP1, EP2, EP3 and EP4. It has been known that EP2 receptor is differentially over-expressed in the epithelia of inflamed human colonic mucosa. However, the significance of the differential expression in altering epithelial barrier function is not known. In this study, we used Caco-2 cells expressing EP2 receptor, either high (EP2S) or low (EP2A), as a model epithelia and determined the barrier function of these cell monolayers by measuring the trans epithelial resistance (TER). Basal TER of EP2A (but not EP2S) monolayer was significantly lower suggesting a loss of colonic epithelial barrier integrity. In comparison, the TER of wild type Caco-2 was decreased in response to an EP2 receptor specific antagonist (AH-6809) indicating an important role for EP2 receptor in the maintenance of epithelial barrier function. The decrease TER in EP2A monolayer corresponded with a significant loss of the tight junction (TJ) protein claudin-4 without affecting other major TJ proteins. Similarly, EP2 receptor antagonism/siRNA based silencing significantly decreased claudin-4 expression in EP2S cells. Surprisingly, alteration in claudin-4 was not transcriptionally regulated in EP2A cells but rather undergoes increased proteosomal degradation. Moreover, among the TER compromising cytokines examined (IL-8, IL-1β, TNF-α, IFN-γ) only IFN-γ was significantly up regulated in EP2A cells. However, IFN-γ did not significantly decreased claudin-4 expression in Caco-2 cells indicating no role for IFN-γ in degrading claudin-4. We conclude that differential down-regulation of EP2 receptor play a major role in compromising colonic epithelial barrier function by selectively increasing proteosomal degradation of claudin-4. PMID:25396731

  13. 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

  14. 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.

  15. Crosstalk between Microbiota-Derived Short-Chain Fatty Acids and Intestinal Epithelial HIF Augments Tissue Barrier Function.

    PubMed

    Kelly, Caleb J; Zheng, Leon; Campbell, Eric L; Saeedi, Bejan; Scholz, Carsten C; Bayless, Amanda J; Wilson, Kelly E; Glover, Louise E; Kominsky, Douglas J; Magnuson, Aaron; Weir, Tiffany L; Ehrentraut, Stefan F; Pickel, Christina; Kuhn, Kristine A; Lanis, Jordi M; Nguyen, Vu; Taylor, Cormac T; Colgan, Sean P

    2015-05-13

    Interactions between the microbiota and distal gut are fundamental determinants of human health. Such interactions are concentrated at the colonic mucosa and provide energy for the host epithelium through the production of the short-chain fatty acid butyrate. We sought to determine the role of epithelial butyrate metabolism in establishing the austere oxygenation profile of the distal gut. Bacteria-derived butyrate affects epithelial O2 consumption and results in stabilization of hypoxia-inducible factor (HIF), a transcription factor coordinating barrier protection. Antibiotic-mediated depletion of the microbiota reduces colonic butyrate and HIF expression, both of which are restored by butyrate supplementation. Additionally, germ-free mice exhibit diminished retention of O2-sensitive dyes and decreased stabilized HIF. Furthermore, the influences of butyrate are lost in cells lacking HIF, thus linking butyrate metabolism to stabilized HIF and barrier function. This work highlights a mechanism where host-microbe interactions augment barrier function in the distal gut. PMID:25865369

  16. 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

  17. Healthcare Burden, Risk Factors, and Outcomes of Mucosal Barrier Injury Laboratory-Confirmed Bloodstream Infections after Stem Cell Transplantation.

    PubMed

    Dandoy, Christopher E; Haslam, David; Lane, Adam; Jodele, Sonata; Demmel, Kathy; El-Bietar, Javier; Flesch, Laura; Myers, Kasiani C; Pate, Abigail; Rotz, Seth; Daniels, Paulina; Wallace, Gregory; Nelson, Adam; Waters, Heather; Connelly, Beverly; Davies, Stella M

    2016-09-01

    Mucosal barrier injury laboratory-confirmed bloodstream infections (MBI-LCBIs) lead to significant morbidity, mortality, and healthcare resource utilization in hematopoietic stem cell transplant (HSCT) patients. Determination of the healthcare burden of MBI-LCBIs and identification of patients at risk of MBI-LCBIs will allow researchers to identify strategies to reduce MBI-LCBI rates. The objective of our study was to describe the incidence, risk factors, timing, and outcomes of MBI-LCBIs in hematopoietic stem cell transplant patients. We performed a retrospective analysis of 374 patients who underwent HSCT at a large free-standing academic children's hospital to determine the incidence, risk factors, and outcomes of patients that developed a bloodstream infection (BSI) including MBI-LCBI, central line-associated BSI (CLABSI), or secondary BSI in the first year after HSCT. Outcome measures included nonrelapse mortality (NRM), central venous catheter removal within 7 days of positive culture, shock, admission to the pediatric intensive care unit (PICU) within 48 hours of positive culture, and death within 10 days of positive culture. One hundred seventy BSIs were diagnosed in 100 patients (27%): 80 (47%) MBI-LCBIs, 68 (40%) CLABSIs, and 22 (13%) secondary infections. MBI-LCBIs were diagnosed at a significantly higher rate in allogeneic HSCT patients (18% versus 7%, P = .007). Reduced-intensity conditioning (OR, 1.96; P = .015) and transplant-associated thrombotic microangiopathy (OR, 2.94; P = .0004) were associated with MBI-LCBI. Nearly 50% of all patients with a BSI developed septic shock, 10% died within 10 days of positive culture, and nearly 25% were transferred to the PICU. One-year NRM was significantly increased in patients with 1 (34%) and more than 1 (56%) BSIs in the first year post-HSCT compared with those who did not develop BSIs (14%) (P ≤ .0001). There was increased 1-year NRM in patients with at least 1 MBI-LCBI (OR, 1.94; P

  18. 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...

  19. 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. PMID:26519558

  20. Wound repair: role of immune–epithelial interactions

    PubMed Central

    Leoni, G; Neumann, P-A; Sumagin, R; Denning, TL; Nusrat, A

    2016-01-01

    The epithelium serves as a highly selective barrier at mucosal surfaces. Upon injury, epithelial wound closure is orchestrated by a series of events that emanate from the epithelium itself as well as by the temporal recruitment of immune cells into the wound bed. Epithelial cells adjoining the wound flatten out, migrate, and proliferate to rapidly cover denuded surfaces and re-establish mucosal homeostasis. This process is highly regulated by proteins and lipids, proresolving mediators such as Annexin A1 protein and resolvins released into the epithelial milieu by the epithelium itself and infiltrating innate immune cells including neutrophils and macrophages. Failure to achieve these finely tuned processes is observed in chronic inflammatory diseases that are associated with non-healing wounds. An improved understanding of mechanisms that mediate repair is important in the development of therapeutics aimed to promote mucosal wound repair. PMID:26174765

  1. IL-22 modulates gut epithelial and immune barrier functions following acute alcohol exposure and burn injury

    PubMed Central

    Rendon, Juan L.; Li, Xiaoling; Akhtar, Suhail; Choudhry, Mashkoor A.

    2012-01-01

    Interleukin (IL)–22 maintains gut epithelial integrity and expression of antimicrobial peptides (AMPs) Reg3β and Reg3γ. Our laboratory has shown that acute alcohol/ethanol (EtOH) exposure prior to burn injury results in increased gut permeability, intestinal T cell suppression and enhanced bacterial translocation. Herein, we determined the effect of combined EtOH intoxication and burn injury on intestinal levels of IL-22 as well as Reg3β and Reg3γ expression. We further examined whether in vivo restitution of IL-22 restores gut permeability, Reg3β and Reg3γ levels, and bacterial load (e.g. gut bacterial growth) within the intestine following EtOH and burn injury. Male mice, ~25g, were gavaged with EtOH (2.9 mg/kg) prior to receiving a ~12.5% total body surface area full thickness burn. Mice were immediately treated with saline control or IL-22 (1 mg/kg) by i.p. injection. One day post injury, there was a significant decrease in intestinal IL-22, Reg3β and Reg3γ expression along with an increase in intestinal permeability and gut bacterial load following EtOH combined with burn injury, as compared to sham injury. Treatment with IL-22 normalized Reg3β and Reg3γ expression, and attenuated the increase in intestinal permeability following EtOH and burn injury. Qualitatively, IL-22 treatment reduced the bacterial load in nearly half of mice receiving EtOH combined with burn injury. Our data indicate that IL-22 maintains gut epithelial and immune barrier integrity following EtOH and burn injury; thus, the IL-22/AMP pathway may provide a therapeutic target for the treatment of patients who sustain burn injury under the influence of EtOH. PMID:23143063

  2. 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

  3. Vascular endothelial growth factor antagonism restores epithelial barrier dysfunction via affecting zonula occludens proteins

    PubMed Central

    YUKSEL, HASAN; YILMAZ, OZGE; KARAMAN, MERAL; FIRINCI, FATIH; TURKELI, AHMET; KANIK, ESRA TOPRAK; INAN, SEVINC

    2015-01-01

    Epithelial barrier dysfunction is important in the pathogenesis of asthma and allergic responses, and is therefore a therapeutic target. The aim of the present study was to investigate the effects of dexamethasone, a classic therapeutic agent, an anti-tumor necrosis factor agent (etanercept), which is used to treat difficult cases of asthma, and an anti-vascular endothelial growth factor (VEGF) agent (bevacizumab), which is an angiogenesis inhibitor, on zonula occludens (ZO) proteins in an experimental asthma model. The experimental model of asthma was developed using intraperitoneal (IP) and inhaled administration of ovalbumin in 38 BALB/c mice, which were divided into four groups. The control group (n=6) did not receive any treatment, while the four remaining groups (n=8 per group) received an IP injection of saline, etanercept, bevacizumab or dexamethasone, respectively. Occludin, claudin and junctional adhesion molecule (JAM) were immunohistochemically stained in the left middle lobe samples using an indirect avidin-peroxidase method, after which the staining was semiquantified with H-scores. Statistically significant differences were observed in the occludin, claudin and JAM H-scores among the four groups (P<0.001). In the untreated asthma, etanercept, bevacizumab and dexamethasone groups, the median H-scores for occludin were 93, 177, 280 and 198, respectively, while the H-scores for claudin were 82, 193.5, 274 and 202.5, respectively, and the median H-scores for JAM were 130, 210, 288 and 210, respectively. Pairwise comparisons revealed that all three ZO protein H-scores were significantly lower in the saline group when compared with each treatment group. However, the H-scores of the ZO proteins were not significantly different between the etanercept and dexamethasone groups. Furthermore, the bevacizumab group exhibited higher H-scores for all the proteins compared with the dexamethasone group. Therefore, antagonism of VEGF with bevacizumab restores the

  4. Epithelial restitution and wound healing in inflammatory bowel disease.

    PubMed

    Sturm, Andreas; Dignass, Axel U

    2008-01-21

    Inflammatory bowel disease is characterized by a chronic inflammation of the intestinal mucosa. The mucosal epithelium of the alimentary tract constitutes a key element of the mucosal barrier to a broad spectrum of deleterious substances present within the intestinal lumen including bacterial microorganisms, various dietary factors, gastrointestinal secretory products and drugs. In addition, this mucosal barrier can be disturbed in the course of various intestinal disorders including inflammatory bowel diseases. Fortunately, the integrity of the gastrointestinal surface epithelium is rapidly reestablished even after extensive destruction. Rapid resealing of the epithelial barrier following injuries is accomplished by a process termed epithelial restitution, followed by more delayed mechanisms of epithelial wound healing including increased epithelial cell proliferation and epithelial cell differentiation. Restitution of the intestinal surface epithelium is modulated by a range of highly divergent factors among them a broad spectrum of structurally distinct regulatory peptides, variously described as growth factors or cytokines. Several regulatory peptide factors act from the basolateral site of the epithelial surface and enhance epithelial cell restitution through TGF-beta-dependent pathways. In contrast, members of the trefoil factor family (TFF peptides) appear to stimulate epithelial restitution in conjunction with mucin glycoproteins through a TGF-beta-independent mechanism from the apical site of the intestinal epithelium. In addition, a number of other peptide molecules like extracellular matrix factors and blood clotting factors and also non-peptide molecules including phospholipids, short-chain fatty acids (SCFA), adenine nucleotides, trace elements and pharmacological agents modulate intestinal epithelial repair mechanisms. Repeated damage and injury of the intestinal surface are key features of various intestinal disorders including inflammatory bowel

  5. Epithelial restitution and wound healing in inflammatory bowel disease

    PubMed Central

    Sturm, Andreas; Dignass, Axel U

    2008-01-01

    Inflammatory bowel disease is characterized by a chronic inflammation of the intestinal mucosa. The mucosal epithelium of the alimentary tract constitutes a key element of the mucosal barrier to a broad spectrum of deleterious substances present within the intestinal lumen including bacterial microorganisms, various dietary factors, gastrointestinal secretory products and drugs. In addition, this mucosal barrier can be disturbed in the course of various intestinal disorders including inflammatory bowel diseases. Fortunately, the integrity of the gastrointestinal surface epithelium is rapidly reestablished even after extensive destruction. Rapid resealing of the epithelial barrier following injuries is accomplished by a process termed epithelial restitution, followed by more delayed mechanisms of epithelial wound healing including increased epithelial cell proliferation and epithelial cell differentiation. Restitution of the intestinal surface epithelium is modulated by a range of highly divergent factors among them a broad spectrum of structurally distinct regulatory peptides, variously described as growth factors or cytokines. Several regulatory peptide factors act from the basolateral site of the epithelial surface and enhance epithelial cell restitution through TGF-β-dependent pathways. In contrast, members of the trefoil factor family (TFF peptides) appear to stimulate epithelial restitution in conjunction with mucin glycoproteins through a TGF-β-independent mechanism from the apical site of the intestinal epithelium. In addition, a number of other peptide molecules like extracellular matrix factors and blood clotting factors and also non-peptide molecules including phospholipids, short-chain fatty acids (SCFA), adenine nucleotides, trace elements and pharmacological agents modulate intestinal epithelial repair mechanisms. Repeated damage and injury of the intestinal surface are key features of various intestinal disorders including inflammatory bowel

  6. The Role of Intestinal Microbiota in the Development and Severity of Chemotherapy-Induced Mucositis

    PubMed Central

    van Vliet, Michel J.; Harmsen, Hermie J. M.; de Bont, Eveline S. J. M.; Tissing, Wim J. E.

    2010-01-01

    Mucositis, also referred to as mucosal barrier injury, is one of the most debilitating side effects of radiotherapy and chemotherapy treatment. Clinically, mucositis is associated with pain, bacteremia, and malnutrition. Furthermore, mucositis is a frequent reason to postpone chemotherapy treatment, ultimately leading towards a higher mortality in cancer patients. According to the model introduced by Sonis, both inflammation and apoptosis of the mucosal barrier result in its discontinuity, thereby promoting bacterial translocation. According to this five-phase model, the intestinal microbiota plays no role in the pathophysiology of mucositis. However, research has implicated a prominent role for the commensal intestinal microbiota in the development of several inflammatory diseases like inflammatory bowel disease, pouchitis, and radiotherapy-induced diarrhea. Furthermore, chemotherapeutics have a detrimental effect on the intestinal microbial composition (strongly decreasing the numbers of anaerobic bacteria), coinciding in time with the development of chemotherapy-induced mucositis. We hypothesize that the commensal intestinal microbiota might play a pivotal role in chemotherapy-induced mucositis. In this review, we propose and discuss five pathways in the development of mucositis that are potentially influenced by the commensal intestinal microbiota: 1) the inflammatory process and oxidative stress, 2) intestinal permeability, 3) the composition of the mucus layer, 4) the resistance to harmful stimuli and epithelial repair mechanisms, and 5) the activation and release of immune effector molecules. Via these pathways, the commensal intestinal microbiota might influence all phases in the Sonis model of the pathogenesis of mucositis. Further research is needed to show the clinical relevance of restoring dysbiosis, thereby possibly decreasing the degree of intestinal mucositis. PMID:20523891

  7. Bacterial-mucosal interactions in inflammatory bowel disease—an alliance gone bad

    PubMed Central

    Chichlowski, Maciej; Hale, Laura P.

    2008-01-01

    The complex interaction of genetic, microbial, and environmental factors may result in continuous activation of the mucosal immune system leading to inflammatory bowel disease (IBD). Most present treatments for IBD involve altering or suppressing the aberrant immune response; however, the role of the intestinal microbiota in the pathophysiology of IBD is becoming more evident. The epithelial layer is essential for the proper functioning of the gastrointestinal tract, and its increased permeability to the luminal antigens may lead to the inflammatory processes and mucosal damage observed in IBD. Factors affecting the efficacy of the epithelial barrier include presence of pathogenic bacteria (e.g., Helicobacter spp.), presence of probiotic bacteria, availability of selected nutrients, and others. Defective function of the mucosal barrier might facilitate the contact of bacterial antigens and adjuvants with innate and adaptive immune cells to generate prolonged inflammatory responses. This review will briefly describe the complex structure of the epithelial barrier in the context of bacterial-mucosal interactions observed in human IBD and mouse models of colitis. PMID:18927210

  8. The methionine precursor DL-2-hydroxy-(4-methylthio)butanoic acid protects intestinal epithelial barrier function.

    PubMed

    Martín-Venegas, Raquel; Brufau, M Teresa; Guerrero-Zamora, Ana Maria; Mercier, Yves; Geraert, Pierre-André; Ferrer, Ruth

    2013-12-01

    DL-2-hydroxy-(4-methylthio)butanoic acid (HMTBA) is a source of dietary methionine (Met) that is widely used in poultry nutrition. We have previously shown that HMTBA is preferentially diverted to the transsulfuration pathway, which gives antioxidant metabolites such as taurine and glutathione. Therefore, here we hypothesize that this Met source can protect epithelial barrier function in an in vitro model of intestinal inflammation of Caco-2 cells. The results show that HMTBA prevents the increase in paracellular permeability induced by H2O2 or tumour necrosis factor-α. This effect can be attributed to the increased production of taurine and reduced glutathione. Similar results were obtained for DL-Met, although the protective role of the amino acid was less pronounced than that of the hydroxy analogue. In conclusion, the diversion to the transsulfuration pathway means that this Met precursor is of greater value than previously thought, due to its capacity to improve intestinal homeostasis and the quality of poultry products destined for human consumption. PMID:23870881

  9. 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.

  10. 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. PMID:25907690

  11. Intestinal mucosal tolerance and impact of gut microbiota to mucosal tolerance

    PubMed Central

    Chistiakov, Dimitry A.; Bobryshev, Yuri V.; Kozarov, Emil; Sobenin, Igor A.; Orekhov, Alexander N.

    2015-01-01

    The mucosal barriers are very sensitive to pathogenic infection, thereby assuming the capacity of the mucosal immune system to induce protective immunity to harmful antigens and tolerance against harmless substances. This review provides current information about mechanisms of induction of mucosal tolerance and about impact of gut microbiota to mucosal tolerance. PMID:25628617

  12. Polyinosinic:polycytidylic acid induces protein kinase D–dependent disassembly of apical junctions and barrier dysfunction in airway epithelial cells

    PubMed Central

    Rezaee, Fariba; Meednu, Nida; Emo, Jason A.; Saatian, Bahman; Chapman, Timothy J.; Naydenov, Nayden G.; De Benedetto, Anna; Beck, Lisa A.; Ivanov, Andrei I.; Georas, Steve N.

    2011-01-01

    Background Disruption of the epithelial barrier might be a risk factor for allergen sensitization and asthma. Viral respiratory tract infections are strongly associated with asthma exacerbation, but the effects of respiratory viruses on airway epithelial barrier function are not well understood. Many viruses generate double-stranded RNA, which can lead to airway inflammation and initiate an antiviral immune response. Objectives We investigated the effects of the synthetic double-stranded RNA polyinosinic:polycytidylic acid (polyI:C) on the structure and function of the airway epithelial barrier in vitro. Methods 16HBE14o- human bronchial epithelial cells and primary airway epithelial cells at an air-liquid interface were grown to confluence on Transwell inserts and exposed to polyI:C. We studied epithelial barrier function by measuring transepithelial electrical resistance and paracellular flux of fluorescent markers and structure of epithelial apical junctions by means of immunofluorescence microscopy. Results PolyI:C induced a profound decrease in transepithelial electrical resistance and increase in paracellular permeability. Immunofluorescence microscopy revealed markedly reduced junctional localization of zonula occludens-1, occludin, E-cadherin, β-catenin, and disorganization of junction-associated actin filaments. PolyI:C induced protein kinase D (PKD) phosphorylation, and a PKD antagonist attenuated polyI:C-induced disassembly of apical junctions and barrier dysfunction. Conclusions PolyI:C has a powerful and previously unsuspected disruptive effect on the airway epithelial barrier. PolyI:C-dependent barrier disruption is mediated by disassembly of epithelial apical junctions, which is dependent on PKD signaling. These findings suggest a new mechanism potentially underlying the associations between viral respiratory tract infections, airway inflammation, and allergen sensitization. PMID:21996340

  13. 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

  14. 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

  15. Efficient Mucosal Delivery of Vaccine Using the FcRn-Mediated IgG Transfer Pathway

    PubMed Central

    Ye, Lilin; Zeng, Rongyu; Bai, Yu; Roopenian, Derry C.; Zhu, Xiaoping

    2011-01-01

    Vaccine strategies to prevent invasive mucosal pathogens are being sought because 80–90% of infectious diseases are initiated at mucosal surfaces. However, our ability to deliver an intact vaccine antigen across a mucosal barrier for induction of effective immunity is limited. The neonatal Fc receptor (FcRn) mediates the transport of IgG across polarized epithelial cells lining mucosal surfaces. By mimicking IgG transfer at mucosal surfaces, intranasal immunization with a model antigen, herpes simplex virus type-2 (HSV-2) glycoprotein gD fused with an IgG Fc fragment, in combination with the adjuvant CpG, resulted in complete protection of wild type, but not FcRn knockout, mice that were intravaginally challenged with virulent HSV-2 186. This immunization strategy induced efficient mucosal and systemic antibody, B and T cell immune responses, including memory immune responses, which remained stable at least 6 months post-vaccination and conferred protection for a majority of animals. These results demonstrate that the FcRn-IgG transcellular transport pathway may represent a novel mucosal vaccine delivery route for a subunit vaccine against abundant mucosal pathogens. PMID:21240266

  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. PMID:26536291

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

    PubMed

    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

  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. Altered intestinal microbial flora and impaired epithelial barrier structure and function in CKD: the nature, mechanisms, consequences and potential treatment.

    PubMed

    Vaziri, Nosratola D; Zhao, Ying-Yong; Pahl, Madeleine V

    2016-05-01

    Chronic kidney disease (CKD) results in systemic inflammation and oxidative stress which play a central role in CKD progression and its adverse consequences. Although many of the causes and consequences of oxidative stress and inflammation in CKD have been extensively explored, little attention had been paid to the intestine and its microbial flora as a potential source of these problems. Our recent studies have revealed significant disruption of the colonic, ileal, jejunal and gastric epithelial tight junction in different models of CKD in rats. Moreover, the disruption of the epithelial barrier structure and function found in uremic animals was replicated in cultured human colonocytes exposed to uremic human plasma in vitro We have further found significant changes in the composition and function of colonic bacterial flora in humans and animals with advanced CKD. Together, uremia-induced impairment of the intestinal epithelial barrier structure and function and changes in composition of the gut microbiome contribute to the systemic inflammation and uremic toxicity by accommodating the translocation of endotoxin, microbial fragments and other noxious luminal products in the circulation. In addition, colonic bacteria are the main source of several well-known pro-inflammatory uremic toxins such as indoxyl sulfate, p-cresol sulfate, trimethylamine-N-oxide and many as-yet unidentified retained compounds in end-stage renal disease patients. This review is intended to provide an overview of the effects of CKD on the gut microbiome and intestinal epithelial barrier structure and their role in the pathogenesis of systemic inflammation and uremic toxicity. In addition, potential interventions aimed at mitigating these abnormalities are briefly discussed. PMID:25883197

  1. 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

  2. Enteral Nutrient Deprivation in Patients Leads to a Loss of Intestinal Epithelial Barrier Function

    PubMed Central

    Ralls, Matthew W.; Demehri, Farokh R.; Feng, Yongjia; Woods Ignatoski, Kathleen M.; Teitelbaum, Daniel H.

    2014-01-01

    Objective To investigate the effect of nutrient withdrawal on human intestinal epithelial barrier function (EBF). We hypothesized that unfed mucosa results in decreased EBF. This was tested in a series of surgical small intestinal resection specimens. Design Small bowel specifically excluding inflamed tissue, was obtained from pediatric patients (aged 2 days to 19 years) undergoing intestinal resection. EBF was assessed in Ussing chambers for transepithelial resistance (TER) and passage of FITC-Dextran (4kD). Tight junction and adherence junction proteins were imaged with immunofluorescence staining. Expression of Toll like receptors (TLR) and inflammatory cytokines were measured in loop ileostomy takedowns in a second group of patients. Results Because TER increased with patient age (p<0.01), results were stratified into infant versus teenage groups. Fed bowel had significantly greater TER versus unfed bowel (p<0.05) in both age populations. Loss of EBF was also observed by an increase in FITC-Dextran permeation in nutrient-deprived segments (p<0.05). Immunofluorescence staining showed marked declines in intensity of ZO-1, occludin, Ecadherin and Claudin-4 in unfed intestinal segments, as well as a loss of structural formation of tight junctions. Analysis of cytokine and TLR expression showed significant increases in TNF-α and TLR4 in unfed segments of bowel compared to fed segments from the same individual. Conclusion EBF declined in unfed segments of human small bowel. This work represents the first direct examination of EBF from small bowel derived from nutrient-deprived humans and may explain the increased infectious complications seen in patients not receiving enteral feeds. PMID:25704423

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

    PubMed

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

    2012-11-01

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

  4. 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

  5. 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

  6. 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

  7. 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

  8. 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

  9. Reinforced Epithelial Barrier Integrity via Matriptase Induction with Sphingosine-1-Phosphate Did Not Result in Disturbances in Physiological Redox Status.

    PubMed

    Pászti-Gere, E; Jerzsele, Á; Balla, P; Ujhelyi, G; Székács, A

    2016-01-01

    Objectives. The relationship among matriptase function, cellular redox status, and maintenance of intestinal barrier integrity has not been established yet. The aim of this study is to reveal if the crosstalk between matriptase activators and intestinal epithelial monolayers can lead to perturbations in physiological redox regulation in vitro. Methods. The effects of suramin and sphingosine-1-phosphate (S1P) were tested on viability of intestinal porcine epithelial IPEC-J2 cells using MTS assay. Measurements of transepithelial electrical resistance (TER) were performed to determine changes in barrier integrity of cell monolayers. Amplex Red assay was used to monitor extracellular hydrogen peroxide production. Occludin distribution pattern was detected prior to and after matriptase activation using immunofluorescent staining technique. Results. TER reduction was observed in suramin-treated IPEC-J2 cell monolayers, which could be attributed to cell cytotoxic properties of 48 hr 50 μM suramin administration. In contrast, S1P treatment increased TER significantly and elevated occludin accumulation in tight junctions. It was also found that extracellular hydrogen peroxide levels were maintained in IPEC-J2 cells exposed to matriptase activators. Discussion. S1P administration not accompanied by redox imbalance might be one of the key strategies in the improvement of barrier function and consequently in the therapy of intestinal inflammations. PMID:26823955

  10. Mucosal vaccines

    PubMed Central

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

    2014-01-01

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