Helicobacter pylori-Induced Disruption of Monolayer Permeability and Proinflammatory Cytokine Secretion in Polarized Human Gastric Epithelial Cells

University of Maryland, Mucosal Biology Research Center, Baltimore.
Infection and immunity (Impact Factor: 3.73). 01/2013; 81(3). DOI: 10.1128/IAI.01406-12
Source: PubMed


Helicobacter pylori infection of the stomach is related to the development of diverse gastric pathologies. The ability of H. pylori to compromise epithelial junctional complexes and to induce proinflammatory cytokines is believed to contribute to pathogenesis.
The purpose of this study was to use an in vitro human gastric epithelial model to investigate the ability of H. pylori to affect permeability and the extent and polarity of the host inflammatory response. NCI-N87 monolayers were cocultured
with live or heat-killed H. pylori or culture supernatants. Epithelial barrier function was measured by transepithelial electric resistance (TEER) analysis,
diffusion of fluorescein isothiocyanate (FITC)-labeled markers, and immunostaining for tight junction proteins. Supernatants
from both apical and basolateral chambers were tested for cytokine production by multiplex analysis. H. pylori caused a significant decrease in TEER, an increased passage of markers through the infected monolayer, and severe disruption
and mislocalization of ZO-1 and claudin-1 proteins. Cell viability was not altered by H. pylori, indicating that loss of barrier function could be attributed to a breakdown of tight junction integrity. Significantly high
levels of cytokine secretion were induced by either viable or heat-killed H. pylori. H. pylori affects monolayer permeability of polarized human gastric epithelial cells. Proinflammatory cytokines were secreted in a
polarized manner, mostly basolaterally. Live bacteria are required for disruption of tight junctions but not for the induction
of cytokine secretion. The NCI-N87 cell line provides an excellent model for the in vitro study of H. pylori pathogenesis and the epithelial cell host response to infection.

Download full-text


Available from: Alessio Fasano, Jun 25, 2014
  • Source
    • "Claudin-5 can be found at the apical membrane of the rat podocytes of puromycin aminonucleoside (PAN) induced nephrosis [16], and claudin-1 can be found in basolateral membrane of rat epididymis [18]. Various factors, such as interleukin-1β [19], helicobacter pylori [20], and dexamethasone [21], are known to affect the expression and localization of claudin. Moreover, Dhawan et al. demonstrated high expression of claudin-1 in human colorectal cancer tissues and that nuclear and cytoplasmic mislocalization of claudin-1 was frequently seen as compared to the normal mucosa [22]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The origin of crescent forming cells in human glomerulonephritis (GN) remains unknown. Some animal studies demonstrated that parietal epithelial cells of Bowman's capsule (PECs) were the main component of proliferating cells and PEC-specific tight junction protein claudin-1 was expressed in crescentic lesions. We investigated the expression of claudin-1 in human GN. Immunohistochemistry for claudin-1 was performed on 17 kidney biopsy samples with crescent formation. Colocalization of claudin-1 with intracellular tight junction protein ZO-1 was also evaluated by immunofluorescence double staining. Claudin-1 is expressed mainly at the cell to cell contact site of proliferating cells in cellular crescentic lesions in patients with these forms of human GN. Small numbers of crescent forming cells showed extrajunctional localization of claudin-1. Colocalization of claudin-1 with ZO-1 was found at cell to cell contact sites of adjacent proliferating cells. In control samples, staining of claudin-1 was positive in PECs, but not in podocytes. Our findings suggest that claudin-1 contributes to crescent formation as a component of the tight junction protein complex that includes ZO-1. Co-localization of claudin-1 with ZO-1 implies the formation of functional tight junction complexes in crescentic lesions to prevent the interstitial damage caused by penetration of filtered molecules from Bowman's space.
    International Journal of Nephrology 04/2014; 2014:598670. DOI:10.1155/2014/598670
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Helicobacter pylori contains a pathogenicity island, cagPAI, with genes homologous to components of the type IV secretion system (T4SS) of Agrobacterium tumefaciens. The T4SS components assemble a structure that transfers CagA protein and peptidoglycan into host epithelial cells, causing the increased release of interleukin 8 (IL8) from the cells. The Toll-like receptors on neutrophils recognize H. pylori, initiating signaling pathways that enhance the activation of NF-κB. However, the roles of cagPAI and T4SS in the inflammatory response of neutrophils are unknown. We evaluated the participation of cagPAI and T4SS in the response of human neutrophils to H. pylori infection. Neutrophils were isolated from the blood of healthy donors and infected with H. pylori cagPAI(+), cagPAI(-), and cagPAI mutant strains virB4 (-) and virD4 (-). Whereas cagPAI(+) strain 26695 induced the greatest IL8 production, a proinflammatory response, cagPAI(-) strain 8822 induced the greatest IL10 production, an anti-inflammatory response. In contrast, the virB4 (-) and virD4 (-) mutant strains produced significantly more of the two proinflammatory cytokines IL1β and tumor necrosis factor αthan the cagPAI(+) strain 26695. We observed that H. pylori downregulated the expression of TLRs 2 and 5 but upregulated TLR9 expression in a cagPAI and T4SS-independent manner. These results show for the first time that the response of human neutrophils to H. pylori may vary from a pro-inflammatory to an anti-inflammatory response, depending on cagPAI and the integrity of T4SS.
    PLoS ONE 06/2013; 8(6):e64623. DOI:10.1371/journal.pone.0064623 · 3.23 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Helicobacter pylori (Hp) infection is an important cause of chronic gastritis and peptic ulcer, but their pathogenesis is unclear. The role of gastric mucosal barrier dysfunction induced by impaired structure and function of tight junction in the pathogenesis of Hp-associated gastric diseases has received considerable attention in recent years. Tight junction is composed of a variety of proteins and molecules, including 3 integral membrane proteins (occludin, claudins, and junctional adhesion molecules) and a cytoplasmic protein (zonula occludens). This paper mainly describes the composition and function of various tight junction proteins, changes in tight junction protein function induced by Hp infection and their relationship with the incidence of gastric diseases, and the significance of enhancing the tight junction protein function in the prevention and treatment of Hp-associated gastric diseases.
    Zhongguo dang dai er ke za zhi = Chinese journal of contemporary pediatrics 03/2014; 16(3):242-7. DOI:10.7499/j.issn.1008-8830.2014.03.005
Show more