Article

Developmental regulation of claudin localization by fetal alveolar epithelial cells.

Department of Physiology, Institute for Environmental Medicine, University of Pennsylvania School of Medicine, B-400 Richards Bldg./6085, 3700 Hamilton Walk, Philadelphia, PA 19104, USA.
AJP Lung Cellular and Molecular Physiology (Impact Factor: 3.52). 12/2004; 287(6):L1266-73. DOI: 10.1152/ajplung.00423.2003
Source: PubMed

ABSTRACT Tight junction proteins in the claudin family regulate epithelial barrier function. We examined claudin expression by human fetal lung (HFL) alveolar epithelial cells cultured in medium containing dexamethasone, 8-bromo-cAMP, and isobutylmethylxanthanine (DCI), which promotes alveolar epithelial cell differentiation to a type II phenotype. At the protein level, HFL cells expressed claudin-1, claudin-3, claudin-4, claudin-5, claudin-7, and claudin-18, where levels of expression varied with culture conditions. DCI-treated differentiated HFL cells cultured on permeable supports formed tight transepithelial barriers, with transepithelial resistance (TER) >1,700 ohm/cm(2). In contrast, HFL cells cultured in control medium without DCI did not form tight barriers (TER <250 ohm/cm(2)). Consistent with this difference in barrier function, claudins expressed by HFL cells cultured in DCI medium were tightly localized to the plasma membrane; however, claudins expressed by HFL cells cultured in control medium accumulated in an intracellular compartment and showed discontinuities in claudin plasma membrane localization. In contrast to claudins, localization of other tight junction proteins, zonula occludens (ZO)-1, ZO-2, and occludin, was not sensitive to HFL cell phenotype. Intracellular claudins expressed by undifferentiated HFL cells were localized to a compartment containing early endosome antigen-1, and treatment of HFL cells with the endocytosis inhibitor monodansylcadaverine increased barrier function. This suggests that during differentiation to a type II cell phenotype, fetal alveolar epithelial cells use differential claudin expression and localization to the plasma membrane to help regulate tight junction permeability.

0 Bookmarks
 · 
99 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Eine intakte alveolar-epitheliale Barriere ist für eine ungestörte Gasaustauschfunktion der Lunge von maßgeblicher Bedeutung. Beeinträchtigungen sowohl der passiven wie auch aktiven Barrierefunktion können hier zu einem folgenschweren Organversagen führen. Der Einfluss alveolärer Hypoxie auf aktive Resorptionsmechanismen des alveolären Epithels wurde bisher unzureichend unter physiologischen Bedingungen untersucht. Im vorliegenden ex-vivo Modell der isolierten, perfundierten und ventilierten Kaninchenlunge wurde der Einfluss von Hypoxie und/oder alveolärer Flüssigkeitsbeladung auf die Funktion des alveolären Epithels untersucht. Der das Epithel auskleidende Flüssigkeitsfilm (epithelial lining fluid (ELF)) wird über aktiven und passiven transmembranösen Flüssigkeitstransport (alveolar fluid clearence (AFC)) reguliert. In dem hier verwendeten Modell konnten wir zeigen, dass der alveoläre Natriumtransport die treibende Kraft des AFC darstellt. Der aktive, durch apikale Amilorid-sensitive Natriumkänale (ENaC) und basolateral lokalisierte Na,KATPase von alveolär nach interstitiell/intravasal gerichtete Natriumgradient führt zu einem konsekutiven Nachströmen von Wasser entlang des Ionengradienten. Die Deposition eines Flüssigkeitsbolus von 2,5 ml per Aerosolierung (engl. excess fluid load) führte im verwendeten Modell zu keiner relevanten Funktionsbeeinträchtigung des isolierten Organs. Es zeigte sich überraschenderweise sogar eine Stimulation des aktiven Natrium- und somit auch Flüssigkeitstransportes, im Sinne einer gesteigerten Ödemresorption. Demgegenüber führte eine alveoläre Hypoxie (zwei Stunden Ventilation mit 3,0 % Sauerstoff) zu einem signifikanten Abfall des AFC und konsekutiver Ödembildung. Ursache hierfür war eine um ca. 60 % reduzierte aktive Natriumresorption mit entsprechender Senkung des Flüssigkeitstransportes. Wir konnten somit erstmals in einem intakten isolierten Lungenmodell (unter Beibehaltung der Ventilation) zeigen, dass alveoläre Hypoxie, nicht jedoch eine iatrogene Flüssigkeitsbeladung des Alveolarraumes zu einer Störung der Ödemresorption der Lunge führt. Diese Beobachtung hat Implikationen für das Verständnis von Krankheitsbildern wie dem akuten Lungenversagen, dem Höhenlungenödem und dem Beinahe-Ertrinken. Alveolar-capillary barrier function is essential to maintain alveolar fluid balance and adequate gas exchange in the lung. Malfunction of either passive and/or active barrier could resolve in a detrimental organ failure So far the influence of alveolar hypoxia on active barrier function and its underlying machinery has been only inadequately described in a physiological environment. Using the ex-vivo model of the isolated, perfused and ventilated rabbit lung, this experimental study evaluated the influence of hypoxia and/or excess fluid load on alveolar epithelium. The epithelial lining fluid (ELF) is balanced via active and passive transmembrane alveolar fluid clearance (AFC). In the present study using the isolated, perfused and ventilated rabbit lung alveolar sodium transport (AST) was evidenced to be the driving force of the AFC. Sodium is actively pumped out of the alveolar epithelial cells into the interstitium by Na,K-ATPase, located on the basolateral membrane of the epithelium, which in turn drives the sodium uptake by amiloride-sensitive sodium channels (ENaCs), located on the apical membrane of the epithelium. This generates an osmotic gradient that drives water out of the alveolar space. The deposition of 2,5 ml excess fluid load resulted in no relevant barrier dysfunction in our isolated organ model. Surprisingly, we found the active sodium- and therefore fluid transport stimulated, ascribed by enhanced edema resorption. In contrast, alveolar hypoxia (2 hours of ventilation with 3,0 % Oxygen) led to a significant decrease of AFC and consecutive alveolar edema formation. Etiologic was the reduction of active AST by about 60 %, correspondingly resulting in a decreased AFC. Taken together, the deleterious effect of alveolar hypoxia, but not experimental alveolar deposition of excess fluid load, on the edema clearence in the distal lung was proofed for the first time in an intact isolated lung model under perpetuated ventilation. Our results establish ramifications for the disease pattern of acute lung injury (ALI), adult respiratory distress syndrome (ARDS), high altitude pulmonary edema (HAPE) and near-drowning syndrome.
    Giessen : VVB Laufersweiler.
  • [Show abstract] [Hide abstract]
    ABSTRACT: Claudin expression is altered in lung cancer, but the pathophysiological role of claudin is not well understood. We examined the effect of claudin-2 expression on cell migration using human adenocarcinoma A549 cells. The mRNA level was measured by real time polymerase chain reaction. To knockdown claudin-2 expression, we made the cells expressing doxycycline-inducible claudin-2 shRNA vector. The protein level was examined by Western blotting. Cell migration was measured by wound-healing assay. The enzymatic activity of MMP-9 was assessed by gelatin zymography. In A549 cells, claudin-2 expression was higher than in normal lung tissue. Claudin-2 knockdown did not affect the expression of other junctional proteins including claudin-1, occludin and E-cadherin. Claudin-2 knockdown decreased cell migration concomitant with a decrease in the mRNA level and enzymatic activity of MMP-9. The expression level of Sp1 in the nuclei was decreased by claudin-2 knockdown. In contrast, the expression levels of c-Fos, c-Jun and NF-kB p65 in the nuclei were not changed by claudin-2 knockdown. The knockdown of Sp1 expression by siRNA decreased cell migration, and the mRNA expression, enzymatic activity, and promoter activity of MMP-9. Claudin-2 may increase the mRNA level and enzymatic activity of MMP-9 mediated by the elevation of nuclear distribution of Sp1, resulting in the up-regulation of A549 cell migration.
    Life sciences 02/2011; 88(13-14):628-33. · 2.56 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The intestinal epithelium represents a critical barrier protecting the host against diverse luminal noxious agents, as well as preventing the uncontrolled uptake of bacteria that could activate an immune response in a susceptible host. The epithelial monolayer that constitutes this barrier is regulated by a meshwork of proteins that orchestrate complex biological function such as permeability, transepithelial electrical resistance, and movement of various macromolecules. Because of its key role in maintaining host homeostasis, factors regulating barrier function have attracted sustained attention from the research community. This paper will address the role of bacteria, bacterial-derived metabolism, and the interplay of dietary factors in controlling intestinal barrier function.
    BioMed research international. 01/2013; 2013:425146.