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  • 1
    ISSN: 1573-904X
    Keywords: tracheal epithelial cells ; air-interfaced culture ; ion transport ; drug transport ; permeability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Purpose. The objective of this study was to investigate how culture conditions would affect the morphological, functional, and permeability characteristics of rabbit tracheal epithelial cell layers being considered for drug transport studies. Methods. Rabbit tracheocytes were isolated by protease treatment and plated onto collagen-treated permeable supports at 1.3 × 106 cells/cm2. After 24 hr, cell layers were cultured either air-interfaced (AIC) on their apical surface or under conventional liquid covered conditions (LCC). Results. Scanning electron microscopy revealed mature cilia in AIC cell layers and ciliated cells denuded of cilia in LCC cell layers. Compared with LCC, AIC cell layers (n = 20) achieved a significantly higher peak equivalent short-circuit current (74.1 ± 6.5 vs. 51.6 ± 3.4 µA/cm2), a higher potential difference (70.9 ± 2.8 vs. 60.5 ± 3.0 mV), and a lower peak transepithelial electrical resistance (1.1 ± 0.03 vs. 1.5 ± 0.02 kohms,cm2). About 70% of the short-circuit current in AIC was amiloride-sensitive whereas 〈10% was furosemide-sensitive, similar to that found in native tissue. The corresponding values in LCC were 50% and 46%. The permeability of both AIC and LCC to lipophilic solutes (dexamethasone and propranolol) was similar, whereas the permeability of hydrophilic solutes (mannitol, sucrose, and albuterol) in AIC was only half that in LCC. Conclusions. Given the striking similarity in morphological and functional characteristics of the AIC to those in the in vivo situation, the AIC is favored as an in vitro model for future drug transport studies.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1573-904X
    Keywords: human alveolar epithelium ; primary cell culture ; lectin binding ; histochemical characterization ; drug transport
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Purpose. To develop a cell culture model of human alveolar epithelial cells in primary culture for the in vitro study of pulmonary absorption and transport. Methods. Type II pneumocytes isolated from normal human distal lung tissue by enzyme treatment and subsequent purification were plated on fibronectin/collagen coated polyester filter inserts, and cultured using a low-serum growth medium. Characterization of the cell culture was achieved by bioelectric measurements, cell-specific lectin binding, immunohistochemical detection of cell junctions, and by assessment of transepithelial transport of dextrans of varying molecular weights. Results. In culture, the isolated cells spread into confluent monolayers, exhibiting peak transepithelial resistance of 2,180 ± 62 Ω X cm2 and potential difference of 13.5 ± 1.0 mV (n = 30−48), and developing tight junctions as well as desmosomes. As assessed by lectin-binding, the cell monolayers consisted of mainly type I cells with some interspersed type II cells, thus well mimicking the situation in vivo. The permeability of hydrophilic macromolecular FITC-dextrans across the cell monolayer was found to be inversely related to their molecular size, with Papp values ranging from 1.7 to 0.2 X 10−8 cm/sec. Conclusions. A primary cell culture model of human alveolar epithelial cells has been established, which appears to be a valuable in vitro model for pulmonary drug delivery and transport studies.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1573-0603
    Keywords: transport ; epithelium ; edema ; beta-agonist ; barrier ; permeability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary To investigate the cell physiologic and biological properties of the alveolar epithelium, we studied rat alveolar epithelial cell monolayers grown on permeable supports in primary culture. Type II alveolar epithelial cells were disaggregated using elastase, and partially purified on a discontinuous metrizamide gradient. These isolated cells were plated onto tissue culture-treated Nuclepore membrane filters at 1.5×106 cells/cm2 and maintained in a humidified incubator (5% CO2 in air, 37° C). After 2 days in culture, the bathing media on both sides of the cell monolayers were changed to fresh culture medium, thus removing nonadherent cells (mostly leukocytes). These monolayers exhibit a high transmonolayer resistance (〉2000 Ω-cm2) and actively transport ions. Radionuclide flux studies indicate that Na+ is the predominant ionic species absorbed actively under baseline conditions, accounting for about 80% of the total active ion transport. Cl− seems to be passively transported across the epithelium. However, when the epithelium is exposed to a beta-agonist (terbutaline), active absorption of Na+ is increased and active absorption of Cl− occurs. Although it is clear that both active Na+ and Cl− transport are dependent on Na+/K+-ATPase activity, and that Na+ enters cells predominantly through channels, the specific mechanisms by which Cl− enters and exits the alveolar epithelial cells remain unclear. The stimulated reabsorption of Na+ and Cl− may be important in helping to remove excess fluid from alveolar air spaces in the lung.
    Type of Medium: Electronic Resource
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