Constitutive Expression of P-Glycoprotein in Normal Lung Alveolar Epithelium and Functionality in Primary Alveolar Epithelial Cultures

Campbell, Lee Ann; Abulrob, Abedelnasser; Kandalaft, Lana E.; Plummer, Steven; Hollins, Andrew John; Gibbs, A. R.; Gumbleton, Mark

doi:10.1124/jpet.102.042994

Cited by 94 publications

(91 citation statements)

References 39 publications

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“…Although it is possible to isolate primary rodent AT1 cells, to our knowledge, there are no methods to isolate human AT1 cells (24). An alternative is to use primary AT2 cells that have lost their phenotype in vitro to resemble AT1 cells (28,36). Recently, AT1-like cells have been obtained from transdifferentiating human fetal primary AT2 cells (37).…”

Section: Discussionmentioning

confidence: 99%

Immortalization of Human Alveolar Epithelial Cells to Investigate Nanoparticle Uptake

Kemp

Thorley

Gorelik

et al. 2008

Am J Respir Cell Mol Biol

131

117

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Primary human alveolar type 2 (AT2) cells were immortalized by transduction with the catalytic subunit of telomerase and simian virus 40 large-tumor antigen. Characterization by immunochemical and morphologic methods demonstrated an AT1-like cell phenotype. Unlike primary AT2 cells, immortalized cells no longer expressed alkaline phosphatase, pro-surfactant protein C, and thyroid transcription factor-1, but expressed increased caveolin-1 and receptor for advanced glycation end products (RAGE). Live cell imaging using scanning ion conductance microscopy showed that the cuboidal primary AT2 cells were approximately 15 mm and enriched with surface microvilli, while the immortal AT1 cells were attenuated more than 40 mm, resembling these cells in situ. Transmission electron microscopy highlighted the attenuated morphology and showed endosomal vesicles in some immortal AT1 cells (but not primary AT2 cells) as found in situ. Particulate air pollution exacerbates cardiopulmonary disease. Interaction of ultrafine, nano-sized particles with the alveolar epithelium and/or translocation into the cardiovasculature may be a contributory factor. We hypothesized differential uptake of nanoparticles by AT1 and AT2 cells, depending on particle size and surface charge. Uptake of 50-nm and 1-mm fluorescent latex particles was investigated using confocal microscopy and scanning surface confocal microscopy of live cells. Fewer than 10% of primary AT2 cells internalized particles. In contrast, 75% immortal AT1 cells internalized negatively charged particles, while less than 55% of these cells internalized positively charged particles; charge, rather than size, mattered. The process was rapid: one-third of the total cellassociated negatively charged 50-nm particle fluorescence measured at 24 hours was internalized during the first hour. AT1 cells could be important in translocation of particles from the lung into the circulation.

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Section: Discussionmentioning

confidence: 99%

Immortalization of Human Alveolar Epithelial Cells to Investigate Nanoparticle Uptake

Kemp

Thorley

Gorelik

et al. 2008

Am J Respir Cell Mol Biol

131

117

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“…In addition, MDR1 is localized to the apical membranes of epithelial cells lining the human bronchus [11], as well as the alveolar epithelium of human and rat lung [12]. MDR1 transports numerous structurally diverse therapeutic drugs, as well as insecticides [13].…”

Section: Introductionmentioning

confidence: 99%

Regulatory mechanisms to control tissue α-tocopherol

Mustacich

Elias

et al. 2007

Free Radical Biology and Medicine

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To test the hypothesis that hepatic regulation of α-tocopherol metabolism would be sufficient to prevent over-accumulation of α-tocopherol in extrahepatic tissues and that administration of high doses of α-tocopherol would up-regulate extrahepatic xenobiotic pathways, rats received daily subcutaneous injections of either vehicle or 0.5, 1, 2, or 10 mg α-tocopherol/100 g body wt for 9 days. Liver α-tocopherol increased 15-fold in rats given 10 mg α-tocopherol/100 g body weight (mg/ 100 g) compared with controls. Hepatic α-tocopherol metabolites increased with increasing α-tocopherol doses, reaching 40-fold in rats given the highest dose. In rats injected with 10 mg/100 g, lung and duodenum α-tocopherol concentrations increased 3-fold, while α-tocopherol concentrations of other extrahepatic tissues increased 2-fold or less. With the exception of muscle, daily administration of less than 2 mg/100 g) failed to increase α-tocopherol concentrations in extrahepatic tissues. Lung cytochrome P450 3A and 1A levels were unchanged by administration of α-tocopherol at any dose. In contrast, lung P-glycoprotein (MDR-1) levels increased dose dependently and expression of this xenobiotic transport protein was correlated with lung α-tocopherol concentrations (R 2 = 0.88, P < 0.05). Increased lung MDR1 may provide protection from exposure to environmental toxins by increasing alveolar space α-tocopherol.

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“…Using these data, a [B-A/A-B] permeability ratio can be determined that is relatively high, averaging 14.6 and 19.7 for the FE and SE diastereoisomers, respectively. Using the classic P-gp substrate vinblastine, we have previously reported [B-A/A-B] permeability ratios of approximately 4 in these Caco-2 cell monolayers (Campbell et al, 2003). Inclusion of the P-gp inhibitor verapamil (50 M) in both apical and basal chambers abolished the profound polarized p-Me pronucleotide transport, with the A-B permeability coefficients for the FE and SE diastereoisomers increasing to 4.7 and 6.7 (ϫ10 Ϫ6 cm/s), respectively, and the B-A permeability coefficients decreasing to 2.1 and 3.1 (ϫ10 Ϫ6 cm/s), respectively (Fig.…”

Section: Resultsmentioning

confidence: 99%

Stereoselective and Concentration-Dependent Polarized Epithelial Permeability of a Series of Phosphoramidate Triester Prodrugs of d4T: An in Vitro Study in Caco-2 and Madin-Darby Canine Kidney Cell Monolayers

Siccardi¹,

Kandalaft²,

Gumbleton³

et al. 2003

J Pharmacol Exp Ther

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Nucleoside analogs are successful, widely used antiviral and anticancer therapeutics. Nucleotide prodrugs (i.e., pronucleotide) have increasingly been used to improve in vivo efficacy of nucleoside analogs. In this study, we evaluated the permeability of a series of phosphoramidate triester prodrugs of the anti-HIV drug 2Ј,3Ј-didehydro-2Ј,3Ј-dideoxythymidine across monolayers of Caco-2, Madin-Darby canine kidney (MDCKII) epithelial cell line, and its recombinant clone containing the human MDR1/P-gp gene (MDR1-MDCKII). Transport was studied in the apical-to-basolateral (A-B) and the basolateral-toapical directions (B-A). The impact upon transport of differences in stereochemistry at the chiral phosphate center was evaluated. In the Caco-2 and MDCK models the A-B permeability was lower than expected based on the lipophilicity of the compounds, suggesting the involvement of a polarized efflux system and/or metabolic degradation in limiting the absorption of these ester-based prodrugs. Average permeability values through cell monolayers obtained in the A-B direction were lower than in the B-A direction. The inclusion of the P-glycoprotein (P-gp) inhibitor verapamil in the transport medium markedly increased the permeability in the A-B direction, whereas decreasing it in the opposite direction, suggesting an efflux mechanism mainly mediated by P-gp. Stereoselective permeability was significant for the most lipophilic compounds, where the diastereoisomer possessing the slower eluting time on a reverse-phase high-performance liquid chromatography column was transported through Caco-2 and MDCK monolayers at higher rate.

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Constitutive Expression of P-Glycoprotein in Normal Lung Alveolar Epithelium and Functionality in Primary Alveolar Epithelial Cultures

Cited by 94 publications

References 39 publications

Immortalization of Human Alveolar Epithelial Cells to Investigate Nanoparticle Uptake

Immortalization of Human Alveolar Epithelial Cells to Investigate Nanoparticle Uptake

Regulatory mechanisms to control tissue α-tocopherol

Stereoselective and Concentration-Dependent Polarized Epithelial Permeability of a Series of Phosphoramidate Triester Prodrugs of d4T: An in Vitro Study in Caco-2 and Madin-Darby Canine Kidney Cell Monolayers

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