Papilloma virus immortalized tracheal epithelial cells retain a well-differentiated phenotype

Abstract

Human airway epithelial cell lines that retain phenotypic properties representative of the native tissue will be useful physiological models. Human papilloma viral (HPV) genes can immortalize human genital keratinocytes and breast and bronchial epithelia. We transfected cystic fibrosis (CF) and normal tracheobronchial epithelial cell cultures with DNA encoding the HPV-18 E6 and E7 genes and characterized phenotypic properties of resultant cell lines. Of the 11 CF clones isolated, 6 developed a polarized phenotype with vectorial ion transport and membrane-specific expression of histamine and purinergic receptors. The ion transport properties of these lines differed from the normal lines and approximated those of primary CF airway epithelial cell cultures more closely than do those of cell lines transformed with the simian virus 40 large T gene. When transplanted into denuded tracheal grafts, these cells can differentiate into ciliated and secretory phenotypes. We conclude that HPV-18 E6 and E7 genes are sufficient to transform human airway epithelial cells and that the resultant cell lines express differentiated phenotypic properties that approximate those of the native epithelium.

DUOX1 mediates persistent epithelial EGFR activation, mucous cell metaplasia, and airway remodeling during allergic asthma

“…Healthy volunteers had no history of rhinitis or asthma, negative skin tests, and negative methacholine challenge tests. Cultured NECs or bronchial epithelial HBE1 cells (59) were analyzed for protein cysteine oxidation and EGFR activation, and the role of DUOX1 was determined using genetic or pharmacologic inhibitor approaches. The importance of DUOX1 in allergic asthma was also evaluated in a mouse model of HDM-induced allergic inflammation using both male and female C57BL/6J wild-type mice (The Jackson Laboratory) and DUOX1-deficient mice, originally generated using a retroviral-based gene-trapping method (Lexicon Pharmaceuticals) and backcrossed onto a C57BL/6J background (60).…”

“…For experiments, NECs were plated on collagen-coated 24-well culture plates (Costar) and cultured in a 1:1 mixture of bronchial epithelial cell basic medium and DMEM-H with SingleQuot supplements (Cambrex), bovine pituitary extracts (13 mg/ml), bovine serum albumin (1.5 μg/ml), and nystatin (20 units). The immortalized bronchial epithelial cell line HBE1 was maintained as described previously (23,59). For in vitro cell studies, cells were grown to confluence in 24-well culture plates (Corning) and incubated overnight in full media without EGF to suppress basal EGFR activity, prior to treatment with indicated stimuli or inhibitors.…”

Extracellular acidification elicits a chloride current that shares characteristics with ICl(swell)

“…14) and human tracheal epithelial (HTE) cells (Ref. 47) were grown in DMEM (GIBCO Life Technologies) with 10% fetal bovine serum (GIBCO Life Technologies) and 1% gentamicin (Sigma) in flasks coated with a solution containing 1 mg of human fibronectin (Stratech), 0.33 mg Vitrogen (Imperial Laboratories), and 10 mg bovine serum albumin (ICN Flow) in 100 ml of MEM with Earle's salts (GIBCO Life Technologies). The NIH3T3-MDR1 cell line, a derivative of the mouse NIH 3T3 fibroblast cell line permanently transfected with the human multidrug resistant gene (MDR1) coding for P-gp (34), was grown in DMEM with Glutamax, 10% FBS, 1% P/S, and 1 g/ml colchicine.…”

Retroviral Gene Transfer Is Inhibited by Chondroitin Sulfate Proteoglycans/Glycosaminoglycans in Malignant Pleural Effusions

“…These cell lines were maintained in RPMI 1640 medium (Life Technologies, Inc.) supplemented with 10% fetal bovine serum and penicillin/ streptomycin (R 10 ). CFT1 cells (8), a human bronchial epithelial cell line that is efficiently transduced by gibbon ape leukemia virus (GALV) and amphotropic retroviral vectors (9), were maintained in defined serumfree medium.…”