CFTR and differentiation markers expression in non-CF and delta F 508 homozygous CF nasal epithelium.

Abstract

Human nasal polyps from non-CF and AF 508 homozygous CF patients were used to compare the expression of CFTR and markers of epithelial differentiation, such as cytokeratins (CK) and desmoplakins (DP), at the transcriptional and translational levels. mRNA expression was assessed by semiquantitative RT/PCR kinetic assays while the expression and distribution of proteins were evaluated by immunofluorescence analysis. In parallel, for each nasal tissue specimen, the importance of surface epithelium remodeling and inflammation was estimated after histological observations. Our results show that the steady-state levels of CFTR, CK13, CK18, CK14, or DP 1 mRNA transcripts in AF 508 CF nasal polyps were not significantly different from those of non-CF tissues. A variability in the CFTR mRNA transcript level and in the pattern of CFTR immunolabeling has been observed between the different tissue samples. However, no relationship was found between the level of CFTR mRNA transcripts and the CFTR protein expression and distribution, either in the non-CF or in the CF group. The histological observations of non-CF and CF nasal polyp tissue indicated that the huge variations in the expression and distribution of the CFTR protein were associated with the variations in the degree of surface epithelium remodeling and inflammation in the lamina propria. A surface epithelium, showing a slight basal cell hyperplasia phenotype associated with diffuse inflammation, was mainly characterized by a CFTR protein distribution at the apex of ciliated cells in both non-CF and CF specimens. In contrast, in a remodeled surface epithelium associated with severe inflammation, CFTR protein presented either a diffuse distribution in the cytoplasm of ciliated cells, or was absent. These results suggest that abnormal expression and distribution of the CFTR protein in CF airways is not only caused by CFTR mutations. Airway surface epithelium remodeling and inflammation could play a critical role in the posttranscriptional and/ or the posttranslational regulation of the CFIR protein expression in non-CF and CF airways. (J. Clin. Invest. 1995Invest. . 95:1601Invest. -1611

α7 Nicotinic Acetylcholine Receptor Regulates Airway Epithelium Differentiation by Controlling Basal Cell Proliferation

“…20 In this model, we observed a progressive regeneration of pseudostratified epithelium after 4 to 6 weeks ( Figure 3A). By using immunofluorescence and RT-PCR techniques, we observed that the expression of ␣7 nAChR in the basal layers of the epithelium was maximal in steps II and III (appearance of the first ciliated cells) and paralleled the expression of CK14, a marker of airway basal cells, 29 suggesting that ␣7 nAChR expression in basal cells is essential during the initial step of epithelial differentiation and the establishment of the basal cell layer. Thereafter, the expression became progressively restricted to one single layer of basal cells for ␣7 nAChR or isolated basal cells for CK14 at a time where the epithelium was pseudostratified and continuously expressed ␤-tubulin at the apical membrane (steps IV and V).…”

Characterization of Wild-Type and ΔF508 Cystic Fibrosis Transmembrane Regulator in Human Respiratory Epithelia

“…Because of the potential for insensitivity of immunolocalization, we supplemented these studies with biochemical studies of ⌬F508 CFTR protein maturation and bioelectric studies of function in freshly excised CF compared with normal tissues. Finally, to control for nonspecific effects of in situ inflammation on ⌬F508 CFTR processing (Dupuit et al, 1995), we repeated key studies in cell culture.…”

“…Second, we applied multiple, complementary techniques to the same specimens. Finally, the ability to culture epithelial cells from aliquots of the same freshly excised specimens used for localization and protein studies provided an alternative strategy to minimize the effects of inflammation on CFTR expression (Dupuit et al, 1995).…”

Cystic Fibrosis F508del Patients Have Apically Localized CFTR in a Reduced Number of Airway Cells

“…Three different and previously used (Brezillon et al, 1995;Crawford et al, 1991;Denning et al, 1992a;1992b;Dupuit et al, 1995;Puchelle et al, 1992) anti-CFTR Abs, raised against different epitopes, were used in the present study to identify the intracellular localization of CFTR and F508del-CFTR in freshly obtained cells from the nasal epithelium. The specificity of the Abs was also confirmed under our immunocytochemistry protocol in CFPAC and CFPAC-PLJ-6-CFTR cell lines (Fig.…”