Expression of CFTR from a ciliated cell-specific promoter is ineffective at correcting nasal potential difference in CF mice

Ostrowski, Lawrence E.; Yin, Weining; Diggs, P S; Rogers, Troy D.; O’Neal, Wanda K.; Grubb, Barbara R.

doi:10.1038/sj.gt.3302994

Cited by 33 publications

(22 citation statements)

References 43 publications

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“…However, for gene transfer studies aimed at correcting ciliated cells, the nasal PD assay may be less useful in ascertaining correction due to the robust contribution of the olfactory cells to the bioelectrics of the nasal PD. In fact, in a previous study, we have reported that expression of CFTR specifically in ciliated cells of CF mice did not alter nasal PD values (26). Because some vectors more readily transduced OE compared with CE (2,17,32) it is critical to know from which cell type a "correction" is being measured.…”

Section: Discussionmentioning

confidence: 96%

“…However, unlike human airways, in the mouse, ϳ50% of the surface area of the nasal cavity is covered by olfactory epithelium (1, 7). On the basis of previous studies, we hypothesized that a significant fraction of the bioelectric signal from the nasal epithelium may arise from the OE (15,22,26). Because defects in ion transport across the ciliated cell are thought to be responsible for the pulmonary phenotype in the human CF patient (18), it is important to determine ion transport characteristics of each epithelium independently.…”

Section: Discussionmentioning

confidence: 98%

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Ion transport across CF and normal murine olfactory and ciliated epithelium

Grubb

Rogers²,

Boucher

et al. 2009

American Journal of Physiology-Cell Physiology

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The nasal epithelium of the cystic fibrosis (CF) mouse has been used extensively in CF research because it exhibits ion transport defects similar to those of human CF airways. This tissue is composed of approximately 50% olfactory (OE) and approximately 50% ciliated epithelium (CE), and on the basis of previous observations, we hypothesized that a significant fraction of the bioelectric signals from murine nasal tissue may arise from OE rather than CE, while CE is the target tissue for CF gene therapy. We compared the bioelectric properties of isolated OE from the nasal cavity and CE from the nasopharynx in Ussing chamber studies. Hyperabsorption of Na(+) [amiloride response; CF vs. wild type (WT)] was approximately 7.5-fold greater in the OE compared with the CE. The forskolin response in native tissues did not reliably distinguish genotypes, likely due to a cyclic nucleotide-gated cation conductance in OE and a calcium-mediated Cl(-) conductance in CE. By potential difference assay, hyperabsorption of Na(+) (CF vs. WT) and the difference in response to apical 0 Cl(-) buffer (CF vs. WT) were approximately 2-fold greater in the nasal cavity compared with the nasopharynx. Our studies demonstrate that in the CF mouse, both the hyperabsorption of Na(+) and the Cl(-) transport defect are of larger magnitude in the OE than in the CE. Thus, while the murine CF nasal epithelium is a valuable model for CF studies, the bioelectrics are likely dominated by the signals from the OE, and assays of the nasopharynx may be more specific for studying the ciliated epithelium.

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Section: Discussionmentioning

confidence: 96%

Section: Discussionmentioning

confidence: 98%

Ion transport across CF and normal murine olfactory and ciliated epithelium

Grubb

Rogers²,

Boucher

et al. 2009

American Journal of Physiology-Cell Physiology

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“…Significant expression of CFTR was more recently also shown in the apical membrane of ciliated cells from healthy human beings, whereas CFTR was absent in the apical membrane of cells from CF patients homozygous for the ⌬F508 mutation (17). Expression of human CFTR (driven by the ciliated cell-specific promoter foxj1) in the trachea of a CF mouse model confirmed targeting of CFTR to the apical membrane of ciliated cells while restoring forskolinstimulated chloride secretion (18). Electrolyte conductance through CFTR is usually associated with chloride transport, but bidirectional conductance of bicarbonate through CFTR has also been demonstrated and is thought to be important in the airway and in pancreatic ducts (14, 19 -21).…”

mentioning

confidence: 87%

Decreased Soluble Adenylyl Cyclase Activity in Cystic Fibrosis Is Related to Defective Apical Bicarbonate Exchange and Affects Ciliary Beat Frequency Regulation

Schmid

Sutto

Schmid

et al. 2010

Journal of Biological Chemistry

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Adenylyl cyclases are generally thought to be transmembrane-, G-protein-, and forskolin-responsive proteins, but a nontransmembrane, soluble adenylyl cyclase (sAC) 2 has been identified (1). Mammalian sAC is not activated by G-proteins or forskolin (1) but by HCO 3 Ϫ /CO 2 in a pH-independent manner (2, 3) and by Ca 2ϩ , which synergizes with HCO 3 Ϫ (4). sAC expression has been described in many human tissues (5). We have shown that it is expressed in the airway epithelium, where it represents the only known adenylyl cyclase localized to cilia (6). sAC is important for flagellar beating in sperm (7-9), and we have shown its importance for regulating ciliary beating in human airway epithelia via cAMP production upon stimulation with HCO 3 Ϫ /CO 2 (6). There are different sources of luminal bicarbonate in the airways: it can be secreted from submucosal glands and ciliated cells (10) and can also be produced de novo from CO 2 and H 2 O by locally secreted carbonic anhydrase (11). Two proteins responsible for transporting HCO 3 Ϫ into cells, Slc26a9 (12, 13) and CFTR (14), have been described in the apical membrane of airway epithelial cells. Slc26a9 is mainly a chloride channel with very low bicarbonate permeability (12); thus, the major apical HCO 3 Ϫ exchange in human airways occurs likely through CFTR (14) or is dependent on it.In 1989, the CFTR gene was linked to cystic fibrosis (15). Initial attempts to localize the protein in the airways were based on mRNA in situ hybridization and pinpointed expression mainly to submucosal gland acini (16). Significant expression of CFTR was more recently also shown in the apical membrane of ciliated cells from healthy human beings, whereas CFTR was absent in the apical membrane of cells from CF patients homozygous for the ⌬F508 mutation (17). Expression of human CFTR (driven by the ciliated cell-specific promoter foxj1) in the trachea of a CF mouse model confirmed targeting of CFTR to the apical membrane of ciliated cells while restoring forskolinstimulated chloride secretion (18). Electrolyte conductance through CFTR is usually associated with chloride transport, but bidirectional conductance of bicarbonate through CFTR has also been demonstrated and is thought to be important in the airway and in pancreatic ducts (14, 19 -21). Regulation of HCO 3 Ϫ transport through CFTR has been related to sAC in Calu3 cells, an airway epithelial cell line (22), and in corneal endothelium (23), but there are no reports of the influence of diminished bicarbonate transport through CFTR on the activity of sAC in primary airway epithelial cells, especially as it * This work was supported, in whole or in part, by National Institutes of Health

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“…Tracheas from adult CF mice also do not exhibit defects in either cAMP-mediated Cl Ϫ secretion nor Na ϩ hyperabsorption (39), likely due to low endogenous Cftr expression in adult tracheal tissue combined with expression of alternative Cl Ϫ channels, e.g., Tmem16a (39). In contrast, neonatal murine tracheas have a more robust Cftr mRNA expression (ϳ2-fold more than adult tracheas; W. K. O'Neal, unpublished observations), and CF neonatal mice exhibit the classic defect in cAMP-mediated Cl Ϫ secretion, with a forskolin response reduced approximately fourfold compared with wild-type (WT) mice (38,39). Nonetheless, neonatal CF mice have no detectable lung disease.…”

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confidence: 92%

Loss of Cftr function exacerbates the phenotype of Na⁺ hyperabsorption in murine airways

Livraghi‐Butrico

Kelly

Wilkinson

et al. 2013

American Journal of Physiology-Lung Cellular and Molecular Phys

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Airway surface hydration depends on the balance between transepithelial Na(+) absorption and Cl(-) secretion. In adult mice, absence of functional cystic fibrosis transmembrane conductance regulator (Cftr) fails to recapitulate human cystic fibrosis (CF) lung disease. In contrast, overexpression of the epithelial Na(+) channel β subunit in transgenic mice (βENaC-Tg) produces unregulated Na(+) hyperabsorption and results in CF-like airway surface dehydration, mucus obstruction, inflammation, and increased neonatal mortality. To investigate whether the combination of airway Na(+) hyperabsorption and absent Cftr-mediated Cl(-) secretion resulted in more severe lung pathology, we generated double-mutant ΔF508 CF/βENaC-Tg mice. Survival of ΔF508 CF/βENaC-Tg mice was reduced compared with βENaC-Tg or ΔF508 CF mice. Absence of functional Cftr did not affect endogenous or transgenic ENaC currents but produced reduced basal components of Cl(-) secretion and tracheal cartilaginous defects in both ΔF508 CF and ΔF508 CF/βENaC-Tg mice. Neonatal ΔF508 CF/βENaC-Tg mice exhibited higher neutrophilic pulmonary inflammation and club cell (Clara cell) necrosis compared with βENaC-Tg littermates. Neonatal ΔF508 CF/βENaC-Tg mice also exhibited spontaneous bacterial infections, but the bacterial burden was similar to that of βENaC-Tg littermates. Adult ΔF508 CF/βENaC-Tg mice exhibited pathological changes associated with eosinophilic crystalline pneumonia, a phenotype not observed in age-matched βENaC-Tg mice. Collectively, these data suggest that the combined abnormalities in Na(+) absorption and Cl(-) secretion produce more severe lung disease than either defect alone. Airway cartilage abnormalities, airway cell necrosis, and exaggerated neutrophil infiltration likely interact with defective mucus clearance caused by βENaC overexpression and absent CFTR-mediated Cl(-) secretion to produce the increased neonatal mortality observed in ΔF508 CF/βENaC-Tg mice.

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Expression of CFTR from a ciliated cell-specific promoter is ineffective at correcting nasal potential difference in CF mice

Cited by 33 publications

References 43 publications

Ion transport across CF and normal murine olfactory and ciliated epithelium

Ion transport across CF and normal murine olfactory and ciliated epithelium

Decreased Soluble Adenylyl Cyclase Activity in Cystic Fibrosis Is Related to Defective Apical Bicarbonate Exchange and Affects Ciliary Beat Frequency Regulation

Loss of Cftr function exacerbates the phenotype of Na⁺ hyperabsorption in murine airways

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Expression of CFTR from a ciliated cell-specific promoter is ineffective at correcting nasal potential difference in CF mice

Cited by 33 publications

References 43 publications

Ion transport across CF and normal murine olfactory and ciliated epithelium

Ion transport across CF and normal murine olfactory and ciliated epithelium

Decreased Soluble Adenylyl Cyclase Activity in Cystic Fibrosis Is Related to Defective Apical Bicarbonate Exchange and Affects Ciliary Beat Frequency Regulation

Loss of Cftr function exacerbates the phenotype of Na+ hyperabsorption in murine airways

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Loss of Cftr function exacerbates the phenotype of Na⁺ hyperabsorption in murine airways