Respiratory epithelium

About: Respiratory epithelium is a research topic. Over the lifetime, 5048 publications have been published within this topic receiving 222304 citations. The topic is also known as: respiratory tract epithelium & Respiratory Mucosa.

Adeno-Associated Virus Type 6 (AAV6) Vectors Mediate Efficient Transduction of Airway Epithelial Cells in Mouse Lungs Compared to That of AAV2 Vectors

Abstract: Transfer of therapeutic genes to the lung may provide a cure for diseases such as cystic fibrosis (CF), which affects 1 in 3,000 Caucasian births Inactivating mutations in the CF transmembrane regulator (CFTR), a chloride ion channel, result in gradual lung destruction, which is the major cause of morbidity Although the normal CFTR protein has been localized to both the apical surface of the airway epithelium (38) and the submucosal glands beneath the epithelium (10), the airway is the site of microbial obstruction associated with mortality This clinical manifestation provides the rationale for targeting the airway epithelium for CF gene therapy Among the many gene transfer systems being investigated are viral vectors such as those based on adeno-associated virus (AAV), a single-stranded DNA parvovirus AAV can integrate and promote persistent gene expression in cultured cells and in dividing and nondividing cells in multiple somatic tissues of animals (19, 20, 22–24, 26, and 32) The ability to transduce nondividing cells is an important feature of AAV vectors for gene transfer to the airway epithelium, which has a low rate of proliferation (3, 21) Additionally, the fact that wild-type (wt) AAV has been isolated from human airways (4, 5, 27) is consistent with the idea that AAV has tropism for the lung epithelium However, although AAV type 2 (AAV2) vectors can transduce multiple cell types in the lung, animal data thus far have shown low to modest rates of transduction by AAV2 vectors in the lung (11, 16, 17) Indeed, even introduction of a high dose of an AAV2 vector (1012 genome-containing particles) resulted in an overall transduction efficiency of 2% in the mouse airway epithelium (2) Animal studies show that the efficiency of AAV transduction is affected by several factors and can be enhanced by various treatments AAV transduction in the developing neonatal rabbit lung was more efficient than that in adult lung and was observed in a variety of airway and alveolar cell types (40) AAV vector-transduced cells in adult mouse lungs were rare, but their numbers could be increased by addition of adenovirus to provide helper functions (11), DNA-damaging reagents (1, 23, 30), or tissue injury (13, 16) Additionally, administration of much higher doses of AAV vector could also increase transduction in the lung epithelium (2) Tissue culture models have shown that proliferation rates (29) and polarity of epithelia influence the efficiency of AAV vectors (9) Indeed, reagents that help AAV vectors to bypass the natural resistance of the apical surface to infection can augment AAV transduction 10- to 100-fold (9, 35) These results show that dosage, proliferation rates, and target cell access are factors involved in efficient transduction by AAV vectors in lung epithelia Although AAV vector expression can persist for months to years, there may still be a need for readministration of vector to increase or replenish the population of modified cells In readministration studies, few to no new transduction events have been detected in the rabbit or mouse lung or in skeletal muscle (12, 16, 17, 37), and these results were associated with the detection of neutralizing antibodies (16, 17) Several approaches have been used to achieve effective readministration These include immune suppression in the lung (17) and muscle (25) and the use of other AAV types in the muscle and liver (36) and the lung (18) We showed that AAV vectors utilizing an AAV6 capsid (AAV6 pseudotype vector) have properties that could be useful for gene therapy, including low immunogenicity and the lack of cross-reactive antibodies generated against AAV2 (18) Quantitation of the number of vector-expressing cells in our previous study indicated that the AAV6 pseudotype vector was as efficient as, if not more efficient than, AAV2 vectors in the mouse lung and that the transduction rates in various cell types were different between AAV2 and AAV6 (18) In this study, we evaluated the transduction of lung cells by vectors based on the AAV6 serotype more thoroughly to determine the combination of vector components that mediated the most efficient transduction of airway epithelia To achieve this goal, we made separate expression plasmids for the three components of the AAV6 vector: rep, cap, and the packaged genome containing terminal repeats (TR) from AAV6 and expressing human placental alkaline phosphatase (AP) In conjunction with similar constructs for AAV2 (2), complementation of AAV2 and AAV6 components to generate infectious virions was assessed and then transduction was evaluated in a mouse model of lung gene transfer Our data show that AAV2 and AAV6 pseudotype vectors could be generated from all combinations of rep, cap, and genome from the two viruses and that transduction efficiencies of these vectors in tissue-cultured cells were primarily determined by the vector pseudotype, defined as the source of the capsid While both AAV2 and AAV6 pseudotype vectors bind heparin, only AAV2 is inhibited by heparin in cell transduction assays, indicating that AAV2 and AAV6 interact with different receptors In mouse lung delivery, the AAV2 vector gave high transduction rates in alveolar cells and much lower rates in airway epithelia, similar to the results obtained in previous studies In contrast, an AAV6 pseudotype vector showed preferential transduction of epithelial cells in large and small airways at rates up to 80% Transduction of the mouse airway epithelium by AAV6 pseudotype vectors was 15 to 74 times more efficient than transduction by an AAV2 vector These results, combined with our previous results showing lower immunogenicity of AAV6 than of AAV2, indicate that AAV6 vectors may provide significant advantages for gene therapy for CF

The innate immune function of airway epithelial cells in inflammatory lung disease

Abstract: The airway epithelium is now considered to be central to the orchestration of pulmonary inflammatory and immune responses, and is also key to tissue remodelling. It acts as the first barrier in the defence against a wide range of inhaled challenges, and is critically involved in the regulation of both innate and adaptive immune responses to these challenges. Recent progress in our understanding of the developmental regulation of this tissue, the differentiation pathways, recognition of pathogens and antimicrobial responses is now exploited to help understand how epithelial cell function and dysfunction contributes to the pathogenesis of a variety of inflammatory lung diseases. Herein, advances in our knowledge of the biology of airway epithelium, as well as its role and (dys)function in asthma, chronic obstructive pulmonary fibrosis and cystic fibrosis will be discussed.

Effects of human neutrophil elastase and Pseudomonas aeruginosa proteinases on human respiratory epithelium.

Abstract: It has been suggested that proteinase enzymes could play an important role in the pathogenesis of chronic bronchial infections including bronchiectasis and cystic fibrosis (CF). Because Pseudomonas aeruginosa frequently colonizes the respiratory tract in bronchiectasis and CF, we examined the in vitro effects of human neutrophil elastase (HNE) and proteinase enzymes produced by P. aeruginosa (elastase: PE; alkaline proteinase: PAP) on the ciliary beat frequency (CBF) and ultrastructure of human nasal ciliated respiratory epithelium. HNE (500 micrograms/ml) progressively reduced CBF and caused marked epithelial disruption; lower concentrations (100 and 20 micrograms/ml) also caused epithelial disruption but without slowing CBF. The effects of HNE (500 micrograms/ml) were completely abolished by adding alpha 1-antitrypsin (5 mg/ml). There was no synergy between HNE and pyocyanin, a product of P. aeruginosa which slows CBF. PE in phosphate-buffered saline also caused epithelial disruption without slowing CBF; however, PE in medium containing divalent metal ions caused CBF slowing as well as epithelial disruption at 100 micrograms/ml. PAP (500 micrograms/ml) had almost no effect on ciliated epithelium. The effects of HNE and PE on nasal and bronchial epithelium obtained from the same patient were similar. Light and transmission electron microscopy revealed that HNE and PE were cytotoxic and caused detachment of epithelial cells from neighboring cells and the basement membrane. There was cytoplasmic blebbing of the cell surface and mitochondrial damage; however, no increase of abnormalities in the ultrastructure of cilia on living cells was seen. These results support the hypothesis that HNE and PE contribute to the delayed mucociliary clearance and epithelial damage that is observed in patients with chronic bronchial infection.

New observations of rat airway epithelium: a quantitative and electron microscopic study.

Abstract: Epithelial thickness, depth of the ciliary layer and concentration of cells has been estimated at 5 levels of the rat airway (3 extrapulmonary and 2 intrapulmonary) and the ultrastructure of the cells described. Extrapulmonary airways have a pseudostratified epithelium, intrapulmonary airways a simple one. The epithelium thins progressively from upper to lower trachea while the epithelium of the lower trachea is thicker than at more peripheral airway levels, all of which are similar. The depth of the ciliary layer decreases peripherally. By electron microscopy 10 cell types could be identified - 8 epithelial and 2 'migratory'. A cell type not described previously has been found that resembles the serous cells of the submucosal glands. The salient features of each cell type are described. The upper trachea has a concentration of cells significantly higher than elsewhere, while the remaining four airway levels have about 230 cells per 10 high power fields (1-8 mm length epithelium). Ciliated cells are sparse proximally and increase in number progressively toward the periphery, while basal and migratory cells are most frequent proximally. At all levels of the airway, between 40% and 50% of cells are non-ciliated. In the upper trachea the most frequent cell type was the 'intermediate', elsewhere the 'epithelial serous' cell. Goblet cells were few at all levels, amounting to less than 1% of the total counted. The Clara cell was not restricted to the terminal bronchioles but was found as far proximally as the hilum.