Characterization of pediatric cystic fibrosis airway epithelial cell cultures at the air-liquid interface obtained by non-invasive nasal cytology brush sampling

Respiratory Research

Research

Fabian Blank¹  Seraina Beyeler²  Aline Schögler³  Marco P. Alves⁴  Thomas Geiser⁵  Carmen Casaulta⁶  Melanie Brügger⁷  Stefan A. Tschanz⁸  Nicolas Regamey⁹ 

[1] Department of Clinical Research, University of Bern, Bern, Switzerland;Department of Pulmonary Medicine, University Hospital of Bern, Bern, Switzerland;Department of Clinical Research, University of Bern, Bern, Switzerland;Department of Pulmonary Medicine, University Hospital of Bern, Bern, Switzerland;Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland;Department of Clinical Research, University of Bern, Bern, Switzerland;Division of Respiratory Medicine, University Children’s Hospital of Bern, Bern, Switzerland;Department of Clinical Research, University of Bern, Bern, Switzerland;Division of Respiratory Medicine, University Children’s Hospital of Bern, Bern, Switzerland;Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland;Institute of Virology and Immunology, Federal Department of Home Affairs, Mittelhäusern, Switzerland;Department of Infectious Diseases and Pathobiology, Vetsuisse Faculty, University of Bern, Bern, Switzerland;Department of Pulmonary Medicine, University Hospital of Bern, Bern, Switzerland;Division of Respiratory Medicine, University Children’s Hospital of Bern, Bern, Switzerland;Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland;Institute of Virology and Immunology, Federal Department of Home Affairs, Mittelhäusern, Switzerland;Department of Infectious Diseases and Pathobiology, Vetsuisse Faculty, University of Bern, Bern, Switzerland;Institute of Anatomy, University of Bern, Bern, Switzerland;Lucerne Children’s Hospital, Lucerne, Switzerland;

关键词: Cystic fibrosis;    Pediatric;    Air-liquid interface;    Airway epithelium;    Cytology brush;    Nasal brushing;   

DOI  :  10.1186/s12931-017-0706-7

 received in 2017-07-28, accepted in 2017-12-18,  发布年份 2017

来源: Springer

【 摘 要 】

BackgroundIn vitro systems of primary cystic fibrosis (CF) airway epithelial cells are an important tool to study molecular and functional features of the native respiratory epithelium. However, undifferentiated CF airway cell cultures grown under submerged conditions do not appropriately represent the physiological situation. A more advanced CF cell culture system based on airway epithelial cells grown at the air-liquid interface (ALI) recapitulates most of the in vivo-like properties but requires the use of invasive sampling methods. In this study, we describe a detailed characterization of fully differentiated primary CF airway epithelial cells obtained by non-invasive nasal brushing of pediatric patients.MethodsDifferentiated cell cultures were evaluated with immunolabelling of markers for ciliated, mucus-secreting and basal cells, and tight junction and CFTR proteins. Epithelial morphology and ultrastructure was examined by histology and transmission electron microscopy. Ciliary beat frequency was investigated by a video-microscopy approach and trans-epithelial electrical resistance was assessed with an epithelial Volt-Ohm meter system. Finally, epithelial permeability was analysed by using a cell layer integrity test and baseline cytokine levels where measured by an enzyme-linked immunosorbent assay.ResultsPediatric CF nasal cultures grown at the ALI showed a differentiation into a pseudostratified epithelium with a mucociliary phenotype. Also, immunofluorescence analysis revealed the presence of ciliated, mucus-secreting and basal cells and tight junctions. CFTR protein expression was observed in CF (F508del/F508del) and healthy cultures and baseline interleukin (IL)-8 and IL-6 release were similar in control and CF ALI cultures. The ciliary beat frequency was 9.67 Hz and the differentiated pediatric CF epithelium was found to be functionally tight.ConclusionIn summary, primary pediatric CF nasal epithelial cell cultures grown at the ALI showed full differentiation into ciliated, mucus-producing and basal cells, which adequately reflect the in vivo properties of the human respiratory epithelium.

【 授权许可】

CC BY   
© The Author(s). 2017