Techniques and Procedures
Expansion of Primary Bronchial Epithelial Cell Cultures
G. Margarit
, J. Belda, P. Casan, J. Sanchis
Corresponding author
gmargarit@hsp.santpau.es
Correspondence: Dra. G. Margarit. Antoni M. Claret, 167. 08025 Barcelona. España
Correspondence: Dra. G. Margarit. Antoni M. Claret, 167. 08025 Barcelona. España
Departamento de Neumología, Hospital de la Santa Creu i de Sant Pau, Facultat de Medicina, Universitat Autònoma de Barcelona, Barcelona, Spain
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López-Viña, R. Agüero-Balbín, J.L. Aller-Álvarez, T. Bazús-González, F.B. García-Cosio, A. de Diego-Damiá, E. Martínez-Moragón, A. Pereira-Vega, V. Plaza-Moral, G. Rodríguez-Trigo, J.R. Villa-Asensi" "autores" => array:12 [ 0 => array:4 [ "preGrado" => "Dr." "nombre" => "A." "apellidos" => "López-Viña" "postGrado" => "coordinator" ] 1 => array:2 [ "nombre" => "R." "apellidos" => "Agüero-Balbín" ] 2 => array:2 [ "nombre" => "J.L." "apellidos" => "Aller-Álvarez" ] 3 => array:2 [ "nombre" => "T." "apellidos" => "Bazús-González" ] 4 => array:2 [ "nombre" => "F.B." "apellidos" => "García-Cosio" ] 5 => array:2 [ "nombre" => "A." "apellidos" => "de Diego-Damiá" ] 6 => array:2 [ "nombre" => "E." "apellidos" => "Martínez-Moragón" ] 7 => array:2 [ "nombre" => "A." "apellidos" => "Pereira-Vega" ] 8 => array:2 [ "nombre" => "V." "apellidos" => "Plaza-Moral" ] 9 => array:2 [ "nombre" => "G." "apellidos" => "Rodríguez-Trigo" ] 10 => array:2 [ "nombre" => "J.R." "apellidos" => "Villa-Asensi" ] 11 => array:1 [ "colaborador" => "Assembly on Asthma of the Spanish Society of Pulmonology and Thoracic Surgery (SEPAR)" ] ] ] ] ] "idiomaDefecto" => "en" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S1579212906602723?idApp=UINPBA00003Z" "url" => "/15792129/0000004100000009/v1_201305150225/S1579212906602723/v1_201305150225/en/main.assets" ] "en" => array:15 [ "idiomaDefecto" => true "cabecera" => "<span class="elsevierStyleTextfn">Techniques and Procedures</span>" "titulo" => "Expansion of Primary Bronchial Epithelial Cell Cultures" "tieneTextoCompleto" => 0 "paginas" => array:1 [ 0 => array:2 [ "paginaInicial" => "524" "paginaFinal" => "527" ] ] "autores" => array:1 [ 0 => array:4 [ "autoresLista" => "G. Margarit, J. Belda, P. Casan, J. Sanchis" "autores" => array:4 [ 0 => array:5 [ "preGrado" => "Dra." "nombre" => "G." "apellidos" => "Margarit" "email" => array:1 [ 0 => "gmargarit@hsp.santpau.es" ] "referencia" => array:1 [ 0 => array:2 [ "etiqueta" => "<span class="elsevierStyleSup">*</span>" "identificador" => "cor1" ] ] ] 1 => array:2 [ "nombre" => "J." "apellidos" => "Belda" ] 2 => array:2 [ "nombre" => "P." "apellidos" => "Casan" ] 3 => array:2 [ "nombre" => "J." "apellidos" => "Sanchis" ] ] "afiliaciones" => array:1 [ 0 => array:1 [ "entidad" => "Departamento de Neumología, Hospital de la Santa Creu i de Sant Pau, Facultat de Medicina, Universitat Autònoma de Barcelona, Barcelona, Spain" ] ] "correspondencia" => array:1 [ 0 => array:3 [ "identificador" => "cor1" "etiqueta" => "*" "correspondencia" => "Correspondence: Dra. G. Margarit. Antoni M. Claret, 167. 08025 Barcelona. España" ] ] ] ] "pdfFichero" => "main.pdf" "tienePdf" => true "fechaRecibido" => "2005-01-14" "fechaAceptado" => "2005-02-01" "PalabrasClave" => array:2 [ "en" => array:1 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Key Words" "identificador" => "xpalclavsec154157" "palabras" => array:5 [ 0 => "Primary cultures" 1 => "Bronchial epithelial cells" 2 => "Culture expansion" 3 => "Explants" 4 => "Cytokeratin 7" ] ] ] "es" => array:1 [ 0 => array:4 [ "clase" => "keyword" "titulo" => "Palabras clave" "identificador" => "xpalclavsec154156" "palabras" => array:5 [ 0 => "Cultivos primarios" 1 => "Células epiteliales bronquiales" 2 => "Amplificación" 3 => "Explantes" 4 => "Citoqueratina 7" ] ] ] ] "tieneResumen" => true "resumen" => array:2 [ "en" => array:1 [ "resumen" => "<span class="elsevierStyleSectionTitle">Objective</span><p class="elsevierStyleSimplePara elsevierViewall">Cell cultures provide a good model for studying lung diseases but they are difficult to reproduce and the number of cells obtained is limited. The aim of this study was to develop a way to increase the production of human bronchial epithelial cells (BEC) in primary cultures.</p> <span class="elsevierStyleSectionTitle">Materialand methods</span><p class="elsevierStyleSimplePara elsevierViewall">A total of 12 samples (9 from surgical specimens and 3 from endoscopic biopsies) were processed on plates coated with type I collagen with growth medium supplemented for BEC. When cell proliferation started, the explants were removed for successive subculturing. The remaining cells were left to proliferate and were trypsinized after 50% confluence. We recorded the number of cells obtained, cell viability, and the percentage positive for cytokeratin 7.</p> <span class="elsevierStyleSectionTitle">Results</span><p class="elsevierStyleSimplePara elsevierViewall">The total number of cells obtained by this method was 3-fold the number of human BEC obtained with simple primary cultures. The maximum number of subcultures was 5, mean (SD) cell viability was 91.9% (11.7%), and the percentage of cells positive for cytokeratin 7 was 30.71% (10.68%).</p> <span class="elsevierStyleSectionTitle">Conclusions</span><p class="elsevierStyleSimplePara elsevierViewall">The described method for expanding primary BEC cultures increases cell production.</p>" ] "es" => array:1 [ "resumen" => "<span class="elsevierStyleSectionTitle">Objetivo</span><p class="elsevierStyleSimplePara elsevierViewall">LOS cultivos celulares son un buen modelo para el estudio de las enfermedades pulmonares, pero son difíciles de reproducir y producen un número limitado de células. El objetivo de este estudio ha sido desarrollar un método que incrementase la producción de células epiteliales bronquiales (CEB) humanas en cultivos primarios.</p> <span class="elsevierStyleSectionTitle">Materialy métodos</span><p class="elsevierStyleSimplePara elsevierViewall">Se procesó un total de 12 muestras (9 procedentes de muestras quirúrgicas y 3 de biopsias endoscópicas) en placas recubiertas de colágeno tipo I con medio suplementado para CEB. Al iniciarse la proliferación celular a su alrededor, los explantes se extrajeron y subcultivaron sucesivamente. Las células restantes se dejaron proliferar y se tripsinizaron tras alcanzar más del 50% de confluencia. Se valoraron el número de células obtenidas, la viabilidad y la citoqueratina 7.</p> <span class="elsevierStyleSectionTitle">Resultados</span><p class="elsevierStyleSimplePara elsevierViewall">El número total de células obtenidas con este método superó en una media de 3 veces el número de CEB humanas obtenidas en cultivos primarios simples. El número máximo de subcultivos fue de 5, la viabilidad media (± desviación estándar) fue de 91,9 ± 11,7% y el porcentaje de células positivas para la citoqueratina 7 del 30,71 ± 10,68%.</p> <span class="elsevierStyleSectionTitle">Conclusiones</span><p class="elsevierStyleSimplePara elsevierViewall">El método descrito para amplificar cultivos primarios de CEB permite incrementar la producción de células obtenidas.</p>" ] ] "NotaPie" => array:1 [ 0 => array:1 [ "nota" => "<p class="elsevierStyleNotepara">This study was partially funded by Red-Respira-ISCIII-RTIC-03/11.</p>" ] ] "bibliografia" => array:2 [ "titulo" => "REFERENCES" "seccion" => array:1 [ 0 => array:1 [ "bibliografiaReferencia" => array:7 [ 0 => array:3 [ "identificador" => "bib1" "etiqueta" => "1" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Bronchial epithelium: morphology, function and pathophysiology in asthma" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:2 [ 0 => "VH Velden" 1 => "HF Versnel" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:6 [ "tituloSerie" => "Eur Cytokine Netw" "fecha" => "1998" "volumen" => "9" "paginaInicial" => "585" "paginaFinal" => "597" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/9889402" "web" => "Medline" ] ] ] ] ] ] ] ] 1 => array:3 [ "identificador" => "bib2" "etiqueta" => "2" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Culture and comparison of human bronchial and nasal epithelial cells in vitro" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:5 [ 0 => "JL Devalia" 1 => "RJ Sapsford" 2 => "CW Wells" 3 => "P Richman" 4 => "RJ Davies" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:6 [ "tituloSerie" => "Respir Med" "fecha" => "1990" "volumen" => "84" "paginaInicial" => "303" "paginaFinal" => "312" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/2236758" "web" => "Medline" ] ] ] ] ] ] ] ] 2 => array:3 [ "identificador" => "bib3" "etiqueta" => "3" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Cell culture of airway epithelia" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:4 [ 0 => "MR van Scott" 1 => "PW Chen" 2 => "DC Henke" 3 => "JR Yankaskas" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "LibroEditado" => array:5 [ "paginaInicial" => "135" "paginaFinal" => "167" "serieVolumen" => "55" "serieTitulo" => "The airway epithelium: physiology, pathophysiology and pharmacology Lung biology in health and disease" "serieFecha" => "1991" ] ] ] ] ] ] 3 => array:3 [ "identificador" => "bib4" "etiqueta" => "4" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Contribution of in vitro culture methods for respiratory epithelial cells to the study of the physiology of the respiratory tract" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:4 [ 0 => "M Jorissen" 1 => "B van der Schueren" 2 => "H van den Bergue" 3 => "JJ Cassiman" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:6 [ "tituloSerie" => "Eur Respir J" "fecha" => "1991" "volumen" => "4" "paginaInicial" => "210" "paginaFinal" => "217" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/2044738" "web" => "Medline" ] ] ] ] ] ] ] ] 4 => array:3 [ "identificador" => "bib5" "etiqueta" => "5" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Growth and differentiation of conducting airway epithelial cells in culture" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:3 [ 0 => "R Wu" 1 => "YH Zhao" 2 => "MMJ Chang" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:6 [ "tituloSerie" => "Eur Respir J" "fecha" => "1997" "volumen" => "10" "paginaInicial" => "2398" "paginaFinal" => "2403" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/9387971" "web" => "Medline" ] ] ] ] ] ] ] ] 5 => array:3 [ "identificador" => "bib6" "etiqueta" => "6" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "OVTL 12/30 (keratin 7 antibody) is a marker of glandular differentiation in lung cancer" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:4 [ 0 => "FJ van der Molengraft" 1 => "CC van Niekerk" 2 => "PH Jap" 3 => "LG Poels" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:6 [ "tituloSerie" => "Histopathology" "fecha" => "1993" "volumen" => "22" "paginaInicial" => "35" "paginaFinal" => "38" "link" => array:1 [ 0 => array:2 [ "url" => "https://www.ncbi.nlm.nih.gov/pubmed/7679658" "web" => "Medline" ] ] ] ] ] ] ] ] 6 => array:3 [ "identificador" => "bib7" "etiqueta" => "7" "referencia" => array:1 [ 0 => array:2 [ "contribucion" => array:1 [ 0 => array:2 [ "titulo" => "Culture and characterisation of human bronchial epithelial cells in vitro" "autores" => array:1 [ 0 => array:2 [ "etal" => false "autores" => array:4 [ 0 => "RJ Sapsford" 1 => "JH Wang" 2 => "JL Devalia" 3 => "RJ Davies" ] ] ] ] ] "host" => array:1 [ 0 => array:1 [ "Revista" => array:4 [ "tituloSerie" => "Eur Respir J" "fecha" => "1995" "volumen" => "8" "paginaInicial" => "237S" ] ] ] ] ] ] ] ] ] ] ] "idiomaDefecto" => "en" "url" => "/15792129/0000004100000009/v1_201305150225/S1579212906602735/v1_201305150225/en/main.assets" "Apartado" => null "PDF" => "https://static.elsevier.es/multimedia/15792129/0000004100000009/v1_201305150225/S1579212906602735/v1_201305150225/en/main.pdf?idApp=UINPBA00003Z&text.app=https://archbronconeumol.org/" "EPUB" => "https://multimedia.elsevier.es/PublicationsMultimediaV1/item/epub/S1579212906602735?idApp=UINPBA00003Z" ]
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