Efficient generation of functional CFTR-expressing airway epithelial cells from human pluripotent stem cells
Functional CFTR-expressing airway epithelial cells are generated via differentiation firstly into progenitors and then into mature epithelia with apical CFTR activity. Airway epithelial cells are of great interest for research on lung development, regeneration and disease modeling. This protocol des...
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| Veröffentlicht in: | Nature protocols 2015-03, Vol.10 (3), p.363-381 |
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| creator | Wong, Amy P Chin, Stephanie Xia, Sunny Garner, Jodi Bear, Christine E Rossant, Janet |
| description | Functional CFTR-expressing airway epithelial cells are generated via differentiation firstly into progenitors and then into mature epithelia with apical CFTR activity.
Airway epithelial cells are of great interest for research on lung development, regeneration and disease modeling. This protocol describes how to generate cystic fibrosis (CF) transmembrane conductance regulator protein (CFTR)-expressing airway epithelial cells from human pluripotent stem cells (PSCs). The stepwise approach from PSC culture to differentiation into progenitors and then mature epithelia with apical CFTR activity is outlined. Human PSCs that were inefficient at endoderm differentiation using our previous lung differentiation protocol were able to generate substantial lung progenitor cell populations. Augmented CFTR activity can be observed in all cultures as early as at 35 d of differentiation, and full maturation of the cells in air-liquid interface cultures occurs in |
| doi_str_mv | 10.1038/nprot.2015.021 |
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Airway epithelial cells are of great interest for research on lung development, regeneration and disease modeling. This protocol describes how to generate cystic fibrosis (CF) transmembrane conductance regulator protein (CFTR)-expressing airway epithelial cells from human pluripotent stem cells (PSCs). The stepwise approach from PSC culture to differentiation into progenitors and then mature epithelia with apical CFTR activity is outlined. Human PSCs that were inefficient at endoderm differentiation using our previous lung differentiation protocol were able to generate substantial lung progenitor cell populations. Augmented CFTR activity can be observed in all cultures as early as at 35 d of differentiation, and full maturation of the cells in air-liquid interface cultures occurs in <5 weeks. This protocol can be used for drug discovery, tissue regeneration or disease modeling.</description><identifier>ISSN: 1754-2189</identifier><identifier>EISSN: 1750-2799</identifier><identifier>DOI: 10.1038/nprot.2015.021</identifier><identifier>PMID: 25654755</identifier><language>eng</language><publisher>London: Nature Publishing Group UK</publisher><subject>631/1647/1407/651 ; 631/1647/767/2201 ; 631/532/1360 ; 631/532/2064/2158 ; Airway (Medicine) ; Analytical Chemistry ; Biological Techniques ; Cell culture ; Cell Culture Techniques - methods ; Cell Differentiation - physiology ; Computational Biology/Bioinformatics ; Cystic fibrosis ; Cystic Fibrosis Transmembrane Conductance Regulator - metabolism ; Epithelial cells ; Health aspects ; Humans ; Life Sciences ; Methods ; Microarrays ; Organic Chemistry ; Physiological aspects ; Pluripotent Stem Cells - cytology ; Proteins ; protocol ; R&D ; Research & development ; Respiratory Mucosa - cytology ; Respiratory Mucosa - metabolism ; Stem cells ; Tissue engineering</subject><ispartof>Nature protocols, 2015-03, Vol.10 (3), p.363-381</ispartof><rights>Springer Nature Limited 2014</rights><rights>COPYRIGHT 2015 Nature Publishing Group</rights><rights>Copyright Nature Publishing Group Mar 2015</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c603t-ac4a03e56876d0a68a73fd42bac2b478fbb41272cbc5b5fed07f90da188191923</citedby><cites>FETCH-LOGICAL-c603t-ac4a03e56876d0a68a73fd42bac2b478fbb41272cbc5b5fed07f90da188191923</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/25654755$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Wong, Amy P</creatorcontrib><creatorcontrib>Chin, Stephanie</creatorcontrib><creatorcontrib>Xia, Sunny</creatorcontrib><creatorcontrib>Garner, Jodi</creatorcontrib><creatorcontrib>Bear, Christine E</creatorcontrib><creatorcontrib>Rossant, Janet</creatorcontrib><title>Efficient generation of functional CFTR-expressing airway epithelial cells from human pluripotent stem cells</title><title>Nature protocols</title><addtitle>Nat Protoc</addtitle><addtitle>Nat Protoc</addtitle><description>Functional CFTR-expressing airway epithelial cells are generated via differentiation firstly into progenitors and then into mature epithelia with apical CFTR activity.
Airway epithelial cells are of great interest for research on lung development, regeneration and disease modeling. This protocol describes how to generate cystic fibrosis (CF) transmembrane conductance regulator protein (CFTR)-expressing airway epithelial cells from human pluripotent stem cells (PSCs). The stepwise approach from PSC culture to differentiation into progenitors and then mature epithelia with apical CFTR activity is outlined. Human PSCs that were inefficient at endoderm differentiation using our previous lung differentiation protocol were able to generate substantial lung progenitor cell populations. Augmented CFTR activity can be observed in all cultures as early as at 35 d of differentiation, and full maturation of the cells in air-liquid interface cultures occurs in <5 weeks. This protocol can be used for drug discovery, tissue regeneration or disease modeling.</description><subject>631/1647/1407/651</subject><subject>631/1647/767/2201</subject><subject>631/532/1360</subject><subject>631/532/2064/2158</subject><subject>Airway (Medicine)</subject><subject>Analytical Chemistry</subject><subject>Biological Techniques</subject><subject>Cell culture</subject><subject>Cell Culture Techniques - methods</subject><subject>Cell Differentiation - physiology</subject><subject>Computational Biology/Bioinformatics</subject><subject>Cystic fibrosis</subject><subject>Cystic Fibrosis Transmembrane Conductance Regulator - metabolism</subject><subject>Epithelial cells</subject><subject>Health aspects</subject><subject>Humans</subject><subject>Life Sciences</subject><subject>Methods</subject><subject>Microarrays</subject><subject>Organic Chemistry</subject><subject>Physiological aspects</subject><subject>Pluripotent Stem Cells - cytology</subject><subject>Proteins</subject><subject>protocol</subject><subject>R&D</subject><subject>Research & development</subject><subject>Respiratory Mucosa - cytology</subject><subject>Respiratory Mucosa - metabolism</subject><subject>Stem cells</subject><subject>Tissue engineering</subject><issn>1754-2189</issn><issn>1750-2799</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNqNks1v1DAQxSMEoqVw5YgicYFDtrbjj-RYrVqoVAmplLPlOOPUVWIH2xHtf4_TLaVFe0A-eGT_3tPM6BXFe4w2GNXNsZuDTxuCMNsggl8Uh1gwVBHRti_va1oR3LQHxZsYbxCioubidXFAGGdUMHZYjKfGWG3BpXIAB0El613pTWkWp9dajeX27Oqygts5QIzWDaWy4Ze6K2G26RpGmwkN4xhLE_xUXi-TcuU8LsHOPq2-McG0I94Wr4waI7x7uI-KH2enV9uv1cW3L-fbk4tKc1SnSmmqUA2MN4L3SPFGidr0lHRKk46KxnQdxUQQ3WnWMQM9EqZFvcJNg1vckvqo-LTzzcv5uUBMcrJx7UA58EuUmHNEEWGM_wfKCOWkaUVGP_6D3vgl5AXtqJo0VJC_1KBGkNYZn4LSq6k8oYgSwRmmmdrsofLpYbLaOzA2vz8TfH4myEyC2zSoJUZ5_v1yr7kOPsYARs7BTircSYzkmhp5nxq5pkbm1GTBh4fJlm6C_hH_E5MMHO-AmL_cAOHJ6PstfwMaUcxg</recordid><startdate>20150301</startdate><enddate>20150301</enddate><creator>Wong, Amy P</creator><creator>Chin, Stephanie</creator><creator>Xia, Sunny</creator><creator>Garner, Jodi</creator><creator>Bear, Christine E</creator><creator>Rossant, Janet</creator><general>Nature Publishing Group UK</general><general>Nature Publishing Group</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>ISR</scope><scope>3V.</scope><scope>7QG</scope><scope>7T5</scope><scope>7T7</scope><scope>7TM</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>P64</scope><scope>PATMY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PYCSY</scope><scope>RC3</scope><scope>7X8</scope><scope>7QO</scope></search><sort><creationdate>20150301</creationdate><title>Efficient generation of functional CFTR-expressing airway epithelial cells from human pluripotent stem cells</title><author>Wong, Amy P ; Chin, Stephanie ; Xia, Sunny ; Garner, Jodi ; Bear, Christine E ; Rossant, Janet</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c603t-ac4a03e56876d0a68a73fd42bac2b478fbb41272cbc5b5fed07f90da188191923</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>631/1647/1407/651</topic><topic>631/1647/767/2201</topic><topic>631/532/1360</topic><topic>631/532/2064/2158</topic><topic>Airway (Medicine)</topic><topic>Analytical Chemistry</topic><topic>Biological Techniques</topic><topic>Cell culture</topic><topic>Cell Culture Techniques - methods</topic><topic>Cell Differentiation - physiology</topic><topic>Computational Biology/Bioinformatics</topic><topic>Cystic fibrosis</topic><topic>Cystic Fibrosis Transmembrane Conductance Regulator - metabolism</topic><topic>Epithelial cells</topic><topic>Health aspects</topic><topic>Humans</topic><topic>Life Sciences</topic><topic>Methods</topic><topic>Microarrays</topic><topic>Organic Chemistry</topic><topic>Physiological aspects</topic><topic>Pluripotent Stem Cells - cytology</topic><topic>Proteins</topic><topic>protocol</topic><topic>R&D</topic><topic>Research & development</topic><topic>Respiratory Mucosa - cytology</topic><topic>Respiratory Mucosa - metabolism</topic><topic>Stem cells</topic><topic>Tissue engineering</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Wong, Amy P</creatorcontrib><creatorcontrib>Chin, Stephanie</creatorcontrib><creatorcontrib>Xia, Sunny</creatorcontrib><creatorcontrib>Garner, Jodi</creatorcontrib><creatorcontrib>Bear, Christine E</creatorcontrib><creatorcontrib>Rossant, Janet</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Gale In Context: Science</collection><collection>ProQuest Central (Corporate)</collection><collection>Animal Behavior Abstracts</collection><collection>Immunology Abstracts</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Nucleic Acids Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Technology Research Database</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>Agricultural & Environmental Science Collection</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>Engineering Research Database</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>ProQuest Biological Science Collection</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biological Science Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Environmental Science Database</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>Environmental Science Collection</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><collection>Biotechnology Research Abstracts</collection><jtitle>Nature protocols</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Wong, Amy P</au><au>Chin, Stephanie</au><au>Xia, Sunny</au><au>Garner, Jodi</au><au>Bear, Christine E</au><au>Rossant, Janet</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Efficient generation of functional CFTR-expressing airway epithelial cells from human pluripotent stem cells</atitle><jtitle>Nature protocols</jtitle><stitle>Nat Protoc</stitle><addtitle>Nat Protoc</addtitle><date>2015-03-01</date><risdate>2015</risdate><volume>10</volume><issue>3</issue><spage>363</spage><epage>381</epage><pages>363-381</pages><issn>1754-2189</issn><eissn>1750-2799</eissn><abstract>Functional CFTR-expressing airway epithelial cells are generated via differentiation firstly into progenitors and then into mature epithelia with apical CFTR activity.
Airway epithelial cells are of great interest for research on lung development, regeneration and disease modeling. This protocol describes how to generate cystic fibrosis (CF) transmembrane conductance regulator protein (CFTR)-expressing airway epithelial cells from human pluripotent stem cells (PSCs). The stepwise approach from PSC culture to differentiation into progenitors and then mature epithelia with apical CFTR activity is outlined. Human PSCs that were inefficient at endoderm differentiation using our previous lung differentiation protocol were able to generate substantial lung progenitor cell populations. Augmented CFTR activity can be observed in all cultures as early as at 35 d of differentiation, and full maturation of the cells in air-liquid interface cultures occurs in <5 weeks. This protocol can be used for drug discovery, tissue regeneration or disease modeling.</abstract><cop>London</cop><pub>Nature Publishing Group UK</pub><pmid>25654755</pmid><doi>10.1038/nprot.2015.021</doi><tpages>19</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | 631/1647/1407/651 631/1647/767/2201 631/532/1360 631/532/2064/2158 Airway (Medicine) Analytical Chemistry Biological Techniques Cell culture Cell Culture Techniques - methods Cell Differentiation - physiology Computational Biology/Bioinformatics Cystic fibrosis Cystic Fibrosis Transmembrane Conductance Regulator - metabolism Epithelial cells Health aspects Humans Life Sciences Methods Microarrays Organic Chemistry Physiological aspects Pluripotent Stem Cells - cytology Proteins protocol R&D Research & development Respiratory Mucosa - cytology Respiratory Mucosa - metabolism Stem cells Tissue engineering |
| title | Efficient generation of functional CFTR-expressing airway epithelial cells from human pluripotent stem cells |
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