Type II Pneumocyte-Restricted Green Fluorescent Protein Expression After Lentiviral Transduction of Lung Epithelial Cells
Type II alveolar epithelial (AT2) cell-specific reporter expression has been highly useful in the study of embryology and alveolar regeneration in transgenic mice. Technologies enabling efficient gene transfer and cell type-restricted transgene expression in AT2 cells would allow for correction of A...
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Veröffentlicht in: | Human gene therapy 2008, Vol.19 (1), p.39-51 |
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creator | WUNDERLICH, Stephanie GRUH, Ina WINKLER, Monica E BEIER, Jennifer RADTKE, Kerstin SCHMIEDL, Andreas GROOS, Stephanie HAVERICH, Axel MARTIN, Ulrich |
description | Type II alveolar epithelial (AT2) cell-specific reporter expression has been highly useful in the study of embryology and alveolar regeneration in transgenic mice. Technologies enabling efficient gene transfer and cell type-restricted transgene expression in AT2 cells would allow for correction of AT2 cell-based diseases such as genetic surfactant deficiencies. Moreover, such approaches are urgently required to investigate differentiation of AT2 cells from adult and embryonic stem cells of other species than mouse. Using a human surfactant protein C (SP-C) promoter fragment, we have constructed lentiviral vectors enabling AT2-restricted transgene expression and identification of stem cell-derived AT2 cells. Lung epithelial cell lines M3E3/C3, H441, RLE-6TN, A549, MLE-12, and MLE-15 were characterized at the molecular and ultrastructural levels to identify cell lines useful to assess the cell type specificity of our vector constructs. After transduction, no green fluorescent protein (GFP) expression was observed in nontarget cells including bronchial H441 cells, pulmonary A549 cells, fibroblasts, smooth muscle cells, and endothelial cells. In contrast, and in correlation with endogenous SP-C expression, lentiviral transduction resulted in stable GFP expression in MLE-12 and MLE-15 AT2 cells. In conclusion, we have constructed a lentiviral vector mediating SP-C promoter-dependent GFP expression. Transgene expression strictly corresponds with an AT2 phenotype of the transduced cells. In particular, the generated vector should facilitate local alveolar gene therapy and investigation of alveolar regeneration and stem cell differentiation. |
doi_str_mv | 10.1089/hum.2006.0180 |
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Technologies enabling efficient gene transfer and cell type-restricted transgene expression in AT2 cells would allow for correction of AT2 cell-based diseases such as genetic surfactant deficiencies. Moreover, such approaches are urgently required to investigate differentiation of AT2 cells from adult and embryonic stem cells of other species than mouse. Using a human surfactant protein C (SP-C) promoter fragment, we have constructed lentiviral vectors enabling AT2-restricted transgene expression and identification of stem cell-derived AT2 cells. Lung epithelial cell lines M3E3/C3, H441, RLE-6TN, A549, MLE-12, and MLE-15 were characterized at the molecular and ultrastructural levels to identify cell lines useful to assess the cell type specificity of our vector constructs. After transduction, no green fluorescent protein (GFP) expression was observed in nontarget cells including bronchial H441 cells, pulmonary A549 cells, fibroblasts, smooth muscle cells, and endothelial cells. In contrast, and in correlation with endogenous SP-C expression, lentiviral transduction resulted in stable GFP expression in MLE-12 and MLE-15 AT2 cells. In conclusion, we have constructed a lentiviral vector mediating SP-C promoter-dependent GFP expression. Transgene expression strictly corresponds with an AT2 phenotype of the transduced cells. In particular, the generated vector should facilitate local alveolar gene therapy and investigation of alveolar regeneration and stem cell differentiation.</description><identifier>ISSN: 1043-0342</identifier><identifier>EISSN: 1557-7422</identifier><identifier>DOI: 10.1089/hum.2006.0180</identifier><identifier>PMID: 18052721</identifier><identifier>CODEN: HGTHE3</identifier><language>eng</language><publisher>Larchmont, NY: Liebert</publisher><subject>Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy ; Animals ; Applied cell therapy and gene therapy ; Biological and medical sciences ; Biotechnology ; Cells, Cultured ; Epithelial Cells - metabolism ; Fundamental and applied biological sciences. Psychology ; Gene targeting ; Gene therapy ; Genetic Vectors ; Green Fluorescent Proteins - genetics ; Green Fluorescent Proteins - metabolism ; Health. Pharmaceutical industry ; Humans ; Industrial applications and implications. Economical aspects ; Lentivirus - genetics ; Medical sciences ; Methods ; Mice ; Phenotype ; Promoter Regions, Genetic ; Pulmonary Alveoli - cytology ; Pulmonary Alveoli - metabolism ; Pulmonary Surfactant-Associated Protein C - genetics ; Rats ; Transduction, Genetic ; Transfusions. Complications. Transfusion reactions. 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Technologies enabling efficient gene transfer and cell type-restricted transgene expression in AT2 cells would allow for correction of AT2 cell-based diseases such as genetic surfactant deficiencies. Moreover, such approaches are urgently required to investigate differentiation of AT2 cells from adult and embryonic stem cells of other species than mouse. Using a human surfactant protein C (SP-C) promoter fragment, we have constructed lentiviral vectors enabling AT2-restricted transgene expression and identification of stem cell-derived AT2 cells. Lung epithelial cell lines M3E3/C3, H441, RLE-6TN, A549, MLE-12, and MLE-15 were characterized at the molecular and ultrastructural levels to identify cell lines useful to assess the cell type specificity of our vector constructs. After transduction, no green fluorescent protein (GFP) expression was observed in nontarget cells including bronchial H441 cells, pulmonary A549 cells, fibroblasts, smooth muscle cells, and endothelial cells. In contrast, and in correlation with endogenous SP-C expression, lentiviral transduction resulted in stable GFP expression in MLE-12 and MLE-15 AT2 cells. In conclusion, we have constructed a lentiviral vector mediating SP-C promoter-dependent GFP expression. Transgene expression strictly corresponds with an AT2 phenotype of the transduced cells. In particular, the generated vector should facilitate local alveolar gene therapy and investigation of alveolar regeneration and stem cell differentiation.</description><subject>Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy</subject><subject>Animals</subject><subject>Applied cell therapy and gene therapy</subject><subject>Biological and medical sciences</subject><subject>Biotechnology</subject><subject>Cells, Cultured</subject><subject>Epithelial Cells - metabolism</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Gene targeting</subject><subject>Gene therapy</subject><subject>Genetic Vectors</subject><subject>Green Fluorescent Proteins - genetics</subject><subject>Green Fluorescent Proteins - metabolism</subject><subject>Health. Pharmaceutical industry</subject><subject>Humans</subject><subject>Industrial applications and implications. Economical aspects</subject><subject>Lentivirus - genetics</subject><subject>Medical sciences</subject><subject>Methods</subject><subject>Mice</subject><subject>Phenotype</subject><subject>Promoter Regions, Genetic</subject><subject>Pulmonary Alveoli - cytology</subject><subject>Pulmonary Alveoli - metabolism</subject><subject>Pulmonary Surfactant-Associated Protein C - genetics</subject><subject>Rats</subject><subject>Transduction, Genetic</subject><subject>Transfusions. Complications. Transfusion reactions. Cell and gene therapy</subject><subject>Transgenes</subject><issn>1043-0342</issn><issn>1557-7422</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2008</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkdGL1DAQxoso3nn66KsERN-6Jm3atI_LsncuLHjI-hyyyeQukjY1SQ_3v3fKLoogSB4mzPzmY2a-onjL6IrRrv_0OA-ritJ2RVlHnxXXrGlEKXhVPcc_5XVJa15dFa9S-k4pq5tWvCyuEG0qUbHr4nQ4TUB2O3I_wjwEfcpQfoWUo9MZDLmLACO59XOIkDSMmdzHkMGNZPtzwlRyYSRrmyGSPVbdk4vKk0NUYzKzzks1WLKfxweynVx-BO-wvgHv0-vihVU-wZtLvCm-3W4Pm8_l_svdbrPel5qzPpctVUofecN73gjRdJxxfeyNsrgK73jXCwVca9aYrjZW8N6CEaI2R9P2fVdBfVN8POtOMfyYcTU5OFzFezVCmJMUeCTBW_pfsMJz9zgFgu_P4IPyIN1oQ45KL7BcM4FqFas5Uqt_UPgMDE6HEazD_F8N5blBx5BSBCun6AYVT5JRuXgt0Wu5eC0Xr5F_d5l3Pg5g_tAXcxH4cAFU0spbdEW79JtDJS6qTtS_AJUIsWo</recordid><startdate>2008</startdate><enddate>2008</enddate><creator>WUNDERLICH, Stephanie</creator><creator>GRUH, Ina</creator><creator>WINKLER, Monica E</creator><creator>BEIER, Jennifer</creator><creator>RADTKE, Kerstin</creator><creator>SCHMIEDL, Andreas</creator><creator>GROOS, Stephanie</creator><creator>HAVERICH, Axel</creator><creator>MARTIN, Ulrich</creator><general>Liebert</general><general>Mary Ann Liebert, Inc</general><scope>IQODW</scope><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>7QO</scope><scope>7U9</scope><scope>8FD</scope><scope>FR3</scope><scope>H94</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope></search><sort><creationdate>2008</creationdate><title>Type II Pneumocyte-Restricted Green Fluorescent Protein Expression After Lentiviral Transduction of Lung Epithelial Cells</title><author>WUNDERLICH, Stephanie ; GRUH, Ina ; WINKLER, Monica E ; BEIER, Jennifer ; RADTKE, Kerstin ; SCHMIEDL, Andreas ; GROOS, Stephanie ; HAVERICH, Axel ; MARTIN, Ulrich</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c419t-60aacb4549457758414cb9daf356484897ae4cc15d83df749fed773dbd69982e3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2008</creationdate><topic>Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy</topic><topic>Animals</topic><topic>Applied cell therapy and gene therapy</topic><topic>Biological and medical sciences</topic><topic>Biotechnology</topic><topic>Cells, Cultured</topic><topic>Epithelial Cells - metabolism</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Gene targeting</topic><topic>Gene therapy</topic><topic>Genetic Vectors</topic><topic>Green Fluorescent Proteins - genetics</topic><topic>Green Fluorescent Proteins - metabolism</topic><topic>Health. Pharmaceutical industry</topic><topic>Humans</topic><topic>Industrial applications and implications. Economical aspects</topic><topic>Lentivirus - genetics</topic><topic>Medical sciences</topic><topic>Methods</topic><topic>Mice</topic><topic>Phenotype</topic><topic>Promoter Regions, Genetic</topic><topic>Pulmonary Alveoli - cytology</topic><topic>Pulmonary Alveoli - metabolism</topic><topic>Pulmonary Surfactant-Associated Protein C - genetics</topic><topic>Rats</topic><topic>Transduction, Genetic</topic><topic>Transfusions. Complications. Transfusion reactions. Cell and gene therapy</topic><topic>Transgenes</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>WUNDERLICH, Stephanie</creatorcontrib><creatorcontrib>GRUH, Ina</creatorcontrib><creatorcontrib>WINKLER, Monica E</creatorcontrib><creatorcontrib>BEIER, Jennifer</creatorcontrib><creatorcontrib>RADTKE, Kerstin</creatorcontrib><creatorcontrib>SCHMIEDL, Andreas</creatorcontrib><creatorcontrib>GROOS, Stephanie</creatorcontrib><creatorcontrib>HAVERICH, Axel</creatorcontrib><creatorcontrib>MARTIN, Ulrich</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Biotechnology Research Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Human gene therapy</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>WUNDERLICH, Stephanie</au><au>GRUH, Ina</au><au>WINKLER, Monica E</au><au>BEIER, Jennifer</au><au>RADTKE, Kerstin</au><au>SCHMIEDL, Andreas</au><au>GROOS, Stephanie</au><au>HAVERICH, Axel</au><au>MARTIN, Ulrich</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Type II Pneumocyte-Restricted Green Fluorescent Protein Expression After Lentiviral Transduction of Lung Epithelial Cells</atitle><jtitle>Human gene therapy</jtitle><addtitle>Hum Gene Ther</addtitle><date>2008</date><risdate>2008</risdate><volume>19</volume><issue>1</issue><spage>39</spage><epage>51</epage><pages>39-51</pages><issn>1043-0342</issn><eissn>1557-7422</eissn><coden>HGTHE3</coden><abstract>Type II alveolar epithelial (AT2) cell-specific reporter expression has been highly useful in the study of embryology and alveolar regeneration in transgenic mice. Technologies enabling efficient gene transfer and cell type-restricted transgene expression in AT2 cells would allow for correction of AT2 cell-based diseases such as genetic surfactant deficiencies. Moreover, such approaches are urgently required to investigate differentiation of AT2 cells from adult and embryonic stem cells of other species than mouse. Using a human surfactant protein C (SP-C) promoter fragment, we have constructed lentiviral vectors enabling AT2-restricted transgene expression and identification of stem cell-derived AT2 cells. Lung epithelial cell lines M3E3/C3, H441, RLE-6TN, A549, MLE-12, and MLE-15 were characterized at the molecular and ultrastructural levels to identify cell lines useful to assess the cell type specificity of our vector constructs. After transduction, no green fluorescent protein (GFP) expression was observed in nontarget cells including bronchial H441 cells, pulmonary A549 cells, fibroblasts, smooth muscle cells, and endothelial cells. In contrast, and in correlation with endogenous SP-C expression, lentiviral transduction resulted in stable GFP expression in MLE-12 and MLE-15 AT2 cells. In conclusion, we have constructed a lentiviral vector mediating SP-C promoter-dependent GFP expression. Transgene expression strictly corresponds with an AT2 phenotype of the transduced cells. In particular, the generated vector should facilitate local alveolar gene therapy and investigation of alveolar regeneration and stem cell differentiation.</abstract><cop>Larchmont, NY</cop><pub>Liebert</pub><pmid>18052721</pmid><doi>10.1089/hum.2006.0180</doi><tpages>13</tpages></addata></record> |
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subjects | Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy Animals Applied cell therapy and gene therapy Biological and medical sciences Biotechnology Cells, Cultured Epithelial Cells - metabolism Fundamental and applied biological sciences. Psychology Gene targeting Gene therapy Genetic Vectors Green Fluorescent Proteins - genetics Green Fluorescent Proteins - metabolism Health. Pharmaceutical industry Humans Industrial applications and implications. Economical aspects Lentivirus - genetics Medical sciences Methods Mice Phenotype Promoter Regions, Genetic Pulmonary Alveoli - cytology Pulmonary Alveoli - metabolism Pulmonary Surfactant-Associated Protein C - genetics Rats Transduction, Genetic Transfusions. Complications. Transfusion reactions. Cell and gene therapy Transgenes |
title | Type II Pneumocyte-Restricted Green Fluorescent Protein Expression After Lentiviral Transduction of Lung Epithelial Cells |
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