Inducible regulation of Runx2-stimulated osteogenesis
Ex vivo gene therapy is a promising approach to orthopedic regenerative medicine. These strategies typically focus on the constitutive overexpression of osteogenic factors to induce osteoblastic differentiation and matrix mineralization. However, the unregulated production of osteoinductive molecule...
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| Veröffentlicht in: | Gene therapy 2006-06, Vol.13 (11), p.873-882 |
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| creator | Gersbach, C A Le Doux, J M Guldberg, R E García, A J |
| description | Ex vivo
gene therapy is a promising approach to orthopedic regenerative medicine. These strategies typically focus on the constitutive overexpression of osteogenic factors to induce osteoblastic differentiation and matrix mineralization. However, the unregulated production of osteoinductive molecules has also resulted in abnormal bone formation and tumorigenesis. To address these limitations, this work describes a retroviral system to deliver the Runx2 osteoblastic transcription factor under control of the tetracycline-inducible (tet-off) promoter in primary skeletal myoblasts. Runx2 expression was tightly regulated by anhydrotetracyline (aTc) concentration in cell culture media. Osteoblastic gene expression, alkaline phosphatase activity, and matrix mineralization were also controlled by aTc in a dose-dependent manner. Additionally, osteoblastic differentiation was temporally regulated by adding and removing aTc from the culture media. Engineered cells were seeded onto collagen scaffolds and implanted intramuscularly in the hind limbs of syngeneic mice.
In vivo
mineralization by these constructs was regulated by supplementing the drinking water with aTc, as demonstrated by micro-computed tomography and histological analyses. Collectively, these results present a novel system for regulating osteoblastic differentiation of a clinically relevant autologous cell source. This system is significant to developing controlled and effective orthopedic gene therapy strategies and studying the regulation of osteoblastic differentiation. |
| doi_str_mv | 10.1038/sj.gt.3302725 |
| format | Article |
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gene therapy is a promising approach to orthopedic regenerative medicine. These strategies typically focus on the constitutive overexpression of osteogenic factors to induce osteoblastic differentiation and matrix mineralization. However, the unregulated production of osteoinductive molecules has also resulted in abnormal bone formation and tumorigenesis. To address these limitations, this work describes a retroviral system to deliver the Runx2 osteoblastic transcription factor under control of the tetracycline-inducible (tet-off) promoter in primary skeletal myoblasts. Runx2 expression was tightly regulated by anhydrotetracyline (aTc) concentration in cell culture media. Osteoblastic gene expression, alkaline phosphatase activity, and matrix mineralization were also controlled by aTc in a dose-dependent manner. Additionally, osteoblastic differentiation was temporally regulated by adding and removing aTc from the culture media. Engineered cells were seeded onto collagen scaffolds and implanted intramuscularly in the hind limbs of syngeneic mice.
In vivo
mineralization by these constructs was regulated by supplementing the drinking water with aTc, as demonstrated by micro-computed tomography and histological analyses. Collectively, these results present a novel system for regulating osteoblastic differentiation of a clinically relevant autologous cell source. This system is significant to developing controlled and effective orthopedic gene therapy strategies and studying the regulation of osteoblastic differentiation.</description><identifier>ISSN: 0969-7128</identifier><identifier>EISSN: 1476-5462</identifier><identifier>DOI: 10.1038/sj.gt.3302725</identifier><identifier>PMID: 16496016</identifier><language>eng</language><publisher>London: Nature Publishing Group UK</publisher><subject>Alkaline phosphatase ; Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy ; Animals ; Anti-Bacterial Agents ; Applied cell therapy and gene therapy ; Biological and medical sciences ; Biomedical and Life Sciences ; Biomedicine ; Biotechnology ; Bone diseases ; Bone growth ; Bone Transplantation ; Bones ; Care and treatment ; Cbfa-1 protein ; Cell Biology ; Cell culture ; Cells ; Cells, Cultured ; Collagen ; Computed tomography ; Core Binding Factor Alpha 1 Subunit - genetics ; Core Binding Factor Alpha 1 Subunit - metabolism ; Culture media ; DNA binding proteins ; Drinking water ; Fundamental and applied biological sciences. Psychology ; Gene Expression ; Gene Expression Regulation ; Gene Therapy ; Genetic aspects ; Genetic Therapy - methods ; Genetic Vectors - administration & dosage ; Growth ; Health aspects ; Health. Pharmaceutical industry ; Hindlimb ; Human Genetics ; Industrial applications and implications. Economical aspects ; Male ; Medical sciences ; Mice ; Mice, Inbred BALB C ; Mineralization ; Myoblasts ; Nanotechnology ; original-article ; Orthopedics ; Osteoblastogenesis ; Osteoblasts ; Osteoblasts - cytology ; Osteoblasts - diagnostic imaging ; Osteoblasts - metabolism ; Osteogenesis ; Physiological aspects ; Regenerative medicine ; Retroviridae - genetics ; Tetracycline ; Tissue engineering ; Tissue Engineering - methods ; Tomography, X-Ray Computed ; Trans-Activators ; Transduction, Genetic - methods ; Transfusions. Complications. Transfusion reactions. Cell and gene therapy ; Tumorigenesis</subject><ispartof>Gene therapy, 2006-06, Vol.13 (11), p.873-882</ispartof><rights>Springer Nature Limited 2006</rights><rights>2006 INIST-CNRS</rights><rights>COPYRIGHT 2006 Nature Publishing Group</rights><rights>Copyright Nature Publishing Group Jun 2006</rights><rights>Nature Publishing Group 2006.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c639t-3011bfedead056c7db5a3277022895c160d850f349c2622bd2e212be9deb57f93</citedby><cites>FETCH-LOGICAL-c639t-3011bfedead056c7db5a3277022895c160d850f349c2622bd2e212be9deb57f93</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=17756324$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/16496016$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Gersbach, C A</creatorcontrib><creatorcontrib>Le Doux, J M</creatorcontrib><creatorcontrib>Guldberg, R E</creatorcontrib><creatorcontrib>García, A J</creatorcontrib><title>Inducible regulation of Runx2-stimulated osteogenesis</title><title>Gene therapy</title><addtitle>Gene Ther</addtitle><addtitle>Gene Ther</addtitle><description>Ex vivo
gene therapy is a promising approach to orthopedic regenerative medicine. These strategies typically focus on the constitutive overexpression of osteogenic factors to induce osteoblastic differentiation and matrix mineralization. However, the unregulated production of osteoinductive molecules has also resulted in abnormal bone formation and tumorigenesis. To address these limitations, this work describes a retroviral system to deliver the Runx2 osteoblastic transcription factor under control of the tetracycline-inducible (tet-off) promoter in primary skeletal myoblasts. Runx2 expression was tightly regulated by anhydrotetracyline (aTc) concentration in cell culture media. Osteoblastic gene expression, alkaline phosphatase activity, and matrix mineralization were also controlled by aTc in a dose-dependent manner. Additionally, osteoblastic differentiation was temporally regulated by adding and removing aTc from the culture media. Engineered cells were seeded onto collagen scaffolds and implanted intramuscularly in the hind limbs of syngeneic mice.
In vivo
mineralization by these constructs was regulated by supplementing the drinking water with aTc, as demonstrated by micro-computed tomography and histological analyses. Collectively, these results present a novel system for regulating osteoblastic differentiation of a clinically relevant autologous cell source. This system is significant to developing controlled and effective orthopedic gene therapy strategies and studying the regulation of osteoblastic differentiation.</description><subject>Alkaline phosphatase</subject><subject>Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy</subject><subject>Animals</subject><subject>Anti-Bacterial Agents</subject><subject>Applied cell therapy and gene therapy</subject><subject>Biological and medical sciences</subject><subject>Biomedical and Life Sciences</subject><subject>Biomedicine</subject><subject>Biotechnology</subject><subject>Bone diseases</subject><subject>Bone growth</subject><subject>Bone Transplantation</subject><subject>Bones</subject><subject>Care and treatment</subject><subject>Cbfa-1 protein</subject><subject>Cell Biology</subject><subject>Cell culture</subject><subject>Cells</subject><subject>Cells, Cultured</subject><subject>Collagen</subject><subject>Computed tomography</subject><subject>Core Binding Factor Alpha 1 Subunit - genetics</subject><subject>Core Binding Factor Alpha 1 Subunit - metabolism</subject><subject>Culture media</subject><subject>DNA binding proteins</subject><subject>Drinking water</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Gene Expression</subject><subject>Gene Expression Regulation</subject><subject>Gene Therapy</subject><subject>Genetic aspects</subject><subject>Genetic Therapy - methods</subject><subject>Genetic Vectors - administration & dosage</subject><subject>Growth</subject><subject>Health aspects</subject><subject>Health. Pharmaceutical industry</subject><subject>Hindlimb</subject><subject>Human Genetics</subject><subject>Industrial applications and implications. Economical aspects</subject><subject>Male</subject><subject>Medical sciences</subject><subject>Mice</subject><subject>Mice, Inbred BALB C</subject><subject>Mineralization</subject><subject>Myoblasts</subject><subject>Nanotechnology</subject><subject>original-article</subject><subject>Orthopedics</subject><subject>Osteoblastogenesis</subject><subject>Osteoblasts</subject><subject>Osteoblasts - cytology</subject><subject>Osteoblasts - diagnostic imaging</subject><subject>Osteoblasts - metabolism</subject><subject>Osteogenesis</subject><subject>Physiological aspects</subject><subject>Regenerative medicine</subject><subject>Retroviridae - genetics</subject><subject>Tetracycline</subject><subject>Tissue engineering</subject><subject>Tissue Engineering - methods</subject><subject>Tomography, X-Ray Computed</subject><subject>Trans-Activators</subject><subject>Transduction, Genetic - methods</subject><subject>Transfusions. Complications. Transfusion reactions. Cell and gene therapy</subject><subject>Tumorigenesis</subject><issn>0969-7128</issn><issn>1476-5462</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2006</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>8G5</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><sourceid>GUQSH</sourceid><sourceid>M2O</sourceid><recordid>eNqF0t1rFDEQAPBFFHutPvqoHIoFH_ZMsvnYPJZS9aAgVH0O2ezsdo-9pGayUP97c9zqeWKRPASS38wwyRTFC0pWlFT1e9ys-rSqKsIUE4-KBeVKloJL9rhYEC11qSirT4pTxA0hhKuaPS1OqORaEioXhVj7dnJDM8IyQj-NNg3BL0O3vJn8PSsxDdvdIbTLgAlCDx5wwGfFk86OCM_n_az49uHq6-Wn8vrzx_XlxXXpZKVTWRFKmw5asC0R0qm2EbZiShHGai0claStBekqrh2TjDUtA0ZZA7qFRqhOV2fF-T7vXQzfJ8BktgM6GEfrIUxopNJ1zVj1X0i1FkoqleGbv-AmTNHnJgyTnEsuKOdZvX5Q0VoRzWp-SNXbEczgu5Cidbu65oLWuU2qKc1q9Q-VVwvbwQUP3ZDPjwLeHQVkk-A-9XZCNOsvN8f2_A97C3ZMtxjGafeJeAzLPXQxIEbozF0ctjb-MJSY3RwZ3Jg-mXmOsn81P8DUbKE96HlwMng7A4vOjl203g14cEoJWTF-aB_zle8hHl7yocov9wHepinC74y_7n8CUobj4w</recordid><startdate>20060601</startdate><enddate>20060601</enddate><creator>Gersbach, C A</creator><creator>Le Doux, J M</creator><creator>Guldberg, R E</creator><creator>García, A J</creator><general>Nature Publishing Group UK</general><general>Nature Publishing Group</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>ISR</scope><scope>3V.</scope><scope>7QP</scope><scope>7TK</scope><scope>7TM</scope><scope>7U9</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>8G5</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>GUQSH</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M2O</scope><scope>M7P</scope><scope>MBDVC</scope><scope>P64</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>Q9U</scope><scope>RC3</scope><scope>7QO</scope><scope>7X8</scope></search><sort><creationdate>20060601</creationdate><title>Inducible regulation of Runx2-stimulated osteogenesis</title><author>Gersbach, C A ; Le Doux, J M ; Guldberg, R E ; García, A J</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c639t-3011bfedead056c7db5a3277022895c160d850f349c2622bd2e212be9deb57f93</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2006</creationdate><topic>Alkaline phosphatase</topic><topic>Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy</topic><topic>Animals</topic><topic>Anti-Bacterial Agents</topic><topic>Applied cell therapy and gene therapy</topic><topic>Biological and medical sciences</topic><topic>Biomedical and Life Sciences</topic><topic>Biomedicine</topic><topic>Biotechnology</topic><topic>Bone diseases</topic><topic>Bone growth</topic><topic>Bone Transplantation</topic><topic>Bones</topic><topic>Care and treatment</topic><topic>Cbfa-1 protein</topic><topic>Cell Biology</topic><topic>Cell culture</topic><topic>Cells</topic><topic>Cells, Cultured</topic><topic>Collagen</topic><topic>Computed tomography</topic><topic>Core Binding Factor Alpha 1 Subunit - genetics</topic><topic>Core Binding Factor Alpha 1 Subunit - metabolism</topic><topic>Culture media</topic><topic>DNA binding proteins</topic><topic>Drinking water</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Gene Expression</topic><topic>Gene Expression Regulation</topic><topic>Gene Therapy</topic><topic>Genetic aspects</topic><topic>Genetic Therapy - methods</topic><topic>Genetic Vectors - administration & dosage</topic><topic>Growth</topic><topic>Health aspects</topic><topic>Health. Pharmaceutical industry</topic><topic>Hindlimb</topic><topic>Human Genetics</topic><topic>Industrial applications and implications. Economical aspects</topic><topic>Male</topic><topic>Medical sciences</topic><topic>Mice</topic><topic>Mice, Inbred BALB C</topic><topic>Mineralization</topic><topic>Myoblasts</topic><topic>Nanotechnology</topic><topic>original-article</topic><topic>Orthopedics</topic><topic>Osteoblastogenesis</topic><topic>Osteoblasts</topic><topic>Osteoblasts - cytology</topic><topic>Osteoblasts - diagnostic imaging</topic><topic>Osteoblasts - metabolism</topic><topic>Osteogenesis</topic><topic>Physiological aspects</topic><topic>Regenerative medicine</topic><topic>Retroviridae - genetics</topic><topic>Tetracycline</topic><topic>Tissue engineering</topic><topic>Tissue Engineering - methods</topic><topic>Tomography, X-Ray Computed</topic><topic>Trans-Activators</topic><topic>Transduction, Genetic - methods</topic><topic>Transfusions. Complications. Transfusion reactions. Cell and gene therapy</topic><topic>Tumorigenesis</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Gersbach, C A</creatorcontrib><creatorcontrib>Le Doux, J M</creatorcontrib><creatorcontrib>Guldberg, R E</creatorcontrib><creatorcontrib>García, A J</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>Gale In Context: Science</collection><collection>ProQuest Central (Corporate)</collection><collection>Calcium & Calcified Tissue Abstracts</collection><collection>Neurosciences Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>Public Health Database</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>Research Library (Alumni Edition)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</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>Research Library Prep</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>Research Library</collection><collection>Biological Science Database</collection><collection>Research Library (Corporate)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><collection>ProQuest Central Basic</collection><collection>Genetics Abstracts</collection><collection>Biotechnology Research Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Gene therapy</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Gersbach, C A</au><au>Le Doux, J M</au><au>Guldberg, R E</au><au>García, A J</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Inducible regulation of Runx2-stimulated osteogenesis</atitle><jtitle>Gene therapy</jtitle><stitle>Gene Ther</stitle><addtitle>Gene Ther</addtitle><date>2006-06-01</date><risdate>2006</risdate><volume>13</volume><issue>11</issue><spage>873</spage><epage>882</epage><pages>873-882</pages><issn>0969-7128</issn><eissn>1476-5462</eissn><abstract>Ex vivo
gene therapy is a promising approach to orthopedic regenerative medicine. These strategies typically focus on the constitutive overexpression of osteogenic factors to induce osteoblastic differentiation and matrix mineralization. However, the unregulated production of osteoinductive molecules has also resulted in abnormal bone formation and tumorigenesis. To address these limitations, this work describes a retroviral system to deliver the Runx2 osteoblastic transcription factor under control of the tetracycline-inducible (tet-off) promoter in primary skeletal myoblasts. Runx2 expression was tightly regulated by anhydrotetracyline (aTc) concentration in cell culture media. Osteoblastic gene expression, alkaline phosphatase activity, and matrix mineralization were also controlled by aTc in a dose-dependent manner. Additionally, osteoblastic differentiation was temporally regulated by adding and removing aTc from the culture media. Engineered cells were seeded onto collagen scaffolds and implanted intramuscularly in the hind limbs of syngeneic mice.
In vivo
mineralization by these constructs was regulated by supplementing the drinking water with aTc, as demonstrated by micro-computed tomography and histological analyses. Collectively, these results present a novel system for regulating osteoblastic differentiation of a clinically relevant autologous cell source. This system is significant to developing controlled and effective orthopedic gene therapy strategies and studying the regulation of osteoblastic differentiation.</abstract><cop>London</cop><pub>Nature Publishing Group UK</pub><pmid>16496016</pmid><doi>10.1038/sj.gt.3302725</doi><tpages>10</tpages><oa>free_for_read</oa></addata></record> |
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| issn | 0969-7128 1476-5462 |
| language | eng |
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| source | MEDLINE; EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection |
| subjects | Alkaline phosphatase Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy Animals Anti-Bacterial Agents Applied cell therapy and gene therapy Biological and medical sciences Biomedical and Life Sciences Biomedicine Biotechnology Bone diseases Bone growth Bone Transplantation Bones Care and treatment Cbfa-1 protein Cell Biology Cell culture Cells Cells, Cultured Collagen Computed tomography Core Binding Factor Alpha 1 Subunit - genetics Core Binding Factor Alpha 1 Subunit - metabolism Culture media DNA binding proteins Drinking water Fundamental and applied biological sciences. Psychology Gene Expression Gene Expression Regulation Gene Therapy Genetic aspects Genetic Therapy - methods Genetic Vectors - administration & dosage Growth Health aspects Health. Pharmaceutical industry Hindlimb Human Genetics Industrial applications and implications. Economical aspects Male Medical sciences Mice Mice, Inbred BALB C Mineralization Myoblasts Nanotechnology original-article Orthopedics Osteoblastogenesis Osteoblasts Osteoblasts - cytology Osteoblasts - diagnostic imaging Osteoblasts - metabolism Osteogenesis Physiological aspects Regenerative medicine Retroviridae - genetics Tetracycline Tissue engineering Tissue Engineering - methods Tomography, X-Ray Computed Trans-Activators Transduction, Genetic - methods Transfusions. Complications. Transfusion reactions. Cell and gene therapy Tumorigenesis |
| title | Inducible regulation of Runx2-stimulated osteogenesis |
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