BDNF and NT-3 expression by using glucocorticoid-induced bicistronic expression vector pGC-BDNF-IRES-NT3 protects apoptotic cells in a cellular injury model
Abstract Spinal cord injury (SCI) is a severe traumatic disease in the central nervous system with high incidence and high morbidity. Recent study demonstrated that cell transplantation therapy may improve local microenvironment of the injury site and promote nerve regeneration to restore spinal cor...
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| Veröffentlicht in: | Brain research 2012-04, Vol.1448, p.137-143 |
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| creator | Wang, Yongxiang Gu, Jiaxiang Wang, Jingcheng Feng, Xingmin Tao, Yuping Jiang, Baichuan He, Jinshan Wang, Qiang Yang, Jiandong Zhang, Shenfei Cai, Jun Sun, Yu |
| description | Abstract Spinal cord injury (SCI) is a severe traumatic disease in the central nervous system with high incidence and high morbidity. Recent study demonstrated that cell transplantation therapy may improve local microenvironment of the injury site and promote nerve regeneration to restore spinal cord functions. In this study, we constructed a glucocorticoid-induced bicistronic eukaryotic expression vector pGC-BDNF-IRES-NT3 by using molecular cloning techniques and examined the protective effect of neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) expressed by this vector in a rat spinal cord injury (SCI) model. We first connected glucocorticoid response element (GRE) to cytomegalovirus (CMV) promoter and then the GRE-CMV gene was inserted into pEGFP-1 vector to construct the eukaryotic expression vector pGC-EGFP. Western blot analysis was used to confirm the expression of EGFP by transfecting this vector in RN-DSC cells. The IRES was used to connect BDNF gene and NT-3 gene and replaced the EGFP gene in pGC-EGFP plasmid to form the bicistronic expression vector-pGC-BDNF-IRES-NT3. After RN-DSC cells were transfected with the plasmid and treated with glucocorticoid, BDNF and NT-3 expression in the culture medium were measured by ELISA method. Finally, we found that combination therapy with the transfection of this vector and glucocorticoid had an anti-apoptotic effect in a cellular SCI model of RN-DSC cells. Therefore, the co-expression of BDNF and NT-3 by using this vector rescued the injured cells. This provided useful information for the gene-modification cell transplantation combined with glucocorticoid for the treatment of SCI. |
| doi_str_mv | 10.1016/j.brainres.2012.02.007 |
| format | Article |
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Recent study demonstrated that cell transplantation therapy may improve local microenvironment of the injury site and promote nerve regeneration to restore spinal cord functions. In this study, we constructed a glucocorticoid-induced bicistronic eukaryotic expression vector pGC-BDNF-IRES-NT3 by using molecular cloning techniques and examined the protective effect of neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) expressed by this vector in a rat spinal cord injury (SCI) model. We first connected glucocorticoid response element (GRE) to cytomegalovirus (CMV) promoter and then the GRE-CMV gene was inserted into pEGFP-1 vector to construct the eukaryotic expression vector pGC-EGFP. Western blot analysis was used to confirm the expression of EGFP by transfecting this vector in RN-DSC cells. The IRES was used to connect BDNF gene and NT-3 gene and replaced the EGFP gene in pGC-EGFP plasmid to form the bicistronic expression vector-pGC-BDNF-IRES-NT3. After RN-DSC cells were transfected with the plasmid and treated with glucocorticoid, BDNF and NT-3 expression in the culture medium were measured by ELISA method. Finally, we found that combination therapy with the transfection of this vector and glucocorticoid had an anti-apoptotic effect in a cellular SCI model of RN-DSC cells. Therefore, the co-expression of BDNF and NT-3 by using this vector rescued the injured cells. This provided useful information for the gene-modification cell transplantation combined with glucocorticoid for the treatment of SCI.</description><identifier>ISSN: 0006-8993</identifier><identifier>EISSN: 1872-6240</identifier><identifier>DOI: 10.1016/j.brainres.2012.02.007</identifier><identifier>PMID: 22373651</identifier><identifier>CODEN: BRREAP</identifier><language>eng</language><publisher>Amsterdam: Elsevier B.V</publisher><subject>Ageing, cell death ; Animals ; apoptosis ; Apoptosis - physiology ; BDNF ; Bicistronic expression vector, Apoptosis ; Biological and medical sciences ; Blotting, Western ; brain ; Brain-Derived Neurotrophic Factor - biosynthesis ; Cell Line ; Cell physiology ; cell transplantation ; Cells, Cultured ; culture media ; Cytomegalovirus ; Cytomegalovirus - genetics ; DNA Primers ; Enzyme-Linked Immunosorbent Assay ; Fundamental and applied biological sciences. Psychology ; Gene Expression - drug effects ; genes ; Genetic Vectors - genetics ; Glucocorticoids - pharmacology ; Green Fluorescent Proteins ; Indicators and Reagents ; Microscopy, Fluorescence ; Molecular and cellular biology ; molecular cloning ; morbidity ; Mutant Chimeric Proteins - genetics ; nerve tissue ; Neurology ; Neurons - physiology ; Neurotrophin 3 - biosynthesis ; Neurotrophin-3 ; plasmids ; Plasmids - genetics ; protective effect ; Rats ; Response Elements - genetics ; spinal cord ; Spinal Cord - cytology ; Spinal Cord - drug effects ; Spinal Cord - metabolism ; Spinal cord injury ; Transfection ; Western blotting</subject><ispartof>Brain research, 2012-04, Vol.1448, p.137-143</ispartof><rights>Elsevier B.V.</rights><rights>2012 Elsevier B.V.</rights><rights>2015 INIST-CNRS</rights><rights>Copyright © 2012 Elsevier B.V. All rights reserved.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c575t-e27ea61a503a329ef6fb8a2f91d53e4bdd8758b35d0c47efa88c87ae524a4e743</citedby><cites>FETCH-LOGICAL-c575t-e27ea61a503a329ef6fb8a2f91d53e4bdd8758b35d0c47efa88c87ae524a4e743</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/j.brainres.2012.02.007$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3550,27924,27925,45995</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=25755159$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/22373651$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Wang, Yongxiang</creatorcontrib><creatorcontrib>Gu, Jiaxiang</creatorcontrib><creatorcontrib>Wang, Jingcheng</creatorcontrib><creatorcontrib>Feng, Xingmin</creatorcontrib><creatorcontrib>Tao, Yuping</creatorcontrib><creatorcontrib>Jiang, Baichuan</creatorcontrib><creatorcontrib>He, Jinshan</creatorcontrib><creatorcontrib>Wang, Qiang</creatorcontrib><creatorcontrib>Yang, Jiandong</creatorcontrib><creatorcontrib>Zhang, Shenfei</creatorcontrib><creatorcontrib>Cai, Jun</creatorcontrib><creatorcontrib>Sun, Yu</creatorcontrib><title>BDNF and NT-3 expression by using glucocorticoid-induced bicistronic expression vector pGC-BDNF-IRES-NT3 protects apoptotic cells in a cellular injury model</title><title>Brain research</title><addtitle>Brain Res</addtitle><description>Abstract Spinal cord injury (SCI) is a severe traumatic disease in the central nervous system with high incidence and high morbidity. Recent study demonstrated that cell transplantation therapy may improve local microenvironment of the injury site and promote nerve regeneration to restore spinal cord functions. In this study, we constructed a glucocorticoid-induced bicistronic eukaryotic expression vector pGC-BDNF-IRES-NT3 by using molecular cloning techniques and examined the protective effect of neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) expressed by this vector in a rat spinal cord injury (SCI) model. We first connected glucocorticoid response element (GRE) to cytomegalovirus (CMV) promoter and then the GRE-CMV gene was inserted into pEGFP-1 vector to construct the eukaryotic expression vector pGC-EGFP. Western blot analysis was used to confirm the expression of EGFP by transfecting this vector in RN-DSC cells. The IRES was used to connect BDNF gene and NT-3 gene and replaced the EGFP gene in pGC-EGFP plasmid to form the bicistronic expression vector-pGC-BDNF-IRES-NT3. After RN-DSC cells were transfected with the plasmid and treated with glucocorticoid, BDNF and NT-3 expression in the culture medium were measured by ELISA method. Finally, we found that combination therapy with the transfection of this vector and glucocorticoid had an anti-apoptotic effect in a cellular SCI model of RN-DSC cells. Therefore, the co-expression of BDNF and NT-3 by using this vector rescued the injured cells. This provided useful information for the gene-modification cell transplantation combined with glucocorticoid for the treatment of SCI.</description><subject>Ageing, cell death</subject><subject>Animals</subject><subject>apoptosis</subject><subject>Apoptosis - physiology</subject><subject>BDNF</subject><subject>Bicistronic expression vector, Apoptosis</subject><subject>Biological and medical sciences</subject><subject>Blotting, Western</subject><subject>brain</subject><subject>Brain-Derived Neurotrophic Factor - biosynthesis</subject><subject>Cell Line</subject><subject>Cell physiology</subject><subject>cell transplantation</subject><subject>Cells, Cultured</subject><subject>culture media</subject><subject>Cytomegalovirus</subject><subject>Cytomegalovirus - genetics</subject><subject>DNA Primers</subject><subject>Enzyme-Linked Immunosorbent Assay</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Gene Expression - drug effects</subject><subject>genes</subject><subject>Genetic Vectors - genetics</subject><subject>Glucocorticoids - pharmacology</subject><subject>Green Fluorescent Proteins</subject><subject>Indicators and Reagents</subject><subject>Microscopy, Fluorescence</subject><subject>Molecular and cellular biology</subject><subject>molecular cloning</subject><subject>morbidity</subject><subject>Mutant Chimeric Proteins - genetics</subject><subject>nerve tissue</subject><subject>Neurology</subject><subject>Neurons - physiology</subject><subject>Neurotrophin 3 - biosynthesis</subject><subject>Neurotrophin-3</subject><subject>plasmids</subject><subject>Plasmids - genetics</subject><subject>protective effect</subject><subject>Rats</subject><subject>Response Elements - genetics</subject><subject>spinal cord</subject><subject>Spinal Cord - cytology</subject><subject>Spinal Cord - drug effects</subject><subject>Spinal Cord - metabolism</subject><subject>Spinal cord injury</subject><subject>Transfection</subject><subject>Western blotting</subject><issn>0006-8993</issn><issn>1872-6240</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2012</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFks9u1DAQxiMEotvCKxRfEL1k8Z84Ti4IWNpSqVokdnu2HGey8pKNg51U7LvwsEzYLSAOII1kj_ybb0b-JknOGZ0zyvLX23kVjOsCxDmnjM8pBlWPkhkrFE9zntHHyYxSmqdFWYqT5DTGLaZClPRpcsK5UCKXbJZ8f_9heUVMV5PlOhUEvvUoGZ3vSLUnY3Tdhmza0Xrrw-Csd3Xqunq0UJPKWReH4Dtn_yy7Bzv4QPrrRTpJpzefL1fpci1IH_yAb5GY3veDRzVioW0jcR0xP69jawJm2zHsyc7X0D5LnjSmjfD8eJ4ld1eX68XH9PbT9c3i3W1qpZJDClyByZmRVBjBS2jypioMb0pWSwFZVdeFkkUlZE1tpqAxRWELZUDyzGSgMnGWvDro4oxfR4iD3rk4TWQ68GPUJS8lzWg-kRf_JFmeU1EooXJE8wNqg48xQKP74HYm7DWjevJQb_WDh3ryUFMMqrDw_NhjrHZQ_yp7MA2Bl0fARGvaJpgOrfjN4adIJkvkXhy4xnhtNgGZuxV2krgIZcYzjsTbAwH4u_cOgo7WQYf2uoBe6dq7_0_75i8J2zrcCdN-gT3ErR9Dh95ppiMW6NW0k9NKMk4pFxg_ACY73P0</recordid><startdate>20120411</startdate><enddate>20120411</enddate><creator>Wang, Yongxiang</creator><creator>Gu, Jiaxiang</creator><creator>Wang, Jingcheng</creator><creator>Feng, Xingmin</creator><creator>Tao, Yuping</creator><creator>Jiang, Baichuan</creator><creator>He, Jinshan</creator><creator>Wang, Qiang</creator><creator>Yang, Jiandong</creator><creator>Zhang, Shenfei</creator><creator>Cai, Jun</creator><creator>Sun, Yu</creator><general>Elsevier B.V</general><general>Elsevier</general><scope>FBQ</scope><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>7TK</scope><scope>7X8</scope></search><sort><creationdate>20120411</creationdate><title>BDNF and NT-3 expression by using glucocorticoid-induced bicistronic expression vector pGC-BDNF-IRES-NT3 protects apoptotic cells in a cellular injury model</title><author>Wang, Yongxiang ; Gu, Jiaxiang ; Wang, Jingcheng ; Feng, Xingmin ; Tao, Yuping ; Jiang, Baichuan ; He, Jinshan ; Wang, Qiang ; Yang, Jiandong ; Zhang, Shenfei ; Cai, Jun ; Sun, Yu</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c575t-e27ea61a503a329ef6fb8a2f91d53e4bdd8758b35d0c47efa88c87ae524a4e743</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2012</creationdate><topic>Ageing, cell death</topic><topic>Animals</topic><topic>apoptosis</topic><topic>Apoptosis - physiology</topic><topic>BDNF</topic><topic>Bicistronic expression vector, Apoptosis</topic><topic>Biological and medical sciences</topic><topic>Blotting, Western</topic><topic>brain</topic><topic>Brain-Derived Neurotrophic Factor - biosynthesis</topic><topic>Cell Line</topic><topic>Cell physiology</topic><topic>cell transplantation</topic><topic>Cells, Cultured</topic><topic>culture media</topic><topic>Cytomegalovirus</topic><topic>Cytomegalovirus - genetics</topic><topic>DNA Primers</topic><topic>Enzyme-Linked Immunosorbent Assay</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Gene Expression - drug effects</topic><topic>genes</topic><topic>Genetic Vectors - genetics</topic><topic>Glucocorticoids - pharmacology</topic><topic>Green Fluorescent Proteins</topic><topic>Indicators and Reagents</topic><topic>Microscopy, Fluorescence</topic><topic>Molecular and cellular biology</topic><topic>molecular cloning</topic><topic>morbidity</topic><topic>Mutant Chimeric Proteins - genetics</topic><topic>nerve tissue</topic><topic>Neurology</topic><topic>Neurons - physiology</topic><topic>Neurotrophin 3 - biosynthesis</topic><topic>Neurotrophin-3</topic><topic>plasmids</topic><topic>Plasmids - genetics</topic><topic>protective effect</topic><topic>Rats</topic><topic>Response Elements - genetics</topic><topic>spinal cord</topic><topic>Spinal Cord - cytology</topic><topic>Spinal Cord - drug effects</topic><topic>Spinal Cord - metabolism</topic><topic>Spinal cord injury</topic><topic>Transfection</topic><topic>Western blotting</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Wang, Yongxiang</creatorcontrib><creatorcontrib>Gu, Jiaxiang</creatorcontrib><creatorcontrib>Wang, Jingcheng</creatorcontrib><creatorcontrib>Feng, Xingmin</creatorcontrib><creatorcontrib>Tao, Yuping</creatorcontrib><creatorcontrib>Jiang, Baichuan</creatorcontrib><creatorcontrib>He, Jinshan</creatorcontrib><creatorcontrib>Wang, Qiang</creatorcontrib><creatorcontrib>Yang, Jiandong</creatorcontrib><creatorcontrib>Zhang, Shenfei</creatorcontrib><creatorcontrib>Cai, Jun</creatorcontrib><creatorcontrib>Sun, Yu</creatorcontrib><collection>AGRIS</collection><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>Neurosciences Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Brain research</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Wang, Yongxiang</au><au>Gu, Jiaxiang</au><au>Wang, Jingcheng</au><au>Feng, Xingmin</au><au>Tao, Yuping</au><au>Jiang, Baichuan</au><au>He, Jinshan</au><au>Wang, Qiang</au><au>Yang, Jiandong</au><au>Zhang, Shenfei</au><au>Cai, Jun</au><au>Sun, Yu</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>BDNF and NT-3 expression by using glucocorticoid-induced bicistronic expression vector pGC-BDNF-IRES-NT3 protects apoptotic cells in a cellular injury model</atitle><jtitle>Brain research</jtitle><addtitle>Brain Res</addtitle><date>2012-04-11</date><risdate>2012</risdate><volume>1448</volume><spage>137</spage><epage>143</epage><pages>137-143</pages><issn>0006-8993</issn><eissn>1872-6240</eissn><coden>BRREAP</coden><abstract>Abstract Spinal cord injury (SCI) is a severe traumatic disease in the central nervous system with high incidence and high morbidity. Recent study demonstrated that cell transplantation therapy may improve local microenvironment of the injury site and promote nerve regeneration to restore spinal cord functions. In this study, we constructed a glucocorticoid-induced bicistronic eukaryotic expression vector pGC-BDNF-IRES-NT3 by using molecular cloning techniques and examined the protective effect of neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) expressed by this vector in a rat spinal cord injury (SCI) model. We first connected glucocorticoid response element (GRE) to cytomegalovirus (CMV) promoter and then the GRE-CMV gene was inserted into pEGFP-1 vector to construct the eukaryotic expression vector pGC-EGFP. Western blot analysis was used to confirm the expression of EGFP by transfecting this vector in RN-DSC cells. The IRES was used to connect BDNF gene and NT-3 gene and replaced the EGFP gene in pGC-EGFP plasmid to form the bicistronic expression vector-pGC-BDNF-IRES-NT3. After RN-DSC cells were transfected with the plasmid and treated with glucocorticoid, BDNF and NT-3 expression in the culture medium were measured by ELISA method. Finally, we found that combination therapy with the transfection of this vector and glucocorticoid had an anti-apoptotic effect in a cellular SCI model of RN-DSC cells. Therefore, the co-expression of BDNF and NT-3 by using this vector rescued the injured cells. This provided useful information for the gene-modification cell transplantation combined with glucocorticoid for the treatment of SCI.</abstract><cop>Amsterdam</cop><pub>Elsevier B.V</pub><pmid>22373651</pmid><doi>10.1016/j.brainres.2012.02.007</doi><tpages>7</tpages></addata></record> |
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| subjects | Ageing, cell death Animals apoptosis Apoptosis - physiology BDNF Bicistronic expression vector, Apoptosis Biological and medical sciences Blotting, Western brain Brain-Derived Neurotrophic Factor - biosynthesis Cell Line Cell physiology cell transplantation Cells, Cultured culture media Cytomegalovirus Cytomegalovirus - genetics DNA Primers Enzyme-Linked Immunosorbent Assay Fundamental and applied biological sciences. Psychology Gene Expression - drug effects genes Genetic Vectors - genetics Glucocorticoids - pharmacology Green Fluorescent Proteins Indicators and Reagents Microscopy, Fluorescence Molecular and cellular biology molecular cloning morbidity Mutant Chimeric Proteins - genetics nerve tissue Neurology Neurons - physiology Neurotrophin 3 - biosynthesis Neurotrophin-3 plasmids Plasmids - genetics protective effect Rats Response Elements - genetics spinal cord Spinal Cord - cytology Spinal Cord - drug effects Spinal Cord - metabolism Spinal cord injury Transfection Western blotting |
| title | BDNF and NT-3 expression by using glucocorticoid-induced bicistronic expression vector pGC-BDNF-IRES-NT3 protects apoptotic cells in a cellular injury model |
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