Green Fluorescent Protein
Human Parvovirus B19 (PVB19) has been associated with myocarditis putative due to endothelial infection. Whether PVB19 infects endothelial cells and causes a modification of endothelial function and inflammation and, thus, disturbance of microcirculation has not been elucidated and could not be visu...
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| Veröffentlicht in: | PloS one 2012-03, Vol.7 (3), p.e33602 |
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| creator | Wurster, Thomas Pölzelbauer, Catharina Schönberger, Tanja Paul, Angela Seizer, Peter Stellos, Konstantinos Schuster, Andreas Botnar, Rene M Gawaz, Meinrad Bigalke, Boris |
| description | Human Parvovirus B19 (PVB19) has been associated with myocarditis putative due to endothelial infection. Whether PVB19 infects endothelial cells and causes a modification of endothelial function and inflammation and, thus, disturbance of microcirculation has not been elucidated and could not be visualized so far. To examine the PVB19-induced endothelial modification, we used green fluorescent protein (GFP) color reporter gene in the non-structural segment 1 (NS1) of PVB19. NS1-GFP-PVB19 or GFP plasmid as control were transfected in an endothelial-like cell line (ECV304). The endothelial surface expression of intercellular-adhesion molecule-1 (CD54/ICAM-1) and extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) were evaluated by flow cytometry after NS-1-GFP or control-GFP transfection. To evaluate platelet adhesion on NS-1 transfected ECs, we performed a dynamic adhesion assay (flow chamber). NS-1 transfection causes endothelial activation and enhanced expression of ICAM-1 (CD54: mean±standard deviation: NS1-GFP vs. control-GFP: 85.3±11.2 vs. 61.6±8.1; P |
| doi_str_mv | 10.1371/journal.pone.0033602 |
| format | Article |
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Whether PVB19 infects endothelial cells and causes a modification of endothelial function and inflammation and, thus, disturbance of microcirculation has not been elucidated and could not be visualized so far. To examine the PVB19-induced endothelial modification, we used green fluorescent protein (GFP) color reporter gene in the non-structural segment 1 (NS1) of PVB19. NS1-GFP-PVB19 or GFP plasmid as control were transfected in an endothelial-like cell line (ECV304). The endothelial surface expression of intercellular-adhesion molecule-1 (CD54/ICAM-1) and extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) were evaluated by flow cytometry after NS-1-GFP or control-GFP transfection. To evaluate platelet adhesion on NS-1 transfected ECs, we performed a dynamic adhesion assay (flow chamber). NS-1 transfection causes endothelial activation and enhanced expression of ICAM-1 (CD54: mean±standard deviation: NS1-GFP vs. control-GFP: 85.3±11.2 vs. 61.6±8.1; P<0.05) and induces endothelial expression of EMMPRIN/CD147 (CD147: mean±SEM: NS1-GFP vs. control-GFP: 114±15.3 vs. 80±0.91; P<0.05) compared to control-GFP transfected cells. Dynamic adhesion assays showed that adhesion of platelets is significantly enhanced on NS1 transfected ECs when compared to control-GFP (P<0.05). The transfection of ECs was verified simultaneously through flow cytometry, immunofluorescence microscopy and polymerase chain reaction (PCR) analysis. GFP color reporter gene shows transfection of ECs and may help to visualize NS1-PVB19 induced endothelial activation and platelet adhesion as well as an enhanced monocyte adhesion directly, providing in vitro evidence of possible microcirculatory dysfunction in PVB19-induced myocarditis and, thus, myocardial tissue damage.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0033602</identifier><language>eng</language><publisher>Public Library of Science</publisher><subject>Analysis ; Endothelium ; Flow (Dynamics) ; Fluorescence ; Genes ; Health aspects ; Infection ; Inflammation</subject><ispartof>PloS one, 2012-03, Vol.7 (3), p.e33602</ispartof><rights>COPYRIGHT 2012 Public Library of Science</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,860,27901,27902</link.rule.ids></links><search><creatorcontrib>Wurster, Thomas</creatorcontrib><creatorcontrib>Pölzelbauer, Catharina</creatorcontrib><creatorcontrib>Schönberger, Tanja</creatorcontrib><creatorcontrib>Paul, Angela</creatorcontrib><creatorcontrib>Seizer, Peter</creatorcontrib><creatorcontrib>Stellos, Konstantinos</creatorcontrib><creatorcontrib>Schuster, Andreas</creatorcontrib><creatorcontrib>Botnar, Rene M</creatorcontrib><creatorcontrib>Gawaz, Meinrad</creatorcontrib><creatorcontrib>Bigalke, Boris</creatorcontrib><title>Green Fluorescent Protein</title><title>PloS one</title><description>Human Parvovirus B19 (PVB19) has been associated with myocarditis putative due to endothelial infection. Whether PVB19 infects endothelial cells and causes a modification of endothelial function and inflammation and, thus, disturbance of microcirculation has not been elucidated and could not be visualized so far. To examine the PVB19-induced endothelial modification, we used green fluorescent protein (GFP) color reporter gene in the non-structural segment 1 (NS1) of PVB19. NS1-GFP-PVB19 or GFP plasmid as control were transfected in an endothelial-like cell line (ECV304). The endothelial surface expression of intercellular-adhesion molecule-1 (CD54/ICAM-1) and extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) were evaluated by flow cytometry after NS-1-GFP or control-GFP transfection. To evaluate platelet adhesion on NS-1 transfected ECs, we performed a dynamic adhesion assay (flow chamber). NS-1 transfection causes endothelial activation and enhanced expression of ICAM-1 (CD54: mean±standard deviation: NS1-GFP vs. control-GFP: 85.3±11.2 vs. 61.6±8.1; P<0.05) and induces endothelial expression of EMMPRIN/CD147 (CD147: mean±SEM: NS1-GFP vs. control-GFP: 114±15.3 vs. 80±0.91; P<0.05) compared to control-GFP transfected cells. Dynamic adhesion assays showed that adhesion of platelets is significantly enhanced on NS1 transfected ECs when compared to control-GFP (P<0.05). The transfection of ECs was verified simultaneously through flow cytometry, immunofluorescence microscopy and polymerase chain reaction (PCR) analysis. GFP color reporter gene shows transfection of ECs and may help to visualize NS1-PVB19 induced endothelial activation and platelet adhesion as well as an enhanced monocyte adhesion directly, providing in vitro evidence of possible microcirculatory dysfunction in PVB19-induced myocarditis and, thus, myocardial tissue damage.</description><subject>Analysis</subject><subject>Endothelium</subject><subject>Flow (Dynamics)</subject><subject>Fluorescence</subject><subject>Genes</subject><subject>Health aspects</subject><subject>Infection</subject><subject>Inflammation</subject><issn>1932-6203</issn><issn>1932-6203</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2012</creationdate><recordtype>article</recordtype><recordid>eNqFzEFLwzAYxvEgCs7pBxA87CR4aE3yZklzHMPNwWCiw2tJ2zddR0ikScGPb2Ee5snT_zn8eAh5YDRnoNjzMQy9Ny7_Ch5zSgEk5RdkwjTwTHIKl2f7mtzEeKR0DoWUE3K_7hH9bOWG0GOs0afZWx8Sdv6WXFnjIt79dkr2q5f98jXb7tab5WKbtVqLzBhlRa2AjtdSNAWXDdaVoqZWtpkr0WhtwUqNoEdnuawYG1tVCMJaW8CUPJ1uW-Ow7HwdfMLv1JohxnLz8V4uhFK0AC7EP3b3-dc-ntkDGpcOMbghdcHHc_gDH5lczg</recordid><startdate>20120315</startdate><enddate>20120315</enddate><creator>Wurster, Thomas</creator><creator>Pölzelbauer, Catharina</creator><creator>Schönberger, Tanja</creator><creator>Paul, Angela</creator><creator>Seizer, Peter</creator><creator>Stellos, Konstantinos</creator><creator>Schuster, Andreas</creator><creator>Botnar, Rene M</creator><creator>Gawaz, Meinrad</creator><creator>Bigalke, Boris</creator><general>Public Library of Science</general><scope>IOV</scope><scope>ISR</scope></search><sort><creationdate>20120315</creationdate><title>Green Fluorescent Protein</title><author>Wurster, Thomas ; Pölzelbauer, Catharina ; Schönberger, Tanja ; Paul, Angela ; Seizer, Peter ; Stellos, Konstantinos ; Schuster, Andreas ; Botnar, Rene M ; Gawaz, Meinrad ; Bigalke, Boris</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-g994-aa7f4c73020364d826decb70ac7fd574d99f3f69e39f4cf26b114cfbbe34fff83</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2012</creationdate><topic>Analysis</topic><topic>Endothelium</topic><topic>Flow (Dynamics)</topic><topic>Fluorescence</topic><topic>Genes</topic><topic>Health aspects</topic><topic>Infection</topic><topic>Inflammation</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Wurster, Thomas</creatorcontrib><creatorcontrib>Pölzelbauer, Catharina</creatorcontrib><creatorcontrib>Schönberger, Tanja</creatorcontrib><creatorcontrib>Paul, Angela</creatorcontrib><creatorcontrib>Seizer, Peter</creatorcontrib><creatorcontrib>Stellos, Konstantinos</creatorcontrib><creatorcontrib>Schuster, Andreas</creatorcontrib><creatorcontrib>Botnar, Rene M</creatorcontrib><creatorcontrib>Gawaz, Meinrad</creatorcontrib><creatorcontrib>Bigalke, Boris</creatorcontrib><collection>Gale In Context: Opposing Viewpoints</collection><collection>Gale In Context: Science</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Wurster, Thomas</au><au>Pölzelbauer, Catharina</au><au>Schönberger, Tanja</au><au>Paul, Angela</au><au>Seizer, Peter</au><au>Stellos, Konstantinos</au><au>Schuster, Andreas</au><au>Botnar, Rene M</au><au>Gawaz, Meinrad</au><au>Bigalke, Boris</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Green Fluorescent Protein</atitle><jtitle>PloS one</jtitle><date>2012-03-15</date><risdate>2012</risdate><volume>7</volume><issue>3</issue><spage>e33602</spage><pages>e33602-</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Human Parvovirus B19 (PVB19) has been associated with myocarditis putative due to endothelial infection. Whether PVB19 infects endothelial cells and causes a modification of endothelial function and inflammation and, thus, disturbance of microcirculation has not been elucidated and could not be visualized so far. To examine the PVB19-induced endothelial modification, we used green fluorescent protein (GFP) color reporter gene in the non-structural segment 1 (NS1) of PVB19. NS1-GFP-PVB19 or GFP plasmid as control were transfected in an endothelial-like cell line (ECV304). The endothelial surface expression of intercellular-adhesion molecule-1 (CD54/ICAM-1) and extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) were evaluated by flow cytometry after NS-1-GFP or control-GFP transfection. To evaluate platelet adhesion on NS-1 transfected ECs, we performed a dynamic adhesion assay (flow chamber). NS-1 transfection causes endothelial activation and enhanced expression of ICAM-1 (CD54: mean±standard deviation: NS1-GFP vs. control-GFP: 85.3±11.2 vs. 61.6±8.1; P<0.05) and induces endothelial expression of EMMPRIN/CD147 (CD147: mean±SEM: NS1-GFP vs. control-GFP: 114±15.3 vs. 80±0.91; P<0.05) compared to control-GFP transfected cells. Dynamic adhesion assays showed that adhesion of platelets is significantly enhanced on NS1 transfected ECs when compared to control-GFP (P<0.05). The transfection of ECs was verified simultaneously through flow cytometry, immunofluorescence microscopy and polymerase chain reaction (PCR) analysis. GFP color reporter gene shows transfection of ECs and may help to visualize NS1-PVB19 induced endothelial activation and platelet adhesion as well as an enhanced monocyte adhesion directly, providing in vitro evidence of possible microcirculatory dysfunction in PVB19-induced myocarditis and, thus, myocardial tissue damage.</abstract><pub>Public Library of Science</pub><doi>10.1371/journal.pone.0033602</doi><tpages>e33602</tpages></addata></record> |
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| source | DOAJ Directory of Open Access Journals; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; PubMed Central; Free Full-Text Journals in Chemistry; Public Library of Science (PLoS) |
| subjects | Analysis Endothelium Flow (Dynamics) Fluorescence Genes Health aspects Infection Inflammation |
| title | Green Fluorescent Protein |
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