Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification.
BACKGROUND: 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...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2012-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC3305306?pdf=render |
| _version_ | 1811274311624818688 |
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| author | Thomas Wurster Catharina Pölzelbauer Tanja Schönberger Angela Paul Peter Seizer Konstantinos Stellos Andreas Schuster Rene M Botnar Rene M Botnar Meinrad Gawaz Boris Bigalke |
| author_facet | Thomas Wurster Catharina Pölzelbauer Tanja Schönberger Angela Paul Peter Seizer Konstantinos Stellos Andreas Schuster Rene M Botnar Rene M Botnar Meinrad Gawaz Boris Bigalke |
| author_sort | Thomas Wurster |
| collection | DOAJ |
| description | BACKGROUND: 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. METHODS AND FINDINGS: 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. CONCLUSIONS: 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. |
| first_indexed | 2024-04-12T23:16:43Z |
| format | Article |
| id | doaj.art-fecc628ca9844e76b723367ceab100cf |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-04-12T23:16:43Z |
| publishDate | 2012-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-fecc628ca9844e76b723367ceab100cf2022-12-22T03:12:40ZengPublic Library of Science (PLoS)PLoS ONE1932-62032012-01-0173e3360210.1371/journal.pone.0033602Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification.Thomas WursterCatharina PölzelbauerTanja SchönbergerAngela PaulPeter SeizerKonstantinos StellosAndreas SchusterRene M BotnarRene M BotnarMeinrad GawazBoris BigalkeBACKGROUND: 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. METHODS AND FINDINGS: 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. CONCLUSIONS: 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.http://europepmc.org/articles/PMC3305306?pdf=render |
| spellingShingle | Thomas Wurster Catharina Pölzelbauer Tanja Schönberger Angela Paul Peter Seizer Konstantinos Stellos Andreas Schuster Rene M Botnar Rene M Botnar Meinrad Gawaz Boris Bigalke Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification. PLoS ONE |
| title | Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification. |
| title_full | Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification. |
| title_fullStr | Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification. |
| title_full_unstemmed | Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification. |
| title_short | Green fluorescent protein (GFP) color reporter gene visualizes parvovirus B19 non-structural segment 1 (NS1) transfected endothelial modification. |
| title_sort | green fluorescent protein gfp color reporter gene visualizes parvovirus b19 non structural segment 1 ns1 transfected endothelial modification |
| url | http://europepmc.org/articles/PMC3305306?pdf=render |
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