Figure 1.
Green Fluorescent Protein to Visualize Transfection of an Endothelial-like Cell Line (ECV304) by NS1-Parvovirus B19 (PVB19).
(A) Sketch of the parvovirus B19 (PVB19) plasmid with its three distinct proteins, non-structural protein segment 1 (NS1) and two structural viral capsid proteins VP1 and VP2, as well as the binding of green fluorescent protein (GFP) color reporter gene to NS1. (B) The transfection of endothelial cells (ECs) was verified simultaneously through flow cytometry, immunofluorescence microscopy and real-time polymerase chain reaction (RT-PCR) analysis after 24, 48, 72, and 96 h. The transfection rate (%) at the subsequent timepoints decreased in NS1-GFP transfected EC faster than in the GFP-control.
Figure 2.
Green Fluorescent Protein NS1 Transfection Induces Endothelial Activation and Platelet Adhesion.
(A) Non-structural protein segment 1 (NS1) transfection of endothelial cells visualized by green fluorescent protein (GFP) causes endothelial activation and enhanced expression of ICAM-1/CD54 and (B) induces endothelial expression of EMMPRIN/CD147 compared to control-GFP transfected cells. (C) Dynamic adhesion assays (flow chamber) showed that adhesion of platelets was enhanced twice as much on NS1 transfected ECs compared to control-GFP. (D) Similar results were found with the number of adherent monocytes. (E) Distinctive images of ECs representative for NS1-GFP, GFP-control, TNF-α/IFN-γ stimulated cells and resting cells 24 h after transfection. *denotes stastical significance (P<0.05). Whiskers represent standard deviation.