A population analysis of VEGF transgene expression and secretion.
The induction of therapeutic angiogenesis with gene therapy approaches has received considerable interest and some limited clinical success. A major drawback to this approach is a lack of understanding of the pharmacokinetics of therapeutic protein delivery. This has become increasingly more relevant as recent studies have illustrated a defined therapeutic window for angiogenic protein secretion into the local microenvironment. For cell based gene therapies, with cells widely distributed throughout the tissue, this implies that any individual cell must attain a specific secretion rate to produce a local angiogenic response. Here we report a reproducible technique enabling the study of growth factor secretion from individual cells following transient plasmid transfection. We demonstrate significant variability in single cell vascular endothelial growth factor (VEGF) secretion with the majority of total protein secretion arising from a small subpopulation of transfected cells. We demonstrate that VEGF secretion is linearly correlated to intracellular plasmid copy number and protein secretion does not appear to reach saturation within the cell population. The selection of gene therapy approaches that optimize individual cell secretion profiles may be essential for the development of effective gene therapies.
Karoubi
Golnaz
G
Division of General Thoracic Surgery, University Hospital Berne, 35 Murtenstrasse, Berne CH3010, Switzerland. golnaz.karoubi@insel.ch
Stewart
Duncan J
DJ
Courtman
David W
DW
D000818:Animals; D002478:Cells, Cultured; D046508:Culture Techniques; D018274:Electroporation; D005347:Fibroblasts; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods; D010957:Plasmids / Q000008:administration & dosage; D051381:Rats; D014162:Transfection / Q000379:methods*; D019076:Transgenes / Q000502:physiology; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*
D042461:Vascular Endothelial Growth Factor A