Hydrodynamics based transfection in normal and fibrotic rats.
OBJECTIVE
Hydrodynamics based transfection (HBT), the injection of a large volume of naked plasmid DNA in a short time is a relatively simple, efficient and safe method for in vivo transfection of liver cells. Though used for quite some time, the mechanism of gene transfection has not yet been elucidated.


METHODS
A luciferase encoding plasmid was injected using the hydrodynamics based procedure into normal and thioacetamide-induced fibrotic Sprague Dawley rats. Scanning and transmission electron microscopy images were taken. The consequence of a dual injection of Ringer solution and luciferase pDNA was followed. Halofuginone, an anti collagen type I inhibitor was used to reduce ECM load in fibrotic rats prior to the hydrodynamic injection.


RESULTS
Large endothelial gaps formed as soon as 10' following hydrodynamic injection; these gradually returned to normal 10 d post injection. Hydrodynamic administration of Ringer 10 or 30 m prior to moderate injection of plasmid did not result in efficient transfection suggesting that endothelial gaps by themselves are not sufficient for gene expression. Gene transfection following hydrodynamic injection in thioacetamide induced fibrotic rats was diminished coinciding with the level of fibrosis. Halofuginone, a specific collagen type I inhibitor, alleviated this effect.


CONCLUSIONS
The hydrodynamic pressure formed following HBT results in the formation of large endothelial gaps. These gaps, though important in the transfer of DNA molecules from the blood to the space of Disse are not enough to provide the appropriate conditions for hepatocyte transfection. Hydrodynamics based injection is applicable in fibrotic rats provided that ECM load is reduced.
Yeikilis
Rita
R
Department of Anatomy and Cell Biology, Faculty of Medicine, POB 9649, Haifa 31096, Israel.
Gal
Shunit
S
Kopeiko
Natalia
N
Paizi
Melia
M
Pines
Mark
M
Braet
Filip
F
Spira
Gadi
G
D000818:Animals; D004727:Endothelium / Q000473:pathology; D017930:Genes, Reporter; D007552:Isotonic Solutions; D008099:Liver / Q000378:metabolism* / Q000473:pathology / Q000648:ultrastructure; D008103:Liver Cirrhosis / Q000378:metabolism* / Q000473:pathology; D008156:Luciferases / Q000235:genetics; D010957:Plasmids; D051381:Rats / Q000235:genetics; D017207:Rats, Sprague-Dawley; D000077331:Ringer's Solution; D014162:Transfection / Q000379:methods*
D007552:Isotonic Solutions; D000077331:Ringer's Solution; D008156:Luciferases