title: Hepatocyte Transfection in Small Pigs After Weaning by Hydrodynamic Intraportal Injection of Naked DNA/Minicircle Vectors.
abstract: Liver is an attractive organ for gene delivery in order to correct various genetic (metabolic) diseases. Hydrodynamic vein injection of naked DNA/minicircles devoid of viral or plasmid backbones was demonstrated in, for example, murine phenylketonuria to allow sustained therapeutic transduction of hepatocytes. Here we show successful hepatocyte transfusion in domestic small pigs immediately after weaning upon portal vein catheterization and hydrodynamic injection of naked DNA/minicircle vectors expressing the luciferase gene from the CMV or a liver-specific promoter. First, we established a surgical method allowing hydrodynamic portal vein pressurization up to 120 mmHg and infusion of naked DNA in pigs (n = 5) with long-term survival. No acute adverse effects such as changes in liver transaminases or signs of liver cell damage were observed. We then showed efficiency of stable hepatocyte transfection at 10 and 28 days in single experiments (n = 7) where we found that up to 60% of samples (45/75) were polymerase chain reaction (PCR)-positive for minicircle-DNA. Of these samples, 13% of the positive specimen (6/45) showed low but stable luciferase expression when driven by a liver-specific promoter, as well as appropriate copy numbers per diploid genome. In conclusion, we accomplished a safe procedure for stable transfection of liver cells upon hydrodynamic gene delivery using minicircle vectors in small pigs as a prerequisite to potentially treat infants with genetic liver diseases.
authors.affiliation: 1 Division of Metabolism and Children's Research Centre, University Children's Hospital Zürich , Zürich, Switzerland .
authors.forename: Fabienne
authors.initials: F
authors.lastname: Stoller
authors.affiliation: 2 Swiss HBP and Transplant Center, Department of Surgery, University Hospital Zürich , Zürich, Switzerland .
authors.forename: Andrea
authors.initials: A
authors.lastname: Schlegel
authors.affiliation: 1 Division of Metabolism and Children's Research Centre, University Children's Hospital Zürich , Zürich, Switzerland .
authors.forename: Hiu Man
authors.initials: HM
authors.lastname: Viecelli
authors.affiliation: 1 Division of Metabolism and Children's Research Centre, University Children's Hospital Zürich , Zürich, Switzerland .
authors.forename: Véronique
authors.initials: V
authors.lastname: Rüfenacht
authors.affiliation: 3 Division of Surgical Research, University Hospital Zürich , Zürich, Switzerland .
authors.forename: Nikola
authors.initials: N
authors.lastname: Cesarovic
authors.affiliation: 1 Division of Metabolism and Children's Research Centre, University Children's Hospital Zürich , Zürich, Switzerland .
authors.forename: Claudio
authors.initials: C
authors.lastname: Viecelli
authors.affiliation: 1 Division of Metabolism and Children's Research Centre, University Children's Hospital Zürich , Zürich, Switzerland .
authors.forename: Sereina
authors.initials: S
authors.lastname: Deplazes
authors.affiliation: 4 Department of Anaesthesia, Vetsuisse-Faculty University of Zürich , Zürich, Switzerland .
authors.forename: Regula
authors.initials: R
authors.lastname: Bettschart
authors.affiliation: 5 Department of Small Animal Surgery, Vetsuisse-Faculty University of Zürich , Zürich, Switzerland .
authors.forename: Karin
authors.initials: K
authors.lastname: Hurter
authors.affiliation: 5 Department of Small Animal Surgery, Vetsuisse-Faculty University of Zürich , Zürich, Switzerland .
authors.forename: Philipp
authors.initials: P
authors.lastname: Schmierer
authors.affiliation: 6 Departement für Nutztiere, Abteilung für Schweinemedizin, Vetsuisse-Fakultät, University of Zürich , Switzerland .
authors.forename: Xaver
authors.initials: X
authors.lastname: Sidler
authors.affiliation: 2 Swiss HBP and Transplant Center, Department of Surgery, University Hospital Zürich , Zürich, Switzerland .
authors.forename: Philipp
authors.initials: P
authors.lastname: Kron
authors.affiliation: 2 Swiss HBP and Transplant Center, Department of Surgery, University Hospital Zürich , Zürich, Switzerland .
authors.forename: Philipp
authors.initials: P
authors.lastname: Dutkowski
authors.affiliation: 2 Swiss HBP and Transplant Center, Department of Surgery, University Hospital Zürich , Zürich, Switzerland .
authors.forename: Rolf
authors.initials: R
authors.lastname: Graf
authors.affiliation: 1 Division of Metabolism and Children's Research Centre, University Children's Hospital Zürich , Zürich, Switzerland .|7 Zürich Center for Integrative Human Physiology , Zürich, Switzerland .|8 Neuroscience Center Zürich , Zürich, Switzerland .
authors.forename: Beat
authors.initials: B
authors.lastname: Thöny
authors.affiliation: 1 Division of Metabolism and Children's Research Centre, University Children's Hospital Zürich , Zürich, Switzerland .|7 Zürich Center for Integrative Human Physiology , Zürich, Switzerland .|8 Neuroscience Center Zürich , Zürich, Switzerland .
authors.forename: Johannes
authors.initials: J
authors.lastname: Häberle
mesh_terms: D000818:Animals; D002404:Catheterization; D004247:DNA / Q000008:administration & dosage*; D005822:Genetic Vectors / Q000008:administration & dosage*; D057446:Hydrodynamics; D008107:Liver Diseases / Q000235:genetics* / Q000628:therapy*; D008659:Metabolic Diseases / Q000235:genetics* / Q000628:therapy*; D011169:Portal Vein / Q000378:metabolism; D013552:Swine; D019076:Transgenes
chemical_list: D004247:DNA