title: Gene transfection into fetal sheep airways in utero using guanidinium-cholesterol cationic lipids.
abstract: BACKGROUND
  Over the last several years, we have developed a novel class of cationic lipids, cholesterol derivatives characterized by polar head groups with guanidinium functions. We have in particular shown that bis(guanidinium)-tren-cholesterol/dioleoylphosphatidylethanolamine (BGTC/DOPE) cationic liposomes can mediate efficient gene transfection into the mouse airways in vivo via direct intratracheal administration or intranasal instillation. As prenatal gene therapy may be necessary for the treatment of a variety of congenital lung diseases, we have explored in the present work the feasibility of BGTC-mediated gene transfection into the respiratory tract of fetal sheep in utero.
  
  
  METHODS
  Thus, BGTC/DOPE liposomes were complexed with plasmids expressing the Escherichia coli chloramphenicol acetyltransferase (CAT) reporter gene and the resulting lipoplexes were administered to fetal sheep at 70 days of gestation via surgical replacement of the airway fluid by the transfection mixture followed by tracheal occlusion. The fetal lungs and tracheas were harvested at 72 h and examined for CAT expression and evidence of toxicity.
  
  
  RESULTS
  CAT expression was detected in both lung and trachea homogenates, no CAT expression being observed in control fetuses receiving naked plasmid DNA. Immunohistochemical analysis showed that airway epithelial cells and some mesenchymal cells were transfected. Pulmonary histopathology of varied severity was however observed under our transfection conditions and manifested as focal epithelial and mesenchymal lesions.
  
  
  CONCLUSIONS
  These results show that BGTC/DOPE liposomes can mediate gene transfection into the fetal sheep airway epithelium. They also invite the development of optimized BGTC-based formulations and administration conditions with a view to future prenatal gene transfer experiments involving therapeutic genes.
authors.affiliation: Maternité, Hôpital Robert Debré, AP-HP, 48 Boulevard Sérurier, 75019 Paris, France. plehn@infobiogen.fr
authors.forename: Dominique
authors.initials: D
authors.lastname: Luton
authors.forename: Noufissa
authors.initials: N
authors.lastname: Oudrhiri
authors.forename: Pascal
authors.initials: P
authors.lastname: de Lagausie
authors.forename: Abderrahim
authors.initials: A
authors.lastname: Aissaoui
authors.forename: Michelle
authors.initials: M
authors.lastname: Hauchecorne
authors.forename: Sébastien
authors.initials: S
authors.lastname: Julia
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Oury
authors.forename: Yves
authors.initials: Y
authors.lastname: Aigrain
authors.forename: Michel
authors.initials: M
authors.lastname: Peuchmaur
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Vigneron
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Lehn
authors.forename: Pierre
authors.initials: P
authors.lastname: Lehn
mesh_terms: D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000032:analysis / Q000235:genetics; D002784:Cholesterol / Q000031:analogs & derivatives*; D005260:Female; D005333:Fetus / Q000378:metabolism*; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors; D006146:Guanidines; D007120:Immunochemistry; D008081:Liposomes / Q000737:chemistry; D008168:Lung / Q000196:embryology / Q000473:pathology; D010714:Phosphatidylethanolamines / Q000737:chemistry; D010957:Plasmids / Q000008:administration & dosage; D011247:Pregnancy; D020545:Respiratory Mucosa / Q000196:embryology / Q000378:metabolism*; D012756:Sheep / Q000196:embryology; D014132:Trachea / Q000473:pathology; D014162:Transfection / Q000379:methods*
chemical_list: C101389:3-(N(4)-(N(1),N(8)-diguanidinospermidine)carbamoyl)cholesterol; D006146:Guanidines; D008081:Liposomes; D010714:Phosphatidylethanolamines; C094877:dioleoyl phosphatidylethanolamine; D002784:Cholesterol; D015500:Chloramphenicol O-Acetyltransferase