Amphotropic and ecotropic retroviral vector viruses transduce midgestational murine fetal liver cells in a dual-chambered cocultivation system.
A beta-glucuronidase cDNA was transferred into fetal liver cells (FLC) from mice affected with mucopolysaccharidosis (MPS) type VII and normal littermates using a retrovirus vector. The cells were transduced by direct cocultivation or by culturing the FLC and vector packaging cells separated by a 0.45 micron filter in a dual-chambered cocultivation system. Gene transduction occurred using an ecotropic or amphotropic vector in FLC obtained from 13.5- and 15.5-day-old murine fetuses. Histochemical staining assays and measurement of enzyme activity demonstrated gene expression in the midgestational FLC. Enzyme secreted into the supernatant from transduced FLC obtained from 13.5-day-old affected fetuses surpassed secreted enzyme from normal, age-matched untransduced FLC. The 13.5 day FLC, transduced by direct cocultivation or by using the dual-chambered system, were transplanted in utero into 13.5-day-old murine fetuses. Proviral sequences were detected in various organs shortly after birth. The results indicate that midgestational FLC can be transduced with an amphotropic vector virus in a dual-chambered cocultivation system without contaminating virus-producing packaging cells and that the transduced cells survive in utero transplantation.
Casal
M L
ML
Department of Pathobiology, School of Veterinary Medicine, Univeristy of Pennsylvania, Philadelphia 19104, USA.
Wolfe
J H
JH
D000818:Animals; D017690:Cell Transplantation; D018920:Coculture Techniques; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D005966:Glucuronidase / Q000235:genetics*; D008099:Liver / Q000196:embryology* / Q000201:enzymology; D051379:Mice; D016538:Mucopolysaccharidosis VII / Q000201:enzymology / Q000628:therapy*; D012190:Retroviridae
D005966:Glucuronidase