These authors contributed equally to this study.
Article first published online: 9 AUG 2011
Copyright © 2011 American Association for the Study of Liver Diseases
Volume 54, Issue 4, pages 1351–1359, October 2011
How to Cite
Hickey, R. D., Lillegard, J. B., Fisher, J. E., McKenzie, T. J., Hofherr, S. E., Finegold, M. J., Nyberg, S. L. and Grompe, M. (2011), Efficient production of Fah-null heterozygote pigs by chimeric adeno-associated virus-mediated gene knockout and somatic cell nuclear transfer. Hepatology, 54: 1351–1359. doi: 10.1002/hep.24490
Potential conflict of interest: Dr. Grompe owns stock in and is a consultant for Yecuris Corporation.
Supported by grants from the National Institute of Diabetes and Digestive and Kidney Diseases, grant numbers R01DK048252-17 and R01DK051592-13 (to M.G.); American Society of Transplant Surgeons-NKF Folkert Belzer, MD Research Award (to J.B.L.).
- Issue published online: 27 SEP 2011
- Article first published online: 9 AUG 2011
- Accepted manuscript online: 14 JUN 2011 08:06AM EST
- Manuscript Accepted: 25 MAY 2011
- Manuscript Received: 11 NOV 2010
Hereditary tyrosinemia type I (HT1) results in hepatic failure, cirrhosis, and hepatocellular carcinoma (HCC) early in childhood and is caused by a deficiency in the enzyme fumarylacetoacetate hydrolase (FAH). In a novel approach we used the chimeric adeno-associated virus DJ serotype (AAV-DJ) and homologous recombination to target and disrupt the porcine Fah gene. AAV-DJ is an artificial chimeric AAV vector containing hybrid capsid sequences from three naturally occurring serotypes (AAV2, 8, and 9). The AAV-DJ vector was used to deliver the knockout construct to fetal pig fibroblasts with an average knockout targeting frequency of 5.4%. Targeted Fah-null heterozygote fibroblasts were used as nuclear donors for somatic cell nuclear transfer (SCNT) to porcine oocytes and multiple viable Fah-null heterozygote pigs were generated. Fah-null heterozygotes were phenotypically normal, but had decreased Fah transcriptional and enzymatic activity compared to wildtype animals. Conclusion: This study is the first to use a recombinant chimeric AAV vector to knockout a gene in porcine fibroblasts for the purpose of SCNT. In using the AAV-DJ vector we observed targeting frequencies that were higher than previously reported with other naturally occurring serotypes. We expect that the subsequent generation of FAH-null homozygote pigs will serve as a significant advancement for translational research in the areas of metabolic liver disease, cirrhosis, and HCC. (HEPATOLOGY 2011;)