RESUMEN
UNLABELLED: Current techniques for the alteration of gene expression in the liver have a number of limitations, including the lack of stable somatic gene transfer and the technical challenges of germline transgenesis. Rapid and stable genetic engineering of the liver would allow systematic, in vivo testing of contributions by many genes to disease. After fumaryl acetoacetate hydrolase (Fah) gene transfer to hepatocytes, selective repopulation of the liver occurs in FAH-deficient mice. This genetic correction is readily mediated with transposons. Using this approach, we show that genes with biological utility can be linked to a selectable Fah transposon cassette. First, net conversion of Fah(-/-) liver tissue to transgenic tissue, and its outgrowth, was monitored by bioluminescence in vivo from a luciferase gene linked to the FAH gene. Second, coexpressed short hairpin RNAs (shRNAs) stably reduced target gene expression, indicating the potential for loss-of-function assays. Third, a mutant allele of human alpha1-antitrypsin (hAAT) was linked to Fah and resulted in protein inclusions within hepatocytes, which are the histopathological hallmark of hAAT deficiency disorder. Finally, oncogenes linked to Fah resulted in transformation of transduced hepatocytes. CONCLUSION: Coexpression with FAH is an effective technique for lifelong expression of transgenes in adult hepatocytes with applicability to a wide variety of genetic studies in the liver.
Asunto(s)
Regulación de la Expresión Génica , Hígado/fisiología , Sustitución de Aminoácidos , Animales , Línea Celular Tumoral , ADN Complementario/genética , Femenino , Genes Reporteros , Humanos , Hidrolasas/deficiencia , Hidrolasas/genética , Neoplasias Intestinales , Hígado/citología , Hígado/patología , Neoplasias Hepáticas/genética , Luciferasas/genética , Ratones , Análisis de Secuencia por Matrices de Oligonucleótidos , Plásmidos , Neoplasias del Cuello Uterino , alfa 1-Antitripsina/genéticaRESUMEN
Advances in drug therapy have made it possible to cure HCV infection. This article describes how best to screen, diagnose, and counsel these patients.