239 A NOVEL PIG MODEL OF CYSTIC FIBROSIS GENERATED BY SEQUENTIAL TARGETING OF CFTR BY BACTERIAL ARTIFICIAL CHROMSOME VECTORS

Abstract

Cystic fibrosis (CF), the most common inherited disease in the Caucasian population, is caused by mutations of the CFTR gene, coding for an anion channel responsible for chloride movement, determining transepithelial salt transport, fluid flow and ion concentrations. Cystic fibrosis is determined by chronic lung disease, marked by airway surface dehydration, reduced mucus clearance and increased mucus obstruction, impaired bacterial killing and inflammation and continuous destruction of the lung tissue. The lack of suitable CF mouse models has so far impeded the understanding of disease mechanisms and the development of novel therapies. Recently, a pig model reflecting the main hallmarks of the human disease in the respiratory, gastrointestinal and genital tracts has been reported (Rogers et al. 2008 Science 321, 1837–1841). In the present study, we generated a new CF pig model by disrupting the CFTR gene using an approach that differs in 3 major aspects from the previously published model: (1) gene targeting was achieved using a modified bacterial artificial chromosome (BAC) vector; (2) transcription of the CFTR gene was completely prevented; and (3) homozygous CFTR mutant pigs were derived by nuclear transfer from cells in which both alleles were inactivated by sequential BAC targeting. We characterised 2 cloned litters with a total of 13 piglets (including 4 stillborns). All piglets showed a characteristic CF multi-organ disease with severe meconium ileus, microcolon, microgallbladder, hypoplasia of the exocrine pancreas, congenital aplasia of the vas deferens and malformations of the trachea. In contrast to the recently published CF pig, where the majority of the intestinal obstructions occurred proximal to the ileocecal junction, we found obstructions exclusively distal to this site. We thus present a novel pig model for CF that reflects the main hallmarks of the human disease. It confirms the suitability of the CFTR mutant pig for cystic fibrosis research and increases the availability of CF models for molecular and translational CF research. The help by Prof. K. Heinritzi and his team from the Pig Clinic, LMU Munich, is gratefully acknowledged. This study was supported by Mukoviszidose e.V.