Abstract
Grafting of testicular tissue into immunodeficient mice makes it possible to obtain functional sperm from immature donor animals that cannot be used for reproduction. We have developed a porcine model of human haemophilia A (haemophilia-A pigs) by nuclear transfer cloning from foetal fibroblasts after disruption of the X-linked coagulation factor VIII (F8) gene. Despite having a recessive condition, female F8+/- cloned pigs died of severe bleeding at an early age, as was the case for male F8-/Y cloned pigs, thus making it impossible to obtain progeny. In this study, therefore, we produced sperm from F8-/Y cloned pigs by grafting their foetal testicular tissue into nude mice. Two F8+/- female pigs were generated from oocytes injected with xenogeneic sperm. Unlike the F8+/- cloned pigs, they remained asymptomatic, and delivered five F8-/Y and four F8+/- pigs after being crossed with wild-type boars. The descendant F8-/Y pigs conserved the haemophilia phenotype. Thus, the present F8+/- pigs show resolution of the phenotypic abnormality, and will facilitate production of F8-/Y pigs as founders of a strain of haemophilia-A pigs for the development of new therapeutics for haemophilia A. This strategy will be applicable to other genetically modified pigs.