(Re)producing transgenic pigs for xenotransplantation

Xenotransplantation is discussed as an alternative treatment for end-stage organ failure. The pig has been widely accepted as a feasible donor species. However, it has to be genetically modified in order to overcome incompatibilities of the human and the porcine immune systems. Although profound experience in long-term breeding of multi-transgenic animals exists for mouse models in biomedical research, the situation in breeding multi-transgenic pigs is far more complicated. In contrast to small animal models, several limitations have to be taken into account for the pig: (i) inbreeding has a detrimental effect on fertility and litter size in pigs while congenic mouse strains are being bred with small limitations; (ii) generation times of approximately 12 months and pregnancies of 115 days in pigs necessitate careful breeding management; (iii) costs and space requirements in pig maintenance far exeed those for mice. In addition, insight into xenograft rejection mechanisms is still limited and further transgenes or novel transgene combinations might be required in the future. Thus, effective breeding schedules have to be designed that accomodate these limitations and allow for adaptation to changing transgene requirements. This thesis describes the design and establishment of multi-transgenic GalKO/CD46/HLA-E and GalKO/CD46/hTM donor herds and the characterisation of novel hTM and LEA29Y transgenic lines for future incorporation into the breeding herd. In a first step, two fertile GalKO/CD46 boars and three fertile HLA-E sows were selected as founders for an initial GalKO/CD46/HLA-E breeding herd into which further xeno-relevant transgenes were to be incorporated later on. Breeding schedules that accommodate the conflicting issues of inbreeding, transgene segregation and time requirements to different extents were designed and the most feasible strategy was identified. Furthermore, options for incorporation of already fertile hTM transgenic founder pigs into the core breeding herd were analysed.