Improved multi-transgenic pig lines for xenotransplantation

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This project involves the generation and characterization of multi-transgenic pigs with organs which resist hyperacute rejection (HAR), acute vascular rejection (AVR) and disseminated intravascular coagulation (DIC) following xeno-transplantation. In the first phase, existing transgenic hCD59 pigs will be mated with pigs expressing TNFa-related apoptosis inducing ligand (TRAIL) - provided by Prof. Wolf of this consortium - to produce fetuses from which double-transgenic nuclear donor cell lines can be cultured. The double-transgenic cell lines will be transfected with gene constructs based on the cytomegalovirus (CMV) promoter and the coding sequences for human heme-oxygenase 1 (hHO-1) and human thrombomodulin (hTM). After screening and clone selection, somatic nuclear transfer will be used to produce poly-transgenic offspring. Additional animals will be produced as required from these original donor cell clones to give homogeneous groups of experimental animals for other members of the consortium. Expression of transgenes will be measured at the mRNA and protein level by established protocols and biological activity will be assessed in vitro. Hearts, lungs and kidneys from the multi-transgenic pigs will be evaluated by other partners in the consortium using their specialized perfusion and transplantation protocols.