Expression of biologically active human TRAIL in transgenic pigs.
BACKGROUND
Xenotransplantation of porcine organs into human recipients is a potential option for overcoming the dramatic shortage of suitable donor organs. To date, transgenic modification of pig organs has achieved partial or temporal reduction of xenograft rejection by inhibition of hyperacute rejection. Expression of human tumor necrosis factor-alpha-related apoptosis-inducing ligand (TRAIL) in transgenic pigs might be a strategy for controlling posthyperacute rejection mechanisms mediated by cellular components of the immune system. The objective of this study was generation of a transgenic pig model to evaluate the potential of this strategy for xenotransplantation.


METHODS
Transgenic pigs were generated by microinjection of an expression vector for human TRAIL under control of the murine H-2K promoter. Expression of the transgene was analyzed by Western blot and immunohistochemistry. Biologic activity of TRAIL on transgenic porcine lymphocytes was evaluated in co-culture experiments using Jurkat and Hut 78.2 cells as targets.


RESULTS
In three lines of transgenic pigs, human TRAIL protein was detected in the membrane fractions of various tissues. Highest expression levels were observed in spleen and lung. Human TRAIL expression on porcine lymphocytes was augmented on activation of cells. Transgenic pig lymphoblasts induced apoptosis in Jurkat and Hut 78.2 cells, which was inhibited by neutralizing anti-TRAIL antibodies, demonstrating a TRAIL-specific effect.


CONCLUSIONS
Ubiquitous expression of human TRAIL was achieved in transgenic pigs without detrimental side effects. Pigs expressing biologically active human TRAIL will be used for future xenotransplantation experiments to modulate primate anti-pig cellular immune responses.
Klose
Regina
R
Lehrstuhl für Molekulare Tierzucht und Biotechnologie, Ludwig-Maximilians-Universität, München, Germany.
Kemter
Elisabeth
E
Bedke
Tanja
T
Bittmann
Iris
I
Kelsser
Barbara
B
Endres
Robert
R
Pfeffer
Klaus
K
Schwinzer
Reinhard
R
Wolf
Eckhard
E
D000818:Animals; D030801:Animals, Genetically Modified; D015703:Antigens, CD / Q000032:analysis / Q000235:genetics; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D015139:Blotting, Southern; D002460:Cell Line; D006801:Humans; D007150:Immunohistochemistry; D008214:Lymphocytes / Q000166:cytology / Q000276:immunology / Q000502:physiology; D008545:Melanoma / Q000235:genetics; D008562:Membrane Glycoproteins / Q000032:analysis / Q000235:genetics*; D013552:Swine; D053221:TNF-Related Apoptosis-Inducing Ligand; D014183:Transplantation, Heterologous / Q000276:immunology*; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis / Q000235:genetics*; D019014:fas Receptor / Q000235:genetics
D015703:Antigens, CD; D051017:Apoptosis Regulatory Proteins; D008562:Membrane Glycoproteins; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor