Skin graft survival in genetically identical cloned pigs.

Nuclear transfer technology allows for the reprogramming of somatic cells, and the production of embryonic stem cells and animals that are genetically identical in terms of nuclear DNA to the parental somatic cell. It is assumed that these products of nuclear transfer technology will be immunologically compatible to each other in spite of the fact that there are data that show differences in the expression patterns and phenotypes between animals produced by nuclear transfer. We have produced a series of cloned pigs from embryonic fibroblasts. Microsatellite analysis was used to confirm that the clones were genetically identical. Skin transplants were performed to assess immunological reactivity. Skin transplants between genetically identical cloned pigs were accepted, whereas third party grafts were rejected. Histological analysis of the grafts showed edema and mononuclear cell infiltrates in the recipient's skin in rejected grafts and not in grafts that were accepted. Our data supports the notion that genetically identical cloned pigs are immunologically compatible.

Pig cells that lack the gene for alpha1-3 galactosyltransferase express low levels of the gal antigen.

BACKGROUND: The major antigen recognized on pig tissue by primate antibodies is a terminal galalpha1-3gal carbohydrate structure (gal antigen) present on glycolipids and glycoproteins. The production of animals from somatic cells allows for the inactivation of specific genes. It is anticipated that the complete inactivation of the gene encoding alpha1-3 galactosyltransferase, the enzyme that synthesizes the galalpha1-3gal linkage, will result in loss of that antigen from pig organs and tissue and will provide a survival benefit in pig-to-primate xenotransplants. METHODS: Positive-negative selection was used to produce fetal-pig fibroblasts that were a heterozygous knockout (+/-) of the alpha1-3 galactosyltransferase gene. Nuclear transfer of these cells generated pig embryos and live born pigs with the appropriate genotype. Using a novel selection method with cells from (+/-) embryos, we produced homozygous (-/-) fetal-pig fibroblast cells. RESULTS: Southern blot analysis of the alpha1-3 galactosyltransferase gene showed that we had produced (+/-) pig embryos, (+/-) live born pigs, and (-/-) pig-fetal fibroblast cells. Fluorescence-activated cell sorter (FACS) analysis with some, but not all, mouse anti-gal monoclonal antibodies and sensitized human serum showed that (-/-) cells still synthesized the gal antigen at 1 to 2% of the level of control heterozygous cells. CONCLUSIONS: Fetal-pig fibroblasts homozygous for the knockout of the alpha1-3 galactosyltransferase gene appear to express low but detectable levels of the gal antigen.