Hyperacute rejection of porcine xenografts in a non-human primate model has been diminished with development of Α1,3-galactosyltransferase knockout (GT-KO) pigs devoid of the galactose Α-1,3-galactose (Gal) antigen. However, xenografts are still susceptible to antibody-mediated rejection caused by antibodies to non-Gal xenoantigens such as N-glycolylneuraminic acid (Neu5Gc). Neu5Gc, or Hanganutziu-Deicher (HD) antigen, is a carbohydrate, which commonly terminates oligosaccharides on glycoproteins in all known mammals except humans. Humans do not express Neu5Gc due to a mutation in the gene for cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH), which hydroxylates N-acetylneuraminic acid to Neu5Gc. This study compares the immunoreactivity of human serum antibody to CMAH/GT-KO and GT-KO peripheral blood mononuclear cells (PBMCs). CMAH and GT genes were disrupted using zinc finger nucleases in porcine liver derived cells. CMAH/GT-KO pigs were produced using somatic cell nuclear transfer. Flow cytometric analyses and confocal microscopy of porcine cells and tissues confirmed the absence of Gal and Neu5Gc. The immunoreactivity of human serum antibody to these cells was evaluated by flow cytometric crossmatch analysis and antibody-mediated complement-dependent cytotoxicity using sera from ten randomly selected healthy human volunteers. The medians of the mean fluorescence intensity (MFI) values for human IgM binding to CMAH/GT-KO and GT-KO cells were 4,411 and 15,059, respectively (n = 10, p = 0.0039, 25% serum). The medians of the MFI values for human IgG binding to CMAH/GT-KO and GT-KO cells were 714 and 2,306, respectively (n = 10, p = 0.002, 25% serum). Further, human serum was less cytotoxic to CMAH/GT-KO than to GT-KO PBMCs. The median percent cytotoxicity of CMAH/GT-KO and GT-KO PBMCs using 50% human serum was 71.63% and 94.45%, respectively (n = 10, p = 0.002). In conclusion, CMAH/GT-KO pig PBMCs show diminished immunoreactivity compared to GT-KO PBMCs. Future studies will determine if CMAH/GT-KO porcine xenografts exhibit improved graft survival.
To cite this abstract in AMA style:Lutz A, Sidner R, Li P, Estrada J, Paris L, Chihara R, Downey S, Tector A. Pigs Deficient in N-Glycolylneuraminic Acid and Galactose α-1,3-Galactose Reduce the Humoral Barrier to Xenotransplantation [abstract]. Am J Transplant. 2013; 13 (suppl 5). https://atcmeetingabstracts.com/abstract/pigs-deficient-in-n-glycolylneuraminic-acid-and-galactose-13-galactose-reduce-the-humoral-barrier-to-xenotransplantation/. Accessed June 12, 2021.
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