Immunogenicity Test of GGTA1, GGTA1/β4GalNT2 Different Genotypes Knockout Pigs (Sus scrofa)
LIU Chun-Chen1, DU Min-Jie2, XING Xiang-Yang2, LU Lin1, *, PAN Deng-Ke3, *
1 Animal Science and Technology College, Beijing University of Agriculture, Beijing 102206, China; 2 Chengdu Clonorgan Biotechnology Co., LTD., Chengdu 610041, China; 3 Institute of Organ Transplantation, Sichuan Academy of Medical Science & Sichuan Provincial People's Hospital, Chengdu 610072, China;
Abstract：Immune rejection is the main reason for limiting xenotransplantation. In this paper, α-1, 3-galactosyltransferase (GGTA1) and GGTA1/β1, 4 N-acetylgalactosaminyltransferase (β4GalNT2) gene knockout cloned pigs (Sus scrofa) were selected for donors of low-immunogenic xenotransplantation organ transplants. The expression of GGTA1/β4GalNT2 gene was identified by integrating peripheral blood mononuclear cells (PBMC) with fluorescein isothiocyanate-griffonia simplicifolia isolectin B4 (FITC-GSIB4) and fluorescein isothiocyanate Dolichos bliflorus agglutinin (FITC-DBA); the effect of GGTA1/β4GalNT2 double gene knockout on the health of Bama mini-pigs was assessed by blood changes in physiological and biochemical indicators; analysis of ideal receptor blood type was by detecting IgG binding of mixed PBMC of different genetically modified pigs and human serum of ABO blood group system after incubation; screening for low immunity primary donor genotype was bdye tecting IgG binding of mixed of PBMC of different genetically modified pigs and human (Homo sapiens) blood serum of AB blood group system after incubation. GSIB4 and DBA staining results showed that the expression of corresponding antigens on the surface of GGTA1/β-4GalNT2 gene knockout pig cells decreased; physiological and biochemical tests showed that GGTA1-/- and GGTA1-/-/β4GalNT2+/- were compared with wild type (WT) Bama mini-pigs, genetically modified pigs had no significant differences in all indicators, GGTA1-/-/β4GalNT2-/- gene modified pigs had significantly reduced neutrophil counts, and significantly increased creatine kinase and creatinine; the results of incubation of serum of different blood types showed that compared with the amount of IgG binding of type A, B and O blood, the binding of IgG in type AB was the lowest, and it was the ideal recipient blood type; GGTA1-/- and GGTA1/β-4GalNT2 knockout pigs, compared with Bama mini-pigs, had the lowest binding amount. The GGTA1/β-4GalNT2 knockout Bama mini-pigs were genetically stable, they also had reduced immunogenicity and immune rejection reactions with AB serum human serum were lowest. This study provides immunorejection data for future xenotransplantation clinical trials.
刘春晨, 杜敏杰, 邢向阳, 鲁琳, 潘登科. GGTA1、GGTA1/β4GalNT2不同基因型敲除猪免疫原性检测[J]. 农业生物技术学报, 2020, 28(9): 1587-1594.
LIU Chun-Chen, DU Min-Jie, XING Xiang-Yang, LU Lin, PAN Deng-Ke. Immunogenicity Test of GGTA1, GGTA1/β4GalNT2 Different Genotypes Knockout Pigs (Sus scrofa). 农业生物技术学报, 2020, 28(9): 1587-1594.
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