• ISSN 1674-8301
  • CN 32-1810/R
Volume 33 Issue 4
Jul.  2019
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Article Contents
Wang Ronggen, Ruan Miaomiao, Zhang Runjie, Chen Lei, Li Xiaoxue, Fang Bin, Li Chu, Ren Xueyang, Liu Jiying, Xiong Qiang, Zhang Lining, Jin Yong, Li Lin, Li Rongfeng, Wang Ying, Yang Haiyuan, Dai Yifan. Antigenicity of tissues and organs from GGTA1/CMAH/β4GalNT2 triple gene knockout pigs[J]. The Journal of Biomedical Research, 2019, 33(4): 235-243. doi: 10.7555/JBR.32.20180018
Citation: Wang Ronggen, Ruan Miaomiao, Zhang Runjie, Chen Lei, Li Xiaoxue, Fang Bin, Li Chu, Ren Xueyang, Liu Jiying, Xiong Qiang, Zhang Lining, Jin Yong, Li Lin, Li Rongfeng, Wang Ying, Yang Haiyuan, Dai Yifan. Antigenicity of tissues and organs from GGTA1/CMAH/β4GalNT2 triple gene knockout pigs[J]. The Journal of Biomedical Research, 2019, 33(4): 235-243. doi: 10.7555/JBR.32.20180018

Antigenicity of tissues and organs from GGTA1/CMAH/β4GalNT2 triple gene knockout pigs

doi: 10.7555/JBR.32.20180018
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  • Corresponding author: Haiyuan Yang and Yifan Dai, Jiangsu Key Laboratory of Xenotransplantation, Nanjing Medical University, 101 Longmian Avenue, Nanjing, Jiangsu 211166, China. E-mails: hyyang@njmu.edu.cn and daiyifan@njmu.edu.cn
  • Received: 2018-02-12
  • Revised: 2018-03-30
  • Accepted: 2018-04-23
  • Published: 2018-05-31
  • Issue Date: 2019-07-01
  • Clinical xenotransplantations have been hampered by human preformed antibody-mediated damage of the xenografts. To overcome biological incompatibility between pigs and humans, one strategy is to remove the major antigens [Gal, Neu5Gc, and Sd(a)] present on pig cells and tissues. Triple gene (GGTA1, CMAH, and β4GalNT2) knockout (TKO) pigs were produced in our laboratory by CRISPR-Cas9 targeting. To investigate the antigenicity reduction in the TKO pigs, the expression levels of these three xenoantigens in the cornea, heart, liver, spleen, lung, kidney, and pancreas tissues were examined. The level of human IgG/IgM binding to those tissues was also investigated, with wildtype pig tissues as control. The results showed that αGal, Neu5Gc, and Sd(a) were markedly positive in all the examined tissues in wildtype pigs but barely detected in TKO pigs. Compared to wildtype pigs, the liver, spleen, and pancreas of TKO pigs showed comparable levels of human IgG and IgM binding, whereas corneas, heart, lung, and kidney of TKO pigs exhibited significantly reduced human IgG and IgM binding. These results indicate that the antigenicity of TKO pig is significantly reduced and the remaining xenoantigens on porcine tissues can be eliminated via a gene targeting approach.


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  • [1]
    McGregor CGA, Byrne GW. Porcine to human heart transplantation: is clinical application now appropriate?[J]. J Immunol Res, 2017, 2017: 2534653.
    Patel MS, Louras N, Vagefi PA. Liver xenotransplantation[J]. Curr Opin Organ Transplant, 2017, 22(6): 535–540. doi: 10.1097/MOT.0000000000000459
    Shah JA, Patel MS, Elias N, et al. Prolonged survival following pig-to-primate liver xenotransplantation utilizing exogenous coagulation factors and costimulation blockade[J]. Am J Transplant, 2017, 17(8): 2178–2185. doi: 10.1111/ajt.2017.17.issue-8
    Iwase H, Kobayashi T. Current status of pig kidney xenotransplantation[J]. Int J Surg, 2015, 23(Pt B): 229–233.
    Fukui A, Yokoo T. Kidney regeneration using developing xenoembryo[J]. Curr Opin Organ Transplant, 2015, 20(2): 160–164. doi: 10.1097/MOT.0000000000000176
    Wilczek P, Lesiak A, Niemiec-Cyganek A, et al. Biomechanical properties of hybrid heart valve prosthesis utilizing the pigs that do not express the galactose-α-1,3-galactose (α-Gal) antigen derived tissue and tissue engineering technique[J]. J Mater Sci Mater Med, 2015, 26(1): 5329.
    Laird C, Burdorf L, Pierson RN 3rd. Lung xenotransplantation: a review[J]. Curr Opin Organ Transplant, 2016, 21(3): 272–278. doi: 10.1097/MOT.0000000000000311
    Sahara H, Watanabe H, Pomposelli T, et al. Lung xenotransplantation[J]. Curr Opin Organ Transplant, 2017, 22(6): 541–548. doi: 10.1097/MOT.0000000000000465
    Abalovich A, Matsumoto S, Wechsler CJ, et al. Level of acceptance of islet cell and kidney xenotransplants by personnel of hospitals with and without experience in clinical xenotransplantation[J]. Xenotransplantation, 2017, 24(4): 1–5.
    Cooper DKC, Matsumoto S, Abalovich A, et al. Progress in clinical encapsulated islet xenotransplantation[J]. Transplantation, 2016, 100(11): 2301–2308. doi: 10.1097/TP.0000000000001371
    Cooper DKC, Ekser B, Ramsoondar J, et al. The role of genetically engineered pigs in xenotransplantation research[J]. J Pathol, 2016, 238(2): 288–299. doi: 10.1002/path.4635
    Dong X, Hara H, Wang Y, et al. Initial study of 1,3- galactosyltransferase gene-knockout/CD46 pig full-thickness corneal xenografts in rhesus monkeys[J]. Xenotransplantation, 2017, 24(1): 1–12.
    Ekser B, Li P, Cooper DKC. Xenotransplantation: past, present, and future[J]. Curr Opin Organ Transplant, 2017, 22(6): 513–521.
    Cooper DKC, Pierson RN 3rd, Hering BJ, et al. Regulation of clinical xenotransplantation-time for a reappraisal[J]. Transplantation, 2017, 101(8): 1766–1769. doi: 10.1097/TP.0000000000001683
    Higginbotham L, Mathews D, Breeden CA, et al. Pre-transplant antibody screening and anti-CD154 costimulation blockade promote long-term xenograft survival in a pig-to-primate kidney transplant model[J]. Xenotransplantation, 2015, 22(3): 221–230. doi: 10.1111/xen.12166
    Cooper DKC, Ezzelarab MB, Hara H. Low anti-pig antibody levels are key to the success of solid organ xenotransplantation: But is this sufficient?[J]. Xenotransplantation, 2017, 24(6): 1–6.
    Lai L, Kolber-Simonds D, Park KW, et al. Production of alpha- 1,3-galactosyltransferase knockout pigs by nuclear transfer cloning[J]. Science, 2002, 295(5557): 1089–1092. doi: 10.1126/science.1068228
    Dai Y, Vaught TD, Boone J, et al. Targeted disruption of the alpha1, 3-galactosyltransferase gene in cloned pigs[J]. Nat Biotechnol, 2002, 20(3): 251–255. doi: 10.1038/nbt0302-251
    McGregor CGA, Ricci D, Miyagi N, et al. Human CD55 expression blocks hyperacute rejection and restricts complement activation in Gal knockout cardiac xenografts[J]. Transplantation, 2012, 93(7): 686–692. doi: 10.1097/TP.0b013e3182472850
    McGregor C, Byrne G, Rahmani B, et al. Physical equivalency of wild type and galactose α 1,3 galactose free porcine pericardium; a new source material for bioprosthetic heart valves[J]. Acta Biomater, 2016, 41: 204–209. doi: 10.1016/j.actbio.2016.06.007
    Mohiuddin MM, Corcoran PC, Singh AK, et al. B-cell depletion extends the survival of GTKO.hCD46Tg pig heart xenografts in baboons for up to 8 months[J]. Am J Transplant, 2012, 12(3): 763–771. doi: 10.1111/j.1600-6143.2011.03846.x
    Chen G, Qian H, Starzl T, et al. Acute rejection is associated with antibodies to non-Gal antigens in baboons using Gal-knockout pig kidneys[J]. Nat Med, 2005, 11(12): 1295–1298. doi: 10.1038/nm1330
    Byrne GW, Du Z, Stalboerger P, et al. Cloning and expression of porcine 1,4 N-acetylgalactosaminyl transferase encoding a new xenoreactive antigen[J]. Xenotransplantation, 2014, 21(6): 543–554. doi: 10.1111/xen.2014.21.issue-6
    Lee W, Hara H, Ezzelarab MB, et al. Initial in vitro studies on tissues and cells from GTKO/CD46/NeuGcKO pigs[J]. Xenotransplantation, 2016, 23(2): 137–150. doi: 10.1111/xen.2016.23.issue-2
    Burlak C, Bern M, Brito AE, et al. N-linked glycan profiling of GGTA1/CMAH knockout pigs identifies new potential carbohydrate xenoantigens[J]. Xenotransplantation, 2013, 20(5): 277–291. doi: 10.1111/xen.2013.20.issue-5
    Petersen B, Frenzel A, Niemann H. Efficient generation of a triple knockout (GGTA1/CMAH/ASGR1) of xenorelevant genes in pig fibroblasts[J]. Xenotransplantation, 2015, 22(Suppl S1): S40.
    Estrada JL, Martens G, Li P, et al. Evaluation of human and non-human primate antibody binding to pig cells lacking GGTA1/CMAH/4GalNT2 genes[J]. Xenotransplantation, 2015, 22(3): 194–202. doi: 10.1111/xen.2015.22.issue-3
    Zhang RJ, Wang Y, Chen L, et al. Reducing immunoreactivity of porcine bioprosthetic heart valves by genetically-deleting three major glycan antigens, GGTA1/4GalNT2/CMAH[J]. Acta Biomater, 2018, 72: 196–205. doi: 10.1016/j.actbio.2018.03.055
    Wang ZY, Martens GR, Blankenship RL, et al. Eliminating xenoantigen expression on swine RBC[J]. Transplantation, 2017, 101(3): 517–523. doi: 10.1097/TP.0000000000001302
    Amano S, Shimomura N, Kaji Y, et al. Antigenicity of porcine cornea as xenograft[J]. Curr Eye Res, 2003, 26(6): 313–318. doi: 10.1076/ceyr.26.5.313.15440
    Hara H, Koike N, Long C, et al. Initial in vitro investigation of the human immune response to corneal cells from genetically engineered pigs[J]. Invest Ophthalmol Vis Sci, 2011, 52(8): 5278–5286. doi: 10.1167/iovs.10-6947
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