Generation of a-1,3-galactosyltransferase knocked-out transgenic cloned pigs with knocked-in five human genes

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Title
Generation of a-1,3-galactosyltransferase knocked-out transgenic cloned pigs with knocked-in five human genes
Author(s)
D J Kwon; Dong-Hwan Kim; I S Hwang; Dong-Ern Kim; Hyung-Joo Kim; Jang Seong Kim; K Lee; G S Im; Jeong Woong Lee; S Hwang
Bibliographic Citation
Transgenic Research, vol. 26, pp. 153-163
Publication Year
2017
Abstract
Recent progress in genetic manipulation of pigs designated for xenotransplantation ha6s shown considerable promise on xenograft survival in primates. However, genetic modification of multiple genes in donor pigs by knock-out and knock-in technologies, aiming to enhance immunological tolerance against transplanted organs in the recipients, has not been evaluated for health issues of donor pigs. We produced transgenic Massachusetts General Hospital piglets by knocking-out the α-1,3-galactosyltransferase (GT) gene and by simultaneously knocking-in an expression cassette containing five different human genes including, DAF, CD39, TFPI, C1 inhibitor (C1-INH), and TNFAIP3 (A20) [GT?(DAF/CD39/TFPI/C1-INH/TNFAIP3)/+] that are connected by 2A peptide cleavage sequences to release individual proteins from a single translational product. All five individual protein products were successfully produced as determined by western blotting of umbilical cords from the newborn transgenic pigs. Although gross observation and histological examination revealed no significant pathological abnormality in transgenic piglets, hematological examination found that the transgenic piglets had abnormally low numbers of platelets and WBCs, including neutrophils, eosinophils, basophils, and lymphocytes. However, transgenic piglets had similar numbers of RBC and values of parameters related to RBC compared to the control littermate piglets. These data suggest that transgenic expression of those human genes in pigs impaired hematopoiesis except for erythropoiesis. In conclusion, our data suggest that transgenic expression of up to five different genes can be efficiently achieved and provide the basis for determining optimal dosages of transgene expression and combinations of the transgenes to warrant production of transgenic donor pigs without health issues
Keyword
Immune rejectionLeucopeniaMulti-transgenic pigsThrombocytopeniaXenotransplantation
ISSN
0962-8819
Publisher
Springer
DOI
http://dx.doi.org/10.1007/s11248-016-9979-8
Type
Article
Appears in Collections:
Division of Biomedical Research > Biotherapeutics Translational Research Center > 1. Journal Articles
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