Generation of genetic modified pigs devoid of GGTA1 and expressing the human leukocyte antigen-G5 / 生物工程学报
Chinese Journal of Biotechnology
; (12): 1096-1111, 2022.
Article
en Zh
| WPRIM
| ID: wpr-927766
Biblioteca responsable:
WPRO
ABSTRACT
Pigs are considered as ideal donors for xenotransplantation because they have many physiological and anatomical characteristics similar to human beings. However, antibody-mediated immunity, which includes both natural and induced antibody responses, is a major challenge for the success of pig-to-primate xenotransplantation. Various genetic modification methods help to tailor pigs to be appropriate donors for xenotransplantation. In this study, we applied transcription activator-like effector nuclease (TALEN) to knock out the porcine α-1, 3-galactosyltransferase gene GGTA1, which encodes Gal epitopes that induce hyperacute immune rejection in pig-to-human xenotransplantation. Meanwhile, human leukocyte antigen-G5 gene HLA-G5, which acts as an immunosuppressive factor, was co-transfected with TALEN into porcine fetal fibroblasts. The cell colonies of GGTA1 biallelic knockout with positive transgene for HLA-G5 were chosen as nuclear donors to generate genetic modified piglets through a single round of somatic cell nuclear transfer. As a result, we successfully obtained 20 modified piglets that were positive for GGTA1 knockout (GTKO) and half of them expressed the HLA-G5 protein. Gal epitopes on the cell membrane of GTKO/HLA-G5 piglets were completely absent. Western blotting and immunofluorescence showed that HLA-G5 was expressed in the modified piglets. Functionally, the fibroblasts from the GTKO/HLA-G5 piglets showed enhanced resistance to complement-mediated lysis ability compared with those from GTKO-only or wild-type pigs. These results indicate that the GTKO/HLA-G5 pigs could be a valuable donor model to facilitate laboratory studies and clinics for xenotransplantation.
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Texto completo:
1
Índice:
WPRIM
Asunto principal:
Porcinos
/
Trasplante Heterólogo
/
Animales Modificados Genéticamente
/
Técnicas de Transferencia Nuclear
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Técnicas de Inactivación de Genes
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Antígenos HLA
Tipo de estudio:
Prognostic_studies
Límite:
Animals
/
Humans
Idioma:
Zh
Revista:
Chinese Journal of Biotechnology
Año:
2022
Tipo del documento:
Article