Human HLA-Ev (147) Expression in Transgenic Animals

R. Matsuura, A. Maeda, R. Sakai, H. Eguchi, P. C. Lo, Hidetoshi Hasuwa, M. Ikawa, K. Nakahata, M. Zenitani, T. Yamamichi, S. Umeda, K. Deguchi, H. Okuyama, S. Miyagawa

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Background In our previous study, we reported on the development of substituting S147C for HLA-E as a useful gene tool for xenotransplantation. In this study we exchanged the codon of HLA-Ev (147), checked its function, and established a line of transgenic mice. Methods A new construct, a codon exchanging human HLA-Ev (147) + IRES + human beta 2-microgloblin, was established. The construct was subcloned into pCXN2 (the chick beta-actin promoter and cytomegalovirus enhancer) vector. Natural killer cell- and macrophage-mediated cytotoxicities were performed using the established the pig endothelial cell (PEC) line with the new gene. Transgenic mice with it were next produced using a micro-injection method. Results The expression of the molecule on PECs was confirmed by the transfection of the plasmid. The established molecules on PECs functioned well in regulating natural killer cell-mediated cytotoxicity and macrophage-mediated cytotoxicity. We have also successfully generated several lines of transgenic mice with this plasmid. The expression of HLA-Ev (147) in each mouse organ was confirmed by assessing the mRNA. The chick beta-actin promoter and cytomegalovirus enhancer resulted in a relatively broad expression of the gene in each organ, and a strong expression in the cases of the heart and lung. Conclusion A synthetic HLA-Ev (147) gene with a codon usage optimized to a mammalian system represents a critical factor in the development of transgenic animals for xenotransplantation.

Original languageEnglish
Pages (from-to)1323-1325
Number of pages3
JournalTransplantation Proceedings
Volume48
Issue number4
DOIs
Publication statusPublished - 2016 May 1
Externally publishedYes

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Genetically Modified Animals
Codon
Transgenic Mice
Heterologous Transplantation
Cytomegalovirus
Natural Killer Cells
Actins
Plasmids
Macrophages
Genes
Transfection
Swine
Endothelial Cells
Gene Expression
Cell Line
Lung
Messenger RNA
Injections

ASJC Scopus subject areas

  • Surgery
  • Transplantation

Cite this

Matsuura, R., Maeda, A., Sakai, R., Eguchi, H., Lo, P. C., Hasuwa, H., ... Miyagawa, S. (2016). Human HLA-Ev (147) Expression in Transgenic Animals. Transplantation Proceedings, 48(4), 1323-1325. https://doi.org/10.1016/j.transproceed.2015.10.069

Human HLA-Ev (147) Expression in Transgenic Animals. / Matsuura, R.; Maeda, A.; Sakai, R.; Eguchi, H.; Lo, P. C.; Hasuwa, Hidetoshi; Ikawa, M.; Nakahata, K.; Zenitani, M.; Yamamichi, T.; Umeda, S.; Deguchi, K.; Okuyama, H.; Miyagawa, S.

In: Transplantation Proceedings, Vol. 48, No. 4, 01.05.2016, p. 1323-1325.

Research output: Contribution to journalArticle

Matsuura, R, Maeda, A, Sakai, R, Eguchi, H, Lo, PC, Hasuwa, H, Ikawa, M, Nakahata, K, Zenitani, M, Yamamichi, T, Umeda, S, Deguchi, K, Okuyama, H & Miyagawa, S 2016, 'Human HLA-Ev (147) Expression in Transgenic Animals', Transplantation Proceedings, vol. 48, no. 4, pp. 1323-1325. https://doi.org/10.1016/j.transproceed.2015.10.069
Matsuura, R. ; Maeda, A. ; Sakai, R. ; Eguchi, H. ; Lo, P. C. ; Hasuwa, Hidetoshi ; Ikawa, M. ; Nakahata, K. ; Zenitani, M. ; Yamamichi, T. ; Umeda, S. ; Deguchi, K. ; Okuyama, H. ; Miyagawa, S. / Human HLA-Ev (147) Expression in Transgenic Animals. In: Transplantation Proceedings. 2016 ; Vol. 48, No. 4. pp. 1323-1325.
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abstract = "Background In our previous study, we reported on the development of substituting S147C for HLA-E as a useful gene tool for xenotransplantation. In this study we exchanged the codon of HLA-Ev (147), checked its function, and established a line of transgenic mice. Methods A new construct, a codon exchanging human HLA-Ev (147) + IRES + human beta 2-microgloblin, was established. The construct was subcloned into pCXN2 (the chick beta-actin promoter and cytomegalovirus enhancer) vector. Natural killer cell- and macrophage-mediated cytotoxicities were performed using the established the pig endothelial cell (PEC) line with the new gene. Transgenic mice with it were next produced using a micro-injection method. Results The expression of the molecule on PECs was confirmed by the transfection of the plasmid. The established molecules on PECs functioned well in regulating natural killer cell-mediated cytotoxicity and macrophage-mediated cytotoxicity. We have also successfully generated several lines of transgenic mice with this plasmid. The expression of HLA-Ev (147) in each mouse organ was confirmed by assessing the mRNA. The chick beta-actin promoter and cytomegalovirus enhancer resulted in a relatively broad expression of the gene in each organ, and a strong expression in the cases of the heart and lung. Conclusion A synthetic HLA-Ev (147) gene with a codon usage optimized to a mammalian system represents a critical factor in the development of transgenic animals for xenotransplantation.",
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AU - Maeda, A.

AU - Sakai, R.

AU - Eguchi, H.

AU - Lo, P. C.

AU - Hasuwa, Hidetoshi

AU - Ikawa, M.

AU - Nakahata, K.

AU - Zenitani, M.

AU - Yamamichi, T.

AU - Umeda, S.

AU - Deguchi, K.

AU - Okuyama, H.

AU - Miyagawa, S.

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N2 - Background In our previous study, we reported on the development of substituting S147C for HLA-E as a useful gene tool for xenotransplantation. In this study we exchanged the codon of HLA-Ev (147), checked its function, and established a line of transgenic mice. Methods A new construct, a codon exchanging human HLA-Ev (147) + IRES + human beta 2-microgloblin, was established. The construct was subcloned into pCXN2 (the chick beta-actin promoter and cytomegalovirus enhancer) vector. Natural killer cell- and macrophage-mediated cytotoxicities were performed using the established the pig endothelial cell (PEC) line with the new gene. Transgenic mice with it were next produced using a micro-injection method. Results The expression of the molecule on PECs was confirmed by the transfection of the plasmid. The established molecules on PECs functioned well in regulating natural killer cell-mediated cytotoxicity and macrophage-mediated cytotoxicity. We have also successfully generated several lines of transgenic mice with this plasmid. The expression of HLA-Ev (147) in each mouse organ was confirmed by assessing the mRNA. The chick beta-actin promoter and cytomegalovirus enhancer resulted in a relatively broad expression of the gene in each organ, and a strong expression in the cases of the heart and lung. Conclusion A synthetic HLA-Ev (147) gene with a codon usage optimized to a mammalian system represents a critical factor in the development of transgenic animals for xenotransplantation.

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