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 language | English |
|---|---|
| Pages (from-to) | 1323-1325 |
| Number of pages | 3 |
| Journal | Transplantation Proceedings |
| Volume | 48 |
| Issue number | 4 |
| DOIs | |
| Publication status | Published - 2016 May 1 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Surgery
- Transplantation
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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 journal › Article
}
TY - JOUR
T1 - Human HLA-Ev (147) Expression in Transgenic Animals
AU - Matsuura, R.
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.
PY - 2016/5/1
Y1 - 2016/5/1
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.
AB - 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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U2 - 10.1016/j.transproceed.2015.10.069
DO - 10.1016/j.transproceed.2015.10.069
M3 - Article
C2 - 27320614
AN - SCOPUS:84975138684
VL - 48
SP - 1323
EP - 1325
JO - Transplantation Proceedings
JF - Transplantation Proceedings
SN - 0041-1345
IS - 4
ER -