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, H.
AU - Ikawa, M.
AU - Nakahata, K.
AU - Zenitani, M.
AU - Yamamichi, T.
AU - Umeda, S.
AU - Deguchi, K.
AU - Okuyama, H.
AU - Miyagawa, S.
N1 - Funding Information:
This work was supported by Grants-in Aid for Scientific Research, and Health and Labor Sciences Research Grants, Japan.
Publisher Copyright:
© 2016 Elsevier Inc. All rights reserved.
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 -