Faculty Profiles - ITO Akira

Papers - ITO Akira

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Medium temperature independent cytotoxicity of cell-adhesive heat-generating nanoparticles named magnetite cationic liposomes and its therapeutic use Reviewed Open Access

Tomio Morino, Hiroshi Takase, Toshiki Etani, Taku Naiki, Noriyasu Kawai, Akira Ito, Takahiro Yasui

Thermal Medicine Vol. 36 ( 1 ) page： 25-34 - 34 2020

Cre-Mediated Transgene Integration in Chinese Hamster Ovary Cells Using Minicircle DNA Vectors. Reviewed International journal

Wang X, Kawabe Y, Hada T, Ito A, Kamihira M

Biotechnology journal Vol. 13 ( 7 ) page： e1800063 2018.7

Language：English Publishing type：Research paper (scientific journal)

Bacterial backbone sequences of conventional plasmid vectors have been reported to exhibit negative effects on transgene expression in mammalian cells, such as cytotoxicity and gene silencing. Minicircle DNA vectors can be employed to overcome these issues and to improve the transfection efficiency because of their smaller size. In this study, transgenes are integrated into the hypoxanthine phosphoribosyltransferase (hprt) locus of Chinese hamster ovary (CHO) cells by the Cre-loxP system using minicircle DNA vectors as transgene donors. The targeted transgene integration efficiency is improved 2-3-fold (≈1.4%) using minicircle DNA vectors compared with conventional plasmid vectors. Moreover, clones with expected structures after transgene integration are obtained with a high frequency. When a transgene together with bacterial sequences derived from a plasmid vector is integrated into the hprt locus, the cell growth rate and antibody titer decrease. These results indicate that minicircle DNA vectors are more suitable than conventional plasmid vectors for transgene delivery in recombinant protein production using CHO cells.

DOI： 10.1002/biot.201800063

PubMed

Targeted knock-in of an scFv-Fc antibody gene into the hprt locus of Chinese hamster ovary cells using CRISPR/Cas9 and CRIS-PITCh systems Reviewed

Yoshinori Kawabe, Shinya Komatsu, Shodai Komatsu, Mai Murakami, Akira Ito, Tetsushi Sakuma, Takahiro Nakamura, Takashi Yamamoto, Masamichi Kamihira

Journal of Bioscience and Bioengineering Vol. 125 ( 5 ) page： 599 - 605 2018.5

Language：English Publishing type：Research paper (scientific journal) Publisher：Elsevier B.V.

Chinese hamster ovary (CHO) cells have been used as host cells for the production of pharmaceutical proteins. For the high and stable production of target proteins, the transgene should be integrated into a suitable genomic locus of host cells. Here, we generated knock-in CHO cells, in which transgene cassettes without a vector backbone sequence were integrated into the hprt locus of the CHO genome using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 and CRISPR-mediated precise integration into target chromosome (CRIS-PITCh) systems. We investigated the efficiency of targeted knock-in of transgenes using these systems. As a practical example, we generated knock-in CHO cells producing an scFv-Fc antibody using the CRIS-PITCh system mediated by microhomology sequences for targeting. We found that the CRIS-PITCh system can facilitate targeted knock-in for CHO cell engineering.

DOI： 10.1016/j.jbiosc.2017.12.003

Scopus

PubMed

Characterization of genetically engineered mouse hepatoma cells with inducible liver functions by overexpression of liver-enriched transcription factors Reviewed

Hideaki Yamamoto, Jane Marie Tonello, Takanori Sambuichi, Yoshinori Kawabe, Akira Ito, Masamichi Kamihira

Journal of Bioscience and Bioengineering Vol. 125 ( 1 ) page： 131 - 139 2018.1

Language：English Publishing type：Research paper (scientific journal) Publisher：Elsevier B.V.

New cell sources for the research and therapy of organ failure could significantly alleviate the shortage of donor livers that are available to patients who suffer from liver disease. Liver carcinoma derived cells, or hepatoma cells, are the ideal cells for developing bioartificial liver systems. Such cancerous liver cells are easy to prepare in large quantities and can be maintained over long periods under standard culture conditions, unlike primary hepatocytes. However, hepatoma cells possess only a fraction of the functions of primary hepatocytes. In a previous study, by transducing cells with liver-enriched transcription factors that could be inducibly overexpressed—hepatocyte nuclear factor (HNF)1α, HNF1β, HNF3β [FOXA2], HNF4α, HNF6, CCAAT/enhancer binding protein (C/EBP)α, C/EBPβ and C/EBPγ—we created mouse hepatoma cells with high liver-specific gene expression called the Hepa/8F5 cell line. In the present study, we performed functional and genetic analyses to characterize the Hepa/8F5 cell line. Further, in three-dimensional cultures, the function of these cells improved significantly compared to parental cells. Ultimately, these cells might become a new resource that can be used in basic and applied hepatic research.

DOI： 10.1016/j.jbiosc.2017.07.011

Scopus

PubMed

温熱誘導型遺伝子発現システムの開発と応用 Invited Reviewed

井藤　彰

放射線生物研究 Vol. 53 ( 2 ) page： 104-114 2018

ハイパーサーミアに伴う免疫賦活のメカニズム Invited Reviewed

井藤彰

放射線生物研究 Vol. 53 ( 2 ) page： 104-114 2018

Three-dimensional culture of a genetically modified hepatoma cell line using macroporous gelatin beads Reviewed Open Access

Jane Marie Tonello, Saori Kawashima, Kazuki Sato, Yoshinori Kawabe, Akira Ito, Masamichi Kamihira

CYTOTECHNOLOGY Vol. 69 ( 6 ) page： 925 - 931 2017.12

Language：English Publishing type：Research paper (scientific journal) Publisher：SPRINGER

Hepatoma cells are a candidate cell source for bio-artificial livers. However, they exhibit reduced liver functions compared with primary hepatocytes. In our previous study, genetically engineered mouse hepatoma cells were created by transduction with vectors mediating inducible overexpression of eight liver-enriched transcription factors. Upon the induction of the liver-enriched transcription factors transduced, the cells expressed both phenotypic and genotypic liver functions at high levels. In the present study, we performed three-dimensional culture of these cells using macroporous gelatin beads. When immobilized on the macroporous gelatin beads, these cells exhibited further enhancement in liver functionality, including increased albumin secretion, ammonia removal and cytochrome P450 activity. The levels of these functions were significantly enhanced compared to monolayer culture. The method is simple and scalable, and provides highly functional cells that can be used in basic and applied fields of hepatic research.

DOI： 10.1007/s10616-017-0117-0

Web of Science

PubMed

Accumulative scFv-Fc antibody gene integration into the &IThprt&IT chromosomal locus of Chinese hamster ovary cells Reviewed

Xue Wang, Yoshinori Kawabe, Risa Kato, Takeshi Hada, Akira Ito, Yoshimasa Yamana, Masako Kondo, Masamichi Kamihira

JOURNAL OF BIOSCIENCE AND BIOENGINEERING Vol. 124 ( 5 ) page： 583 - 590 2017.11

Language：English Publishing type：Research paper (scientific journal) Publisher：SOC BIOSCIENCE BIOENGINEERING JAPAN

We have previously developed an accumulative site-specific gene integration system (AGIS) using Cre-recombinase and mutated loxP sites. AGIS enables repeated transgene integration into a predetermined chromosomal site in mammalian cells. However, the process of establishing cells with multiple integrated copies of the transgene is still time-consuming. In the present study, we describe an improved version of AGIS that facilitates and accelerates the establishment of high-producer Chinese hamster ovary (CHO) cells. Two donor vectors were simultaneously introduced into the cells in a single transfection. Cells with successfully targeted transgene integration were screened based on a change in the color of the reporter fluorescent protein that they express. Repeated rounds of integration allowed the transgene copy number to be increased. As a model, an scFv-Fc antibody gene was integrated into the hprt locus of the CHO cell genome. After three rounds of integration, a high-producer CHO cell clone with six copies of the scFv-Fc gene was successfully established. scFv-Fc productivity was approximately four-fold greater than a control cell line harboring a single copy of the transgene. This newly designed AGIS procedure should facilitate the development of producer cells suitable for biopharmaceutical protein production. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.

DOI： 10.1016/j.jbiosc.2017.05.017

Web of Science

PubMed

Hypoxia-responsive transgene expression system using RTP801 promoter and synthetic transactivator fused with oxygen-dependent degradation domain Reviewed

Akihiko Ono, Akira Ito, Tomonaga Sato, Masaki Yamaguchi, Taiga Suzuki, Yoshinori Kawabe, Masamichi Kamihira

JOURNAL OF BIOSCIENCE AND BIOENGINEERING Vol. 124 ( 1 ) page： 115 - 124 2017.7

Language：English Publishing type：Research paper (scientific journal) Publisher：SOC BIOSCIENCE BIOENGINEERING JAPAN

Precise control of gene expression using an artificial gene circuit is a major challenge in the application of synthetic biology. Here, we designed a hypoxia-responsive transgene expression system by combining a hypoxia-inducible RTP801 promoter and a tetracycline-responsive transactivator fused with an oxygen-dependent degradation domain (TA-ODD). The reporter gene expression was highly induced by hypoxia when a transactivator-expression plasmid, pRTP801/TA-ODD, harboring a TA-ODD gene driven by the RTP801 promoter, was cotransfected with a reporter plasmid, pTRE/EGFP, harboring an EGFP gene controlled under the transactivator-responsive promoter. A stable cell line into which the expression units RTP801/TA-ODD and THE/EGFP had been introduced responded to hypoxia and expressed the reporter gene in an oxygen-concentration-dependent manner. Moreover, the cells demonstrated potential as sensors to detect hypoxic conditions in a three-dimensional tissue culture in vitro. These results indicate that the hypoxia-responsive transgene expression system is useful for constructing cell-based hypoxia detection systems. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.

DOI： 10.1016/j.jbiosc.2017.02.012

Web of Science

PubMed

Novel therapy for pancreatic fistula using adipose-derived stem cell sheets treated with mannose Reviewed

Kaneko Hirokazu, Kokuryo Toshio, Yokoyama Yukihiro, Yamaguchi Junpei, Yamamoto Tokunori, Shibata Rei, Gotoh Momokazu, Murohara Toyoaki, Ito Akira

SURGERY Vol. 161 ( 6 ) page： 1561 - 1569 2017.6

Effects of heat stimulation and L-ascorbic acid 2-phosphate supplementation on myogenic differentiation of artificial skeletal muscle tissue constructs Reviewed Open Access

Kazushi Ikeda, Akira Ito, Masanori Sato, Shota Kanno, Yoshinori Kawabe, Masamichi Kamihira

JOURNAL OF TISSUE ENGINEERING AND REGENERATIVE MEDICINE Vol. 11 ( 5 ) page： 1322 - 1331 2017.5

Language：English Publishing type：Research paper (scientific journal) Publisher：WILEY

Although skeletal muscle tissue engineering has been extensively studied, the physical forces produced by tissue-engineered skeletal muscles remain to be improved for potential clinical utility. In this study, we examined the effects of mild heat stimulation and supplementation of a L-ascorbic acid derivative, L-ascorbic acid 2-phosphate (AscP), on myoblast differentiation and physical force generation of tissue-engineered skeletal muscles. Compared with control cultures at 37 degrees C, mouse C2C12 myoblast cells cultured at 39 degrees C enhanced myotube diameter (skeletal muscle hypertrophy), whereas mild heat stimulation did not promote myotube formation (differentiation rate). Conversely, AscP supplementation resulted in an increased differentiation rate but did not induce skeletal muscle hypertrophy. Following combined treatment with mild heat stimulation and AscP supplementation, both skeletal muscle hypertrophy and differentiation rate were enhanced. Moreover, the active tension produced by the tissue-engineered skeletal muscles was improved following combined treatment. These findings indicate that tissue culture using mild heat stimulation and AscP supplementation is a promising approach to enhance the function of tissue-engineered skeletal muscles. Copyright (C) 2015 John Wiley & Sons, Ltd.

DOI： 10.1002/term.2030

Open Access

Web of Science

PubMed

Novel therapy for pancreatic fistula using adipose-derived stem cell sheets treated with mannose. Reviewed

Hirokazu Kaneko, Toshio Kokuryo, Yukihiro Yokoyama, Junpei Yamaguchi, Tokunori Yamamoto, Rei Shibata, Momokazu Gotoh, Toyoaki Murohara, Akira Ito, Masato Nagino

Surgery Vol. 161 ( 6 ) page： 1561–1569 2017

Multilayered adipose-derived regenerative cell sheets created by a novel magnetite tissue engineering method for myocardial infarction. Reviewed

Masakazu Ishii, Rei Shibata, Yuuki Shimizu, Takashi Yamamoto, Kazuhisa Kondo, Yoko Inoue, Noriyuki Ouchi, Tohru Tanigawa, Noriyoshi Kanemura, Akira Ito, Honda Honda, Toyoaki Murohara

International Journal of Cardiology. Vol. 175 ( 3 ) page： 545-553 2014

TLR4 and NLRP3 inflammasome activation 1 in monocytes by N-propionyl cysteaminylphenol-maleimid-dextran (NPCMD). Reviewed

Yu Mizote, Kazumasa Wakamatsu, Shosuke Ito, Akiko Uenaka, Yoshihiro Ohue, Koji Kurose, Midori Isobe, Akira Ito, Yasuaki Tamura, Hiroyuki Honda, Toshiharu Yamashita, Satoshi Nohara, Mikio Oka, Kowichi Jimbow, Eiichi Nakayama

Journal of Dermatological Science Vol. 73 ( 3 ) page： 209-215 2014

Heat-inducible gene expression system by applying alternating magnetic field to magnetic nanoparticles. Reviewed

Masaki Yamaguchi, Akira Ito, Akihiko Ono, Yoshinori Kawabe, Masamichi Kamihira

ACS Synthetic Biology Vol. 3 ( 5 ) page： 273-279 2014

Enhancement of contractile force generation of artificial skeletal muscle tissues by mild and transient heat treatment. Reviewed

Masanori Sato, Kazushi Ikeda, Shota Kanno, Akira Ito, Yoshinori Kawabe, Masamichi Kamihira

Current Pharmaceutical Biotechnology. Vol. 14 ( 13 ) page： 1083-1087 2014

T-cell receptor repertoires of tumor-infiltrating lymphocytes after hyperthermia using functionalized magnetite nanoparticles. Reviewed

Akira Ito, Masaki Yamaguchi, Noriaki Okamoto, Yuji Sanematsu, Yoshinori Kawabe, Kazumasa Wakamatsu, Shosuke Ito, Hiroyuki Honda, Takeshi Kobayashi, Eiichi Nakayama, Yasuaki Tamura, Masae Okura, Toshiharu Yamashita, Kowichi Jimbow, Masamichi Kamihira

Nanomedicine Vol. 8 ( 6 ) page： 891-902 2013

Melanoma-targeted chemo-thermotherapy and in situ peptide immunotherapy through HSP production by using melanogenesis substrate, NPrCAP and magnetite nanoparticles. Reviewed

Kowichi Jimbow, Yoshie Ishii-Osai, Shosuke Ito, Yasuaki Tamura, Akira Ito, Akihiro Yoneta, Takafumi Kamiya, Toshiharu Yamashita, Hiroyuki Honda, Kazumasa Wakamatsu, Katsutoshi Murase, Satoshi Nohara, Eiichi Nakayama, Takeo Hasegawa, Itsuo Yamamoto, Takeshi Kobayashi.

Journal of Skin Cancer Vol. 2013:742925. 2013

Effects of B-cell lymphoma 2 gene transfer to myoblast cells on skeletal muscle tissue formation using magnetic force-based tissue engineering. Reviewed Open Access

Masanori Sato, Akira Ito, Hirokazu Akiyama, Yoshinori Kawabe, Masamichi Kamihira

Tissue Engineering Part A Vol. 19 ( 1-2 ) page： 307-315 2013

Conjugation of magnetite nanoparticles with melanogenesis substrate, NPrCAP provides melanoma targeted, in situ peptide vaccine immunotherapy through HSP production by chemo-thermotherapy. Reviewed

Kowichi Jimbow, Yasuaki Tamura, Akihiro Yoneta, Takafumi Kamiya, Ichiro Ono, Toshiharu Yamashita, Akira Ito, Hiroyuki Honda, Kazumasa Wakamatsu, Shosuke Ito, Satoshi Nohara, Eiichi Nakayama and Takeshi Kobayashi

Journal of Biomaterials and Nanobiotechnology Vol. 3 page： 140-153 2012

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Papers - ITO Akira