Embryonal Carcinoma and Glioblastoma Cell Lines Derived from Monkey Induced Pluripotent Stem Cells
Induced pluripotent stem cells (iPSCs) are useful for the development of therapies in regenerative medicine, analysis of pathogenesis, and exploration of candidate drugs. We developed an alternative usage of iPSCs of which the MHC haplotype is matched to
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Embryonal Carcinoma and Glioblastoma Cell Lines Derived from Monkey Induced Pluripotent Stem Cells Hirohito Ishigaki and Yasushi Itoh Abstract Induced pluripotent stem cells (iPSCs) are useful for the development of therapies in regenerative medicine, analysis of pathogenesis, and exploration of candidate drugs. We developed an alternative usage of iPSCs of which the MHC haplotype is matched to transplantable recipients in a cynomolgus macaque model. We established two cancer cell lines, embryonal carcinoma, and glioblastoma cell lines from cynomolgus monkey iPSCs. Here, we describe a method to induce the cancer cell lines including a technique for culture of the monkey iPSCs on feeder cells and the induction of hematopoietic stem cells and neural progenitor cells from monkey iPSCs. Keywords iPSC, Cynomolgus macaque, Cancer cell line, Oncogene, Retrovirus, Embryonal carcinoma, Glioblastoma
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Introduction Induced pluripotent stem cells (iPSCs) are useful not only for treatment in regenerative medicine but also for establishment of disease models to study the pathogenesis and to evaluate drug efficacy and toxicity [1–3]. We developed an alternative usage of iPSCs of which the MHC haplotype is matched to transplantable recipients in a cynomolgus macaque model. We have established two cancer cell lines, embryonal carcinoma and glioblastoma cell lines, from cynomolgus monkey iPSCs that have a homozygous haplotype of MHC. Somatic progenitor cells derived from iPSCs were transduced with oncogenes specific for each cancer to examine biological characteristics of the cancers and to develop anticancer drugs for cancers with high-malignant potential such as glioblastoma in in vitro and in vivo studies [4, 5]. We used iPSCs derived from fibroblasts of cynomolgus macaques, in which the structures of biologically relevant molecules are similar to those of biologically relevant molecules in humans [6– 10]. Indeed, more than half of the antibodies against human molecules react to molecules of cynomolgus monkeys [6, 7]. In addition, monkey bronchial epithelial cells were transformed to lung
Hirohito Ishigaki and Yasushi Itoh
adenocarcinoma cells by induction of human lung cancer-specific oncogenes, and the morphology and histological type of the carcinoma cells were the same as those of human lung cancer cells ([8] and unpublished data). Using iPSCs instead of somatic cells for establishment of cancer cell lines allows us to develop various types of cancer cell lines, and the use of iPSCs may reveal the pathogenesis of various cancers [4].
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Materials
2.1 Culture of iPSCs (See Note 1)
1. iPSCs: iPSCs were established from skin fibroblasts of a 3-yearold female cynomolgus macaque. For iPSC induction, the fibroblasts were transduced with human OCT3/4, SOX2, KLF4, and c-MYC genes by using lentivirus vectors after transduction of the mouse solute carrier family 7 (cationic amino acid transporter, y+, system), member 1 (Slc7a1) gene [12, 13]. Store in liquid nitrogen. 2. Feeder cells: Mouse embryonic fibroblasts (MEFs, ReproCELL
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