Immortalized prairie vole-derived fibroblasts (VMF-K4DTs) can be transformed into pluripotent stem cells and provide a useful tool with which to determine optimal reprogramming conditions

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  • KATAYAMA Masafumi
    Graduate School of Agricultural Science, Tohoku University, Sendai 981-8555, Japan National Institute for Environmental Studies, Japan, Center for Environmental Biology and Ecosystem Studies, Ibaraki 305-8506, Japan Wildlife Genome Collaborative Research Group, National Institute for Environmental Studies, Ibaraki 305-8506, Japan
  • HIRAYAMA Takashi
    Department of Obstetrics and Gynecology, Juntendo University, Tokyo 113-8421, Japan
  • KIYONO Tohru
    Division of Carcinogenesis and Cancer Prevention, National Cancer Center Research Institute, Tokyo 104-0045, Japan
  • ONUMA Manabu
    National Institute for Environmental Studies, Japan, Center for Environmental Biology and Ecosystem Studies, Ibaraki 305-8506, Japan Wildlife Genome Collaborative Research Group, National Institute for Environmental Studies, Ibaraki 305-8506, Japan
  • TANI Tetsuya
    Laboratory of Animal Reproduction, Department of Agriculture, Kindai University, Nara 3327-204, Japan
  • TAKEDA Satoru
    Department of Obstetrics and Gynecology, Juntendo University, Tokyo 113-8421, Japan
  • NISHIMORI Katsuhiko
    Graduate School of Agricultural Science, Tohoku University, Sendai 981-8555, Japan
  • FUKUDA Tomokazu
    Wildlife Genome Collaborative Research Group, National Institute for Environmental Studies, Ibaraki 305-8506, Japan United Graduate School of Agricultural Sciences, Iwate University, Morioka 020-8551, Japan

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<p> The cellular conditions required to establish induced pluripotent stem cells (iPSCs), such as the number of reprogramming factors and/or promoter selection, differ among species. The establishment of iPSCs derived from cells of previously unstudied species therefore requires the extensive optimization of programming conditions, including promoter selection and the optimal number of reprogramming factors, through a trial-and-error approach. While the four Yamanaka factors Oct3/4, Sox2, Klf4, and c-Myc are sufficient for iPSC establishment in mice, we reported previously that six reprogramming factors were necessary for the creation of iPSCs from primary prairie vole-derived cells. Further to this study, we now show detailed data describing the optimization protocol we developed in order to obtain iPSCs from immortalized prairie vole-derived fibroblasts. Immortalized cells can be very useful tools in the optimization of cellular reprogramming conditions, as cellular senescence is known to dramatically decrease the efficiency of iPSC establishment. The immortalized prairie vole cells used in this optimization were designated K4DT cells as they contained mutant forms of CDK4, cyclin D, and telomerase reverse transcriptase (TERT). We show that iPSCs derived from these immortalized cells exhibit the transcriptional silencing of exogenous reprogramming factors while maintaining pluripotent cell morphology. There were no observed differences between the iPSCs derived from primary and immortalized prairie vole fibroblasts. Our data suggest that cells that are immortalized with mutant CDK4, cyclin D, and TERT provide a useful tool for the determination of the optimal conditions for iPSC establishment.</p>

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