ORCID Profile
0000-0002-3530-3015
Current Organisations
Keio University
,
Elixirgen Therapeutics
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Publisher: Oxford University Press (OUP)
Date: 05-06-2014
Abstract: Keratinocytes represent an easily accessible cell source for derivation of human induced pluripotent stem (hiPS) cells, reportedly achieving higher reprogramming efficiency than fibroblasts. However, most studies utilized a retroviral or lentiviral method for reprogramming of keratinocytes, which introduces undesirable transgene integrations into the host genome. Moreover, current protocols of generating integration-free hiPS cells from keratinocytes are mostly inefficient. In this paper, we describe a more efficient, simple-to-use, and cost-effective method for generating integration-free hiPS cells from keratinocytes. Our improved method using lipid-mediated transfection achieved a reprogramming efficiency of ∼0.14% on average. Keratinocyte-derived hiPS cells showed no integration of episomal vectors, expressed stem cell-specific markers and possessed potentials to differentiate into all three germ layers by in vitro embryoid body formation as well as in vivo teratoma formation. To our knowledge, this represents the most efficient method to generate integration-free hiPS cells from keratinocytes.
Publisher: Bioscientifica
Date: 07-2006
DOI: 10.1530/REP.1.01059
Abstract: A series of Ca 2+ oscillations during mammalian fertilization is necessary and sufficient to stimulate meiotic resumption and pronuclear formation. It is not known how effectively development continues in the absence of the initial Ca 2+ signal. We have triggered parthenogenetic egg activation with cycloheximide that causes no Ca 2+ increase, with ethanol that causes a single large Ca 2+ increase, or with Sr 2+ that causes Ca 2+ oscillations. Eggs were co-treated with cytochalasin D to make them diploid and they formed pronuclei and two-cell embryos at high rates with each activation treatment. However, far fewer of the embryos that were activated by cycloheximide reached the blastocyst stagecompared tothose activated by Sr 2+ orethanol. Any cycloheximide-activated embryos that reached the blastocyst stage had a smaller inner cell mass number and a greater rate of apoptosis than Sr 2+ -activated embryos. The poor development of cycloheximide-activated embryos was due to the lack of Ca 2+ increase because they developed to blastocyst stages at high rates when co-treated with Sr 2+ or ethanol. Embryos activated by either Sr 2+ or cycloheximide showed similar signs of initial embryonic genome activation (EGA) when measured using a reporter gene. However, microarray analysis of gene expression at the eight-cell stage showed that activation by Sr 2+ leads to a distinct pattern of gene expression from that seen with embryos activated by cycloheximide. These data suggest that activation of mouse eggs in the absence of a Ca 2+ signal does not affect initial parthenogenetic events, but can influence later gene expression and development.
Publisher: Oxford University Press (OUP)
Date: 05-05-2013
No related grants have been discovered for Minoru Ko.