Label-free characterization of living human induced pluripotent stem cells by subcellular topographic imaging technique using full-field quantitative phase microscopy coupled with interference reflection microscopy
Sugiyama N.,Hamamatsu Photonics K K |
Asai Y.,ReproCELL |
Yamauchi T.,Hamamatsu Photonics K K |
Kataoka T.,Hamamatsu Photonics K K |
And 4 more authors.
Biomedical Optics Express | Year: 2012
There is a need for a noninvasive technique to monitor living pluripotent stem cell condition without any labeling. We present an optical imaging technique that is able to capture information about optical path difference through the cell and cell adhesion properties simultaneously using a combination of quantitative phase microscopy (QPM) and interference reflection microscopy (IRM) techniques. As a novel application of QPM and IRM, this multimodal imaging technique demonstrated its ability to distinguish the undifferentiated status of human induced pluripotent stem (hiPS) cells quantitatively based on the variation of optical path difference between the nucleus and cytoplasm as well as hiPS cellspecific cell adhesion properties. © 2012 Optical Society of America.
Minami I.,Kyoto University |
Yamada K.,Kyoto University |
Yamada K.,Kanazawa University |
Otsuji T.G.,Kyoto University |
And 12 more authors.
Cell Reports | Year: 2012
Human pluripotent stem cells (hPSCs), including embryonic stem cells and induced pluripotent stem cells, are potentially useful in regenerative therapies for heart disease. For medical applications, clinical-grade cardiac cells must be produced from hPSCs in a defined, cost-effective manner. Cell-based screening led to the discovery of KY02111, a small molecule that promotes differentiation of hPSCs to cardiomyocytes. Although the direct target of KY02111 remains unknown, results of the present study suggest that KY02111 promotes differentiation by inhibiting WNT signaling in hPSCs but in a manner that is distinct from that of previously studied WNT inhibitors. Combined use of KY02111 and WNT signaling modulators produced robust cardiac differentiation of hPSCs in a xeno-free, defined medium, devoid of serum and any kind of recombinant cytokines and hormones, such as BMP4, Activin A, or insulin. The methodology has potential as a means for the practical production of human cardiomyocytes for regeneration therapies.
Miyazaki T.,Yokohama City University |
Fujita H.,Yokohama City University |
Fujimaki K.,Yokohama City University |
Hosoyama T.,ReproCELL |
And 12 more authors.
Leukemia Research | Year: 2012
We analyzed minimal residual disease (MRD) by multidimensional flow cytometry (MFC) after allogeneic stem cell transplantation in 41 patients with acute myeloid leukemia (AML) (n= 31) or acute lymphoblastic leukemia (ALL) (n= 10). Aberrant antigen expression was compared with the results of quantitative PCR for WT1 mRNA (n= 41) and leukemia-specific fusion transcripts (n= 12; AML in seven, ALL in five). There was a significant correlation between detection of MRD by MFC and WT1 mRNA, as well as between MFC and fusion transcripts. Serial monitoring of MRD by the three techniques correlated in parallel to the clinical course in most of the patients, but three patients were only positive for WT1 during hematological remission. The overall survival time of patients with complete remission was significantly associated with the appearance of aberrant expression after transplantation. In conclusion, MFC is valuable for clinical management decisions after transplantation. © 2012 Elsevier Ltd.
Murakami G.,Kyoto University |
Inoue H.,Kyoto University |
Inoue H.,Japan Science and Technology Agency |
Tsukita K.,Kyoto University |
And 9 more authors.
Journal of Biomolecular Screening | Year: 2011
Familial amyotrophic lateral sclerosis (fALS) accounts for 10% of ALS cases, and about 25% of fALS cases are due to mutations in superoxide dismutase 1 (SOD1). Mutant SOD1-mediated ALS is caused by a gain of toxic function of the mutant protein, and the SOD1 level in nonneuronal neighbors, including astrocytes, determines the progression of ALS (non-cell-autonomous toxicity). Therefore, the authors hypothesized that small molecules that reduce SOD1 protein levels in astrocytes might slow the progression of mutant SOD1-mediated ALS. They developed and optimized a cell-based, high-throughput assay to identify low molecular weight compounds that decrease SOD1 expression transcriptionally in human astrocyte-derived cells. Screening of a chemical library of 9600 compounds with the assay identified two hit compounds that selectively and partially downregulate SOD1 expression in a dose-dependent manner, without any detectable cellular toxicity. Western blot analysis showed that one hit compound significantly decreased the level of endogenous SOD1 protein in H4 cells, with no reduction in expression of ß-actin. The assay developed here provides a powerful strategy for discovering novel lead molecules for treating familial SOD1-mediated ALS. (Journal of Biomolecular Screening 2011;16:405-414) © 2011 Society for Laboratory Automation and Screening.
Asai Y.,ReproCELL |
Tada M.,Japan Building Research Institute |
Tada M.,Kyoto University |
Otsuji T.G.,Japan Building Research Institute |
And 2 more authors.
Current Stem Cell Research and Therapy | Year: 2010
Human pluripotential stem cells including both embryonic stem cells (hESC) and induced pluripotent stem cells (hiPSC) possess self-renewing potency and pluripotentency and can differentiate into virtually any somatic cell type. These features are a distinct advantage for the generation of specific types of human tissue cells in vitro for continuous use in drug development. Recently, an assay system for drug-induced QT interval prolongation using hESC/hiPSC-derived cardiomyocytes and microelectrode arrays (MEA) has been developed. Drug-induced QT interval prolongation (DIQTIP) can lead to sudden cardiac death and is a major safety concern for the drug industry. Regulatory authorities such as the US FDA and the European Medicines Agency require in-vitro testing of all drug candidates to identify potential risk of DIQTIP prior to clinical trials. To reduce the risk of DIQTIP, a routine assay system for in vitro electrophysiological properties using cell-based assays is effective and necessary in early phase of drug discovery. This review discusses developments over the last couple of years for a qualified drug testing method and provides some examples of how hESC/hiPSC-derived cardiomyocytes are beginning to find a practical use for drug discovery and development. © 2010 Bentham Science Publishers Ltd.