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TISSUE FRAGMENT UPDATE

外国特許コード F180009641
整理番号 4601
掲載日 2018年11月21日
出願国 世界知的所有権機関(WIPO)
国際出願番号 2015JP079364
国際公開番号 WO 2016060260
国際出願日 平成27年10月16日(2015.10.16)
国際公開日 平成28年4月21日(2016.4.21)
優先権データ
  • 特願2014-212008 (2014.10.16) JP
  • 特願2014-212010 (2014.10.16) JP
  • 特願2014-212014 (2014.10.16) JP
発明の名称 (英語) TISSUE FRAGMENT UPDATE
発明の概要(英語) The present invention provides a method for producing a tissue fragment, particularly a myocardial tissue fragment, which contains cultured cells having an oriented configuration, said method comprising culturing cells on an oriented fiber sheet. The present invention also provides a tissue fragment produced by the method according to the present invention. The present invention further provides: a device for evaluating an electrophysiological function of a cell; and a method for evaluating an electrophysiological function of a cell. The present invention still further provides a sheet for culturing cells.
従来技術、競合技術の概要(英語) BACKGROUND ART
Severe heart failure including myocardial infarction patient is about 80 million people in Japan of the disease and, about 18 million people die each year. At present the only effective treatment for heart transplantation but, in Japan and extremely serious shortage of donors, such as immune rejection also suffers. Pluripotent stem cells (for example, embryonic pluripotent stem cells (ES) or induced pluripotent stem cell (iPS) ) established by, as a therapeutic alternative to heart transplantation, myocardial cell regeneration treatment and can be formed to restore myocardial function expected. Is for use in therapy in clinical transplantation, to imitate the structure having the same sequence as in vivo, the mature high safe myocardial tissue piece is required to be obtained.
A method for inducing human ES/iPS cells into cardiac muscle cells have been proposed various. Used to induce a current of heart muscle cells is the most general method, cytokine of embryoid bodies (for example, DKK1, bFGF, activin A (ActivinA), and BMP4) and the suspension culture, or mouse END2 (the proximal endoderm-like cells (visceralendoderm) ) co-culture is the removal of the (non-patent document 1-4).
The inventors of the present invention, novel low-molecular-weight compound is used as an efficient stable myocardial differentiation induction method (-98%) clinical grade proposed (non-patent document 5, patent document 1). Myocardial cells induced by such a method, human differentiated cell is an important maturation marker of heart muscle cells is the amount of β-MHC gene expression is less than the amount of expression of the adult human heart (-10%), sequence does not have the myocardial tissue structure, sufficient mature electrophysiological there occurs a problem cannot be obtained.
Myocardial cell transplantation method has been proposed as the method can be divided roughly into the following 2 types: (1) a disease of heart failure and cardiac cells by direct injection of the cells in a sheet form of the method (2), to a method for implanting a disease.
Derived from human ES cells by direct injection of heart muscle cells, improve myocardial infarction in guinea pig has succeeded (non-patent document 6) report. However direct injection method, into a cell solution which flows out from the myocardial tissue, which are applied to the systemic blood flow such as this happens, the low efficiency of cell transplantation is pointed out. The number of the cell engraftment % or less, is not sufficient therapeutic effect is considered to be.
In addition, the temperature-responsive culture dish as a cell sheet technique of the cells with a technology in which a sheet has been proposed (non-patent document 7). And using such a technique myocardial sheet which, in clinical trials to transplant it has been started. However, this constructed using the techniques of the myocardial cell is a single-layered sheet, orientation is not randomly distributed, which mimic the structure of the myocardium within the living body cannot be. Therefore, myocardial cell contraction force is weak, the degree of maturity is low, a problem in that the handling is difficult to lower strength still remains. In addition, remains formed on a support sheet for the production of implantable cardiac muscle cells has not been reported yet.
More effective for use in transplantation and assays, in biological cells which mimic the structure and the fabrication of a piece of tissue structure has been desired, micro-engineering technique in biological cells which mimic the structure studies have been attracting attention. For example in vivo, and a cardiac muscle cells are an array structure, the microline or, punch or micro, fiber or other technique, attempts are sequenced cardiomyocyte culture have been reported (non-patent document 8-14).
The evaluation of the development of a drug safety in conventional, animal-derived primary culture cells or an animal has been used, and the functions in cells of the tissue human bio species difference, various problems are caused by a lot of the differences in the are. In addition, at present, all areas of the electrocardiogram QT extension for a medicament for cardiotoxicity in non-clinical experimental practice guideline is obtained. Such as iPS cells induced pluripotent stem cells into cardiac muscle cells, myocardial tissue obtained and cultured to evaluate the toxicity of the drug piece applied in the evaluation with respect to the behavior or a high demand of the industry.
Is the electrophysiological evaluation of cells, intracellular recording the field potential can be patch-clamp method is have been frequently used. Patch-clamp method is however, an infested cells by patch electrodes, a long-time measurement difficult. In addition, the procedure is not suitable for screening difficult.
Owing to the development of multi-electrode technique, the human ES/iPS cell-derived multi-electrode on the cultured myocardial cells, their electrophysiology function and it is possible to evaluate the (non-patent document 15 and 16). This technique for measuring activity extracellular potential, without damaging the cells, while the function observed for a long period of cell culture, can be measured. Essential to this technology in drug discovery as well as evaluate the toxicity of the drug, such as human ES cells and iPS cells from pluripotent stem cell into a cardiomyocyte differentiation induction and maturity can be possible to monitor the processes respectively.
Myocardial differentiation induction obtained in conventional myocardial cell tissue of heart muscle cells are aligned in a direction not scattered muscle contraction force is weak, and obtaining an electrocardiogram QT extension experiment or the like to arrhythmia occurs frequently, weak adhesion between the electrodes, the long-term culture cannot be set, for use in drug discovery applications, in the evaluation system is provided as a stable but still has a problem. In the first place, the cardiomyocytes random, or lumps had grown to electrical signals from myocardial tissue, myocardial tissue in vivo behavior difficult to say that sufficiently mimic.
  • 出願人(英語)
  • ※2012年7月以前掲載分については米国以外のすべての指定国
  • KYOTO UNIVERSITY
  • 発明者(英語)
  • LIU, Li
  • LI, Junjun
  • MINAMI, Itsunari
  • CHEN, Yong
  • NAKATSUJI, Norio
国際特許分類(IPC)
指定国 National States: AE AG AL AM AO AT AU AZ BA BB BG BH BN BR BW BY BZ CA CH CL CN CO CR CU CZ DE DK DM DO DZ EC EE EG ES FI GB GD GE GH GM GT HN HR HU ID IL IN IR IS JP KE KG KN KP KR KZ LA LC LK LR LS LU LY MA MD ME MG MK MN MW MX MY MZ NA NG NI NO NZ OM PA PE PG PH PL PT QA RO RS RU RW SA SC SD SE SG SK SL SM ST SV SY TH TJ TM TN TR TT TZ UA UG US UZ VC VN ZA ZM ZW
ARIPO: BW GH GM KE LR LS MW MZ NA RW SD SL SZ TZ UG ZM ZW
EAPO: AM AZ BY KG KZ RU TJ TM
EPO: AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR
OAPI: BF BJ CF CG CI CM GA GN GQ GW KM ML MR NE SN ST TD TG
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