JP7479466B2 - 血管内皮細胞純粋分離方法、血管内皮細胞特性維持培地及びそれを含む培養方法 - Google Patents
血管内皮細胞純粋分離方法、血管内皮細胞特性維持培地及びそれを含む培養方法 Download PDFInfo
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Description
図5aは、本発明の一実施例に係る血管内皮細胞純粋分離方法でフィルタの有無によるマーカー発現及び顕微鏡イメージを結果を示したものである。
図5bは、本発明の一実施例に係る血管内皮細胞純粋分離方法でフィルタの有無によるマーカー発現及び顕微鏡イメージを結果を示したものである。
図6aは、本発明の一実施例に係る血管内皮細胞純粋分離方法によって分離された血管内皮細胞のマーカー発現の結果を示したものである。
図11aから図11cは、本発明の一実施例に係る血管内皮細胞特性維持培養でマーカーに対する陽性血管内皮細胞の相対的発現水準を示す結果である。
Claims (14)
- ヒト万能幹細胞から分化された内皮細胞系統の細胞株を分化培地から収得するステップ;
収得された前記細胞株をフィルタを利用してろ過するステップ;
ろ過された前記細胞株を基質上に培養するステップ、及び
培養された前記細胞株から基質上に付着された同質性(homogenous)の内皮細胞を分離するステップを含む、血管内皮細胞純粋分離方法。 - 前記フィルタは、
空隙間隔が20~40μmの範囲である、請求項1に記載の血管内皮細胞純粋分離方法。 - 前記基質は、
コラーゲン、フィブリン、フィブロネクチン、ビトロネクチン、マトリゲル、ゼラチン、ラミニン、ヘパリン、ポリリジン及びヒアルロン酸のうち少なくとも一つを含む、請求項1または2に記載の血管内皮細胞純粋分離方法。 - 前記基質は、
コラーゲンであり、0.1mg/mlの前記コラーゲンを含む、請求項3に記載の血管内皮細胞純粋分離方法。 - 前記培養するステップは、
細胞成長因子及びアスコルビン酸を含むDMEM/F-12培地で培養する、請求項1から4のいずれか一項に記載の血管内皮細胞純粋分離方法。 - 前記細胞成長因子は、
FGF-1(fibroblast growth factor-1)、FGF-2(bFGF)、FGF-3、FGF-4、FGF-5、FGF-6、EGF(epidermal growth factor)、KGF(keratinocyte growth factor)、HGF(hepatocyte growth factor)、TGF-α(transforming growth factor-α)、TGF-β、アンジオポエチン1(angipoietin 1)、アンジオポエチン2、エリスロポエチン(erythropoietin)、ニューロピリン、IGF-1、オステオポンチン、プレイオトロフィン、アクチビン、エンドセリン01及びVEGF-A(vascular endothelial growth factor-A)のうち少なくとも一つを含む、請求項5に記載の血管内皮細胞純粋分離方法。 - 前記培養するステップは、
ろ過された前記細胞株を2個の基質上に播種するステップを含む、請求項1から6のいずれか一項に記載の血管内皮細胞純粋分離方法。 - 前記培養するステップは、
4時間から20時間の間培養される、請求項1から7のいずれか一項に記載の血管内皮細胞純粋分離方法。 - 前記同質性の内皮細胞は、
CDH5及びVWFを発現する、請求項1に記載の血管内皮細胞純粋分離方法。 - 前記CDH5の遺伝子発現水準は、
分離される前より12倍高い、請求項9に記載の血管内皮細胞純粋分離方法。 - 前記VWFの遺伝子発現水準は、
分離される前より2倍高い、請求項9または10に記載の血管内皮細胞純粋分離方法。 - 請求項1から11のいずれか一項に記載の血管内皮細胞純粋分離方法で分離する工程を含み、当該工程により分離されたCDH5及びVWFを発現する同質性(homogenous)の内皮細胞を98%以上含有する、血管内皮細胞の細胞株の製造方法。
- 請求項12に記載の血管内皮細胞の細胞株の製造方法により製造する工程を含み、当該工程により製造された98%以上の純度を有する血管内皮細胞を含む、心血管系疾患の予防用または治療用細胞治療剤組成物の製造方法。
- 前記心血管系疾患は、
虚血性心疾患、心不全、高血圧性心疾患、不整脈、心臓管膜症、心室中隔欠損、先天性心疾患、心筋症、心嚢疾患、脳卒中、末梢血管疾患、動脈瘤、動脈硬化症、高血圧、狭心症及び心筋梗塞症のうち少なくとも一つである、請求項13に記載の心血管系疾患の予防用または治療用細胞治療剤組成物の製造方法。
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US7247477B2 (en) * | 2002-04-16 | 2007-07-24 | Technion Research & Development Foundation Ltd. | Methods for the in-vitro identification, isolation and differentiation of vasculogenic progenitor cells |
EP2401364B1 (en) * | 2009-02-27 | 2015-04-22 | Cellular Dynamics International, Inc. | Differentiation of pluripotent cells |
BR112013030166A2 (pt) * | 2011-06-09 | 2017-03-21 | Hoffmann La Roche | método de diferenciação de células haste pluripotentes em células leito vasculares, células endoteliais ou células dos músculos moles vasculares, banco biológico e uso de células endoteliais ou células dos músculos moles vasculares, composição terapêutica, métodos e usos. |
KR101812979B1 (ko) * | 2015-07-10 | 2017-12-29 | 가톨릭대학교 산학협력단 | 동질성을 가지는 줄기세포 분획의 분리방법 |
US10669529B2 (en) * | 2015-07-17 | 2020-06-02 | Kyoto University | Method for inducing vascular endothelial cells |
CN115927199A (zh) * | 2015-11-04 | 2023-04-07 | 菲特治疗公司 | 用于诱导造血细胞分化的方法和组合物 |
CN109153970A (zh) * | 2016-05-17 | 2019-01-04 | 新加坡科技研究局 | 人类干细胞衍生的内皮细胞、内皮-肝细胞共培养***以及其用途 |
WO2019025826A1 (en) * | 2017-08-02 | 2019-02-07 | Semmelweis Egyetem | METHODS FOR PRODUCING ENDOTHELIAL CELLS AND VASCULAR ENDOTHELIAL CONSTRUCTS DERIVED FROM INDUCED HUMAN PLURIPOTENT STEM CELLS |
-
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Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2010131373A (ja) | 2008-10-30 | 2010-06-17 | Japan Health Science Foundation | 血管平滑筋細胞増殖を抑制する血管狭窄部挿入用基材 |
JP2015519890A (ja) | 2012-04-24 | 2015-07-16 | ダン エス. カウフマン, | 幹細胞よりナチュラルキラー細胞を発生させる方法 |
Non-Patent Citations (2)
Title |
---|
Circulation,2017年,Vol.136,pp.1939-1954 |
Curr. Cardiol. Rep.,2018年,Vol.20, No.6,Article number 45 |
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US20220378845A1 (en) | 2022-12-01 |
JP2023501419A (ja) | 2023-01-18 |
EP4060024A4 (en) | 2023-12-06 |
CA3156948A1 (en) | 2021-05-20 |
WO2021096218A1 (ko) | 2021-05-20 |
EP4060024A1 (en) | 2022-09-21 |
CN114746543A (zh) | 2022-07-12 |
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