成果報告書詳細
管理番号20120000000792
タイトル*平成22年度中間年報「次世代機能代替技術の研究開発/次世代再生医療技術の研究開発/生体内で自己組織の再生を促すセルフリー型再生デバイスの開発(幹細胞ニッチ制御による自己組織再生型心血管デバイスの基盤開発)国立大学法人大阪大学、ニプロ株式会社)
公開日2012/7/28
報告書年度2010 - 2010
委託先名国立大学法人大阪大学 ニプロ株式会社
プロジェクト番号P10004
部署名バイオテクノロジー・医療技術部
和文要約和文要約等以下本編抜粋:
1.研究開発の内容及び成果等
1) 幹細胞ニッチの探索と再構築技術の開発
(担当:国立大学法人 大阪大学 蛋白質研究所)
1)?1 幹細胞ニッチを構築する基底膜分子の同定とその機能評価
幹細胞ニッチを構築する細胞外マトリックスの実体を解明するために、対象臓器の幹細胞の局在部位を調べる技術開発として、組織幹細胞を可視化する “ラベル保持細胞(label-retaining cell)”標識法を応用し、心臓組織の幹細胞の可視化技術を開発した。具体的には、BrdUに替わりEdU(5-ethynyl-2'-deoxy-uridine)を用いてラベル保持細胞を可視化する方法に加えて、GFPを利用する可視化方法を開発した。
1)?2 増殖因子・分化誘導因子を組み込んだ人工幹細胞ニッチの構築
心筋再生に関連するactivin、BMPなどの液性因子をニッチ分子とともにデバイス表面に固相化するため、これら液性因子と主要基底膜蛋白質との相互作用を網羅的に解析し、activin前駆体がヘパラン硫酸プロテオグリカン依存的に基底膜に結合することを明らかにした。また、I型コラーゲンを主成分とするデバイス表面にラミニン等の基底膜分子を固相化するための組換えタンパク質の作製を行った。
英文要約Title; Research and Development of Next-generation Regenerative Technology / Basic research and development of cell-free devices for regenerative medicine

Author; Osaka University, Nipro Corporation

1. Searching and restructuring of stem cell niche
Two novel protocols to visualize tissue stem cells as "label-retaining cells" have been elaborated by utilizing either 5-ethynyl-2'-deoxyl-uridine (for labeling DNA) or histone-green fluorescent protein chimeric protein. The protocol has been successfully applied to label tissue stem cells in the heart and the liver.
2. Development of recruitment and differentiation factor of stem cell
Using microarray analysis of gene expression, HMGB1 increases the expression of CXCR4in bone marrow-derived mesenchymal stem cells (lineage-/PDGFR+; L-P+), which results in the accumulation of those cells in injured tissues enriched in SDF-1.
It is also elucidated that A-box of HMGB1 is responsible the function above. We have established the system for mass production of recombinant HMGB1 using cultured human cells.
In order to establish an efficient strategy to identify cardiomyogenic growth factors, we established a screening system in which adipose tissue-derived mesenchymal stem cells are used to detect a cardiomyogenic activity of soluble factors. Using this system, we have isolated several candidate molecules that may be used to induce cardiomyocyte differentiation of stem/progenitor cell populations.
3.Development of self-regenerative cardiovascular device
In this year, the molecular design of hydrogels is mainly focused by using gelatin of a biodegradable polymer clinically applicable for the controlled release of drugs and proteins. In particular, we prepared gelatin hydrogels with different physicochemical properties, which allow different drugs to efficiently incorporate into the hydrogel via the physicochemical interaction.
Gelatin hydrogels in different shapes of sponge and microsphere were fabricated as the scaffold for both the attachment of stem cells mobilized and the controlled release of growth factors.
4. Development for safety and efficacy evaluation
 We perform standardization of tissues by comprehensive gene expression analysis and detection of genomic alterations by comparative genomic hybridization (CGH) to validate safety index of regenerative tissue. We finished CGH analysis and are ready to detect genomic abnormalities by bio-informatics.
The slow-releasing form of ONO-1301 was injected directly into the canine DCM model. The slow-releasing form of ONO-1301 increased capillary density and ameliorated left ventricular enlargement. In conclusion, our results indicate that ONO-1301 up-regulated endogenous growth factors and promoted angiogenesis in response. Therefore ONO-1301 might have a therapeutic potential in treating heart failure.
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