光造型技術と胎児肝細胞を用いた血管構造を持つin vitro肝組織再構築

利用立体光刻技术和胎儿肝细胞进行具有血管结构的体外肝组织重建

• 批准号: 02F00161
• 负责人: 酒井 康行
• 金额: $ 1.92万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for JSPS Fellows
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02F00161/
• 关键词: 三次元造型; 生体吸収性樹脂; 流路ネットワーク; シート積層造型法; 胎児肝細胞; 三次元培養; in vitro分化誘導; 肝組織移植; 微細流路ネットワーク; 肝組織再構築; マウス胎児肝細胞; 増殖; 成熟化

(1)複雑な内部構造を持つ生体吸収性樹脂の三次元微細造型,および,(2)胎児肝細胞の増殖と分化の制御,の2つの側面から検討を行った.(1)については,前年度までに確立したポリ乳酸多孔質シート積層・流路切削加工法によってマウスの全肝とほぼ同じ1.3mLの担体を作成し,それを用いてin vitroでヒト肝ガン由来Hep G2細胞の灌流培養を行った.その結果,マクロな流路を切削加工することで,細胞の増殖と機能発現を大きく向上させることを確かめた.(2)については,前年度までに確立した胎児マウス肝細胞の培養条件下(ニコチンアミド・DMSO・オンコスタチンMの三因子共存)でポリ乳酸三次元担体を用いて作成した組織を,70%肝切除同系マウスに移植,そのin vivoでの性能を評価した.その結果,2週間程度のin vitro分化誘導にて肝前駆細胞集団を格段に増幅していくことが,移植1ヵ月後の肝組織の再構築を大幅に改善することが明らかとなった.(2)については,確立したin vitro分化誘導条件のラット胎児肝細胞・購入した継代ヒト胎児肝細胞について,その適用可能性を調べた.その結果,ラットに関してはマウスと同様の培養条件が,同じくin vitroでの肝前駆細胞の分化誘導にとって極めて効果的であった.一方,ヒトについては,オンコスタチンMと三次元培養との組み合わせが有効であった.以上,マウスやラット胎児肝由来の肝前駆細胞を利用した新規in vitro再構築型肝組織開発にとって,重要な基礎的知見を得ることができた.

The main results are as follows: (1) the three-dimensional micromodeling of liposuction liposuction, (2) the control of the differentiation of fetal liver cells, and the control of the differentiation of fetal liver cells. (1) the production of lactic acid porous lactic acid in the previous year, the method of active flow path cutting, the whole liver and the same 1.3mL carrier. Hep G2 cells were perfused with in vitro to induce liver injury. According to the results of the experiment, the flow path was cut and machined, and the cellular colonization machine was able to make sure that it was successful. (2) in the previous year, under the condition of fetal liver cell culture (the coexistence of three factors), the triple carrier of lactic acid was used to form tissue tissue. 70% hepatectomy was performed in homologous liver transplantation, and in vivo was performed in patients with poor performance. The results showed that after 2 months of in vitro differentiation, the cell segment of pre-fetal fetal liver cell collection showed significant changes. One month after transplantation, the fetal liver tissue was significantly improved. (2) make sure that fetal in vitro differentiation leads to the inclusion of fetal fetal liver cells into fetal fetal liver cells. It is not possible to use the possibility. The results showed that the conditions were the same as that of in vitro, and the differentiation of prehepatic cells led to the positive effect of the results. On the other hand, there are some problems in the three-dimensional training system. Above, the fetal liver is caused by the fetal liver, and the pre-hepatic tissue is used to carry out the pregnancy by using the new in vitro liver tissue, and the knowledge of the important genes is important.

项目成果

Hanada S., Kayano H., Jiang J., Kojima N., Miyajima A., Sakoda A., Sakai Y.: "Enhanced in vitro maturation of subcultivated fetal human hepatocytes in three dimensional culture using poly-L-lactic acid scaffolds in the presence of oncostatin M"Internation

Hanada S.、Kayano H.、Jiang J.、Kojima N.、Miyajima A.、Sakoda A.、Sakai Y.：“使用聚 L-乳酸支架在三维培养中增强传代培养的胎儿人肝细胞的体外成熟

Y.Sakai, M.Otsuka, S.Hanada, Y.Nishiyama, Y.Konishi, A.Yamashita: "A novel poly-L-lactic acid scaffold that possesses a macroporous structure and a branching/joining three-dimensional flow channel network : Its fabrication and application to perfusion cul

Y.Sakai、M.Otsuka、S.Hanada、Y.Nishiyama、Y.Konishi、A.Yamashita：“一种新型聚左旋乳酸支架，具有大孔结构和分支/连接三维流动通道网络

J.Jinlan, N.Kojima, A.Miyajima, W.Yan, Y.Sakai: "Cultivation and induction of fetal liver cells in poly-L-lactic acid scaffolds"Materials Science and Engineering C. 24・3. 361-363 (2004)

J.Jinlan，N.Kojima，A.Miyajima，W.Yan，Y.Sakai：“聚L-乳酸支架中胎儿肝细胞的培养和诱导”材料科学与工程C. 24・3。 (2004)

立石哲也, 田中順三(分担執筆:酒井康行): "図解 再生医療工学(分担執筆,3.8.マイクロファブリケーション・三次元造型,PP.89-95)"工業調査会. 318 (2004)

Tetsuya Tateishi、Junzo Tanaka（合著者：Yasuyuki Sakai）：“再生医学工程图解（合著者，3.8. 微加工/三维建模，PP.89-95）”工业研究协会 318 (2004)。

J.Jinlan, N.Kojima, A.Miyajima, W Yan, Y.Sakai: "Cultivation and induction of fetal liver cells in poly-L-lactic acid scaffolds"Material Science and Engineering C. (印刷中). (2003)

J.Jinlan，N.Kojima，A.Miyajima，W Yan，Y.Sakai：“聚L-乳酸支架中胎儿肝细胞的培养和诱导”材料科学与工程C.（出版中）。