光駆動型分化に向けた金ナノ粒子含有ヒドロゲルへの胚様体包接

2013 Fiscal Year Annual Research Report

• Project/Area Number: 12F02388
• Research Institution: Kyoto University
• Principal Investigator: 中辻 憲夫 京都大学, 物質―細胞統合システム拠点, 教授
• Co-Investigator(Kenkyū-buntansha): WANG Lin 京都大学, 物質―細胞統合システム拠点, 外国人特別研究員
• Project Period (FY): 2012 – 2014-03-31
• Keywords: iPS cells / Controlled differentiation / Algniate 2D matrix

Research Abstract

The goal of this project is to develop a 3D spherical core-shell structure hydrogel-based induced pluripotent stem (iPS) cell culture platform for controlled iPS propagation and diflerentiation. Our system allows the in situ generation of embryo body (EB) at the center (core) of hydrogel beads and the control of extracellular environment by adjusting alginate inner core composition, in such way todirect the differentiation of EB in a controlled, and in-vivo mimic manner. This proposed system is expected to become a more effective, efficient, and biomimetic platform for iPS cell differentiation, which might ultimately benefit the regenerative medicine research community, and serve as a valuabletool for developmental biologist for better understanding of the development of embryo.
ln 2013 fiscal year, two folds of studies have been carried out : 1) development of modified hydrogel matrixes for iPS cells encapsulation, which allow controlling EB formation and differentiation, 2) characteri ze EB formation, cell Proliferation and differentiation.
Specifically, for part 1, we have achieved synthesizing modified alginate gel by covalently attaching oxtracollular matrix (ECM) protein motifs such as RGD and YIGSR peptide to the chain of alginate molecules. We were able to successfully got core shell structure hydrogel beads with modified inner core composition. For the second part, we have found that by changing the inner core composition, we were able to control the size of EB fomed inside our beads. Cells inside 0.5% alginate core formed the biggest~600um (diameter) EBs and cells inside 1% alginate mixed with 0.5%HA at 1:1 ratio core formed smallest~250 um (diameter) EBs. The inner core liquefying and modification can affect EB differentiation morphology and trajectory. PCR results suggested that liquefying inner core enhanced EB differentiation towards endodermal lineage, decreasing alginate concentration from 1 percent to 0.5 peroent suppressed hiPS cell differentiation, and modifying alginate with RGD peptide induced differentiation towards ectodermal lineage.

Strategy for Future Research Activity

We have completed this project by Dec. 15th of 2013.