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Combinatorial development of biomaterials for clonal growth of human pluripotent stem cells

Abstract

Both human embryonic stem cells and induced pluripotent stem cells can self-renew indefinitely in culture; however, present methods to clonally grow them are inefficient and poorly defined for genetic manipulation and therapeutic purposes. Here we develop the first chemically defined, xeno-free, feeder-free synthetic substrates to support robust self-renewal of fully dissociated human embryonic stem and induced pluripotent stem cells. Material properties including wettability, surface topography, surface chemistry and indentation elastic modulus of all polymeric substrates were quantified using high-throughput methods to develop structure–function relationships between material properties and biological performance. These analyses show that optimal human embryonic stem cell substrates are generated from monomers with high acrylate content, have a moderate wettability and employ integrin αvβ3 and αvβ5 engagement with adsorbed vitronectin to promote colony formation. The structure–function methodology employed herein provides a general framework for the combinatorial development of synthetic substrates for stem cell culture.

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Figure 1: High-throughput screening of biomaterials for clonal growth.
Figure 2: Diverse hES cell behaviour on primary polymer arrays.
Figure 3: Mapping hES cell behaviour to polymer properties using primary arrays.
Figure 4: Correlating hES cell behaviour to polymer properties using primary arrays.
Figure 5: Mapping cell behaviour to surface chemistry using secondary arrays.
Figure 6: Short- and long-term feeder-free culture on hit polymer arrays.

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Acknowledgements

We thank C. Beard, R. Alagappan, P. Xu, P. Wisniewski, C. Araneo, K. Wood, J. Daussman, R. Flannery, D. Fu, E. Luther and Compucyte for technical support. We thank all the members of the Langer laboratory and Jaenisch laboratory for helpful discussions and comments on the manuscript. R.J. was supported by NIH grants R37-CA084198, RO1-CA087869 and RO1-HD045022. R.L., R.J. and D.G.A. are advisors to Stemgent and R.L. and R.J. are cofounders of Fate Therapeutics. Financial support for J.Y. and A.H. is from the Wellcome Trust 085246. K.S. is supported by the Society in Science: the Branco Weiss Fellowship. D.G.A., R.L. and Y.M. are supported by NIH DE016516. Z.I.K. was supported by the US Army through the Institute for Soldier Nanotechnologies, under Contract W911NF-07-D-0004 with the US Army Research Office.

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All of the authors developed experiments, participated in the generation and analysis of data and assisted in the writing of the manuscript.

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Correspondence to Robert Langer, Rudolf Jaenisch or Daniel G. Anderson.

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Mei, Y., Saha, K., Bogatyrev, S. et al. Combinatorial development of biomaterials for clonal growth of human pluripotent stem cells. Nature Mater 9, 768–778 (2010). https://doi.org/10.1038/nmat2812

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