Abstract
Direct reprogramming strategies enable rapid conversion of somatic cells to cardiomyocytes or cardiomyocyte-like cells without going through the pluripotent state. A recently described protocol couples Yamanaka factor induction with pluripotency inhibition followed by BMP4 treatment to achieve rapid reprogramming of mouse fibroblasts to beating cardiomyocyte-like cells. The original study was performed using Matrigel-coated tissue culture polystyrene (TCPS), a stiff material that also non-specifically adsorbs serum proteins. Protein adsorption-resistant poly(ethylene glycol) (PEG) materials can be covalently modified to present precise concentrations of adhesion proteins or peptides without the unintended effects of non-specifically adsorbed proteins. Here, we describe an improved protocol that incorporates custom-engineered materials. We first reproduced the Efe etal. protocol on Matrigel-coated TCPS (the original material), reprogramming adult mouse tail-tip mouse fibroblasts (TTF) and mouse embryonic fibroblasts (MEF) to cardiomyocyte-like cells that demonstrated striated sarcomeric α-actinin staining, spontaneous calcium transients, and visible beating. We then designed poly(ethylene glycol) culture substrates to promote MEF adhesion via laminin and RGD-binding integrins. PEG hydrogels improved proliferation and reprogramming efficiency (evidenced by beating patch number and area, gene expression, and flow cytometry), yielding almost twice the number of sarcomeric α-actinin positive cardiomyocyte-like cells as the originally described substrate. These results illustrate that cellular reprogramming may be enhanced using custom-engineered materials.
Original language | English (US) |
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Pages (from-to) | 6559-6571 |
Number of pages | 13 |
Journal | Biomaterials |
Volume | 34 |
Issue number | 28 |
DOIs | |
State | Published - Sep 2013 |
Funding
The authors would like to thank Drs. Jeanne Nerbonne, Shelly Sakiyama-Elbert, Robert Mecham, and Guy Genin for helpful discussions relating to these results. We would also like to thank Mitchell Manar for cell culture assistance, and Dr. Casey Donahoe, Patrick Blanner, Dr. Scott Marrus, Dr. Nithya Jesuraj, Laura Marquardt, and Hao Xu for technical advice. This work was funded by an American Heart Association predoctoral fellowship 11PRE7690043 (AWS), the National Institutes of Health NIH R01 HL085364 (DLE), R01 HL085369 (IRE), the Lucy and Stanley Lopata endowment (IRE), the Career Award for Medical Scientists from the Burroughs Wellcome Fund (SR), and K08 HL107449 (SR).
Keywords
- Cardiomyocyte
- Direct reprogramming
- Laminin
- Poly(ethylene glycol)
- Stem cell microenvironment
- Stem cell niche
ASJC Scopus subject areas
- Biophysics
- Bioengineering
- Ceramics and Composites
- Biomaterials
- Mechanics of Materials