title: Temporal impact of substrate mechanics on differentiation of human embryonic stem cells to cardiomyocytes.
abstract: A significant clinical need exists to differentiate human pluripotent stem cells (hPSCs) into cardiomyocytes, enabling tissue modeling for in vitro discovery of new drugs or cell-based therapies for heart repair in vivo. Chemical and mechanical microenvironmental factors are known to impact the efficiency of stem cell differentiation, but cardiac differentiation protocols in hPSCs are typically performed on rigid tissue culture polystyrene (TCPS) surfaces, which do not present a physiological mechanical setting. To investigate the temporal effects of mechanics on cardiac differentiation, we cultured human embryonic stem cells (hESCs) and their derivatives on polyacrylamide hydrogel substrates with a physiologically relevant range of stiffnesses. In directed differentiation and embryoid body culture systems, differentiation of hESCs to cardiac troponin T-expressing (cTnT+) cardiomyocytes peaked on hydrogels of intermediate stiffness. Brachyury expression also peaked on intermediate stiffness hydrogels at day 1 of directed differentiation, suggesting that stiffness impacted the initial differentiation trajectory of hESCs to mesendoderm. To investigate the impact of substrate mechanics during cardiac specification of mesodermal progenitors, we initiated directed cardiomyocyte differentiation on TCPS and transferred cells to hydrogels at the Nkx2.5/Isl1+ cardiac progenitor cell stage. No differences in cardiomyocyte purity with stiffness were observed on day 15. These experiments indicate that differentiation of hESCs is sensitive to substrate mechanics at early stages of mesodermal induction, and proper application of substrate mechanics can increase the propensity of hESCs to differentiate to cardiomyocytes.
authors.affiliation: Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Laurie B
authors.initials: LB
authors.lastname: Hazeltine
authors.affiliation: Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Mehmet G
authors.initials: MG
authors.lastname: Badur
authors.affiliation: Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Lian
authors.affiliation: Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Amritava
authors.initials: A
authors.lastname: Das
authors.affiliation: Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI 53706, USA.
authors.forename: Wenqing
authors.initials: W
authors.lastname: Han
authors.affiliation: Department of Chemical and Biological Engineering, University of Wisconsin-Madison, Madison, WI 53706, USA. Electronic address: palecek@engr.wisc.edu.
authors.forename: Sean P
authors.initials: SP
authors.lastname: Palecek
mesh_terms: D000180:Acrylic Resins / Q000737:chemistry* / Q000494:pharmacology*; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D048430:Cell Shape / Q000187:drug effects; D055119:Elastic Modulus / Q000187:drug effects; D058732:Embryoid Bodies / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D004707:Endoderm / Q000166:cytology / Q000187:drug effects; D005434:Flow Cytometry; D006801:Humans; D020100:Hydrogels / Q000737:chemistry / Q000494:pharmacology; D008648:Mesoderm / Q000166:cytology / Q000187:drug effects; D051379:Mice; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D054852:Small Molecule Libraries / Q000494:pharmacology; D013997:Time Factors; D020107:Troponin T / Q000378:metabolism; D000099166:Polyacrylamides
chemical_list: D000180:Acrylic Resins; D015415:Biomarkers; D020100:Hydrogels; D054852:Small Molecule Libraries; D020107:Troponin T; D000099166:Polyacrylamides
keywords: Cardiomyocytes; Differentiation; Human embryonic stem cells; Substrate mechanics