Extracellular matrix promotes highly efficient cardiac differentiation of human pluripotent stem cells

The matrix sandwich method

Jianhua Zhang, Matthew Klos, Gisela F. Wilson, Amanda M. Herman, Xiaojun Lian, Kunil K. Raval, Matthew R. Barron, Luqia Hou, Andrew G. Soerens, Junying Yu, Sean P. Palecek, Gary E. Lyons, James A. Thomson, Todd J. Herron, José Jalife, Timothy J. Kamp

Research output: Contribution to journalArticle

242 Citations (Scopus)

Abstract

RATIONALE: Cardiomyocytes (CMs) differentiated from human pluripotent stem cells (PSCs) are increasingly being used for cardiovascular research, including disease modeling, and hold promise for clinical applications. Current cardiac differentiation protocols exhibit variable success across different PSC lines and are primarily based on the application of growth factors. However, extracellular matrix is also fundamentally involved in cardiac development from the earliest morphogenetic events, such as gastrulation. OBJECTIVE: We sought to develop a more effective protocol for cardiac differentiation of human PSCs by using extracellular matrix in combination with growth factors known to promote cardiogenesis. METHODS AND RESULTS: PSCs were cultured as monolayers on Matrigel, an extracellular matrix preparation, and subsequently overlayed with Matrigel. The matrix sandwich promoted an epithelial-to-mesenchymal transition as in gastrulation with the generation of N-cadherin-positive mesenchymal cells. Combining the matrix sandwich with sequential application of growth factors (Activin A, bone morphogenetic protein 4, and basic fibroblast growth factor) generated CMs with high purity (up to 98%) and yield (up to 11 CMs/input PSC) from multiple PSC lines. The resulting CMs progressively matured over 30 days in culture based on myofilament expression pattern and mitotic activity. Action potentials typical of embryonic nodal, atrial, and ventricular CMs were observed, and monolayers of electrically coupled CMs modeled cardiac tissue and basic arrhythmia mechanisms. CONCLUSIONS: Dynamic extracellular matrix application promoted epithelial-mesenchymal transition of human PSCs and complemented growth factor signaling to enable robust cardiac differentiation.

Original languageEnglish (US)
Pages (from-to)1125-1136
Number of pages12
JournalCirculation research
Volume111
Issue number9
DOIs
StatePublished - Oct 12 2012

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Pluripotent Stem Cells
Extracellular Matrix
Cardiac Myocytes
Intercellular Signaling Peptides and Proteins
Gastrulation
Epithelial-Mesenchymal Transition
Bone Morphogenetic Protein 4
Cell Line
Myofibrils
Fibroblast Growth Factor 2
Cadherins
Action Potentials
Cardiac Arrhythmias
Research

All Science Journal Classification (ASJC) codes

  • Physiology
  • Cardiology and Cardiovascular Medicine

Cite this

Zhang, Jianhua ; Klos, Matthew ; Wilson, Gisela F. ; Herman, Amanda M. ; Lian, Xiaojun ; Raval, Kunil K. ; Barron, Matthew R. ; Hou, Luqia ; Soerens, Andrew G. ; Yu, Junying ; Palecek, Sean P. ; Lyons, Gary E. ; Thomson, James A. ; Herron, Todd J. ; Jalife, José ; Kamp, Timothy J. / Extracellular matrix promotes highly efficient cardiac differentiation of human pluripotent stem cells : The matrix sandwich method. In: Circulation research. 2012 ; Vol. 111, No. 9. pp. 1125-1136.
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title = "Extracellular matrix promotes highly efficient cardiac differentiation of human pluripotent stem cells: The matrix sandwich method",
abstract = "RATIONALE: Cardiomyocytes (CMs) differentiated from human pluripotent stem cells (PSCs) are increasingly being used for cardiovascular research, including disease modeling, and hold promise for clinical applications. Current cardiac differentiation protocols exhibit variable success across different PSC lines and are primarily based on the application of growth factors. However, extracellular matrix is also fundamentally involved in cardiac development from the earliest morphogenetic events, such as gastrulation. OBJECTIVE: We sought to develop a more effective protocol for cardiac differentiation of human PSCs by using extracellular matrix in combination with growth factors known to promote cardiogenesis. METHODS AND RESULTS: PSCs were cultured as monolayers on Matrigel, an extracellular matrix preparation, and subsequently overlayed with Matrigel. The matrix sandwich promoted an epithelial-to-mesenchymal transition as in gastrulation with the generation of N-cadherin-positive mesenchymal cells. Combining the matrix sandwich with sequential application of growth factors (Activin A, bone morphogenetic protein 4, and basic fibroblast growth factor) generated CMs with high purity (up to 98{\%}) and yield (up to 11 CMs/input PSC) from multiple PSC lines. The resulting CMs progressively matured over 30 days in culture based on myofilament expression pattern and mitotic activity. Action potentials typical of embryonic nodal, atrial, and ventricular CMs were observed, and monolayers of electrically coupled CMs modeled cardiac tissue and basic arrhythmia mechanisms. CONCLUSIONS: Dynamic extracellular matrix application promoted epithelial-mesenchymal transition of human PSCs and complemented growth factor signaling to enable robust cardiac differentiation.",
author = "Jianhua Zhang and Matthew Klos and Wilson, {Gisela F.} and Herman, {Amanda M.} and Xiaojun Lian and Raval, {Kunil K.} and Barron, {Matthew R.} and Luqia Hou and Soerens, {Andrew G.} and Junying Yu and Palecek, {Sean P.} and Lyons, {Gary E.} and Thomson, {James A.} and Herron, {Todd J.} and Jos{\'e} Jalife and Kamp, {Timothy J.}",
year = "2012",
month = "10",
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doi = "10.1161/CIRCRESAHA.112.273144",
language = "English (US)",
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Zhang, J, Klos, M, Wilson, GF, Herman, AM, Lian, X, Raval, KK, Barron, MR, Hou, L, Soerens, AG, Yu, J, Palecek, SP, Lyons, GE, Thomson, JA, Herron, TJ, Jalife, J & Kamp, TJ 2012, 'Extracellular matrix promotes highly efficient cardiac differentiation of human pluripotent stem cells: The matrix sandwich method', Circulation research, vol. 111, no. 9, pp. 1125-1136. https://doi.org/10.1161/CIRCRESAHA.112.273144

Extracellular matrix promotes highly efficient cardiac differentiation of human pluripotent stem cells : The matrix sandwich method. / Zhang, Jianhua; Klos, Matthew; Wilson, Gisela F.; Herman, Amanda M.; Lian, Xiaojun; Raval, Kunil K.; Barron, Matthew R.; Hou, Luqia; Soerens, Andrew G.; Yu, Junying; Palecek, Sean P.; Lyons, Gary E.; Thomson, James A.; Herron, Todd J.; Jalife, José; Kamp, Timothy J.

In: Circulation research, Vol. 111, No. 9, 12.10.2012, p. 1125-1136.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Extracellular matrix promotes highly efficient cardiac differentiation of human pluripotent stem cells

T2 - The matrix sandwich method

AU - Zhang, Jianhua

AU - Klos, Matthew

AU - Wilson, Gisela F.

AU - Herman, Amanda M.

AU - Lian, Xiaojun

AU - Raval, Kunil K.

AU - Barron, Matthew R.

AU - Hou, Luqia

AU - Soerens, Andrew G.

AU - Yu, Junying

AU - Palecek, Sean P.

AU - Lyons, Gary E.

AU - Thomson, James A.

AU - Herron, Todd J.

AU - Jalife, José

AU - Kamp, Timothy J.

PY - 2012/10/12

Y1 - 2012/10/12

N2 - RATIONALE: Cardiomyocytes (CMs) differentiated from human pluripotent stem cells (PSCs) are increasingly being used for cardiovascular research, including disease modeling, and hold promise for clinical applications. Current cardiac differentiation protocols exhibit variable success across different PSC lines and are primarily based on the application of growth factors. However, extracellular matrix is also fundamentally involved in cardiac development from the earliest morphogenetic events, such as gastrulation. OBJECTIVE: We sought to develop a more effective protocol for cardiac differentiation of human PSCs by using extracellular matrix in combination with growth factors known to promote cardiogenesis. METHODS AND RESULTS: PSCs were cultured as monolayers on Matrigel, an extracellular matrix preparation, and subsequently overlayed with Matrigel. The matrix sandwich promoted an epithelial-to-mesenchymal transition as in gastrulation with the generation of N-cadherin-positive mesenchymal cells. Combining the matrix sandwich with sequential application of growth factors (Activin A, bone morphogenetic protein 4, and basic fibroblast growth factor) generated CMs with high purity (up to 98%) and yield (up to 11 CMs/input PSC) from multiple PSC lines. The resulting CMs progressively matured over 30 days in culture based on myofilament expression pattern and mitotic activity. Action potentials typical of embryonic nodal, atrial, and ventricular CMs were observed, and monolayers of electrically coupled CMs modeled cardiac tissue and basic arrhythmia mechanisms. CONCLUSIONS: Dynamic extracellular matrix application promoted epithelial-mesenchymal transition of human PSCs and complemented growth factor signaling to enable robust cardiac differentiation.

AB - RATIONALE: Cardiomyocytes (CMs) differentiated from human pluripotent stem cells (PSCs) are increasingly being used for cardiovascular research, including disease modeling, and hold promise for clinical applications. Current cardiac differentiation protocols exhibit variable success across different PSC lines and are primarily based on the application of growth factors. However, extracellular matrix is also fundamentally involved in cardiac development from the earliest morphogenetic events, such as gastrulation. OBJECTIVE: We sought to develop a more effective protocol for cardiac differentiation of human PSCs by using extracellular matrix in combination with growth factors known to promote cardiogenesis. METHODS AND RESULTS: PSCs were cultured as monolayers on Matrigel, an extracellular matrix preparation, and subsequently overlayed with Matrigel. The matrix sandwich promoted an epithelial-to-mesenchymal transition as in gastrulation with the generation of N-cadherin-positive mesenchymal cells. Combining the matrix sandwich with sequential application of growth factors (Activin A, bone morphogenetic protein 4, and basic fibroblast growth factor) generated CMs with high purity (up to 98%) and yield (up to 11 CMs/input PSC) from multiple PSC lines. The resulting CMs progressively matured over 30 days in culture based on myofilament expression pattern and mitotic activity. Action potentials typical of embryonic nodal, atrial, and ventricular CMs were observed, and monolayers of electrically coupled CMs modeled cardiac tissue and basic arrhythmia mechanisms. CONCLUSIONS: Dynamic extracellular matrix application promoted epithelial-mesenchymal transition of human PSCs and complemented growth factor signaling to enable robust cardiac differentiation.

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U2 - 10.1161/CIRCRESAHA.112.273144

DO - 10.1161/CIRCRESAHA.112.273144

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