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Efficient Derivation of Human Cardiac Precursors and Cardiomyocytes from Pluripotent Human Embryonic Stem Cells with Small Molecule Induction
Published on: November 3, 2011
Cardiomyocyte enrichment from human embryonic stem cell cultures by selection of ALCAM surface expression
William Rust1, Thavamalar Balakrishnan, Robert Zweigerdt
1Lonza Walkersville, Inc., 8830 Biggs Ford Road, Walkersville, MD 21793, USA. william.rust@lonza.com
Insights
Researchers identified activated leukocyte cell-adhesion molecule (ALCAM) as a marker to isolate human cardiomyocytes from stem cells. This method enables expansion of these cells for potential cardiac tissue regeneration.
Area of Science:
- Cardiovascular Biology
- Stem Cell Research
- Regenerative Medicine
Background:
- Cardiac disease and injury lead to significant loss of heart tissue.
- Developing methods for generating expandable human cardiomyocyte populations is crucial for tissue replacement therapies.
Purpose of the Study:
- To evaluate activated leukocyte cell-adhesion molecule (ALCAM) as a marker for isolating cardiomyocytes from differentiating human embryonic stem cells (hESCs).
- To assess the developmental stage and in vitro expansion potential of ALCAM-selected cardiomyocytes.
Main Methods:
- Human embryonic stem cells (hESCs) were differentiated into cardiomyocytes.
- Activated leukocyte cell-adhesion molecule (ALCAM) expression was analyzed using RT-qPCR and immunohistochemistry.
- DNA methylation patterns were studied to determine the developmental age of hESC-derived cardiomyocytes.
Main Results:
- hESC-derived cardiomyocytes express ALCAM, enabling selection of a cardiomyocyte-enriched population.
- ALCAM-selected cardiomyocytes exhibit an embryonic phenotype, indicated by contractile protein and ion channel expression, and DNA methylation patterns.
- Selected cardiomyocytes survived sorting, adhered to culture surfaces, and proliferated in short-term cultures.
Conclusions:
- ALCAM is a viable surface marker for isolating and enriching cardiomyocytes from hESC cultures.
- ALCAM-selected cardiomyocytes possess an embryonic phenotype and can be expanded in vitro.
- Long-term in vitro survival and proliferation of cardiomyocytes were achieved using 3D aggregate cultures.
Aims:
The production of a homogenous population of human cardiomyocytes that can be expanded in vitro may facilitate development of replacement tissue lost as a result of cardiac disease and injury.
Materials And Methods:
We evaluated the utility of activated leukocyte cell-adhesion molecule, CD166 (ALCAM) expression as a marker for isolating cardiomyocytes from differentiating cultures of human embryonic stem cells (hESCs). Using RT-qPCR, immunohistochemistry and DNA methylation studies, we evaluated the developmental age of hESC-derived cardiomyocytes.
Results And Conclusions:
We demonstrate that cardiomyocytes derived from hESC cultures express ALCAM and that this surface antigen can be used to select a population of differentiated cells that are enriched for cardiomyocytes. Expression of contractile proteins and ion channels, and DNA methylation patterns, suggest that ALCAM-enriched cardiomyocytes have an embryonic phenotype. Selected cardiomyocyte populations survive sorting, adhere to collagen-coated tissue culture plastic and proliferate in short-term culture. Long-term in vitro survival of cardiomyocytes was achieved by culturing cells in 3D aggregates.
