Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques.
Human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes are a valuable resource for cardiac therapeutic development; however, generation of these cells in large numbers and high purity is a limitation in widespread adoption. Here, design of experiments (DOE) is used to investigate the car...
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2020-01-01
|
| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0230001 |
| _version_ | 1818891817054109696 |
|---|---|
| author | Cassady E Rupert Chinedu Irofuala Kareen L K Coulombe |
| author_facet | Cassady E Rupert Chinedu Irofuala Kareen L K Coulombe |
| author_sort | Cassady E Rupert |
| collection | DOAJ |
| description | Human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes are a valuable resource for cardiac therapeutic development; however, generation of these cells in large numbers and high purity is a limitation in widespread adoption. Here, design of experiments (DOE) is used to investigate the cardiac differentiation space of three hiPSC lines when varying CHIR99027 concentration and cell seeding density, and a novel image analysis is developed to evaluate plate coverage when initiating differentiation. Metabolic selection via lactate purifies hiPSC-cardiomyocyte populations, and the bioenergetic phenotype and engineered tissue mechanics of purified and unpurified hiPSC-cardiomyocytes are compared. Findings demonstrate that when initiating differentiation one day after hiPSC plating, low (3 μM) Chiron and 72 x 103 cells/cm2 seeding density result in peak cardiac purity (50-90%) for all three hiPSC lines. Our results confirm that metabolic selection with lactate shifts hiPSC-cardiomyocyte metabolism towards oxidative phosphorylation, but this more "mature" metabolic phenotype does not by itself result in a more mature contractile phenotype in engineered cardiac tissues at one week of culture in 3D tissues. This study provides widely adaptable methods including novel image analysis code and parameters for refining hiPSC-cardiomyocyte differentiation and describes the practical implications of metabolic selection of cardiomyocytes for downstream tissue engineering applications. |
| first_indexed | 2024-12-19T17:46:50Z |
| format | Article |
| id | doaj.art-29218fb74be64648bc75160a126e189c |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-19T17:46:50Z |
| publishDate | 2020-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-29218fb74be64648bc75160a126e189c2022-12-21T20:12:02ZengPublic Library of Science (PLoS)PLoS ONE1932-62032020-01-01153e023000110.1371/journal.pone.0230001Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques.Cassady E RupertChinedu IrofualaKareen L K CoulombeHuman induced pluripotent stem cell (hiPSC)-derived cardiomyocytes are a valuable resource for cardiac therapeutic development; however, generation of these cells in large numbers and high purity is a limitation in widespread adoption. Here, design of experiments (DOE) is used to investigate the cardiac differentiation space of three hiPSC lines when varying CHIR99027 concentration and cell seeding density, and a novel image analysis is developed to evaluate plate coverage when initiating differentiation. Metabolic selection via lactate purifies hiPSC-cardiomyocyte populations, and the bioenergetic phenotype and engineered tissue mechanics of purified and unpurified hiPSC-cardiomyocytes are compared. Findings demonstrate that when initiating differentiation one day after hiPSC plating, low (3 μM) Chiron and 72 x 103 cells/cm2 seeding density result in peak cardiac purity (50-90%) for all three hiPSC lines. Our results confirm that metabolic selection with lactate shifts hiPSC-cardiomyocyte metabolism towards oxidative phosphorylation, but this more "mature" metabolic phenotype does not by itself result in a more mature contractile phenotype in engineered cardiac tissues at one week of culture in 3D tissues. This study provides widely adaptable methods including novel image analysis code and parameters for refining hiPSC-cardiomyocyte differentiation and describes the practical implications of metabolic selection of cardiomyocytes for downstream tissue engineering applications.https://doi.org/10.1371/journal.pone.0230001 |
| spellingShingle | Cassady E Rupert Chinedu Irofuala Kareen L K Coulombe Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques. PLoS ONE |
| title | Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques. |
| title_full | Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques. |
| title_fullStr | Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques. |
| title_full_unstemmed | Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques. |
| title_short | Practical adoption of state-of-the-art hiPSC-cardiomyocyte differentiation techniques. |
| title_sort | practical adoption of state of the art hipsc cardiomyocyte differentiation techniques |
| url | https://doi.org/10.1371/journal.pone.0230001 |
| work_keys_str_mv | AT cassadyerupert practicaladoptionofstateofthearthipsccardiomyocytedifferentiationtechniques AT chineduirofuala practicaladoptionofstateofthearthipsccardiomyocytedifferentiationtechniques AT kareenlkcoulombe practicaladoptionofstateofthearthipsccardiomyocytedifferentiationtechniques |