Table 1.
Selection of hiPSC lines regarding their expansion compatibility and cardiac differentiation efficiency in MC cultures. (A) Measurement of the highest flow cytometry population percentage of NKX2-5, Troponin T, MLC2a, CD44 and HNF4a on day 14 with a 24 h CHIR99021 induction of 4–14 μM on 5 cell lines in a monolayer attachment culture (n = 3). (B) Effects of continuous stirring on cell lines in MC-based spinner flask cultures. Cell yield, aggregate size and the expression of pluripotent (Oct4a and Tra-1-60) were measured (n = 3). (C) Selection of the best cell line for the cardiac differentiation on MC in a stirred spinner. Cell yield and the expression of cardiac marker Troponin T were measured (n = 3)
| DF6 | CB6 | BM1 | IMR90 | FR202 | |
|---|---|---|---|---|---|
| (A) Cardiac differentiation efficiency with CHIR99021 in MNL cultures (Maximum flow cytometry population expression at 4-14 μM CHIR99021 on day14) | |||||
| NKX2–5 (%) | 21.8 ± 17.1 | 1.7 ± 0.1 | 78.8 ± 25.5 | 82.9 ± 8.4 | 57.1 ± 7.2 |
| Troponin T (%) | 29.7 ± 24.6 | 2.1 ± 0.4 | 81.0 ± 31.2 | 83.1 ± 8.9 | 80.6 ± 2.1 |
| MLC2a (%) | 34.9 ± 25.7 | 1.95 ± 0.3 | 70.4 ± 21.9 | 64.9 ± 0.1 | 64.9 ± 9.4 |
| CD44 (%) | 40.5 ± 9.2 | 32.1 ± 8.4 | 16.5 ± 11.7 | 37.1 ± 14.9 | 3.7 ± 3.7 |
| HNF4a (%) | 38.8 ± 14.8 | 7.4 ± 1.9 | 13.6 ± 1.5 | 20.7 ± 5.9 | 4.4 ± 4.4 |
| (B) Cell growth on MC in stirred spinner cultures (day 6) | |||||
| Cells/mL (×106) | No cell growth | 1.7 ± 0.3 | 1.9 ± 0.6 | ||
| Expansion fold | 14.0 ± 0.2 | 16.0 ± 0.5 | |||
| Aggregate size (mm2) | 0.42 ± 0.1 | 0.30 ± 0.1 | |||
| Oct4a | 94.3 ± 1.1 | 91.0 ± 0.1 | |||
| Tra-1-60 | 93.0 ± 0.01 | 96.4 ± 0.1 | |||
| (C) Cardiac differentiation on MC in stirred spinner cultures (day 12 of differentiation) | |||||
| Cells/mL (×106) | 2.1 ± 0.4 | 2.3 ± 0.2 | |||
| Troponin T (%) | 14.4 ± 8.5 | 83.2 ± 0.13 | |||
| CM yields (cells/mL × 106) | 0.32 ± 0.2 | 1.9 ± 0.03 | |||