Nutrient supplemented serum-free medium increases cardiomyogenesis efficiency of human pluripotent stem cells

Table 1 List of nutritional supplements

Supplements	Concentration/addition	Source
MEM vitamin solution (× 100)	30 μL/mL	Gibco 11120
Yeastolate ultrafiltrate (× 50)	20 μL/mL	Gibco 18200
Vitamin E	70 μg/mL	Sigma T3376
Synthechol (cholesterol) (× 500)	12 μL/mL	Sigma S5442
Yeast extract	50 μg/mL	Sigma Y1625
Bovine serum albumin	50 μg/mL	Gibco A10008-01
Soy hydrolysate	50 μg/mL	Kerry Bio-science 5X59022

Table 2 Quantitative real-time reverse transcription-polymerase chain reaction primers

Primers for qPCR	Forward (5’→3’)	Reverse (5’→3’)
Nanog	GAAAAACAACTGGCCGAAGAAT	GGTGCTGAGGCCTTCTGC
4-Oct	AACGACCATCTGCCGCTTT	GGCCGCAGCTTACACATGTT
T-bra	AATTTGGTCCAGCCTTGGAAT	CGTTGCTCACAGACCACAG
Mesp 1	GACGTGCTGGCTCTGTTG	TGTCACTTGGGCTCCTCAG
Nkx 2.5	CAAGTGTGCGTCTGCCTTT	TTGTCCGCCTCTGTCTTCTC
MHC	ATTGCTGAAACCGAGAATGG	CGCTCCTTGAGGTTGAAAAG
GAPDH	GTCGGAGTCAACGGATTTGG	AAAAGCAGCCCTGGTGACC

qPCR: Quantitative polymerase chain reaction; MHC: Myosin heavy chain; GAPDH: Glyceraldehyde 3-phosphate dehydrogenase.

Table 3 Summary of results: Robustness of SupSB vs SB media for cell lines HES-3, H1 and Siu1-human induced pluripotent stem cells

Conditions		Cell expansion		Differentiation efficiency		CM yield
Cell type	Medium	Cell yield (106 cells/mL)	Cell expansion fold	Sarcomeric actinin	MHC	Cardiomyocyte/hESC
HES-3	SupSB	2.27 ± 0.58	1.65 ± 0.47	14.70% ± 6.30%	13.59% ± 3.40%	0.21 ± 0.08
	SB	1.66 ± 0.48	1.25 ± 0.36	9.25% ± 6.19%	6.17% ± 2.95%	0.07 ± 0.03
H1	SupSB	3.14 ± 0.24	2.35 ± 0.18	26.70% ± 2.09%	25.20% ± 5.41%	0.59 ± 0.01
	SB	1.10 ± 0.19	0.82 ± 0.14	20.23% ± 1.87%	4.76% ± 1.62%	0.04 ± 0.01
Siu1-hiPSC	SupSB	1.22 ± 0.15	0.92 ± 0.11	20.76% ± 4.74%	2.11% ± 0.73%	0.04 ± 0.04
	SB	0.67 ± 0.26	0.51 ± 0.20	16.90% ± 1.95%	1.01% ± 0.20%	0.004 ± 0.001

MHC: Myosin heavy chain. Cultures with SupSB medium were compared against SB medium for cell expansion and differentiation efficiency at the end of culture (day 16). The results were summarized by calculating the cardiomyocyte yield [cardiomyocytes (CM) yield] which takes into account the ratio of cardiomyocytes at day 16 compared to the initial human embryonic stem cells (hESC) seeded (n = 3 to 9).