Recombinant human serum albumin promotes differentiation of human pluripotent stem cells into cardiomyocytes <i>in vitro</i> _Chinese Journal of Clinical Thoracic and Cardiovascular Surgery

Authors：

LIU Taoyan , CUI Wei , LI Hongxia , WU Fujian ,  LAN Feng

Beijing Institute of Heart, Lung and Blood Vessel Diseases, Beijing Anzhen Hospital, Capital Medical University, Beijing, 100029, P.R.China;

Corresponding?author：

LAN Feng, Email: fenglan@ccmu.edu.cn

Keywords：

Recombinant human serum albumin; human pluripotent stem cell; differentiation; cardiomyocyte; ultrastructure

DOI：

10.7507/1007-4848.201705038

Video：

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Objective To investigate whether recombinant human serum albumin (rHSA) can replace traditional B27 as a basic medium for differentiation of human pluripotent stem cells (hPSCs) into cardiomyocytes. Methods hPSCs were seeded at a cell density of 1.2×10⁴/cm²; until up to 75% confluency hPSCs were induced by differentiation medium containing various concentration of rHSA (0, 50, 100, 200 g/L). Light microscope and fluorescence microscope recorded the whole process of stem cells differentiating into myocardium. Flow cytometry was used to detect the cardiac differentiation efficiency at different concentrations of rHSA. Immunofluorescence staining was used to detect the cardiac specific protein α-actinin and troponin T (cTnT) and electron microscope to observe the ultrastructure of human pluripotent stem cell-derived cardiomyocytes (hPSC-CM) and beating rates of hPSC-CMs response to drugs. Results A large number of spontaneous beating cardiomyocytes were observed 9 days after induction and differentition. The percentage of colonies showing beating cardiomyocytes was 60.4% at the concentration of 200 g/L of rice derived-rHSA. Beating cardiomyocytes were α-actinin and cTnT positive. Ultrastructural analysis showed scattered sarcomeres and mitochondrial. hPSC-CMs were dose-dependent on isopropyl adrenaline and verapamil. Conclusion Using such simple media to differentiate hPSCs into functional cardiomyocytes is cost-effective and highly efficient, and can be used in the clinical research.

Citation： LIU Taoyan, CUI Wei, LI Hongxia, WU Fujian, LAN Feng. Recombinant human serum albumin promotes differentiation of human pluripotent stem cells into cardiomyocytes in vitro . Chinese Journal of Clinical Thoracic and Cardiovascular Surgery, 2018, 25(7): 604-609. doi: 10.7507/1007-4848.201705038 Copy

1.	Chong JJ, Yang X, Don CW, et al. Human embryonic-stem-cell-derived cardiomyocytes regenerate non-human primate hearts. Nature, 2014, 510(7504): 273-277.
2.	Wang Y, Liang P, Lan F, et al. Genome editing of isogenic human induced pluripotent stem cells recapitulates long QT phenotype for drug testing. J Am Coll Cardiol, 2014, 64(5): 451-459.
3.	Liang P, Lan F, Lee AS, et al. Drug screening using a library of human induced pluripotent stem cell-derived cardiomyocytes reveals disease-specific patterns of cardiotoxicity. Circulation, 2013, 127(16): 1677-1691.
4.	Lan F, Lee AS, Liang P, et al. Abnormal calcium handling properties underlie familial hypertrophic cardiomyopathy pathology in patient-specific induced pluripotent stem cells. Cell Stem Cell, 2013, 12(1): 101-113.
5.	Burridge PW, Keller G, Gold JD, et al. Production of de novo cardiomyocytes: human pluripotent stem cell differentiation and direct reprogramming. Cell Stem Cell, 2012, 10(1): 16-28.
6.	Lian X, Hsiao C, Wilson G, et al. Robust cardiomyocyte differentiation from human pluripotent stem cells via temporal modulation of canonical Wnt signaling. Proc Natl Acad Sci USA, 2012, 109(27): E1848-E1857.
7.	Kadari A, Mekala S, Wagner N, et al. Robust generation of cardiomyocytes from human iPS cells requires precise modulation of BMP and WNT signaling. Stem Cell Rev, 2015, 11(4): 560-569.
8.	Laflamme MA, Chen KY, Naumova AV, et al. Cardiomyocytes derived from human embryonic stem cells in pro-survival factors enhance function of infarcted rat hearts. Nat Biotechnol, 2007, 25(9): 1015-1024.
9.	Burridge PW, Matsa E, Shukla P, et al. Chemically defined generation of human cardiomyocytes. Nat Methods, 2014, 11(8): 855-860.
10.	Elliott DA, Braam SR, Koutsis K, et al. NKX2-5(eGFP/w) hESCs for isolation of human cardiac progenitors and cardiomyocytes. Nat Methods, 2011, 8(12): 1037-1040.
11.	Burridge PW, Thompson S, Millrod MA, et al. A universal system for highly efficient cardiac differentiation of human induced pluripotent stem cells that eliminates interline variability. PLoS One, 2011, 6(4): e18293.
12.	Cao N, Liu Z, Chen Z, et al. Ascorbic acid enhances the cardiac differentiation of induced pluripotent stem cells through promoting the proliferation of cardiac progenitor cells. Cell Res, 2012, 22(1): 219-236.
13.	Takahashi T, Lord B, Schulze PC, et al. Ascorbic acid enhances differentiation of embryonic stem cells into cardiac myocytes. Circulation, 2003, 107(14): 1912-1916.
14.	楊梁, 張曉剛, 魏新偉. 維生素C體外誘導多能干細胞向心肌細胞的分化. 中國組織工程研究與臨床康復, 2011, 15(23): 4229-4232.
15.	穆軍升, 李獻帥, 袁樹民, 等. 直接貼壁法誘導人胚胎干細胞分化為心肌細胞. 中國胸心血管外科臨床雜志, 2013, 20(5): 564-569.
16.	李獻帥, 袁樹民, 穆軍升, 等. 懸浮培養誘導人胚胎干細胞分化為心肌細胞的研究. 心肺血管病雜志, 2014, 33(1): 93-97.

1. Chong JJ, Yang X, Don CW, et al. Human embryonic-stem-cell-derived cardiomyocytes regenerate non-human primate hearts. Nature, 2014, 510(7504): 273-277.
2. Wang Y, Liang P, Lan F, et al. Genome editing of isogenic human induced pluripotent stem cells recapitulates long QT phenotype for drug testing. J Am Coll Cardiol, 2014, 64(5): 451-459.
3. Liang P, Lan F, Lee AS, et al. Drug screening using a library of human induced pluripotent stem cell-derived cardiomyocytes reveals disease-specific patterns of cardiotoxicity. Circulation, 2013, 127(16): 1677-1691.
4. Lan F, Lee AS, Liang P, et al. Abnormal calcium handling properties underlie familial hypertrophic cardiomyopathy pathology in patient-specific induced pluripotent stem cells. Cell Stem Cell, 2013, 12(1): 101-113.
5. Burridge PW, Keller G, Gold JD, et al. Production of de novo cardiomyocytes: human pluripotent stem cell differentiation and direct reprogramming. Cell Stem Cell, 2012, 10(1): 16-28.
6. Lian X, Hsiao C, Wilson G, et al. Robust cardiomyocyte differentiation from human pluripotent stem cells via temporal modulation of canonical Wnt signaling. Proc Natl Acad Sci USA, 2012, 109(27): E1848-E1857.
7. Kadari A, Mekala S, Wagner N, et al. Robust generation of cardiomyocytes from human iPS cells requires precise modulation of BMP and WNT signaling. Stem Cell Rev, 2015, 11(4): 560-569.
8. Laflamme MA, Chen KY, Naumova AV, et al. Cardiomyocytes derived from human embryonic stem cells in pro-survival factors enhance function of infarcted rat hearts. Nat Biotechnol, 2007, 25(9): 1015-1024.
9. Burridge PW, Matsa E, Shukla P, et al. Chemically defined generation of human cardiomyocytes. Nat Methods, 2014, 11(8): 855-860.
10. Elliott DA, Braam SR, Koutsis K, et al. NKX2-5(eGFP/w) hESCs for isolation of human cardiac progenitors and cardiomyocytes. Nat Methods, 2011, 8(12): 1037-1040.
11. Burridge PW, Thompson S, Millrod MA, et al. A universal system for highly efficient cardiac differentiation of human induced pluripotent stem cells that eliminates interline variability. PLoS One, 2011, 6(4): e18293.
12. Cao N, Liu Z, Chen Z, et al. Ascorbic acid enhances the cardiac differentiation of induced pluripotent stem cells through promoting the proliferation of cardiac progenitor cells. Cell Res, 2012, 22(1): 219-236.
13. Takahashi T, Lord B, Schulze PC, et al. Ascorbic acid enhances differentiation of embryonic stem cells into cardiac myocytes. Circulation, 2003, 107(14): 1912-1916.
14. 楊梁, 張曉剛, 魏新偉. 維生素C體外誘導多能干細胞向心肌細胞的分化. 中國組織工程研究與臨床康復, 2011, 15(23): 4229-4232.
15. 穆軍升, 李獻帥, 袁樹民, 等. 直接貼壁法誘導人胚胎干細胞分化為心肌細胞. 中國胸心血管外科臨床雜志, 2013, 20(5): 564-569.
16. 李獻帥, 袁樹民, 穆軍升, 等. 懸浮培養誘導人胚胎干細胞分化為心肌細胞的研究. 心肺血管病雜志, 2014, 33(1): 93-97.

Chinese Journal of Clinical Thoracic and Cardiovascular Surgery

Recombinant human serum albumin promotes differentiation of human pluripotent stem cells into cardiomyocytes in vitro

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