Optimization of cardiomyocyte differentiation of human pluripotent stem cells
Haponen, Markus (2010)
Tampereen ammattikorkeakoulu
2010
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Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:amk-2010112215066
https://urn.fi/URN:NBN:fi:amk-2010112215066
Tiivistelmä
Human pluripotent stem cells can be differentiated into cardiomyocytes in vitro. Differentiated cardiomyocytes are at an important role in the future clinical applications of treating cardiac diseases. The purpose of this study was to optimize the cardiomyocyte differentiation of human pluripotent stem cells using four different culture media and to characterize the differentiated cardiomyocytes. The aim of the study was to find a best suited culture medium for cardiomyocyte differentiation.
Human embryonic stem cell line H7 and two induced pluripotent stem cell lines, h1/12 and 0/gG/25 were differentiated, and on day 22 the beating areas were counted. Cardiomyocytes were characterized by evaluating the beating rate and morphology, with immunocytochemical stainings and quantitative polymerase chain reaction (q-PCR).
The results show only small differences in the amount of beating areas acquired with each medium. 0% KO-SR hes medium gave the best result whereas the smallest amount of beating areas was acquired with Reges medium. The analysis with q-PCR showed positive gene expression of stem cell markers and high expression of cardiac markers in the differentiated cardiomyocytes with each tested medium and cell line. The highest gene expression in cardiomyocytes was observed in day 30 samples grown in 0%-medium.
According to the results, the best medium for the cardimyocyte differentiation would be 0% KO-SR hes medium. However, this medium should be studied even further with more different cell lines. Considering the q-PCR results, it would be interesting to study cardiomyocytes even older than 30 days, which are differentiated in 0% KO-SR medium.
Human embryonic stem cell line H7 and two induced pluripotent stem cell lines, h1/12 and 0/gG/25 were differentiated, and on day 22 the beating areas were counted. Cardiomyocytes were characterized by evaluating the beating rate and morphology, with immunocytochemical stainings and quantitative polymerase chain reaction (q-PCR).
The results show only small differences in the amount of beating areas acquired with each medium. 0% KO-SR hes medium gave the best result whereas the smallest amount of beating areas was acquired with Reges medium. The analysis with q-PCR showed positive gene expression of stem cell markers and high expression of cardiac markers in the differentiated cardiomyocytes with each tested medium and cell line. The highest gene expression in cardiomyocytes was observed in day 30 samples grown in 0%-medium.
According to the results, the best medium for the cardimyocyte differentiation would be 0% KO-SR hes medium. However, this medium should be studied even further with more different cell lines. Considering the q-PCR results, it would be interesting to study cardiomyocytes even older than 30 days, which are differentiated in 0% KO-SR medium.