The acceleration of cardiomyogenesis in embryonic stem cells in
vitro by serum depletion does not increase the number of
developed cardiomyocytes

J 2017

The acceleration of cardiomyogenesis in embryonic stem cells in vitro by serum depletion does not increase the number of developed cardiomyocytes

RADASZKIEWICZ, Katarzyna Anna, Dominika SÝKOROVÁ, Lucia BINÓ, Jana KUDOVÁ, Markéta BÉBAROVÁ et. al.

Basic information

Original name

The acceleration of cardiomyogenesis in embryonic stem cells in vitro by serum depletion does not increase the number of developed cardiomyocytes

Authors

RADASZKIEWICZ, Katarzyna Anna (616 Poland, belonging to the institution), Dominika SÝKOROVÁ (203 Czech Republic, belonging to the institution), Lucia BINÓ (703 Slovakia, belonging to the institution), Jana KUDOVÁ (203 Czech Republic, belonging to the institution), Markéta BÉBAROVÁ (203 Czech Republic, belonging to the institution), Jiřina PROCHÁZKOVÁ (203 Czech Republic), Hana KOTASOVÁ (203 Czech Republic, belonging to the institution), Lukáš KUBALA (203 Czech Republic) and Jiří PACHERNÍK (203 Czech Republic, guarantor, belonging to the institution)

Edition

Plos ONE, San Francisco, Public Library of Science, 2017, 1932-6203

Other information

Language

English

Type of outcome

Článek v odborném periodiku

Field of Study

10601 Cell biology

Country of publisher

United States of America

Confidentiality degree

není předmětem státního či obchodního tajemství

References:

URL

Impact factor

Impact factor: 2.766

RIV identification code

RIV/00216224:14310/17:00096334

Organization unit

Faculty of Science

DOI

http://dx.doi.org/10.1371/journal.pone.0173140

UT WoS

000396092400008

Keywords in English

NEURAL DIFFERENTIATION; RETINOIC ACID; CARCINOMA-CELLS; DEFINED MEDIUM; EXPRESSION; DEPRIVATION; GENERATION; MONOLAYER; MESODERM; REPORTER

Abstract

V originále

The differentiation of pluripotent embryonic stem (ES) cells into various lineages in vitro represents an important tool for studying the mechanisms underlying mammalian embryogenesis. It is a key technique in studies evaluating the molecular mechanisms of cardiomyogenesis and heart development and also in embryotoxicology. Herein, modest modifications of the basic protocol for ES cell differentiation into cardiomyocytes were evaluated in order to increase the yield and differentiation status of developed cardiomyocytes. Primarily, the data show that ES cell cultivation in the form of non-adherent embryoid bodies (EBs) for 5 days compared to 8 days significantly improved cardiomyogenic differentiation. This is illustrated by the appearance of beating foci in the adherent EBs layer at earlier phases of differentiation from day 10 up to day 16 and by the significantly higher expression of genes characteristic of cardiomyogenic differentiation (sarcomeric alpha actinin, myosin heavy chain alpha and beta, myosin light chain 2 and 7, and transcriptional factor Nkx2.5) in EBs cultivated under non-adherent conditions for 5 days. The ratio of cardiomyocytes per other cells was also potentiated in EBs cultivated in non-adherent conditions for only 5 days followed by cultivation in adherent serum-free culture conditions. Nevertheless, the alteration in the percentage of beating foci among these two tested cultivation conditions vanished at later phases and also did not affect the total number of cardiomyocytes determined as myosin heavy chain positive cells at the end of the differentiation process on day 20. Thus, although these modifications of the conditions of ES cells differentiation may intensify cardiomyocyte differentiation, the final count of cardiomyocytes might not change. Thus, serum depletion was identified as a key factor that intensified cardiomyogenesis. Further, the treatment of EBs with N-acetylcysteine, a reactive oxygen species scavenger, did not affect the observed increase in cardiomyogenesis under serum depleted conditions. Interestingly, a mild induction of the ventricular-like phenotype of cardiomyocytes was observed in 5-day-old EBs compared to 8-day-old EBs. Overall, these findings bring crucial information on the mechanisms of ES cells differentiation into cardiomyocytes and on the establishment of efficient protocols for the cardiomyogenic differentiation of ES cells. Further, the importance of determining the absolute number of formed cardiomyocyte-like cells per seeded pluripotent cells in contrast to the simple quantification of the ratios of cells is highlighted.

Citovat

RADASZKIEWICZ, Katarzyna Anna, Dominika SÝKOROVÁ, Lucia BINÓ, Jana KUDOVÁ, Markéta BÉBAROVÁ, Jiřina PROCHÁZKOVÁ, Hana KOTASOVÁ, Lukáš KUBALA and Jiří PACHERNÍK. The acceleration of cardiomyogenesis in embryonic stem cells in vitro by serum depletion does not increase the number of developed cardiomyocytes. Plos ONE. San Francisco: Public Library of Science, 2017, vol. 12, No 3, p. nestránkováno, 17 pp. ISSN 1932-6203. Available from: https://dx.doi.org/10.1371/journal.pone.0173140.

@article{1376215,
   author = {Radaszkiewicz, Katarzyna Anna and Sýkorová, Dominika and Binó, Lucia and Kudová, Jana and Bébarová, Markéta and Procházková, Jiřina and Kotasová, Hana and Kubala, Lukáš and Pacherník, Jiří},
   article_location = {San Francisco},
   article_number = {3},
   doi = {http://dx.doi.org/10.1371/journal.pone.0173140},
   keywords = {NEURAL DIFFERENTIATION; RETINOIC ACID; CARCINOMA-CELLS; DEFINED MEDIUM; EXPRESSION; DEPRIVATION; GENERATION; MONOLAYER; MESODERM; REPORTER},
   language = {eng},
   issn = {1932-6203},
   journal = {Plos ONE},
   title = {The acceleration of cardiomyogenesis in embryonic stem cells in vitro by serum depletion does not increase the number of developed cardiomyocytes},
   url = {http://dx.plos.org/10.1371/journal.pone.0173140},
   volume = {12},
   year = {2017}
}

TY  - JOUR
ID  - 1376215
AU  - Radaszkiewicz, Katarzyna Anna - Sýkorová, Dominika - Binó, Lucia - Kudová, Jana - Bébarová, Markéta - Procházková, Jiřina - Kotasová, Hana - Kubala, Lukáš - Pacherník, Jiří
PY  - 2017
TI  - The acceleration of cardiomyogenesis in embryonic stem cells in vitro by serum depletion does not increase the number of developed cardiomyocytes
JF  - Plos ONE
VL  - 12
IS  - 3
SP  - nestránkováno
EP  - nestránkováno
PB  - Public Library of Science
SN  - 19326203
KW  - NEURAL DIFFERENTIATION
KW  - RETINOIC ACID
KW  - CARCINOMA-CELLS
KW  - DEFINED MEDIUM
KW  - EXPRESSION
KW  - DEPRIVATION
KW  - GENERATION
KW  - MONOLAYER
KW  - MESODERM
KW  - REPORTER
UR  - http://dx.plos.org/10.1371/journal.pone.0173140
L2  - http://dx.plos.org/10.1371/journal.pone.0173140
N2  - The differentiation of pluripotent embryonic stem (ES) cells into various lineages in vitro represents an important tool for studying the mechanisms underlying mammalian embryogenesis. It is a key technique in studies evaluating the molecular mechanisms of cardiomyogenesis and heart development and also in embryotoxicology. Herein, modest modifications of the basic protocol for ES cell differentiation into cardiomyocytes were evaluated in order to increase the yield and differentiation status of developed cardiomyocytes. Primarily, the data show that ES cell cultivation in the form of non-adherent embryoid bodies (EBs) for 5 days compared to 8 days significantly improved cardiomyogenic differentiation. This is illustrated by the appearance of beating foci in the adherent EBs layer at earlier phases of differentiation from day 10 up to day 16 and by the significantly higher expression of genes characteristic of cardiomyogenic differentiation (sarcomeric alpha actinin, myosin heavy chain alpha and beta, myosin light chain 2 and 7, and transcriptional factor Nkx2.5) in EBs cultivated under non-adherent conditions for 5 days. The ratio of cardiomyocytes per other cells was also potentiated in EBs cultivated in non-adherent conditions for only 5 days followed by cultivation in adherent serum-free culture conditions. Nevertheless, the alteration in the percentage of beating foci among these two tested cultivation conditions vanished at later phases and also did not affect the total number of cardiomyocytes determined as myosin heavy chain positive cells at the end of the differentiation process on day 20. Thus, although these modifications of the conditions of ES cells differentiation may intensify cardiomyocyte differentiation, the final count of cardiomyocytes might not change. Thus, serum depletion was identified as a key factor that intensified cardiomyogenesis. Further, the treatment of EBs with N-acetylcysteine, a reactive oxygen species scavenger, did not affect the observed increase in cardiomyogenesis under serum depleted conditions. Interestingly, a mild induction of the ventricular-like phenotype of cardiomyocytes was observed in 5-day-old EBs compared to 8-day-old EBs. Overall, these findings bring crucial information on the mechanisms of ES cells differentiation into cardiomyocytes and on the establishment of efficient protocols for the cardiomyogenic differentiation of ES cells. Further, the importance of determining the absolute number of formed cardiomyocyte-like cells per seeded pluripotent cells in contrast to the simple quantification of the ratios of cells is highlighted.
ER  -

RADASZKIEWICZ, Katarzyna Anna, Dominika SÝKOROVÁ, Lucia BINÓ, Jana KUDOVÁ, Markéta BÉBAROVÁ, Jiřina PROCHÁZKOVÁ, Hana KOTASOVÁ, Lukáš KUBALA and Jiří PACHERNÍK. The acceleration of cardiomyogenesis in embryonic stem cells in vitro by serum depletion does not increase the number of developed cardiomyocytes. \textit{Plos ONE}. San Francisco: Public Library of Science, 2017, vol.~12, No~3, p.~nestránkováno, 17 pp. ISSN~1932-6203. Available from: https://dx.doi.org/10.1371/journal.pone.0173140.

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