DNA double-strand breaks in human induced pluripotent stem cell reprogramming and long-term in vitro culturing

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216224%3A14330%2F17%3A00094705" target="_blank" >RIV/00216224:14330/17:00094705 - isvavai.cz</a>

Výsledek na webu

<a href="https://stemcellres.biomedcentral.com/articles/10.1186/s13287-017-0522-5" target="_blank" >https://stemcellres.biomedcentral.com/articles/10.1186/s13287-017-0522-5</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1186/s13287-017-0522-5" target="_blank" >10.1186/s13287-017-0522-5</a>

Jazyk výsledku

angličtina

Název v původním jazyce

DNA double-strand breaks in human induced pluripotent stem cell reprogramming and long-term in vitro culturing

Popis výsledku v původním jazyce

BACKGROUND: Human induced pluripotent stem cells (hiPSCs) play roles in both disease modelling and regenerative medicine. It is critical that the genomic integrity of the cells remains intact and that the DNA repair systems are fully functional. In this article, we focused on the detection of DNA double-strand breaks (DSBs) by phosphorylated histone H2AX (known as gammaH2AX) and p53-binding protein 1 (53BP1) in three distinct lines of hiPSCs, their source cells, and one line of human embryonic stem cells (hESCs). METHODS: We measured spontaneously occurring DSBs throughout the process of fibroblast reprogramming and during long-term in vitro culturing. To assess the variations in the functionality of the DNA repair system among the samples, the number of DSBs induced by gamma-irradiation and the decrease over time was analysed. The foci number was detected by fluorescence microscopy separately for the G1 and S/G2 cell cycle phases. RESULTS: We demonstrated that fibroblasts contained a low number of non-replication-related DSBs, while this number increased after reprogramming into hiPSCs and then decreased again after long-term in vitro passaging. The artificial induction of DSBs revealed that the repair mechanisms function well in the source cells and hiPSCs at low passages, but fail to recognize a substantial proportion of DSBs at high passages. CONCLUSIONS: Our observations suggest that cellular reprogramming increases the DSB number but that the repair mechanism functions well. However, after prolonged in vitro culturing of hiPSCs, the repair capacity decreases.

Název v anglickém jazyce

DNA double-strand breaks in human induced pluripotent stem cell reprogramming and long-term in vitro culturing

Popis výsledku anglicky

BACKGROUND: Human induced pluripotent stem cells (hiPSCs) play roles in both disease modelling and regenerative medicine. It is critical that the genomic integrity of the cells remains intact and that the DNA repair systems are fully functional. In this article, we focused on the detection of DNA double-strand breaks (DSBs) by phosphorylated histone H2AX (known as gammaH2AX) and p53-binding protein 1 (53BP1) in three distinct lines of hiPSCs, their source cells, and one line of human embryonic stem cells (hESCs). METHODS: We measured spontaneously occurring DSBs throughout the process of fibroblast reprogramming and during long-term in vitro culturing. To assess the variations in the functionality of the DNA repair system among the samples, the number of DSBs induced by gamma-irradiation and the decrease over time was analysed. The foci number was detected by fluorescence microscopy separately for the G1 and S/G2 cell cycle phases. RESULTS: We demonstrated that fibroblasts contained a low number of non-replication-related DSBs, while this number increased after reprogramming into hiPSCs and then decreased again after long-term in vitro passaging. The artificial induction of DSBs revealed that the repair mechanisms function well in the source cells and hiPSCs at low passages, but fail to recognize a substantial proportion of DSBs at high passages. CONCLUSIONS: Our observations suggest that cellular reprogramming increases the DSB number but that the repair mechanism functions well. However, after prolonged in vitro culturing of hiPSCs, the repair capacity decreases.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10601 - Cell biology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2017

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Stem Cell Research & Therapy

ISSN

1757-6512

e-ISSN

—

Svazek periodika

8

Číslo periodika v rámci svazku

73

Stát vydavatele periodika

DE - Spolková republika Německo

Počet stran výsledku

13

Strana od-do

1-13

Kód UT WoS článku

000396978500002

EID výsledku v databázi Scopus

2-s2.0-85016053500