Optimisation of the cell cultivation methods in the embryonic stem cell test results in an increased differentiation potential of the cells into strong beating myocard cells

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• 作者：Ann De Smedt ; Margino Steemans ; Marlies De Boeck ; Annelieke K. Peters ; Bas-jan van der Leede ; Freddy Van Goethem ; Ann Lampo ; Philippe Vanparys
• 关键词：Embryonic stem cell ; Mouse ; Optimisation
• 刊名：Toxicology in Vitro
• 出版年：2008
• 出版时间：October 2008
• 年：2008
• 卷：22
• 期：7
• 页码：1789-1796
• 全文大小：637 K

文摘

In order to support drug research in the selection process for non-embryotoxic pharmaceutical compounds, a screening method for embryotoxicity is needed. The murine embryonic stem cell test (EST) is a validated in vitro test based on two permanent mouse cell lines and delivering results in 10-days.

Implementation of this test within our laboratory, revealed variability in the differentiation potential of the embryonic stem cells and, as a consequence, a lot of assays needed to be rejected due the fact the acceptance criteria were not reached. In order to gain a better yield of contracting myocardial cells, we used (1) a stringent control of the cell growth during subcultivation and a standardised hanging drop culture method and (2) a non-enzymatic cell harvest instead of a trypsin/EDTA cell harvest.

Implementing of these cell culture modifications resulted in a decreased variability in the size of embryonic bodies, an increase of the number of acceptable tests and a significant increase of the differentiation potential of embryonic cells into strong beating myocardium, which made scoring less time consuming. Testing of 6 reference compounds in the optimized EST showed that the cell culture modifications did not changed the in vitro classification.