Key result
NKX2.5 R141C mutation transiently impairs cardiomyogenesis in vitro and embryonic models.
Why the study?
An in vitro model to study early embryonic heart development changes caused by the human CHD-associated NKX2.5 R141C mutation was needed to understand mechanisms of congenital heart disease.
Supports mESC models for NKX2.5 R141C; leaves open mechanistic role in human congenital heart disease.
We have established an in vitro model of the human congenital heart defect (CHD)–associated mutation NKX2.5 R141C. We describe the use of the hanging drop method to differentiate Nkx2.5R141C/+ murine embryonic stem cells (mESCs) along with Nkx2.5+/+ control cells. This method allows us to recapitulate the early stages of embryonic heart development in tissue culture. We also use qRT‐PCR and immunofluorescence to examine samples at different time points during differentiation to validate our data. The in vivo model is a mouse line with a knock‐in of the same mutation. We describe the isolation of RNA from embryonic day 8.5 (E8.5) embryos and E9.5 hearts of wild‐type and mutant mice. We found that the in vitro model shows reduced cardiomyogenesis, similar to Nkx2.5R141C/+ embryos at E8.5, indicating a transient loss of cardiomyogenesis at this time point. These results suggest that our in vitro model can be used to study very early changes in heart development that cause CHD.
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Zakariyah et al. (2018) studied Congenital Heart Disease. NKX2.5 R141C mutation vs. Wild-type Nkx2.5+/+ control cells and mice was evaluated on Cardiomyogenesis at different differentiation time points. The in vitro NKX2.5 R141C mutation model demonstrated reduced cardiomyogenesis, similar to mutant embryos at embryonic day 8.5, indicating a transient loss of cardiomyogenesis.
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