Related Experiment Video
Updated: Jul 8, 2025
En Face Endocardial Cushion Preparation for Planar Morphogenesis Analysis in Mouse Embryos
Published on: July 27, 2022
The role of c-Jun for beating cardiomycyte formation in prepared embryonic body
Lide Su1, Guofu Zhang1, Lili Jiang2
1Department of Cardiovascular Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, 150001, Heilongjiang, China.
Insights
Removing c-Jun in mouse stem cells promotes their differentiation into beating cardiomyocytes. This finding reveals c-Jun
Area of Science:
- Molecular Biology
- Stem Cell Biology
- Cardiovascular Research
Background:
- Cardiovascular diseases are a leading cause of death, partly due to the limited regenerative capacity of cardiomyocytes.
- The precise molecular mechanisms hindering cardiomyocyte regeneration remain incompletely understood.
- c-Jun, a known cell cycle regulator, influences various cellular processes but its role in early cardiac development is largely unexplored.
Purpose of the Study:
- To investigate the role of c-Jun in the early stages of cardiac differentiation.
- To determine if manipulating c-Jun affects the development of cardiomyocytes from stem cells.
Main Methods:
- Utilized CRISPR-Cas9 gene editing to create c-Jun knockout (ko) mouse embryonic stem cells (mESCs).
- Employed the hanging drop method for embryoid body formation.
- Assessed cardiac differentiation up to 9 days post-knockout using immunofluorescence, flow cytometry, and qPCR.
Main Results:
- c-Jun knockout mESCs exhibited spontaneous beating by day 6, significantly earlier than wild-type controls.
- Myocardial marker expression was substantially increased in c-Jun knockout cells.
- Upregulation of mesoderm and endoderm lineage markers was observed, indicating enhanced differentiation potential.
Conclusions:
- Disruption of c-Jun function promotes the differentiation of mouse embryonic stem cells towards a meso-endodermal lineage.
- This lineage commitment ultimately leads to the generation of functional, beating cardiomyocytes.
- c-Jun plays a critical regulatory role in the early developmental stages of cardiac cells.
Background:
Morbidity and mortality associated with cardiovascular diseases, such as myocardial infarction, stem from the inability of terminally differentiated cardiomyocytes to regenerate, and thus repair the damaged myocardial tissue structure. The molecular biological mechanisms behind the lack of regenerative capacity for those cardiomyocytes remains to be fully elucidated. Recent studies have shown that c-Jun serves as a cell cycle regulator for somatic cell fates, playing a key role in multiple molecular pathways, including the inhibition of cellular reprogramming, promoting angiogenesis, and aggravation of cardiac hypertrophy, but its role in cardiac development is largely unknown. This study aims to delineate the role of c-Jun in promoting early-stage cardiac differentiation.
Methods:
The c-Jun gene in mouse embryonic stem cells (mESCs) was knocked out with CRISPR-Cas9, and the hanging drop method used to prepare the resulting embryoid bodies. Cardiac differentiation was evaluated up to 9 days after c-Jun knockout (ko) via immunofluorescence, flow cytometric, and qPCR analyses.
Results:
Compared to the wild-type control group, obvious beating was observed among the c-Jun-ko mESCs after 6 days, which was also associated with significant increases in myocardial marker expression. Additionally, markers associated with mesoderm and endoderm cell layer development, essential for further differentiation of ESCs into cardiomyocytes, were also up-regulated in the c-Jun-ko cell group.
Conclusions:
Knocking out c-Jun directs ESCs toward a meso-endodermal cell lineage fate, in turn leading to generation of beating myocardial cells. Thus, c-Jun plays an important role in regulating early cardiac cell development.
