Poster #056, Stanford University
MEF2A Knockdown in Cardiomyocytes for Reversing Their Maturation
Mentors: Yena Oh, PhD and Sean Wu, MD, PhD
The non-regenerative nature of the adult heart can be attributed to the limited regenerative capacity of adult cardiomyocytes: the specialized muscle cells which compose the heart, synchronously contracting to form heartbeats. While immature cardiomyocytes retain modest regenerative potential, this capacity becomes extremely limited upon maturation into adult cardiomyocytes. Promoting regeneration in adult cardiomyocytes may enable repair and functional recovery following cardiac injury.
Myocyte enhancer factor 2A (MEF2A) is a critical transcription factor involved in cardiomyocyte maturation. We hypothesized that MEF2A knockdown would promote a shift towards an immature phenotype, characterized by decreased expression of maturation genes and increased expression of immature genes.
To explore this hypothesis, cardiomyocytes were cultured from induced pluripotent stem cells (iPSC-CMs) carrying a CRISPR interference (CRISPRi) construct. Mature D82 iPSC-CMs were electroporated with either a nontargeting scramble guide (scramble sgRNA) or a MEF2A-targeting guide RNA (MEF2A sgRNA). Gene expression changes were subsequently assessed by real-time quantitative PCR (RT-qPCR), 3 days following electroporation.
RT-qPCR confirmed a ~80% knockdown of MEF2A expression in MEF2A sgRNA cells compared to control iPSC-CMs. We found that MEF2A knockdown significantly reduced the expression of maturation markers, including MYH7 and MYL2, suggesting impaired cardiomyocyte maturation. However, expression of the immature marker, MYL7 was not significantly altered, although there was a trend. These findings suggest that while MEF2A is essential for maintaining cardiomyocyte maturation, it is not sufficient to induce a reversal of maturity in iPSC-cardiomyocytes.
The results from this study further clarify the role of MEF2A in cardiomyocyte maturation and provide insights into the molecular pathways relevant to cardiac regeneration.