New discovery could aid regenerative heart therapies

After birth, heart muscle cells undergo extensive changes to become fully mature adult cells, altering their form, function and physiology. However, the regulatory processes governing this maturation have been poorly understood thus far. For regenerative therapies in particular, this lack of understanding has proven a major limitation as efforts to grow stem cell-derived heart muscle cells have not been successful at producing mature adult cells, capable of restoring or improving heart function.

Publishing in Circulation, the research team used transcriptomic analysis to pinpoint an RNA splicing regulator named RBFox1 that was highly elevated soon after birth in a newborn heart. Analyses of published single-cell data also showed dramatic RBFox1 increase in maturing heart cells.

"This represents the first evidence that RNA splicing control contributes significantly to heart cell maturation," said lead author Dr Huang Jijun, who performed the preclinical study during his postdoctoral work at UCLA. "While RBFox1 alone may not be sufficient to mature fetal heart muscle cells all the way to fully matured adult cells, our findings uncover a new RNA-based internal network that can substantially drive this maturation process beyond other available approaches."

By expressing RBFox1 in immature human stem cell-derived heart cells, the researchers saw enhancements in key indicators of maturation, including cell size, sarcomere structure, contraction, calcium handling and oxygen usage. RBFox1 expression also led to the development of characteristic electrical properties seen in adult cells.

Further analyses showed RBFox1 regulated splicing of RNA transcripts linked to heart cell contraction and sarcomere components.

"This work demonstrates for the first time that altering RNA splicing alone can stimulate significant maturation of human stem cell-derived heart cells," said senior author Professor Wang Yibin, Director of the Cardiovascular & Metabolic Disorders Programme at Duke-NUS. "Our results uncover a promising molecular approach to improve heart cell maturation, which could overcome a major limitation in cardiac regenerative therapy and disease modelling."

While further research is needed to explore the mechanisms linking RBFox1-mediated RNA splicing with downstream maturation processes and phenotype, the study provides proof-of-concept that modulating RNA splicing can significantly impact heart muscle cell, or cardiomyocyte, maturation. This opens possibilities for regulating maturation that could eventually be translated to therapeutic strategies.

The study brought together researchers from leading research institutions across Singapore and the US. Collaborating groups hail from the Agency for Science, Technology and Research (A*STAR)'s Institute of Molecular and Cell Biology in Singapore, and Baylor College of Medicine, Forcyte Biotechnologies, the Greater Los Angeles VA Healthcare System, Meharry Medical College, the Stanford Cardiovascular Institute, the University of Cincinnati, the University of North Carolina, and Vanderbilt University School of Medicine in the US.

"Duke-NUS' partnerships with leading global institutions continue to foster impactful translational research that advances scientific knowledge and ultimately improves clinical outcomes for patients," said Professor Patrick Tan, Senior Vice-Dean for Research at Duke-NUS. "This study's findings provide a new understanding of the intrinsic regulatory network controlling heart cell maturation and reveal a promising molecular strategy that could potentially be harnessed to progress cell-based therapies and cardiac regenerative medicine."

Moving forward, the researchers will investigate how RBFox1 coordinates splicing to direct the functional and morphological changes underlying maturation. Their long-term aim is to identify druggable targets that can boost heart cell maturation efficiency for regenerative medicine use.