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Suppression of Pro-fibrotic Signaling Potentiates Factor-mediated Reprogramming of Mouse Embryonic Fibroblasts into Induced Cardiomyocytes
Published on: June 3, 2018
Sustained β-catenin activation via GSK3 inhibition promotes direct fibroblast-to-cardiomyocyte reprogramming
Xinghua Wang1, Iqra Anwar1, Richard E Pratt1
1Mandel Center for Heart and Vascular Research, and the Duke Cardiovascular Research Center, Duke University Medical Center, Durham, North Carolina, USA.
Abstract:
During iPSC differentiation, cardiomyocyte formation requires transient activation of canonical Wnt/β-catenin signaling followed by pathway inhibition. Inspired by the mechanisms governing iPSC differentiation into cardiomyocytes, this study sought to explore whether these pathways similarly affect direct fibroblast-to-cardiomyocyte reprogramming. In contrast to iPSC differentiation, direct reprogramming required early inhibition of glycogen synthase kinase-3 (GSK3), which leads to activation of β-catenin signaling, without a subsequent requirement for pathway suppression. Pharmacological GSK3 inhibition enhanced cardiomyocyte yield and improved functional maturation, as demonstrated by optical calcium mapping. Reporter assays confirmed increased β-catenin transcriptional activity, while non-canonical Wnt signaling was unaffected. Transcriptomic analysis revealed enhanced enrichment of muscle-associated gene ontology terms and modulation of immune and signaling pathways following GSK3 inhibition. Together, these findings suggest that although indirect and direct cardiac reprogramming share elements of Wnt pathway involvement, their temporal and mechanistic requirements differ substantially.
Insights
Direct cardiac reprogramming requires early inhibition of glycogen synthase kinase-3 (GSK3) to activate Wnt/β-catenin signaling, unlike iPSC differentiation. This approach enhances cardiomyocyte yield and maturation.
Area of Science:
- Cardiovascular Research
- Stem Cell Biology
- Regenerative Medicine
Background:
- Cardiomyocyte generation from induced pluripotent stem cells (iPSCs) involves transient Wnt/β-catenin signaling.
- Understanding cardiac reprogramming pathways is crucial for regenerative therapies.
Purpose of the Study:
- To investigate if Wnt/β-catenin signaling dynamics in iPSC differentiation apply to direct fibroblast-to-cardiomyocyte reprogramming.
- To explore the role of glycogen synthase kinase-3 (GSK3) in direct cardiac reprogramming.
Main Methods:
- Pharmacological inhibition of GSK3.
- Reporter assays to assess β-catenin transcriptional activity.
- Optical calcium mapping for functional maturation assessment.
- Transcriptomic analysis.
Main Results:
- Direct reprogramming requires early GSK3 inhibition for β-catenin activation, unlike iPSC differentiation.
- GSK3 inhibition improved cardiomyocyte yield and functional maturation.
- Transcriptomic analysis showed enrichment of muscle-related genes and altered immune/signaling pathways.
Conclusions:
- Direct and indirect cardiac reprogramming share Wnt pathway involvement but differ significantly in temporal and mechanistic requirements.
- Early GSK3 inhibition is a key factor for efficient direct cardiac reprogramming.
