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Updated: Jul 19, 2025
Isolation and Characterization of Adult Cardiac Fibroblasts and Myofibroblasts
Published on: March 12, 2020
Heterogeneity and Functional Analysis of Cardiac Fibroblasts in Heart Development
Insights
Cardiac fibroblasts exhibit significant heterogeneity and play crucial roles in early heart development, but not in neonatal growth. Their functions are influenced by developmental stage and interactions with cardiomyocytes.
Area of Science:
- Cardiovascular Biology
- Developmental Biology
- Cellular Heterogeneity
Background:
- Cardiac fibroblasts are essential for heart function but their diverse origins and roles remain unclear.
- Previous studies have not systematically compared transcriptional profiles of cardiac fibroblasts from different sources.
- The overall function of fibroblasts as a cell type during heart development is largely uninvestigated.
Approach:
- Utilized single-cell mRNA sequencing (scRNA-seq) to analyze genome-wide and extracellular matrix gene expression in fibroblasts.
- Employed single molecular in situ hybridization to examine fibroblast subpopulation-specific gene expression patterns.
- Used a Diphtheria toxin fragment A (DTA) system to ablate fibroblasts at distinct developmental phases.
Key Points:
- Identified significant fibroblast heterogeneity across 18 developmental stages, with preserved lineage gene expression.
- Discovered differential expression of Wt1, Tbx18, and Aldh1a2 in fibroblast clusters, with Wt1+ and Tbx18+ fibroblasts originating from epicardial cells.
- Demonstrated the critical role of fibroblasts in early embryonic and heart growth, but not neonatal growth, via DTA-mediated ablation.
- Revealed zone- and stage-associated expression of extracellular matrix genes and fibroblast-cardiomyocyte ligand-receptor interactions.
Conclusions:
- Cardiac fibroblasts display heterogeneity from embryonic to neonatal stages, retaining lineage gene expression.
- Fibroblast ablation studies highlight their distinct developmental roles, modulated by stage-specific extracellular matrix genes and ligand-receptor interactions.
Background:
As one of the major cell types in the heart, fibroblasts play critical roles in multiple biological processes. Cardiac fibroblasts are known to develop from multiple sources, but their transcriptional profiles have not been systematically compared. Furthermore, while the function of a few genes in cardiac fibroblasts has been studied, the overall function of fibroblasts as a cell type remains uninvestigated.
Methods:
Single-cell mRNA sequencing (scRNA-seq) and bioinformatics approaches were used to analyze the genome-wide genes expression and extracellular matrix genes expression in fibroblasts, as well as the ligand-receptor interactions between fibroblasts and cardiomyocytes. Single molecular in situ hybridization was employed to analyze the expression pattern of fibroblast subpopulation-specific genes. The Diphtheria toxin fragment A (DTA) system was utilized to ablate fibroblasts at each developmental phase.
Results:
Using RNA staining of Col1a1 at different stages, we grouped cardiac fibroblasts into four developmental phases. Through the analysis of scRNA-seq profiles of fibroblasts at 18 stages from two mouse strains, we identified significant heterogeneity, preserving lineage gene expression in their precursor cells. Within the main fibroblast population, we found differential expressions of Wt1, Tbx18, and Aldh1a2 genes in various cell clusters. Lineage tracing studies showed Wt1- and Tbx18-positive fibroblasts originated from respective epicardial cells. Furthermore, using a conditional DTA system-based elimination, we identified the crucial role of fibroblasts in early embryonic and heart growth, but not in neonatal heart growth. Additionally, we identified the zone- and stage-associated expression of extracellular matrix genes and fibroblast-cardiomyocyte ligand-receptor interactions. This comprehensive understanding sheds light on fibroblast function in heart development.
Conclusion:
We observed cardiac fibroblast heterogeneity at embryonic and neonatal stages, with preserved lineage gene expression. Ablation studies revealed their distinct roles during development, likely influenced by varying extracellular matrix genes and ligand-receptor interactions at different stages.
