miR-145-5p modulates collagen production and fibroblast behaviour after cardiac injury partially via COL5A1
H Wang, J Paavola, S Perttunen, K Immonen, I Hagerstrom, S Narumanchi, K Kalervo, J Sinisalo, M Mayranpaa, H Ruskoaho, R Kosonen, V Olkkonen, M Laine, I Tikkanen, P LakkistoAbstract
Background
In contrast to humans, upon cryoinjury zebrafish hearts develop transient fibrotic scar that subsequently resolves and renews with functional cardiomyocytes.
Purpose
We aimed to understand the molecular mechanisms underlying the phenotypic transitions from cardiac scarring to fibrotic resolution in regenerating zebrafish hearts.
Methods
Speckle-tracking echocardiography was used to evaluate the cardiac function of adult zebrafish following cryoinjury. Bulk RNA-seq data were generated from healthy and cryoinjured hearts at 14 days post-injury. Bioinformatics analysis was performed to identify differentially expressed genes and miRNAs, enriched biological pathways, and interaction networks of the differentially expressed miRNAs and their target genes. The expression of these miRNAs and genes was validated by RT-qPCR in regenerating zebrafish hearts and in failing hearts of patients with ischemic cardiomyopathy. Human primary cardiac fibroblasts were employed to investigate the potential role of miR-145-5p and COL5A1 in cardiac scar remodelling.
Results
Principal component analyses revealed distinct transcriptome clusters corresponding to the healthy and the injured hearts, highlighting significant changes during cardiac regeneration. Gene set enrichment analyses indicated that extracellular matrix organization and inflammatory response pathways were activated in cryoinjured hearts, while mitochondrial organization and oxidative phosphorylation pathways were downregulated. Notably, miR-145-5p was downregulated and col5a1 was upregulated in the cryoinjured hearts, contrasting with the upregulation of miR-145-5p and downregulation of COL5A1 in the left ventricles of ischemic cardiomyopathy patients. Prediction of miRNA-mRNA interaction and a dual luciferase assay indicated that COL5A1 mRNA is a target of miR-145-5p. Overexpression of miR-145-5p in human cardiac fibroblasts decreased COL5A1 and α-SMA, increased COL1A, and attenuated fibroblast differentiation, proliferation, and migration. Conversely, inhibition of miR-145-5p enhanced COL5A1 and α-SMA, reduced COL1A, and promoted fibroblast proliferation and migration. Furthermore, downregulation of COL5A1 suppressed fibroblast proliferation and migration, which could be rescued by inhibition of miR-145-5p. Interestingly, downregulation of COL5A1 diminished the expression of integrin subunits ITGβ3 and ITGβ5.
Conclusions
miR-145-5p modulates collagen production and fibroblast behaviour partially via targeting COL5A1, potentially through integrin-mediated pathways. Our study highlights that a fibroblast state with low collagen secretion and high proliferation and migration is essential for cardiac scar remodelling in regenerating hearts.