The basic helix–loop–helix transcription factor scleraxis regulates fibroblast collagen synthesis

Abstract 

The transcription factor scleraxis has been implicated in regulating the development of collagen-rich tissues such as tendons and cardiac valves, but its role in general collagen synthesis in the heart is unknown. Scleraxis expression in cardiac fibroblasts was examined, and its ability to regulate gene expression of collagen Iα2, the predominant cardiac collagen isoform, was assayed. Using real-time PCR, we demonstrate here that scleraxis mRNA is up-regulated by the profibrotic agonist TGF-β₁ in rat cardiac myofibroblasts, and that phenoconversion of fibroblasts to myofibroblasts similarly increases scleraxis expression. Over-expression of scleraxis in NIH-3T3 or primary rat cardiac fibroblasts by adenoviral gene delivery is sufficient to significantly increase collagen Iα2 gene expression. Using luciferase reporter assays, we demonstrate that scleraxis transactivates the human collagen Iα2 promoter in a DNA- and protein-binding dependent manner. Intriguingly, examination of infarcted rat hearts reveals a nearly four-fold increase in scleraxis expression in the infarct scar, but not in non-infarcted tissue. These data support a novel and previously unknown role for scleraxis in the regulation of collagen gene expression in the heart, including in post-infarct scar formation.

Abbreviations: αSMA, α-smooth muscle actin, bHLH, basic helix–loop–helix, DMEM, Dulbecco's Modified Eagle's Medium, ECM, extracellular matrix, MOI, multiplicity of infection, qPCR, quantitative real-time polymerase chain reaction, TGF-β₁, transforming growth factor β₁

Keywords: Cardiac myofibroblasts, Cardiac fibroblasts, Fibrosis, Extracellular matrix, Gene expression