# A fibroblast-like endothelial cell state promotes atherosclerosis via C/EBPβ-activated TGF-β signaling

## TL;DR

A fibroblast-like endothelial cell state worsens atherosclerosis by activating TGF-β signaling through C/EBPβ, linking inflammation to vascular disease.

## Contribution

Identifies C/EBPβ as a novel driver of fibroblast-like endothelial cell states and TGF-β signaling in atherosclerosis.

## Key findings

- A fibroblast-like EC population is enriched in atherosclerotic lesions and marked by ECM remodeling and inflammation.
- C/EBPβ activates TGFBR1 expression, enhancing TGF-β signaling and promoting atherosclerosis progression.
- Endothelial C/EBPβ overexpression increases plaque severity and vascular inflammation in vivo.

## Abstract

Endothelial cell (EC) dysfunction is a critical driver of chronic vascular inflammation and atherosclerosis. However, the molecular details of EC state dynamics during vascular disease progression remain ill-defined. Here, we used in-depth single-cell RNA sequencing to map transcriptional landscapes and molecular signatures of EC phenotypic plasticity during atherosclerosis in the mouse arota. This analysis identified a unique fibroblast-like EC population in atherosclerotic blood vessels, characterized by high expression of endothelial activation markers and extracellular matrix (ECM) remodeling, which increased with disease severity. Pseudotime trajectory analysis revealed that these fibroblast-like ECs represent terminal states of endothelial-mesenchymal transition (EndMT) during atherosclerosis. Further, the transcription factor C/EBPβ was identified as prominent driver of this phenotype transition as evidenced in vivo and in vitro. Mechanistically, inflammatory cytokines induce C/EBPβ, triggering TGF-β signaling and subsequent regulation of downstream genes via upregulation of TGF-β receptor type I (TGFBR1) through direct interaction with its promoter. Endothelial overexpression of C/EBPβ in vivo exacerbated atherosclerotic plaques, increased vascular inflammation, and elevated endothelial TGFBR1 levels. These findings highlight endothelial C/EBPβ as a novel regulator of TGF-β signaling and pathological fibroblast-like EC phenotypes during atherosclerosis, linking cytokine-driven inflammation with TGF-β-mediated endothelial dysfunction.

Endothelial cell (EC) dysfunction and TGF-β signaling-induced cell state dynamics drive chronic vascular fibrosis. This single-cell atlas maps transcriptional landscapes of EC phenotypic plasticity during atherosclerosis in the mouse aorta, and highlights a fibroblast-like EC population uniquely enriched in inflammatory lesions.

Single-cell RNA sequencing identifies a fibroblast-like EC population in atherosclerosis marked by endothelial-mesenchymal transition (EndMT), inflammation and extracellular matrix remodeling signatures.The transcription factor C/EBPβ drives this fibroblast-like EC state by increasing expression of mesenchymal markers and TGFBR1, enhancing TGF-β signaling.Overexpression of C/EBPβ in ECs exacerbates atherosclerosis by promoting plaque progression, vascular inflammation, and TGFBR1 levels in vivo.

Single-cell RNA sequencing identifies a fibroblast-like EC population in atherosclerosis marked by endothelial-mesenchymal transition (EndMT), inflammation and extracellular matrix remodeling signatures.

The transcription factor C/EBPβ drives this fibroblast-like EC state by increasing expression of mesenchymal markers and TGFBR1, enhancing TGF-β signaling.

Overexpression of C/EBPβ in ECs exacerbates atherosclerosis by promoting plaque progression, vascular inflammation, and TGFBR1 levels in vivo.

Combined single-cell profiling and functional analyses identify a C/EBPβ-dependent fibroblast-like endothelial cell population uniquely enriched during vascular inflammation.