CEBPB drives a novel endothelial pathological phenotype and promotes atherosclerosis by directly upregulating TGFBR1 expression

Endothelial cell (EC) dysfunction plays a crucial role in chronic vascular inflammation and diseases such as atherosclerosis. However, detailed transcriptional landscape, regulatory network and molecular signature of endothelial pathological phenotypes during atherosclerosis are not comprehensively understood. We hypothesized that novel molecular targets associated with endothelial pathology could be identified by respectively comparing the genetic profiles of functional ECs and dysfunctional ECs in normal and atherosclerotic aortae in mice of the same genetic background using single-cell RNA sequencing. In this study, we aimed to investigate the gene expression and transcriptional profiles of atherosclerotic aortic ECs, identify novel regulators driving endothelial pathological phenotype, and provide mechanistic insights into atherosclerotic progression. To test our hypothesis, single-cell RNA sequencing was performed on endothelial cell-enriched single cells of aortas from non-atherosclerotic mice, mild-atherosclerotic mice, and server-atherosclerotic mice, respectively. Human aortic endothelial cells were used to elucidate the role and underlying mechanism of the newly identified regulator in vitro. Adeno-associated virus was employed to specifically induce endothelial-overexpression of the novel regulator in Apoe -/- mice, allowing us to explore its impact on vascular inflammation and atherogenesis in vivo. Following clustering analysis, an intriguing endothelial pathological phenotype was defined by high expression of fibroblast markers, extracellular matrix, and inflammatory genes, concurrently with a rapid decline in endothelial markers. Notably, these cells did not express smooth muscle cell markers, such as Acta2 and Tagln, indicating a potentially distinct pathological phenotype from typical endothelial-to-mesenchymal transition (EndMT). Further analysis revealed that increased CEBPB contributes to the new pathological phenotype, which was further confirmed in cultured human aortic ECs, where upregulation of CEBPB led to elevated mRNA level of fibroblast markers and inflammation-related genes, while reducing endothelial markers. Additionally, CEBPB promoted EC adhesion to monocytes and worse barrier function. Mechanistically, CEBPB triggers TGF-β signaling and subsequent regulation of downstream genes through upregulation of TGF-β receptor type I (TGFBR1). The inhibition of TGFBR1, either by the inhibitor SB431542 or TGFBR1 siRNA, rescued CEBPB induced pathological transition. In depth, this occurs through direct interaction of CEBPB with the promotor region of TGFBR1, resulting in its upregulation. Furthermore, exacerbating atherosclerotic plaque area, enhanced vascular inflammation and increased endothelial TGFBR1 expression were confirmed by endothelial overexpression of CEBPB in vivo. In conclusion, these findings reveal that endothelial CEBPB functions as a novel transcription factor of TGFBR1, driving a new endothelial pathological phenotype and promoting vascular inflammation and atherosclerosis. Targeting endothelial CEBPB may offer new therapeutic avenues for atherosclerotic diseases. This work was supported by the National Key Research and Development Program of China [2021YFA1101100]; the National Natural Science Foundation of China [81970372]. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.