Abstract Body: Abstract
Objectives: Choline/ethanolamine phosphotransferase 1 (CEPT1) is essential for de novo phospholipogenesis and for endothelial cell (EC) activation. CEPT1 catalyzes the formation of phospholipids, which subsequently activate the transcription factor peroxisome proliferator-activated receptor α (PPARα). We previously observed that CEPT1 and PPARα are upregulated in arterial tissue of patients with peripheral arterial disease (PAD). We hypothesized that they play an important role in macrophage recruitment and atherosclerotic disease progression.
Methods: We developed a Cre-induced conditional EC-specific Cept1 overexpression model in C57BL/6J mice. ECs were isolated and pooled from both Cept1^fl/flCre^- and Cept1^fl/flCre⁺ mice (n=3; Figure 1A), and single-cell RNA sequencing (scRNA-seq) was performed to identify unique molecular markers. Western blotting and immunostaining were conducted on lower extremity arterial structures stratified in minimally (Min) and maximally (Max) disease, from patients with and without PAD (n=3-9). All experimental conditions were conducted in triplicate and analyzed using a one-way ANOVA test.
Results: scRNA-seq from Cept1^fl/flCre⁺ mice demonstrated dense clustering of macrophages (Figure 1B-D), and elevated expression of inflammatory and endocytosis-associated genes (Figure 1E-G), such as Perilipin2 (Plin2; p<0.01; Figure 1H). In human arterial tissue, CEPT1, PECAM1, and PLIN2 content were elevated in arterial segments with Max disease, but not in healthy arterial segments (p<0.05, Figure 2A-D). Similarly, immunostaining demonstrated higher PLIN2 content in CD68+ macrophages in Max diseased arterial segments of patients with PAD (yellow arrows; p<0.05; Figure 2E-G).
Conclusion: Cept1 is crucial for inducing endothelial cell (EC) activation in arterial structures within the context of peripheral arterial disease (PAD). This finding underscores the significant connection between de novo phospholipid synthesis in EC physiology and the progression of atherosclerosis. Future research will be necessary to elucidate the impact of CEPT1 on the advancement of atherosclerosis in PAD patients.