Endothelial CEPT1 Promotes Angiogenesis Through PPARα and VEGF-A Signaling.

BACKGROUND

Cept1 is essential for de novo phopholipogenesis and is impacted by diabetes. We previously demonstrated that conditional knockdown of Cept1 in the endothelium leads to reduced tissue recovery. Therefore, we hypothesized that Cept1 overexpression may also be sufficient in promoting postischemic angiogenesis and recovery in the setting of diabetes.

METHODS

CEPT1 (choline-ethanolamine phosphotransferase 1) content was evaluated in the peripheral arteries of human patients with peripheral artery disease and with or without diabetes. We also engineered a conditional endothelial cell (EC)-specific Cept1 overexpression mouse (Cept1fl/fl Cre+) in adult C57BL/6J (C57 black 6J) mice and performed unilateral hindlimb ischemia to assess the role of Cept1 in promoting angiogenesis. Murine aortae and ECs were harvested for single-cell RNA sequencing and molecular pathway analysis.

RESULTS

In human arterial intima, CEPT1 was elevated in the setting of peripheral artery disease and diabetes, along with ACOX1 (acyl-coenzyme A oxidase 1), VEGF (vascular endothelial growth factor) R2, p-Akt (phosphorylated Akt), and p-eNOS (phosphorylated endothelial nitric oxide synthase). In mice, single-cell RNA sequencing demonstrated that ECs with Cept1 overexpression were enriched with wound healing, angiogenesis, sprouting, and cell migration pathways. Diabetic Cept1fl/flCre+ mice that underwent hindlimb ischemia demonstrated improved hindlimb perfusion and angiogenesis, and their aortic rings had increased ex vivo capillary sprouting. Cept1 overexpression in ECs significantly increased migration, tubule formation, and proliferation as predicted by single-cell RNA sequencing. Cept1 overexpression in ECs also led to increased expression of Pparα, Acox1, Vegfa, and Vegfr2. Similarly, treatment with siPparα and inhibitors for PPARα (peroxisome proliferator-activated receptor α; GW6471), VEGFR2 (ZM323881), Akt (LY294002), and eNOS (L-NAME [nitro-L-arginine methyl ester]) abrogated CEPT1-induced EC migration.

CONCLUSIONS

Cept1 overexpression promotes EC function and postischemic recovery. The impact of CEPT1 on ECs is at least in part dependent on p-Akt/p-eNOS angiogenic signaling and PPARα. Because CEPT1 is elevated in diseased human peripheral arterial tissue, these findings suggest that CEPT1 may be playing an important compensatory role in vascular recovery and reperfusion following ischemic injury in the setting of diabetes.