SIRTUIN 3 REGULATES THE ENDOTHELIAL-TO-MESENCHYMAL TRANSITION BY IMPROVING AUTOPHAGIC DEGRADATION PYRUVATE KINASE M2

Abstract

Objective: Sirtuin 3 (SIRT3), an NAD-dependent deacetylase, is a major cellular sensor of energy metabolism. The aim of this work was therefore to determine the role of SIRT3-mediated autophagy in cellular metabolism and the processes of endothelial-to-mesenchymal transition (EndoMT). Design and method: Experiments were preformed on widetype (WT), SIRT3 knockout (SIRT3 KO) and SIRT3 transgenic (SIRT3 Tg) mice infused with Angiotension II (Ang II) or saline for 2 weeks. Murine aortic endothelial cells (MAECs) were cultured with Ang II, while the protein levels of mesenchymal markers, autophagy related protein and glycolytic enzymes were evaluated by Western blot analysis or immunofluorescence. Glycolysis was determined by measuring lactic acid release (ECAR) using an XF24 XF analyser. Results: We demonstrated that Ang II led to defective autophagy flux and high levels of glycolysis during EndoMT. The loss of SIRT3 further induced the hyperacetylation of endogenous ATG5, which in turn inhibited autophagosome maturation and increased pyruvate kinase M2 (PKM2) dimer expression. The M2 dimer is the less active form of PKM2, which drives glucose through the route of aerobic glycolysis. Additionally, TEPP-46, a selective PKM2 tetramer activator, had lower concentrations of lactate and led to the reduction of EndoMT both in vitro and in vivo. In parallel, the blockade of lactate influx from ECs into vascular smooth muscle cells (VSMCs) downregulated synthetic VSMC markers. EC-specific SIRT3-Tg mice exhibited a reduction in their endothelial cell transition, while partially rescuing vascular fibrosis and collagen accumulation. Conclusions: Taken together, these findings reveal that SIRT3 regulates EndoMT by improving the autophagic degradation of PKM2. Pharmacological targeting of glycolysis metabolism may therefore represent an effective therapeutic strategy for hypertensive vascular remodelling.