Plin5/p-Plin5 Guards Diabetic CMECs by Regulating FFAs Metabolism Bidirectionally

Background: Hyper-free fatty academia (HFFA) impairs cardiac capillaries, as well as type 2 diabetes mellitus (T2DM). Perilipin 5 (Plin5) maintains metabolic balance of free fatty acids (FFAs) in high oxidative tissues via the states of non-phosphorylation and phosphorylation. However, when facing to T2DM-HFFA, Plin5's role in cardiac microvascular endothelial cells (CMECs) is not defined. Methods: In mice of WT or Plin5-/-, T2DM models were rendered by high-fat diet combined with intraperitoneal injection of streptozocin. CMECs isolated from left ventricles were incubated with high glucose (HG) and high FFAs (HFFAs). Plin5 phosphorylation was stimulated by isoproterenol. Plin5 expression was knocked down by siRNA. We determined cardiac function by small animal ultrasound, apoptotic rate by flow cytometry, microvessel quantity by immunohistochemistry, microvascular integrity by scanning electron microscopy, intracellular FFAs by spectrophotometry, lipid droplets (LDs) by nile red staining, mRNAs by quantitative real time polymerase chain reaction, proteins by western blots, nitric oxide (NO) and reactive oxygen species (ROS) by fluorescent dye staining and enzyme-linked immunosorbent assay kits.   Results: In CMECs, HFFAs aggravated cell injury induced by HG and activated Plin5 expression. In mice with T2DM-HFFA, Plin5 deficiency reduced number of cardiac capillaries, worsened structural incompleteness and enhanced diastolic dysfunction. Moreover, in CMECs treated with HG-HFFAs, both ablation and phosphorylation of Plin5 reduced LDs content, increased intracellular FFAs, stimulated mitochondrial β-oxidation and added ROS generation, reduced the expression and activity of endothelial nitric oxide synthase (eNOS), eventually leading to increased apoptotic rate and decreased NO content, all of which were reversed by acetylcysteine. Conclusion: Plin5 preserves lipid balance and cell survival in diabetic CMECs by regualting FFAs metabolism bidirectionally via the states of non-phosphorylation and phosphorylation. Funding Statement: This work was supported by grants from Science Fund for Distinguished Young Scholars of Sichuan Province (No.2017JQ0012), National Science Funds of China (No.81500208), China Postdoctoral Science Foundation (No.2017M613429). Declaration of Interests: The authors declare: "No competing financial interests exist." Ethics Approval Statement: All experiments were conducted in adherence with National Institutes of Health Guidelines on the Use of Laboratory Animals, and were approved by Institutional Animal Care and Use Committee of Chengdu Military General Hospital.