Abstract 15814: Novel Protective Mechanism of Cu Transporter ATP7A for Extracellular SOD Against Endothelial Dysfunction in Type2 Diabetes Vascular Complications

Abstract

Endothelial dysfunction induced by oxidative stress is the major vascular complication in diabetes mellitus (DM). Extracellular SOD (ecSOD), a secretory copper (Cu) enzyme, preserves endothelial function by scavenging extracellular superoxide, and its full activity requires Cu transporter ATP7A. We reported that reduced ATP7A-ecSOD pathway in vascular smooth muscle cells (VSMCs) is involved in endothelial dysfunction in type1 DM with hypoinsulinemia, but its role and mechanism in type2 DM with impaired insulin-Akt signaling is entirely unknown. Here we show that in VSMC, insulin increases ATP7A protein expression (1.8 fold) without affecting transcription by reducing ATP7A ubiquitination in a PI3K/Akt dependent manner. In organoid culture of mouse aorta, insulin increases ecSOD specific activity (47%) and ATP7A expression (1.9-fold). Insulin stimulation (2 hrs) promotes ATP7A translocation from TGN to caveolin (Cav) enriched lipid raft (C/LR) where it binds to ecSOD in PI3K/Akt dependent manner, reflecting Cu delivery to ecSOD through ATP7A at C/LR. Mechanistically, insulin increases pAkt binding to ATP7A to induce pSer-ATP7A, which is required for ATP7A translocation to C/LR. Mass spectrometry analysis identified Akt phosphorylation site of ATP7A at S1424/S1432/S1466. Of note, vascular ATP7A protein expression, ecSOD specific activity and endothelial function are impaired in Cav1 KO or Akt2 KO or ATP7Amut mice, but not Akt1 KO mice, suggesting the importance of Akt2-mediated ATP7A protein stabilization and its translocation to C/LR for ecSOD activation. Moreover, in aorta or VSMCs of high fat diet-induced type2 DM mice with impaired insulin-Akt pathway, ATP7A expression (57%) and ecSOD specific activity (53%), but not Cu/Zn SOD, are decreased. Acetylcholine-induced vasorelaxation is significantly impaired in resistant vessels of type2 DM mice, which is rescued in ATP7A-Tg/Type2 DM mice. In summary, insulin-Akt dependent ATP7A protein stabilization and translocation to C/LR promotes ecSOD activation, which prevents endothelial dysfunction in type2 DM. Thus, enhancing Akt2-ATP7A-ecSOD axis is potential therapeutic strategy for protecting against endothelial dysfunction in diabetes vascular complications.