Copper Ion from Cu(2)O Crystal Induces AMPK-Mediated Autophagy via Superoxide in Endothelial Cells

Copper is an essential element required for a variety of functions exerted by cuproproteins. An alteration of the copper level is associated with multiple pathological conditions including chronic ischemia, atherosclerosis and cancers. Therefore, copper homeostasis, maintained by a combination of two copper ions (Cu(+) and Cu(2+)), is critical for health. However, less is known about which of the two copper ions is more toxic or functional in endothelial cells. Cubic-shaped Cu(2)O and CuO crystals were prepared to test the role of the two different ions, Cu(+) and Cu(2+), respectively. The Cu(2)O crystal was found to have an effect on cell death in endothelial cells whereas CuO had no effect. The Cu(2)O crystals appeared to induce p62 degradation, LC3 processing and an elevation of LC3 puncta, important processes for autophagy, but had no effect on apoptosis and necrosis. Cu(2)O crystals promote endothelial cell death via autophagy, elevate the level of reactive oxygen species such as superoxide and nitric oxide, and subsequently activate AMP-activated protein kinase (AMPK) through superoxide rather than nitric oxide. Consistently, the AMPK inhibitor Compound C was found to inhibit Cu(2)O-induced AMPK activation, p62 degradation, and LC3 processing. This study provides insight on the pathophysiologic function of Cu(+) ions in the vascular system, where Cu(+) induces autophagy while Cu(2+) has no detected effect.