Inhibition of autophagy by chloroquine stimulates nitric oxide production and protects endothelial function during serum deprivation.

BACKGROUND/AIMS Autophagy plays a fundamental role in cell survival under stress conditions such as nutrient deprivation. Decreased nitric oxide (NO) production, which may contribute to vascular dysfunction, is one of the consequences of autophagy in endothelial cells. The antimalarial drug chloroquine (CLQ) inhibits autophagy by blocking autophagosome formation and has been proposed as adjuvant chemotherapy in other diseases. METHODS Autophagy was induced by serum deprivation in Human Umbilical Vascular Endothelial Cells (HUVEC) as demonstrated by formation of Acidic Vesicular Organelles (AVOs), conversion of Microtubule-associated protein 1 light chain (LC3), and Sequestosome-1 (SQTM1/p62) degradation. Using endothelium-dependent vasorelaxation assays, intracellular NO production in an ex vivo rat aortic ring model pre-constricted with phenylephrine was estimated along with DAF-2 DA cell membrane-permeable NO sensitive fluorescent dye. RESULTS The inhibition of autophagy by CLQ restored NO levels, protected against superoxide generation and preserved morphology as well as proliferation of HUVEC under serum deprivation. Interestingly, the incubation of rat aortic rings with CLQ resulted in endothelium-dependent relaxation mediated by the increase of NO. CONCLUSION These findings emphasize the importance of autophagy in endothelial function and demonstrate the potential use of autophagy inhibitors to protect vascular function during nutrient deprivation.