OBJECTIVE The present study was desighed to determine the causative links and precise correlations among nitrosative stress, the cellular autophagy-lysosome process, and neurovascular damage during cerebral ischemia.METHODS To mimic ischemic-like conditions in vitro, endothelial cell cultures were exposed to permanent glucose deprivation and hypoxia for 2, 6, 12 and 24 h.The male rats were subjected to general anesthesia, after which 1000 non-radioactive microspheres were injected into the left carotid arter.The ipsilateral hemisphere was dissected out for morphological and biochemical analysis.RESULTS In parallelled with OGD-induced peroxynitrite formation, a rapid induction of microtubule-associated protein 1 light chain 3 (LC3)-Ⅰ / Ⅱ conversion and green fluorescent protein-LC3 punctae accumulation were observed in endothelial cells.The Western blot analyses indicated that OGD induced elevations in Lamp2 and cathepsin B protein levels.Furthermore, cultured endothelial cells treated with peroxynitrite (1-50 μmol· L-1) exhibited a concentration-dependent change in the pattern of autophagy-lysosome signaling.Intriguingly, OGD-induced autophagy-lysosorne processes were attenuated by PEP-19 overexpression and by a small-interfering RNA (siRNA)-mediated knockdown of eNOS.The current study also focused on the morphological and biochemical aspects of autophagy-lysosome process at miceovessels during the pathological process of cerebral microemboli in vivo.The importance of nitrosative stress in ischemia-induced autophagy Lyso-some cascades is further supported by our finding that pharmacological inhibition of nitrosative stress by melatonin partially inhibits the ischemia-induced autophagy-lysosome cascade and the degradation of the tight junction proteins.CONCLUSION Based on our observations, we predict that peroxynitrite-mediated nitrosative stress at least partially potentiates autophagy-lysosome signaling during sustained ischemic insult-induced endothelial cell damage.