Summary:Researches have shown that melatonin is neuroprotectant inischemia/reperfusion-media-ted injury. Although melatonin is known as an effective antioxidant, the mechanism of the protection cannot be explained merely by antioxidation. This study was devoted to exploreother existing mecha-nisms by investigating whether melatonin protects ischemia/reperfusion-injured neurons through ele-vating autophagy, since autophagy has been frequently suggested to play a crucial role in neuron sur-vival.To find it out, an ischemia/reperfusion model in N2acells was established for examinations. The results showed that autophagy was significantly enhanced in N2a cells treated with melatonin at reper-fusiononset following ischemia and greatly promoted cell survival, whileautophagy blockageby 3-MAled to theshortened N2a cell survival as assessed by MTT,transmission electron microscopy, and laser confocal scanning microscopy. Besides, the protein levels of LC3II and Beclin1 were remarkably in-creased in ischemia/reperfusion-injured N2a in the presence of melatonin, http://www.51lunwen.org/blx/ whereas the expression of p-PKB, key kinase in PI3K/PKBsignaling pathway,showed adecrease when compared with untreated subjects as accessed by immunoblotting. Taken together these data suggest that autophagy is possibly oneof themechanisms underlying neuroprotection ofmelatonin.
Keywords: melatonin; autophagy; ischemia/reperfusion; rapamycin; 3-MA;LC3; Beclin1; PKB;N2a
Cerebral ischemia is the most common type of cerebrovascular diseases[1, 2]. Cerebral ischemia caused by multiple reasons results in neuronal death involving many pathophysiologic mechanisms including severe failureof metabolicenergy support,elevated intracellular Ca2+, damage to macromolecules and cytoskeleton, mi- tochondrial injury and activation of inflammatory reac- tion[3]. Reperfusion and reoxygenation of the ischemic tissue re-established by prompt medical treatment in an effort to prevent severe neurologic damage and favor survival of individuals, also may provide chemical sub- strates for further increasing cellular alterations,neuronal death and neurologic deficits[4]. Overproduction of freeradicalsduringcerebralischemiaandreperfusion,among other pathophysiologic mechanisms, is known to con- tribute to functional disruption and neuronal death[3]. Additionally, a cascade of pro-apoptotic phenomena perpetrates abnormal cell conditions leading to perma- nentfunctional and structuraldamageand cell death[5,6]. Yanchun GUO,E-mail:[email protected],
△Theauthors contributed equally to this wore.
#Corresponding author, E-mail: [email protected]
*This project was supportedby agrant from thePhD Programs F fM y f f (N)oundationo inistr o Educationo China o.20070487101.
Experimental data from numerous studies clearly showed that melatonin, a biogenic amine produced by the pineal gland, is a highly effective agent in reducing neuronal loss and neurophysiologic deficits associated with brain ischemiaand reperfusion (I/R) both in animal models and at cell level[7-12].Yet thereal mechanism for this neuronal protection is not fully elucidated. Re- searches concerning the mechanism mainly focused on melatonin’s antioxidation, which, however, cannot pro- vide sufficient explanation, suggesting many other mechanisms maycontributetotheneuronalprotectionof melatonin as well. In particular, our previous study showed that melatonin did not suppress reactive oxygen species (ROS) immediately after its application but raised ROSlevel,whereasa stronger antioxidation effect of 6-OH melatonin was observed. However, the neuro- protectiveeffectof 6-OH melatoninispoorer than thatof melatonin[13].
Autophagy is a process in which eukaryotic cells self-digest part of their cytosoliccomponents so as to de-grade proteins and organelles to survive starvation and eliminate oxidatively damaged, aberrant macromolecules andorganellesto keep homeostasisinresponsetostress.Itis acrucial process for
