D'Alessandro Angelo, Cervia Davide, Catalani Elisabetta, Gevi Federica, Zolla Lello, Casini Giovanni
Dipartimento di Scienze Ecologiche e Biologiche, Università della Tuscia, L.go dell'Università snc, I-01100 Viterbo, Italy.
Mol Biosyst. 2014 Jun;10(6):1290-304. doi: 10.1039/c3mb70362b. Epub 2014 Feb 11.
Ischemia is a primary cause of neuronal death in retinal diseases and the somatostatin subtype receptor 2 agonist octreotide (OCT) is known to decrease ischemia-induced retinal cell death. Using a recently optimized ex vivo mouse model of retinal ischemia, we tested the anti-ischemic potential of two additional neuropeptides, pituitary adenylate cyclase activating peptide (PACAP) and substance P (SP), and monitored the major changes occurring at the metabolic level. Metabolomics analyses were performed via fast HPLC online using a microTOF-Q MS instrument, a workflow that is increasingly becoming the gold standard in the field of metabolomics. The metabolomic approach allowed detection of the most significant alterations induced in the retina by ischemia and of the significance of the protective effects exerted by OCT, PACAP or SP. All treatments were shown to reduce ischemia-induced cell death, vascular endothelial growth factor over-expression and glutamate release. The metabolomic analysis showed that OCT and, to a lesser extent, also PACAP or SP, were able to counteract the ischemia-induced oxidative stress and to promote, with various efficacies, (i) decreased accumulation of glutamate and normalization of glutathione homeostasis; (ii) reduced build-up of α-ketoglutarate, which might serve as a substrate for the enhanced biosynthesis of glutamate in response to ischemia; (iii) reduced accumulation of peroxidized lipids and inflammatory mediators; (iv) the normalization of glycolytic fluxes and thus preventing the over-accumulation of lactate or either promoting the down-regulation of the glyoxalate anti-oxidant system; (v) a reduced metabolic shift from glycolysis towards the PPP or either a blockade at the non-oxidative phase of the PPP; and (vi) tuning down of purine metabolism. In addition, OCT seemed to stimulate nitric oxide production. None of the treatments was able to restore ATP production, although ATP reservoirs were partly replenished by OCT, PACAP or SP. These data indicate that, in addition to that of somatostatin, peptidergic systems such as those of PACAP and SP deserve attention in view of peptide-based therapies to treat ischemic retinal disorders.
缺血是视网膜疾病中神经元死亡的主要原因，已知生长抑素亚型受体2激动剂奥曲肽（OCT）可减少缺血诱导的视网膜细胞死亡。利用最近优化的视网膜缺血离体小鼠模型，我们测试了另外两种神经肽——垂体腺苷酸环化酶激活肽（PACAP）和P物质（SP）的抗缺血潜力，并监测了代谢水平上发生的主要变化。代谢组学分析通过使用microTOF-Q质谱仪的快速高效液相色谱在线进行，这种工作流程正日益成为代谢组学领域的金标准。代谢组学方法能够检测缺血在视网膜中引起的最显著变化，以及OCT、PACAP或SP所发挥的保护作用的重要性。所有治疗均显示可减少缺血诱导的细胞死亡、血管内皮生长因子过度表达和谷氨酸释放。代谢组学分析表明，OCT以及在较小程度上PACAP或SP能够抵消缺血诱导的氧化应激，并以不同的效力促进：（i）谷氨酸积累减少和谷胱甘肽稳态正常化；（ii）α-酮戊二酸积累减少，α-酮戊二酸可能作为缺血时谷氨酸生物合成增强的底物；（iii）过氧化脂质和炎症介质积累减少；（iv）糖酵解通量正常化，从而防止乳酸过度积累或促进乙醛酸抗氧化系统的下调；（v）从糖酵解向磷酸戊糖途径的代谢转变减少或磷酸戊糖途径非氧化阶段的阻断；以及（vi）嘌呤代谢下调。此外，OCT似乎能刺激一氧化氮的产生。尽管OCT、PACAP或SP部分补充了ATP储备，但没有一种治疗能够恢复ATP的产生。这些数据表明，除生长抑素外，鉴于基于肽的疗法可治疗缺血性视网膜疾病，像PACAP和SP这样的肽能系统也值得关注。
