Dipartimento di Scienze Biochimiche A. Rossi Fanelli, Istituto di Biologia e Patologia Molecolari del CNR, Sapienza Università di Roma, P.le Aldo Moro 5, 00185 Rome, Italy.
Amino Acids. 2012 Aug;43(2):875-84. doi: 10.1007/s00726-011-1147-0. Epub 2011 Nov 24.
Neutrophils play a major role in acute inflammation by generating reactive oxygen/nitrogen species. Opioid peptides, including enkephalins, are present at inflammation sites. Neutrophils contribute to protect against inflammatory pain by releasing opioid peptides. In this investigation, the ability of human polymorphonuclear cells to induce oxidative and nitrative modifications of Leu-enkephalin has been investigated in vitro. Activated human neutrophils mediate the oxidation of Leu-enkephalin resulting in the production of dienkephalin. In the presence of nitrite at concentrations observed during inflammatory and infectious process (10-50 μM), nitroenkephalin, a nitrated derivative of Leu-enkephalin, is additionally formed. The yield of nitroenkephalin increases with nitrite concentration and is significantly inhibited by the addition of catalase or 4-aminobenzoic acid hydrazide (ABAH), a specific inhibitor of peroxidases. These results suggest that neutrophils induce nitration of Leu-enkephalin by a mechanism that is dependent on myeloperoxidase activity and hydrogen peroxide. Oxidative/nitrative modifications of Leu-enkephalin have been also evidenced when cells were treated with the NO-donor molecule, DEANO. The nitrated enkephalin has been examined for its effect on leukocyte functional responses. The data reveal that nitroenkephalin at micromolar concentrations inhibits superoxide anion generation and degranulation of azurophilic granules of human polymorphonuclear cells. Moreover, nitroenkephalin inhibits spontaneous apoptosis of neutrophils, as evaluated by measuring caspase-3 activity. Collectively, our data indicate that the nitrated enkephalin attenuates neutrophil activation and promotes the short-term survival of these cells, suggesting a possible role of the nitrocompound in the efficiency and resolution of inflammatory processes.
中性粒细胞通过生成活性氧/氮物种在急性炎症中发挥主要作用。阿片肽，包括脑啡肽，存在于炎症部位。中性粒细胞通过释放阿片肽有助于防止炎症性疼痛。在本研究中，研究了人多形核细胞在体外诱导亮氨酸脑啡肽氧化和硝化修饰的能力。激活的人中性粒细胞介导亮氨酸脑啡肽的氧化，导致二烯脑啡肽的产生。在炎症和感染过程中观察到的浓度（10-50 μM）的亚硝酸盐存在下，亮氨酸脑啡肽的硝化衍生物硝基脑啡肽另外形成。随着亚硝酸盐浓度的增加，硝基脑啡肽的产率增加，并且通过添加过氧化氢酶或 4-氨基苯甲酰肼（ABAH）（过氧化物酶的特异性抑制剂）显著抑制。这些结果表明，中性粒细胞通过依赖髓过氧化物酶活性和过氧化氢的机制诱导亮氨酸脑啡肽的硝化。当用 NO 供体分子 DEANO 处理细胞时，也已经证明了亮氨酸脑啡肽的氧化/硝化修饰。已经检查了硝化的脑啡肽对白细胞功能反应的影响。数据显示，在微摩尔浓度下，硝基脑啡肽抑制人多形核细胞的超氧阴离子生成和嗜中性粒细胞的脱粒。此外，硝基脑啡肽抑制中性粒细胞的自发凋亡，如通过测量半胱天冬酶-3 活性来评估。总的来说，我们的数据表明，硝化的脑啡肽减弱了中性粒细胞的激活，并促进了这些细胞的短期存活，表明该硝基化合物在炎症过程的效率和解决中可能起作用。
