Departamento de Bioquímica and Instituto de Química, Universidade de São Paulo, São Paulo, Brazil.
Free Radic Biol Med. 2012 Nov 15;53(10):1942-53. doi: 10.1016/j.freeradbiomed.2012.09.001. Epub 2012 Sep 14.
Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl) and other cyclic nitroxides have been shown to inhibit the chlorinating activity of myeloperoxidase (MPO) in vitro and in cells. To examine whether nitroxides inhibit MPO activity in vivo we selected acute carrageenan-induced inflammation on the rat paw as a model. Tempol and three more hydrophobic 4-substituted derivatives (4-azido, 4-benzenesulfonyl, and 4-(4-phenyl-1H-1,2,3-triazol-1-yl)) were synthesized, and their ability to inhibit the in vitro chlorinating activity of MPO and carrageenan-induced inflammation in rat paws was evaluated. All of the tested nitroxides inhibited the chlorinating activity of MPO in vitro with similar IC(50) values (between 1.5 and 1.8 μM). In vivo, the attenuation of carrageenan-induced inflammation showed some correlation with the lipophilicity of the nitroxide at early time points but the differences in the effects were small (<2-fold) compared with the differences in lipophilicity (>200-fold). No inhibition of MPO activity in vivo was evident because the levels of MPO activity in rat paws correlated with the levels of MPO protein. Likewise, paw edema, levels of nitrated and oxidized proteins, and levels of plasma exudation correlated with the levels of MPO protein in the paws of the animals that were untreated or treated with the nitroxides. The effects of the nitroxides in vivo were compared with those of 4-aminobenzoic hydrazide and of colchicine. Taken together, the results indicate that nitroxides attenuate carrageenan-induced inflammation mainly by reducing neutrophil migration and the resulting MPO-mediated damage. Accordingly, tempol was shown to inhibit rat neutrophil migration in vitro.
替普瑞酮（4-羟基-2,2,6,6-四甲基哌啶-1-氧自由基）和其他环状氮氧化物已被证明可以抑制髓过氧化物酶（MPO）的氯化活性在体外和细胞中。为了检查氮氧化物是否抑制体内 MPO 活性，我们选择了急性卡拉胶诱导的大鼠爪炎症作为模型。合成了替普瑞酮和另外三种疏水性 4-取代衍生物（4-叠氮基、4-苯磺酰基和 4-(4-苯基-1H-1,2,3-三唑-1-基))，并评估了它们抑制 MPO 体外氯化活性和卡拉胶诱导的大鼠爪炎症的能力。所有测试的氮氧化物都能抑制 MPO 在体外的氯化活性，IC50 值相似（在 1.5 和 1.8 μM 之间）。在体内，卡拉胶诱导的炎症的衰减与氮氧化物的亲脂性在早期时间点有一定的相关性，但与亲脂性的差异相比（>200 倍），效果的差异较小（<2 倍）。由于大鼠爪 MPO 活性与 MPO 蛋白水平相关，因此体内 MPO 活性没有明显抑制。同样，爪水肿、硝化和氧化蛋白水平以及血浆渗出水平与未处理或用氮氧化物处理的动物爪中 MPO 蛋白的水平相关。体内氮氧化物的作用与 4-氨基苯甲酰肼和秋水仙碱的作用进行了比较。总之，这些结果表明，氮氧化物通过减少中性粒细胞迁移和由此产生的 MPO 介导的损伤来减轻卡拉胶诱导的炎症。因此，替普瑞酮被证明可以抑制大鼠中性粒细胞在体外的迁移。
