Tamura Eduardo Koji, Cecon Erika, Monteiro Alex Willian Arantes, Silva Cláudia Lúcia Martins, Markus Regina Pekelmann
Departamento de Fisiologia, Instituto de Biociências, Universidade de São Paulo, São Paulo, Brazil.
J Pineal Res. 2009 Apr;46(3):268-74. doi: 10.1111/j.1600-079X.2008.00657.x. Epub 2009 Feb 9.
Endothelial cells produce NO by activation of constitutive nitric oxide synthase (NOS) and transcription of inducible NOS (iNOS). We have previously shown that melatonin, in the nanomolar range, inhibits activation of constitutive NOS, and in the present paper, we evaluated whether it could interfere with the expression of iNOS, which is activated by lipopolysaccharide (LPS), a major component of gram-negative bacteria cell walls. Primary cultures of rat endothelial cells were loaded with fluorescent probe for NO detection. Nuclear factor kappa B (NF-kappaB) translocation in endothelial cells elicited by LPS was measured by electromobility shift assay, and the vasodilation of aortic rings was accessed by recording isometric contraction. Melatonin in a micromolar but not in a nanomolar range inhibits the NO production induced by LPS. This effect is not dependent on the activation of G protein-coupled melatonin receptors. The nuclear NF-kappaB translocation is a process necessary for iNOS transcription, and melatonin also inhibits its translocation. LPS induced vasodilation only in endothelium-intact aortic rings, and melatonin (10 mum) inhibits the vasodilation. Here, we show that concentrations compatible with nocturnal melatonin surge (nm) did not interfere with the activity of iNOS. Considering that micromolar melatonin concentrations could be locally achieved through production by activated immune competent cells, extra-pineal melatonin could have a protective effect against tissue injury. We propose that melatonin blocked the LPS-induced vasodilation by inhibiting the NF-kappaB pathway. Finally, we propose that the effect of melatonin on vascular reactivity is one of the mechanisms that underlies the protective effect of this indolamine against LPS.
内皮细胞通过组成型一氧化氮合酶（NOS）的激活和诱导型NOS（iNOS）的转录产生一氧化氮（NO）。我们之前已经表明，纳摩尔浓度范围的褪黑素会抑制组成型NOS的激活，在本文中，我们评估了它是否会干扰iNOS的表达，iNOS由革兰氏阴性菌细胞壁的主要成分脂多糖（LPS）激活。用用于检测NO的荧光探针加载大鼠内皮细胞原代培养物。通过电泳迁移率变动分析测量LPS引起的内皮细胞中核因子κB（NF-κB）的易位，并通过记录等长收缩来评估主动脉环的血管舒张情况。微摩尔浓度而非纳摩尔浓度范围的褪黑素会抑制LPS诱导的NO产生。这种作用不依赖于G蛋白偶联褪黑素受体的激活。核NF-κB易位是iNOS转录所必需的过程，褪黑素也会抑制其易位。LPS仅在内皮完整的主动脉环中诱导血管舒张，而褪黑素（10μM）会抑制这种血管舒张。在这里，我们表明与夜间褪黑素激增（纳摩尔浓度）相当的浓度不会干扰iNOS的活性。考虑到微摩尔浓度的褪黑素可以通过活化的免疫活性细胞产生而在局部达到，松果体外的褪黑素可能对组织损伤具有保护作用。我们提出褪黑素通过抑制NF-κB途径阻断LPS诱导的血管舒张。最后，我们提出褪黑素对血管反应性的影响是这种吲哚胺对LPS具有保护作用的潜在机制之一。
