Department of Pharmacology, Faculty of Medicine of Ribeirao Preto, University of Sao Paulo, Av. Bandeirantes, 3900, 14049-900, Ribeirao Preto, SP, Brazil.
Department of Physics, Faculty of Philosophy, Sciences and Letters of Ribeirao Preto. Av. Bandeirantes, 3900, 14040-901, Ribeirao Preto, SP, Brazil.
Free Radic Biol Med. 2013 Dec;65:446-455. doi: 10.1016/j.freeradbiomed.2013.07.032. Epub 2013 Jul 23.
Orally administered nitrite exerts antihypertensive effects associated with increased gastric nitric oxide (NO) formation. While reducing agents facilitate NO formation from nitrite, no previous study has examined whether antioxidants with reducing properties improve the antihypertensive responses to orally administered nitrite. We hypothesized that TEMPOL (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) could enhance the hypotensive effects of nitrite in hypertensive rats by exerting antioxidant effects (and enhancing NO bioavailability) and by promoting gastric nitrite-derived NO generation. The hypotensive effects of intravenous and oral sodium nitrite were assessed in unanesthetized freely moving rats with L-NAME (N(ω)-nitro-L-arginine methyl ester; 100mg/kg; po)-induced hypertension treated with TEMPOL (18mg/kg; po) or vehicle. While TEMPOL exerted antioxidant effects in hypertensive rats, as revealed by lower plasma 8-isoprostane and vascular reactive oxygen species levels, this antioxidant did not affect the hypotensive responses to intravenous nitrite. Conversely, TEMPOL enhanced the dose-dependent hypotensive responses to orally administered nitrite, and this effect was associated with higher increases in plasma nitrite and lower increases in plasma nitrate concentrations. In vitro experiments using electrochemical and chemiluminescence NO detection under variable pH conditions showed that TEMPOL enhanced nitrite-derived NO formation, especially at low pH (2.0 to 4.0). TEMPOL signal evaluated by electron paramagnetic resonance decreased when nitrite was reduced to NO under acidic conditions. Consistent with these findings, increasing gastric pH with omeprazole (30mg/kg; po) attenuated the hypotensive responses to nitrite and blunted the enhancement in plasma nitrite concentrations and hypotensive effects induced by TEMPOL. Nitrite-derived NO formation in vivo was confirmed by using the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (C-PTIO), which blunted the responses to oral nitrite. Our results showed that TEMPOL promotes nitrite reduction to NO in the stomach and enhanced plasma nitrite concentrations and the hypotensive effects of oral sodium nitrite through mechanisms critically dependent on gastric pH. Interestingly, the effects of TEMPOL on nitrite-mediated hypotension cannot be explained by increased NO formation in the stomach alone, but rather appear more directly related to increased plasma nitrite levels and reduced nitrate levels during TEMPOL treatment. This may relate to enhanced nitrite uptake or reduced nitrate formation from NO or nitrite.
经口给予的亚硝酸盐通过增加胃中的一氧化氮（NO）形成发挥降压作用。虽然还原剂促进亚硝酸盐形成 NO，但以前没有研究检查具有还原性质的抗氧化剂是否可以改善经口给予的亚硝酸盐的降压反应。我们假设 TEMPOL（4-羟基-2,2,6,6-四甲基哌啶-N-氧自由基）可以通过发挥抗氧化作用（并增强 NO 生物利用度）和促进胃中亚硝酸盐衍生的 NO 生成来增强高血压大鼠中硝普钠的降压作用。用 L-NAME（N（ω）-硝基-L-精氨酸甲酯；100mg/kg；po）预处理诱导高血压的未麻醉自由活动大鼠中评估静脉内和口服亚硝酸钠的降压作用，并给予 TEMPOL（18mg/kg；po）或载体。虽然 TEMPOL 在高血压大鼠中发挥了抗氧化作用，如血浆 8-异前列腺素和血管反应性氧物种水平降低所表明的，但这种抗氧化剂不影响静脉内硝普钠的降压反应。相反，TEMPOL 增强了口服给予的硝普钠的剂量依赖性降压反应，并且这种作用与血浆中硝酸盐浓度的升高有关。在可变 pH 条件下使用电化学和化学发光 NO 检测进行的体外实验表明，TEMPOL 增强了亚硝酸盐衍生的 NO 形成，特别是在低 pH（2.0 至 4.0）下。在酸性条件下将亚硝酸盐还原为 NO 时，TEMPOL 的电子顺磁共振评估信号减少。这些发现与以下发现一致，即用奥美拉唑（30mg/kg；po）增加胃 pH 会减弱对硝普钠的降压反应，并减弱 TEMPOL 诱导的血浆中亚硝酸盐浓度升高和降压作用。体内通过使用 NO 清除剂 2-（4-羧基苯基）-4,4,5,5-四甲基咪唑啉-1-氧-3-氧化物（C-PTIO）证实了亚硝酸盐衍生的 NO 形成，这削弱了对口服硝普钠的反应。我们的结果表明，TEMPOL 通过机制促进胃中亚硝酸盐还原为 NO，并通过胃 pH 依赖性机制增强了血浆中亚硝酸盐浓度和口服亚硝酸钠的降压作用。有趣的是，TEMPOL 对硝普钠介导的低血压的影响不能仅用胃中 NO 形成的增加来解释，而是似乎与 TEMPOL 治疗期间血浆中亚硝酸盐水平升高和硝酸盐水平降低直接相关。这可能与亚硝酸盐摄取增加或 NO 或亚硝酸盐形成的硝酸盐形成减少有关。
