School of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, SP, Brazil.
Faculty of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Ribeirão Preto, SP, Brazil.
Nitric Oxide. 2019 May 1;86:12-20. doi: 10.1016/j.niox.2019.02.004. Epub 2019 Feb 14.
This study investigated the intracellular mechanisms involved in the vasodilatation induced by the classic NO donor SNP and the non-classic NO donor cis-Ru(bpy)2(py)(NO2) (or RuBPY) in mesenteric resistance arteries obtained from renal hypertensive (2K-1C) and normotensive (2K) rats.
On the basis of fluorimetric assays in cultured vascular smooth muscle cells (VSMCs) isolated from 2K-1C and 2K rats, we measured NO release from SNP and RuBPY, cytosolic Ca concentration ([Ca]c), and reactive oxygen species (ROS) with the selective probes DAF-2DA, Fluo-3AM and the more selective probe for peroxynitrite (7-CBA), respectively. We determined isometric tension in mesenteric arteries to assess SNP- and RuBPY-induced relaxation.
SNP and RuBPY released NO in comparable amounts in cultured aortic VSMCs from hypertensive 2K-1C and normotensive 2K rats. The NO scavenger hydroxocobalamin blunted NO release. Sarco/endoplasmic reticulum Ca ATPase (SERCA) inhibition with thapsigargin reduced [Ca]c in normotensive 2K rat VSMCs only. ROS amounts were greater in hypertensive 2K-1C than in normotensive 2K rat VSMCs, but neither SNP nor RuBPY altered ROS concentrations in any of the groups. SNP and RuBPY induced similar relaxation in hypertensive 2K-1C and normotensive 2K rat mesenteric resistance arteries. The SNP and RuBPY-induced relaxation involves sGC and PKG activation. On the other hand, SNP but not RuBPY activates K channels. Interestingly, SERCA inhibition reduces SNP induced relaxation only in normotensive 2K rat mesenteric arteries whereas RuBPY-induced relaxation does not involve SERCA activation in both normotensive and hypertensive arteries.
Our results indicate that SNP and RuBPY-induced mesenteric resistance artery relaxation involves NO/sGC/cGMP/PKG pathway activation. K channels and SERCA activation is required to SNP but not for RuBPY-induced relaxation. Moreover, SERCA seems to be impaired in hypertensive 2K-1C rat mesenteric resistance arteries although it does not impact SNP- or RuBPY-induced relaxation.
本研究旨在探讨经典一氧化氮供体 SNP 和非经典一氧化氮供体 cis-Ru(bpy)2(py)(NO2)（或 RuBPY）在肾高血压（2K-1C）和正常血压（2K）大鼠肠系膜阻力血管中诱导血管舒张的细胞内机制。
基于从 2K-1C 和 2K 大鼠分离的培养血管平滑肌细胞（VSMCs）的荧光测定，我们分别用 DAF-2DA、Fluo-3AM 和更特异的过氧亚硝酸盐探针（7-CBA）测量 SNP 和 RuBPY 的 NO 释放、胞浆 Ca 浓度（[Ca]c）和活性氧（ROS）。我们测定肠系膜动脉的等长张力，以评估 SNP 和 RuBPY 诱导的舒张。
SNP 和 RuBPY 在来自高血压 2K-1C 和正常血压 2K 大鼠的培养主动脉 VSMCs 中释放相当数量的 NO。NO 清除剂羟钴胺素削弱了 NO 释放。用他帕昔琼抑制肌浆/内质网 Ca ATP 酶（SERCA）仅减少正常血压 2K 大鼠 VSMCs 的[Ca]c。ROS 含量在高血压 2K-1C 大鼠 VSMCs 中高于正常血压 2K 大鼠 VSMCs，但 SNP 和 RuBPY 在任何组中都没有改变 ROS 浓度。SNP 和 RuBPY 在高血压 2K-1C 和正常血压 2K 大鼠肠系膜阻力血管中诱导相似的舒张。SNP 和 RuBPY 诱导的舒张涉及 sGC 和 PKG 激活。另一方面，SNP 但不是 RuBPY 激活 K 通道。有趣的是，SERCA 抑制仅在正常血压 2K 大鼠肠系膜动脉中降低 SNP 诱导的舒张，而 RuBPY 诱导的舒张在正常血压和高血压血管中均不涉及 SERCA 激活。
我们的结果表明，SNP 和 RuBPY 诱导的肠系膜阻力动脉舒张涉及 NO/sGC/cGMP/PKG 途径的激活。K 通道和 SERCA 的激活对于 SNP 诱导的舒张是必需的，但不是 RuBPY 诱导的舒张所必需的。此外，尽管不影响 SNP 或 RuBPY 诱导的舒张，但 SERCA 似乎在高血压 2K-1C 大鼠肠系膜阻力血管中受损。
