Department of Anesthesiology, Affiliated Hospital of Zunyi Medical University, Zunyi, Guizhou 563000, P.R. China.
Zunyi Medical University, Zunyi, Guizhou 563000, P.R. China.
Mol Med Rep. 2021 Jun;23(6). doi: 10.3892/mmr.2021.12072. Epub 2021 Apr 13.
Previous studies have confirmed that 50 mol/l pinacidil postconditioning (PPC) activates the nuclear factor‑E2 related factor 2 (Nrf2)‑antioxidant responsive element (ARE) pathway, which protects the myocardium from ischemia‑reperfusion (IR) injury; however, whether this is associated with reactive oxygen species (ROS) generation remains unclear. In the present study, a Langendorff rat model of isolated myocardial IR was established to investigate the mechanism of PPC at different concentrations, as well as the association between the rat myocardial Nrf2‑ARE signaling pathway and ROS. A total of 48 rats were randomly divided into the following six groups (n=8 per group): i) Normal; ii) IR iii) 10 mol/l PPC (P10); iv) 30 mol/l PPC (P30); v) 50 mol/l PPC (P50); and vi) N‑(2‑mercaptopropionyl)‑glycine (MPG; a ROS scavenger) + 50 mol/l pinacidil (P50 + MPG). At the end of reperfusion (T3), compared with the IR group, the P10, P30 and P50 groups exhibited improved cardiac function, such as left ventricular development pressure, heart rate, left ventricular end‑diastolic pressure, +dp/dtmax, myocardial cell ultrastructure and mitochondrial Flameng score. Furthermore, the P10 and P50 groups demonstrated the weakest and most marked improvements, respectively. Additionally, in the P10, P30 and P50 groups, the residual ROS content at the end of reperfusion was highly negatively correlated with relative expression levels of Nrf2 gene and protein. Higher pinacidil concentration was associated with higher ROS generation at 5 min post‑reperfusion (T2), although this was significantly lower compared with the IR group, as well as with increased expression levels of antioxidant proteins and phase II detoxification enzymes downstream of the Nrf2 and Nrf2‑ARE pathways. This result was associated with a stronger ability to scavenge ROS during reperfusion, leading to lower levels of ROS at the end of reperfusion (T3) and less myocardial damage. The optimal myocardial protective effect was achieved by 50 mmol/l pinacidil. However, cardiac function of the P50 + MPG group was significantly decreased, ultrastructure of cardiomyocytes was significantly impaired and the relative expression levels of genes and proteins in the Nrf2‑ARE pathway were decreased. The aforementioned results confirmed that different PPC concentrations promoted early generation of ROS and activated the Nrf2‑ARE signaling pathway following reperfusion, regulated expression levels of downstream antioxidant proteins and alleviated myocardial IR injury in rats. Treatment with 50 mmol/l pinacidil resulted in the best myocardial protection.
先前的研究已经证实，50mmol/L 吡那地尔后处理（PPC）可激活核因子-E2 相关因子 2（Nrf2）-抗氧化反应元件（ARE）通路，从而保护心肌免受缺血再灌注（IR）损伤；然而，其是否与活性氧（ROS）的产生有关仍不清楚。在本研究中，建立了离体心肌 IR 的 Langendorff 大鼠模型，以研究不同浓度 PPC 的作用机制，以及大鼠心肌 Nrf2-ARE 信号通路与 ROS 之间的关系。将 48 只大鼠随机分为以下六组（每组 8 只）：i）正常组；ii）IR 组；iii）10mol/L PPC（P10）组；iv）30mol/L PPC（P30）组；v）50mol/L PPC（P50）组；和 vi）N-（2-巯基丙酰基）-甘氨酸（MPG；ROS 清除剂）+50mol/L 吡那地尔（P50+MPG）组。再灌注结束时（T3），与 IR 组相比，P10、P30 和 P50 组的左心室发展压、心率、左心室舒张末期压、+dp/dtmax、心肌细胞超微结构和线粒体 Flameng 评分等心功能均得到改善。此外，P10 和 P50 组的改善程度最弱和最强。此外，在 P10、P30 和 P50 组中，再灌注结束时残留 ROS 含量与 Nrf2 基因和蛋白的相对表达水平呈高度负相关。再灌注后 5min（T2）时，较高的吡那地尔浓度与较高的 ROS 生成相关，尽管与 IR 组相比，这一水平明显降低，并且 Nrf2 和 Nrf2-ARE 通路下游的抗氧化蛋白和 II 相解毒酶的表达水平也升高。这一结果与再灌注期间清除 ROS 的能力增强有关，导致再灌注结束时 ROS 水平较低，心肌损伤较少。50mmol/L 吡那地尔实现了最佳的心肌保护作用。然而，P50+MPG 组的心脏功能显著降低，心肌细胞超微结构严重受损，Nrf2-ARE 通路下游基因和蛋白的相对表达水平降低。上述结果证实，不同浓度的 PPC 在再灌注后促进早期 ROS 的产生并激活 Nrf2-ARE 信号通路，调节下游抗氧化蛋白的表达水平，减轻大鼠心肌 IR 损伤。使用 50mmol/L 吡那地尔治疗可获得最佳的心肌保护作用。
