Department of Cardiology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, Sichuan 400016, P.R. China.
Center for Perinatal Biology, Division of Pharmacology, Department of Basic Sciences, Loma Linda University School of Medicine, Loma Linda, CA 92350, USA.
Int J Mol Med. 2018 Mar;41(3):1693-1703. doi: 10.3892/ijmm.2017.3318. Epub 2017 Dec 12.
Accumulating evidence suggests that glucagon-like peptide-1 (GLP-1) and its analogues exert cardioprotective effects via modulating cardiomyocyte metabolism. Mitochondria play a pivotal role in the regulation of cell metabolism. It was hypothesized that treatment with exenatide, a GLP-1 analogue, may exert cardioprotective effects by improving mitochondrial function in an in vitro model of hypoxia/reoxygenation (H/R). H9c2 cells were employed to establish an in vitro model of H/R. Exenatide was added to the cells for 30 min prior to exposure to hypoxia. The GLP-1 receptor antagonist exendin‑(9‑39), the cyclic adenosine monophosphate (cAMP) inhibitor Rp-cAMPS and the protein kinase A (PKA) inhibitor H-89 were added to the cells for 10 min prior to treatment with exenatide. The release of lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) and cardiomyocyte apoptosis were evaluated. The characteristics of mitochondrial morphology and functions, including ATP synthesis, membrane potential (ΔΨm), mitochondrial permeability transition pore (mPTP), mitochondrial ATPase activity and oxidative stress, were determined. the mitochondrial uncoupling protein-3 (UCP-3) and nuclear respiratory factor-1 (Nrf-1) were also investigated by western blot analysis. Exenatide pretreatment significantly decreased LDH and CK-MB release and cardiomyocyte apoptosis in H9c2 cells subjected to H/R. More importantly, to the best of our knowledge, this is the first report of exenatide pretreatment decreasing mitochondrial abnormalities and reducing oxidative stress, while enhancing ATP synthesis, mitochondrial ATPase activity and ΔΨm in H9c2 cells subjected to H/R. Exenatide pretreatment also decreased mitochondrial calcium overload and inhibited the opening of mPTP in H9c2 cells subjected to H/R. Furthermore, exenatide pretreatment upregulated UCP-3 and Nrf-1 expression in H9c2 cells subjected to H/R. However, the abovementioned observed effects of exenatide were all abolished when exenatide was co-administered with exendin‑(9‑39), Rp-cAMPS and̸or H-89. Therefore, the GLP-1 analogue exenatide was found to exert cardioprotective effects in an in vitro model of H/R, and this cardioprotection may be attributed to the improvement of mitochondrial function. These effects are most likely associated with the activation of the GLP-1 receptor/cAMP/PKA signaling pathway.
越来越多的证据表明，胰高血糖素样肽-1（GLP-1）及其类似物通过调节心肌细胞代谢发挥心脏保护作用。线粒体在细胞代谢调节中起着关键作用。据推测，在缺氧/复氧（H/R）的体外模型中，用 GLP-1 类似物 exenatide 治疗可能通过改善线粒体功能发挥心脏保护作用。使用 H9c2 细胞建立 H/R 的体外模型。在暴露于缺氧之前，将 exenatide 添加到细胞中 30 分钟。在使用 exenatide 治疗之前，将 GLP-1 受体拮抗剂 exendin-（9-39）、环磷酸腺苷（cAMP）抑制剂 Rp-cAMPS 和蛋白激酶 A（PKA）抑制剂 H-89 添加到细胞中 10 分钟。评估乳酸脱氢酶（LDH）和肌酸激酶-MB（CK-MB）的释放以及心肌细胞凋亡。测定线粒体形态和功能的特征，包括三磷酸腺苷（ATP）合成、膜电位（ΔΨm）、线粒体通透性转换孔（mPTP）、线粒体 ATP 酶活性和氧化应激。通过 Western blot 分析还研究了线粒体解偶联蛋白-3（UCP-3）和核呼吸因子-1（Nrf-1）。与 H/R 处理的 H9c2 细胞相比，exenatide 预处理可显著降低 LDH 和 CK-MB 的释放和心肌细胞凋亡。更重要的是，据我们所知，这是首次报道 exenatide 预处理可降低 H/R 处理的 H9c2 细胞中的线粒体异常并减少氧化应激，同时增强 ATP 合成、线粒体 ATP 酶活性和 ΔΨm。Exenatide 预处理还可减少 H/R 处理的 H9c2 细胞中线粒体钙超载并抑制 mPTP 的开放。此外，exenatide 预处理还可上调 H/R 处理的 H9c2 细胞中 UCP-3 和 Nrf-1 的表达。然而，当 exenatide 与 exendin-（9-39）、Rp-cAMPS 和/或 H-89 联合使用时，上述 exenatide 的观察到的作用均被消除。因此，在 H/R 的体外模型中发现 GLP-1 类似物 exenatide 具有心脏保护作用，这种心脏保护作用可能归因于线粒体功能的改善。这些作用很可能与 GLP-1 受体/cAMP/PKA 信号通路的激活有关。
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