Department of Ecological and Biological Sciences, University of Tuscia, Viterbo, Italy.
Department of Experimental Medicine, University La Sapienza, Rome, Italy.
Biochem Pharmacol. 2017 Mar 1;127:60-70. doi: 10.1016/j.bcp.2016.12.017. Epub 2016 Dec 23.
Recent evidence has shown the cardioprotective effect of PDE5 inhibition in myocardial ischemia/reperfusion injury, heart failure and cardiac hypertrophy. To investigate the biochemical changes that occur during PDE5 inhibition in cardiac cells, this study assessed the metabolic profile of the HL1 cell line, a murine atrial cell line with adult cardiomyocyte properties. After one hour of treatment with sildenafil, glycolysis was moderately but selectively stimulated, unlike the pentose phosphate pathway and the Krebs cycle. Moreover, malate and a-Ketoglutarate accumulated, paralleled by a decrease in aspartate and glutamate. Interestingly, increased activity of malate dehydrogenase (MDH) was also detected in these cells after sildenafil treatment. Thus, we hypothesized that sildenafil stimulates the malate-aspartate shuttle (MAS) with the final effect of transferring electrons and protons from glycolysis-derived cytosolic NADH into the matrix for use by the electron transport chain, using malate as an electron carrier. Through this metabolic modification, sildenafil may counteract what is often observed in ischemia, i.e. reduced MAS flux as well as a dramatic acceleration of glycolysis, which switches to lactate production. Additionally, the results observed in HL1 cells were also found in isolated mouse hearts. The documented metabolic alteration in cardiomyocytes upon treatment with sildenafil occurred by stimulating cGMP production, which did not activate PKG (cGMP-PKG signaling), since the addition of DT-2, a PKG inhibitor, did not block malate accumulation and increased MDH activity. Conversely, the addition of chelerythrine, a PKC inhibitor, counteracted both malate accumulation and MAS activation, supporting previous evidence that, upon the addition of sildenafil, some PKC isoforms may be implicated in cardioprotection (cGMP-PKC signaling). Interestingly, an increase in cGMP, driven by sildenafil, another cGMP stimulator such as nitroprusside (SNP), or a C-type natriuretic peptide (CNP) which does not inhibit PDE5, led to MAS stimulation and increased MDH activity.
最近的证据表明，PDE5 抑制在心肌缺血/再灌注损伤、心力衰竭和心肌肥厚中具有心脏保护作用。为了研究 PDE5 抑制在心脏细胞中引起的生化变化，本研究评估了 HL1 细胞系的代谢谱，HL1 细胞系是一种具有成年心肌细胞特性的鼠心房细胞系。在西地那非治疗 1 小时后，与戊糖磷酸途径和三羧酸循环不同，糖酵解被适度但选择性地刺激。此外，苹果酸和α-酮戊二酸积累，伴随着天冬氨酸和谷氨酸的减少。有趣的是，在西地那非处理后，这些细胞中的苹果酸脱氢酶（MDH）活性也增加。因此，我们假设西地那非刺激苹果酸-天冬氨酸穿梭（MAS），最终将电子和质子从糖酵解衍生的细胞质 NADH 转移到基质中，以供电子传递链使用，苹果酸作为电子载体。通过这种代谢修饰，西地那非可能抵消缺血时常观察到的现象，即 MAS 通量减少以及糖酵解急剧加速，从而转向乳酸生成。此外，在 HL1 细胞中观察到的结果也在分离的小鼠心脏中发现。在心肌细胞中用西地那非处理时记录到的代谢改变是通过刺激 cGMP 产生而发生的，这并没有激活 PKG（cGMP-PKG 信号），因为添加 PKG 抑制剂 DT-2 并没有阻止苹果酸积累和增加 MDH 活性。相反，添加 PKC 抑制剂 Chelerythrine 可拮抗苹果酸积累和 MAS 激活，支持先前的证据表明，在用西地那非处理时，一些 PKC 同工型可能参与心脏保护（cGMP-PKC 信号）。有趣的是，由西地那非、另一种 cGMP 刺激物如硝普钠（SNP）或不抑制 PDE5 的 C 型利钠肽（CNP）驱动的 cGMP 增加导致 MAS 刺激和 MDH 活性增加。
