Peleli Maria, Aggeli Ioanna-Katerina, Matralis Alexios N, Kourounakis Angeliki P, Beis Isidoros, Gaitanaki Catherine
Department of Animal and Human Physiology, School of Biology, University of Athens, University Campus, Athens 157 84, Greece.
Department of Medicinal Chemistry, School of Pharmacy, University of Athens, Athens 157 71, Greece.
Bioorg Med Chem. 2015 Feb 1;23(3):390-400. doi: 10.1016/j.bmc.2014.12.046. Epub 2014 Dec 29.
Excessive levels of reactive oxygen species (ROS) result in numerous pathologies including muscle disorders. In essence, skeletal muscle performance of daily activities can be severely affected by the redox imbalances occurring after muscular injuries, surgery, atrophy due to immobilization, dystrophy or eccentric muscle contraction. Therefore, research on the potential beneficial impact of antioxidants is of outmost importance. In this context, aiming at further exploring the mechanisms of action of our newly synthesized antioxidant compounds (AK1 and AK2) in a skeletal muscle experimental setting, we initially investigated their scavenging effect on 2,2-diphenyl-1-picrylhydrazyl (DPPH) and subsequently assessed their effect on the viability of C2 skeletal myoblasts in the presence of two pro-oxidants: H2O2 and curcumin (MTT assay). Interestingly, while both compounds reversed the detrimental effect of H2O2, only AK2 was cytoprotective in curcumin-treated C2 cells. We next confirmed the immediate activation of extracellular signal-regulated kinases (ERKs) and the more delayed activation profile of c-Jun NH2-terminal kinases (JNKs) in C2 skeletal myoblasts exposed to curcumin, by Western blotting. In correlation with the aforementioned results, only AK2 blocked the curcumin-induced activation of JNKs pathway. Furthermore, JNKs were revealed to mediate curcumin-induced apoptosis in C2 cells and only AK2 to effectively suppress it (by detecting its effect on poly(ADP-ribose) polymerase fragmentation). Overall, we have shown that two similar in structure novel antioxidants confer differential effects on C2 skeletal myoblasts viability under oxidative stress conditions. This result may be attributed to these antioxidants respective diverse mode of interaction with the signaling effectors involved in the observed responses. Future studies should further evaluate the mechanism of action of these compounds in order to support their potential application in therapeutic protocols against ROS-related muscle disorders.
活性氧（ROS）水平过高会导致包括肌肉疾病在内的多种病理状况。本质上，日常活动中的骨骼肌表现会受到肌肉损伤、手术、因固定不动导致的萎缩、营养不良或离心性肌肉收缩后发生的氧化还原失衡的严重影响。因此，对抗氧化剂潜在有益影响的研究至关重要。在此背景下，为了在骨骼肌实验环境中进一步探索我们新合成的抗氧化化合物（AK1和AK2）的作用机制，我们首先研究了它们对2,2-二苯基-1-苦基肼基（DPPH）的清除作用，随后在两种促氧化剂H2O2和姜黄素存在的情况下评估了它们对C2骨骼肌成肌细胞活力的影响（MTT法）。有趣的是，虽然两种化合物都逆转了H2O2的有害作用，但只有AK2在姜黄素处理的C2细胞中具有细胞保护作用。接下来，我们通过蛋白质印迹法证实了暴露于姜黄素的C2骨骼肌成肌细胞中细胞外信号调节激酶（ERK）的立即激活以及c-Jun氨基末端激酶（JNK）更延迟的激活模式。与上述结果相关，只有AK2阻断了姜黄素诱导的JNKs通路激活。此外，研究发现JNKs介导姜黄素诱导的C2细胞凋亡，只有AK2能有效抑制它（通过检测其对聚（ADP-核糖）聚合酶片段化的影响）。总体而言，我们已经表明两种结构相似的新型抗氧化剂在氧化应激条件下对C2骨骼肌成肌细胞活力具有不同的影响。这一结果可能归因于这些抗氧化剂与所观察到的反应中涉及的信号效应器各自不同的相互作用模式。未来的研究应进一步评估这些化合物的作用机制，以支持它们在针对ROS相关肌肉疾病的治疗方案中的潜在应用。
