Babylon Lukas, Grewal Rekha, Stahr Pascal-L, Eckert Ralph W, Keck Cornelia M, Eckert Gunter P
Biomedical Research Center Seltersberg (BFS), Laboratory for Nutrition in Prevention and Therapy, Institute of Nutritional Sciences, Justus Liebig University, Schubertstr. 81, 35392 Giessen, Germany.
Department of Pharmaceutics and Biopharmaceutics, Philipps Universität, Robert-Koch-Str. 4, 35037 Marburg, Germany.
Antioxidants (Basel). 2021 Jun 23;10(7):1003. doi: 10.3390/antiox10071003.
Mitochondrial dysfunction represents a hallmark of both brain aging and age-related neurodegenerative disorders including Alzheimer disease (AD). AD-related mitochondrial dysfunction is characterized by an impaired electron transport chain (ETC), subsequent decreased adenosine triphoshpate (ATP) levels, and elevated generation of reactive oxygen species (ROS). The bioactive citrus flavanone hesperetin (Hst) is known to modulate inflammatory response, to function as an antioxidant, and to provide neuroprotective properties. The efficacy in improving mitochondrial dysfunction of Hst nanocrystals (HstN) with increased bioavailability has not yet been investigated. Human SH-SY5Y cells harboring neuronal amyloid precursor protein (APP) acted as a model for the initial phase of AD. MOCK-transfected cells served as controls. The energetic metabolite ATP was determined using a luciferase-catalyzed bioluminescence assay. The activity of mitochondrial respiration chain complexes was assessed by high-resolution respirometry using a Clarke electrode. Expression levels of mitochondrial respiratory chain complex genes were determined using quantitative real-time polymerase chain reaction (qRT-PCR). The levels of amyloid β-protein (Aβ) were measured using homogeneous time-resolved fluorescence (HTRF). ROS levels, peroxidase activity, and cytochrome c activity were determined using a fluorescence assay. Compared to pure Hst dissolved in ethanol (HstP), SH-SY5Y-APP cells incubated with HstN resulted in significantly reduced mitochondrial dysfunction: ATP levels and respiratory chain complex activity significantly increased. Gene expression levels of RCC I, IV, and V were significantly upregulated. In comparison, the effects of HstN on SY5Y-MOCK control cells were relatively small. Pure Hst dissolved in ethanol (HstP) had almost no effect on both cell lines. Neither HstN nor HstP led to significant changes in Aβ levels. HstN and HstP were both shown to lower peroxidase activity significantly. Furthermore, HstN significantly reduced cytochrome c activity, whereas HstP had a significant effect on reducing ROS in SH-SY5Y-APP cells. Thus, it seems that the mechanisms involved may not be linked to altered Aβ production. Nanoflavonoids such as HstN have the potential to prevent mitochondria against dysfunction. Compared to its pure form, HstN showed a greater effect in combatting mitochondrial dysfunction. Further studies should evaluate whether HstN protects against age-related mitochondrial dysfunction and thus may contribute to late-onset AD.
线粒体功能障碍是大脑衰老和包括阿尔茨海默病（AD）在内的与年龄相关的神经退行性疾病的一个标志。与AD相关的线粒体功能障碍的特征是电子传递链（ETC）受损、随后三磷酸腺苷（ATP）水平降低以及活性氧（ROS）生成增加。生物活性柑橘类黄酮橙皮素（Hst）已知可调节炎症反应、作为抗氧化剂发挥作用并具有神经保护特性。具有提高生物利用度的Hst纳米晶体（HstN）改善线粒体功能障碍的功效尚未得到研究。携带神经元淀粉样前体蛋白（APP）的人SH-SY5Y细胞用作AD初始阶段的模型。转染空载体的细胞用作对照。使用荧光素酶催化的生物发光测定法测定能量代谢物ATP。使用克拉克电极通过高分辨率呼吸测定法评估线粒体呼吸链复合物的活性。使用定量实时聚合酶链反应（qRT-PCR）测定线粒体呼吸链复合物基因的表达水平。使用均相时间分辨荧光（HTRF）测量淀粉样β蛋白（Aβ）的水平。使用荧光测定法测定ROS水平、过氧化物酶活性和细胞色素c活性。与溶解在乙醇中的纯Hst（HstP）相比，用HstN处理的SH-SY5Y-APP细胞导致线粒体功能障碍显著减轻：ATP水平和呼吸链复合物活性显著增加。RCC I、IV和V的基因表达水平显著上调。相比之下，HstN对SY5Y-MOCK对照细胞的影响相对较小。溶解在乙醇中的纯Hst（HstP）对这两种细胞系几乎没有影响。HstN和HstP均未导致Aβ水平发生显著变化。HstN和HstP均显示出显著降低过氧化物酶活性。此外，HstN显著降低细胞色素c活性，而HstP对降低SH-SY5Y-APP细胞中的ROS有显著作用。因此，似乎所涉及的机制可能与Aβ生成的改变无关。诸如HstN之类的纳米类黄酮有潜力防止线粒体功能障碍。与其纯形式相比，HstN在对抗线粒体功能障碍方面显示出更大的效果。进一步的研究应评估HstN是否能预防与年龄相关的线粒体功能障碍，从而可能对晚发性AD有作用。
