含氮氧化物自由基的氧化还原纳米颗粒可保护神经母细胞瘤 SH-SY5Y 细胞免受 6-羟多巴胺毒性的影响。
Nitroxide Radical-Containing Redox Nanoparticles Protect Neuroblastoma SH-SY5Y Cells against 6-Hydroxydopamine Toxicity.
机构信息
Department of Analytical Biochemistry, Institute of Food Technology and Nutrition, College of Natural Sciences, Rzeszow University, Zelwerowicza Street 4, 35-601 Rzeszow, Poland.
Department of Molecular Biophysics, Faculty of Biology and Environmental Protection, University of Lodz, Pomorska Street 141/143, 90-236 Lodz, Poland.
出版信息
Oxid Med Cell Longev. 2020 Apr 24;2020:9260748. doi: 10.1155/2020/9260748. eCollection 2020.
Parkinson's disease (PD) patients can benefit from antioxidant supplementation, and new efficient antioxidants are needed. The aim of this study was to evaluate the protective effect of selected nitroxide-containing redox nanoparticles (NRNPs) in a cellular model of PD. Antioxidant properties of NRNPs were studied in cell-free systems by protection of dihydrorhodamine 123 against oxidation by 3-morpholino-sydnonimine and protection of fluorescein against bleaching by 2,2-azobis(2-amidinopropane) hydrochloride and sodium hypochlorite. Model blood-brain barrier penetration was studied using hCMEC/D3 cells. Human neuroblastoma SH-SY5Y cells, exposed to 6-hydroxydopamine (6-OHDA), were used as an model of PD. Cells were preexposed to NRNPs or free nitroxides (TEMPO or 4-amino-TEMPO) for 2 h and treated with 6-OHDA for 1 h and 24 h. The reactive oxygen species (ROS) level was estimated with dihydroethidine 123 and Fluorimetric Mitochondrial Superoxide Activity Assay Kit. Glutathione level (GSH) was measured with -phtalaldehyde, ATP by luminometry, changes in mitochondrial membrane potential with JC-1, and mitochondrial mass with 10-Nonyl-Acridine Orange. NRNP1, TEMPO, and 4-amino-TEMPO (25-150 M) protected SH-SY5Y cells from 6-OHDA-induced viability loss; the protection was much higher for NRNP1 than for free nitroxides. NRNP1 were better antioxidants and permeated better the model BBB than free nitroxides. Exposure to 6-OHDA decreased the GSH level after 1 h and increased it considerably after 24 h (apparently a compensatory overresponse); NRNPs and free nitroxides prevented this increase. NRNP1 and free nitroxides prevented the decrease in ATP level after 1 h and increased it after 24 h. 6-OHDA increased the intracellular ROS level and mitochondrial superoxide level. Studied antioxidants mostly decreased ROS and superoxide levels. 6-OHDA decreased the mitochondrial potential and mitochondrial mass; both effects were prevented by NRNP1 and nitroxides. These results suggest that the mitochondria are the main site of 6-OHDA-induced cellular damage and demonstrate a protective effect of NRNP1 in a cellular model of PD.
帕金森病（PD）患者可以从抗氧化补充剂中获益，因此需要新的高效抗氧化剂。本研究的目的是评估选定的含硝氧化物的氧化还原纳米颗粒（NRNP）在 PD 细胞模型中的保护作用。NRNP 的抗氧化特性通过以下方法在无细胞系统中进行研究：用 3-吗啉代-sydnonimine 氧化二氢罗丹明 123 并保护荧光素免于 2,2-偶氮双（2-脒基丙烷）盐酸盐和次氯酸钠漂白，保护二氢罗丹明 123；用 hCMEC/D3 细胞研究模型血脑屏障穿透性。将暴露于 6-羟多巴胺（6-OHDA）的人神经母细胞瘤 SH-SY5Y 细胞用作 PD 模型。细胞先用 NRNPs 或游离硝氧化物（TEMPO 或 4-氨基-TEMPO）预处理 2 小时，然后用 6-OHDA 处理 1 小时和 24 小时。用二氢乙啶 123 和荧光线粒体超氧化物活性测定试剂盒估计活性氧（ROS）水平。用对-羟醛测量谷胱甘肽（GSH）水平，用发光计测量 ATP，用 JC-1 测量线粒体膜电位变化，用 10-壬基吖啶橙测量线粒体质量。NRNP1、TEMPO 和 4-氨基-TEMPO（25-150 μM）可保护 SH-SY5Y 细胞免受 6-OHDA 诱导的活力丧失；与游离硝氧化物相比，NRNP1 的保护作用更高。NRNP1 是更好的抗氧化剂，比游离硝氧化物更好地穿透模型 BBB。暴露于 6-OHDA 会在 1 小时后降低 GSH 水平，在 24 小时后显著增加（显然是一种代偿性过度反应）；NRNP1 和游离硝氧化物可防止这种增加。NRNP1 和游离硝氧化物可防止 1 小时后 ATP 水平降低，并在 24 小时后增加。6-OHDA 增加细胞内 ROS 水平和线粒体超氧化物水平。研究的抗氧化剂大多降低了 ROS 和超氧化物水平。6-OHDA 降低了线粒体电位和线粒体质量；这两种作用均被 NRNP1 和硝氧化物所预防。这些结果表明，线粒体是 6-OHDA 诱导的细胞损伤的主要部位，并证明了 NRNP1 在 PD 细胞模型中的保护作用。
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