Bezine Maryem, Debbabi Meryam, Nury Thomas, Ben-Khalifa Rym, Samadi Mohammad, Cherkaoui-Malki Mustapha, Vejux Anne, Raas Quentin, de Sèze Jérôme, Moreau Thibault, El-Ayeb Mohamed, Lizard Gérard
Univ Bourgogne Franche-Comté, Team 'Biochemistry of the Peroxisome, Inflammation and Lipid Metabolism' EA 7270/INSERM, Dijon, France; Univ. Tunis El Manar-Pasteur Institut, Lab. 'Venoms & Therapeutic Biomolecules', Tunis, Tunisia.
Univ Bourgogne Franche-Comté, Team 'Biochemistry of the Peroxisome, Inflammation and Lipid Metabolism' EA 7270/INSERM, Dijon, France; Univ Monastir, LR12ES05, Lab-NAFS 'Nutrition-Functional Food & Vascular Health', Monastir, Tunisia.
Chem Phys Lipids. 2017 Oct;207(Pt B):135-150. doi: 10.1016/j.chemphyslip.2017.03.006. Epub 2017 Mar 18.
Imbalance in the homeostasis of K ions has been reported to contribute to the pathogenesis of neurodegenerative diseases. 7-ketocholesterol (7KC), 24S-hydroxycholesterol (24S-OHC), and tetracosanoic acid (C24:0), often found at increased levels in patients with Alzheimer's disease, Multiple Sclerosis and X-ALD, are able to trigger numerous nerve cell dysfunctions. We therefore studied the impact of 7KC, 24S-OHC, and C24:0 on 158N murine oligodendrocytes, and determined their impact on K homeostasis. The effects of 7KC, 24S-OHC and C24:0 on lipid membrane organization and membrane potential were examined with merocyanine 540 (MC540) and bis-(1,3-diethylthiobarbituric acid) trimethine oxonol (DiSBAC2(3)), respectively. The intracellular concentration of K ([K]i) was measured by flame photometry and the ratiometric approach using the PBFI-AM fluorescence indicator. To determine the relationships between [K]i and lipotoxicity, 158N cells were pre-treated with a universal Kv channels blocker, 4-aminopyridine (4-AP), without or with 7KC, 24S-OHC or C24:0. Cell adhesion, cell growth, mitochondrial depolarization, cytoplasmic membrane integrity, the presence of SubG1 and the morphological aspect of the nuclei were determined with various microscopy, flow cytometry and biochemistry methods. 7KC, 24S-OHC and C24:0 induced changes in lipid content and polarization of the cytoplasmic membrane. These events were associated with increased [K]i. Blocking Kv channels with 4-AP exacerbated 7KC-, 24S-OHC- and C24:0-induced cell dysfunction. 4-AP exacerbated loss of cell adhesion and cell growth inhibition, amplified mitochondrial depolarization and cytoplasmic membrane damage, and increased the percentage of SubG1 cells. The positive correlation between [K]i and cell death supports the potential involvement of K in 7KC-, 24S-OHC-, and C24:0-induced cytotoxicity.
据报道，钾离子稳态失衡与神经退行性疾病的发病机制有关。在阿尔茨海默病、多发性硬化症和X-连锁肾上腺脑白质营养不良患者中，经常发现7-酮胆固醇（7KC）、24S-羟基胆固醇（24S-OHC）和二十四烷酸（C24:0）水平升高，它们能够引发众多神经细胞功能障碍。因此，我们研究了7KC、24S-OHC和C24:0对158N小鼠少突胶质细胞的影响，并确定了它们对钾稳态的影响。分别用部花青540（MC540）和双（1,3-二乙基硫代巴比妥酸）三甲川草酚（DiSBAC2(3)）检测了7KC、24S-OHC和C24:0对脂质膜组织和膜电位的影响。通过火焰光度法和使用PBFI-AM荧光指示剂的比率法测量细胞内钾浓度（[K]i）。为了确定[K]i与脂毒性之间的关系，用通用的钾通道阻滞剂4-氨基吡啶（4-AP）对158N细胞进行预处理，同时或不同时添加7KC、24S-OHC或C24:0。用各种显微镜、流式细胞术和生化方法测定细胞黏附、细胞生长、线粒体去极化、细胞质膜完整性、亚G1期的存在以及细胞核的形态。7KC、24S-OHC和C24:0诱导脂质含量和细胞质膜极化发生变化。这些事件与[K]i升高有关。用4-AP阻断钾通道会加剧7KC、24S-OHC和C24:0诱导的细胞功能障碍。4-AP加剧了细胞黏附丧失和细胞生长抑制，放大了线粒体去极化和细胞质膜损伤，并增加了亚G1期细胞的百分比。[K]i与细胞死亡之间的正相关支持了钾可能参与7KC、24S-OHC和C24:0诱导的细胞毒性作用。
