Independent Laboratory of Genetics and Molecular Biology, Kaczkowski Military Institute of Hygiene and Epidemiology, Kozielska 4, 01-163 Warsaw, Poland.
Tumor Cell Death Laboratory, Cancer Research UK, Beatson Institute, Garscube Estate, Switchback Road, Glasgow G61 1BD, UK.
Int J Mol Sci. 2019 Mar 24;20(6):1481. doi: 10.3390/ijms20061481.
In this study we attempted to verify the hypothesis that the mevalonate pathway affects amyloid beta precursor protein (AβPP) processing and regulates clusterin protein levels. expression was monitored by green fluorescence (FL) and Western blot (WB). WB showed soluble amyloid protein precursor alpha (sAβPPα) presence in -wt cells and Aβ expression in -sw cells. Nerve growth factor (NGF)-differentiated rat neuronal pheochromocytoma PC-12 cells were untreated/treated with statins alone or together with non-sterol isoprenoids. Co-treatment with mevalonate, dolichol, ubiquinol, farnesol, geranylgeraniol, or water-soluble cholesterol demonstrated statin-dependent neurotoxicity resulted from the attenuated activity of mevalonate pathway rather than lower cholesterol level. Atorvastatin (50 μM) or simvastatin (50 μM) as well as cholesterol chelator methyl-β-cyclodextrin (0.2 mM) diminished cell viability ( < 0.05) and clusterin levels. Interestingly, co-treatment with mevalonate, dolichol, ubiquinol, farnesol, geranylgeraniol, or water-soluble cholesterol stimulated ( < 0.05) clusterin expression. Effects of non-sterol isoprenoids, but not water soluble cholesterol (Chol-PEG), were the most significant in mock-transfected cells. Geranylgeraniol (GGOH) overcame atorvastatin (ATR)-dependent cytotoxicity. This effect does not seem to be dependent on clusterin, as its level became lower after GGOH. The novelty of these findings is that they show that the mevalonate (MEV) pathway rather than cholesterol itself plays an important role in clusterin expression levels. In mock-transfected, rather than in AβPP-overexpressing cells, GGOH/farnesol (FOH) exerted a protective effect. Thus, protein prenylation with GGOH/FOH might play substantial role in neuronal cell survival.
在这项研究中，我们试图验证这样一个假设，即甲羟戊酸途径会影响淀粉样前体蛋白（AβPP）的加工，并调节簇蛋白的水平。通过绿色荧光（FL）和 Western blot（WB）监测表达。WB 显示在 -wt 细胞中存在可溶性淀粉样蛋白前体 alpha（sAβPPα），在 -sw 细胞中存在 Aβ表达。未处理/单独用他汀类药物或与非甾体异戊二烯一起处理神经生长因子（NGF）分化的大鼠嗜铬细胞瘤 PC-12 细胞。与甲羟戊酸、鲨烯醇、泛醌、法呢醇、香叶醇或水溶性胆固醇一起进行共处理表明，他汀类药物依赖性神经毒性是由甲羟戊酸途径活性减弱引起的，而不是胆固醇水平降低。阿托伐他汀（50 μM）或辛伐他汀（50 μM）以及胆固醇螯合剂甲基-β-环糊精（0.2 mM）降低细胞活力（<0.05）和簇蛋白水平。有趣的是，与甲羟戊酸、鲨烯醇、泛醌、法呢醇、香叶醇或水溶性胆固醇共处理刺激（<0.05）簇蛋白表达。非甾体异戊二烯的作用，但不是水溶性胆固醇（Chol-PEG），在模拟转染细胞中最为显著。香叶醇（GGOH）克服了阿托伐他汀（ATR）依赖性细胞毒性。这种作用似乎不依赖于簇蛋白，因为在 GGOH 之后其水平降低。这些发现的新颖之处在于它们表明甲羟戊酸（MEV）途径而不是胆固醇本身在簇蛋白表达水平中起着重要作用。在模拟转染细胞中，而不是在 AβPP 过表达细胞中，GGOH/法呢醇（FOH）发挥了保护作用。因此，GGOH/FOH 的蛋白质 prenylation 可能在神经元细胞存活中发挥重要作用。
