Cigarette smoke induced autophagy-impairment regulates AMD pathogenesis mechanisms in ARPE-19 cells.
Cigarette smoke induced autophagy-impairment regulates AMD pathogenesis mechanisms in ARPE-19 cells.
复制标题
香烟烟雾诱导的自噬损伤调节 ARPE-19 细胞中的 AMD 发病机制。
DOI：
10.1371/journal.pone.0182420
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Vij N
中科院分区：
文献类型：
--
作者：
Govindaraju VK;Bodas M;Vij N
年龄相关性黄斑变性（AMD）是致盲的主要原因之一。遗传、环境损害和年龄相关因素都在改变蛋白质平衡（调节蛋白质合成、降解和加工的稳态过程）中发挥关键作用。这些因素也在AMD的发病机制中发挥作用，并且已经确定吸烟（CS）启动AMD的发病机制。本研究的主要目的是阐明CS是否可以诱导视网膜色素上皮（RPE）细胞的蛋白质停滞/自噬损伤。在初步分析中，我们发现香烟烟雾提取物（CSE）诱导了不溶性蛋白质部分中泛素化蛋白的积累（p < 0.01），随后通过半胱胺（p < 0.01）或非瑟酮（p < 0.05）处理减轻了泛素化蛋白的积累。进一步证实，这些CSE诱导的泛素化蛋白在核周围空间积累（p<0.05），并被半胱胺（p<0.05）或非赛特酮（p<0.05）清除。此外，半胱胺（p<0.05）或非瑟酮（p<0.05）处理显著减轻了cse诱导的聚合体形成（LC3B-GFP和Ub-RFP共定位）和自噬通量损伤（p<0.01），表明cse介导的自噬损伤恢复。CSE处理还可以诱导细胞内活性氧（ROS, p < 0.001），同时影响细胞活力（p < 0.001），分别用CMH2DCFDA-dye （ROS）和MTS（增殖）或propodium iodiine染色（细胞活力）测定。此外，半胱胺和非瑟酮处理改善了cs介导的ROS生成（p < 0.05），降低了细胞活力（p < 0.05）。最后，发现CSE诱导细胞衰老（p < 0.001），半胱胺（p < 0.001）或非瑟酮（p < 0.001）显著改善细胞衰老。总之，我们的研究表明，CS诱导的蛋白质停滞/自噬损伤调节了与AMD发病相关的机制。此外，自噬诱导药物如半胱胺或非瑟酮可以改善AMD的发病机制，值得在临床前小鼠模型中进一步研究。
Age related macular degeneration (AMD) is one of the leading causes of blindness. Genetics, environmental insult, and age-related factors all play a key role in altering proteostasis, the homeostatic process regulating protein synthesis, degradation and processing. These factors also play a role in the pathogenesis of AMD and it has been well established that cigarette smoking (CS) initiates AMD pathogenic mechanisms. The primary goal of this study is to elucidate whether CS can induce proteostasis/autophagy-impairment in retinal pigment epithelial (RPE) cells. In our preliminary analysis, it was found that cigarette smoke extract (CSE) induces accumulation of ubiquitinated proteins in the insoluble protein fraction (p < 0.01), which was subsequently mitigated through cysteamine (p < 0.01) or fisetin (p < 0.05) treatment. Further, it was verified that these CSE induced ubiquitinated proteins accumulated in the peri-nuclear spaces (p<0.05) that were cleared- off with cysteamine (p < 0.05) or fisetin (p < 0.05). Moreover, CSE-induced aggresome-formation (LC3B-GFP and Ub-RFP co-localization) and autophagy-flux impairment was significantly (p<0.01) mitigated by cysteamine (p<0.05) or fisetin (p<0.05) treatment, indicating the restoration of CSE-mediated autophagy-impairment. CSE treatment was also found to induce intracellular reactive oxygen species (ROS, p < 0.001) while impacting cell viability (p < 0.001), which was quantified using CMH2DCFDA-dye (ROS) and MTS (proliferation) or propodium iodide staining (cell viability) assays, respectively. Moreover, cysteamine and fisetin treatment ameliorated CS-mediated ROS production (p < 0.05) and diminished cell viability (p < 0.05). Lastly, CSE was found to induce cellular senescence (p < 0.001), which was significantly ameliorated by cysteamine (p < 0.001) or fisetin (p < 0.001). In conclusion, our study indicates that CS induced proteostasis/autophagy-impairment regulates mechanisms associated with AMD pathogenesis. Moreover, autophagy-inducing drugs such as cysteamine or fisetin can ameliorate AMD pathogenesis mechanisms that warrant further investigation in pre-clinical murine models.
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影响因子：
3.7
作者：
Fujihara, Masashi;Nagai, Norihiro;Sussan, Thomas E.;Biswal, Shyam;Handa, James T.
通讯作者：
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影响因子：
3.5
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