Elevated hydrostatic pressure triggers mitochondrial fission and decreases cellular ATP in differentiated RGC-5 cells
Elevated hydrostatic pressure triggers mitochondrial fission and decreases cellular ATP in differentiated RGC-5 cells
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DOI：
10.1167/iovs.06-0573
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发表时间：
2007-05-01
影响因子：
4.4
通讯作者：
Weinreb, Robert N.
中科院分区：
文献类型：
--
作者：
Ju, Won-Kyu;Liu, Quan;Weinreb, Robert N.
目的。线粒体裂变是细胞对应激的反应，在神经退行性疾病中的神经细胞死亡中发挥重要作用。本研究的目的是确定升高的静水压是否会诱导培养的视网膜神经节细胞中的线粒体裂变和功能障碍。 RGC-5细胞用琥珀酰刀豆球蛋白A(50μg/mL)分化并转移至加压培养箱中，在其中施加30mmHg的压力1、2或3天。作为对照，来自相同传代的分化细胞在每个时间点在常规培养箱中同时培养。然后用红色荧光线粒体染料标记活 RGC-5 细胞，并通过荧光显微镜和电子显微镜评估线粒体形态。静水压升高后，还通过基于荧光素酶的测定法测量细胞三磷酸腺苷 (ATP) 水平。 结果。线粒体裂变的特征是管状融合线粒体转化为孤立的小细胞器，在静水压升高后 3 天，> 74.3% +/- 1.9% 的线粒体被触发。 3 天后，只有 4.7% +/- 1.4% 的非加压对照细胞显示线粒体裂变。电子显微镜显示，持续 3 天升高的静水压会导致嵴异常耗竭，并缩短线粒体长度。当静水压升高时，裂变连接蛋白 Drp-1 从细胞质转移到线粒体。静水压升高还会导致细胞 ATP 显着且随时间的减少。结论。升高的静水压引发分化的 RGC-5 细胞中的线粒体裂变、异常嵴耗竭、Drp-1 易位和细胞 ATP 减少。增加对调节细胞对高压反应（包括线粒体裂变）的分子机制的了解，可能会为保护 RGC 免受静水压升高的影响提供新的治疗靶点。
PURPOSE. Mitochondrial fission is a cellular response to stress that has an important role in neuronal cell death in neurodegenerative diseases. The purpose of this study was to determine whether elevated hydrostatic pressure induces mitochondrial fission and dysfunction in cultured retinal ganglion cells.METHODS. RGC-5 cells were differentiated with succinyl concanavalin A (50 mu g/mL) and transferred to a pressurized incubator in which 30 mm Hg of pressure was applied for 1, 2, or 3 days. As a control, differentiated cells from an identical passage were incubated simultaneously in a conventional incubator at each of the time points. Live RGC-5 cells were then labeled with a red fluorescent mitochondrial dye and mitochondrial morphology was assessed by fluorescence microscopy and electron microscopy. After elevated hydrostatic pressure, the cellular adenosine triphosphate (ATP) levels were also measured by a luciferase-based assay.RESULTS. Mitochondrial fission, characterized by the conversion of tubular fused mitochondria into isolated small organelles, was triggered in > 74.3% +/- 1.9% of mitochondria at 3 days after elevated hydrostatic pressure. Only 4.7% +/- 1.4% of nonpressurized control cells displayed mitochondrial fission after 3 days. Electron microscopy showed that elevated hydrostatic pressure for 3 days induced abnormal cristae depletion and decreased the length of the mitochondria. On elevation of hydrostatic pressure, the fission-linked protein, Drp-1 was translocated from the cytosol to the mitochondria. Elevated hydrostatic pressure also resulted in a significant, time-dependent reduction of cellular ATP.CONCLUSIONS. Elevated hydrostatic pressure triggered mitochondrial fission, abnormal cristae depletion, Drp-1 translocation, and cellular ATP reduction in differentiated RGC-5 cells. Increased understanding of the molecular mechanisms that regulate the cellular response to elevated pressure including mitochondrial fission may provide new therapeutic targets for protecting RGCs from elevated hydrostatic pressure.