Allani Pramod K, Sum Tak, Bhansali Suraj G, Mukherjee Suman K, Sonee Manisha
Pharmaceutical Sciences, College of Pharmacy, South Dakota State University, Brookings, SD 57007, USA.
Toxicol Appl Pharmacol. 2004 Apr 1;196(1):29-36. doi: 10.1016/j.taap.2003.12.010.
Cytoskeleton disruption is a process by which oxidative stress disrupts cellular function. This study compares and contrasts the effect of oxidative stress on the three major cytoskeleton filaments, microfilaments (MFs), microtubule (MT), and vimentin in human cortical neuronal cell line (HCN2). HCN2 cells were treated with 100 microM tertiary butylhydroperoxide (t-BuOOH), a free radical generating neurotoxin for 1, 3, or 6 h. Cell viability studies demonstrated significant cell death although the morphology studies showed that there was a substantial loss in neurites of neurons treated with t-BuOOH for 6 h. Because the cytoskeleton plays a role in neurite outgrowth, the effect of oxidative stress on the cytoskeletal was studied. In neurons subjected to oxidative stress for 30 min or 1 h, there were no major changes in microfilament distribution though there was altered distribution of microtubule and vimentin filaments as compared to controls. However, loss and disruption of all the three cytoskeletal filaments was observed at later times (3 and 6 h), which was confirmed by Western Blot analysis. Further studies were done to measure the gene expression levels of actin, tubulin, and vimentin. Results indicated that the overall loss of the cytoskeletal proteins in neurons treated with free radical generating toxin might not be a direct result of the downregulation of the cytoskeletal genes. This study shows that free radical generation in human neurons leads to the disruption of the cytoskeleton, though there may be a difference in the susceptibility to oxidative stress among the individual components of the cytoskeletal filaments.
细胞骨架破坏是氧化应激破坏细胞功能的一个过程。本研究比较并对比了氧化应激对人皮质神经元细胞系（HCN2）中三种主要细胞骨架丝，即微丝（MFs）、微管（MT）和波形蛋白的影响。HCN2细胞用100微摩尔叔丁基过氧化氢（t-BuOOH，一种产生自由基的神经毒素）处理1、3或6小时。细胞活力研究表明存在显著的细胞死亡，尽管形态学研究显示，用t-BuOOH处理6小时的神经元的神经突有大量损失。由于细胞骨架在神经突生长中起作用，因此研究了氧化应激对细胞骨架的影响。在经受氧化应激30分钟或1小时的神经元中，微丝分布没有重大变化，尽管与对照相比，微管和波形蛋白丝的分布发生了改变。然而，在后期（3小时和6小时）观察到所有三种细胞骨架丝的丢失和破坏，这通过蛋白质免疫印迹分析得到证实。进一步开展研究以测量肌动蛋白、微管蛋白和波形蛋白的基因表达水平。结果表明，用产生自由基的毒素处理的神经元中细胞骨架蛋白的总体丢失可能不是细胞骨架基因下调的直接结果。本研究表明，人神经元中自由基的产生会导致细胞骨架的破坏，尽管细胞骨架丝的各个组分对氧化应激的敏感性可能存在差异。
