Unitat de Farmacologia i Farmacognosia, Institut de Biomedicina, Centro de Investigacion Biomedica en Red de Enfermedades Neurodegenerativas, Facultat de Farmacia, Universitat de Barcelona, Nucli Universitari de Pedralbes, Barcelona, Spain.
Free Radic Res. 2009 Oct;43(10):985-94. doi: 10.1080/10715760903159188. Epub 2009 Aug 5.
Reactive oxygen species and oxidative stress are associated with neuronal cell death in many neurodegenerative conditions. However, the exact molecular mechanisms triggered by oxidative stress in neurodegeneration are still unclear. This study used the B65 rat neuroblastoma cell line as a model to study the molecular events that occur after H(2)O(2) treatment. Treatment of B65 cells with H(2)O(2) rapidly up-regulated the DNA damage pathway involved in double-strand breakage. Subsequently, proteins involved in p53 regulation, such as sirtuin 1 and STAT1, were modified. In addition, H(2)O(2) treatment altered the pattern of cell cycle protein expression. Specifically, a decrease was found in the expression of cyclin D1, cdk4 and surprisingly the levels of cyclin A and the retinoblastoma protein phosphorylated at ser780 were increased. Furthermore, this study shows that pre-treatment of B65 cells with 50 microM trolox confers almost total protection against apoptotic cell death and restores the cell cycle. Likewise, the increase in retinoblastoma phosphorylation was attenuated by KU-55993, a selective ATM inhibitor, and also by trolox. These observations indicate that DNA damage and oxidative stress are responsible for cell cycle regulation. In summary, this study describes the molecular mechanisms involved in cell cycle alterations induced by oxidative stress in B65 cells. These findings highlight the relevance of ATM in the regulation of cell cycle after oxidative stress.
活性氧和氧化应激与许多神经退行性疾病中的神经元细胞死亡有关。然而，氧化应激在神经退行性变中引发的确切分子机制仍不清楚。本研究使用 B65 大鼠神经母细胞瘤细胞系作为模型，研究了 H2O2 处理后发生的分子事件。用 H2O2 处理 B65 细胞可迅速上调涉及双链断裂的 DNA 损伤途径。随后，p53 调节相关的蛋白质，如 SIRT1 和 STAT1，被修饰。此外，H2O2 处理改变了细胞周期蛋白表达模式。具体来说，发现细胞周期蛋白 D1、cdk4 的表达减少，而令人惊讶的是，细胞周期蛋白 A 和磷酸化丝氨酸 780 的视网膜母细胞瘤蛋白的水平增加。此外，本研究表明，用 50 μM trolox 预处理 B65 细胞可提供几乎完全的抗细胞凋亡死亡保护，并恢复细胞周期。同样，KU-55993（一种选择性 ATM 抑制剂）和 trolox 也可减弱视网膜母细胞瘤磷酸化的增加。这些观察结果表明，DNA 损伤和氧化应激是细胞周期调节的原因。总之，本研究描述了氧化应激诱导的 B65 细胞细胞周期改变所涉及的分子机制。这些发现强调了 ATM 在氧化应激后细胞周期调节中的相关性。
