Department of Biochemistry, College of Medicine, The Catholic University of Korea, Seoul 137-701, Korea.
Korean J Physiol Pharmacol. 2011 Jun;15(3):149-56. doi: 10.4196/kjpp.2011.15.3.149. Epub 2011 Jun 30.
Golgi SNAP receptor complex 1 (GS28) has been implicated in vesicular transport between intra-Golgi networks and between endoplasmic reticulum (ER) and Golgi. Additional role(s) of GS28 within cells have not been well characterized. We observed decreased expression of GS28 in rat ischemic hippocampus. In this study, we examined the role of GS28 and its molecular mechanisms in neuronal (SK-N-SH) cell death induced by hydrogen peroxide (H(2)O(2)). GS28 siRNA-transfected cells treated with H(2)O(2) showed a significant increase in cytotoxicity under glutathione (GSH)-depleted conditions after pretreatment with buthionine sulfoximine, which corresponded to an increase of intracellular reactive oxygen species (ROS) in the cells. Pretreatment of GS28 siRNA-transfected cells with p38 chemical inhibitor significantly inhibited cytotoxicity; we also observed that p38 was activated in the cells by immunoblot analysis. We confirmed the role of p38 MAPK in cotransfected cells with GS28 siRNA and p38 siRNA in the cell viability assay, flow cytometry, and immunoblot. Involvement of apoptotic or autophagic processes in the cells was not shown in the cell viability, flow cytometry, and immunoblot analyses. However, pretreatment of the cells with necrostatin-1 completely inhibited H(2)O(2)-induced cytotoxicity, ROS generation, and p38 activation, indicating that the cell death is necroptotic. Collectively these data imply that H(2)O(2) induces necroptotic cell death in the GS28 siRNA-transfected cells and that the necroptotic signals are mediated by sequential activations in RIP1/p38/ROS. Taken together, these results indicate that GS28 has a protective role in H(2)O(2)-induced necroptosis via inhibition of p38 MAPK in GSH-depleted neuronal cells.
高尔基 SNAP 受体复合物 1（GS28）已被牵连到内高尔基网络和内质网（ER）与高尔基体之间的囊泡运输中。GS28 在细胞内的其他作用尚未得到很好的描述。我们观察到在大鼠缺血性海马中 GS28 的表达降低。在这项研究中，我们研究了 GS28 及其分子机制在过氧化氢（H2O2）诱导的神经元（SK-N-SH）细胞死亡中的作用。在用谷胱甘肽（GSH）耗尽预处理后，用丁硫氨酸亚砜胺处理的转染了 GS28 siRNA 的细胞在 H2O2 处理下显示出明显的细胞毒性增加，这与细胞内活性氧（ROS）的增加相对应。用 p38 化学抑制剂预处理转染了 GS28 siRNA 的细胞显著抑制了细胞毒性；我们还通过免疫印迹分析观察到 p38 在细胞中被激活。我们在细胞活力测定、流式细胞术和免疫印迹中证实了 p38 MAPK 在转染了 GS28 siRNA 和 p38 siRNA 的共转染细胞中的作用。在细胞活力、流式细胞术和免疫印迹分析中，未显示凋亡或自噬过程参与细胞。然而，在用 necrostatin-1 预处理细胞后，完全抑制了 H2O2 诱导的细胞毒性、ROS 生成和 p38 激活，表明细胞死亡是坏死性的。总之，这些数据表明 H2O2 诱导转染了 GS28 siRNA 的细胞发生坏死性细胞死亡，并且坏死性信号是通过 RIP1/p38/ROS 的顺序激活来介导的。综上所述，这些结果表明 GS28 通过抑制 GSH 耗尽的神经元细胞中的 p38 MAPK 在 H2O2 诱导的坏死性细胞死亡中起保护作用。
