NanoDrugs, S.L. Parque Científico y Tecnológico, Albacete, Spain.
J Neurochem. 2012 Jan;120(2):259-68. doi: 10.1111/j.1471-4159.2011.07556.x. Epub 2011 Nov 24.
Autophagy is an important process which plays a key role in cellular homeostasis by degrading cytoplasmic components in the lysosomes, which facilitates recycling. Alterations to normal autophagy have been linked to excitotoxicity, but the mechanisms governing its signal transduction remain unclear. The aim of this study was to explore the role of autophagy in neuronal excitotoxic death by delivering small interfering RNA (siRNA) to rat cortical neurons, using a dendrimer to silence the autophagy-related gene 6 (beclin 1) and to determine the role of autophagy in excitotoxicity. We have found that the dendrimer is very efficient to deliver siRNA to rat cortical neurons, leading to almost complete removal of the target protein Beclin 1. In addition, NMDA increases autophagy markers, such as the protein levels of Beclin 1, the microtubule-associated light chain 3 (LC3) B-II/LC3B-I ratio, and monodansylcadaverine (MDC) labeling in rat cortical neurons. Moreover, NMDA also increases the formation of autophagosomes observed under a transmission electron microscope. Silencing beclin 1 expression blocked NMDA-induced autophagy. Moreover, Beclin 1 removal potentiated NMDA-induced neuronal death indicating that autophagy plays a protective role during excitotoxicity and suggesting that targeting autophagy might be a helpful therapeutic strategy in neurodegenerative diseases.
自噬是一种重要的过程，通过溶酶体降解细胞质成分在细胞内稳态中起着关键作用，有助于回收利用。正常自噬的改变与兴奋毒性有关，但控制其信号转导的机制仍不清楚。本研究旨在通过向大鼠皮质神经元递送小干扰 RNA（siRNA），使用树枝状聚合物沉默自噬相关基因 6（beclin 1），探索自噬在神经元兴奋毒性死亡中的作用，并确定自噬在兴奋毒性中的作用。我们发现树枝状聚合物非常有效地将 siRNA 递送至大鼠皮质神经元，导致靶蛋白 Beclin 1 几乎完全被去除。此外，NMDA 增加自噬标志物，如 Beclin 1 的蛋白水平、微管相关轻链 3（LC3）B-II/LC3B-I 比值和单丹磺酰尸胺（MDC）标记在大鼠皮质神经元中的表达。此外，NMDA 还增加了透射电子显微镜下观察到的自噬体的形成。沉默 beclin 1 表达阻断了 NMDA 诱导的自噬。此外，Beclin 1 的去除增强了 NMDA 诱导的神经元死亡，表明自噬在兴奋毒性中起保护作用，并提示靶向自噬可能是神经退行性疾病的一种有帮助的治疗策略。
