Institute of Neuroscience, Brain Research Center, National Yang-Ming University, Taipei 11221, Taiwan.
J Neuroinflammation. 2010 Apr 29;7:28. doi: 10.1186/1742-2094-7-28.
Overactivated microglia that cluster at neuritic plaques constantly release neurotoxins, which actively contribute to progressive neurodegeneration in Alzheimer's disease (AD). Therefore, attenuating microglial clustering can reduce focal neuroinflammation at neuritic plaques. Previously, we identified CCL5 and CCL2 as prominent chemokines that mediate the chemotaxis of microglia toward beta-amyloid (Abeta)aggregates. Although transforming growth factor-beta1 (TGF-beta1) has been shown to down-regulate the expression of chemokines in activated microglia, whether TGF-beta1 can reduce the chemotaxis of microglia toward neuritic plaques in AD remains unclear.
In the present study, we investigated the effects of TGF-beta1 on Abeta-induced chemotactic migration of BV-2 microglia using time-lapse recording, transwell assay, real-time PCR, ELISA, and western blotting.
The cell tracing results suggest that the morphological characteristics and migratory patterns of BV-2 microglia resemble those of microglia in slice cultures. Using this model system, we discovered that TGF-beta1 reduces Abeta-induced BV-2 microglial clustering in a dose-dependent manner. Chemotactic migration of these microglial cells toward Abeta aggregates was significantly attenuated by TGF-beta1. However, these microglia remained actively moving without any reduction in migration speed. Pharmacological blockade of TGF-beta1 receptor I (ALK5) by SB431542 treatment reduced the inhibitory effects of TGF-beta1 on Abeta-induced BV-2 microglial clustering, while preventing TGF-beta1-mediated cellular events, including SMAD2 phosphorylation and CCL5 down-regulation.
Our results suggest that TGF-beta1 reduces Abeta-induced microglial chemotaxis via the SMAD2 pathway. The down-regulation of CCL5 by TGF-beta1 at least partially contributes to the clustering of microglia at Abeta aggregates. The attenuating effects of SB431542 upon TGF-beta1-suppressed microglial clustering may be mediated by restoration of CCL5 to normal levels. TGF-beta1 may ameliorate microglia-mediated neuroinflammation in AD by preventing activated microglial clustering at neuritic plaques.
过度激活的小胶质细胞在神经突斑块处聚集，并不断释放神经毒素，这积极促成了阿尔茨海默病（AD）的进行性神经退行性变。因此，减轻小胶质细胞的聚集可以减少神经突斑块处的局灶性神经炎症。以前，我们发现 CCL5 和 CCL2 是介导小胶质细胞向β-淀粉样蛋白（Abeta）聚集物趋化的主要趋化因子。尽管转化生长因子-β1（TGF-β1）已被证明可下调激活的小胶质细胞中趋化因子的表达，但 TGF-β1 是否可以减少 AD 中神经突斑块处小胶质细胞的趋化性尚不清楚。
在本研究中，我们使用延时记录、Transwell 测定、实时 PCR、ELISA 和 Western blot 研究了 TGF-β1 对 Abeta 诱导的 BV-2 小胶质细胞趋化性迁移的影响。
细胞示踪结果表明，BV-2 小胶质细胞的形态特征和迁移模式类似于切片培养中的小胶质细胞。使用该模型系统，我们发现 TGF-β1 以剂量依赖性方式减少 Abeta 诱导的 BV-2 小胶质细胞聚集。TGF-β1 显著减弱了这些小胶质细胞向 Abeta 聚集物的趋化性迁移。然而，这些小胶质细胞仍然保持活跃的运动，而没有任何迁移速度的降低。用 SB431542 抑制 TGF-β1 受体 I（ALK5）的药理阻断降低了 TGF-β1 对 Abeta 诱导的 BV-2 小胶质细胞聚集的抑制作用，同时阻止了 TGF-β1 介导的细胞事件，包括 SMAD2 磷酸化和 CCL5 的下调。
我们的结果表明，TGF-β1 通过 SMAD2 途径减少 Abeta 诱导的小胶质细胞趋化性。TGF-β1 下调 CCL5 至少部分导致小胶质细胞在 Abeta 聚集物处聚集。SB431542 对 TGF-β1 抑制的小胶质细胞聚集的减弱作用可能是通过将 CCL5 恢复到正常水平介导的。TGF-β1 可通过防止神经突斑块处激活的小胶质细胞聚集来改善 AD 中的小胶质细胞介导的神经炎症。
