Smad proteins are known to transduce the actions of the transforming growth factor-β (TGF-β) family including TGF-βs, activins, and bone morphogenetic proteins (BMPs). We previously reported that Smad1/5/9 immunoreactivity was observed in astrocytes of various rat brain regions including the hippocampus, suggesting that Smad1/5/9 may be associated with the physiology of astrocytes. However, the Smad1/5/9 expression and activation in the hippocampal astrocytes after global cerebral ischemia has not been yet elucidated. In this study, we examined temporal changes in the expression and phosphorylation of Smad1/5/9 in the hippocampus using a rat model of global cerebral ischemia. Furthermore, we examined the candidate ligand involved in the phosphorylation of Smad1/5/9 in the hippocampus after ischemia. Pyramidal neuronal cell death in the CA1 regions was visible at 3 days, and maximum death occurred within 7 days after ischemia. At 7 days after ischemia, astrocytes that showed strong immunoreactivity for Smad1/5/9 were frequently observed in the CA1 region. Additionally, there was an increase in phosphorylated Smad1/5/9 (phospho-Smad1/5/9) -immunopositive astrocytes in the CA1 region 7 days after ischemia. Real-time PCR analysis showed an increase in the expression level of TGF-β1 mRNA in the hippocampus after ischemia. Intracerebroventricular injection of SB525334, an inhibitor of TGF-β/Smad signaling, reduced immunoreactivity for phospho-Smad1/5/9 in astrocytes. These results suggest that TGF-β1 may be a key molecule for ischemia-induced Smad1/5/9 phosphorylation in astrocytes, and TGF-β1-Smad1/5/9 signaling may play a role in post-ischemic events, including brain inflammation or tissue repair rather than neuroprotection of the hippocampus.

已知 Smad 蛋白可转导转化生长因子-β (TGF-β) 家族的作用，包括 TGF-β、激活素和骨形态发生蛋白 (BMP)。我们之前报道过在包括海马在内的不同大鼠大脑区域的星形胶质细胞中观察到 Smad1/5/9 免疫反应性，这表明 Smad1/5/9 可能与星形胶质细胞的生理机能有关。然而，尚未阐明全脑缺血后海马星形胶质细胞中 Smad1/5/9 的表达和激活。在这项研究中，我们使用全脑缺血大鼠模型检查了海马中 Smad1/5/9 表达和磷酸化的时间变化。此外，我们检查了参与缺血后海马中 Smad1/5/9 磷酸化的候选配体。CA1 区域的锥体神经元细胞死亡在第 3 天可见，最大死亡发生在缺血后 7 天内。缺血后 7 天，在 CA1 区经常观察到对 Smad1/5/9 表现出强烈免疫反应性的星形胶质细胞。此外，在缺血 7 天后，CA1 区域中磷酸化的 Smad1/5/9 (phospho-Smad1/5/9) 免疫阳性星形胶质细胞有所增加。Real-time PCR 分析显示缺血后海马中 TGF-β1 mRNA 的表达水平增加。脑室内注射 SB525334（一种 TGF-β/Smad 信号传导抑制剂）可降低星形胶质细胞中磷酸-Smad1/5/9 的免疫反应性。这些结果表明 TGF-β1 可能是星形胶质细胞缺血诱导的 Smad1/5/9 磷酸化的关键分子，