17β-Estradiol (E2) is produced from androgens via the action of the enzyme aromatase. E2 is known to be made in neurons in the brain, but the functions of neuron-derived E2 in the ischemic brain are unclear. Here, we used a forebrain neuron-specific aromatase KO (FBN-ARO-KO) mouse model to deplete neuron-derived E2 in the forebrain and determine its roles after global cerebral ischemia. We demonstrated that ovariectomized female FBN-ARO-KO mice exhibited significantly attenuated astrocyte activation, astrocytic aromatization, and decreased hippocampal E2 levels compared with FLOX mice. Furthermore, FBN-ARO-KO mice had exacerbated neuronal damage and worse cognitive dysfunction after global cerebral ischemia. Similar results were observed in intact male mice. RNA-seq analysis revealed alterations in pathways and genes associated with astrocyte activation, neuroinflammation, and oxidative stress in FBN-ARO-KO mice. The compromised astrocyte activation in FBN-ARO-KO mice was associated with robust downregulation of the astrocyte-derived neurotrophic factors, BDNF and IGF-1, as well as the astrocytic glutamate transporter, GLT-1. Neuronal FGF2, which acts in a paracrine manner to suppress astrocyte activation, was increased in FBN-ARO-KO neurons. Interestingly, blocking FGF2 signaling by central injection of FGFR3-neutralizing antibody was able to reverse the diminishment in neuroprotective astrocyte reactivity, and attenuate neuronal damage in FBN-ARO-KO mice. Moreover, in vivo E2 replacement suppressed FGF2 signaling and rescued the compromised reactive astrogliosis and cognitive deficits. Collectively, our data provide novel genetic evidence for a beneficial role of neuron-derived E2 in astrocyte activation, neuroprotection, and cognitive preservation following ischemic injury to the brain.

SIGNIFICANCE STATEMENT Following cerebral ischemia, astrocytes become highly reactive and can exert neuroprotection through the release of neurotrophic factors and clearance of neurotoxic glutamate. The current study advances our understanding of this process by demonstrating that neuron-derived 17β-estradiol (E2) is neuroprotective and critical for induction of reactive astrocytes and their ability to produce astrocyte-derived neurotrophic factors, BDNF and IGF-1, and the glutamate transporter, GLT-1 after ischemic brain damage. These beneficial effects of neuron-derived E2 appear to be due, at least in part, to suppression of neuronal FGF2 signaling, which is a known suppressor of astrocyte activation. These findings suggest that neuron-derived E2 is neuroprotective after ischemic brain injury via a mechanism that involves suppression of neuronal FGF2 signaling, thereby facilitating astrocyte activation.

17β-雌二醇（E2）是通过芳香化酶的作用从雄激素产生的。已知E2是在大脑的神经元中产生的，但尚不清楚缺血性脑中神经元衍生的E2的功能。在这里，我们使用前脑神经元特异性芳香化酶KO（FBN-ARO-KO）小鼠模型消耗前脑中神经元衍生的E2并确定其在全脑缺血后的作用。我们证明，与FLOX小鼠相比，卵巢切除的雌性FBN-ARO-KO小鼠表现出显着减弱的星形胶质细胞活化，星形细胞芳香化和海马E2水平降低。此外，FBN-ARO-KO小鼠在全脑缺血后加剧了神经元损伤和更严重的认知功能障碍。在完整的雄性小鼠中观察到类似的结果。RNA-seq分析揭示了FBN-ARO-KO小鼠中与星形胶质细胞激活，神经炎症和氧化应激相关的途径和基因的改变。FBN-ARO-KO小鼠中星形胶质细胞活化受损与星形胶质细胞衍生的神经营养因子BDNF和IGF-1以及星形细胞谷氨酸转运蛋白GLT-1的强烈下调有关。FBN-ARO-KO神经元中以旁分泌方式抑制星形胶质细胞活化的神经元FGF2增加。有趣的是，通过集中注射FGFR3中和抗体来阻断FGF2信号传导能够逆转神经保护性星形胶质细胞反应性的减弱，并减轻FBN-ARO-KO小鼠的神经元损伤。此外，FBN-ARO-KO小鼠中星形胶质细胞活化受损与星形胶质细胞衍生的神经营养因子BDNF和IGF-1以及星形细胞谷氨酸转运蛋白GLT-1的强烈下调有关。FBN-ARO-KO神经元中以旁分泌方式抑制星形胶质细胞活化的神经元FGF2增多。有趣的是，通过集中注射FGFR3中和抗体来阻断FGF2信号传导能够逆转神经保护性星形胶质细胞反应性的减弱，并减轻FBN-ARO-KO小鼠的神经元损伤。此外，FBN-ARO-KO小鼠中受损的星形胶质细胞活化与星形胶质细胞衍生的神经营养因子BDNF和IGF-1以及星形细胞谷氨酸转运蛋白GLT-1的强烈下调有关。FBN-ARO-KO神经元中以旁分泌方式抑制星形胶质细胞活化的神经元FGF2增加。有趣的是，通过集中注射FGFR3中和抗体来阻断FGF2信号传导能够逆转神经保护性星形胶质细胞反应性的减弱，并减轻FBN-ARO-KO小鼠的神经元损伤。此外，通过集中注射FGFR3中和抗体来阻断FGF2信号传导能够逆转神经保护性星形胶质细胞反应性的减弱，并减轻FBN-ARO-KO小鼠的神经元损伤。此外，通过集中注射FGFR3中和抗体来阻断FGF2信号传导能够逆转神经保护性星形胶质细胞反应性的减弱，并减轻FBN-ARO-KO小鼠的神经元损伤。此外，体内E2替代抑制了FGF2信号传导并挽救了受损的反应性星形胶质增生和认知缺陷。总的来说，我们的数据为神经元衍生的E2在脑缺血性损伤后星形胶质细胞激活，神经保护和认知保存中的有益作用提供了新的遗传证据。

重要性声明脑缺血后，星形胶质细胞变得高度活跃，并可以通过释放神经营养因子和清除神经毒性谷氨酸来发挥神经保护作用。本研究通过证明神经元来源的17β-雌二醇（E2）对诱导反应性星形胶质细胞及其产生星形胶质细胞源性神经营养因子BDNF和IGF-1以及谷氨酸的能力具有神经保护作用和关键作用，从而增进了我们对该过程的理解。转运蛋白，缺血性脑损伤后的GLT-1。神经元来源的E2的这些有益作用似乎至少部分是由于抑制了神经元FGF2信号传导，这是星形胶质细胞活化的已知抑制剂。这些发现表明，神经元衍生的E2通过抑制神经元FGF2信号传导的机制，在缺血性脑损伤后具有神经保护作用，