Progranulin通过TNFR参与焦虑发生发展的分子机制研究

Anxiety is one of the most common psychiatric disorders. Because of its complex pathogenesis, the existing anxiolytic drugs cannot meet the clinical requirements. Therefore, it is urgent to explore new therapeutic strategies for clinical treatment. Progranulin (PGRN) is a multifunctional secreted growth factor, which is widely expressed in the central nervous system. PGRN as a new ligand of tumor necrosis factor receptor (TNFR) has attracted much attention. Our previous study found that PGRN highly expressed in the anxiety-related amygdala region. Amygdala local infusion of PGRN could significantly inhibit the peripheral inflammation induced anxiety like behavior. Morphological studies found that PGRN significantly enhanced the development of neural spines, increased the percentage of mature spines process significantly, and enhanced cortex-amygdala synaptic plasticity. These results prompt the potential value of PGRN in the treatment of anxiety. It is the first time to propose that PGRN regulates the inhibitory and excitatory synaptic transmission in the amygdala, which may be a new target of anxiolytic drugs through binding TNFR. Multiple methods such as genetic modulation, molecular biology, electrophysiology, and behavioral test will be used to explore the role of PGRN in amygdala neural circuits. This project will be helpful to elucidate the mechanisms underlying the pathogenesis of anxiety disorders and to provide a theoretical basis for target of novel anxiolytics.

焦虑症发病机制复杂，现有药物尚不能满足临床需要。因此，发掘新的治疗策略成为临床迫切需求。颗粒蛋白前体（Progranulin，PGRN）是一种多功能的分泌性生长因子，广泛表达于中枢，参与中枢炎症调节。PGRN 作为肿瘤坏死因子受体（TNFR）的一种新配体而备受关注。我们前期研究发现，与焦虑相关的杏仁核区域高表达PGRN；杏仁核局部给予PGRN可显著抑制外周炎症引起的小鼠焦虑样行为；形态学研究发现PGRN可显著增强杏仁核神经棘突的发育，成熟型棘突显著增加，并增强皮层-杏仁核通路突触可塑性；提示PGRN在焦虑治疗中的潜在价值。本项目首次提出PGRN能够快速调节杏仁核兴奋性/抑制性突触传递平衡，通过TNFR干扰中枢炎症反应，可望成为抗焦虑药物作用的新靶点。本研究拟采用基因干涉、电生理和行为学等方法，探讨PGRN对杏仁核神经环路的调节，为阐明焦虑症发病机制及发掘药物新靶点提供理论依据。

Progranulin（PGRN）由GRN基因编码，为Granulin的前体蛋白，是一个关键的中枢抗炎因子和神经营养因子。我们利用脆性X综合征（FXS）模型小鼠，即FMR1基因敲除（KO）小鼠，发现此小鼠脑中PGRN水平显著升高，利用shRNA降低PGRN可显著改善前扣带回皮层突触可塑性（LTP），并改善神经元树突发育。而在正常小鼠，PGRN可促进突触发育，增强突触传递，此作用与其结合肿瘤坏死因子2（TNFα）受体（TNFR2）介导。利用慢性束缚应急模型，我们发现在杏仁核和前额叶（mPFC）脑区PGRN水平升高，考虑可能是由于代偿反应性增加，同时检测发现小鼠焦虑样行为增加。为验证这一假说，我们采用正常小鼠，检测小鼠天然焦虑样行为，即抑制PGRN是否增加焦虑样行为。利用shRNA阻断技术，在杏仁核侧核脑定位注射，7天后检测发现可诱导显著的焦虑样行为；同时发现PGRN-shRNA杏仁核侧核脑定位后，制备300微米厚脑薄片，采用MED64点阵记录采集突触可塑性（LTP），发现皮层-杏仁核侧核通路LTP诱导显著降低。进一步采用PGRN过表达病毒升高mPFC和杏仁核侧核脑区PGRN水平，可显著缓解小鼠焦虑样行为。利用免疫组织化学技术发现，杏仁核PGRN-shRNA注射后小鼠星型胶质细胞激活程度。本研究，首次发现PGRN通过作用于TNFR2受体，可显著阻断炎症因子TNFα介导的中枢炎症反应，恢复突触可塑性，发挥抗焦虑作用。目前，我们利用计算机药物辅助设计手段，以TNFR2受体为靶点，筛选得到具有TNFR2选择性激动作用的候选化合物，其选择性高于TNFR1激动作用60-100倍，为进一步优化TNFR2选择性激动剂奠定了基础。