The role of endozepine diazepam binding inhibitor on the structural plasticity of dendrite spines in neurodegenerative diseases

在中枢神经系统(CNS)中，树突棘的结构可塑性是高度动态的，反映突触的建立或破坏。新出现的证据表明，突触连接的这种结构变化可能不仅是对生理刺激的认知过程的基础，也是许多神经退行性疾病(如果不是全部的话)的基础，如阿尔茨海默病(AD)和突触核病。到目前为止，虽然已经确定了一些树枝晶刺结构可塑性的内在和外在调节因素，但还在不断地发现更多的调控因素。最近，我们观察到在WT小鼠中服用苯二氮卓类药物后TSPO升高，或者在AD小鼠模型中观察到斑块相关小胶质细胞中TSPO的升高，分别导致树突棘可塑性的整体或局部下降。这些证据激发了我们对内源性TSPO配体的兴趣，内源性TSPO配体被称为内源性内切西平，它可能调节健康状况和神经退行性疾病中树突棘的动力学。最著名的内毒素之一是安定结合抑制物(DBI)，它是一种9 kDa的多肽，主要由中枢神经系统中的星形胶质细胞产生和释放，并与TSPO具有高亲和力。在我们的初步工作中，我们观察到在DBI敲除WT小鼠后，使用由U6(细胞型非特异性)启动子驱动的shRNA的病毒载体，WT小鼠的树突棘密度增加，这暗示了DBI在调节树突棘中的作用。最重要的是，在AD小鼠模型中，我们观察到斑块相关星形胶质细胞的DBI显著增加。因此，基于这些数据，很容易假设星形胶质细胞和小胶质细胞通过DBI-TSPO信号通路在神经退行性变的大脑中交织在一起，这可能协同影响树突棘。在这个项目中，我们计划(WP1)表征星形细胞DBI在调节树突棘结构可塑性以及对小胶质细胞形态和功能的作用。之后，我们将探索(WP2)树突棘和小胶质细胞是否以及如何在星形细胞DBI耗竭引起的神经退行性疾病中受到影响。为了以翻译的方式扩展我们的研究，我们将验证(WP3)我们在死后人类组织中的发现，调查神经退行性疾病患者与健康对照组相比星形细胞DBI信号和随后的小胶质突触吞噬的变化。综上所述，我们预计这些实验将破译星形细胞DBI作为树突棘的生理调节器的作用，并可能成为恢复各种神经退行性疾病突触病理的常见治疗靶点。

In the central nervous system (CNS), the structural plasticity of dendritic spines, reflecting the establishment or destruction of synapses, is highly dynamic. Emerging evidence suggests that such structural alterations in synaptic connectivity, are likely the underpinning of cognitive processes in response to not only physiological stimuli but also many, if not all, neurodegenerative diseases, such as Alzheimer's disease (AD) and synucleinopathies. So far, though a few intrinsic and extrinsic modulators of the structural plasticity of dendrite spines have been identified, more are kept being discovered. Recently, we observed elevation of TSPO either upon benzodiazepine administration in WT mice or in plaque-associated microglia in the AD mouse model, leading to a global or regional decrease of dendritic spine plasticity, respectively. Such evidence prompted our interest in endogenous TSPO ligands designated by the generic term ‘endozepines’ that may regulate the dendritic spine dynamics across healthy condition and neurodegenerative diseases. One of the best-known endozepines is the diazepam binding inhibitor (DBI), a 9 kDa peptide primarily produced and released by astrocytes in the CNS, and prominently bind to TSPO with high affinity. In our preliminary work, we have observed increased dendritic spine density in WT mice upon DBI knockdown using viral vectors with shRNA driven by a U6 (cell-type unspecific) promoter, hinting DBI's role in regulating dendritic spines. On top of that, we observed a significant increase of DBI from plaque-associated astrocytes in an AD mouse model. Hence, based on these data, it is tempting to hypothesize that astrocytes and microglia intertwine in the neurodegenerative brain via the DBI-TSPO signaling pathway, which might collaboratively affect dendritic spines. In this project, we plan to (WP1) characterise the role of astrocytic DBI in regulating the structural plasticity of dendritic spines, as well as on microglial morphology and functions. After that, we will explore (WP2) whether and how the dendritic spines and microglia are to be affected in the neurodegenerative diseases upon astrocytic DBI depletion. To extend our study in a translational manner, we will then validate (WP3) our findings in post-mortem human tissues, investigating alterations of astrocytic DBI signalling and subsequent microglial synaptic engulfment in patients with neurodegenerative diseases in comparison to healthy controls. In summary, we expect these experiments will decipher the role of astrocytic DBI as a physiological modulator of dendritic spines, and may as a common therapeutic target for restoring synaptic pathology in a variety of neurodegenerative diseases.