Sigma 1 Receptor: a novel therapeutic target for retinal degeneration

项目摘要 致盲性视网膜病变构成了一种迫切的未满足的医疗保健需求，需要新的治疗策略。 视网膜退行性疾病是世界范围内无法治愈的失明的主要原因， 感光细胞（PRC）死亡。最近，我们报道了在哺乳动物中，锥体PRC功能的戏剧性拯救。 Pde 6 brd 10/J（rd 10）小鼠视网膜病模型，当用（+）-喷他佐辛（（+）- PTZ），其是σ 1受体（σ 1 R）的高亲和力配体。σ 1 R是一种跨膜蛋白， 参与内质网应激反应的分子伴侣。研究支持σ 1 R调节ER的观点 视网膜中应激和Ca ~（2+）信号。然而，新的数据表明，σ 1 R也定位于核膜 几种视网膜细胞类型（RGC，PRC和Müller细胞）的变化表明ER应激的减弱可能不 完全解释了σ 1 R介导视网膜神经保护的机制。我们假设一部小说 由σ 1 R激活提供的强大的视网膜神经保护的潜在机制是调节Nrf 2- Keap 1途径。nrf 2可以说是细胞中最重要的抗氧化分子，因为它调节 超过500个抗氧化/细胞保护基因的转录。在没有压力的情况下，Nrf 2被保留在 细胞质中的Keap 1和过量的Nrf 2被蛋白酶体降解。然而，在细胞压力下， Keap 1释放Nrf 2，然后转移到细胞核激活“抗氧化反应元件”（ARE） 编码大量细胞防御蛋白和酶的基因。氧化应激是一种主要的致病因素 PRC退化的潜在因素。它显著增加（+）-PTZ与σ 1 R的结合。我们知道（+）-PTZ rd 10小鼠的治疗调节视网膜Nrf 2水平、抗氧化剂基因表达和蛋白质/脂质 然而，我们不知道Nrf 2是否是这些视网膜神经保护作用的核心。要求1 将通过评估Nrf 2在多大程度上对rd 10 PRC救援是足够和必要的来测试这一点。我们知道 在原代Müller细胞中，σ 1 R与Nrf 2-Keap 1在功能上相互作用;然而，我们不知道其程度， 从而改变途径中蛋白质的功能或与其相互作用。目标2将通过评估 （+）-PTZ处理改变Nrf 2-Keap 1基因/蛋白表达、Nrf 2核转位、Nrf 2 ARE WT和σ 1 R-/- Müller细胞中的活化、Keap 1抑制和Nrf 2蛋白酶体降解，并将评价 σ 1 R与Nrf 2-Keap 1通路成员的相互作用。最后，我们知道（+）-PTZ可以拯救圆锥PRC 出生后第42天（P）的rd 10小鼠中;然而，我们不知道（+）-PTZ治疗可以在多大程度上 延迟超过该年龄的rd 10小鼠的视锥细胞死亡，也不确定该效应是否可推广到其他σ 1 R配体。 目的3将通过评价（+）-PTZ治疗的rd 10小鼠在延长的时间内的视网膜功能/结构来测试这一点 并将评估其他两种σ 1 R配体的疗效。总之，有希望的数据构成了我们的基础。 该提案探索了σ 1 R激活介导PRC拯救的新机制，并研究了 这种现象在何种程度上是可持续的，并可推广到其他σ 1 R配体。

PROJECT SUMMARY Blinding retinopathies constitute an urgent unmet healthcare need demanding novel therapeutic strategies. Retinal degenerative diseases are the major cause of untreatable blindness worldwide and frequently involve photoreceptor cell (PRC) death. Recently, we reported dramatic rescue of cone PRC function in the Pde6brd10/J (rd10) mouse model of retinopathy when mice were treated systemically with (+)-pentazocine ((+)- PTZ), a high affinity ligand for sigma 1 receptor (σ1R). σ1R is a transmembrane protein considered a molecular chaperone involved in the ER stress response. Studies support the notion that σ1R modulates ER stress and Ca2+ signaling in retina. However, new data show that σ1R localizes also to the nuclear membrane of several retinal cell types (RGC, PRC, and Müller cells) suggesting that attenuation of ER stress may not explain entirely the mechanism(s) by which σ1R mediates retinal neuroprotection. We postulate that a novel mechanism underlying the robust retinal neuroprotection afforded by σ1R activation is modulation of the Nrf2- Keap1 pathway. Nrf2 is arguably the most important antioxidant molecule in cells because it regulates transcription of more than 500 antioxidant/cytoprotective genes. In the absence of stress, Nrf2 is retained in the cytoplasm by Keap1 and excess Nrf2 is degraded by the proteasome. However, under cellular stress, Keap1 releases Nrf2, which then translocates to the nucleus to activate ‘antioxidant response elements’ (ARE) of genes that encode numerous cell defense proteins and enzymes. Oxidative stress is a major pathogenic factor underlying PRC degeneration. It significantly increases binding of (+)-PTZ to σ1R. We know that (+)-PTZ treatment of rd10 mice modulates retinal Nrf2 levels, antioxidant gene expression and protein/lipid peroxidation, however we do not know whether Nrf2 is central to these retinal neuroprotective effects. Aim 1 will test this by evaluating the extent to which Nrf2 is sufficient and essential to rd10 PRC rescue. We know that σ1R interacts functionally with Nrf2-Keap1 in primary Müller cells; however we do not know the extent to which it alters function of or interacts with proteins in the pathway. Aim 2 will test this by evaluating the extent to which (+)-PTZ treatment alters Nrf2-Keap1 gene/protein expression, Nrf2 nuclear translocation, Nrf2 ARE activation, Keap1 inhibition, and Nrf2 proteasomal degradation in WT and σ1R-/- Müller cells and will evaluate σ1R interaction with members of the Nrf2-Keap1 pathway. Finally, we know that (+)-PTZ can rescue cone PRC in rd10 mice through post-natal day (P)42; however we do not know the extent to which (+)-PTZ treatment can delay cone death in rd10 mice beyond this age nor whether the effects are generalizable to other σ1R ligands. Aim 3 will test this by evaluating retinal function/structure in (+)-PTZ-treated rd10 mice over an extended time course and will assess efficacy of two other σ1R ligands. In summary, promising data form the basis of our proposal, which explores a novel mechanism by which σ1R activation mediates PRC rescue and investigates the extent to which the phenomenon is sustainable and generalizable to other σ1R ligands.