Sigma-1 Chaperone-Mediated in vivo Neuroprotection in the Retina

描述(申请人提供)：神经退行性眼病，如青光眼，在美国导致大量人口失明，但缺乏有效的治疗方法。该项目的最终目标是通过开发一种独特的内源性神经保护剂σ1R伴侣来开发拯救视网膜神经退行性变的新方法，其抗凋亡特性正在被发现。我们在体内的研究表明，σ1R是ROS抑制因子，在神经节细胞和光感受器中对视网膜退行性变具有保护作用。本项目是通过σ1R基因治疗、通过在干细胞来源的光感受器中过表达σ1R以提高其移植后存活以及通过将σ-1受体敲打模型(σ1KO)与慢性σ/2J青光眼模型或RD10模型相结合来测试σ1R介导的视网膜神经保护的有效性的第一项研究。在特定目的1中，我们将分别在DBA/2J青光眼模型和RD10 RP模型中检测σ1R缺失对在体神经节细胞和光感受器活性的影响。由于我们已经发现σ1KO小鼠的神经节细胞在急性模型中比野生型更容易发生视网膜变性，我们将使用与σ1KO和DBA/2J慢性青光眼模型杂交的新模型进一步检验我们的发现。我们还将继续研究σ1R在视网膜光感受器中的神经保护作用，这一作用尚未定义，在我们通过杂交σ1K0和RD10产生的新模型中。在这些模型中，视网膜ROS水平将在存在和不存在σ1R的情况下进行比较。在特定的目标2中，我们将测试σ1R基因疗法，以挽救使用慢病毒的视网膜变性。为了增加σ1R在视网膜中的丰度，我们将利用视网膜下注射技术将携带σ1R基因和Opsin启动子的慢病毒转导RD10小鼠光感受器。过度表达σ1R的转基因眼睛的视网膜功能将与注射σ1R阴性模拟病毒的眼睛进行比较。在具体目标3中，我们将探讨σ1R对干细胞来源的光感受器移植后存活的神经保护作用。在干细胞治疗中，由于视杆细胞的死亡而导致的氧化应激增加，视杆细胞是视网膜中活跃的氧气消耗者，对移植的光感受器的生存构成了有害的威胁。我们将在人胚胎干细胞来源的光感受器中过表达σ1R，以提高其移植到RD10视网膜后的存活率。σ1R对移植的光感受器的促存活作用将与转导σ1R阴性模拟病毒的对照移植细胞进行比较。最终，我们的发现将导致新的σ1R靶向治疗策略，以拯救毁灭性的视网膜神经退行性疾病。

DESCRIPTION (provided by applicant): Neurodegenerative eye diseases such as glaucoma cause blindness in a large population in the USA, yet effective treatments are lacking. The ultimate goal of this project is to develop new methods for rescue of retinal neurodegeneration by exploiting a unique endogenous neuroprotective agent, the σ1R chaperone, whose anti-apoptotic properties are being uncovered. Our in vivo studies suggest that the σ1R is ROS (reactive oxygen species)-suppressing and is protective against retinal degeneration in ganglion cells as well as in photoreceptors. This project represents the first study to test the efficacy of the σ1R-mediated retinal neuroprotection by σ1R gene therapy, by overexpressing the σ1R in stem cell-derived photoreceptors to enhance their post-transplantation survival, and by using the sigma-1 receptor knock model (σ1ko) combined with the chronic DBA/2J glaucoma model or rd10 model. In Specific Aim 1, we will examine the impact of the absence of the σ1R for the viability of ganglion cells and photoreceptors in vivo in the DBA/2J model of glaucoma and the rd10 model of RP, respectively. Since we have discovered that ganglion cells of the σ1ko mice are more susceptible to retinal degeneration than the wild type in an acute model, we will further test our finding using a new model crossed with the σ1ko and the chronic glaucoma model of DBA/2J. We will also extend our investigation into a neuroprotective role of the σ1R in retinal photoreceptors, which has yet to be defined, in a new model that we have generated by crossing σ1ko and rd10. Retinal ROS levels will be compared in the presence and absence of the σ1R in these models. In Specific Aim 2, we will test σ1R gene therapy for rescue of retinal degeneration using lentiviruses. To increase the σ1R abundance in the retina, we will transduce the rd10 mouse photoreceptors with lentiviruses harboring the σ1R gene and the Opsin promoter using a technique of subretinal injection. Retinal functions of the transgenic eye overexpressing the σ1R will be compared to that injected with the σ1R-negative mock virus. In Specific Aim 3, we will explore the neuroprotective effect of the σ1R on the post-transplantation survival of stem cell-derived photoreceptors. In stem cell therapy, elevated oxidative stress due to the death of rods, which are active major consumers of oxygen in the retina, imposes detrimental threat to the survival of transplanted photoreceptors. We will overexpress the σ1R in human embryonic stem cell-derived photoreceptors to enhance their survival after transplantation into the rd10 retina. The pro-survival effect of the σ1R on transplanted photoreceptors will be assessed in comparison to the control transplant cells that are transduced with the σ1R-negative mock virus. Ultimately, our finding will lead to new σ1R-targeted therapeutic strategies for rescue of the devastating retinal neurodegenerative diseases.