Axonal cytoskeletal changes in experimental glaucoma

项目概要/摘要： 该项目提出，在青光眼中视网膜神经节细胞（RGC）死亡之前，以及在永久性丧失视网膜神经节细胞之前， 在视觉中，存在以轴突微管（MT）降解为特征的RGC功能障碍的阶段。 新出现的证据表明，MT降解最初可能发生在轴突未发生实质性变化的情况下。 口径的因此，我们认为，早期RGC功能障碍，涉及MT降解，应 优选地，通过视网膜神经纤维层（RNFL）的扫描激光偏振测定法（SLP）可检测， RNFL厚度的变化。这是因为SLP的基本光学原理是基于检测 偏振光的相位延迟，这是由于在RNFL中产生的光学性质双折射， 细长的圆柱形MT。初步研究表明，RNFL延迟下降之前，和 在几种不同的RGC损伤实验模型中， 青光眼（EG）。在没有RNFL厚度变化的情况下，通过SLP对轴突MT破坏的临床检测， 可能代表青光眼损伤的早期且可能可逆的阶段，并提供临床治疗 用于治疗调整的可检测标记物。因此，这一提议的核心假设是， 视乳头周围RNFL轴突内的MT是神经胶质瘤损害的早期指标， 轴突直径的变化和轴突的物理损失。从这一假设得出的预测是 使用EG的非人灵长类动物（NHP）模型在三个特定目的中进行测试。具体目标1：测试 预测视乳头周围RNFL延迟将在RNFL厚度变化之前下降， 谱域光学相干断层扫描（sd-OCT）和视神经乳头（ONH）表面变化之前 通过共焦扫描激光断层扫描（CSLT）测量NHP眼EG;具体目标2：测试 预测视乳头周围RNFL MT破坏的组织学证据将比 组织学定义的RNFL厚度变化和球后视神经轴突缺失;具体目标3：测试 RGC功能异常与RGC中期相关的预测 以异常轴突MT为特征的变性。为了实现这些目标，将通过激光诱导EG 小梁网光凝术导致24名NHP中度、单侧慢性IOP升高。 将每周测量视乳头周围RNFL延迟、RNFL厚度和ONH表面形貌， 在为期4周的激光前治疗期间，分别使用SLP、sd-OCT和CSLT在每只NHP的双眼中进行 基线期和EG发作后长达8个月（目标1）。对于每个参数，具有统计学显著性 变化被定义为超过每只眼睛的基线会话间变异性的任何变化，两次 确认一旦每只动物进展到其终点，将其处死以收集组织学数据， 分析（目标2）。在针对Aim 1的每周体内结构测试期间，还将评估RGC功能， 使用三种经证实的视网膜电描记法（Aim 3）对双眼进行检查。

Project Summary/Abstract: This project proposes that prior to retinal ganglion cell (RGC) death in glaucoma, and before permanent loss of vision, there exists a stage of RGC dysfunction characterized by degradation of axonal microtubules (MTs). Emerging evidence suggests that MT degradation can occur initially without substantial changes in axonal caliber. Therefore, it is proposed that early stage RGC dysfunction involving MT degradation should be preferentially detectable by scanning laser polarimetry (SLP) of the retinal nerve fiber layer (RNFL) prior to changes in RNFL thickness. This is because the fundamental optical principle of SLP is based on detecting phase retardance of polarized light, which is due to the optical property birefringence produced in the RNFL by the long, thin cylindrical MTs. Preliminary studies demonstrate that RNFL retardance declines prior to, and faster than RNFL thickness in several different experimental models of RGC injury, including experimental glaucoma (EG). Clinical detection of axonal MT disruption by SLP, in the absence of RNFL thickness changes, might represent an early and potentially reversible phase of glaucomatous damage and provide a clinically detectable marker for therapeutic adjustment. Thus the central hypothesis of this proposal is that disruption of MTs within the axons of the peripapillary RNFL is an early indicator of glaucomatous damage, preceding both changes in axonal caliber and physical loss of those axons. Predictions arising from this hypothesis are tested in three Specific Aims using a non-human primate (NHP) model of EG. Specific Aim 1: To test the prediction that peripapillary RNFL retardance will decline prior to RNFL thickness changes measured by spectral domain optical coherence tomography (sd-OCT) and prior to optic nerve head (ONH) surface changes measured by confocal scanning laser tomography (CSLT) in NHP eyes with EG; Specific Aim 2: To test the predictions that histological evidence of peripapillary RNFL MT disruption will be more pronounced than histologically-defined RNFL thickness changes and retrobulbar optic nerve axon loss; Specific Aim 3: To test the prediction that RGC functional abnormalities are associated with the intermediate stage of RGC degeneration characterized by abnormal axonal MTs. To achieve these Aims, EG will be induced via laser photocoagulation of the trabecular meshwork to cause moderate, unilateral chronic IOP elevation in 24 NHPs. Weekly measurements of peripapillary RNFL retardance, RNFL thickness and ONH surface topography will be made in both eyes of each NHP using SLP, sd-OCT and CSLT, respectively, during a 4-week pre-laser baseline period and for up to 8 months after onset of EG (Aim 1). For each parameter, statistically significant change is defined as any change exceeding the baseline intersession variability for each individual eye, twice confirmed. Once each animal progresses to its endpoint, it is sacrificed for histological data collection and analysis (Aim 2). During each week of in vivo structural testing for Aim 1, RGC function will also be assessed in both eyes using three proven forms of electroretinography (Aim 3).