Axonal cytoskeletal changes in experimental glaucoma

描述(申请人提供)：本项目提出，在青光眼视网膜神经节细胞(RGC)死亡之前，在永久性失明之前，存在一个以轴突微管(MTS)降解为特征的RGC功能障碍阶段。新出现的证据表明，MT的降解最初可以发生在轴突口径没有实质性变化的情况下。因此，建议在视网膜神经纤维层(RNFL)厚度改变之前，优先通过扫描激光偏振仪(SLP)检测涉及MT降解的早期RGC功能障碍。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，这是由于长而细的柱面MTS在RNFL中产生的光学性质的双折射。初步研究表明，在包括实验性青光眼(EG)在内的几种不同的RGC损伤实验模型中，RNFL的延迟性在RNFL厚度之前下降，并且比RNFL厚度下降得更快。在无RNFL厚度变化的情况下，SLP临床检测到轴突MT的破坏，可能代表了青光眼损害的早期和潜在的可逆性阶段，并为治疗调整提供了临床可检测的标志物。因此，这一建议的中心假设是，乳头周围RNFL轴突内MTS的破坏是青光眼损害的早期指标，先于轴突口径的变化和这些轴突的物理损失。使用EG的非人灵长类动物(NHP)模型，在三个特定的目标上检验了这一假说产生的预测。具体目的1：验证在伴有EG的NHP眼中，在光谱域光学相干断层扫描(SD-OCT)测量到的RNFL厚度改变之前，以及在用共聚焦扫描激光断层扫描(CSLT)测量到视神经头(ONH)表面改变之前，RNFL延迟性将下降的预测；具体目的2：测试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丢失更明显的预测RNFL周围MT破坏的预测；具体目标3：测试RGC功能异常与以轴突异常为特征的RGC变性中期相关的预测。为了达到这些目的，将通过激光光凝小梁网诱导EG，以引起24例NHP的中度、单侧慢性眼压升高。每只NHP的双眼将分别使用SLP、SD-OCT和CSLT在4周的激光前基线期间和EG开始后的8个月内每周测量RNFL的乳头周围RNFL延迟、RNFL厚度和ONH表面形貌。对于每个参数，统计学上显著的变化被定义为超过每只眼的基线间隔变异性的任何变化，两次确认。一旦每只动物发展到其终点，它就被牺牲用于组织数据收集和分析(目标2)。在每周AIM 1的活体结构测试期间，还将使用三种已证实的视网膜电描记术(AIM 3)来评估双眼的RGC功能。与公共健康相关：青光眼是美国和世界各地最常见的致盲原因之一。这是一种慢性疾病，目前尚无治愈方法，但前瞻性纵向试验发现，降低眼压的治疗可以降低进展性视力丧失的比率。因此，早期诊断可以及时进行治疗干预，减少青光眼的总体影响。然而，要获得及时的诊断，需要临床检测青光眼对视神经头(ONH)和视网膜神经纤维层(RNFL)的损害的发生和发展，这仍然是每个青光眼患者临床护理的中心挑战。该项目评估并改进了检测青光眼早期损害和进展的临床工具。

DESCRIPTION (provided by applicant): 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). PUBLIC HEALTH RELEVANCE: Glaucoma is one of the most common causes of blindness in the United States and around the world. It is a chronic disease with no known cure, but prospective longitudinal trials have found that treatment to lower intraocular pressure decreases the rate of progressive vision loss. Thus, early diagnosis enables timely therapeutic intervention and reduces the overall impact of glaucoma. However, achieving a timely diagnosis requires clinical detection of the onset and progression of glaucomatous damage to the optic nerve head (ONH) and retinal nerve fiber layer (RNFL), which remain a central challenge in the clinical care of every glaucoma patient. This project evaluates and advances clinical tools for detecting early damage and progression of glaucoma.