Inflammatory demyelination induces axonal injury and retinal ganglion cell apoptosis in experimental optic neuritis

Optic neuritis is an inflammatory disease of the optic nerve that often occurs in patients with multiple sclerosis and leads to permanent visual loss mediated by retinal ganglion cell (RGC) damage. Optic neuritis occurs with high frequency in relapsing-remitting experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, with significant loss of RGCs. In the current study, mechanisms of RGC loss in this model were examined to determine whether inflammation-induced axonal injury mediates apoptotic death of RGCs. RGCs were retrogradely labeled by injection of fluorogold into superior colliculi of 6-7 week old female SJL/J mice. EAE was induced one week later by immunization with proteolipid protein peptide. Optic neuritis was detected by inflammatory cell infiltration on histological examination as early as 9 days after immunization, with peak incidence by day 12. Demyelination occurred 1-2 days after inflammation began. Loss of RGC axons was detected following demyelination, with significant axonal loss occurring by day 13 post-immunization. Axonal loss occurred prior to loss of RGC bodies at day 14. Apoptotic cells were also observed at day 14 in the ganglion cell layer of eyes with optic neuritis, but not in control eyes. Together these results suggest that inflammatory cell infiltration mediates demyelination and leads to direct axonal injury in this model of experimental optic neuritis. RGCs die by an apoptotic mechanism triggered by axonal injury. Potential neuroprotective therapies to prevent permanent RGC loss from optic neuritis will likely need to be initiated prior to axonal injury to preserve neuronal function.     

Long-term suppression of neurodegeneration in chronic experimental optic neuritis: antioxidant gene therapy

PURPOSE: To test in mice with experimental autoimmune encephalomyelitis (EAE) a strategy designed to treat patients at risk for axonal degeneration and persistent visual loss from optic neuritis and multiple sclerosis. METHODS: The authors cloned the human extracellular superoxide dismutase (ECSOD) or catalase (CAT) gene into recombinant adenoassociated virus (AAV). Transgene expression was evaluated by immunochemistry of infected RGC-5 cells and after intravitreal injection of AAV-ECSOD or AAV-CAT, or both, into the right eyes of DBA/1J mice. Control cells and left eyes were inoculated with AAV-GFP. Animals were sensitized for EAE, followed by serial contrast-enhanced MRI for 6 months, and then were euthanatized. The effects of ECSOD and CAT modulation on the EAE optic nerve were gauged by computerized analysis of optic nerve volume, myelin fiber area, axonal cell loss, and retinal ganglion cell (RGC) loss. RESULTS: Western blot analysis of infected RGC-5 cells revealed that expression of ECSOD increased 15-fold and that of CAT increased 3.5-fold. One month after intraocular injections, transgene expression increased 4-fold for AAV-ECSOD and 3.3-fold for AAV-CAT. Six months after intraocular injections and EAE sensitization, combination therapy with ECSOD and catalase decreased RGC loss by 29%, optic nerve demyelination by 36%, axonal loss by 44%, and cellular infiltration by 34% compared with the contralateral control eyes inoculated with AAV-GFP. Compared with the normal optic nerve, it limited RGC loss to 9%. CONCLUSIONS: Viral-mediated delivery of antioxidant genes provides long-lasting suppression against neuronal and axonal loss associated with permanent visual disability in patients with optic neuritis and multiple sclerosis.     

Evolving management of optic neuritis and multiple sclerosis

PURPOSE: To review the relation of optic neuritis to multiple sclerosis (MS) and the indications, modalities, and results of therapy for optic neuritis, for both visual and general neurologic function. DESIGN: Literature review and author's experience. METHODS: Analysis of both laboratory and clinical evidence supporting the use of corticosteroids, immunomodulation agents, and other modalities in the treatment of optic neuritis and MS. RESULTS: Although treatment of optic neuritis with corticosteroids may hasten visual recovery to a minor degree, it has no long-term beneficial effect on visual outcome. Optic neuritis is frequently the initial manifestation of multiple sclerosis. The risk of later development of clinically definite MS (CDMS) correlates with white matter demyelinative lesions on magnetic resonance imaging (MRI). The role of corticosteroid therapy alone in reducing the risk of subsequent MS is unclear, but recent studies suggest that the combination of immunomodulation agents (IMAs) and corticosteroids significantly reduces the later development of MS. Current research indicates that, contrary to previous doctrine, axonal damage is an early finding in MS. CONCLUSIONS: The risk of MS after optic neuritis may be predicted. The use of corticosteroids and IMAs, particularly in those at substantial risk, reduces the frequency and severity of developing CDMS. Earlier, more aggressive therapy in optic neuritis may be proven to reduce permanent axonal injury and progressive disability in MS.     

Suppression of mitochondrial oxidative stress provides long-term neuroprotection in experimental optic neuritis

PURPOSE: Axonal loss is thought to contribute to the persistence of visual loss in optic neuritis and multiple sclerosis (MS). The mechanisms of injury are poorly understood. The authors investigated the contribution of mitochondrial oxidative stress and the effects of modulating mitochondrial antioxidant gene expression in the optic nerves of mice induced with experimental allergic encephalomyelitis (EAE), with a focus on long-term neuroprotection. METHODS: Optic nerves from mice with EAE were probed for reactive oxygen species (ROS) with the use of dichlorofluorescein diacetate (DCFDA), dihydroethidium, and cerium chloride. To modulate mitochondrial oxidative stress, recombinant AAV containing the human SOD2 gene or a ribozyme targeting murine SOD2 was injected into the vitreous. Control eyes received the recombinant virus without a therapeutic gene. Mice were sensitized for EAE and were monitored by serial contrast-enhanced MRI. The effects of SOD2 modulation on the EAE optic nerve were gauged by computerized analysis of optic nerve volume, myelin fiber area, and retinal ganglion cell loss at 1, 3, and 12 months after sensitization for EAE. RESULTS: ROS were detected in the EAE optic nerve as early as 3 days after antigenic sensitization. Colocalization suggested mitochondria as the source of ROS activity in the absence of inflammation. The ribozyme suppressing SOD2 gene expression increased myelin fiber injury by 27%. Increasing SOD2 levels twofold in the optic nerve by virally mediated gene transfer ameliorated myelin fiber injury by 51% and RGC loss fourfold, limiting it to 7% in EAE at 1 year. CONCLUSIONS: Amelioration of mitochondrial oxidative stress by SOD2 gene delivery may be a therapeutic strategy for suppressing neurodegeneration in optic neuritis.     

Management of optic neuritis and multiple sclerosis.

Optic neuritis and multiple sclerosis (MS) are common causes of visual and neurologic dysfunction in young adults. Advances in magnetic resonance imaging, molecular genetics, and neuroimmunology have increased our understanding of the pathophysiology underlying both disorders. Corticosteroids remain the mainstay of treatment of optic neuritis, but alternate dosages and routes of administration are undergoing investigation. The available therapies for MS have expanded, and there is evidence that early intervention is beneficial. Treatments for MS show sustained efficacy, but are not curative, and adjunctive treatments may prove valuable in patients with progressive visual and neurologic disability.     

Acute optic neuritis associated with immunization with the CNS myelin proteolipid protein.

Optic nerve tissue for SJL/J mice immunized with the central nervous system (CNS) myelin-specific proteolipid protein (PLP) was examined for histopathologic evidence of optic neuritis. Optic nerves isolated 17 d after immunization with PLP revealed an interstitial and submeningeal inflammatory infiltrate consisting of neutrophils and monocytes. In all cases, histologic evidence of optic nerve involvement correlated serologically with the presence of circulating anti-PLP antibodies. Control animals had no histopathologic evidence of disease or anti-PLP antibody. In many respects, the observed histopathologic profile of PLP-induced optic neuritis is similar to that associated with human inflammatory demyelinating diseases such as multiple sclerosis (MS). Because optic neuritis frequently is associated with some of the earliest clinical symptoms of MS, the acute nature of optic nerve involvement in this animal model suggests that immune recognition of the myelin PLP may play a significant role in the pathophysiology of optic nerve damage associated with sensitization to CNS-specific antigens.     

辅助性T细胞在脱髓鞘性视神经炎发病中的作用

背景 视神经炎是神经眼科临床常见疾病之一,易反复发作并可造成视神经轴索损害及不可逆性视力损伤,且无有效的防治方法,其发病机制被认为与机体免疫调节失衡有关. 目的 研究辅助性T细胞（Th）亚群调节性T细胞（Treg）、Th17特异性细胞因子及其转录因子在脱髓鞘性视神经炎早期发病中的表达,以明确实验性自身免疫性脑脊髓炎（EAE）小鼠脱髓鞘性视神经炎的免疫反应启动及免疫炎症维持机制. 方法 采用随机数字表法将6～9周龄清洁级C57BL/6（H-2b）雌性小鼠84只随机分为正常对照组12只和免疫后7、11、14、19、23、28 d组（模型组亚分）各12只.模型组小鼠分别于背部两侧皮下注射相应抗原200 μg,免疫当天记作第0天,于免疫后第0、2天给予模型组小鼠腹腔内注射百日咳疫苗400 ng,对照组注射生理盐水.观察视神经炎小鼠的眼部表现及闪光视觉诱发电位（F-VEP）变化;苏木精-伊红染色观察视神经的组织学改变;免疫组织化学法检测视神经轴索损伤标志物β-淀粉样前体蛋白（β-APP）表达量的变化;逆转录PCR（RT-PCR）法检测视神经组织中转化生长因子-β（TGF-β）、白细胞介素-1β（IL-1β）、IL-17、IL-10及叉头蛋白3（Foxp3）等的动态表达.结果 免疫后11d,模型组小鼠陆续出现神经系统症状,苏木精-伊红染色可见模型组视神经组织内出现炎性因子浸润,免疫组织化学法检测发现模型组轴索损伤标志物β-APP阳性染色增强;F-VEP检测结果显示,免疫后7、11、14、19、23、28 d组小鼠P1波潜时值较正常对照组小鼠延长,差异均有统计学意义（t=4.487、15.203、16.364、11.540、11.959、16.163,P＜0.05）;免疫后7d,正常对照组与模型组小鼠N1-P1振幅的差异无统计学意义（t=-0.992,P=0.378）;而免疫后11、14、19、23、28 d组小鼠N1-P1振幅明显低于正常对照组,差异均有统计学意义（t=-13.161、-31.401、-16.109、-7.025、-7.257,P＜0.05）.小鼠视神经中TGF-β、IL-1β、IL-17、Foxp3、IL-10 mRNA表达各时间点间整体比较差异有统计学意义（F=12.721、15.015、14.343、69.374、68.290,均P=0.000）,与正常对照组比较,免疫后11d、14d组,TGF-β mRNA水平明显升高,免疫后19d、23 d组,IL-1β mRNA水平明显升高,免疫后7d、11d组IL-17 mRNA水平明显升高,免疫后7、11、14、19、23、28 d组Foxp3 mRNA水平均明显降低,免疫后19、23、28 d组IL-10 mRNA水平明显降低,差异均有统计学意义（P＜0.05）.结论 Th17细胞亚群参与了脱髓鞘性视神经炎免疫损伤的启动,而Th1细胞亚群维持了炎症损伤,Treg亚群较Th2亚群更早出现异常.Th17/Treg比例失衡可能参与了脱髓鞘性视神经炎的发生.     

Optic neuritis and demyelination

The Optic Neuritis Treatment Trial continues to generate information and controversy on the visual and neurologic outcome and treatment of optic neuritis. At the same time, other researchers explored cerebrospinal fluid parameters in multiple sclerosis, treatment of experimental optic neuritis, corticosteroid treatment of multiple sclerosis, and variations and mimickers of optic neuritis.     

Management of optic neuritis

Optic neuritis is an inflammatory condition of the optic nerve characterized by a sudden onset of unilateral visual loss, usually affecting young females. Demyelination associated with multiple sclerosis (MS) is the most common cause in regions where MS is prevalent; while in other places, there are a substantial proportion of cases where infective or autoimmune causes are seen. Optic Neuritis Treatment Trial (ONTT) was the first major study that provided information on the natural history, role of steroids in treatment and risk of development of MS. Subsequently, numerous clinical trials have evaluated different modalities of management of optic neuritis and MS. The Controlled High-Risk Subjects Avonex Multiple Sclerosis Prevention Study (CHAMPS); the Prevention of Relapses and Disability by Interferon β-1a Subcutaneously in Multiple Sclerosis (PRISMS) Trial; and, most recently, the Betaferon in Newly Emerging Multiple Sclerosis for Initial Treatment (BENEFIT) Study have provided large amount of information on the natural history of optic neuritis and management options available. However, due to the low prevalence of MS reported in Asian studies, high cost of therapy and indefinite time period of treatment, it may not be cost effective to start interferon therapy in most cases.     

Neuroprotective effect of latanoprost on rat retinal ganglion cells

Background To investigate the neuroprotective effect of intravitreal administration of latanoprost on retinal ganglion cell (RGC) damage induced by N-methyl-D-aspartic acid (NMDA) or optic nerve axotomy.Methods Using Sprague-Dawley rats, retinal ganglion cell damage was induced by either intravitreal administration of NMDA or optic nerve axotomy. Latanoprost at doses of 0.03, 0.3, 3, 30 and 300 pmol was administered intravitreally before NMDA injection or optic nerve axotomy. Retinal damage was evaluated by counting the number of surviving RGCs retrogradely labeled with fluorogold under the microscope.Results Seven days after the NMDA injury, the number of surviving RGCs was significantly increased at doses of more than 30 pmol atanoprost (846±178 cells/mm² P=0.0166) compared with vehicle control (556±122 cells/mm²). Ten days after the optic nerve axotomy, the number of surviving RGC was significantly increased even at a dose of 0.3 pmol (815±239 cells/mm², P=0.0359) compared with control (462±75 cells/mm²).Conclusions Intravitreal administration of latanoprost has a neuroprotective effect on rat RGC damage induced by either NMDA or optic nerve axotomy, while its pharmacological features are different.     

CNTF和Ad-BDNF对视神经夹伤后视网膜神经节细胞存活的影响

目的:观察大鼠视神经夹伤后玻璃体腔内注射睫状神经营养因子(CNTF)和腺病毒介导脑源性神经营养因子(Ad-BDNF)对视神经损伤后视网膜神经节细胞(RGC)存活的影响。方法:制作大鼠视神经定量夹伤模型,玻璃体腔内注射CNTF和Ad-BDNF,经上丘荧光金(FG)逆行标记RGC,计数视网膜铺片上的RGC并行统计学分析。结果:正常SD大鼠视网膜上RGC密度为2155±265个/mm2(n=12),视神经夹伤后RGC在1~2wk内下降速率最快,到3,4wk时RGC细胞数量虽仍有减少但下降速度已经明显减慢。CNTF组在视神经夹伤后1wk时视网膜RGC数显著高于对照组,但2~4wk的结果和对照组比较差异不明显。Ad-BDNF组视神经夹伤后1~4wk视网膜RGC数均显著高于对照组。结论:CNTF治疗组玻璃体腔内一次性注射CNTF可以在损伤早期2wk内为损伤的RGC提供神经营养因子,减少RGC的早期死亡。Ad-BDNF治疗组的这种保护作用可以持续到损伤后4wk,能够为RGC提供长时间地营养支持,但这种作用比较局限,可能与单一营养因子作用有关。     

Optic neuritis in cat scratch disease

A patient with optic neuritis is described whose associated lymphadenopathy, cat exposure, and positive cat scratch antigen skin test suggested the diagnosis of cat scratch disease. Optic neuritis and other neurologic abnormalities, especially encephalopathy, may be associated with, or the presenting manifestation of, cat scratch disease. To our knowledge, this is the fourth case of cat scratch disease-related optic neuritis to be reported.     

Optic neuritis in adults and children

Most ophthalmologists will encounter optic neuritis in the course of their practice. The disease behaves very differently in children versus adults. The ONTT and CHAMPS trials have given us important data regarding prognosis and management in adults, including probability of developing MS. Optic neuritis in children, while less common, must be considered in the differential diagnosis of subacute visual loss, especially in children with bilateral impairment. Secondary causes of optic neuritis are more common in children and an investigation should be performed to assess for associated disease states. In both populations, the ophthalmologist will play a central role in the diagnosis, treatment, and management of these patients, whether it is an isolated case of optic neuritis or the initial presentation of a long-term struggle with MS. With the advent of immunomodulating therapy to minimize both the subsequent incidence and severity of MS, early diagnosis and appropriate management by the ophthalmologist has become even more critical.     

Optic neuritis in adults and children

Most ophthalmologists will encounter optic neuritis in the course of their practice. The disease behaves very differently in children versus adults. The ONTT and CHAMPS trials have given us important data regarding prognosis and management in adults, including probability of developing MS. Optic neuritis in children, while less common, must be considered in the differential diagnosis of subacute visual loss, especially in children with bilateral impairment. Secondary causes of optic neuritis are more common in children and an investigation should be performed to assess for associated disease states. In both populations, the ophthalmologist will play a central role in the diagnosis, treatment, and management of these patients, whether it is an isolated case of optic neuritis or the initial presentation of a long-term struggle with MS. With the advent of immunomodulating therapy to minimize both the subsequent incidence and severity of MS, early diagnosis and appropriate management by the ophthalmologist has become even more critical.     

Current concepts of the pathogenesis of optic neuritis associated with multiple sclerosis

Optic neuritis may occur as an isolated entity or as a manifestation of multiple sclerosis (MS), a widespread central nervous system demyelinating disease. Clinical, electrophysiological, magnetic resonance imaging and neuropathologic data support the hypothesis that "idiopathic" optic neuritis represents a restricted form of MS. The current evidence for viral, cell mediated, and antibody-induced etiologies of MS and optic neuritis are reviewed.     

Myelin/oligodendrocyte glycoprotein-specific T-cells induce severe optic neuritis in the C57BL/6 mouse

PURPOSE: The optic nerve is a common site of tissue damage in multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE). To determine the relationship between optic neuritis (ON) and EAE, we examined the incidence of ON in C57BL/6 (B6) mice immunized with a myelin oligodendrocyte/glycoprotein (MOG)-derived peptide or injected with MOG-specific T cells, which are known to induce EAE. METHODS: Mice were immunized with MOG35-55 or MOG40-54 peptides emulsified in complete Freund's adjuvant (CFA). Pertussis toxin (PTX) was injected intraperitoneally 1 day before and after immunization. For disease induction by adoptive transfer of primed cells, donor C57BL/6 mice were received with T-cell blasts (1-6 x 10(6)/mouse). Both EAE and ON were observed by either clinical signs or histology. RESULTS: ON developed in a high proportion of B6 mice treated with either protocol. The most severe inflammation was observed in the adoptively transferred mice. The induced ON was most frequently bilateral. In either actively or adoptively transferred diseases, both association and dissociation of EAE and ON was observed. CONCLUSIONS: Different MOG-specific T-cell subsets might be involved in the pathogenesis of EAE and ON. A better understanding of the pathogenesis of ON after induction by MOG may have important diagnostic and therapeutic implications.     

Optic Neuritis: An Update. II. Optic Neuritis and Multiple Sclerosis

Demyelinating optic neuritis (DON) is an acute inflammatory demyelinating disorder affecting the optic nerve. The risk of having a subsequent clinical episode elsewhere in the central nervous system leading to a diagnosis of clinically definite multiple sclerosis (CDMS) varies in different parts of the world. In countries with a high prevalence of MS, most patients with DON will eventually develop CDMS. The presence of oligoclonal bands in cerebrospinal fluid and asymptomatic lesions on magnetic resonance imaging (MRI) of the brain at presentation are strong predictors of progression to CDMS. The Optic Neuritis Treatment Trial reported that DON patients treated with intravenous methylprednisolone had a delay in progression to CDMS, but only up to two years. Interferon-β and glatiramer actetate have now been shown to reduce the risk of developing CDMS after two to three years when compared with placebo. The BENEFIT study has demonstrated that delaying interferon-β1b therapy in patients presenting with a clinically isolated syndrome (CIS) such as optic neuritis leads to a slightly higher risk for sustained disability progression compared with patients who started interferon-β1b after a CIS. Controversy remains if these small benefits of reducing conversion to CDMS from CIS and slowing disability with interferon-β treatment outweigh the adverse effects of the medication and its costs as the overall disability progression off treatment is mild, at least in the short-term, and particularly in patients in whom optic neuritis is the initial event.     

神经生长因子对脱髓鞘性视神经炎小鼠视网膜神经组织保护作用的实验研究

目的 研究神经生长因子（NGF）对脱髓鞘性视神经炎小鼠视网膜神经组织的保护作用.方法 实验研究.50只C57BL/6小鼠按随机数字表法分为对照组（10只）、BSS组（20只）和NGF组（20只）,取右眼为实验眼.后两组应用MOG35-55多肽加完全弗氏佐剂皮下注射建立实验性视神经炎小鼠模型,每天对各组小鼠进行称重及神经功能评分.NGF组和BSS组分别于免疫后的第4天和第10天,对右眼进行玻璃体腔注射3μg/μl NGF或2 μl BSS.分别于免疫当天、免疫后的第7天和免疫后的第14天,对每组小鼠进行闪光视觉诱发电位（f-VEP）和闪光视网膜电图（f-ERG）检查;采用HE染色、LFB染色、Bielschowsky银染分别评估视神经炎性细胞浸润、髓鞘脱失、轴突病理改变;使用TUNEL法检测视网膜神经节细胞凋亡并计算凋亡指数.对两组实验数据采用t检验进行统计学分析.结果 NGF组与BSS组小鼠发病时间和临床评分比较,差异均无统计学意义（t=-1.844,P=0.079;t=-2.012,P=0.059）.在不同时间点,NGF组和BSS组的f-VEP差异均无统计学意义（P＞0.05）.在免疫后的第14天,NGF组f-ERG b波潜伏期较BSS组缩短,振幅较BSS组增大,两组差异有统计学意义（t=5.909,P=0.000;t=3.602,P=0.043）.LFB染色示,NGF组和BSS组视神经的脱髓鞘面积占横截面比例分别为（31.50±8.72）%、（29.91±10.00）%,差异无统计学意义（t=0.298,P=0.709）.TUNEL检测结果示,NGF组小鼠视网膜神经节细胞凋亡指数[（15.18±3.36）%]低于BSS组[（34.14±3.83）%],差异有统计学意义（t=11.790,P=0.000）.结论 NGF可以促进脱髓鞘性视神经炎小鼠视网膜神经节细胞的存活,对其视网膜神经组织可能具有一定的保护作用.     

Spontaneous ocular and neurologic deficits in transgenic mouse models of multiple sclerosis and noninvasive investigative modalities: a review

Multiple sclerosis (MS) is an autoimmune, inflammatory, neurodegenerative, demyelinating disease of the central nervous system, predominantly involving myelinated neurons of the brain, spinal cord, and optic nerve. Optic neuritis is frequently associated with MS and often precedes other neurologic deficits associated with MS. A large number of patients experience visual defects and have abnormalities concomitant with neurologic abnormalities. Transgenic mice manifesting spontaneous neurologic and ocular disease are unique models that have revolutionized the study of MS. Spontaneous experimental autoimmune encephalomyelitis (sEAE) presents with spontaneous onset of demyelination, without the need of an injectable immunogen. This review highlights the various models of sEAE, their disease characteristics, and applicability for future research. The study of optic neuropathy and neurologic manifestations of demyelination in sEAE will expand our understanding of the pathophysiological mechanisms underlying MS. Early and precise diagnosis of MS with different noninvasive methods has opened new avenues in managing symptoms, reducing morbidity, and limiting disease burden. This review discusses the spectrum of available noninvasive techniques, such as electrophysiological and behavioral assessment, optical coherence tomography, scanning laser polarimetry, confocal scanning laser ophthalmoscopy, pupillometry, magnetic resonance imaging, positron emission tomography, gait, and cardiovascular monitoring, and their clinical relevance.