Chronic experimental glaucoma in primates. II. Effect of extended intraocular pressure elevation on optic nerve head and axonal transport

Intraocular pressure (IOP) elevations lasting from 2 to 42 days were produced in 13 primate eyes by anterior chamber injections of autologous, fixed red blood cells. The retina, optic nerve head, and optic nerves were studied by electron microscopy, and ganglion cell rapid axonal transport was examined after IOP elevations for various durations. Transport of axonal material was blocked at the scleral lamina cribrosa by IOP elevations to 50 mm Hg. With IOP elevation for less than 1 week, return to normal IOP restored normal transport in some axons. However, in other axons IOP elevation for less than 1 week intiated ganglion cell degeneration. The process of cellular death involved a rapid ascending degeneration from nerve head to brain, followed 3 to 4 weeks later by descending degeneration of the ganglion cell body and its attached axon. Axons of the superior and inferior optic nerve head and nerve seem to be damaged more extensively than those in the nasal and temporal optic nerve. Two to four days after IOP elevation, axons of the superficial optic nerve head were swollen with accumulating axonal material, leading to histologic disk edema. In those eyes with IOP elevation longer than 1 week, the loss of anterior disk nerve fibers combined with posterior and lateral movement of the lamina cribrosa lead to an increase in optic disk cupping. Astrocytes and capillaries of the optic nerve head seem to tolerate elevated IOP well and were relatively spared.     

Obstructed axonal transport of BDNF and its receptor TrkB in experimental glaucoma

PURPOSE: In both animal model system and in human glaucoma, retinal ganglion cells (RGCs) die by apoptosis. To understand how RGC apoptosis is initiated in these systems, the authors studied RGC neurotrophin transport in experimental glaucoma using acute intraocular pressure (IOP) elevations in rats and chronic IOP elevation and unilateral optic nerve transections in monkeys. METHODS: Eyes were studied in masked fashion by light and electron microscopy and by immunohistochemistry with antibodies directed against the tyrosine kinase receptors (TrkA, B, and C) and against brain-derived neurotrophic factor (BDNF), as well as by autoradiography to identify retrograde axonal transport of 125I-BDNF injected into the superior colliculus. RESULTS: With acute glaucoma in the rat, RGC axons became abnormally dilated, accumulating vesicles presumed to be moving in axonal transport at the optic nerve head. Label for TrkB, but not TrkA, was relatively increased at and behind the optic nerve head with IOP elevation. Abnormal, focal labeling for TrkB and BDNF was identified in axons of monkey optic nerve heads with chronic glaucoma. With acute IOP elevation in rats, radiolabeled BDNF arrived at cells in the RGC layer at less than half the level of control eyes. CONCLUSIONS: Interruption of BDNF retrograde transport and accumulation of TrkB at the optic nerve head in acute and chronic glaucoma models suggest a role for neurotrophin deprivation in the pathogenesis of RGC death in glaucoma.     

High intraocular pressure-induced ischemia and reperfusion injury in the optic nerve and retina in rats

• Purpose: The purpose of this paper is to describe the damage caused to the retina and the axons of the optic nerve by acute ischemia-reperfusion injury and the extent to which optic nerve damage correlates with the duration if ischemia due to high intraocular pressure (IOP). • Methods: Acute ischemia in the retina and optic disc was induced in albino rats by increasing the IOP to 110 mmHg for a period of 45–120 min. Thereafter, the eyes were reperfused at normal IOP after 7 days. The retina and optic nerve were examined by light and electron microscopy, and morphometrical counts of the optic nerve axons were performed. • Results: After 45 min of ischemia, electron microscopic examination revealed swelling of mitochondria and degeneration of neurotubules on axons in cross sections of the optic nerve. The axonal counts in eyes subjected to 45 min of ischemia were 29% lower than in control eyes. After 60 min of ischemia, there were distinct disruptions of mitochondria and degeneration of the axons. After 90 min of ischemia, numerous axons showed degeneration with disordered myelin sheaths. Neuronal cell death was seen in the retina, mainly in the ganglion cell layer. • Conclusion: Damage to the retinal ganglion cell layer and the optic nerve was evident after only 45 min of ischemia in normal eyes. This experiment suggests that seriously injured eyes must be protected from high IOP; if IOP elevation is required during vitrectomy, it is essential to reduce the duration of interruption of blood flow to a minimum.     

Optic nerve dynein motor protein distribution changes with intraocular pressure elevation in a rat model of glaucoma

Acute intraocular pressure (IOP) elevation causes accumulation of retrogradely-transported brain derived neurotrophic factor and its receptor at the optic nerve head (ONH) in rats and monkeys. Obstruction of axonal transport may therefore be involved in glaucoma pathogenesis, but it is unknown if obstruction is specific to certain transported factors or represents a generalized failure of retrograde axonal transport. The dynein motor complex mediates retrograde axonal transport in retinal ganglion cells (RGC). Our hypothesis was that elevated IOP interferes with dynein-mediated axonal transport. We studied the distribution of dynein subunits in the retina and optic nerve after acute and chronic experimental IOP elevation in the rat. IOP was elevated unilaterally in 54 rats. Dynein subunit distribution was compared in treated and control eyes by immunohistochemistry and Western blotting at 1 day (n=12), 3 days (n=4), 1 week (n=15), 2 weeks (n=12) and 4 weeks (n=11). For immunohistochemistry, sections through the ONH were probed with an anti-dynein heavy chain (HC) antibody and graded semi-quantitatively by masked observers. Other freshly enucleated eyes were microdissected for separate Western blot quantification of dynein intermediate complex (IC) in myelinated and unmyelinated optic nerve, ONH and retina. Immunohistochemistry showed accumulation of dynein HC at the ONH in IOP elevation eyes compared to controls (P<0.001, Wilcoxon paired sign-rank test, n=29). ONH dynein IC was elevated by 46.5% in chronic IOP elevation eyes compared to controls by Western blotting (P<0.001, 95% CI=25.9% to 67.8%, n=17). The maximum increase in ONH dynein IC was 78.7% after 1 week (P<0.05, n=5), but significant increases were also detected after 4 h and 4 weeks of IOP elevation (P<0.05, n=4 rats per group). Total retinal dynein IC was increased by 8.7% in chronic IOP elevation eyes compared to controls (P<0.03, 95% CI 1.4% to 16.1%, n=24). In the retina, IOP elevation particularly affected the 72 kD subunit of dynein IC, which was 100.7% higher in chronic IOP elevation eyes compared to controls (P<0.00001, 95% CI 71.0% to 130.4%, n=21). Dynein IC changes in myelinated and unmyelinated optic nerve were not significant (P>0.05). We conclude that dynein accumulates at the ONH with experimental IOP elevation in the rat, supporting the hypothesis that disrupted axonal transport in RGC may be involved in the pathogenesis of glaucoma. The effect of IOP elevation on other motor proteins deserves further investigation in the future.     

Understanding mechanisms of pressure-induced optic nerve damage

Patients with glaucoma can suffer progressive vision loss, even in the face of what appears to be excellent intraocular pressure (IOP) control. Some of this may be secondary to non-pressure-related (pressure-independent) factors. However, it is likely that chronically elevated IOP produces progressive changes in the optic nerve head, the retina, or both that alter susceptibility of remaining optic nerve fibers to IOP. In order to understand the nature of these progressive changes, relevant, cost-effective animal models are necessary. Several rat models are now used to produce chronic, elevated IOP, and methods exist for measuring the resulting IOP and determining the extent of the damage this causes to the retina and optic nerve. A comparison of damage, pressure and duration shows that these models are not necessarily equivalent. These tools are beginning to uncover clear evidence that elevated IOP produces progressive changes in the optic nerve head and retina. In the optic nerve head, these include axonal and non-axonal effects, the latter pointing to involvement of extracellular matrix and astrocyte responses. In the retina, retinal ganglion cells appear to undergo changes in neurotrophin response as well as morphologic changes prior to actual cell death. These, and other, as yet uncovered, abnormalities in the optic nerve head and retina may influence relative susceptibility to IOP and explain progressive optic nerve damage and visual field loss, in spite of apparent, clinically adequate IOP control. Ultimately, this knowledge may lead to the development of new treatments designed to preserve vision in these difficult patients.     

Retrograde axonal transport of BDNF in retinal ganglion cells is blocked by acute IOP elevation in rats

PURPOSE: To determine whether acute experimental glaucoma in rats obstructs retrograde transport of brain-derived neurotrophic factor (BDNF) to retinal ganglion cells (RGCs). METHODS: Forty rats had unilateral injection of either (125)I-BDNF (20 animals) or a mixture of (125)I-BDNF and 100-fold excess nonradiolabeled BDNF (20 animals). In each group of 20 animals, eyes contralateral to injection had either normal intraocular pressure (IOP; 10 animals) or IOP elevated to 25 mm Hg below the systolic blood pressure of the eye (10 animals). In each group of 20 rats, ipsilateral eyes had IOP set at systolic blood pressure (4 eyes), had optic nerve transection (10 eyes), or had normal IOP (6 eyes). Six hours after injection, animals were killed and tissues were fixed, embedded, and sectioned for autoradiography. Grain counts were performed over retina and optic nerve using automated image analysis. RESULTS: IOP elevation to 25 mm Hg below systolic blood pressure (perfusion pressure [PP] 25) decreased median retinal nerve fiber layer (NFL) grains by 38% compared with controls (P: < 0.001). Competition by cold BDNF reduced NFL grains by 28% (P: = 0.013). Considering only the radioactivity representing specific retrograde transport of BDNF, IOP elevation to PP25 reduced transport by 74%, whereas elevation to PP0 (equaling systolic blood pressure) reduced specific transport by 83%. CONCLUSIONS: BDNF is transported retrogradely from the superior colliculus in adult rats, and this transport is substantially inhibited by acute IOP elevation. Deprivation of BDNF among RGCs may contribute to neuron loss in glaucoma.     

负压吸引对兔眼视网膜和视神经的影响

探讨准分子激光原位角膜磨镶术(1aser in situ keratomileusis,LASIK)中眼压急剧升高对视网膜和视神经的影响。方法将LASIK术中使用的巩膜负压环置于兔眼,瞬时压力达到65mm Hg(1mm Hg=0.133kPa),分别持续30s、1min及3min,在光镜和电镜下观察负压吸引后即刻摘除的(即刻组)和2周后摘除的(修复组)兔眼视神经和视网膜组织,并与正常兔眼(对照组)进行比较。结果负压吸引30s,视神经和视网膜细胞改变轻微;负压吸引1min,部分视神经纤维改变,视网膜视细胞出现异常;负压吸引3min,神经纤维和视网膜视细胞结构明显异常。即刻组和修复组组织改变基本相同。结论LASIK术中眼压急剧升高,可导致视网膜和视神经细胞的亚细胞结构改变,且持续时间越长改变越明显。     

Distribution of axonal transport blockade by acute intraocular pressure elevation in the primate optic nerve head.

We studied the degree of axonal transport blockade in various areas of the optic nerve head with acute intraocular pressure (IOP) elevation in 19 squirrel monkey eyes. When IOP was raised to 20 to 50 mm. Hg for 7 hr., mild axonal transport blockade occurred in each area of the disk, most prominently in nerve fiber bundles of the superior pole. With 7 hr. IOP elevations between 50 and 90 mm. Hg, a somewhat greater degree of transport blockade occurred throughout the nerve head, although again the superior and inferior poles were somewhat more affected. The distribution of short-term transport blockade over the entire nerve head corresponds to the diffuse damage of acute glaucoma, but the pattern hints at the preference for damage near the poles of the disk seen in chronic glaucoma. However, before these results can be fully evaluated, further information is needed on axonal pathways through the optic nerve head and on the relationship between transport obstruction and ganglion cell death.     

Increase in dephosphorylation of the heavy neurofilament subunit in the monkey chronic glaucoma model

PURPOSE: To investigate the phosphorylation of the heavy neurofilament subunit (NF-H), which could be deeply involved in axonal transport of retinal ganglion cells (RGCs), in an experimental glaucoma model of chronic elevation of intraocular pressure (IOP) in monkeys. METHODS: One eye in adult monkeys was randomly selected for laser treatment, and IOP was maintained between 30 and 40 mm Hg throughout the experiment. The eyeballs with the optic nerve and optic chiasm were enucleated as one tissue and were subject to immunocytochemical observation, using two NF-H-specific antibodies, NF-200 and SMI31. NF-200 reacts with both phosphorylated and dephosphorylated NF-H, whereas SMI reacts only with phosphorylated NF-H. Ratios of SMI31-positive to NF-200-positive areas were calculated for quantitative evaluation of phosphorylation status. Specimens from the retina, lamina cribrosa (LC), post-LC, and optic chiasm were evaluated separately. Phosphorylation of NF-H at the retina and optic nerve head was compared between specimens from temporal retina and nasal retina, or between temporal and nasal regions of the optic disc. The status of phosphorylation was confirmed by Western blot analysis. RESULTS: An enlargement of the disc cup was observed on the temporal side, and the superior and inferior poles were preferentially involved in the neuronal damage in laser-treated eyes. Most NF-Hs in the control eyes were phosphorylated in all investigated regions, whereas those in the glaucomatous eyes were significantly dephosphorylated, and NF-Hs in the temporal region were significantly dephosphorylated compared with those in the nasal region. At the optic chiasm, NF-Hs in axons traveling from laser-treated eyes were highly dephosphorylated, and the extent of NF-H dephosphorylation corresponded to the degree of glaucoma-induced axonal damage. Western blot analysis showed the change in the phosphorylation of NF-Hs. CONCLUSIONS: NF-Hs in RGC axons are dephosphorylated by elevated IOP, which may be deeply involved in glaucoma-induced damage to axonal transport.     

Neurotrophin roles in retinal ganglion cell survival: Lessons from rat glaucoma models

The neurotrophin (NT) hypothesis proposes that the obstruction of retrograde transport at the optic nerve head results in the deprivation of neurotrophic support to retinal ganglion cells (RGC) leading to apoptotic cell death in glaucoma. An important corollary to this concept is the implication that appropriate enhancement of neurotrophic support will prolong the survival of injured RGC indefinitely. This hypothesis is, perhaps, the most widely recognized theory to explain RGC loss resulting from exposure of the eye to elevated intraocular pressure (IOP). Recent studies of NT signaling using rat glaucoma models, have examined the endogenous responses of the retina to pressure exposure as well as studies designed to augment NT signaling in order to rescue RGC from apoptosis following pressure-induced injury. The examination of these studies in this review reveals a number of consistent observations and provides direction for further investigations of this hypothesis.     

The mechanism of optic nerve damage in experimental acute intraocular pressure elevation

We produced intraocular pressure (IOP) elevations in 32 primate eyes and studied retinal ganglion cell rapid axonal transport with autoradiography and electron microscopy. Animals breathing room air at sea level pressure were compared to animals breathing 100% oxygen at 3 atm pressure in a hyperbaric chamber. Despite major increases in arterial oxygen levels in the hyperbarically oxygenated animals, both groups had axonal transport blockade at the optic nerve head. Anoxia appears not to be the most important cause of acute axonal damage induced by elevated IOP. The pattern of axonal abnormality within individual fiber bundles at the optic nerve head provides support for mechanical compression as a more likely alternative cause for induced neural damage.     

Efficacy and safety of memantine treatment for reduction of changes associated with experimental glaucoma in monkey, II: Structural measures

PURPOSE: To determine, using anatomic measurements, whether daily oral dosing with memantine is both safe and effective to reduce the injury associated with experimental glaucoma in primates. METHODS: Argon laser treatment of the anterior chamber angle was used to induce chronic ocular hypertension (COHT) in the right eyes of 18 macaque monkeys. Nine animals were daily orally dosed with 4 mg/kg memantine while the other nine animals received vehicle only. Measurements of intraocular pressure (IOP) from both eyes of all animals were made at regular intervals. Appearance of the optic nerve head, retinal vessels, and surrounding retina was documented with stereo fundus photographs obtained at multiple time points throughout the study. Measurements of optic nerve head topography were obtained from confocal laser scans made from animals with the highest IOPs at approximately 3, 5, and 10 months after elevation of IOP. At approximately 16 months after IOP elevation, animals were killed and histologic counts of cells in the retinal ganglion cell (RGC) layer were made. RESULTS: Histologic measurements showed that, for animals with moderate elevation of IOP, memantine treatment was associated with an enhanced survival of RGCs in the inferior retina. Measurements of optic nerve head topography showed less IOP-induced change in memantine-treated animals. This effect was seen in measurements of both the cup and the neuroretinal rim. A comparison of these same histologic and morphologic measurements in normotensive eyes from the two treatment groups showed that memantine treatment was not associated with any significant effects on these eyes. CONCLUSIONS: Histologic measurements of RGC survival as well as tomographic measurements of nerve head topography show that systemic treatment with memantine, a compound which does not lower intraocular pressure, is both safe and effective to reduce changes associated with experimental glaucoma.     

Time-dependent effects of elevated intraocular pressure on optic nerve head axonal transport and cytoskeleton proteins

PURPOSE: To study the time-dependent effects of elevated intraocular pressure (IOP) on axonal transport and cytoskeleton proteins in the porcine optic nerve head. METHODS: Fifteen pigs were used for this study. Rhodamine-beta-isothiocyanate was injected into the vitreous of each eye to study axonal transport. IOP in the left eye was elevated to 40 to 45 mm Hg, and IOP in the right eye was maintained between 10 and 15 mm Hg. Cerebrospinal fluid pressure was also continually monitored. IOP was elevated for 3 hours (n = 7) or 12 hours (n = 8) before animal euthanatization. Antibodies to phosphorylated neurofilament heavy (NFHp), phosphorylation-independent neurofilament heavy (NFH), neurofilament light, neurofilament medium (NFM), microtubule, and microtubule-associated protein (MAP) were used to study the axonal cytoskeleton. Confocal microscopy was used to compare axonal transport and cytoskeleton change between control and high IOP eyes in different laminar regions and quadrants of the optic nerve head. Results from these experiments were also compared with 6-hour elevated IOP data from an earlier study. RESULTS: Three hours of IOP elevation caused a decrease in NFH, NFHp, and NFM within laminar regions, with no demonstrable change in axonal transport. Changes to MAP and microtubules were only seen after 12 hours of IOP elevation. Axonal transport change occurred in a time-dependent manner with peripheral nerve bundle changes occurring earlier and being greater than central nerve bundle changes. CONCLUSIONS: Time-dependent changes in axonal transport and cytoskeletal structure in the optic nerve head provide further pathogenic evidence of axonal damage caused by elevated IOP.     

Understanding glaucomatous damage: anatomical and functional data from ocular hypertensive rodent retinas

Glaucoma, the second most common cause of blindness, is characterized by a progressive loss of retinal ganglion cells and their axons, with a concomitant loss of the visual field. Although the exact pathogenesis of glaucoma is not completely understood, a critical risk factor is the elevation, above normal values, of the intraocular pressure. Consequently, deciphering the anatomical and functional changes occurring in the rodent retina as a result of ocular hypertension has potential value, as it may help elucidate the pathology of retinal ganglion cell degeneration induced by glaucoma in humans. This paper predominantly reviews the cumulative information from our laboratory’s previous, recent and ongoing studies, and discusses the deleterious anatomical and functional effects of ocular hypertension on retinal ganglion cells (RGCs) in adult rodents. In adult rats and mice, perilimbar and episcleral vein photocauterization induces ocular hypertension, which in turn results in devastating damage of the RGC population. In wide triangular sectors, preferentially located in the dorsal retina, RGCs lose their retrograde axonal transport, first by a functional impairment and after by mechanical causes. This axonal damage affects up to 80% of the RGC population, and eventually causes their death, with somal and intra-retinal axonal degeneration that resembles that observed after optic nerve crush. Importantly, while ocular hypertension affects the RGC population, it spares non-RGC neurons located in the ganglion cell layer of the retina. In addition, functional and morphological studies show permanent alterations of the inner and outer retinal layers, indicating that further to a crush-like injury of axon bundles in the optic nerve head there may by additional insults to the retina, perhaps of ischemic nature.     

Effect of intraocular pressure on optic disc topography,electroretinography, and axonal loss in a chronic pressure-induced rat model of optic nerve damage

PURPOSE: To characterize the effect of intraocular pressure (IOP) on optic disc topography, retinal function, and axonal survival in a model of IOP-induced optic nerve damage in rat. METHODS: Hypertonic (1.75 M) saline was injected into an episcleral vein of one eye of 49 Brown Norway rats, with the fellow untreated eye serving as the control. During the 1 to 3 months of follow-up, IOP was measured twice weekly in conscious animals with a handheld tonometer, and changes in disc topography and retinal function were monitored with scanning laser tomography and electroretinography (ERG), respectively. Peak IOP elevation in the experimental eye compared with the fellow control eye (peak deltaIOP), integral of IOP elevation over time (deltaIOP integral), and days of IOP elevation were calculated. Axon counts were obtained from electron micrographs of the sectioned optic nerves. RESULTS: Progressive cupping was found in 9 (56.3%) of 16 eyes with peak deltaIOP of more than 15 mm Hg and in none of 21 eyes with peak deltaIOP less than 15 mm Hg. A strong correlation between deltaIOP integral and progressive cupping was also found, but not with days of IOP elevation. ERG abnormalities (limited to the b-wave) were found in 11 (64.7%) of 17 eyes with peak deltaIOP of more than 15 mm Hg and in 2 (8.7%) of 23 eyes with peak deltaIOP of less than 15 mm Hg. Neither of the other IOP parameters was predictive of ERG damage. The proportion of surviving axons was negatively correlated to both deltaIOP and deltaIOP integral (P 相似文献   

Axonal transport blockage by acute intraocular pressure elevation in monkeys]

The distribution of orthograde rapid axonal transport blockage in the optic nerve head by acute intraocular pressure (IOP) elevation in monkeys was studied by autoradiography. Tritiated leucine was injected intravenously two hours before IOP elevation, and the IOP was elevated for five hours, maintaining a perfusion pressure of 30mmHg. Serial step cross sections from the optic nerve head at the level of the lamina cribrosa were prepared for light microscopic autoradiography, and the accumulation and distribution of grains were quantitatively analyzed using computerized image analysis. The area of focal grain accumulation was expressed as a percent of the defined optic nerve area and each eight sectors, divided by axially intersecting vertical, horizontal, and diagonal lines, respectively. In eyes with IOP elevation, the mean area of focal accumulation of grains in the temporal half of the optic nerve was significantly larger, compared with the nasal half. Within axonal bundles, the focal high accumulation of grains was frequently identified to be in association with trabecular beams of connective tissue in the peripheral portion of the axonal bundle. These results suggest temporal dominant blockage of axonal transport in the optic nerve head, which may be induced by mechanical compression of the axonal bundles during IOP elevation.     

The optic nerve head in glaucoma: role of astrocytes in tissue remodeling

Primary open angle glaucoma is a common eye disease characterized by loss of the axons of the retinal ganglion cells leading to progressive loss of vision. The site of damage to the axons is at the level of the lamina cribrosa in the optic nerve head. The mechanism of axonal loss is unknown but elevated intraocular pressure and age are the most common factors associated with the disease. Previous studies in human glaucoma and in experimental glaucoma in monkeys have established a relationship between chronic elevation of intraocular pressure and remodeling of the optic nerve head tissues known clinically as cupping of the optic disc. This review focuses on the astrocytes, the major cell type in the optic nerve head. Astrocytes participate actively in the remodeling of neural tissues during development and in disease. In glaucomatous optic neuropathy, astrocytes play a major role in the remodeling of the extracellular matrix of the optic nerve head, synthesize growth factors and other cellular mediators that may affect directly, or indirectly, the axons of the retinal ganglion cells. Due to the architecture of the lamina cribrosa, formed by the cells and the fibroelastic extracellular matrix, astrocytes may respond to changes in intraocular pressure in glaucoma, leading to some of the detrimental events that underlie axonal loss and retinal ganglion cell degeneration.     

急性眼压升高所致视网膜、视神经及视交叉胶质细胞的早期反应

背景 中枢神经系统以及视网膜中的胶质细胞与神经元关系十分紧密,胶质细胞在神经元损伤和修复过程中发挥着重要作用.急性眼压升高引起的视网膜、视神经及视交叉各部位胶质细胞的早期反应特点以及其与视神经损伤的关系目前尚不清楚. 目的 探讨大鼠视网膜、视神经及视交叉的胶质细胞对急性高眼压的早期反应,同时观察神经前体细胞标志物巢蛋白( nestin)在反应性胶质细胞中的表达. 方法 成年雌性Wistar大鼠9只,分为正常对照组3只和急性高眼压组6只,急性高眼压组大鼠采用右眼前房灌注生理盐水的方法升高大鼠眼压至110 mmHg,持续60 min.于术后第3天和第7天用过量麻醉法处死各组动物各3只,摘出眼球分离视神经和大脑标本,并制作冰冻切片.利用Nissl染色的方法测量高眼压眼视网膜内层厚度,观察视网膜和视交叉的大体形态.用βⅢ-tubulin免疫荧光染色法标记视神经内的视网膜神经节细胞(RGCs)轴突,用胶质纤维酸性蛋白(GFAP)和nestin双重标记显示视网膜、视神经及视交叉的胶质细胞反应,并对两组结果进行比较.结果 正常大鼠的视网膜、视神经以及视交叉内均可见到一定量的GFAP阳性胶质细胞,但nestin的表达量很低.急性眼压升高后的第3天,视网膜内丛状层厚度明显变薄,RGCs数目较损伤前减少约46％.视网膜内胶质细胞GFAP的表达显著增加,细胞突起由神经纤维层伸展至整个视网膜,增生的胶质细胞内可见nestin的明显表达.视神经内RGCs轴突发生变性样改变,GFAP阳性胶质细胞内nestin的表达较眼压升高前明显增加.同损伤眼相对应的一侧视交叉的横断面积减小,出现大量星状GFAP和nestin共表达的胶质细胞.以上改变在眼压升高后第7天更趋明显.结论 急性眼压升高早期即可引起RGCs的丢失及轴突的变性,视觉神经元改变的同时伴随胶质细胞的反应,增生的胶质细胞表达神经前体细胞的标志物.视网膜与视神经和视交叉的改变在时间上具有一定的同步性.     

TNF-alpha-induced optic nerve degeneration and nuclear factor-kappaB p65

PURPOSE: To characterize a model of optic nerve axonal degeneration induced by tumor necrosis factor (TNF)-alpha and to determine the role of nuclear factor (NF)-kappaB p65 in axonal degeneration. METHODS: Groups of rats were euthanatized at 1 day, 1 or 2 weeks, or 1 or 2 months after intravitreal injection of TNF-alpha. Morphometric analyses of neurofilament- or Thy-1-positive cells, retinal ganglion cells (flat preparations stained with cresyl violet or retrograde labeling with a neurotracer), the number of axons, immunostaining for myelin basic protein, and TUNEL assays were performed. Levels of NF-kappaB p65 protein in retina and optic nerve were determined by Western blot analysis and immunohistochemistry. The effects of antisense oligodeoxynucleotide (AS ODN) against NF-kappaB p65 and helenalin, an inhibitor of NF-kappaB p65 activation, on TNF-alpha-induced optic nerve degeneration were determined by counting the number of axons. RESULTS: Intravitreal injections of TNF-alpha induced obvious axonal loss and extensive degeneration of the axons from 2 weeks to 2 months after injection, whereas significant retinal ganglion cell loss was noted only at 2 months after injection. NF-kappaB p65 was increased in the optic nerve but not in the retina and was found to colocalize with ED-1 and Iba1, markers of microglia. Inhibition of NF-kappaB p65 with AS ODN or helenalin significantly ameliorated the effects of TNF-alpha-mediated axonal loss. CONCLUSIONS: TNF-alpha causes axonal degeneration with probable delayed loss of retinal ganglion cell bodies. NF-kappaB p65 may play a pivotal role in axonal degeneration, with the possible involvement of microglial cells.     

神经营养素家族因子对视网膜神经细胞保护作用的研究进展

现有大量研究表明神经营养素家族因子对视网膜神经细胞具有重要的保护作用。在所有家族成员中,神经生长因子能够有效保护视网膜光感受器细胞;阻断视网膜神经节细胞中神经生长因子与受体p75的结合,可以抑制神经节细胞的凋亡。脑源性神经营养因子可以防止光感受器细胞变性,增强光感受器细胞损伤后的修复。在刺激神经节细胞轴突生长和突触形成方面,脑源性神经营养因子、神经营养素-3及神经营养素-4/5均具有显著效应。通过对神经营养素家族因子的研究,了解其作用机制,以期能够应用于视网膜神经细胞变性及损伤性疾病的治疗中。