Ultrahigh resolution optical coherence tomography in non-exudative age related macular degeneration

AIM: To describe the appearance of the non-exudative forms of age related macular degeneration (AMD) as imaged by ultrahigh resolution optical coherence tomography (UHR-OCT). METHODS: A UHR-OCT ophthalmic imaging system, which utilises a femtosecond laser light source capable of approximately 3 mum axial resolution, was employed to obtain retinal cross sectional images of patients with non-exudative AMD. Observational studies of the resulting retinal images were performed. RESULTS: 52 eyes of 42 patients with the clinical diagnosis of non-exudative AMD were imaged using the UHR-OCT system. 47 of the 52 (90%) eyes had the clinical diagnosis of drusen and/or retinal pigment epithelial (RPE) changes. In these patients, three patterns of drusen were apparent on UHR-OCT: (1) distinct RPE excrescences, (2) a saw toothed pattern of the RPE, and (3) nodular drusen. On UHR-OCT, three eyes (6%) with a clinical diagnosis of non-exudative AMD had evidence of fluid under the retina or RPE. Two of these three patients had findings suspicious for subclinical choroidal neovascularisation on UHR-OCT. CONCLUSION: With the increased resolution of UHR-OCT compared to standard OCT, the involvement of the outer retinal layers are more clearly defined. UHR-OCT may allow for the detection of early exudative changes not visible clinically or by angiography.     

Semenogelins in the human retina: Differences in distribution and content between AMD and normal donor tissues

The two cellular targets of interest in age-related macular degeneration (AMD) are the photoreceptors and the RPE. However, the mechanisms involved in AMD pathology are not yet fully understood. In the present report, we extend our previous studies on semenogelin proteins (Sgs) in normal human retina and compare these with the distribution in retinas from AMD donor eyes. Semenogelins I (SgI) and II (SgII) are the major structural protein components of semen coagulum, but have been recently found in non-genital tissues as well. Cryo and paraffin sections of human retina were processed for both immunofluorescence and DAB reaction with a specific antibody. The presence of SgI was analyzed in retina and RPE total lysates and SgI was detected by western blot in human retina and RPE. The intensity of immunoreactivity was significantly reduced in the AMD eyes. SgI is expressed in the normal human retina and in the retina of AMD donor eyes, where localization was detected in the photoreceptors and in a few ganglion cells. We find the distribution of SgI in the AMD retinas substantially lower than observed in normal retina. SgI localization to photoreceptors and the RPE suggests a possible function related to the ability of these cells to sequester zinc.     

Studies on the retina and the pigment epithelium in hereditary canine ceroid lipofuscinosis, I. The distribution of enzymes in the whole retina and pigment epithelium

The massive accumulation of autofluorescent lipopigments, representative of autoxidation, is a key morphological feature in canine ceroid lipofuscinosis (CCL). In the eye peroxidase, catalase, and four acid hydrolases were compared with regard to aged and clinical condition in a series of English setters affected with CCL. In unaffected English setters "soluble" peroxidase increased in the RPE to adult levels at 2 yr of age. Affected dogs had higher RPE peroxidase activity earlier in life, which then decline with age. The soluble retinal peroxidase of both unaffected and CCL dogs increased steadily with age, but the latter group of dogs were much lower in activity. By 2 yr of age, RPE and retinal peroxidase values were only 25% and 47% of unaffected dog levels. Although the soluble enzyme of unaffected dogs exhibited a maturational profile, membrane-bound RPE peroxidase showed a hyperbolic curve reaching a maximum at 10 mo of age. By 2 yr of age, the "bound" enzyme in affected dogs was below unaffected levels in the RPE and retina. Three acid hydrolases were slightly increased in the RPE and retina of affected dogs. Acid lipase activity, however, was similar in both unaffected and CCL dogs. Catalase was not found in the RPE of either group of dogs. The catalase activity in the retina of both affected and unaffected dogs was at similar levels. Since catalase is not present in the RPE, the major defense against peroxidase accumulation and peroxide toxicity probably depends upon peroxidase. The present study indicates that a decrease in this key regulating enzyme may be related to the formation of lipopigments in the retina and RPE of dogs with CCL.     

Contribution of calpain to cellular damage in human retinal pigment epithelial cells cultured with zinc chelator

PURPOSE: We previously showed involvement of calpains in neural retina degeneration induced by hypoxia and ischemia-reperfusion. Age-related macular degeneration (AMD) is one of the leading causes for loss of vision. AMD showed degeneration of neural retina due to dysfunction and degeneration of the retinal pigment epithelium (RPE). RPE performs critical functions in neural retina, such as phagocytosis of shed rod outer segments. The purpose of the current study was to determine the contribution of calpain-induced proteolysis to damage in cultured human RPE cells. Zinc chelator TPEN was used to induce cellular damage as zinc deficiency is a suspected risk factor for AMD. METHODS: In RPE/choroid preparations from normal and AMD patients, calpain mRNAs were measured by qPCR, and calpain activity was assessed by casein zymography. Third- to fifth-passage cells from human RPE cells were cultured with TPEN. Cell damage was morphologically assessed under the phase-contrast microscope, and TUNEL staining was performed to detect apoptosis. Leakage of lactate dehydrogenase (LDH) into the medium was measured as a marker of RPE cell damage. Activation of calpains and proteolysis of the known calpain substrate alpha -spectrin were assessed by immunoblotting. To further confirm calpain-induced proteolysis, calpain in homogenized RPE was also activated directly by addition of calcium. RESULTS: RPE/choroid from normal patients expressed mRNAs for calpain 1, calpain 2, and calpastatin moderately, and calpain 2 activity tended to be lower in AMD patients. TPEN caused RPE cell damage with positive TUNEL staining. TPEN also caused leakage of LDH into the medium from RPE cells, and calpain inhibitor SJA6017 inhibited the leakage. Caspase-3 inhibitors z-VAD and z-DEVD also showed inhibitory effects. Immunoblotting for calpain and alpha -spectrin showed activation of calpain in RPE cells cultured with TPEN. Proteolysis by activated calpain was confirmed by addition of calcium to homogenized RPE. CONCLUSIONS: These results suggested that activation of calpain contributed to cellular damage induced by TPEN in cultured human RPE cells.     

Clinical patterns and electrophysiological findings in retinal pigment epithelium diseases

At present it is difficult to distinguish those human chorioretinal diseases in which the retinal pigment epithelium (RPE) is the primary site of dysfunction. This difficulty is caused by several factors such as scarcity of biochemical and histological information and a lack of correlation of basic science information available with the clinical body of knowledge.In the present study we examined 134 eyes at early or late stages of hereditary diseases involving the RPE. We tried to distinguish primary RPE involvement by using standard ERG (a- and b-wave) and EOG testing.We conclude that in general primary RPE damage can be better assessed by current electrophysiology in those diseases which seem to remain localized at the RPE level for a long time.     

The retinal pigment epithelium: Development,injury responses,and regenerative potential in mammalian and non-mammalian systems

Diseases that result in retinal pigment epithelium (RPE) degeneration, such as age-related macular degeneration (AMD), are among the leading causes of blindness worldwide. Atrophic (dry) AMD is the most prevalent form of AMD and there are currently no effective therapies to prevent RPE cell death or restore RPE cells lost from AMD. An intriguing approach to treat AMD and other RPE degenerative diseases is to develop therapies focused on stimulating endogenous RPE regeneration. For this to become feasible, a deeper understanding of the mechanisms underlying RPE development, injury responses and regenerative potential is needed. In mammals, RPE regeneration is extremely limited; small lesions can be repaired by the expansion of adjacent RPE cells, but large lesions cannot be repaired as remaining RPE cells are unable to functionally replace lost RPE tissue. In some injury paradigms, RPE cells proliferate but do not regenerate a morphologically normal monolayer, while in others, proliferation is pathogenic and results in further disruption to the retina. This is in contrast to non-mammalian vertebrates, which possess tremendous RPE regenerative potential. Here, we discuss what is known about RPE formation during development in mammalian and non-mammalian vertebrates, we detail the processes by which RPE cells respond to injury, and we describe examples of RPE-to-retina and RPE-to-RPE regeneration in non-mammalian vertebrates. Finally, we outline barriers to RPE-dependent regeneration in mammals that could potentially be overcome to stimulate a regenerative response from the RPE.     

Retinal pigment epithelium in the pathogenesis of age‐related macular degeneration and photobiomodulation as a potential therapy?

The retinal pigment epithelium (RPE) comprises a monolayer of cells located between the neuroretina and the choriocapillaries. The RPE serves several important functions in the eye: formation of the blood‐retinal barrier, protection of the retina from oxidative stress, nutrient delivery and waste disposal, ionic homeostasis, phagocytosis of photoreceptor outer segments, synthesis and release of growth factors, reisomerization of all‐trans‐retinal during the visual cycle, and establishment of ocular immune privilege. Age‐related macular degeneration (AMD) is the leading cause of blindness in developed countries. Dysfunction of the RPE has been associated with the pathogenesis of AMD in relation to increased oxidative stress, mitochondrial destabilization and complement dysregulation. Photobiomodulation or near infrared light therapy which refers to non‐invasive irradiation of tissue with light in the far‐red to near‐infrared light spectrum (630–1000 nm), is an intervention that specifically targets key mechanisms of RPE dysfunction that are implicated in AMD pathogenesis. The current evidence for the efficacy of photobiomodulation in AMD is poor but its safety profile and proposed mechanisms of action motivate further research as a novel therapy for AMD.     

Retinal pigment epithelium transplantation: concepts,challenges, and future prospects

The retinal pigment epithelium (RPE) is a single layer of cells that supports the light-sensitive photoreceptor cells that are essential for retinal function. Age-related macular degeneration (AMD) is a leading cause of visual impairment, and the primary pathogenic mechanism is thought to arise in the RPE layer. RPE cell structure and function are well understood, the cells are readily sustainable in laboratory culture and, unlike other cell types within the retina, RPE cells do not require synaptic connections to perform their role. These factors, together with the relative ease of outer retinal imaging, make RPE cells an attractive target for cell transplantation compared with other cell types in the retina or central nervous system. Seminal experiments in rats with an inherited RPE dystrophy have demonstrated that RPE transplantation can prevent photoreceptor loss and maintain visual function. This review provides an update on the progress made so far on RPE transplantation in human eyes, outlines potential sources of donor cells, and describes the technical and surgical challenges faced by the transplanting surgeon. Recent advances in the understanding of pluripotent stem cells, combined with novel surgical instrumentation, hold considerable promise, and support the concept of RPE transplantation as a regenerative strategy in AMD.     

老年性黄斑变性光学相干断层扫描观察

目的 观察萎缩型，渗出型老年黄斑变性（Age-related macular degeneration AMD）患者的光学相干断层扫描图像特征。比较光学相干断层扫描（Optical Coherence Tomography OCT）和荧光血管造影（Fluorescein angiography FFA）的特点，对脉络膜新生血管（Choroidal neovascularization CNV）进行OCT分型。方法 经FFA确诊的AMD57例76只眼进行OCT检查。结果 AMD患者色素上皮萎缩，软性玻璃膜疣，神经上皮和色素上皮脱离具有特有的OCT特征，OCT图像中视网膜神经上皮增厚、隆起反映视网膜下，视网膜层间积液。神经上皮或色素上皮（Retinal pigment epithelium，RPE）隆起，其下低反射区反映神经上皮或RPE层脱离。CNV的OCT图像分为边界清晰的CNV，边界模糊的CNV，纤维血管性RPE脱离。FFA中的典型性CNV相当于OCT图像中边界清晰CNV，隐匿性CNV相当于OCT图像中边界模糊CNV和纤维血管性RPE脱离。结论 OCT能特征性显示AMD中视网膜神经上皮隆起，视网膜层间积液，出血，神经上皮和RPE的脱离，且显示不同类型CNV的OCT特征。     

Comparison with indocyanine green angiography and immunohistochemical study of choroidal neovascular membrane in age-related macular degeneration

PURPOSE: We classified choroidal neovascular membranes (CNMs) (48 eyes) in age-related macular degeneration (AMD) into four types using indocyanine green angiography. METHODS: Surgically extracted CNMs were examined immunohistochemically. Specimens were stained with glial fibrillary acidic protein (GFAP), von Willebrand factor, Ki-67 antigen, actin, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (b-FGF), and transforming growth factor (TGF-beta 1). RESULTS: All types of CNM were positively stained with GFAP on the side opposite the retinal pigment epithelial (RPE) cells. This suggests the adherence of neurosensory retina to the CNM. Type I membranes which showed hyperfluorescence in the early and late phase of ICG angiography were significantly stained with Ki-67 antigen. CONCLUSION: These results indicate that CNMs in AMD are extracted with neurosensory retina and RPE cells during surgery. Type I membranes have a high capability for proliferation.     

Clinical electrophysiology of the retinal pigment epithelium

There is no ideal electrophysiological test for retinal pigment epithelial (RPE) function. The light-induced responses (EOG, c-wave, fast oscillation) that require photoreception are not pure RPE signals, and even the widely-used EOG has not been associated with any specific physiological disturbance of the RPE or retina. The discovery of non-photic RPE responses (hyperosmolarity, acetazolamide and bicarbonate) has enhanced the possibility of finding tissue-specific RPE tests, but these responses have yet to be correlated with specific RPE functional activity or pathology. We may face a dilemma in our search for RPE tests, insofar as electrophysiology measures membrane changes, but RPE membrane activity is related only indirectly to many functions of the RPE cell. These concerns notwithstanding, RPE electrophysiology can be a valuable clinical tool if one accounts for the physiological limitations and assets of the procedures.Presented at the 28th ISCEV Symposium, Guangzhou, China, 10–14 March 1990.     

Iron induced oxidative damage as a potential factor in age-related macular degeneration: the Cogan Lecture

Iron is a potent generator of oxidative damage whose levels increase with age, potentially exacerbating age-related diseases. Several lines of evidence suggest that iron accumulation may be a factor in age-related macular degeneration (AMD). AMD retinas have more iron within the photoreceptors, RPE, and drusen than do age-matched control retinas. Accelerated AMD-like maculopathy develops in patients with retinal iron overload from the hereditary disease aceruloplasminemia. Mice with retinal iron overload resulting from knockout of ceruloplasmin and its homologue hephaestin exhibit retinal degeneration with some features of AMD, including subretinal neovascularization, accumulation of RPE lipofuscin and sub-RPE deposits, and RPE/photoreceptor death. Increased understanding of the mechanisms of retinal iron homeostasis may help in the development of therapies to prevent iron overload. For example, herein it is shown that one regulator of systemic iron homeostasis, HFE, is expressed in the RPE. Thus, patients with the common disease hereditary hemochromatosis, which is often caused by an HFE mutation, may have retinal iron overload predisposing to AMD. Preliminary data suggest that iron chelation can reduce RPE iron overload in mice and protect them from degeneration, suggesting that iron-binding drugs may one day prove useful in reducing RPE oxidative stress and decreasing the risk of AMD progression.     

激光光凝后视网膜色素上皮、Bruch膜和脉络膜超微结构改变

目的观察激光光凝后视网膜色素上皮(RPE)、Bruch膜和脉络膜超微结构的改变和修复过程。方法收集眼眶恶性肿瘤患者7例,其眼球的视网膜及脉络膜结构基本正常。均经患者签署知情同意书后,于眶内容物剜除术前1、3、7d,用Ⅰ、Ⅱ、Ⅲ级光斑光凝视网膜。摘除眼球后,将其视网膜和脉络膜组织制成超薄切片、醋酸铀和枸橼酸铅染色,于透射电镜下观察其超微结构。结果激光光凝后1d,经Ⅰ级光斑光凝视网膜组织见部分RPE细胞肿胀、坏死、数量减少,微绒毛部分消失;Bruch膜完整;脉络膜毛细血管内皮肿胀,少数管腔闭塞。经Ⅱ级光斑光凝的视网膜组织见较多RPE细胞溶解破坏,Bruch膜空泡形成,脉络膜毛细血管广泛闭塞。经Ⅲ级光斑光凝的视网膜组织见大量RPE溶解消失,Bruch膜部分破坏,脉络膜血管受损。激光光凝后3d,RPE细胞和脉络膜水肿减退,破坏区内出现吞噬细胞,RPE细胞和脉络膜纤维组织开始增生。激光光凝后7d,增生的RPE细胞覆盖破坏区,脉络膜血管减少和基质纤维化。结论激光光凝可引起RPE细胞水肿、坏死,Bruch膜和脉络膜血管不同程度破坏,随后RPE细胞和纤维组织增生修复破坏区。不同参数的激光治疗可修复病变的RPE和脉络膜组织。（中华眼科杂志,2004,40：692-695)     

Fundus autofluorescence after selective RPE laser treatment

BACKGROUND: The selective RPE laser treatment is a new technique which selectively damages the RPE and avoids adverse effects to the neural retina. A problem is the ophthalmoscopically non-visibility of the laser lesions. The aim of the study was to investigate whether fundus autofluorescence (AF), which is derived from the lipofuscin contained by the RPE cells, is changed due to the RPE damage, and thus may be used for non-invasive treatment control. METHODS: A total of 26 patients with macular diseases, i.e. diabetic maculopathy (DMP), soft drusen maculopathy (AMD) and central serous retinopathy (CSR), were treated with repetitive short laser pulses (800 ns) from a green Nd:YAG laser (parameters: 532 nm, 100 and 50 pulses at 500 and 125 Hz, retinal spot diameter 200 micrometer, pulse energies 70-175 microJ). AF was excited by 488 nm and detected by a barrier filter at 500 nm (HRA, Heidelberg engineering). Patients were examined by ophthalmoscopy, fluorescein angiography and autofluorescence measurements at various times after treatment (i.e. 1 h, 1 and 6 weeks, 3, 6 and 12 months). RESULTS: None of the laser lesions was ophthalmoscopically visible during treatment although fluorescein angiography showed leakage of the irradiated areas. Identification of the lesions was possible by AF imaging showing an intensity decay in the irradiated area in 22 out of 26 patients, predominantly in patients with CSR and AMD. Lesions could be identified as hypoautofluorescent spots 1 h after treatment. During follow-up the laser spots became hyperautofluorescent. In patients with DMP some AF images were less helpful due to diffuse edema and larger retinal thickness. CONCLUSION: Imaging of non-visible selective RPE laser effects can be achieved by AF measurements predominantly in patients without retinal edema. Thus AF may replace invasive fluorescein angiography in many cases to verify therapeutic laser success.     

Localization of collagen XVIII and the endostatin portion of collagen XVIII in aged human control eyes and eyes with age-related macular degeneration

PURPOSE: Endostatin, a C-terminal fragment of collagen XVIII (coll XVIII) formed by proteolysis, specifically inhibits endothelial cell migration and proliferation in vitro and potently inhibits angiogenesis and tumor growth in vivo. The purpose of this study was to examine the immunolocalization of endostatin and coll XVIII in the retina and choroid of human donor tissue sections from aged control donor eyes and to determine whether the localization or relative levels are changed in age-related macular degeneration (AMD). METHODS: Ocular tissues were obtained from six aged control donors (age range, 75-86 years; mean age, 80.5 years) without evidence or history of chorioretinal disease and from nine donors with AMD (age range, 74-105 years; mean age, 88.6 years). Tissues were cryopreserved, and streptavidin alkaline phosphatase immunohistochemistry was performed with goat anti-human and mouse anti-human endostatin antibodies and rabbit anti-mouse coll XVIII. Blood vessels were identified with mouse anti-human CD-34 antibody in adjacent sections. Pigment in RPE and choroidal melanocytes was bleached. Three independent observers scored the immunohistochemical reaction product. RESULTS: In aged control eyes, coll XVIII and endostatin (the endostatin portion of coll XVIII) immunoreactivity was observed in large retinal blood vessels and in capillaries in some individuals, but the internal limiting membrane (ILM) had the most intense retinal immunostaining. There was no significant difference in immunoreactivity to both antibodies in retinal blood vessels in aged control eyes. In the choroid, endostatin and coll XVIII were localized to blood vessels, Bruch's membrane, and RPE basal lamina. AMD retina and choroid had a similar pattern and intensity of coll XVIII immunostaining, as observed in control eyes but reaction product was more diffuse in the choroid. Endostatin immunoreactivity was significantly higher in ILM (P = 0.037) in AMD retina and significantly lower in the choriocapillaris, Bruch's membrane, and RPE basal lamina of AMD choroids (P < 0.05) and completely negative in some areas of AMD choroids. CONCLUSIONS: These data suggest that reduced levels of the endostatin portion of coll XVIII in Bruch's membrane, RPE basal lamina, intercapillary septa, and choriocapillaris in eyes with AMD may be permissive for choroidal neovascularization.     

Autofluorescence imaging after selective RPE laser treatment in macular diseases and clinical outcome: a pilot study

AIM: Selective retinal pigment epithelium (RPE) laser treatment is a new technique which selectively damages the RPE while sparing the neural retina. One difficulty is the inability to visualise the laser lesions. The aim of the study was to investigate whether fundus autofluorescence (AF) is changed because of the RPE damage, and thus might be used for treatment control. Additionally, the clinical course of patients with various macular diseases was evaluated. METHODS: 26 patients with macular diseases (diabetic maculopathy (DMP), soft drusen maculopathy (AMD), and central serous retinopathy (CSR)) were treated and followed up for at least 6 months. Treatment was performed with a train of repetitive short laser pulses (800 ns) of a frequency doubled Nd:YAG laser (parameters: 532 nm, 50 and 500 pulses at 100 and 500 Hz, retinal spot diameter 200 micro m, pulse energies 75-175 micro J). AF was excited by 488 nm and detected by a barrier filter at 500 nm (HRA, Heidelberg Engineering, Germany). Patients were examined by ophthalmoscopy, fluorescein angiography, and autofluorescence measurements at various times after treatment (10 minutes, 1 hour, 1 and 6 weeks, 3, 6, and 12 months). RESULTS: Fluorescein angiography showed leakage from the irradiated areas for about 1 week after treatment. None of the laser lesions was ophthalmoscopically visible during treatment. Identification of the lesions was possible by AF imaging showing an intensity decay in the irradiated area in 22 out of 26 patients, predominantly in patients with CSR and AMD. Lesions could be identified 10 minutes after treatment as hypoautofluorescent spots, which were more pronounced 1 hour later. During follow up the laser spots became hyperautofluorescent. In patients with DMP some AF images were less helpful because of diffuse oedema and larger retinal thickness. In these cases ICG angiography was able to confirm therapeutic success very well. Most of the patients have had benefit from the treatment, with best results obtained for CSR patients. CONCLUSION: Imaging of non-visible selective RPE laser effects can be achieved by AF measurements predominantly in patients without retinal oedema. Therefore, AF may replace invasive fluorescein angiography in many cases to verify therapeutic laser success. Selective laser treatment has the potential to improve the prognosis of macular diseases without the risk of laser scotomas.     

Insulin-like growth factor-I and its receptor in neovascular age-related macular degeneration

PURPOSE: The insulin-like growth factor (IGF)-I protein is a growth-promoting polypeptide that can act as an angiogenic agent in the eye. The purpose of the current study was to localize the expression of IGF-I and its receptor (IGF-IR) mRNA and IGF-IR protein in situ in the normal human eye and to examine the presence of expression in eyes with neovascular age-related macular degeneration (AMD). METHODS: Formalin-fixed, paraffin-embedded slides of 4 normal control eyes and 14 eyes with choroidal neovascularization (CNV) secondary to AMD were examined. Three eyes with proliferative diabetic retinopathy were studied as the positive control. IGF-I and IGF-IR mRNA was detected by in situ hybridization with digoxigenin-labeled RNA probes. IGF-IR protein was studied by immunohistochemistry. RESULTS: In the normal retina, IGF-I and IGF-IR mRNA expression was found throughout the neuroretinal layers, in the retinal pigment epithelium (RPE), and in some choriocapillary and retinal capillary endothelial cells. In eyes with CNV we found IGF and IGF-IR mRNA in capillary endothelial cells, some transdifferentiated RPE, and fibroblast-like cells. IGF-IR protein was found in normal eyes in all neuroretinal layers, in the RPE, and in the choroidal vessels. In eyes with CNV, IGF-IR protein was present in the RPE monolayer, in transdifferentiated RPE, and in newly formed vessels. CONCLUSIONS: The colocalization of protein and receptor indicates an autocrine function of IGF-I in the normal human retina. Because IGF-I participates in ocular neovascularization, synthesis of IGF-IR and IGF-I in endothelial cells, RPE cells, and fibroblast-like cells in CNV may point toward a role for this growth factor in the pathogenesis of neovascular AMD.     

Proteomics of the retinal pigment epithelium reveals altered protein expression at progressive stages of age-related macular degeneration

PURPOSE: Age-related macular degeneration (AMD) is characterized clinically by changes in the retinal pigment epithelium (RPE), formation of drusen between the RPE and the underlying vasculature, geographic atrophy, and choroidal neovascularization. Later clinical stages are accompanied by impaired central vision. A limited understanding of the molecular events responsible for AMD has constrained the development of effective treatments. A proteomics approach was used to investigate the underlying mechanisms of AMD and to identify proteins exhibiting significant changes in expression with disease onset and progression. METHODS: Human donor eyes were categorized into one of four progressive stages of AMD. Proteins from the RPE were resolved and quantified by two-dimensional (2-D) gel electrophoresis. Proteins exhibiting significant expression changes at different disease stages were identified by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. 2-D and semiquantitative one dimensional (1-D) Western blot analyses were used to determine whether changes identified by the proteomic analysis were specific for a protein subpopulation or representative of the entire protein population. RESULTS: Proteins were identified from several critical pathways that changed at early and late disease stages, indicating potential causal mechanisms and secondary consequences of AMD, respectively. Proteins involved in protecting from stress-induced protein unfolding and aggregation, mitochondrial trafficking and refolding, and regulating apoptosis changed early in the disease process. Late-stage changes occurred in proteins that regulate retinoic acid and regeneration of the rhodopsin chromophore. CONCLUSIONS: These results provide the first direct evidence of AMD stage-specific changes in human RPE protein expression and provide a basis for functional investigation of AMD that may ultimately suggest new therapeutic strategies.     

Adult vitelliform macular degeneration: a clinicopathological study

AIMS/BACKGROUND: The yellow lesions of adult vitelliform macular degeneration (AVMD) slowly fade, progressing to hyperpigmentation or atrophy. This study aims to provide further observations on the location and nature of the vitelliform material. METHODS: This report describes the clinicopathological correlation of four eyes with AVMD. A retrospective histopathological study of a further 526 aged eyes previously graded for the stage of age-related macular degeneration (AMD) found another 10 eyes with similar pathology. RESULTS: The predominant finding was a collection of extracellular material beneath the sensory retina at the fovea. This material was derived internally from photoreceptor outer segments and externally from the retinal pigment epithelium (RPE), the latter first undergoing hypertrophy and then disruption and attenuation. Fallout of foveal cones occurred over these lesions and the inner retina was thinned, which may explain macular hole formation in this condition. All affected eyes showed histopathological evidence of AMD. CONCLUSIONS: This study confirms that the vitelliform lesions of AVMD lie beneath the sensory retina. In contrast to previous reports, however, it is proposed that the lesions comprise mainly extracellular material consisting of photoreceptor debris, possibly the result of faulty phagocytosis by the RPE, mixed with pigment liberated as the RPE undergoes disruption. The vitelliform lesions therefore are a marker for the area of maximal RPE disturbance.     

Stem cell based therapies for age-related macular degeneration: The promises and the challenges

Age-related macular degeneration (AMD) is the leading cause of blindness among the elderly in developed countries. AMD is classified as either neovascular (NV-AMD) or non-neovascular (NNV-AMD). Cumulative damage to the retinal pigment epithelium, Bruch's membrane, and choriocapillaris leads to dysfunction and loss of RPE cells. This causes degeneration of the overlying photoreceptors and consequential vision loss in advanced NNV-AMD (Geographic Atrophy). In NV-AMD, abnormal growth of capillaries under the retina and RPE, which leads to hemorrhage and fluid leakage, is the main cause of photoreceptor damage. Although a number of drugs (e.g., anti-VEGF) are in use for NV-AMD, there is currently no treatment for advanced NNV-AMD. However, replacing dead or dysfunctional RPE with healthy RPE has been shown to rescue dying photoreceptors and improve vision in animal models of retinal degeneration and possibly in AMD patients. Differentiation of RPE from human embryonic stem cells (hESC-RPE) and from induced pluripotent stem cells (iPSC-RPE) has created a potentially unlimited source for replacing dead or dying RPE. Such cells have been shown to incorporate into the degenerating retina and result in anatomic and functional improvement. However, major ethical, regulatory, safety, and technical challenges have yet to be overcome before stem cell-based therapies can be used in standard treatments. This review outlines the current knowledge surrounding the application of hESC-RPE and iPSC-RPE in AMD. Following an introduction on the pathogenesis and available treatments of AMD, methods to generate stem cell-derived RPE, immune reaction against such cells, and approaches to deliver desired cells into the eye will be explored along with broader issues of efficacy and safety. Lastly, strategies to improve these stem cell-based treatments will be discussed.