BTS clinical statement for the diagnosis and management of ocular tuberculosis.

The BTS clinical statement for the diagnosis and management of ocular tuberculosis (TB) draws on the expertise of both TB and and ophthalmic specialists to outline the current understanding of disease pathogenesis, diagnosis and management in adults. Published literature lacks high-quality evidence to inform clinical practice and there is also a paucity of data from animal models to elucidate mechanisms of disease. However, in order to improve and standardise patient care, this statement provides consensus points with the currently available data and agreed best practice.

Identification of ganglion cell neurites in human subretinal and epiretinal membranes.

AIM: To determine whether neural elements are present in subretinal and epiretinal proliferative vitreoretinopathy (PVR) membranes as well as in diabetic, fibrovascular membranes removed from patients during vitrectomy surgery. METHODS: Human subretinal and epiretinal membranes of varying durations were immunolabelled with different combinations of antibodies to glial fibrillary acidic protein, vimentin, neurofilament protein and laminin. RESULTS: Anti-neurofilament-labelled neurites from presumptive ganglion cells were frequently found in epiretinal membranes and occasionally found in subretinal membranes. In addition, the neurites were only observed in regions that also contained glial processes. CONCLUSIONS: These data demonstrate that neuronal processes are commonly found in human peri-retinal cellular membranes similar to that demonstrated in animal models. These data also suggest that glial cells growing out of the neural retina form a permissive substrate for neurite growth and thus may hold clues to factors that support this growth.

Glial remodeling and neural plasticity in human retinal detachment with proliferative vitreoretinopathy.

PURPOSE: To investigate glial remodeling and neuronal plasticity in adult human retinal detachment complicated by proliferative vitreoretinopathy (PVR) and to grade pathologic changes with a severity scoring system. METHODS: Sixteen full-thickness retinectomy specimens obtained at retinal relaxing surgery for PVR were fixed in 4% paraformaldehyde immediately after excision and compared to similarly processed normal donor retinas. Agarose-embedded sections (100-microm-thick) were double labeled for immunohistochemistry by confocal microscopy, with antibodies against rod opsin and GFAP; vimentin and M/L-cone opsin; calbindin D and S-cone opsin; and cytochrome oxidase and synaptophysin. These staining patterns formed the basis of a retinal pathology scoring system, and immunohistochemistry was also used to detect CD68, neurofilaments, protein kinase C, growth-associated protein-43, and a pan-cone-specific enzymatic marker. Morphology was also assessed by light microscopy of resin-embedded semithin sections. RESULTS: Prolonged detachment was characterized by photoreceptor degeneration and intracellular redistribution of opsin proteins to the plasma membrane in the outer nuclear layer (ONL). Remodeling of rod synaptic terminals was characterized by terminal retraction and also by axon extension to the inner retina in most specimens. Rod bipolar cell dendrites extended into the ONL, as did fine, horizontal cell processes. Large ganglion cells showed upregulated neurofilament and GAP-43 expression, with neurites sprouting from somata and axon collaterals. Anti-cytochrome oxidase labeling of surviving inner segments was reduced but detectable in all specimens, as was anti-calbindin D labeling of horizontal and amacrine cells. All specimens demonstrated a marked upregulation of Muller cell and astrocyte expression of GFAP and vimentin. More severe degenerative changes correlated with trauma and prolonged detachment duration when scored according to this system. CONCLUSIONS: The neural and glial components of detached neurosensory retina complicated by PVR exhibit pathology that changes characteristically with increasing detachment severity. Even in advanced degeneration, most of the structural motifs necessary for functional recovery are retained. Evidence of remodeling in the first-, second-, and third-order neurons of detached adult human retina may represent an attempt to re-establish synaptic connectivity.

Microglial cell activation following retinal detachment: a comparison between species.

PURPOSE: To compare the activation of microglia in response to retinal detachment in four species. METHODS: Experimental detachments were created in cats, rabbits, and ground squirrels and the retinas harvested 1, 3, 7, or 28 days later. Retinal reattachments of 28 days in duration were also performed in cats following a 3-day detachment. Human tissue was obtained during reattachment surgery. Microglia and macrophages were labeled with the lectins Griffonia simplicifolia and Ricinus communis and the antibody CD11b. Müller cell and photoreceptor responses were followed immunocytochemically on the same tissue sections labeled with microglial markers. Images were collected by laser scanning confocal microscopy. RESULTS: Lightly labeled microglia were observed primarily in the inner retina of control tissue. In the cat and rabbit, a progressive increase in the number of labeled cells occurred in the outer retina beginning at 1 day of detachment. In both long term human and cat detachments numbers of microglia were elevated throughout the retina. This is in contrast to the rabbit and ground squirrel retinas where microglial activation was dramatically diminished in longer term detachments. Presumptive macrophages (anti-CD11b labeled cells) occurred only in the subretinal space. Retinal reattachment in cats significantly attenuated the response except in areas of poor outer segment regeneration. CONCLUSIONS: The robust microglial response to retinal detachment is an indicator of the importance of this cell type in the overall response of the retina. Our data suggest that the feline retina is a particularly appropriate model system for understanding this response in humans. Inhibiting the microglial response in that species should help us understand more precisely its potential role in photoreceptor survival in human pathology.

Staining and peeling of the internal limiting membrane in the cat eye.

PURPOSE: To investigate the cat vitreomacular interface using trypan blue (TB) and indocyanine green (ICG) and to determine the validity of the cat model in terms of staining and peeling of the internal limiting membrane (ILM). METHODS: Lensectomy and vitrectomy were performed in four eyes of two cats. The ILM of two eyes was stained with TB (0.15%). ILM peeling was performed in one eye. Two eyes were stained with ICG (0.5%). One eye was illuminated for 3 min. Light and transmission electron microscopy and confocal microscopy were performed. RESULTS: Clinically, both dyes stained the cat ILM similar to human ILM. TB staining resulted in a normal ultrastructure and antigenity of the retina. ILM peeling was associated with intraretinal bleeding. There were fragments of Müller cells adherent to the retinal side of the ILM, and Müller cell endfeet were ruptured and avulsed. ICG staining of the ILM followed by illumination caused severe inner retinal damage. ICG without illumination resulted in focal ILM detachments associated with tearing of Müller cell endfeet. CONCLUSIONS: The cat can be used as a model to study the effect of TB and ICG on the central area of the cat retina, as previous results from clinical and experimental postmortem settings in human eyes were confirmed in the current study. Peeling of the ILM as a sheet as performed in human macular surgery is not feasible. Differences in the ultrastructure of the ILM and a strong adhesion of the ILM to Müller cell endfeet may account for this observation.

Experimental retinal reattachment: a new perspective.

In the feline model, retinal detachment initiates a cascade of changes that include photoreceptor- cell "deconstruction," apoptotic death of some photoreceptors, neurite outgrowth from second- and third-order neurons, remodeling of photoreceptor synaptic terminals, and Müller-cell gliosis. We have previously shown that reattachment within 24 h halts or reverses many of these presumed detrimental changes. However, in patients with retinal detachments, reattachment cannot always be performed within this 24-h window. Moreover, recovery of vision following successful reattachment surgery in the macula is often imperfect. Here, we examine the ability of relatively long-term reattachment (28 d) to stop or reverse several cellular events that occur at 3 d of detachment. In contrast to earlier studies of reattachment, which focused on the regeneration of outer segments, we focus our attention here on other cellular events such as neuronal remodeling and gliosis. Some of these changes are reversed by reattachment, but reattachment itself appears to stimulate other changes that are not associated with detachment. The implications of these events for the return of vision are unknown, but they do indicate that simply reattaching the retina does not return the retina to its pre-detachment state within 28 d.

Posterior vitreous detachment induced by microplasmin.

PURPOSE: To demonstrate the efficacy of microplasmin in inducing posterior vitreous detachment (PVD) and to evaluate the human and the feline retina after treatment. METHODS: Thirteen human donor eyes were injected with 62.5, 125, or 188 micro g microplasmin. The 13 fellow eyes received balanced salt solution. Four of the microplasmin-treated eyes received an additional intravitreal gas injection. After incubation at 37 degrees C for 30 minutes, all globes were placed in 4% paraformaldehyde. Retinal specimens were processed for scanning (SEM) and transmission (TEM) electron microscopy. Five feline eyes were injected with 14.5- or 25- micro g microplasmin. Animals were killed after 1 day, 3 days, or 3 weeks, and retinal specimens were evaluated by electron and confocal microscopy. RESULTS: In all control eyes, SEM demonstrated the cortical vitreous covering the inner limiting membrane (ILM). Intravitreal injection of 125 or 188 micro g microplasmin resulted in complete PVD. After treatment with 62.5 micro g microplasmin, SEM revealed collagen fibrils covering the ILM. Additional gas injection did not change the dose necessary for PVD. In vivo in cats, 25 micro g microplasmin resulted in complete PVD after 3 days. After 3 weeks, there was complete PVD with both doses of microplasmin. The retina and the ILM were well preserved in all eyes. CONCLUSIONS: Both after death and in vivo, microplasmin induces a dose-dependent cleavage between the vitreous cortex and the ILM without morphologic alterations of the retina. In the feline eye, there is no cellular response of retinal glial cells or neurons.

The ability of rapid retinal reattachment to stop or reverse the cellular and molecular events initiated by detachment.

PURPOSE: To determine the effects of reattachment on the molecular and cellular events initiated by a retinal detachment lasting 1 hour or 1 day. METHODS: Experimental retinal detachments were created in the right eyes of nine cats. Reattachments were performed 1 hour (n = 3) or 1 day (n = 3) after the detachment, and the animals were killed 3 days after detachment. Three-day detached (n = 3) and normal (n = 3) retinas were used for comparisons. Agarose-embedded sections were double labeled with a panel of antibodies. Some sections were also probed with the TUNEL technique to detect apoptotic cells. Wax-embedded sections were labeled with the MIB-1 antibody to the Ki67 protein to detect proliferating cells. RESULTS: The 1-hour and 1-day detachments followed by reattachment showed a very similar and consistent reduction in photoreceptor deconstruction and the Müller cell gliotic response when compared with 3-day retinal detachments without reattachment. Light microscopy and immunolabeling with opsin antibodies showed a significant reduction in both rod and cone outer segment (OS) degeneration, even though OS length was shorter than normal. The reattachments also showed a reduction in opsin redistribution, retraction of rod terminals, TUNEL-labeled photoreceptors, loss of cytochrome oxidase staining in photoreceptors, neurite outgrowth from second-order neurons, the number of proliferating cells, and the increase in intermediate filaments and loss of soluble proteins from Müller cells. The apparent re-ensheathing of the OS by the apical processes of the retinal pigment epithelium had begun but was not completely normal. CONCLUSIONS: These data indicate that, even though the length of the OS is less than normal, retinal reattachment within 1 day of detachment can either greatly retard or reverse many of the molecular and cellular changes initiated by detachment.

Matrix metalloproteinases in disease and repair processes in the anterior segment.

The pathogenesis of many anterior segment disorders and ocular complications following surgery are secondary to the wound healing response. The extent of clinical damage observed is closely related to the amount of scarring and tissue contraction. Matrix metalloproteinases (MMPs) are a family of enzymes that play a vital role in all stages of the wound healing process. They degrade all extracellular matrix components and also have the ability to synthesize collagen and extracellular matrix members, and are therefore important in the remodeling of a wound. Overexpression of MMPs results in excessive extracellular matrix degradation, leading to tissue destruction and loss of organ function. In the case of the anterior segment, this may mean the loss of visual function. This review focuses on the role MMPs have in the development of various anterior segment disorders. The importance of MMPs in the wound healing response and its potential modulation to manipulate the scarring response is being recognized, and current developments will be described.

A standardized digital photography system with computerized eyelid measurement analysis.

BACKGROUND: The authors have developed a slit-lamp mounted digital photography system. This ensures that the patient's head is consistently positioned in the same plane in relation to the camera and generates standardized images of the eyelids, which are then analyzed using commercially available computer measurement software. The aim of this study was to assess the repeatability and reproducibility of this system compared with traditional methods of eyelid measurement. METHODS: Both eyes of 10 patients were photographed and then measured by three clinicians using a handheld ruler for five eyelid parameters (i.e., palpebral aperture, superior marginal reflex distance, inferior scleral show, levator function, and upper eyelid skin crease height). The photographs were then assessed using computer software by the same clinicians (twice by a consultant and once by a clinical fellow and a specialist registrar). At all times, each observer was masked to their colleagues' results. Data were analyzed using Bland-Altman plots. RESULTS: There was a good level of agreement between measurements obtained by computer analysis of digital photographs and measurements obtained by traditional methods. A higher level of reproducibility (interobserver variability) was demonstrated in all digital measurements compared with those obtained by handheld ruler. Repeatability (intraobserver variability) with the digital photograph technique was also high and consistent for each of the five eyelid parameters. CONCLUSIONS: These results suggest that a digital photography system with computer analysis is comparable to, and offers advantages over, traditional methods of measurement. This system offers a simple, standardized, and rapid method of patient assessment with important applications in electronic patient records, audit, and research.

Intraretinal and periretinal pathology in anterior proliferative vitreoretinopathy.

OBJECTIVES: To determine the intraretinal and periretinal pathological changes in early anterior proliferative vitreoretinopathy (APVR). DESIGN: Observational case series. PARTICIPANTS: Eighteen patients undergoing retinectomy for APVR. METHODS: Retinectomy specimens removed at vitrectomy surgery were analysed by (a) semithin light microscopy, (b) immunohistochemistry and (c) electron microscopy. RESULTS: The specimens showed consistent outer retinal degenerative changes, marked Muller cell hypertrophy and glial continuity to epiretinal membranes. Photoreceptor outer and inner segments were markedly disrupted and occasional photoreceptor nuclear had pyknosis and chromatin clumping consistent with apoptosis. Muller cells expressed upregulated levels of glial fibrillary acid protein (GFAP) and extended through glial bridges to complex epiretinal membranes which in some areas had a bilaminar structure with a glial-negative inner lamina. CONCLUSION: Retinal degeneration and photoreceptor apoptosis occur in retinal detachment complicated by proliferative vitreoretinopathy (PVR), although during the early stages of the process neural retinal cells remain present, suggesting potential for recovery. The intraretinal glial response appears to be centrally involved in the formation of contractile epiretinal membranes. The retina retains the capacity for a degree of functional recovery following surgery for PVR. Surgical separation of anterior epiretinal membranes in PVR may be difficult and incomplete and alternative surgical strategies may be necessary to prevent recurrence.

Complement factor H deficiency in aged mice causes retinal abnormalities and visual dysfunction.

Age-related macular degeneration is the most common form of legal blindness in westernized societies, and polymorphisms in the gene encoding complement factor H (CFH) are associated with susceptibility to age-related macular degeneration in more than half of affected individuals. To investigate the relationship between complement factor H (CFH) and retinal disease, we performed functional and anatomical analysis in 2-year-old CFH-deficient (cfh(-/-)) mice. cfh(-/-) animals exhibited significantly reduced visual acuity and rod response amplitudes on electroretinography compared with age-matched controls. Retinal imaging by confocal scanning laser ophthalmoscopy revealed an increase in autofluorescent subretinal deposits in the cfh(-/-) mice, whereas the fundus and vasculature appeared normal. Examination of tissue sections showed an accumulation of complement C3 in the neural retina of the cfh(-/-) mice, together with a decrease in electron-dense material, thinning of Bruch's membrane, changes in the cellular distribution of retinal pigment epithelial cell organelles, and disorganization of rod photoreceptor outer segments. Collectively, these data show that, in the absence of any specific exogenous challenge to the innate immune system, CFH is critically required for the long-term functional health of the retina.

Acute macular pucker.

PURPOSE: To describe the presenting features, histopathology, and surgical outcome in a group of patients with rapidly progressive macular pucker. DESIGN: Retrospective interventional noncomparative case series. PARTICIPANTS: Five patients. METHODS: Review of case notes and the existing literature. RESULTS: All five patients had rapidly progressive visual loss and metamorphopsia over 2 weeks to 3 months, secondary to macular pucker after retinal tears or detachment. Vitrectomy and epiretinal membrane removal was performed within 1 month of diagnosis. In the absence of complications, there was rapid recovery of the visual acuity with resolution of metamorphopsia within 6 weeks to 3 months. Surgical complications limited the visual outcome in two cases. Histopathologic examination of epiretinal membrane removed from two of the cases suggests that these tend to form tubuloacinar structures and contain more retinal pigment epithelium-derived cells than tissue excised from cases with idiopathic macular pucker. CONCLUSIONS: Patients with acute macular pucker have precipitous visual loss caused by epiretinal membrane formation after retinal tear or detachment. Early surgery in these patients results in rapid recovery of visual acuity and resolution of metamorphopsia. The clinical features and comparative immunohistochemistry suggest that acute macular pucker is a distinct clinicopathologic entity.

The Leber congenital amaurosis gene product AIPL1 is localized exclusively in rod photoreceptors of the adult human retina.

Leber congenital amaurosis (LCA) is the most severe inherited retinal dystrophy resulting in markedly impaired vision or blindness at birth. LCA is characterized by an extinguished electroretinogram in infancy, which is thought to be indicative of an early and severe impairment of both the rod and cone photoreceptors in the human retina. Recently, the aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) gene was identified as the fourth causative gene of LCA. AIPL1 encodes a 384 amino acid protein of unknown function. We have generated a polyclonal antibody against a peptide from a unique region within the primate AIPL1 protein, which detects a protein of approximately 43 kDa in human retinal extracts. A screen of human tissues and immortalized cell lines with this antibody reveals AIPL1 to be specific to human retina and cell lines of retinal origin (Y79 retinoblastoma cells). Within the retina, AIPL1 was detected only in the rod photoreceptor cells of the peripheral and central human retina. The AIPL1 staining pattern extended within the rod photoreceptor cells from the inner segments, through the rod nuclei to the rod photoreceptor synaptic spherules in the outer plexiform layer. AIPL1 was not detected in the cone photoreceptors of peripheral or central human retina. This study is the first to suggest that AIPL1 performs a function essential to the maintenance of rod photoreceptor function.