G.P. Lewis

Mechanisms of photoreceptor death and survival in mammalian retina

The mammalian retina, like the rest of the central nervous system, is highly stable and can maintain its structure and function for the full life of the individual, in humans for many decades. Photoreceptor dystrophies are instances of retinal instability. Many are precipitated by genetic mutations and scores of photoreceptor-lethal mutations have now been identified at the codon level. This review explores the factors which make the photoreceptor more vulnerable to small mutations of its proteins than any other cell of the body, and more vulnerable to environmental factors than any other retinal neurone. These factors include the highly specialised structure and function of the photoreceptors, their high appetite for energy, their self-protective mechanisms and the architecture of their energy supply from the choroidal circulation. Particularly important are the properties of the choroidal circulation, especially its fast flow of near-arterial blood and its inability to autoregulate. Mechanisms which make the retina stable and unstable are then reviewed in three different models of retinal degeneration, retinal detachment, photoreceptor dystrophy and light damage. A two stage model of the genesis of photoreceptor dystrophies is proposed, comprising an initial depletion stage caused by genetic or environmental insult and a second late stage during which oxygen toxicity damages and eventually destroys any photoreceptors which survive the initial depletion. It is a feature of the model that the second late stage of retinal dystrophies is driven by oxygen toxicity. The implications of these ideas for therapy of retinal dystrophies are discussed.

Basic fibroblast growth factor: a potential regulator of proliferation and intermediate filament expression in the retina

Proliferation of astrocytes, and a concomitant increase of intermediate filaments in astrocytes are two fundamental responses of the CNS to injury. We have previously identified these two events in the retinas response to detachment of the neural retina from the adjoining monolayer of retinal pigmented epithelium. In order to analyze the potential role of basic fibroblast growth factor (bFGF) in these responses, we studied cellular proliferation and intermediate filament protein expression in the retinas of cats and rabbits 4 d and 4 weeks after a single intravitreal injection of 1 microgram of bFGF. Our results show that bFGF stimulates both of these processes in an otherwise normal eye. The eyes that received bFGF had significantly elevated numbers of 3H-thymidine-labeled Muller cells, astrocytes, vascular cells, retinal pigmented epithelial cells, microglia, and macrophages by comparison to control eyes. This proliferation was apparent at 4 d after the injection of bFGF but not after 4 weeks. In control eyes, antibodies to glial fibrillary acidic protein and vimentin labeled intermediate filaments only in the inner (vitread) portion of the Muller cells, the specialized radial astrocytes that span the width of the retina. In eyes that had been injected with bFGF, almost the entire Muller cell cytoplasm was labeled at 4 d after injection; after 4 weeks, the cytoplasmic labeling intensity had increased significantly. Release or activation of endogenous stores of bFGF after injury or disease may be involved in the control of cellular proliferation and intermediate filament expression in the retina and elsewhere in the CNS.

Proliferative vitreoretinopathy—developments in adjunctive treatment and retinal pathology

Proliferative vitreoretinopathy (PVR) remains a difficult management problem despite advances in vitreoretinal surgery. There is still a significant incidence of PVR in rhegmatogenous retinal detachment and other forms of retinal disease. Surgery for PVR now has a high anatomical success rate although visual results are often disappointing. The use of adjunctive treatments to prevent cellular proliferation holds promise for the prevention of PVR or recurrences after surgery. Control of proliferation and strategies aimed at improving visual outcome are important areas of future research in PVR and other forms of retinal disease. Studies of the intraretinal and peri-retinal pathology of PVR have demonstrated characteristic changes which may have a significant influence on visual outcome and surgical management.

Animal models of retinal detachment and reattachment: identifying cellular events that may affect visual recovery

Retinal detachment continues to be a significant cause of visual impairment, either through the direct effects of macular detachment or through secondary complications such as subretinal fibrosis or proliferative vitreoretinopathy. Animal models can provide us with an understanding of the cellular mechanisms at work that account for the retinopathy induced by detachment and for the generation of secondary effects. As we understand the mechanisms involved, animal models can also provide us with opportunities to test therapeutic agents that may reduce the damaging effects of detachment or improve the outcome of reattachment surgery. They may also reveal information of use to understanding other causes of blindness rooted in retinal defects or injuries. Understanding the effects of detachment (and reattachment) are likely to become even more important as surgeons gain skills in subretinal surgical techniques and macular translocation, both of which will generate short-lived detachments. Here we discuss the fundamental events that occur after detachment, present changes associated with reattachment, and discuss retinal changes that may affect the return of vision.

Brilliant violet fluorophores: A new class of ultrabright fluorescent compounds for immunofluorescence experiments †

The Nobel Prize in Chemistry was awarded in 2000 for the discovery of conductive organic polymers, which have subsequently been adapted for applications in ultrasensitive biological detection. Here, we report the first use of this new class of fluorescent probes in a diverse range of cytometric and imaging applications. We demonstrate that these “Brilliant Violet” reporters are dramatically brighter than other UV‐violet excitable dyes, and are of similar utility to phycoerythrin (PE) and allophycocyanin (APC). They are thus ideally suited for cytometric assays requiring high sensitivity, such as MHC‐multimer staining or detection of intracellular antigens. Furthermore, these reporters are sensitive and spectrally distinct options for fluorescence imaging, two‐photon microscopy and imaging cytometry. These ultra‐bright materials provide the first new high‐sensitivity fluorescence probes in over 25 years and will have a dramatic impact on the design and implementation of multicolor panels for high‐sensitivity immunofluorescence assays. Published 2012 Wiley Periodicals, Inc.

Ganglion cell neurites in human idiopathic epiretinal membranes

Aim: To identify and confirm the presence of neural elements in idiopathic epiretinal membranes removed from patients’ eyes during vitrectomy with epiretinal membrane peeling. Methods: Human epiretinal membranes from patients with no other known eye disease and of varying durations were labelled immunohistochemically with antibodies for neurofilament protein, laminin and either vimentin or GFAP; proteins expressed in ganglion cells, the inner limiting membrane (ILM), and Muller cells, respectively. Results: Anti-neurofilament labelled neurites, presumed to originate from ganglion cells, were found in all 32 idiopathic epiretinal membranes examined. The neurites were only observed in regions of anti-vimentin or -GFAP labelled glial cells, both of which were observed embedded in anti-laminin labelled material assumed to originate from the ILM. Conclusions: We show that neurofilamentous processes, presumed to originate from retinal ganglion cells, are found universally in idiopathic epiretinal membranes, suggesting that the presence of these membranes is sufficient to stimulate neurite growth in the absence of trauma or disease. In addition, since neurites were invariably found in association with glial cells, the glia may play a permissive role in neurite growth both within the retina and into extra-retinal glial membranes.

Clinicopathological case series of four patients with inherited macular disease

PURPOSEnTo correlate the phenotype of four patients with inherited macular disease with the immunohistopathology of retinal tissue collected at the time of retinal pigment epithelium (RPE)-choroidal transplantation.nnnMETHODSnA clinicopathologic case series describing the phenotype of four patients, including confocal immunohistochemistry and electron microscopy (EM), and the results of genetic testing.nnnRESULTSnIn Case 1, electrophysiology showed only macular dysfunction. Confocal microscopy revealed minor abnormalities. EM showed abnormal cone inner segments with swollen mitochondria. In case 2 (R172W mutation in RDS), electrophysiology demonstrated generalized cone system dysfunction with severe macular involvement. Peripherin labeling of outer segments was nonuniform, and EM showed discs arranged in whorllike structures. Case 3 showed severe central macular dysfunction on multifocal electroretinogram (ERG). Peripherin staining was irregular and disorganized. EM revealed abnormal inner segment morphology, particularly in rods, and disorganized irregular outer segments. Case 4 had localized central macular dysfunction on multifocal ERG. Confocal microscopy was grossly normal, with evidence of early redistribution of cone opsin to the inner segment. EM showed variable rod morphology and normal cones.nnnCONCLUSIONSnRPE transplantation provides a unique opportunity to gain insight into retinal disorders by enabling phenotypic correlation with the immunohistopathology of retinal tissue collected during surgery.

Histological analysis of retinas sampled during translocation surgery: a comparison with normal and transplantation retinas.

Aims: To carry out a histopathological analysis of retinal specimens of patients undergoing translocation surgery for age-related macular degeneration (ARMD). Methods: A histopathological analysis, using confocal microscopy, was performed on six retinal specimens. Results were compared with those from two further retinal specimens, collected during RPE transplantation, to control for the effects of vitrectomy and ARMD. In addition, a third control specimen from a cadaver with no history of ophthalmic disease was also analysed. Results: In the translocation specimens, rods and cones were relatively well preserved but showed reduced density and outer segment length. In four specimens, there were focal areas of rod opsin redistribution to the inner segment, but this was not observed in the controls. Staining with calbindin was decreased in cones compared with controls but normal in horizontal and amacrine cells. Rod bipolar cells were mildly disorganised, and in one there was evidence of neurite sprouting. Glial fibrillar acidic protein was raised in both translocation and transplantation retinae but not in the cadaver control. Conclusions: In this study, there was little evidence of cellular injury following iatrogenic detachment; however, the rate of PVR following translocation surgery infers that cellular events set in motion may continue despite early reattachment.

Injury and Repair Responses: Retinal Detachment

Injury to the mammalian retina produces a number of cellular wound-healing types of responses, with similarities and differences to injury responses in the brain and spinal cord. Retinal detachment is one such serious, sight-threatening injury that produces responses from almost all cell types including cellular degeneration and death, neuronal remodeling, some poorly understood immune responses, and glial cell reactivity. Glial cell reactivity is particularly important in glial scar formation which may, in turn, have future downstream effects on visual recovery. The glial cell response is prominent in all injuries to the central nervous system and, yet, its exact purpose and regulatory mechanisms are poorly understood.