Gary E. Korte

Epithelium-Capillary Interactions in the Eye: The Retinal Pigment Epithelium and the Choriocapillaris

Publisher Summary This chapter describes the evidence for epithelium–capillary interactions where the two are apposed, emphasizing structural and functional manifestations such as capillary permeability and cell polarity. The observations derived from human ocular histopathology and experimental animal models are focused in which interactions are evident between (1) the retinal-pigment epithelium (RPE) and its apposed capillary plexus, the choriocapillaris and (2) RPE and retinal capillaries experimentally brought into apposition with RPE, from which they are normally isolated. These observations are relevant to the pathogenesis of chorioretinal diseases like age-related macular degeneration and retinitis pigmentosa. The biology of the RPE-choriocapillaris interactions determines the relative contributions of ECM components, soluble factors, and phenotypically different types of RPE cells to observations made in situ. They are the two causes of reduced vision and blindness that arise at the RPE and lead to complicating secondary changes in the adjacent choriocapillaris and neural retina. The possible mechanisms of RPE–choriocapillaris interactions are also discussed in the chapter.

Choriocapillaris Atrophy after Experimental Destruction of the Retinal Pigment Epithelium in the Rat

Rats that receive intravenous injections of sodium iodate develop a retinopathy characterized by the partial loss of the retinal pigment epithelium (RPE). In thin sections examined by transmission electron microscopy the choriocapillaris atrophied adjacent to areas of RPE destruction. The endothelial cells thickened and lost their fenestrae and the lumen of the capillary was reduced. At sites where the RPE remained normal in appearance the choriocapillaris did not atrophy. Scanning electron microscopy of vascular casts of the choriocapillaris showed the coexistence of atrophic and normal choriocapillaris throughout the retina, presumably adjacent to sites where the RPE was destroyed or spared, respectively. Our observations support the concept that the RPE exerts some control over the structure and function of the choriocapillaris.

Regeneration of Mammalian Retinal Pigment Epithelium

Publisher Summary This chapter focuses on the Regeneration of mammalian retinal pigment epithelium (RPE). By regeneration it mean the replacement of the damaged RPE sheet by a new one, involving cell proliferation and migration to produce new cells that differentiate into epithelium with the structural and functional characteristics of normal RPE. This is opposed to the recovery of surviving but functionally compromised RPE cells, for example, which may produce retinal edema. Observations in mammals most directly elucidate the RPE response to injury during human retinal disease and the biology of RPE and photoreceptors transplanted into diseased retinas. The success of these efforts depends on the reestablishment of normal interactions between RPE, photoreceptors, and choriocapillaris. For example, normal RPE cells may exert a trophic effect that helps “rescue” photoreceptors when they are transplanted into the retina of rats with dysfunctional RPE. The plasma membrane reorganizes, or remodels, during RPE regeneration, This is seen as changes in surface specializations, the binding of plasma membrane probes such as lectins and ruthenium red, and the redistribution of plasma membrane components as the cells mature, such as the gradual enrichment of the apical plasma membrane in Na+K+-ATPase.

Remodelling of the retinal pigment epithelium in response to intraepithelial capillaries: evidence that capillaries influence the polarity of epithelium

SummaryLight- and urethane-induced retinopathies in rats are characterized by loss of photoreceptors. Retinal capillaries subsequently become incorporated into the normally avascular retinal pigment epithelium. These models provided an opportunity to study the response of epithelial cells to closely apposed capillaries, in order to determine if capillaries contribute to the polar organization of epithelial cells. Pigment epithelial cells reorganized their lateral plasma membrane where the latter faced intraepithelial capillaries. This normally flat, undifferentiated membrane developed attachment sites, folds and intracytoplasmic tubules, and exhibited endocytosis and putative basal lamina secretion. These structural and functional specializations are normally restricted to the basal plasma membrane — the normal vascular front of the cell facing the dense meshwork of capillaries constituting the choriocapillaris. We conclude that RPE cells, and perhaps epithelia in general, polarize in response to an adjacent capillary bed.

New ultrastructure of rat RPE cells: Basal intracytoplasmic tubules

Retinal pigment epithelial cells have prominent basal folds facing Bruchs membrane. In addition to folds I have observed intracytoplasmic tubules 60-90 nm in diameter in the basal cytoplasm of rat pigment epithelial cells. The tubules arise from the basal plasma membrane and open to the extracellular space. The tubules are most evident when intravenous horseradish peroxidase is used as a tracer. The tracer leaks out of the fenestrated choriocapillaris, into the extracellular space of the pigment epithelium and into the tubules. Electron microscopy at 1000 KV (High Voltage Electron Microscopy) confirms the tubular nature of these structures and their continuity with folds or the plasma membrane facing Bruchs membrane. The tubules are also observed in tissue not infiltrated with peroxidase. Morphometry shows that the tubules occupy about 21% of the surface area of the basal plasma membrane. Tubules appear plentiful where folds are reduced, and reduced where folds are plentiful; the tubules may be a different conformation of the normally slit-like fold extracellular space. The tubules are observed in all quadrants of the retina; centrally and peripherally; in young and adult rats and in pigmented and albino rats. The tubules function may be linked to that of the folds, from which many of them arise.

Choriocapillaris Regeneration in the Rabbit: A Study with Vascular Casts

Vascular casts were made of the choriocapillaris (CC) of rabbits that received sodium iodate intravenously 6-28 days prior to examination, in order to augment studies of CC regeneration in sectioned material. Regeneration of CC was evident 6 days after administration of iodate where zones of spared CC bordered zones of atrophic CC. Venular as well as capillary sprouts created foci of regenerating CC at this border. These foci to create an extended capillary plexus similar to mature CC. The observations corroborate those obtained in sectioned material as regards the geography of the CC response and the origins of new CC during the sodium iodate retinopathy, and provide new information on the way in which CC regenerates.

Reorganization of actin microfilaments and microtubules in regenerating retinal pigment epithelium

Retinal pigment epithelium (RPE) regenerating after experimental damage in rabbits exhibits major changes in cell shape, polarity and junctions--features that depend on the cytoskeleton. This report correlates these changes with the redistribution of actin microfilaments and microtubules, using electron microscopy and confocal laser scanning microscopy. We compare immature cells with the more mature cells that form the new epithelial monolayer. Two populations of immature RPE cells are interspersed at the edge of the regenerating RPE sheet. One population of immature cells makes few junctions with their neighbors or the basement membrane. They form pseudopodia and exhibit a prominent network of actin microfilaments beneath the plasma membrane. These cells are probably motile and advance the epithelial sheet. Another population of immature cells contains numerous stress fibers that insert into large basement membrane attachments. The cells make focal adhesions with their neighbors, rather than the junctional complexes characteristic of mature RPE cells. These cells are probably not motile and mature into the cells forming the new monolayer--cuboidal cells with numerous basal folds and apical villi and a complete belt of intercellular junctions. Stress fibers are lost as the circumferential bundle associated with the zonula adherens re-forms. Microtubules, which form prominent longitudinal bundles running through the processes of immature cells, take on the meshwork organization characteristic of mature RPE as the immature cells differentiate.

Expression of Plasma Membrane Alkaline Phosphatase in Normal and Regenerating Choriocapillaris in the Rabbit

Ultrastructural histochemistry for plasma membrane nonspecific alkaline phosphatase was performed on the normal and regenerating choriocapillaris (CC) of rabbits. In normal animals the CC endothelium expressed little or no staining, whereas in regenerating CC the endothelium exhibited staining. The staining was most intense at the unfenestrated plasma membrane. As the capillaries matured and the fenestrated plasma membrane became more extensive, the staining was reduced and eventually eliminated. Pericytes did not stain in normal or regenerating CC.

Permeability of Regenerating and Atrophic Choriocapillaris in the Rabbit

When rabbits receive intravenous injections of sodium iodate, large expanses of the retinal pigment epithelium are destroyed. The adjacent capillary bed, the choriocapillaris, atrophies in response to the loss of the pigment epithelium and then regenerates. This provides a model of the permeability of regenerating and atrophic choriocapillaris, which we studied using intravenously injected horseradish peroxidase and catalase. Regenerating capillaries were permeable to peroxidase but not catalase. The permeability to peroxidase was probably due to endothelial fenestrations, since catalase (which is larger than peroxidase and does not penetrate endothelial fenestrae) was retarded at interendothelial junctional complexes, indicating that they were intact. Atrophic choriocapillaries were impermeable to catalase but displayed a heterogeneous permeability to peroxidase. This was correlated with the presence or absence of fenestrae; capillary profiles lacking fenestrae retained peroxidase in their lumina, whereas if fenestrae were present the tracer penetrated into the pericapillary space. The observations indicate that: (1) the permeability of the regenerating choriocapillaris is qualitatively similar to the mature choriocapillaris, and (2) the atrophic choriocapillaris undergoes changes in permeability that are primarily correlated with the loss of endothelial fenestrae. The observations provide a functional correlate - change in permeability - for structural changes in choriocapillaris endothelium (thickening, loss of fenestrae) in response to destruction of the retinal pigment epithelium, which has been postulated to exert a trophic effect on these capillaries.

Repair of Retinal Pigment Epithelium and Choriocapillaries after Laser Photocoagulation: Correlations between Scanning Electron, Transmission Electron and Light Microscopy

Cell-to-cell interactions between retinal pigment epithelial (RPE) cells and vascular endothelium were examined by scanning electron microscopy, which demonstrates many facets of surface topography. The repair process after laser photocoagulation was characterized by a morphologically heterogeneous population of regenerated RPE cells, including normal-looking cells, macrophage-like cells and large elongated RPE cells. The macrophage-like and normal-looking RPE cells were predominantly seen adjacent to growing new choroidal vessels. Large cells appeared mainly in the vicinity of the extracellular matrix, where no viable new vessels were seen. The observed association between a specific phenotype of regenerated RPE cells and neovascularization may further support the hypothesis that regenerated RPE cells have a dual function, exerting both stimulatory and inhibitory effects on endothelial cells.