Experimental Approaches to Diabetic Retinopathy

H. -P. Hammes, M. Porta

• Publisher: S. Karger
• Year: 2009
• ISBN:
  9783805592765
• Topic: Medicine
• Subtopic: Endocrinology & Metabolism

This volume sets the stage for clinical experts working with diabetic patients as well as for researchers by describing the clinical presentations of retinopathy and their anatomical and functional correlates. It reviews currently available experimental models in animals. The impact of retinal pericytes, neuroglia and, specifically, Müller cells are discussed in detail. The volume addresses a variety of current scientific discussions about mechanisms of damage such as growth factors and the VEGF/PEDF balance in the diabetic eye, the ocular renin-angiotensin system, and leukocyte interactions with the microvasculature among others. Stem and progenitor cells in the retina are discussed as potential directions for future investigation. The final chapters return to emerging clinical aspects, including current approaches to retinopathy as a predictor of cardiovascular risk and how knowledge can be translated from bench to bedside.

Hammes H-P, Porta M (eds): Experimental Approaches to Diabetic Retinopathy.
Front Diabetes. Basel, Karger, 2010, vol 20, pp 20–41

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Retinal Vascular Permeability in Health and Disease

Vassiliki Poulaki

Retina Research Laboratory, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, Mass., USA

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Abstract

Homeostasis in the retina microenvironment is maintained by the proper function of the blood-retinal barrier (BRB), which regulates the movement of chemicals and cells between the intravascular compartment and the retina. The BRB consists of two major topographically distinct components: the endothelium of the retinal vessels (inner BRB) and the retinal pigment epithelium (outer BRB). The barrier function of the retinal vascular endothelium depends on its lack of fenestrations, whereas the ability of the retinal pigment epithelium to regulate solute transport depends on the apical tight junctions. The tight junctions are membrane fusion areas between adjacent cells that serve as a diffusion barrier for paracellular transport and as a ‘molecular fence’, restricting the free movement of transmembrane proteins, and thus maintaining cell polarity and the asymmetric distribution of transmembrane proteins. Among the most important proteins that are associated with tight junctions are occludin, zonula occludens and claudins. Pathologic increase in blood retinal permeability can be caused by endothelial or pericyte cell death, tight junction disassembly, or cytokines such as vascular endothelial growth factor. Several assays have been developed to allow detection, quantification and monitoring of BRB breakdown in experimental and clinical settings. Assays used in animal models include the injection of chromophores, such as Evans blue, horseradish peroxidase, and fluorescein; the imaging techniques include electron microscopy and MRI. In humans, fluorescein angiography, vitreous fluorophotometry and optical coherence tomography are most commonly used. The disruption of the BRB contributes to the pathophysiology of several retinal diseases such as diabetic retinopathy, age-related macula degeneration, retinopathy of prematurity, central serous chorioretinopathy, vascular occlusive and inflammatory diseases. Several medical and surgical treatments have been developed to restore normal BRB function. Traditional procedures such as laser photocoagulation and corticosteroids have been recently supplemented with vascular endothelial growth factor pathway inhibitors, anti-TNF-α agents, mammalian target of rapamycin inhibitors and PKCβ inhibitors. Early results from clinical trials offer hope for effective vision-preserving therapies.

Copyright © 2010 S. Karger AG, Basel

Although the mammalian retina is constantly exposed to the rich choroidal circulation, it maintains a high level of electrolyte and metabolite balance that is crucial for the proper retinal function and ultimately vision. This homeostasis is maintained by the proper function of the blood-retinal barrier (BRB) that regulates the transport of cells and chemical substances from the circulation to the retina, therefore the retinal microenvironment. The molecular basis for the BRB are tight junctions (TJs) between endothelial cells in the inner retina, and between pigmented epithelial cells in the outer retina. The disruption of the BRB in the retinal vas-culature or in neovessels underlies the pathophysiology of a variety of vision-threatening diseases of the retina. Restoration of the vascular stability and integrity improves visual outcomes and is currently a therapeutic goal for many ocular conditions.

Physiology of the Retinal Vascular Network

The retina is a highly specialized neural tissue that consists of seven layers: the nerve fiber layer, the ganglion cell layer, the inner plexiform layer, the inner nuclear layer, the outer plexiform layer, the outer nuclear layer and the photoreceptors (rods and cones). The majority of the retina blood supply (85%) is derived from the choroidal blood vessels, whereas the central retinal artery provides the remaining 15%. The central retinal artery gives out four main vessels as it runs through the optic nerve head and supplies three capillary networks: the radial peripapillary, the inner and the outer network. The most superficial capillary network is the radial peripapillary one, which runs in the inner part of the nerve fiber layer along the major arterial arcades. The inner capillary network runs in the ganglion cell layer, whereas the outer capillary network runs throughout the inner nuclear layer. The three networks form multiple anastomoses between them. The retinal area responsible for central vision is located in the center of the macula, called the fovea; it is avascular and the retinal vessels arc around it. The choroidal vasculature consists of fan-shaped lobules of capillaries derived from the long and short posterior ciliary arteries and from branches of the peripapillary arterial network.

Physiology of the Blood-Retinal Barrier

The BRB maintains a constant milieu by regulating the exchange of water, nutrients, metabolites, proteins and neurotransmitters, and the efflux of toxic byproducts of metabolism. Moreover, it shields the neural retina from the circulating blood by restricting the entry of toxins, inflammatory cytokines, antibodies and circulating immune cells. The concept of the existence of the blood-tissue barrier in neural tissues was first introduced in the literature in 1885 by Goodman who demonstrated that trypan blue injected intravenously in the rat stained all tissues except the brain [1, 2]. In 1965, Ashton and Cuhna-Vaz demonstrated that intravenously injected histamine increased the vascular permeability of various ocular tissues except the retina [3], leading to the concept of the BRB [2]. Subsequent morphological studies showed that the retinal endothelial cells demonstrate an epithelial-like structure with ‘zonnulae occludentes’ between them.

Maurice and Cunha-Vaz performed morphological studies and permeability measurements and proposed that the BRB consists of two major components: the endothelium of the retinal vessels (inner BRB) and the retinal pigment epithelium (RPE; outer BRB) [2]. These two components are topographically distinct (the former is responsible for BRB functions in the inner retina, whereas the latter for the outer retina) and mechanistically independent. Therefore, it should be emphasized that the two different yet parallel sources of perfusion in the retina (the choroidal blood vessels and the central retina artery) are dependent on different mechanisms of the BRB: the endothelial cells of the choroidal capillaries have fenestrations similar to those of endothelial cells elsewhere in the body and rely entirely on the adjacent RPEs for BRB functions. In contrast, the endothelial cells of the retinal network capillaries lack fenestrations and exhibit all the specialized barrier properties of the BRB, while their surrounding pericytes, which contribute to a second line of defense in the blood-brain barrier, are approximately four times as numerous in the retina as in the brain [4].

There are no diffusional barriers between the extracellular fluid of the retina and the adjacent vitreous, and the vitreous body does not hinder significantly the diffusion of solutes. It should be emphasized that not all aspects of the physiology of BRB have been well studied in a retina-related model. Several conclusions are derived from extrapolation based on observations in other natural barriers, such as the blood-brain barrier.

Molecular Biology of the Blood-Retinal Barrier

The main routes used by water, solutes and proteins to move across endothelial and epithelial cell layers can be classified as transcellular vs. paracellular flux. Transcellular (transfer across the cell) can be via passive diffusion, facilitated diffusion (channel-facilitated transport), active transport (receptor-mediated uptake), endocytosis/pinocytosis (membrane invaginations across the cell surface that pinch off to form vesicles that move to the cell interior and are released on the other side, allowing nonspecific transport of material), and finally via pores or fenestrations. It should be noted that RPE cells and endothelial cells in the BBB and BRB lack fenestrations [5] and have profoundly decreased pinocytosis activity, while the choriocapillaris is fenestrated [6]. It is possible that the choriocapillaris endothelial cell fenestrations are regulated by vascular endothelial growth factor (VEGF), as intravitreal injection of the anti-VEGF antibody bevacizumab in cynomolgus monkeys significantly reduced these fenestrations, an effect that may be of clinical relevance in the treatment of macular edema [7]. Because the choriocapillaris is fenestrated, it is the RPE cells that form the outer BRB and regulate the environment of the outer retina. Like all epithelia and endothelia, the ability of RPE to regulate transepithelial transport depends upon two properties: apical TJs to resist diffusion through the paracellular spaces of the monolayer, and an asymmetric distribution of proteins to regulate vectorial transport across the monolayer [8]. During development, these properties form gradually. Initially, the TJs are leaky, and the RPE exhibits only partial polarity. As the neural retina and choriocapillaris develop, there are progressive changes in the composition of the apical junctional complexes, the expression of cell adhesion proteins, and the distribution of membrane and cytoskeletal proteins [8]. Another aspect of RPE function is the active transport of water out of the retina into the choriocapillaris. This flow of water out of the retina helps maintain retinal attachment.

In addition to controlling the influx of solutes, the BRB also actively transports potentially noxious compounds out of the retina in order to maintain the ideal microenvironment for its function. Lactate is actively transported from the RPE cells to the choroid [9, 10]. The P-glycoprotein is present in the BRB and actively pumping lipophilic toxins and drugs out of the endothelial or RPE cell, back to the bloodstream [5,...