Ординатура / Офтальмология / Английские материалы / The Retina and its Disorders_Besharse, Bok_2011

protection of rat photoreceptors from a subsequent acute light damage independent of preconditioning. Additional work from Wen and Ash has similarly found that injection of LIF, CNTF, or BDNF protects mouse photoreceptors from a subsequent acute light damage. Of these, LIF and CNTF have tremendous therapeutic potential, given their ability to protect photoreceptors not only from acute light damage but also from inherited retinal degenerations.

Studies using intravitreal injections have demonstrated that multiple factors can induce protection, but did not demonstrate which factors were required for chronic light stress-induced protection. We have recently shown that LIF, CNTF, or CLC are the most likely candidates. This was demonstrated using an antagonist (LIF05) to the LIF receptor (LIFR), which blocks the activity of LIF, CNTF, and CLC. Intravitreal injection of LIF05 during light stress preconditioning greatly diminishes stress-induced protection from acute light damage. We also used conditional knock-out mice for the gp130 receptor and found that mice lacking gp130 expression in retinal photoreceptors also lose chronic light stress-induced protection. These recent studies demonstrate that the LIFR and gp130 expression in photoreceptors is essential for stress-induced protection and that the likely ligands are LIF, CLC, or CNTF, since these upregulated cytokines all utilize the LIFR and gp130. Our studies suggest that LIF may be the more important ligand since its expression is induced more than 100-fold following chronic light stress-induced preconditioning.

As described above, stress-induced protection was accompanied by upregulation of oxidant defense enzymes as well as photostasis. More recently, we have injected different doses of LIF into the vitreous of mice to determine whether LIF could induce both events. We found that at lower doses, LIF could induce protection from light damage without reducing photoreceptor sensitivity to light flashes. At higher concentrations, LIF not only induced protection, but also induced photostasis through decreased mRNA and protein expression of opsin, transducin (a and b subunits), and cyclic guanosine monophosphate (GMP) phosphodiesterase (PDE6A and PDE6B). Because of decreased expression of genes required for phototransduction, photoreceptors exhibited reduced efficiency in photon capture. These results demonstrate that LIF is upregulated by chronic bright-light stress and its induced expression is necessary for protection. LIF also has the ability to upregulate both photostasis and oxidant defense mechanisms. These studies clearly establish that in mice, LIF receptor, gp130, and perhaps LIF are essential players in chronic light stress-induced endogenous protection from acute light damage.

Many questions remain to be answered. In particular: Is LIF the essential factor? Which cells induce expression of LIF? Which signal transduction pathways are required for protection? What changes in gene expression are

required for protection? In a series of studies, we have shown that intravitreal injection of LIF results in activation of ERK1/2 and STAT3. The pattern of activation changes over time. Within the first 30 min of injection, both phosphorylated ERK1/2 and STAT3 were detected only in Mu¨ller’s cells and some ganglion cells. Within 4 h, however, all retinal cells were positive for STAT3, while ERK1/2 activation returned to basal levels. Detectable phosphorylated STAT3 remained up to 6 days following a single injection of LIF. Since phosphorylated STAT3 is a transcription factor, these results suggested that LIF could induce gene expression changes in all retinal cells including Mu¨ller’s cells and photoreceptors.

Concluding Remarks

Light-damage models have led to major advances in our molecular understanding of retinal degeneration through oxidative injury mechanisms and, in the future, lightdamage studies will be used to advance our understanding of injury induced by inflammation. Light-damage models have also led to the discovery of two independent mechanisms of endogenous neuroprotection, as well as to the discovery of promising new therapies to prevent or delay inherited retinal degenerations. Indeed, phase I clinical trials using encapsulated cells expressing CNTF have been completed with promising early results, and the study has since progressed into phase II trials. The protective effect of CNTF was initially described in the retina using the light-damage model. This alone should clearly establish the relevance for using light damage to identify potential therapeutic agents. By further defining the mechanisms of cell injury or protection, light-damage studies will likely lead to the development of new and more specific therapies.

See also: Injury and Repair: Retinal Remodeling; Phototransduction: Phototransduction in Rods; Phototransduction: Rhodopsin; Phototransduction: The Visual Cycle; Primary Photoreceptor Degenerations: Retinitis Pigmentosa; Primary Photoreceptor Degenerations: Terminology; Retinal Pigment Epithelium: Cytokine Modulation of Epithelial Physiology; Secondary Photoreceptor Degenerations: Age-Related Macular Degeneration.

Further Reading

Boulton, M., Rozanowska, M., and Rozanowski, B. (2001). Retinal photodamage. Journal of Photochemistry and Photobiology 64: 144–161.

Chen, L., Wu, W., Dentchev, T., et al. (2004). Light damage induced changes in mouse retinal gene expression. Experimental Eye Research 79: 239–247.

Marc, R. E., Jones, B. W., Watt, C. B., et al. (2008). Extreme retinal remodeling triggered by light damage: Implications for age related macular degeneration. Molecular Vision 14: 782–806.

Noell, W. K., Organisciak, D. T., Ando, H., Braniecki, M. A., and Durlin, C. (1987). Ascorbate and dietary protective mechanisms in retinal light damage of rats: Electrophysiological, histological and DNA measurements. Progress in Clinical and Biological Research 247: 469–483.

Penn, J. S. and Anderson, R. E. (1992). Effects of light history on the rat retina. In: Osborne, N. and Chader, G. (eds.) Progress in Retinal Research, vol. 11, pp. 75–98. New York: Pergamon Press.

Penn, J. S. and Thum, L. A. (1987). A comparison of the retinal effects of light damage and high illuminance light history. Progress in Clinical and Biological Research 247: 425–438.

Ranchon, I., Chen, S., Alvarez, K., and Anderson, R. E. (2001). Systemic administration of phenyl-N-tert-butylnitrone protects the retina from light damage. Investigative Ophthalmology and Visual Science 42:

1375–1379.

Reme, C. E., Grimm, C., Hafezi, F., Marti, A., and Wenzel, A. (1998). Apoptotic cell death in retinal degenerations. Progress in Retinal and Eye Research 17: 443–464.

Tanito, M., Agbaga, M. P., and Anderson, R. E. (2007). Upregulation of thioredoxin system via Nrf2-antioxidant responsive element pathway in adaptive-retinal neuroprotection in vivo and in vitro. Free Radical Biology and Medicine 42: 1838–1850.

Tanito, M., Kaidzu, S., Ohira, A., and Anderson, R. E. (2008). Topography of retinal damage in light-exposed albino rats.

Experimental Eye Research 87: 292–295. Wenzel, A., Grimm, C., Marti, A., et al. (2000). C-fos

controls the ‘‘private pathway’’ of light-induced apoptosis of retinal photoreceptors. Journal of Neuroscience 20: 81–88.

Wenzel, A., Grimm, C., Samardzija, M., and Reme, C. E. (2005). Molecular mechanisms of light-induced photoreceptor apoptosis and neuroprotection for retinal degeneration. Progress in Retinal and Eye Research 24: 275–306.

Wu, J., Seregard, S., and Algvere, P. V. (2006). Photochemical damage of the retina. Survey of Ophthalmology 51: 461–481.

Glossary

Complement – A complex molecular cascade that directly removes pathogens and activates the host immune system.

Cytokines – The secreted proteins that mediate cellular communication through a specific set of cell-surface receptors.

Extracellular matrix – The extracellular material that supports cell structure and intercellular communication.

Laser-induced injury – A reproducible model of tissue injury with the application of a high-energy laser photocoagulation to the fundus.

Neovascularization – The formation of new blood vessels.

Vascular endothelial growth factor – A potent cytokine that is critical for the process of neovascularization.

Introduction

The retina is a highly organized multicellular system designed to efficiently convert light into electrical signals which can then be transmitted to the brain for cortical integration and, ultimately, visual perception. In an acute setting of retinal injury, the cellular components of the neural retina, retinal pigment epithelium (RPE), and choroid get activated in order to contain the wound, destroy any invading pathogens, and initiate the repair process. As a result of the subsequent proinflammatory surge, blood vessels invade the wound bed to revascularize injured and hypoxic tissue and provide a new source of nutrients to the remodeling tissues. Typically, as in several other peripheral tissues in the human body, such as skin, this vascular growth allows for tissue healing and scar remodeling; however, in the delicate architecture of the retina, these abnormal vessels grow in an unregulated fashion and are susceptible to leakage and rupture. These traits are hardly suitable for the retina’s highly specialized function of vision and often lead to neovascularization (NV) as a cause of blindness in the context of retinal disease and injury. NV also contributes to other cellular responses in the injured retina, including gliosis and fibrosis. Through clinical observations and scientific investigation, critical insights into the

biology of NV in injury and repair in the retina, RPE, and choroid have modernized our current understanding of the cellular and molecular pathways that drive this pathological process.

Mechanisms of Injury

Retinal, RPE, and choroidal injury may be caused by a plethora of mechanical, cellular, physiologic, photochemical, and iatrogenic processes that are addressed in detail in other sections. Each of these instigating factors, either individually or in concert with one another, is capable of setting off the intricate cascade of factors leading to subretinal NV. A list of some of the common causes of injuryinduced NV is provided next.

1.Mechanical trauma. Blunt trauma to the eye disorganizes the retinal architecture to varying degrees with displacement of reactive glial cells and RPE to the vitreo-retinal interface. These cells can form a fibrotic scar at the surface of the internal-limiting membrane (ILM), in the case of epiretinal membrane, and even grow into the vitreous as in the case of proliferative vitreo-retinopathy. More ominous is the presence of a choroidal rupture, a finding that is very likely to induce neovascular invasion of the retina through fractures in Bruch’s membrane. Over time, these lesions eventually involute but do leave behind a residual fibrotic scar that significantly decreases visual acuity in the area.

2.Age-related macular degeneration. One of the most common forms of retinal injury is due to a combination of inflammation, oxidative stress, and photochemical damage secondary to age-related macular degeneration (AMD), a disease which accounts for the epidemic loss of vision in people over 60 years of age in the developed world. Between 10% and 20% of people with AMD will progress to the neovascular or wet form, which is responsible for about 90% of vision loss in patients with AMD. Recent evidence suggests that the gradual accumulation of inflammatory debris in drusen beneath the RPE and within the choroid ultimately leads to the production of proangiogenic cytokines and retinal invasion by abnormal and leaky vasculature. Some of these inflammatory components are comprised of photochemically altered proteins that are linked to intermediate or terminal products of the vitamin-A cycle or lipid-breakdown products released

by dying photoreceptors and RPE. Neovascular AMD occurs either below the RPE in its occult form or invades the neural retinal in its classic form. Choroidal NV (CNV) can wreak havoc through leakage into the intraretinal or vitreous spaces which acutely decreases visual acuity (Figure 1). Overtime, these NV membranes remodel into fibrotic scars that may still harbor abnormal vasculature and become sites of active NV recurrence.

3.Diabetic retinopathy. Diabetic retinopathy (DR) remains the primary cause of vision loss in patients aged less than 65 years. Hyperglycemia induces multiple microvascular abnormalities in the retina, eventually leading to retinal NV.

4.Vaso-occlusive disease. The retina receives dual blood supply with the central retinal artery feeding the inner layers and the choroidal vasculature feeding the outer layers, including the highly metabolic photoreceptor segments. Numerous pathologic processes can choke the retinal circulation, including atherosclerosis, vasculitis, thrombosis, and emboli. Retinal venous outflow is through the central retinal vein which is also at risk for collapse, stasis, or thrombosis. Choroidal venous flow exits through the vortex veins where thrombus may also form, albeit in a less-dramatic presentation than the central retinal vein occlusion. Another vascular disease that can trigger massive retinal ischemic injury is giantcell arteritis, an inflammatory disorder that can occlude the central retinal artery. Ischemic damage to the retina is irreversible approximately 90–100 min after initial insult and can result in a widespread neovascular response in the retina, iris, and angle. These sequelae are the leading causes of enucleation after such catastrophic vascular events in the eye.

5.Choroidal disease. Histoplasma capsulatum is a fungal organism endemic to the Ohio and Mississippi river

valleys that infects the choroidal tissue and causes localized areas of subretinal NV, which eventually scar over with RPE and glial cells. The organism is rarely isolated; however, patients often show a positive skin-allergy test thus earning this condition the title of presumed ocular histoplasmosis (POHS). Choroidopathies are a group of rare disorders that affect the tissues around the RPE due to either immune dysregulation or infectious disease, most often associated with nematode invasion. Although it is difficult to discern among the different forms of choroidopathy, several of them may lead to the formation of CNV. Multifocal choroiditis (MFC), multiple evanescent white-dot syndrome (MEWDS), bird-shot choroidopathy, serpiginous choroidopathy, and diffuse unilateral subacute neuroretinitis (DUSN) have all been associated with NV, hypothesized to be secondary to inflammation or immune dysregulation.

6.Cryogenic injury. Cryotherapy is a widely used treatment modality in vitreoretinal surgery to seal peripheral retinal tears in order to prevent rhegmatogenous retinal detachments. It is also used in combination with scleral buckling to destroy large areas of ischemic retina in order to prevent further NV. In some instances, cryotherapy itself can lead to subretinal NV.

7.Laser-induced injury. The use of laser photocoagulation for targeted retinal injury in the treatment of extrafoveal NVand proliferative DR has been widely used with success for several decades. In the treatment of CNV, focal laser injury is able to inhibit growth and decrease leakage from abnormal microvasculature. With panretinal photocoagulation, laser is used to ablate large swaths of the peripheral retina in DR that would otherwise be ischemic, producing proangiogenic factors. It is through the destruction of this pathologic tissue, that NV is spared in the precious visual real estate in the macula. Laser-injured tissues will commonly

remodel into small areas of RPE hyperpigmentation with fibrosis and only go on to develop CNV when high-energy laser photocoagulation is able to break through Bruch’s membrane. It is critical to note that laser-induced injury which fractures Bruch’s membrane serves as a very well-described animal model of CNV in many different laboratory animals, which is addressed in the following section.

Animal Models of NV after

Laser-Induced Injury

In order to study the natural history of injury-induced NV in vivo, several experimental models have been developed to mimic the disease process which occurs in humans. Over 25 years ago, Stephen Ryan described a laser-induced injury model of subretinal NV in rhesus monkeys. The same technology has since been used to develop similar models in a variety of research animals, including the mouse. In this model, laser photocoagulation is used to fracture Bruch’s membrane, which results in the formation of CNV (Figure 2). The laser-induced model captures many of the important features of the human condition including: migration of choroidal endothelial cells into the subretinal space via defects in Bruch’s membrane (Figure 3), accumulation of subretinal fluid,

congregation of leukocytes adjacent to neovascular tufts, gliosis (Figure 4), leakage of fluorescein from immature new vessels into the subretinal space, and increased expression of angiogenic growth factors and their receptors in cells and ocular tissues. The cellular and molecular details of retinal injury and repair are discussed below.

Acute Responses to Retinal Injury

Blood–Retina Barrier Breakdown

In a normal uninjured retina, the blood–retina barrier (BRB), comprised of RPE and retinal endothelial cell tight junctions, serves to prevent the influx of circulating proinflammatory cells and proteins where their presence and actions would compromise vision; however, in animal models of high-energy laser injury, there is an immediate disruption in the neural retinal, RPE tight junctions, and Bruch’s membrane, leading to BRB breakdown.

Acute Release of Cytokines

With laser injury, thousands of retinal and RPE cells suffer instantaneous thermal damage or death and disperse their intracellular contents into the interstitium. Many proand antiangiogenic cytokines which are harbored in cytoplasmic granules are immediately released

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inducing the influx of circulating leukocytes into the retinal tissues (Figure 5). There is a paucity of evidence on which resident cells are the sources of these cytokines. RPE cells constitutively produce a wide array of cytokines that are usually secreted in a polarized fashion either toward choriocapillaris or the photoreceptor layer to maintain their relative functions. For example, pigment-epithelium- derived factor (PEDF), a potent antiangiogenic mediator, is secreted from the apical RPE surface toward the photoreceptor layer, while vascular endothelial growth factor-A (VEGF-A), a predominant proangiogenic cytokine, is secreted from the basal RPE toward the choriocapillaris. With injury, this directionality is lost allowing for the disinhibition of angiogenesis in normally avascular tissue planes.

VEGF-A belongs to the highly conserved plateletderived growth factor (PDGF) family along with several other related proteins that constitute the VEGF subfamily, including placental growth factor (PlGF), VEGF-B, VEGF-C, and VEGF-D. For nomenclature purposes, the original VEGF protein was designated VEGF-A. Multiple isoforms of VEGF-A exist due to alternative splicing of the Vegfa gene; however, the 165-amino-acid variant in humans (VEGF-A165) is the most potent and prevalent among the major isoforms expressed during pathologic NV. Many other important components within the VEGF family contribute to injury-related NV. Several genes important in VEGF regulation and activation, such as the coreceptors neuropilin-1 and-2 as well as PlGF, are

acutely upregulated in the setting of laser injury and in some forms of human subretinal NV.

An acute elevation of VEGF-A released from various resident cells types, including RPE, microglia, Mu¨ller cells, and retinal astrocytes, transiently induces vascular permeability and upregulates endothelial cell expression of intercellular adhesion molecule (ICAM)-1. In response, there is a marked leukocytosis of injured tissues within 24 h. Resident cells also spew monocyte-chemoattractant protein (MCP)-1 into the wound matrix which acts as a homing device for circulating monocytes expressing its cognate receptor (CCR2). Interleukin-1b (IL-1b), tumor necrosis factor-a (TNF-a), and interleukin-6 (IL-6), all potent cytokines that sequester a wide range of inflammatory cells and induce their proliferation, are increased within 24 h of injury. IL-6 is instrumental in promoting continued expression of VEGF-A, ICAM-1, and MCP-1, which regulate the specific sequence of leukocyte chemotaxis that occurs over the initial 72 h after injury.

Injury-Induced Complement Activation

Complement factors are also a major participant in the robust and rapid retinal tissue response to injury. The complement pathways are comprised of multiple, complex, domino-like cascades engineered to generate massive proinflammatory induction through cytokine expression and leukocyte invasion in addition to promoting the selfassembly of molecular components that are capable of directly lysing bacteria and virus-infected cells. The two main active byproducts of complement activation, C3a and C5a, are predominantly responsible for the anaphylactic response. After laser injury, C3a and C5a levels are selectively and swiftly increased in the RPE and choroid within 4 h after photocoagulation with peak concentrations at 12 h. With blockade of C3a and C5a, as in mice that are deficient in C3a or C5a receptors, VEGF-A production is downregulated which may also inhibit leukocyte trafficking. Functional blockade of complement activation may be a valuable preventative therapy to arrest NV progression in patients with retinal disease and injury.

Other Angiogenic Mediators in the Retinal Response to Injury

Several other molecular factors have been implicated in the development of injury-induced NV through descriptive expression analyses in human specimens, including transforming growth factor-beta 1(TGF-b1), PDGF, fibroblast growth factor-2 (FGF-2), insulin-like growth factor-1 (IGF-1), and estrogen. Hepatocyte growth factor (HGF) is produced by RPE, upregulated as early as 6 h after laser injury, and hypothesized to mediate RPE proliferation during tissue repair.

Endogenous antiangiogenic molecules have also been identified such as PEDF and thrombospondin-1 that are decreased in CNV lesions and are capable of regulating NV responses in animal studies. These factors provide yet another level of control in this increasingly complex system of vascular regulation.

Modulation of the Extracellular Matrix

The biologic activities of VEGF-A and other cytokines, which acutely increased after injury, are influenced by the myriad of interactions between cells and the extracellular matrix (ECM). ECM proteins transmit cell–ECM communication and modulate tissue-remodeling events including NV. CNV membranes contain an abundance of ECM proteins similar to those found in granulation tissue during epithelial wound healing. Fibrin, fibronectin, and integrin receptors, all of which are important in ECM-guided angiogenesis, are present in surgically excised CNV membranes. The pathogenesis of injury-induced NV requires invasion of pre-existing ECM and the formation of new vascular conduits (Figure 6). Matrix metalloproteinases (MMP-2 and 9), which are capable of ECM dissolution, are upregulated in the acute-phase response. Tissue inhibitors of MMPs (TIMPs), which prevent excessive degradation of ECM by MMPs, are also expressed in CNV membranes thus offering a novel therapeutic target. Another proteinase mechanism, known to be involved in CNV progression, is the urokinase plasminogen activator pathway which is also being evaluated as an alternative treatment modality for pathologic NV.

More recently characterized ECM proteins may also be involved in VEGF-A signaling and NV response after injury. The matricellular protein SPARC (secreted protein, acidic, and rich in cysteine) is involved in tissue remodeling, cellular migration, and angiogenesis through its interaction with VEGF-A. After injury, SPARC expression is acutely decreased thus allowing VEGF-A to activate VEGFR-2 inducing endothelial cell proliferation and migration. In a paradoxical molecular mechanism, intravitreous administration of VEGF-A is able to suppress CNV when delivered after laser injury, whereas VEGF-A treatment prior to laser injury expectedly augments proangiogenic tissue response. These data may help improve our understanding of VEGF-A duality in NV induction and enhance the timing of administration of our targeted therapeutics to suppress this unwanted growth.

Cellular Response in Injury-Induced NV

Similar to epithelial wound healing, retinal injury is succeeded by an orchestrated arrival of various inflammatory cell types, a process that is determined by the acute response to injury discussed above. From other

wound-healing studies, it is known that inflammatory cell recruitment is directed by the expression of cytokines such as MCP-1, GROa (also known as CXCL1), and macrophage inflammatory protein-1a and-2 (MIP-1a, MIP-2). Many of these counterparts have been shown to be involved in NV progression in retinal injury.

Neutrophils

With breakdown of the BRB and acute VEGF-A-driven expression of ICAM-1 on the retinal and choroidal endothelia, there is an immediate and rampant neutrophil extravasation into the injured tissues which is maximal at 24 h. In addition to C5a, acute elevations in IL-1b and TNF-a may upregulate interleukin-8 (IL-8) which also acts as a potent neutrophil chemoattractant. These cells serve as a potent proinflammatory stimulus by secreting more VEGF-A along with numerous proangiogenic cytokines into the wound. Moreover, neutrophils are capable of engulfing invading pathogens and destroying them through the release of hypochlorous acid. Without neutrophil participation in wound healing, there is only a partial abrogation of the NV response to laser injury.

Macrophages

The next major influx of circulating proinflammatory leukocytes is the macrophage, which is driven by the enhanced expression of MCP-1 that peaks at 2 days after injury. These professional inflammatory cells home to sites of increased MCP-1 gradients through the expression of its cognate receptor, CCR2. This signaling axis is responsible for maximal macrophage infiltration into the choroid at 3 days post injury. Macrophages are able to respond to the proinflammatory milieu of vascular growth factors and secrete even more VEGF-A in order to aid revascularization

and tissue repair. After the arrival of macrophages, endothelial cells continue to increase their mitotic activity with peak proliferation at 5 days post injury. Approximately 1 week after injury, organized subretinal NV membranes are formed, which can be visualized in vivo with fluorescein angiography. Without macrophage influx, the ability of laser-injured areas to fully develop CNV is completely eliminated signifying the critical importance of this infiltrating cell in injury-induced NV.

Progenitor/Stem Cells

Pluripotent stem/progenitor cells are also recruited to sites of NV during injury and repair and incorporate into pathologic vasculature. Animal studies have now demonstrated that interfering with stem/progenitor cell homing to the sites of CNV through the modulation of the hypoxia responsive cytokine stromal-derived factor (SDF)-1a and its receptor, CXCR4, inhibits neovascular growth, a finding that offers another potential treatment for the human condition.

Other Infiltrating Cell Types

Although mast cells and eosinophils are able to generate significant proinflammatory stimuli, their contribution to injury-induced NV in the retina is believed to be negligible. B- and T-lymphocytes as well as natural killer cells are also able to infiltrate retina injuries, yet their particular effects and contribution to NVare still unclear but likely to be negligible.

Resident Tissue Cells

Among the resident cells of the retinal, RPE, and choroidal tissues that are involved in injury-induced NV, the

effects of RPE cells, microglia, retinal astrocytes, and Mu¨ller cells are the most evident.

Microglia

Microglial cells are tissue macrophages dispersed throughout the normal retina, choroid, and central nervous system. Their primary functions are to respond to the local invasion of pathogens through activation of the innate immune system, promotion of inflammation, and phagocytosis of bacteria and virus-laden cells. Microglia express a unique set of surface markers, as well as the chemokine receptor CX3CR1 which binds to fractalkine (or CX3CL1), a cytokine that is secreted preferentially by inflamed retinal and endothelial cells. CX3CR1 is present on a number of differentiated cell types derived from myeloid progenitor cells, such as dendritic cells, infiltrating macrophages, neutrophils, and some endothelial cells, thus providing another signaling axis for the infiltration of professional inflammatory cells after laser injury.

As part of their immune-surveillance capabilities, microglia express a series of innate immune receptors, called toll-like receptors (TLRs), that recognize pathogenassociated molecular patterns and respond by alarming other immune system components, mediating cellular infiltration, and selectively inducing apoptosis to prevent infectious spread. Several TLRs, including TLR 2–9, all of which are expressed on microglia, are likely to be involved in modulating angiogenesis in the setting of injury, thus creating a significant link between the innate immune response and NV. This immunovascular phenomenon is an area of great interest at this time given the strong evidence that several neovascular-related diseases, including AMD, may be driven by immune-system activation.

VEGF-A, once thought to be a pure vascular mediator, has been discovered to impart significant cellular effects on neural cells including microglia. Similar to bloodderived macrophages, microglia express VEGF receptor-1 (VEGFR-1) enabling their migration toward areas of VEGF-A production. With injury, microglia switch from their quiescent state to become activated, demonstrated by increased cytokine secretion, immune receptor expression, and proliferation.

Retinal astrocytes

Activated microglia may also initiate the reparative process by inducing the chemotaxis and proliferation of retinal astrocytes through TGF-b1, IL-1b, and TNF-a. Unlike microglia, astrocytes may be far more robust in the promotion of immune-mediated inflammation and contribution to glial scarification after retinal injury.

Mu¨ller cells

Mu¨ller cells are specialized glia that provide structural and trophic support, among numerous other functions, in the retina. In the setting of injury, Mu¨ller cells are

able to proliferate and dedifferentiate into progenitorlike cells thus providing a potential source for retinal regeneration. Whereas some vertebrates (avian, amphibian, and fish) have been found to demonstrate such reparative potential, there is still controversial evidence on whether Mu¨ller cells of the mammalian retina are capable of this phenomenon.

Cellular Responses to VEGF-A Receptor Binding

VEGF-A is a multifaceted cytokine that influences multiple endothelial cell pathways promoting growth, survival, and vascular permeability. Its signal transduction is mediated primarily through two receptor tyrosine kinases, VEGFR-1 and VEGFR-2; however, the latter is implicated as the principal proangiogenic transducer. Both VEGFR-1 and R-2 are expressed in normal human eyes and surgically excised CNV membranes. There is a continuing controversy on the role of VEGFR-1 as a decoy receptor designed to sequester free VEGF-A and prevent excessive VEGFR-2 activation. In multiple models of injury-induced NV, VEGFR-1 has been shown to function bilaterally as both a proand antivascular mediator. Importantly, it does not appear that VEGFR-1 does this by simply serving as a decoy, as VEGFR-1 can repress VEGFR-2 mediated endothelial cell proliferation through active signaling after retinal injury. Thus, VEGF-A may possess the ability to function dichotomously in both proand antiangiogenic capacities.

Conclusion

NV in the context of injury and repair is designed to aid the healing process, but, in the fragile and transparent cellular layers of the retina, it wreaks havoc on neural function and visual acuity. Several decades of dedicated molecular science and centuries of clinical observations have yielded an exhaustive foundation of knowledge that has significantly improved our understanding of injuryinduced blood vessel growth. These mechanisms are currently being elucidated with such resolution and speed that a detailed molecular map of this disease process may be within our reach in the near future. More importantly, the work has been a tremendous benefit to society, as several targeted therapeutics are now available to the millions of people around the world suffering from neovascular diseases. It is our hope that scientific investigation of the unique biologic responses to retinal injury will continue to reveal critical facets in vasomolecular medicine both in the eye and elsewhere that will aid in the design of advanced therapeutics.

See also: Breakdown of the Blood–Retinal Barrier; Breakdown of the RPE Blood–Retinal Barrier; Central Retinal Vein

Occlusion; Immunobiology of Age-Related Macular Degeneration; Injury and Repair: Retinal Remodeling; Pathological Retinal Angiogenesis; Primary Photoreceptor Degenerations: Terminology; Retinal Pigment Epithelium: Cytokine Modulation of Epithelial Physiology; Retinal Vasculopathies: Diabetic Retinopathy; Secondary Photoreceptor Degenerations: Age-Related Macular Degeneration; Secondary Photoreceptor Degenerations.

Further Reading

Ambati, J., Ambati, B. K., Yoo, S. H., et al. (2003). Age-related macular degeneration: Etiology, pathogenesis, and therapeutic strategies.

Survey Ophthalmology 48(3): 257–293.

Carmeliet, P. (2005). Angiogenesis in life, disease and medicine. Nature 438(7070): 932–936.

Eter, N., Engel, D. R., Meyer, L., et al. (2008). In vivo visualization of dendritic cells, macrophages, and microglial cells responding to laser-induced damage in the fundus of the eye. Investigative Ophthalmology and Visual Science 49(8): 3649–4358.

Fisher, S. K., Lewis, G. P., Linberg, K. A., et al. (2005). Cellular remodeling in mammalian retina: Results from studies of experimental retinal detachment. Progress in Retinal and Eye Research 24(3): 395–431.

Friedlander, M. (2007). Fibrosis and diseases of the eye. Journal of Clinical Investigation 117(3): 576–586.

Holtkamp, G. M., Kijlstra, A., Peek, R., et al. (2001). Retinal pigment epithelium–immune system interactions: Cytokine production and cytokine-induced changes. Progress in Retinal and Eye Research

20(1): 29–48.

Nozaki, M., Raisler, B. J., Sakurai, E., et al. (2006). Drusen complement components C3a and C5a promote choroidal neovascularization.

Proceedings of the National Academy of Sciences of the United States of America 103(7): 2328–2333.

Nozaki, M., Sakurai, E., Raisler, B. J., et al. (2006). Loss of sparc-mediated Vegfr-1 suppression after injury reveals a novel antiangiogenic activity of Vegf-A. Journal of Clinical Investigation 116(2): 422–429.

Osborne, N. N., Casson, R. J., Wood, J. P., et al. (2004). Retinal ischemia: Mechanisms of damage and potential therapeutic strategies. Progress in Retinal and Eye Research 23(1): 91–147.

Pournaras, C. J., Rungger-Brandle, E., Riva, C. E., et al. (2008). Regulation of retinal blood flow in health and disease. Progress in Retinal and Eye Research 27(3): 284–330.

Rattner, A. and Nathans, J. (2005). The genomic response to retinal disease and injury: Evidence for endothelin signaling from photoreceptors to glia. Journal of Neuroscience 25(18): 4540–4549.

Ryan, S. J. (1982). Subretinal neovascularization. Natural history of an experimental model. Archives of Ophthalmology 100(11): 1804–1809.

Ryan, S. J. (2006). Retina. Philadelphia, PA: Elsevier. Vazquez-Chona, F. R., Khan, A. N., Chan, C. K., et al. (2005).

Genetic networks controlling retinal injury. Molecular Visision 11: 958–970.

Wu, J., Seregard, S., and Algvere, P. V. (2006). Photochemical damage of the retina. Survey Ophthalmology 51(5): 461–481.