Control of Chemokine Gradients by the Retinal Pigment Epithelium

The retinal pigment epithelium (RPE) is a monolayer of polarized pigmented cells that form the outer blood-retina barrier.1-3 In the subretinal space (SRS) of the intact eye, the RPE apical membrane is in close physical proximity to the distal ends of the photoreceptors, whereas the basolateral membrane is closely apposed to Bruch’s membrane and the choroidal blood supply. The RPE is remarkable in its ability to transport fluid and to secrete a variety of growth and neurotrophic factors and proinflammatory cytokines and chemokines in a polarized manner.4-9 In vitro, RPE cells have been shown to produce monocyte chemotactic protein (MCP)-1, interleukin (IL)-8, IL-6,10 regulated on activation of normal T-cell expressed and secreted (RANTES),11 interferon (IFN)-γ induced protein (IP) of 10 kDa (IP-10),12 and growth-related oncogene (GRO)-α,13 with or without stimulation by other proinflammatory cytokines. Together, the secretion of proinflammatory cytokines and chemokines14-16 and fluid transport17,18 could act in concert to change the course of inflammatory diseases. Cytokines are soluble proteinaceous substances (5-20 kDa) produced by a wide variety of cell types that secrete humoral factors and act to modulate the activities of other cells or tissues in normal or pathologic conditions. The functions of cytokines are pleiotropic and redundant. Chemokines are a subfamily of low-molecular-weight cytokines (8-10 kDa) identified by their ability to attract and activate leukocytes. CC and CXC are two structural variants that differ according to variations in a shared cysteine (C) motif. CC chemokines contain adjacent cysteines and are chemoattractants for lymphocytes, monocytes, eosinophils, and basophils, whereas CXC chemokines contain paired cysteines separated by a different amino acid and are chemoattractants for neutrophils. CXC chemokines can be categorized in terms of the “ELR” motif, which has been shown to associate the presence or absence of glutamic acid-leucine-arginine immediately preceding the first cysteine residue of the primary structure of these molecules with proangiogenic activity or its absence with antiangiogenic activity.19 In addition, some CC chemokines are highly angiogenic.20,21 Some chemokines are constitutively expressed in the eye, participate in the normal physiological activities of ocular cells and tissues,10 and undergo a dramatic increase in expression in diseases such as uveitis,15,22 diabetic proliferative retinopathy,23 and age-related macular degeneration (AMD).24 There is growing evidence that the immunologic properties of the RPE play a critical role in the accumulation of drusen and the progression to choroidal neovascularization (CNV) in AMD.25-32 The present study utilizes a previously described hfRPE primary culture model that has the morphology, polarization, and other physiologic properties of intact native tissue.7 A mixture of three proinflammatory cytokines—IL-1β, IFN-γ, and tumor necrosis factor (TNF)-α—was used to stimulate confluent monolayers of hfRPE. These proinflammatory cytokines have been found to be elevated in animal models of experimental autoimmune uveitis (EAU)14 and in patients with uveitis.33,34 Similarly, increased levels of TNF-α and IL-1β are also detected in the vitreous and blood of patients with proliferative diabetic retinopathy (PDR),35 and in those with AMD with CNV.36 Our data show that hfRPE expresses the receptors for all three cytokines and that activation of these receptors can dramatically affect the polarized secretion of cytokines/chemokines, alter transepithelial resistance, and increase net epithelial fluid absorption from the retinal to the choroidal side of the monolayer. These experiments suggest how the RPE in vivo can actively control the inflammatory environment in the SRS and choroid.