photoreceptor protection, further suggesting a cell-autonomous mechanism of photoreceptor protection through STAT3 activation (Ueki et al. 2008). In this current study, we observed loss of both STAT3 and ERK1/2 signaling in at least 50% of Müller cells in the gp130f/f/VMD2-cre+ retina (Fig. 75.2; ERK1/2 data not shown). However, this loss of signaling did not affect the gp130-mediated photoreceptor protection against light-induced cell death (Figs. 75.2 and 75.3). These data support the hypothesis that the protection is mediated by cell autonomous mechanism in photoreceptors.

In summary, impaired activation of gp130 in Müller cells does not diminish gp130-mediated photoreceptor protection from light damage. A caveat to this study is that our conditional gp130 KO is not complete and some Müller cells still express gp130. Data from these VMD2-cre mice cannot be used to conclusively rule out the possibility that remaining activated Müller cells compensate for the loss of gp130 in gp130-negative Müller cells. However, in another study to be published separately, knockout of gp130 in photoreceptors does impair gp130-mediated protection. In total, these studies provide clear evidence that gp130 activation in photoreceptors is the direct mechanism of LIF-mediated protection.

Acknowledgments This work was supported by funding from National Institute of Health (R01 EY016459, P20 RR017703, P30 EY012190), The Foundation Fighting Blindness, and an Unrestricted grant from Research to Prevent Blindness.

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intracellular signaling pathways in Muller cells and other cells of the inner retina, but not photoreceptors. Invest Ophthalmol Vis Sci 41:927–936

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Chapter 76

Neuroprotectin D1 Modulates the Induction

of Pro-Inflammatory Signaling and Promotes

Retinal Pigment Epithelial Cell Survival During

Oxidative Stress

Jorgelina M. Calandria and Nicolas G. Bazan

Abstract Retinal pigment epithelial (RPE) cells are the most restrictive layer of the three components of the outer Blood-Retina Barrier, preventing the passage of biomolecules in relation to size and charge and thus preserving a controlled environment for the photoreceptors. The retinal pigment epithelium is a tight structure that, when disrupted as a cause or consequence of pathological conditions, deeply affects the neural retina. Since adult human RPE cells are not replicative cells, their preservation is of major interest for the biomedical field due to their loss in many retino-degenerative pathologies. There are several triggers that elicit reactive oxygen species (ROS) formation in normal and pathological circumstances. When the production of these species overwhelms the scavenging and detoxifying systems, their activity results in programmed cell death. Docosahexaenoic acid (DHA) is an essential lipid that is conspicuously accumulated in photoreceptors and RPE cells in the retina. DHA and its oxygenation product, neuroprotectin D1 (NPD1), are major players in the protection of these cells and the retina. NPD1 promotes the synthesis of anti-apoptotic proteins of certain members of the Bcl-2 family and blocks the expression of pro-inflammatory proteins like cyclooxygenase-2.

76.1The Importance of RPE Cell Function and Integrity for Photoreceptor Survival

The outer Blood-Retinal Barrier (oBRB) mediates the exchange of small molecules and solutes and other metabolites from the blood stream to the neural-retina (Strauss 2005). Specifically, the retinal pigment epithelium is the most restrictive layer of the three components of the oBRB, preventing the passage of biomolecules regarding size and charge and thus preserving a controlled environment for the

N.G. Bazan (B)

Neuroscience Center of Excellence and Department of Ophthalmology, Louisiana State University Health Sciences Center, 2020 Gravier Street, Suite D, New Orleans, LA 70112, USA

e-mail: nbazan@lsuhsc.edu

Medicine and Biology 664, DOI 10.1007/978-1-4419-1399-9_76,C Springer Science+Business Media, LLC 2010

Fig. 76.1 RPE role in the preservation of the retinal structure and NPD1 signaling. Retinal pigment epithelial (RPE) cells interact with photoreceptors in the neural retina. (a) They are actively involved in the remodeling and permeability of blood vessels in the choriocapillaries, which in turn provides them with nutrients such as docosahexaenoic acid (DHA). (b) Within the retinal pigment epithelium, neuroprotectin D1 (NPD1) signaling is transmitted and proand anti-apoptotic cues are integrated to decide the cell fate. (c) RPE cells also contribute to daily maintenance by recycling oxidized and bleached pigments and membranes, and then returning DHA, other lipids, and pro-survival factors, such as NPD1

photoreceptors (see Fig. 76.1). The retinal pigment epithelium is a tight structure in which cells are communicated laterally through tight junctions (see Fig. 76.1B). These cells present an elaborate trans-cellular transport system and a high polarization that allows the two different surfaces to have different functions (Pournaras et al. 2008). The selective permeability of the oBRB depends on its integrity, and retinal pigment epithelial (RPE) cells are also involved in the preservation of this structure by interacting reciprocally in cell formation and maintenance. TRP2-FGF9 transgenic mice, in which embryonic RPE cells are forced to become neural retinal cells through the ectopic expression of FGF9, fail to form blood vessels in the choroid layers adjacent to those regions where RPE cells are absent. Instead, blood vessels are found in the vicinity of patch where RPE cells are present (Zhao and Overbeek 2001). The dependency between RPE cells and endothelial vascular cells continues during the adult life through the regulation of neovascularization.

Compelling evidence links RPE cells with the secretion of several angiogenicrelated factors. In particular, RPE cells from transgenic ApoE2 mice, which express human ApoE2 protein and whose eyes present several common features with those of age-related macular degeneration (AMD) patients, show reciprocal unbalanced expression of pigment epithelium-derived factor (PEDF) and vascular endothelial growth factor (VEGF), indicating that neovascularization may be increased (Lee et al. 2007). Furthermore, autocrine VEGF signaling in RPE cells stimulate VEGFrelated gene expression as well as PEDF modulation, which is a potent angiogenic inhibitor.

Apart from these, RPE cells are capable of producing a wide variety of growth factors (Tanihara et al. 1997), including ciliary neurotrophic factor (CNTF), platelet-derived growth factor (PDGF), insulin-like growth factor-1 (IGF-1) and transforming growth factor-beta (TGF-β) and other molecules such as different types of tissue inhibitor of matrix metalloproteases (TIMPs) (Strauss 2005).

In addition, the structural apico-basal polarization evidenced by deep basal folds and apical microvilli, which have functionally pronounced differences, are found on both sides of the epithelium (see Fig. 76.1). The long apical processes are specialized phagocytic surfaces that engulf shed photoreceptor outer segments in a daily rhythmic cycle to scavenge bleached photopigments, proteins and lipids. During this cycle, a large amount of docosahexaenoic acid (DHA) is incorporated into the RPE cells and is either transformed into neuroprotectin D1 (NPD1), which acts in a paracrine and autocrine manner, or is returned to the photoreceptor to serve as a substrate in the biogenesis of disc membranes (Bazan 2007). Concomitantly with the photoreceptor survival effect of outer segment phagocytosis, the retinal pigment epithelium benefits from this process, which provides resistance to oxidative stress (Mukherjee et al. 2007b). Furthermore, the dependence between RPE cells and photoreceptors is significant in Usher type 1B syndrome. The lack of myosin VIIa in this progressive disease affects the ability of RPE cells to phagocytize the photoreceptor outer segment, which resulted in retinal degeneration in a mouse model (Gibbs et al. 2003). Similar to what was proposed to happen in the Blood Brain Barrier in the neurovascular hypothesis of Alzheimer’s disease (AD) (Zlokovic 2005), defective clearance of certain molecules across the barrier may initiate a series of faulty maintenance functions that could lead to a retino-vascular inflammatory response, contributing to the development of AMD.

It was recently proposed that some reactive oxygen species (ROS) may have specific targets that affect certain signaling pathways, a property that makes these reactive species signaling molecules, as is the case of Nitric Oxide (NO) (D’Autreaux and Toledano 2007). Accordingly, redox systems are a part of the normal cell milieu, and thus are tightly regulated. If this were not the case, the disturbed equilibrium may lead to fatal consequences for the cell.

The context in which RPE cells are immersed is highly destructive due to the presence of disposal sub-products of retinal activity. Photochemical reactions take place in the retina under normal conditions to produce ROS by the absorption of photons in the presence of oxygen (Boulton et al. 2001). Moreover, unlike in other cell types in the retina, oxygen consumption and oxidative enzymatic