Age-related macular degeneration (AMD) is the leading cause of acquired blindness in the United States, yet patients with early disease have no preventative and treatment options. To address this shortcoming, this proposal focuses on early disease. A key early event in AMD is apoptosis of the retinal pigment epithelium (RPE). Apoptosis has an early stage, when dying cells are removed without inflammation, and a late stage, when apoptotic cells incite tissue-damaging inflammation. Complement plays a pivotal role because it can i) initiate apoptosis, and ii) label dying cells that need to be removed. Complement is tightly controlled by cell membrane (i.e. local) and fluid-phase (i.e. systemic) regulators. When this regulation is disturbed, disease can develop. The RPE cell membrane regulator CD46 is reduced by oxidized lipids, and genetic abnormalities to the fluid-phase regulator factor H (CFH) are linked with AMD risk. What remains unclear is how these regulators work together to protect the RPE, how impaired regulation influences RPE apoptosis, the origin of CFH, and how the CFH H402 variant causes AMD pathology. The objective of this proposal is to define how local RPE and systemic complement regulators interact to protect the RPE from oxidized lipids that trigger complement and apoptosis. The hypothesis to be tested is that both local (RPE) and systemic complement regulators protect the macula from uncontrolled complement activation initiated by oxidized lipids, and that impaired complement control from the RPE and systemic circulation contributes to both RPE apoptosis and impaired recognition of apoptotic debris. The following specific aims are proposed: 1. Determine the extent to which local regulators control complement activation and early stage RPE apoptosis after exposure to oxidized lipids;2. Define i) the origin of CFH that regulates complement activation in the RPE, and ii) the regulatory role of PTX3 on CFH within Bruch's membrane;3. Determine the extent to which CFH protects the RPE from excessive complement activation after its cell membrane complement regulators are impaired. Novel genetically modified mice targeting the RPE regulator CD46 (BEST1-cre-Crry-/- mice) and the CFH risk variant (transgenic human H402 and Y402 CFH mice) will be given different oxidized lipid challenges. The degree of complement activation, stage of apoptosis, associated inflammation, and recruitment of phagocytic cells will be quantified. The origin of CFH will be determined by topographically knocking out CFH using MX1-cre-CFH-/- mice in either the eye and/or liver. These contributions are significant because they will provide treatment targets that will maintain a protective complement response, limit apoptosis, and remove apoptotic debris without pathologic inflammation. The research is innovative because it will study the interplay between RPE and systemic regulators, considers the stage of RPE apoptosis, and utilizes novel genetic mouse models. Targeted therapy that maintains a protective complement response and limits apoptosis without unwanted inflammation is expected to result from this work.

National Institute of Health (NIH)
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Shen, Grace L
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Johns Hopkins University
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Cano, Marisol; Wang, Lei; Wan, Jun et al. (2014) Oxidative stress induces mitochondrial dysfunction and a protective unfolded protein response in RPE cells. Free Radic Biol Med 69:1-14
Chen, Chen; Cano, Marisol; Wang, Joshua J et al. (2014) Role of unfolded protein response dysregulation in oxidative injury of retinal pigment epithelial cells. Antioxid Redox Signal 20:2091-106
Wang, Lei; Cano, Marisol; Handa, James T (2014) p62 provides dual cytoprotection against oxidative stress in the retinal pigment epithelium. Biochim Biophys Acta 1843:1248-58
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Valapala, Mallika; Edwards, Malia; Hose, Stacey et al. (2014) Increased Lipocalin-2 in the retinal pigment epithelium of Cryba1 cKO mice is associated with a chronic inflammatory response. Aging Cell 13:1091-4
Fujihara, Masashi; Cano, Marisol; Handa, James T (2014) Mice that produce ApoB100 lipoproteins in the RPE do not develop drusen yet are still a valuable experimental system. Invest Ophthalmol Vis Sci 55:7285-95
Sachdeva, Mira M; Cano, Marisol; Handa, James T (2014) Nrf2 signaling is impaired in the aging RPE given an oxidative insult. Exp Eye Res 119:111-4

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