Abstract
Purpose: :
Age-related macular degeneration (AMD) is the leading cause of legal blindness in the elderly population. Debris (drusen) below the retinal pigment epithelium (RPE) is a risk factor for dry AMD and its progression to wet AMD, which is characterized by choroidal neovascularization (CNV). The underlying mechanism of how drusen might elicit CNV remains undefined. Cigarette smoking, oxidative damage to the RPE and inflammation might be involved in the pathophysiology of the disease. To better understand the cellular mechanism(s) linking oxidative stress and inflammation to AMD, we examined the expression of pro-inflammatory monocyte chemoattractant protein-1 (MCP-1), pro-angiogenic vascular endothelial growth factor (VEGF) and anti-angiogenic pigment epithelial derived factor (PEDF) in RPE from smoker patients with AMD. We also evaluated the effects of hydroquinone (HQ), a major pro-oxidant in cigarette smoke on MCP-1, VEGF and PEDF expression in vitro and in vivo.
Methods: :
Protein was extracted from RPE lysates from smoker patients with AMD and control patients. Confluent ARPE-19 cells were treated with either HQ 10µM for 24 hours for 5 consecutive days or HQ 50µM for 24 hours every 4 days for 3 weeks. Supernatants were collected and cells were harvested for RNA extraction. RNA and protein were extracted from RPE-choroid complexes dissected from C57BL6/J mice treated with HQ 0.8% in drinking water for 5 days and 3 weeks. MCP-1, VEGF and PEDF expression was examined by Western blot analysis, real-time PCR and ELISA.
Results: :
Low levels of MCP-1 protein were detected in RPE from AMD smoker patients relative to controls. Both MCP-1 mRNA and protein were downregulated in ARPE-19 cells and RPE/choroids from C57BL/6 mice after exposure to HQ-induced oxidative injury. VEGF protein expression was increased and PEDF protein expression was decreased in RPE from smoker patients with AMD compared with controls resulting in an increased VEGF/PEDF ratio. Treatment with HQ increased the VEGF-to-PEDF ratio in ARPE-19 cells and RPE/choroids from HQ-treated mice.
Conclusions: :
We propose that impaired RPE-derived MCP-1-mediated scavenging macrophages recruitment and phagocytosis might lead to incomplete clearance of proinflammatory debris and infiltration of proangiogenic macrophages which along with increased VEGF/PEDF ratio favoring angiogenesis might promote drusen accumulation and progression to CNV in smoker patients with AMD.
Keywords: retinal pigment epithelium • oxidation/oxidative or free radical damage • age-related macular degeneration