Department of Ophthalmology and Visual Science, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
Biochem Biophys Res Commun. 2012 Aug 10;424(4):704-9. doi: 10.1016/j.bbrc.2012.07.014. Epub 2012 Jul 14.
Subretinally-deposited amyloid β (Aβ) is a main contributor of developing age-related macular degeneration (AMD). However, the mechanism causing Aβ deposition in AMD eyes is unknown. Hypercholesterolemia is a significant risk for developing AMD. Thus, we investigated the effects of cholesterol on Aβ production in retinal pigment epithelial (RPE) cells in vitro and in the mouse retina in vivo. RPE cells isolated from senescent (12-month-old) C57BL/6 mice were treated with 10μg/ml cholesterol for 48h. Aβ amounts in culture supernatants were measured by ELISA. Activity and expression of enzymes and proteins that regulate Aβ production were examined by activity assay and real time PCR. The retina of mice fed cholesterol-enriched diet was examined by transmission electron microscopy. Cholesterol significantly increased Aβ production in cultured RPE cells. Activities of Aβ degradation enzyme; neprilysin (NEP) and anti-amyloidogenic secretase; α-secretase were significantly decreased in cell lysates of cholesterol-treated RPE cells compared to non-treated cells, but there was no change in the activities of β- or γ-secretase. mRNA levels of NEP and α-secretase (ADAM10 and ADAM17) were significantly lower in cholesterol-treated RPE cells than non-treated cells. Senescent (12-month-old) mice fed cholesterol-enriched chow developed subRPE deposits containing Aβ, whereas age-matched mice fed standard rodent chow diet did not. Activities and mRNA levels of NEP and α-secretase were significantly lower in native RPE cells freshly isolated from cholesterol-enriched chow fed mice compared to standard rodent chow fed mice. These findings suggest that cholesterol enhances subretinal Aβ accumulation by modulating the activities of enzymes degrading and processing Aβ in RPE cells in senescent subjects.
视网膜下沉积的淀粉样β（Aβ）是导致年龄相关性黄斑变性（AMD）的主要原因。然而，导致 AMD 眼中 Aβ沉积的机制尚不清楚。高胆固醇血症是发生 AMD 的重要危险因素。因此，我们研究了胆固醇对体外培养的视网膜色素上皮（RPE）细胞和体内小鼠视网膜中 Aβ产生的影响。从衰老（12 个月大）C57BL/6 小鼠中分离的 RPE 细胞用 10μg/ml 胆固醇处理 48h。通过 ELISA 测量培养上清液中的 Aβ 量。通过活性测定和实时 PCR 检查调节 Aβ产生的酶和蛋白质的活性和表达。用透射电子显微镜检查喂食富含胆固醇饮食的小鼠的视网膜。胆固醇显著增加了培养的 RPE 细胞中的 Aβ 产生。与未处理的细胞相比，胆固醇处理的 RPE 细胞的细胞裂解物中 Aβ 降解酶；neprilysin（NEP）和抗淀粉样分泌酶；α-分泌酶的活性显著降低，但β-或γ-分泌酶的活性没有变化。与未处理的细胞相比，胆固醇处理的 RPE 细胞中 NEP 和 α-分泌酶（ADAM10 和 ADAM17）的 mRNA 水平显着降低。喂食富含胆固醇的 Chow 的衰老（12 个月大）小鼠出现含有 Aβ 的 subRPE 沉积物，而喂食标准啮齿动物饲料的同龄对照小鼠则没有。从喂食富含胆固醇的 Chow 的小鼠中新鲜分离的天然 RPE 细胞中的 NEP 和 α-分泌酶的活性和 mRNA 水平明显低于喂食标准啮齿动物 Chow 的小鼠。这些发现表明，胆固醇通过调节衰老受试者 RPE 细胞中降解和处理 Aβ 的酶的活性来增强视网膜下 Aβ 的积累。
