Liu Chaoqi, Cao Lining, Yang Shuai, Xu Linxinyu, Liu Pei, Wang Fang, Xu Ding
Department of Ophthalmology, Shanghai Tenth People's Hospital Affiliated to Tongji University School of Medicine, Shanghai 200072, P.R. China.
Int J Mol Med. 2015 Jan;35(1):169-76. doi: 10.3892/ijmm.2014.1993. Epub 2014 Nov 10.
Drusen are considered a hallmark characteristic of age-related macular degeneration (AMD). In our previous study, we found that amyloid-β (Aβ) peptide, a component of drusen, induced the cells of the retinal pigment epithelium (RPE; RPE cells) to enter senescence; however, its effects in vivo remain unknown. Thus, the present study was carried out to explore the in vivo effects of Aβ peptide on RPE cell senescence and senescence-associated inflammation in C57BL/6 mice. C57BL/6 mice received a subretinal injection of Aβ(1-42) peptide; on day 7 post-injection, the mice were anesthetized and subjected to whole-body perfusion with 4% paraformaldehyde (PFA) in PBS and the whole eyes were then enucleated. Retinal function was assessed by electroretinography (ERG), and the morphological characteristics of the retina were examined by light and electron microscopy. Fundus autofluorescence (FAF) was examined by confocal scanning laser ophthalmoscopy (cSLO). The expression of p16INK4a, a marker of cellular senescence, was examined by immunofluorescence staining and western blot analysis. The RPE-choroid was analyzed for cytokine expression by RT-PCR. In Aβ(1-42)-injected mice, scotopic ERG responses declined. Degenerative alterations, including the disruption of the inner segment (IS)/outer segment (OS) junction and extensive vacuolation and thickness of Bruch's membrane (BrM) were observed under a a light microscope. The accumulation of vacuoles and the loss of basal infoldings in the RPE were identified using an electron microscope. FAF and p16INK4a expression increased in Aβ(1-42)-injected mice. In addition, Aβ(1-42) upregulated interleukin (IL)-6 and IL-8 gene expression in the RPE-choroid. In conclusion, our results confirm the effects of Aβ(1-42) peptide on RPE senescence in vivo. The Aβ-injected mice developed AMD-like ocular pathology. It is thus suggested that RPE cell senescence is a potential mechanistic link between inflammation and retinal degeneration.
玻璃膜疣被认为是年龄相关性黄斑变性（AMD）的一个标志性特征。在我们之前的研究中，我们发现玻璃膜疣的一种成分淀粉样β（Aβ）肽可诱导视网膜色素上皮（RPE；RPE细胞）细胞进入衰老状态；然而，其在体内的作用仍不清楚。因此，本研究旨在探讨Aβ肽对C57BL/6小鼠RPE细胞衰老和衰老相关炎症的体内影响。C57BL/6小鼠接受视网膜下注射Aβ（1-42）肽；注射后第7天，将小鼠麻醉并用PBS中的4%多聚甲醛（PFA）进行全身灌注，然后摘除整个眼球。通过视网膜电图（ERG）评估视网膜功能，并通过光学显微镜和电子显微镜检查视网膜的形态特征。通过共聚焦扫描激光眼科显微镜（cSLO）检查眼底自发荧光（FAF）。通过免疫荧光染色和蛋白质印迹分析检测细胞衰老标志物p16INK4a的表达。通过RT-PCR分析RPE-脉络膜中的细胞因子表达。在注射Aβ（1-42）的小鼠中，暗视ERG反应下降。在光学显微镜下观察到退行性改变，包括内段（IS）/外段（OS）连接的破坏以及布鲁赫膜（BrM）的广泛空泡化和增厚。使用电子显微镜鉴定RPE中液泡的积累和基底褶皱的丧失。在注射Aβ（1-42）的小鼠中，FAF和p16INK4a表达增加。此外，Aβ（1-42）上调了RPE-脉络膜中白细胞介素（IL）-6和IL-8基因的表达。总之，我们的结果证实了Aβ（1-42）肽在体内对RPE衰老的影响。注射Aβ的小鼠出现了类似AMD的眼部病理变化。因此，提示RPE细胞衰老可能是炎症与视网膜变性之间潜在的机制联系。
