Xu Yi-Ting, Wang Ye, Chen Peng, Xu Hai-Feng
School of Medicine and Life Sciences of Shandong Academy of Medical Sciences, University of Jinan, Jinan 250022, Shandong Province, China.
Int J Ophthalmol. 2012;5(2):125-32. doi: 10.3980/j.issn.2222-3959.2012.02.02. Epub 2012 Apr 18.
Age-related macular degeneration (AMD) is a multifactorial disease and a prevalent cause of visual impairment in developed countries. Many studies suggest that age-related maculopathy susceptibility 2 (ARMS2) is a second major susceptibility gene for AMD. At present, there is no functional information on this gene. Therefore, the purpose of the present study was to detect the expression of ARMS2 in retinal pigment epithelium (RPE) cells and to investigate the effect of ARMS2 on the phagocytosis function of RPE cells.
Immunofluorescence and reverse transcriptase PCR were used to demonstrate the presence and location of ARMS2 in ARPE-19 (human retinal pigment epithelial cell line, ATCC, catalog No.CRL-2302) cells. siRNA was used to knock down ARMS2 mRNA, and the effects of the knockdown on the phagocytosis function of the ARPE-19 cells were evaluated via Fluorescence Activated Cell Sorting (FACS).
ARMS2 was present in ARPE-19 cells, localized in the cytosol of the perinuclear region. The expression of ARMS2 mRNA (messenger RNA) in ARPE-19 cells transfected with ARMS2-siRNA (small interfering RNA, 0.73±0.08) was decreased compared with normal cells (1.00±0.00) or with cells transfected with scrambled siRNA (0.95±0.13) (P<0.05). After incubation of RPE cells with a latex beads medium for 12, 18, or 24 hours, the fluorescence intensities were 38.04±1.02, 68.92±0.92, and 78.00±0.12 in the ARMS2-siRNA-transfected groups, respectively, and 77.98±5.43, 94.87±0.60, and 98.30±0.11 in the scrambled siRNA-transfected groups, respectively. The fluorescent intensities of the same time points in the two groups were compared using Student's t-test, and the p values were all less than 0.001 at the three different time points.
There is endogenous expression of ARMS2 in ARPE-19 cells. ARMS2 plays a role in the phagocytosis function of RPE cells, and this role may be one of the mechanisms that participates in the development of AMD.
年龄相关性黄斑变性（AMD）是一种多因素疾病，是发达国家视力损害的常见原因。许多研究表明，年龄相关性黄斑病变易感基因2（ARMS2）是AMD的第二个主要易感基因。目前，关于该基因尚无功能信息。因此，本研究的目的是检测ARMS2在视网膜色素上皮（RPE）细胞中的表达，并研究ARMS2对RPE细胞吞噬功能的影响。
采用免疫荧光和逆转录聚合酶链反应来证实ARMS2在ARPE-19（人视网膜色素上皮细胞系，美国典型培养物保藏中心，编号CRL-2302）细胞中的存在及定位。使用小干扰RNA（siRNA）敲低ARMS2信使核糖核酸（mRNA），并通过荧光激活细胞分选（FACS）评估敲低对ARPE-19细胞吞噬功能的影响。
ARMS2存在于ARPE-19细胞中，定位于核周区域的细胞质中。与正常细胞（1.00±0.00）或转染乱序siRNA的细胞（0.95±0.13）相比，转染ARMS2-siRNA（小干扰RNA）的ARPE-19细胞中ARMS2 mRNA的表达（0.73±0.08）降低（P<0.05）。用乳胶珠培养基孵育RPE细胞12、18或24小时后，ARMS2-siRNA转染组的荧光强度分别为38.04±1.02、68.92±0.92和78.00±0.12，乱序siRNA转染组分别为77.98±5.43、94.87±0.60和98.30±0.11。两组相同时间点的荧光强度采用学生t检验进行比较，在三个不同时间点的P值均小于0.001。
ARPE-19细胞中存在ARMS2的内源性表达。ARMS2在RPE细胞的吞噬功能中起作用，且该作用可能是参与AMD发生发展的机制之一。
