Wang Yong, Li Fei, Zhang Guowei, Kang Lihua, Qin Bai, Guan Huaijin
Eye Institute, Affiliated Hospital of Nantong University , Nantong, Jiangsu Province , China and.
Curr Eye Res. 2015;40(8):815-21. doi: 10.3109/02713683.2014.957778. Epub 2014 Oct 13.
Oxidative stress and DNA damage contribute to the pathogenesis of age-related cataract (ARC). Most oxidative DNA lesions are repaired via the base excision repair (BER) proteins including 8-oxoguanine DNA glycosylase 1 (OGG1). This study examined DNA methylation of CpG islands upstream of OGG1 and their relation to the gene expression in lens cortex from ARC patients.
The clinical case-control study consisted of 15 cortical type of ARC patients and 15 age-matched non-ARC controls who received transparent lens extraction due to vitreoretinal diseases. OGG1 expression in lens cortex was analyzed by qRT-PCR and Western blot. The localization and the proportion of cells positive for OGG1 were determined by immunofluorescence. Bisulfite-sequencing PCR (BSP) was performed to evaluate the methylation status of CpG islands near OGG1 in DNA extracted from lens cortex. To test relationship between the methylation and the expression of the gene of interest, 5-Aza-2'-deoxycytidine (5-Aza-dC) was used to induce demethylation of cultured human lens epithelium B-3 (HLE B-3). To test the role of OGG1 in the repair of cellular damage, HLE B-3 was transfected with OGG1 vector, followed by ultraviolet radiation b (UVB) exposure to induce apoptosis.
The mRNA and protein levels of OGG1 were significantly reduced in the lens cortex of ARC. Immunofluorescence showed that the proportion of OGG1-positive cells decreased significantly in ARC cortex in comparison with the control. The CpG island in first exon of OGG1 displayed hypermethylation in the DNA extracted from the lens cortex of ARC. Treatment of HLEB-3 cells with 5-Aza-dC upregulated OGG1 expression. UVB-induced apoptosis was attenuated after transfection with OGG1.
A reduced OGG1 expression was correlated with hypermethylation of a CpG island of OGG1 in lens cortex of ARC. The role of epigenetic change in OGG1 gene in the susceptibility to oxidative stress induced cortical ARC is warranted to further study.
氧化应激和DNA损伤在年龄相关性白内障（ARC）的发病机制中起作用。大多数氧化性DNA损伤通过包括8-氧代鸟嘌呤DNA糖基化酶1（OGG1）在内的碱基切除修复（BER）蛋白进行修复。本研究检测了ARC患者晶状体皮质中OGG1上游CpG岛的DNA甲基化及其与基因表达的关系。
临床病例对照研究包括15例皮质型ARC患者和15例年龄匹配的非ARC对照，后者因玻璃体视网膜疾病接受透明晶状体摘除术。通过qRT-PCR和蛋白质印迹分析晶状体皮质中OGG1的表达。通过免疫荧光确定OGG1阳性细胞的定位和比例。采用亚硫酸氢盐测序PCR（BSP）评估从晶状体皮质提取的DNA中OGG1附近CpG岛的甲基化状态。为了测试甲基化与目的基因表达之间的关系，使用5-氮杂-2'-脱氧胞苷（5-Aza-dC）诱导培养的人晶状体上皮B-3（HLE B-3）去甲基化。为了测试OGG1在细胞损伤修复中的作用，用OGG1载体转染HLE B-3，然后暴露于紫外线b（UVB）以诱导细胞凋亡。
ARC晶状体皮质中OGG1的mRNA和蛋白水平显著降低。免疫荧光显示，与对照组相比，ARC皮质中OGG1阳性细胞的比例显著降低。从ARC晶状体皮质提取的DNA中，OGG1第一外显子中的CpG岛显示高甲基化。用5-Aza-dC处理HLEB-3细胞可上调OGG1表达。转染OGG1后，UVB诱导的细胞凋亡减弱。
ARC晶状体皮质中OGG1表达降低与OGG1的一个CpG岛高甲基化相关。OGG1基因表观遗传变化在氧化应激诱导的皮质型ARC易感性中的作用值得进一步研究。
