| Purpose To examine the promotor methylation and histone modification of WRN(Werner syndrome gene), a DNA repair gene, and their relationship with the gene expression in age-related cataract(ARC) lens.Methods A clinical case-control study was conducted in the department of ophthalmology from August 2013 to August 2014 in the Affiliated Hospital of Nantong University. The case group consisted of three subgroups: cortical cataract; nuclear cataract and posterior cataract. Each subgroup contained 39 eyes from 39 patients. And the control group included 39 age-matched patients who received transparent lens extraction due to vitreoretinal diseases. RT-PCR and Western blot were used to analyze the WRN expression. Methylation of Cp G islands within the WRN promoter were assessed by Bisulfite-sequencing PCR(BSP). Chromatin immunoprecipitation(Ch IP) was conducted to detect histone modification around the Cp G island region of WRN. Human lens epithelium cell line(HLEB-3) were treated with 5-Aza-2′-deoxycytidine(5-aza-d C) and trichostatin A(TSA) to observe the relationship between methylation and acetylation status of the cells and the expression of WRN.Results The m RNA and protein levels of WRN were significantly decreased in the ARC LECs comparing with the control LECs(F=162.82, P<0.01; F=68.72, P<0.01) while WRN were not detectable in lens cortex. The Cp G island of WRN promoter in the ARC LECs displayed hypermethylation comparing with the controls(F=104.01, P<0.01). The WRN promotor was almost fully methylated in the cortex of ARC and control lens. Acetylated H3 was lower while methylated H3-K9 was higher in ARC LECs than that of the controls(F=67.00, P<0.01; F=21.40, P<0.01). The expression of WRN in HLEB-3 increased after 5-aza-d C and TSA treatment(t=0.32, P<0.01; t=0.36, P<0.01).Conclusions A reduced WRN expression may be associated with the development of ARC. A hypermethylation in WRN promoter and altered histone modification in LECs might adjust its expression and contribute to the ARC mechanism. |
