Kresge Eye Institute, Wayne State University, Detroit, Michigan, USA.
Invest Ophthalmol Vis Sci. 2012 Jul 24;53(8):4881-8. doi: 10.1167/iovs.12-9732.
Diabetic retinopathy fails to halt after cessation of hyperglycemic insult, and a vicious cycle of mitochondria damage continues. The aim of our study was to investigate the effect of termination of hyperglycemia on retinal mtDNA replication, and elucidate the mechanism responsible for the continued mtDNA damage.
Polymerase gamma 1 (POLG1), the catalytic subunit of the mitochondrial DNA replication enzyme, and the damage to the displacement loop region of mtDNA (D-loop) were analyzed in the retina from streptozotocin-diabetic rats maintained in poor glycemic control (PC, glycated hemoglobin ∼11%) or in good glycemic control (GC, glycated hemoglobin ∼6%) for 6 months, or in PC for three months followed by GC for three months (Rev). To understand the mechanism DNA methylation status of POLG1 promoter was investigated by methylation-specific PCR. The key parameters were confirmed in the isolated retinal endothelial cells exposed to high glucose, followed by normal glucose.
POLG1 continued to be down-regulated, the D-loop region damaged, and the CpG islands at the regulatory region of POLG hyper-methylated even after three months of GC that had followed three months of PC (Rev group). Similar results were observed in the retinal endothelial cells exposed to normal glucose after being exposed to high glucose.
Continued hypermethylation of the CpG sites at the regulatory region of POLG affects its binding to the mtDNA, compromising the transcriptional activity. Modulation of DNA methylation using pharmaceutic or molecular means could help maintain mitochondria homeostasis, and prevent further progression of diabetic retinopathy.
高血糖损伤停止后，糖尿病性视网膜病变仍无法得到控制，线粒体损伤继续恶性循环。本研究旨在探讨终止高血糖对视网膜线粒体 DNA 复制的影响，并阐明导致持续的 mtDNA 损伤的机制。
分析链脲佐菌素诱导的糖尿病大鼠视网膜中聚合酶γ 1（POLG1），即线粒体 DNA 复制酶的催化亚基，以及线粒体 DNA（D 环）的位移环区域（D-loop）的损伤情况。这些大鼠分别维持在血糖控制不良（PC，糖化血红蛋白约 11%）或血糖控制良好（GC，糖化血红蛋白约 6%）6 个月，或血糖控制不良 3 个月后转为血糖控制良好 3 个月（Rev）。为了了解 POLG1 启动子的 DNA 甲基化状态，通过甲基化特异性 PCR 进行了分析。在高糖暴露后，对分离的视网膜内皮细胞进行了关键参数的确认，随后恢复为正常葡萄糖。
即使在 GC 治疗三个月（Rev 组）后，POLG1 仍持续下调，D 环区域受损，调控区的 CpG 岛高度甲基化。在正常葡萄糖暴露后，高糖暴露的视网膜内皮细胞也观察到了类似的结果。
调控区 POLG 的 CpG 位点的持续高甲基化影响其与 mtDNA 的结合，从而损害转录活性。使用药物或分子手段调节 DNA 甲基化可能有助于维持线粒体的稳态，防止糖尿病性视网膜病变的进一步进展。
