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Editors Selection IGR 10-4

Basic Research: Single-stranded DNA breaks

Comment by George Lambrou on:

22075 Comet assay analysis of single-stranded DNA breaks in circulating leukocytes of glaucoma patients, Mozaffarieh M; Schoetzau A; Sauter M et al., Molecular Vision, 2008; 14: 1584-1588

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The pathophysiology of glaucoma is certainly multi-faceted with many of these facets still hidden from us and occasional glimpses to them caught sometimes through unexpected ways. In this short paper, Mozaffarieh et al.(1365) report their findings from applying a standard toxicology procedure, the 'Comet Assay', to investigate the amount of DNA breaks in circulating lymphocytes of POAG patients. What is the 'Comet Assay'? It is a very simple technique for estimating the overall amount of genome damage (more specifically:of single-strand DNA breaks) present at a given time in a cell population. DNA being quite vulnerable to external insult, the cells have developed extremely efficient repair mechanisms which normally keep the amount of DNA damage at an amazingly low steady-state of 1.65 parts-per-million. In a nutshell, the technique operates like this: if isolated cells are stained, subjected to lysis and then to electrophoresis, the massive DNA molecules will barely budge, appearing as a solid nucleus or 'head' on the electrophoresis gel, whereas broken DNA fragments will migrate away from the nucleus, creating a comet-like 'tail' flowing away from it. Simple quantification algorithms permit to estimate the amount of (broken) DNA in the tail versus the (unbroken) DNA in the head. One such estimate is the 'tail moment'. The authors have evaluated tail moments from circulating lymphocytes of healthy volunteers and age-&-sex matched glaucoma patients with and without primary vascular dysregulation (PVD), the latter being defined as the combination of positive patient history and abnormal nailfold capillaroscopy. Twenty-eight subjects were enrolled, divided in four groups with six to eight patients each (POAG patients with and without PVD, healthy volunteers with and without PVD). Of these four groups, only glaucoma patients with PVD had significantly high rates of DNA breaks.

The role of oxidative stress in the pathogenesis of glaucoma is being increasingly contemplated by various authors, and this study is adding an indirect argument in its favor

This finding supports a number of possible explanations, hinting at potential pathophysiological mechanisms, with two of them being corroborated by independent evidence. The first is increased oxidative stress in glaucoma patients with PVD, resulting either from a higher rate of oxidative events (mild ischemia-reperfusion episodes may contribute to this) and/or an age-dependent decline of the capacity to cope with the free radicals generated during such events. The role of oxidative stress in the pathogenesis of glaucoma is being increasingly contemplated by various authors, and this study is adding an indirect argument in its favor. Alternatively (and possibly concomitantly) increased DNA breaks may reflect an impaired DNArepair capacity, a hypothesis supported by findings of modified gene expression patterns in the lymphocytes of glaucoma patients with PVD, using molecular biology methods. Although this study brings indirect evidence only of putative pathophysiological mechanisms, it is a fine example by the Basel group of how tools and techniques developed in distant areas of medicine may help shed light onto unexpected facets of ongoing controversies, like the pathogenesis of glaucoma.

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