Oxygen-induced changes in mitochondrial DNA and DNA repair enzymes in aging rat lens

• Published in: Mechanisms of ageing and development 2010-11, Vol.131 (11), p.666-673
• Main Authors: Zhang, Yi, OuYang, Shan, Zhang, Lan, Tang, XianLing, Song, Zhen, Liu, Ping
• Format: Article
• Language: English
• Subjects: Aging - drug effects; Aging - genetics; Aging - pathology; Animals; Base excision repair; Biological and medical sciences; Cataract - chemically induced; Cataract - enzymology; Cataract - pathology; Deoxyguanosine - analogs & derivatives; Deoxyguanosine - analysis; Deoxyguanosine - metabolism; Development. Metamorphosis. Moult. Ageing; DNA damage; DNA Glycosylases - metabolism; DNA Repair; DNA Repair Enzymes - metabolism; DNA, Mitochondrial - drug effects; DNA-Directed DNA Polymerase - metabolism; Enzyme-Linked Immunosorbent Assay; Fundamental and applied biological sciences. Psychology; Gene Expression - drug effects; Lens, Crystalline - drug effects; Lens, Crystalline - enzymology; Lens, Crystalline - pathology; Male; Oxidative stress; Oxidative Stress - genetics; Oxygen; Oxygen - toxicity; Polymerase Chain Reaction; Rats; Rats, Wistar; Reactive oxygen species; Vertebrates: anatomy and physiology, studies on body, several organs or systems
• ISSN: 0047-6374; 1872-6216
• DOI: 10.1016/j.mad.2010.09.003

▶ Our study is the first to examine young and aged rats’ lens for mtDNA and nDNA damage with QPCR assay and examined mtBER enzymes under hypoxic (11% O 2), normoxic (21% O 2) and hyperoxic (60% O 2) conditions. ▶ We found that only 14 month old rats group exposed to hyperoxic condition for 3 weeks developed nuclear cataract. There was a significant reduction of mtDNA amplification and a significantly increased 8-OHdG level in DNA at 14 month old rats in the hyperoxic group. There was significantly increased expression of OGG1, APE1, and Polγ both in 2 month and 14 month old rats exposed to hyperoxic condition, and there was morphological abnormalities of mitochondria of fiber cells in rats exposed to hyperoxic condition. ▶ We use a new dsDNA reagent- EvaGreen. It is stable both under PCR condition and during routine storage and handling. EvaGreen had a good binding affinity for dsDNA and low or no affinity for ssDNA. Therefore, we replaced PicoGreen and SYBR Green with EvaGreen to perform QPCR and RT-PCR in this work. ▶ The addition of a hyperoxic group was a useful “control” group that is often lacking in studies of this type and another positive was that intact animals, rather than isolated lenses were used for the exposures. High levels of oxygen were found to affect mtDNA but not nDNA and the effect was age-related. The treatment of patients with hyperbaric oxygen (HBO), vitrectomy and loss of vitreous gel during aging is associated with a high risk of subsequent development of nuclear cataract. Many studies proved that oxidation is the key reason of nuclear cataract. Reactive oxygen species (ROS) are formed in mitochondria as a by-product of normal metabolism and as a consequence of exposure to environmental compounds. Therefore, mitochondrial DNA (mtDNA) is at particularly high risk of ROS-induced damage. Oxidative damage to mtDNA has been implicated as a causative factor in a wide variety of degenerative diseases and aging. However, the effect of mtDNA damage to the lens has not been studied. The goals of the study were to identify if there was increased mtDNA damage in lens when the eye were exposed to hyperoxic or hypoxic conditions and also to evaluate the changes in gene expression of mtDNA base excision repair (mtBER) enzymes. Our data have shown that the damage of mtDNA, the expression of mtBER enzymes and the level of 8-OHdG in lens increased after inspired hyperoxia, which is likely associated with oxidative stress. However, there was no eff