Effects of hydrogen peroxide on wound healing in mice in relation to oxidative damage.

It has been established that low concentrations of hydrogen peroxide (H(2)O(2)) are produced in wounds and is required for optimal healing. Yet at the same time, there is evidence that excessive oxidative damage is correlated with poor-healing wounds. In this paper, we seek to determine whether topical application of H(2)O(2) can modulate wound healing and if its effects are related to oxidative damage. Using a C57BL/6 mice excision wound model, H(2)O(2) was found to enhance angiogenesis and wound closure at 10 mM but retarded wound closure at 166 mM. The delay in closure was also associated with decreased connective tissue formation, increased MMP-8 and persistent neutrophil infiltration. Wounding was found to increase oxidative lipid damage, as measured by F(2)-isoprostanes, and nitrative protein damage, as measured by 3-nitrotyrosine. However H(2)O(2) treatment did not significantly increase oxidative and nitrative damage even at concentrations that delay wound healing. Hence the detrimental effects of H(2)O(2) may not involve oxidative damage to the target molecules studied.

Mitochondrial changes in ageing Caenorhabditis elegans--what do we learn from superoxide dismutase knockouts?

One of the most popular damage accumulation theories of ageing is the mitochondrial free radical theory of ageing (mFRTA). The mFRTA proposes that ageing is due to the accumulation of unrepaired oxidative damage, in particular damage to mitochondrial DNA (mtDNA). Within the mFRTA, the "vicious cycle" theory further proposes that reactive oxygen species (ROS) promote mtDNA mutations, which then lead to a further increase in ROS production. Recently, data have been published on Caenorhabditis elegans mutants deficient in one or both forms of mitochondrial superoxide dismutase (SOD). Surprisingly, even double mutants, lacking both mitochondrial forms of SOD, show no reduction in lifespan. This has been interpreted as evidence against the mFRTA because it is assumed that these mutants suffer from significantly elevated oxidative damage to their mitochondria. Here, using a novel mtDNA damage assay in conjunction with related, well established damage and metabolic markers, we first investigate the age-dependent mitochondrial decline in a cohort of ageing wild-type nematodes, in particular testing the plausibility of the "vicious cycle" theory. We then apply the methods and insights gained from this investigation to a mutant strain for C. elegans that lacks both forms of mitochondrial SOD. While we show a clear age-dependent, linear increase in oxidative damage in WT nematodes, we find no evidence for autocatalytic damage amplification as proposed by the "vicious cycle" theory. Comparing the SOD mutants with wild-type animals, we further show that oxidative damage levels in the mtDNA of SOD mutants are not significantly different from those in wild-type animals, i.e. even the total loss of mitochondrial SOD did not significantly increase oxidative damage to mtDNA. Possible reasons for this unexpected result and some implications for the mFRTA are discussed.

Chronic resveratrol intake reverses pro-inflammatory cytokine profile and oxidative DNA damage in ageing hybrid mice.

Thymic involution and shrinkage of secondary lymphoid organs are leading causes of the deterioration of the T-cell compartment with age. Inflamm-aging, a sustained inflammatory status, has been associated with chronic diseases and shortened longevity. This is the first study to investigate the effect of treating aging hybrid mice with long-term, low-dose resveratrol (RSV) in drinking water by assessing multiple immunological markers and profiles in the immune system. We found that hybrid mice exhibited marked age-related changes in the CD3+CD4+, C3+CD8+, CD4+CD25+, CD4M and CD8M surface markers. RSV reversed surface phenotypes of old mice to that of young mice by maintaining the CD4+ and CD8+ population in splenocytes as well as reducing CD8+CD44+ (CD8M) cells in the aged. RSV also enhanced the CD4+CD25+ population in old mice. Interestingly, pro-inflammatory status in young mice was transiently elevated by RSV but it consequently mitigated the age-dependent increased pro-inflammatory cytokine profile while preserving the anti-inflammatory cytokine condition in the old mice. Age-dependent increase in 8OHdG, an oxidative DNA damage marker was ameliorated by RSV. Immunological-focused microarray gene expression analysis showed that only the CD72 gene was significantly downregulated in the 12-month RSV-treated mice compared to age-matched controls. Our study indicates that RSV even at low physiological relevant levels is able to affect the immune system without causing marked gene expression changes.

Elevation of oxidative-damage biomarkers during aging in F2 hybrid mice: protection by chronic oral intake of resveratrol.

Resveratrol (RSV), a naturally occurring phytoalexin that can be found in red wine, berries, and peanuts, has been shown to extend both mean and maximum life span in model organisms. RSV has also been reported to shift the physiology of middle-aged mice on a high-calorie diet toward that of mice on a standard diet. These beneficial effects of RSV have been suggested to resemble caloric restriction. Our study in F2 four-way cross-hybrid mice was the first to evaluate the effects of aging and long-term RSV treatment (14.09+/-3.4 mg/L in drinking water for 6 or 12 months) on biomarkers of oxidative damage to DNA, 8-hydroxy-2'-deoxyguanosine (8OHdG); lipid, 8-iso-prostaglandin(2 alpha) (8-iso-PGF(2 alpha)); and protein, protein carbonyl content (PCC). There was a significant age-dependent accumulation of oxidative damage to DNA, lipid, and protein as well as a clear increase in urine 8-iso-PGF(2 alpha) levels in the majority of mouse tissues. Rates of age-dependent increases in damage biomarkers varied between tissues. Chronic RSV treatment elevated total RSV plasma levels and reduced the observed age-dependent accumulation of (1) 8OHdG in liver and heart, (2) 8-iso-PGF(2 alpha) in heart and urine, and (3) PCC in liver and kidney. However, a 12-month RSV intake resulted in significant elevation of 8-iso-PGF(2 alpha) and PCC in kidney. Our studies demonstrate that RSV treatment consistently attenuated oxidative damage in tissues where age-related oxidative damage accumulation was prominent, but also suggested that chronic RSV treatment may induce nephrotoxicity.

Age-related changes in mitochondrial function and antioxidative enzyme activity in fischer 344 rats.

We have previously reported the changes of mitochondrial function and/or antioxidative enzyme efficiency in a few organs of rats as a result of aging. However, there is a further need to reach a conclusion about their interactions in biological functions based on other evaluation tips like the usage of advanced methods and the exploring of crucial biochemical parameters. Therefore, we investigated the mitochondrial inner membrane functional integrity by the analysis of respiration control ratio and membrane potential in the liver and brain of young (8 months) and old (26 months) Fischer 344 rats. The disintegration of mitochondrial membrane integrity was determined higher in the liver of old rats than that of young rats. This was well correlated with the decrease of total superoxide dismutase (SOD), Cu/Zn-SOD, Mn-SOD and glutathione peroxidase activities in most of the organs, except for the increase of catalase activity in heart of old rats. Similarly, the protein expressions of these enzymes were down regulated in the liver and kidney of old rats. Taken together, we suggest that the mitochondrial malfunction in old rats is associated with the decrease of antioxidative enzyme efficiency. And the data are also discussed with changes in the results from inter-laboratories.

Relationship between levels of oxidative DNA damage, lipid peroxidation and mitochondrial membrane potential in young and old F344 rats.

The extent of in vivo oxidative damage has been known to be cumulative over the period of the life of mammals. Our hypothesis is that there should be a positive correlation between the levels of 8-hydroxy-2'-deoxyguanosine (8OHdG) and 8-iso-prostaglandin F(2alpha) (8-iso-PGF(2alpha)) in major rat tissues. We also investigated whether increased level of oxidative stress causes a decrease in the mitochondrial membrane potential of peripheral lymphocytes of old rats using the MitoTracker Red fluorochrome. Our results show positive correlations between 8OHdG and 8-iso-PGF(2alpha) for liver, brain and kidney measured by HPLC-UV-ECD (electrochemical detector) and EIA methods, respectively. However, heart tissues show a negative correlation. The mitochondrial membrane potential of old rat lymphocytes records significant decrease compared with the young lymphocytes. Based on our results, we conclude that in ageing studies, specific tissues need to be examined in order to measure the localised DNA damage and lipid peroxidation as different tissues display different extent of oxidative damage. We believe this approach of using combined markers is useful to verify the true efficacy of health intervention studies in animals and humans. In addition, the isoprostane assay can be further developed looking at lipid peroxidation as a potential marker in ageing studies.