Kazuyoshi Tamae

Occupational and lifestyle factors and urinary 8-hydroxydeoxyguanosine

The amount of 8‐hydroxydeoxyguanosine (8‐OH‐dG) excreted in urine can be used not only as an indicator of DNA repair capacity, but also as a potential marker of oxidative DNA damage. To clarify the oxidation‐related factors, in consideration of cancer risk, this study investigated how urinary 8‐OH‐dG was associated with occupational and lifestyle factors in 372 healthy workers. The creatinine‐adjusted urinary 8‐OH‐dG level was significantly higher in male subjects, smokers and drinkers compared with their counterparts. There were significant positive correlations of the 8‐OH‐dG level with average number of working hours, involvement in work, average number of cigarettes smoked, average volume of alcohol consumed and serum cortisol level, and there were significant negative correlations of the 8‐OH‐dG level with body mass index (BMI) and consumption of soybean products, rice and light‐colored vegetables. Multiple regression analysis showed that average number of working hours and average number of cigarettes smoked were significant predictors of increased 8‐OH‐dG levels, whereas being female and BMI were significant predictors of decreased 8‐OH‐dG levels. Working hours, BMI and smoking were significant predictors of urinary 8‐OH‐dG in male subjects, whereas age and BMI were significant predictors in female subjects. We suggest that several occupational and lifestyle factors, particularly long working hours and cigarette smoking, are linked to the formation of 8‐OH‐dG in workers. (Cancer Sci 2005; 96: 600 – 606)

Effect of age, smoking and other lifestyle factors on urinary 7‐methylguanine and 8‐hydroxydeoxyguanosine

Urinary 8‐hydroxydeoxyguanosine (8‐OH‐dG) and 7‐methylguanine (m7Gua) were measured by a column‐switching high performance liquid chromatography method as markers of oxidative and methylating DNA damage, respectively. We investigated the associations between urinary 8‐OH‐dG or m7Gua and various lifestyle and demographic factors, such as age and sex. The urinary 8‐OH‐dG excretion level was positively correlated with cigarette smoking, but inversely correlated with fruit consumption, physical activity and total energy consumed per day. A multiple regression analysis revealed that daily physical activity and healthy meal combinations decreased the urinary 8‐OH‐dG level, whereas alcohol consumption increased it. In terms of the urinary m7Gua measurement, cigarette smoking, age and consumption of meat, fish, egg, soybean, etc. were positively correlated with the urinary m7Gua level, whereas body weight, BMI, physical activity, and dietary index score, which indicates good nutritional balance, were negatively correlated with the amount of m7Gua. Based on a multiple regression analysis, cigarette smoking and age correlated with the m7Gua level, while high BMI and healthy meal combinations have significant reducing effects on m7Gua level. Therefore, the urinary m7Gua level is considered to be a useful marker of DNA methylation, not only from smoking, but also from aging and unhealthy dietary habits. (Cancer Sci 2009; 100: 715–721)

Suppression of hyperemia and DNA oxidation by indomethacin in cerebral ischemia

We investigated antioxidative activity and the effect of indomethacin, an agent that inhibits cyclooxygenase, on extracellular glutamate and cerebral blood flow in cerebral ischemia in gerbils. Pre-ischemic administration of indomethacin (5 mg/kg, i.p.) significantly rescued hippocampal CA1 neurons (9+/-6 cells/mm in the ischemia, 87+/-43 cells/mm in the indomethacin group, P<0.001). DNA fragmentation induced by ischemia was also examined using the terminal deoxynucleotidyl transferase-mediated UTP nick end labeling (TUNEL) method and indomethacin reduced TUNEL positive cells (140+/-21 in the ischemia, 99+/-31 in the indomethacin group, P<0.01). In addition, indomethacin attenuated the increase in hippocampal blood flow during reperfusion, but not increased extracellular glutamate by ischemia. Eight-hydroxydeoxyguanosine (8-OH-dG), a highly sensitive marker of DNA oxidation, was measured 90 min following ischemia using high-pressure liquid chromatography. Indomethacin significantly decreased the level of ischemia-induced 8-OH-dG in the hippocampus (P<0.05). These results suggest that indomethacin may protect neurons by attenuating oxidative stress and reperfusion injury in ischemic insult.

Heavy Metal-Induced Oxidative DNA Damage in Earthworms: A Review

Earthworms can be used as a bio-indicator of metal contamination in soil, Earlier reports claimed the bioaccumulation of heavy metals in earthworm tissues, while the metal-induced mutagenicity reared in contaminated soils for long duration. But we examined the metal-induced mutagenicity in earthworms reared in metal containing culture beddings. In this experiment we observed the generation of 8-oxoguanine (8-oxo-Gua) in earthworms exposed to cadmium and nickel in soil. 8-oxo-Gua is a major premutagenic form of oxidative DNA damage that induces GC-to-TA point mutations, leading to carcinogenesis.

8-Hydroxyguanine levels and repair capacity during mouse embryonic stem cell differentiation.

Abstracts To evaluate the defence capacities of embryonic stem (ES) cells against gene impairment, this study measured the levels of 8-hydroxyguanine (8-OH-Gua), a well-known marker of oxidative stress in DNA, and its repair capacity during differentiation. Undifferentiated ES cells (EB3) were cultured without leukaemia inhibitory factor (LIF) for 0, 4 and 7 days and are referred to as ES-D0, ES-D4 and ES-D7, respectively. These three cell lines were treated with 300 μM hydrogen peroxide (H2O2) for 48 and 72 h. After treatment, the amounts of 8-OH-Gua in the cells were determined by the high-performance liquid chromatography (HPLC)-electrochemical detector (ECD) method. The levels of 8-OH-Gua in ES-D7 treated with H2O2 were higher than those in ES-D0 and ES-D4, suggesting that the DNA in the undifferentiated cells was protected against gene impairment, as compared to that in the differentiated cells. To examine the repair capacity for 8-OH-Gua, this study analysed the expression of 8-OH-Gua repair-associated genes, 8-oxoguanine DNA glycosylase 1 (OGG1), MutY homolog (MUTYH) and Mut T homolog 1 (MTH1), in ES-D0, ES-D4 and ES-D7. The mRNA levels of MUTYH and MTH1 showed no significant change, whereas OGG1 mRNA was significantly decreased in ES-D7 treated with H2O2. Moreover, it was observed that ES-D7 treated with H2O2 readily underwent apoptosis, in comparison to its undifferentiated counterparts, ES-D0 and ES-D4. Taken together, ES cells are more resistant to DNA oxidative stresses than differentiated cells.

Differentiation of Embryonic Stem Cells and Oxidative DNA Damage /DNA Repair Systems

To maintain genomic integrity, cells are equipped with a defense capacity against DNA damage generated in genomic DNA. The avoidance of mutations in genomic DNA is especially critical for undifferentiated cells, such as Embryonic stem (ES) cells. Among the many factors damaging DNA, Reactive Oxidative Species (ROS) are crucial and unavoidable, and thus the roles of ROS in cell differentiation and cellular functions have been extensively studied. However, the biological significance of ROS generation in cell differentiation remains a matter of debate. Among the types of DNA damage due to ROS, 8-oxoguanine (8-oxo-Gua) has been well studied and is known to generate GC-to-TA point mutations in genomic DNA. We recently analyzed the differences in the resistance to 8-oxo-Gua generation between undifferentiated cells and their differentiated counterparts. Our studies indicated that undifferentiated cells were more resistant to 8-oxo-Gua generation, in comparison to differentiated cells. In this short review, we describe the relationship between ES cell differentiation and oxidative DNA damage / DNA repair systems, by summarizing our previous work and that of other researchers.

Enhancement of adipogenesis induction by conditioned media obtained from cancer cells

White adipose tissue is a multifunctional endocrine organ that synthesizes and secretes cytokine-like proteins termed adipokines. In the present study, the effects of cancer-derived medium on adipogenesis were examined. We prepared conditioned media from cancer cell lines, and cultured preadipocytes in the conditioned media. After 10 days of culture, intracellular lipid droplet accumulation was measured. We observed that the conditioned media significantly induced adipogenesis or enhanced the adipogenesis induced by adipogenesis inducers. Although we could not define the factors, these undefined factors derived from cancer cells may induce adipogenesis, and the resulting adipogenesis may affect cancer development.

Analyses of Associations Between Reactive Oxygen Metabolites and Antioxidant Capacity and Related Factors Among Healthy Adolescents

Evidence based on epidemiologic investigations using biochemical parameter is meaningful for health promotion and administration among adolescents. We conducted Reactive Oxygen Metabolites (ROM) and Biological Antioxidant Potentials (BAP) tests, along with a questionnaire survey, for a sample of 74 high school students (16.51±0.11 years of aged mean±SE), to investigate the associations between ROM, BAP, and related factors, including BMI and blood biochemical data. Venous blood samples (approximately 7cc) were collected. At the same time, each individuals information was obtained from the questionnaire. The mental health status was investigated using the Center for Epidemiologic Study Depression scale (CES-D) included in the same questionnaire. The mean values and standard errors of all variables were calculated. In addition, the relationships between ROM and BAP with these factors were analyzed. The results revealed the preferred levels of ROM (261.95 ± 9.52 U.CARR) and, BAP (2429.89±53.39 µmol/L) and blood biochemical data. Few significant relationships between two markers and related factors were found. So, we detected a cluster with an imbalance between ROM and BAP, which means low antioxidant ability, whereas the other clusters had conditions with moderate balance or good balance between them. Moreover, we determined the Oxidative stress-Antioxidant capacity ratio (OAR), using the ROM and BAP values, in order to clarify the characteristic of the detected clusters.However, comparative analyses across the three clusters did not yield significant differences in all related factors. No correlations between ROM, BAP and related factors were indicated, although significant association between ROM and BAP was observed (R2=0.1156, R=0.340, P=0.013). The reason for these results can be explained by the influences of good health and young age. On the other hand, present study suggests that some latent problems among adolescents may be related to unhealthy conditions in the future. Also, this study indicated that ROM and BAP may be useful as markers of the oxidative stress status. After this, further investigations using biomarkers based on epidemiologic design should be conducted, to reveal the reliability of the present results.