Shinya Asami

Psychosocial Factors as a Potential Trigger of Oxidative DNA Damage in Human Leukocytes

Although numerous studies have been carried out on the stress‐cancer linkage, the results are still inconclusive. One of the useful, but rarely applied, methods to assess this linkage is to examine the relationship between psychosocial stress and cancer‐predisposing genetic alterations simultaneously. We investigated whether various psychosocial factors can be associated with the levels of 8‐hydroxydeoxyguanosine (8‐OH‐dG), a biomarker of cancer‐related oxidative DNA damage, in peripheral blood leukocytes in 362 healthy workers (276 males and 86 females). After adjustments for age, body mass index, cigarette smoking, and alcohol use, female subjects showed positive relationships between the amount of 8‐OH‐dG and the Tension‐Anxiety, Depression‐Rejection, Anger‐Hostility, Fatigue, and Confusion scores of the Profile of Mood States, respectively. The levels of 8‐OH‐dG also increased reliably in the female subjects who had poor stress‐coping behaviors, particularly wishful thinking strategy, in the NIOSH general job stress instrument. There were positive relationships of the 8‐OH‐dG levels to average working hours, a self‐blame coping strategy, and recent loss of a close family member in male subjects. These findings in a nonclinical sample of healthy adults not only provide evidence of a stress‐cancer linkage, but also suggest possible sex differences in the mechanisms of stress‐related cancer initiation.

Reduction of 8-hydroxyguanine in human leukocyte DNA by physical exercise

We investigated the effect of physical exercise on the level of 8-hydroxyguanine (8-OH-Gua), a form of oxidative DNA damage, and its repair activity in human peripheral leukocytes. Whole blood samples were collected by venipuncture from 21 healthy male volunteers (10 trained athletes and 13 untrained men), aged 19-50 years, both before and after physical exercise. Trained athletes showed a lower level of 8-OH-Gua (2.4+/-0.5/10(6) Gua, p = 0.0032) before exercise when compared to that of untrained men (6.2+/-3.5). The mean levels of 8-OH-Gua of untrained subjects decreased significantly (p = 0.0057) from 6.2+/-3.5/10(6) Gua (mean+/-SD/10(6) Gua) to 3.3+/-1.4/10(6) Gua after physical exercise. On the other hand, the mean levels of repair activity of untrained subjects significantly increased after exercise (p = 0.0093) from 0.037+/-0.024 (mean DNA cleavage ratio+/-SD) to 0.056+/-0.036. In the trained athletes 8-OH-Gua level and its repair activity were not changed before and after the exercise. We also observed inter-individual differences in 8-OH-Gua levels and its repair activities. These results suggest that physical exercise causes both rapid and long-range reduction of oxidative DNA damage in human leukocytes, with individually different efficiencies.

Classical conditioning of oxidative DNA damage in rats.

This study investigated whether the formation of 8-hydroxydeoxyguanosine (8-OH-dG), a known oxidative DNA modification relevant to carcinogenicity, can be classically conditioned to a novel taste in order to clarify the possible role of the central nervous system (CNS) or psychological stress on cancer initiation via a classical conditioning mechanism. Male Wistar rats underwent one or two conditioned taste aversion (CTA) experiments in which ferric nitrilotriacetate (Fe-NTA), which has renal toxicity and can induce renal cell carcinoma, served as a visceral unconditioned stimulus (US), and a saccharin solution (SAC) was used as a conditioned stimulus (CS). The 8-OH-dG levels in the group conditioned with the combination of SAC and Fe-NTA significantly increased as compared to those of the uncombined groups by two repeats of the conditioning procedure (P=0.013). The rats that showed a painful response at the Fe-NTA administration had significantly higher values of 8-OH-dG than those without pain (P=0. 003). These results not only provide the first evidence regarding classical conditioning of oxidative DNA damage using the CTA procedure, but also suggest the involvement of the CNS and psychological stress in the pathogenesis of cancer via oxidative DNA damage.

Psychological Mediation of a Type of Oxidative DNA Damage, 8-Hydroxydeoxyguanosine, in Peripheral Blood Leukocytes of Non-Smoking and Non-Drinking Workers

Background: The present study investigates whether the formation of 8-hydroxydeoxyguanosine (8-OH-dG), a known oxidative DNA damage relevant to carcinogenicity, can be associated with psychological factors, in order to clarify the possible stress-cancer linkage from a genetic viewpoint. Methods: We performed a cross-sectional study in which we examined the relationships of the levels of 8-OH-dG in peripheral blood leukocytes to various psychological factors, including the Profile of Mood States (POMS) and the Center for Epidemiologic Studies Depression Scale (CES-D) in 38 non-smoking and non-drinking workers (19 males and 19 females). Results: The levels of 8-OH-dG in male subjects were negatively correlated with the Tension-Anxiety scores of the POMS. In contrast, the levels of 8-OH-dG in female subjects were positively correlated with the Depression-Rejection scores of the POMS and the CES-D scores, and negatively associated with the Vigor scores of the POMS, respectively. Male subjects who had self-blame coping strategy displayed significantly high levels of 8-OH-dG. Moreover, the worse the subjective closeness to parents in childhood, the higher the levels of 8-OH-dG became in male subjects. The levels of 8-OH-dG increased reliably in subjects who had experienced the loss of a close family member within 3 years, when compared with non-bereaved subjects. Conclusions: Psychological distress may be associated with cancer risk, although sex difference influences them. Inadequate coping styles, possibly resulting from a poor interpersonal relationship with parents since childhood, and experience of a relatively recent loss of a close family member also appear to influence the pathogenesis of cancer.

Increased 8-Hydroxyguanine in DNA and Its Repair Activity in Hamster and Rat Lung after Intratracheal Instillation of Crocidolite Asbestos

Asbestos and man‐made‐mineral fibers are known to increase one type of oxidative DNA damage, 8‐hydroxyguanine (8‐OH‐Gua), in vitro. In this study, we analyzed the 8‐OH‐Gua level in DNA and its repair activity after a single intratracheal instillation of fibers (crocidolite or glass) or saline to Syrian hamsters or Wistar rats. The 8‐OH‐Gua level was measured with a high‐performance liquid chromatography‐electrochemical detector (HPLC‐ECD) system. The 8‐OH‐Gua repair enzyme activity was determined with an endonuclease nicking assay using a 32P‐labeled or fluorescently labeled 22mer DNA that contains 8‐OH‐Gua at a specific position. A significant increase in the 8‐OH‐Gua level in the lung DNA was observed 1 day after the exposure to crocidolite, as compared to the saline control. The repair activity was increased significantly at 7 days. On the other hand, after exposure to glass fibers, little or no increase of these carcinogenicity indicators was detected. These assays of 8‐OH‐Gua and its repair activity in short‐term animal experiments will be useful for evaluating the carcinogenicity of fibers. This is the first report of the increase of 8‐OH‐Gua and its repair activity in the animal lung after the instillation of asbestos fibers.

Increase in 8-hydroxyguanine and its repair activity in the esophagi of rats given long-term ethanol and nutrition-deficient diet

Epidemiological studies have shown that an increased risk of esophageal cancer is associated with the chronic consumption of alcoholic beverages, although alcohol itself is not a carcinogen in animal models. Reactive oxygen species produced by the metabolism of ethanol or by chronic inflammation may play an important role in the carcinogenic process. In this study, we analyzed one type of oxidative DNA damage, 8‐hydroxyguanine (8‐OH‐Gua), and its repair activity in the esophagus as indicators of cellular oxidative stress in rats given long‐term ethanol and an autoclaved diet (nutrition‐deficient diet). Three‐week‐old male Sprague‐Dawley rats were fed an ethanol beverage whose concentration was increased from 12 to 70% over 20 weeks. When the concentration reached 50%, the diet of one group was changed from the regular diet to an autoclaved diet. At the feeding periods of 20, 25, 30, and 35 weeks, the rats were sacrificed and the 8‐OH‐Gua levels and repair activities within the esophagi were measured. After 30 weeks of ethanol‐ and autoclaved diet‐feeding, significant increases of 8‐OH‐Gua and its repair activity were observed in the esophagi, but not in those of the ethanol‐ and normal diet‐fed rats. This result indicates that the combined effects of long‐term ethanol consumption and nutritional deficiency may be involved in inducing oxidative stress in the rat esophagus.

2-Hydroxyadenine, a mutagenic form of oxidative DNA damage, is not repaired by a glycosylase type mechanism in rat organs

Oxygen radicals are known to play a role in causing cellular DNA damage, which is involved in carcinogenesis. 8-Hydroxyguanine (8-OH-Gua) is a major form of oxidative DNA damage and is known as a useful marker of DNA oxidation. Recently, we found another type of oxidative DNA damage, 2-hydroxyadenine (2-OH-Ade), which has a mutation frequency comparable to that of 8-OH-Gua. We compared the repair activities for two types of oxidative DNA damage, 8-OH-Gua and 2-OH-Ade, in 7-week-old male Sprague-Dawley (SD) rat organs. The repair activities were measured by an endonuclease nicking assay using 22 mer [32P]-end-labeled double-stranded DNA substrates, which contained either 8-OH-Gua (opposite C) or 2-OH-Ade (opposite T or C). In all of the SD rat organs we studied, the nicking activity for 2-OH-Ade was not detected, while that for 8-OH-Gua was clearly detected with the same conditions. Moreover, the 2-OH-Ade nicking activity was not induced in Wistar rat kidney extracts prepared after ferric nitrilotriacetate (Fe-NTA) treatment, which is known to increase 8-OH-Gua repair activity. These results suggest that 2-OH-Ade might not be repaired by the glycosylase type mechanism in mammalian cells.

Increased 8-Hydroxyguanine Repair Activity in Animal Livers and Human Leukocytes Induced by Oxidative Stress: Effects of Diet, Physical Exercise, and Cigarette Smoking

8-Hydroxyguanine (8-OH-Gua) is a major indicator of oxidative DNA damage. Because 8-OH-Gua repair activity is induced when cells are exposed to oxidative stress due to ionizing radiation, measuring 8-OH-Gua repair activity is useful for assessing cellular oxidative stress. Increased 8-OH-Gua repair activity was observed in the livers of alcohol-and autoclaved-diet-fed rats but not in alcohol- or normal-diet-fed rats. It was found that various vitamins decomposed during the autoclaving. Therefore the level of vitamins may play an important role in reducing oxidative stress in the livers of alcohol-fed rats. An increase in 8-OH-Gua repair activity was observed in human leukocytes after physical exercise, and each individual had a different level of increase. A higher level of 8-OH-Gua repair activity was detected in the leukocytes of smokers compared to that in nonsmokers. A sevenfold interindividual difference was observed in the 8-OH-Gua repair activities of smokers. It is possible to estimate the cancer risk due to oxidative stress of each individual based on the 8-OH-Gua repair assay with human leukocytes.