Yasuhiko Wada

Perceived mental stress and mortality from cardiovascular disease among Japanese men and women : the Japan Collaborative Cohort Study for Evaluation of Cancer Risk Sponsored by Monbusho (JACC Study)

Background—Perceived mental stress has been associated with risk of coronary heart disease (CHD) in white men, but no prospective data are available for other ethnic groups. Methods and Results—From 1988 to 1990, a total of 73 424 Japanese (30 180 men and 43 244 women), aged 40 to 79 years, without a history of stroke, CHD, or cancer completed a lifestyle questionnaire including perception of mental stress under the Japan Collaborative Cohort Study for Evaluation of Cancer Risk Sponsored by Monbusho (JACC Study). Systematic surveillance was completed until the end of 1997, with a 580 378 person-year follow-up, and the underlying causes of death were determined according to the International Classification of Diseases, 10th revision. For women, there were 316 with total stroke, 113 with CHD, and 643 with total cardiovascular disease (CVD); for men, there were 341, 168, and 778, respectively. Women who reported high stress had a 2-fold higher age-adjusted risk of mortality from total stroke and CHD and 1.5-fold higher risk of total CVD compared with those who reported low stress. Further adjustment for known cardiovascular risk factors and selected psychological variables did not alter the associations materially. The multivariate relative risk for women who perceived high stress versus low stress was 2.24 (95% CI 1.52 to 3.31, P <0.001) for total stroke, 2.28 (95% CI 1.17 to 4.43, P =0.02) for CHD, and 1.64 (95% CI 1.25 to 2.16, P <0.001) for total CVD. For men, these relations were generally weaker but suggestive of myocardial infarction. Conclusions—Perceived mental stress was associated with increased mortality from stroke for women and with CHD for men and women.

A tumor preventive effect of dietary restriction is antagonized by a high housing temperature through deprivation of torpor

Energy restriction (ER) has proven to be the only effective means of retarding aging in mice. The mechanisms of multiplicity of effects of ER on aging remain, however, fragmentary. ER induces daily torpor, the induction of which is reduced by increasing the ambient temperature to 30 degrees C. The effects of preventing hypothermia in ER animals were studied in terms of the expected consequences of ER on survival, disease pattern and a number of physiological parameters in autoimmune prone MRL/lpr mice and lymphoma prone C57BL, 6 mice. The results demonstrate that torpor plays a crucial role in the prevention of lymphoma development but does not have an affect on other aspects of ER, such as prevention of autoimmune diseases.

Evidence of an increased risk of hearing loss in heterozygous carriers in a Wolfram syndrome family

Wolfram syndrome (MIM 222300) is characterized by juvenile-onset diabetes mellitus and optic atrophy. Previous linkage analyses in the United States and UK families have indicated that the gene for Wolfram syndrome (WFS) is localized on the short arm of chromosome 4. We herein confirm the linkage of the WFS locus to D4S3023 on 4p with a two-point LOD score of 3.42 in a large Japanese family with Wolfram syndrome. Multipoint linkage analysis revealed the maximum LOD score of 4.82 between D4S3023 and D4S394. We also evaluated putative health risks in carriers by multiple logistic analysis with independent variables, age, gender, and numbers of affected haplotypes and with dependent variables, such as hearing loss, diabetes mellitus, polyuria, incontinence, psychological illness, and visual acuity. The results showed that the putative disease haplotype increased a risk of hearing loss (odds ratio =35.68, 95% confidence interval =4.12–308.95) and diabetes mellitus (odds ratio =7.57, 95% confidence interval =2.03–28.23) independently. This is the first report of an increased health risk of illness in carriers, other than for psychiatric disease.

SLC7A7 genomic structure and novel variants in three Japanese lysinuric protein intolerance families

Lysinuric protein intolerance (LPI) is a rare inherited disease caused by defective transport of the dibasic amino acids at the basolateral membranes of epithelial cells in the renal tubules and small intestine. The metabolic defect leads to brain dysfunction caused by hyperammonemia with a functional impairment of the urea cycle. Recently, mutations in the human SLC7A7 cDNA coding for y+LAT‐1, which express dibasic amino acid transport activity, were reported to be responsible for LPI. In the present study, we examined the genomic structure of SLC7A7 by DNA sequencing of PCR products, and determined that the gene had 11 exons and 10 introns spanning about 18 kb of genomic DNA. We also identified an alternative RNA splicing at the 5′ untranslated region of the SLC7A7 mRNA in human peripheral blood leukocytes, cultured lymphoblasts, and fibroblasts. As a result of mutational analysis of SLC7A7 in three Japanese LPI families, we found a nonsense mutation (R410X), a splicing mutation(911+1G>A) in intron 4, and four silent polymorphisms (201C/T, 445A/G, 784C/T, 946T/C). Identification of the genomic structure of SLC7A7 may provide a molecular basis for a genetic survey for LPI. Hum Mutat 15:367–372, 2000.

Evidence on n -acetyltransferase Allele-associated Metabolism of Hydrazine in Japanese Workers

Hydrazine (N2H4), which has been categorized as a weak carcinogen, is a chemical with the one of the largest production rates in Japan. We have investigated the effects of acetylation phenotypes on the metabolism of hydrazine. Genotypes of N-acetyl transferases, NAT2*, were determined using polymerase chain reaction for 297 male workers. Biological and exposure monitoring were also conducted. The rapid and intermediate acetylators accounted for 45% each, and the slow acetylators accounted for 10%. Biological half-lives were significantly different among the three acetylation phenotypes (analysis of variance, P < 0.05): 3.94+/-1.70 hours for slow acetylators, 2.25+/-0.37 hours for intermediate acetylators, and 1.86+/-0.67 hours for rapid acetylators. Among Japanese, rapid and intermediate acetylators are the major phenotypes, which is in sharp contrast with those among Caucasians. We conclude that biological monitoring should take genetic factors, which may vary dramatically among different populations, into account.

Low blood glucose levels and small islets of Langerhans in the pancreas of calorie-restricted mice

In the present study we investigated the influence of long-term caloric restriction on blood glucose levels and the volume of the islets of Langerhans in the pancreases of mice. Thirty female F1 mice (SHN female × C3H male) were divided into two equal groups and raised for 11 months on either a slightly restricted “control” diet (95 Kcal per week) or a more severely restricted diet (50 Kcal per week), starting at time of weaning (4 weeks of age). These diets, designated as “isonutrient” (1), were designed so that both groups of animals received approximately equal amounts of minerals, proteins, fats and vitamins, but with carbohydrates adjusted to provide the desired calorie differences. Calorie-restricted animals (CR mice) were fed 3 days a week (Monday, Wednesday and Friday) with the restricted diet at 9:00 a.m., while control animals (C mice) were fed the control diets at 9:00 a.m. every day from Monday to Friday. On Sunday any leftover food was removed from the cages at 9:00 a.m. and both CR and C animals were allowed to fast. Fasting blood glucose levels were determined in the blood collected between 9:00 and 10:00 a.m. in 12-month-old mice from both groups on Saturday and on Monday 8 days later, at which time the mice were sacrificed for histological examination. The Saturday morning blood glucose levels (in mg/100 ml of blood) were lower in CR mice than in C mice (112±12 in CR mice and 145±16 in the C mice: p<0.01). A similar trend was noted in the Monday morning samples: 80±19 in the CR and 108±28 in the C mice. In agreement with the blood glucose trends, histological examination of the pancreases demonstrated that the volume of the islets of Langerhans were significantly smaller in CR mice (Geometric mean 76646 μ3/islet, Geometric Standard deviation 2.10) than in C mice (Geometric mean 235578 μ m3/islet, Geometric Standard deviation 3.81)(p<0.01). We conclude that caloric restriction decreases not only blood glucose levels, but reduces volumes of the islets of Langerhans.

Caloric restriction perturbs the pituitary-ovarian axis and inhibits mouse mammary tumor virus production in a high-spontaneous-mammary-tumor-incidence mouse strain (C3H/SHN)

Dietary restriction (DR) retards aging and extends maximum life span. It is also known to decrease the incidence of hormone-dependent tumors. In the present investigation, we focussed primarily on the influence of DR on the pituitary-ovarian axis, and subsequently on gene expression of the mouse mammary tumor virus. F1 females from the mating of SHN female and C3H male mice were used in this study, since these hybrids display a very high incidence of mouse mammary tumors. The mice weaned at 3 weeks were raised on either a calorically-restricted diet (DR: 50 kcal/week; N = 5) or on a control diet (C: 95 kcal/week N = 5) for 5 weeks. Three C57BL/6J Jcl ad libitum-fed female mice, 8 weeks of age, were used as reference animals since this strain has a very low incidence of mammary tumors. The mean cellular contents of prolactin (PRL) and growth hormone (GH) in the pituitary, as determined by immunohistochemistry, were found to be reduced in mice raised on the DR diet. The decrease in the mean cellular content of PRL (50% of the mean control value) was accompanied by a decrease in the number of lactotrophs (17% of the mean value of control diet mice). However, the decrease in cellular content of GH (53% of the mean control value) was not accompanied by a decrease in the number of somatotrophs (no. of somatotrophs in DR = 103% of mean control value). Histologically, ovaries from DR mice showed many growing and atretic follicles, with few corpora lutea. In contrast, both control-diet and reference (C57BL/6J Jcl) mice showed two or three corpora lutea per ovarian section. In accordance with this finding, DR mice had not established stable estrus cycles by 8 weeks of age, in contrast to both control-diet and reference mice. Since caloric restriction has been shown to decrease mammary tumor virus (MMTV) gene expression, MMTV production was investigated by electron microscopy to confirm the validity of our experimental conditions. In DR or reference C57BL/6J Jcl mice, MMTV particles were rarely found in the mammary gland samples, but were always found in samples from control mice. The development of mammary glands, as indicated by the number of villi or the development of the rough endoplasmic reticulum, was delayed in DR mice. Thus, it was concluded that caloric restriction decreases the number of lactrotrophs, inhibits ovulation and delays mammary gland development. This immature status is considered to be due to perturbations in the pituitary-ovarian axis by caloric restriction.

A Linear Pharmacokinetic Model Predicts Usefulness of N-Methyl-2-Pyrrolidone (NMP) in Plasma or Urine as a Biomarker for Biological Monitoring for NMP Exposure

A Linear Pharmacokinetic Model Predicts Usefulness of N‐Methyl‐2‐Pyrrolidone (NMP) in Plasma or Urine as a Biomarker for Biological Monitoring for NMP Exposure: Xiaofei E, et al. Department of Hygiene, Akita University School of Medicine—N‐methyl‐2‐pyrrolidone (NMP: C5H9NO:CAS number 872‐50‐4) is an increasingly used solvent due to the lack of ozone depleting activity. The aim of this study is to construct a simple pharmacokinetic model for NMP. In factory A, four workers who were exposed to NMP at 0.09‐0.69 ppm for 12 h by time weighted average (TWA) were followed up for an entire workweek. Their NMP concentrations in plasma and urine were monitored during the observation period. Five volunteers were exposed to NMP during the observation of workers in the factory A for eight hours. NMP kinetics in plasma and urine were monitored for 2 d after exposure. Concentrations of NMP in plasma and urine as standardized by creatinine concentrations were used to construct a one compartment pharmacokinetic model. The model successfully simulated the kinetics in four workers and five volunteers. In the next step, the model was applied to eight workers in another factory: they were exposed to NMP for 12 h at 0.04 to 0.59 ppm by TWA. The model could successfully predict kinetics of NMP levels in plasma and urine at the end of work. The model was then applied to experimental exposure cases in the literature. The model successfully predicted the concentrations of NMP in plasma and urine at the exposure intensity level of 12 ppm x 8 h. These results imply that metabolic saturation does not occur up to the exposure intensity of 12 ppm x 8 h and demonstrate the usefulness of determinations of NMP in plasma and urine for biological monitoring.

Demonstration of a new mouse mammary tumor virus locus in the genome of the mammary tumor phone strain SHN mouse

Until this report, there have been no detailed analyses of the mouse mammary tumor virus (MMTV) loci in the mammary tumor prone strain SHN. Using a probe, which hybridizes to the env sequence of MMTV, Southern blotting of genomic DNA from the brain after digestion with EcoRI revealed 5 endogenous proviruses: Mtv-1 (4.5kb), Mtv-2 (11 kb), Mtv-8 (6.7kb), Mtv-17 (8.3kb) and a newly-found 6.5-kb fragment. F1-hybrid mice (C3H/He female x SHN male) also possessed the 6.5-kb fragment. Thus, we conclude that the 6.5-kb fragment is unique to SHN mice. Genomic DNA from mammary tumors of SHN mice showed MMTV insertions, suggesting that activation of an oncogene(s) occurred in this strain.

Energy restriction suppresses microsomal Ca2+-ATPase activities in various organs in C57BL/6 female mice in both euthermic and torpor states

C57BL/6 female mice were fed a daily control diet (n = 5, 3.9 g/day, 95 kcal/week) or ER diet (n = 5, 2.3 g/day, 48 kcal/week) at 1800 h from 6 weeks of age. Telemetry, conducted at 6 months of age, confirmed that all ER mice entered daily torpor (core body temperature less than 31 degrees C) for 6.63 +/- 2.34 h/day while control mice were euthermic (> 35 degrees C). In vitro activities of microsomal Ca(2+)-ATPase were determined in the brain, liver, salivary gland and kidney from these mice at 6 months of age. Assays were performed at three incubation temperatures of 37 degrees C, 31 degrees C and 25 degrees C. In assays at 37 degrees C, the activities of Ca(2+)-ATPase in the brain and salivary gland from ER mice were lower than those in corresponding organs from control mice. The suppression became profound as the incubation temperature decreased. On the other hand, at 37 degrees C Ca(2+)-ATPase activities in the liver and kidney from ER mice were not lower than those in corresponding organs from control mice, but decreased significantly at low temperatures. Microsomal Ca(2+)-ATPase activities thus appeared to be reduced in ER mice, although it remains unknown whether the present results represent reduced in vivo capacities to regulate cytosolic Ca2+ concentrations.