Persistent organic pollutants in red-crowned cranes (Grus japonensis) from Hokkaido, Japan.

The red-crowned crane (Grus japonensis) from eastern Hokkaido is classified as a Special Natural Monument in Japan. In this study, we determined the concentrations of persistent organic pollutants (POPs) in red-crowned crane muscle tissues (n = 47). Polychlorinated biphenyls (PCBs) had the highest median concentration (240ng/g lipid weight), followed by dichlorodiphenyltrichloroethane and its metabolites (DDTs) (150ng/g lipid weight), chlordane-related compounds (CHLs) (36ng/g lipid weight), hexachlorobenzene (HCB) (16ng/g lipid weight), hexachlorocyclohexanes (HCHs) (4.4ng/g lipid weight), polybrominated diphenyl ethers (PBDEs) (1.8ng/g lipid weight), and finally, Mirex (1.5ng/g lipid weight). Additionally, a positive correlation was found among POP concentrations. No sex differences beyond body parameters were observed. Additionally, red-crowned cranes exhibited a high enantiomeric excess of (+)-alpha-HCH, with enantiomer fractions varying from 0.51 to 0.87 (average: 0.69).

Fetal exposure to diesel exhaust affects X-chromosome inactivation factor expression in mice.

Several studies have shown effects of diesel exhaust (DE) on the central nervous system, but the mechanism is unclear. Fetal mice were exposed to whole DE (contains gases and particles) in an inhalation chamber, and cerebrum gene expression changes were examined by gene assay (microarray and quantitative real-time PCR). By microarray, upregulation of Xist, B-raf and Drwms2 were detected. Especially, mRNA expression of Xist was increased in a concentration-dependent manner in male and female mice. Xist (X-inactive specific transcript) is a major effector of the X-inactivation process, and X-linked genes are highly expressed in brain tissue and consistent with a role in brain developments. By quantitative real-time PCR, Tsix (crucial noncoding antisense partner of Xist) and other X-linked genes (Mecp2, Hprt1, and Sts) were examined; Tsix was upregulated, and other X-linked genes were unaffected in the male and female mice. Our findings suggest that exposure to DE increases Xist and Tsix gene expression in utero without influencing X-linked gene expression. An examination of Xist gene expression changes may provide an important biomarker for DE-induced effects. The possibility of avoiding X-chromosome inactivation (XCI) mechanisms by minimizing exposure to DE is expected.

Acute effects of diesel emission from the urea selective catalytic reduction engine system on male rats.

Short-term inhalation experiments were performed using Fischer 344 rats exposed to emission from the urea selective catalytic reduction (SCR) diesel engine system to identify health effects and compare them to those of the conventional diesel engine system. Rats were exposed to high-, middle-, or low-concentration emission (dilution ratio 1:29, 1:290, or 1:580) or clean air (control) for 1, 3, or 7 days (6 h/day), under driving conditions at a speed of 1320 rpm and a torque of 840 Nm. For the high-concentration group, the major components of the urea SCR emission were 0.04 mg/m(3) particulate matter (PM) and 0.78 ppm nitrogen dioxide (NO(2)); those of the conventional emission were 0.95 mg/m(3) PM and 0.31 ppm NO(2). The authors evaluated the respiratory effects of each emission on rats. Lymphocytes for 3-day exposure of both emissions significantly increased in bronchoalveolar lavage fluid, but there were slight differences. With an increase in potential antioxidant (PAO), 8-hydroxy-2'-deoxyguanosine for the urea SCR emission was significantly decreased, but that of the conventional emission was highest among all groups and did not show a response to PAO. In lungs, heme oxygenase (HO)-1 and tumor necrosis factor (TNF)-alpha mRNA expressions for the urea SCR emission showed a tendency to increase compared to those of the conventional emission. Thus, gene analysis results suggested that NO(2) from the urea SCR emission affected the expressions of mRNAs in lungs. However, as a whole, the results suggested that the health effects of the urea SCR emission might be less than the conventional emission on rats.

Toxicological evaluation of diesel emissions on A549 cells.

To evaluate the health effects of diesel emissions (DE) using an in vitro experiment, A549 cells were exposed to emission from a diesel engine on an engine dynamo, using a culture-cell-exposure device. Three groups were set according to cell exposure to high concentrations of particulate matter (PM) and/or nitrogen dioxide (NO(2)). The emissions of each group was dilution rate 1:100 and 1:10, and PM was 0.8mg/m(3) and/or NO(2) was 80ppm at dilution rate 1:10. After 1h, exposed cells were analyzed for cell viability and gene expression. Fifty percent of cell viability in the high-PM/high-NO(2) exposure group occurred at a dilution rate of 1:14, based on the concentration of CO(2). Heme oxygenase-1 mRNA expression significantly increased at 1:100 dilution of the high-PM/high-NO(2) group and 1:100 and 1:10 dilutions of the high-PM/low-NO(2) group, compared to background air. By DNA microarray, all gene expressions at a dilution rate of 1:10 in each group were observed to be higher than those at 1:100, and some cancer-related genes up-regulated. We concluded that screening methods for evaluating health effects could be established using this cell-exposure system because the effects of DE on A549 cells were shown by cell viability and gene expression.

Perinatal exposure to diesel exhaust affects gene expression in mouse cerebrum.

Many environmental toxins alter reproductive function and affect the central nervous system (CNS). Gonadal steroid hormones cause differentiation of neurons and affect brain function and behavior during the perinatal period, and the CNS is thought to be particularly susceptible to toxic insult during this period. It was, therefore, hypothesized that inhalation of diesel exhaust (DE) during the fetal or suckling period would disrupt the sexual differentiation of brain function in mice, and the effects of exposure to DE during the perinatal period on sexual differentiation related gene expression of the brain were investigated. In the fetal period exposure group, pregnant ICR mice were exposed to DE from 1.5 days post-coitum (dpc) until 16 dpc. In the neonatal period exposure group, dams and their offspring were exposed to DE from the day of birth [postnatal day (PND)-0] until PND-16. Then, the cerebrums of males and females at PND-2, -5, and -16 from both groups were analyzed for expression level of mRNA encoding stress-related proteins [cytochrome P450 1A1 (CYP1A1), heme oxygenase-1 (HO-1)] and steroid hormone receptors [estrogen receptor alpha (ER alpha), estrogen receptor beta (ER beta), androgen receptor (AR)]. Expression levels of ER alpha and ER beta mRNA were increased in the cerebrum of newborns in the DE exposure groups as well as mRNA for CYP1A1 and HO-1. Results indicate that perinatal exposure to DE during the critical period of sexual differentiation of the brain may affect endocrine function.

Detrimental effects of prenatal exposure to filtered diesel exhaust on mouse spermatogenesis.

We recently showed that prenatal exposure to diesel exhaust (DE) disrupts spermatogenesis in mouse offspring. This study was undertaken to determine whether filtered DE in which 99.97% of diesel exhaust particles >0.3 microm in diameter were removed affects spermatogenesis in growing mice. After prenatal exposure to filtered DE for 2-16 days postcoitum, we examined daily sperm production (DSP), testicular histology, serum testosterone levels and mRNA expression of hormone synthesis process-related factors. In the filtered DE exposed group, DSP was markedly reduced at 12 weeks compared with the control group; clean air exposed group. Histological examination showed multinucleated giant cells and partial vacuolation in the seminiferous tubules of the exposed group. Testosterone was elevated significantly at 5 weeks. Moreover, luteinizing hormone receptor mRNA at 5 and 12 weeks, 17alpha-hydroxylase/C17-20-lyase and 17beta-hydroxysteroid dehydrogenase mRNAs at 12 weeks were significantly elevated. These results suggest that filtered DE retains its toxic effects on the male reproductive system following prenatal exposure.

Prenatal exposure to diesel exhaust impairs mouse spermatogenesis.

The effect of prenatal exposure to diesel exhaust (DE) was investigated. Twenty pregnant ICR mice were exposed to DE at the particle concentration of 1.0 mg/m3, from d 2 until d 16 postcoitum. Male offspring were kept alive until 12 wk of age, and then male reproductive organ weight, daily sperm production (DSP), serum testosterone level, and mRNA expression of sex steroid hormone synthesis process-related factors were measured. Serum testosterone levels of the exposed group were reduced significantly at 3 wk, whereas they were elevated significantly at 12 wk. DSP was also markedly reduced at 5 and 12 wk. Histological examination showed multinucleated giant cells in the seminiferous tubules of the exposed group as well as partial vacuolation of the seminiferous tubules. Follicle-stimulating hormone receptor (FSHR) mRNA expression and steroidogenesis acute regulatory (StAR) protein were significantly increased at 5 wk and 12 wk, respectively. This study suggests that prenatal exposure to DE has detrimental effects on mouse spermatogenesis in offspring.

In utero exposure to diesel exhaust increased accessory reproductive gland weight and serum testosterone concentration in male mice.

The effects of in utero exposure to diesel exhaust (DE) on the male mouse reproductive system were examined. Pregnant ICR mice inhaled DE at soot concentrations of 0.3, 1.0, or 3.0 mg DEP/m3 or clean air as the control, for 2-16 days postcoitum. On postnatal day (PND) 28, the weights of the testes and accessory glands and testosterone concentration in serum were significantly higher in the DE-exposed male pups. Testosterone concentration correlated significantly (P<0.01) with the expression levels of steroidogenic enzyme mRNAs, weights of the testes and male reproductive accessory glands, and daily sperm production. These findings indicate that very early stage mouse embryo exposure to DE leads to endocrine disruption after birth and acceleration of male puberty.

Effect of diesel exhaust particles on mRNA expression of viral and bacterial receptors in rat lung epithelial L2 cells.

Epidemiological studies have shown that particulate matter (PM) is associated with adverse respiratory health effects. Although infection in the respiratory organ is one of the most important health risks the association of infection with PM is not fully understood. As we had hypothesized that diesel exhaust particles (DEP), one of the major component of PM, may induce the expression of receptors for viruses and bacteria at invasion sites, we studied the effect of DEP on the mRNA expression of intercellular adhesion molecule-1 (ICAM-1), low-density lipoprotein (LDL) and platelet-activating factor (PAF) receptors, which are invasion sites of virus and bacteria, on rat lung epithelial cells. The real-time quantitative polymerase chain reaction (PCR) method was used for the evaluation. All of these mRNAs were up-regulated by 3, 10, and 30 microg/ml of DEP in a concentration-dependent manner. The up-regulation of each was associated with the mRNA expression of heme oxygenase-1 (HO-1), a marker of oxidative stress. Our present results show that DEP up-regulated the mRNA expression of viral and bacterial receptors. This up-regulation might be associated with DEP-induced oxidative stress. These results thus suggest that DEP may enhance the risk of pneumonia by increasing the density of bacterial and viral invasion sites in the lungs.

Diesel exhaust affects immunological action in the placentas of mice.

We investigated the effect of diesel exhaust (DE) on pregnancy and fetal development in mice at day 14 postcoitum (pc) with a special focus on the placenta. The number of absorbed fetuses increased in groups exposed to DE, and congestion was observed in histological sections of placentas. During placental absorption expression of CYP1A1 mRNA decreased to undetectable levels, whereas expression of TNF alpha mRNA increased approximately twofold over that of the control. Levels of CYP1A1 mRNA in normal placentas from DE-exposed mice were unchanged. mRNA levels of inflammatory cytokines IL-2, IL-5, IL-12 alpha, IL-12 beta, and GM-CSF increased in placentas exposed to DE (0.3 and 3.0 mg diesel exhaust particles (DEP)/m3). Expression of IL-5 mRNA was markedly increased in DE-exposed placentas, although levels were barely detectable in control placentas. IL-6 mRNA expression was increased approximately 10-fold in placentas exposed to DE (3.0 mg DEP/m3). It has been reported that expression of mRNA encoding proteins involved in immune function in the placenta is increased during fetal absorption in mice. In the present study, expression of such mRNA by the placenta was increased by DE exposure. Because it is believed that expression of mRNA in the placenta also affects fetal development, DE may promote fetal absorption. These findings suggest that in mice exposure to DE affects fetal absorption and placental function by modifying expression of immune-related genes during early gestation and expression of endocrine-related genes during late gestation.

Endocrine-disrupting activity of chemicals in diesel exhaust and diesel exhaust particles.

Diesel exhaust (DE) is known as the main cause of air pollution. DE is a complex mixture of particulate and vapor-phase compounds. The soluble organic fraction of the particulate materials in DE contains thousands of compounds including a variety of polycyclic aromatic hydrocarbons and heavy metals. To clarify the endocrine-disrupting activities of DE, we have reviewed the reports about the effects of DE on the reproductive and brain-nervous systems, and the endocrine-disrupting action of diesel exhaust particles (DEP). In utero exposure to low levels (0.1 mg DEP/m3) of DE from day 2 postcoitum (p.c.) until day 13 p.c. reduced the expression level of Ad4BP/SF-1 mRNA and thereby might affect the development of gonads. Low levels of DE also reduced the expression of several genes known to play key roles in gonadal development, including an enzyme necessary for testosterone synthesis. Mature male rats exposed to DE during the fetal period showed an irreversible decrease in daily sperm production due to an insufficient number of Sertoli cells. DE exposure during the fetal period influenced the brain tissue in newborn mice. In the 3 mg DEP/m3 exposure group at 10 weeks of age, a significant reduction in performance was observed in the passive avoidance learning test in both male and female mice. In addition, the fetal exposure of mice to DE affected the emotional behaviors associated with the serotonergic and dopaminergic systems in the mouse brain. In toluidine blue-stained specimens from the DE-exposed group, edema around the vessels where fluorescent granular perithelial (FGP) cells exist and degenerated granules within the FGP cytoplasm were observed; similar findings were obtained by electron microscopic examination. DEP contain many substances that stimulate Ah receptors, such as the polycyclic aromatic hydrocarbon containing benzo[a]pyrene. DEP also contain substances with estrogenic, antiestrogenic and antiandrogenic activities. The neutral substance fraction of DEP has the causal substance that reduces estrogen receptor mRNA expression. Evaluating the influence of these chemicals present in the environment on human health is an important task.

Diesel exhaust affects the abnormal delivery in pregnant mice and the growth of their young.

To clarify the toxic effects of diesel exhaust (DE) on delivery in mice and on growth of young, C57Bl-strain females were exposed to 0.3, 1.0, or 3.0 mg diesel exhaust particles (DEP)/m3 or filtered clean air (control) for 4 mo (12 h/day, 7 days/wk). After exposure, some females from each group were examined by necropsy, and the remainders were mated with unexposed males. Estrous females for necropsy who had been exposed to 1.0 mg DEP/m3 had significantly lower uterine weights than the control estrous females. In the mated females, 9.1, 10.0, or 25.0% (0.3, 1.0, or 3.0 mg DEP/m3 of the pregnancies resulted in abnormal deliveries (abortion and unable delivery), but this was not significant. The rate of good nest construction by delivered females exposed to 3.0 mg DEP/m3 was significantly lower. Young were weighed at 11, 14, and 21 days, and weekly from wk 4 to 9 after birth. Body weights of male young of dams exposed to 1.0 or 3.0 mg DEP/m3 were significantly lower at 6 and 8 wk of age. Body weights of female young of dams exposed to 1.0 or 3.0 mg DEP/m3 were also significantly lower at 6, 8, and 9 wk. Vaginal orifices of young female mice whose dams were exposed to 0.3 and 1.0 mg DEP/m3 opened significantly earlier. The young were killed at 30 or 70 days during deep anesthesia, and their body weights, organ weights, and body lengths were measured. Anogenital distance (AGD) of 30-day-old males whose dams were exposed to 0.3 mg DEP/m3 was significantly shorter than that of the controls. Weights of thymus and ovary in 30-day-old females whose dams were exposed to 3.0 mg DEP/m3 were significantly lower. In 70-day-old males of dams exposed to 3.0 mg DEP/m3, body weights were significantly lower and AGD was significantly shorter. Weights of adrenals, testes, and seminal vesicles in 70-day-old males with dams exposed to 1.0 mg DEP/m3 were significantly lower. In 70-day-old females with dams exposed to DE, body weights in the 3.0-mg DEP/m3 group were significantly lower, and weights of adrenals, liver, and thymus in the 1.0-mg DEP/m3 group were significantly lower. Thymus weights in 70-day-old females with dams exposed to 0.3 mg DEP/m3 were significantly lower. Crown-rump length (CR) in 70-day-old females with dams exposed to 1.0 or 3.0 mg DEP/m3 was significantly shorter. These results show that toxic substances in DE might cause abnormal delivery in mice, and that exposed females affected the growth and sexual maturation of their young.