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Nitric oxide (NO) is one of the neurotransmitters in the cerebellum. Activities of NO synthase and densities of granule cells were determined in the cerebellum in an attempt to elucidate a mechanism of biochemical changes on the anatomical basis during aging. Activities of Ca2+/calmoduline dependent NO synthase were measured in the cerebellum in male and female mice, from 18 gestation days to 28 months of age. There were significant differences in NO synthase activities among the groups of prenatal (18 gestation days), middle aged (5-18 months) and old aged (21-28 months) mice. Between groups of old and middle ages, a significant decrease with aging was found in the mean density of granular cells, which express NO synthase. Changes in NO synthase activities and granule cell densities occurred to a similar degree between old and middle ages. The present data suggest that an age-associated rise and fall of its activities likely occur in parallel with those of granule cell densities, and thus may represent impairment of cerebellar function during aging.

High consumption of soybean products has been associated with a reduced risk of hormone-sensitive tumors. Soybean products contain phytoestrogens, such as daidzein, and sesame seeds contain secoisolariciresinol. These compounds are further metabolized to equol, enterodiol, and enterolactone by intestinal bacteria. However, individual differences in the metabolizing potential remain unclear. The aim of this study was to evaluate the urinary daidzein, equol, enterodiol, and enterolactone concentrations in women from several different regions of Japan according to age group. Five hundred urine samples collected from Japanese women living in Sapporo, Sendai, Kyoto, Kochi, and Naha were analyzed for daidzein, equol, enterodiol, and enterolactone concentration by gas chromatography–mass spectrometry. The urinary isoflavone and lignan polyphenol levels did not differ significantly among the sampling sites, except for daidzein, which was highest in urine collected at Naha. The prevalence of equol producers was 39 % in the total study cohort. In equol producers, a positive correlation was observed between the urinary daidzein and equol levels (r = 0.399, p

A gas chromatography-mass spectrometry and electron-capture negative ionization (ECNI) method was developed to quantify perfluorinated carboxylic acids (PFCAs) in composite dietary samples. Benzyl esterification was used for pretreatment before PFCAs analysis. This stabilized the benzyl radical leaving group preferentially, and gave carboxylic anions of the PFCAs with ECNI. The method had a low detection limit (0.3-10 pg g(-1)) and good recoveries (98-90%) for PFCAs with 8-14 carbon atoms (C8 to C14). The method was applied to 24-h dietary samples from subjects in Japan (Hokkaido, Kyoto, and Okinawa; 1992 to 2007, and 2009), Korea (Seoul; 1994 and 2007), and China (Beijing; 1993 and 2009). The levels of the PFCAs were between 39 and 169 ng day(-1) in Korea, 58 and 71 ng day(-1) in China, and 56 and 67 ng day(-1) in Japan. Between the two sampling years, the total levels of PFCAs (C8 to C14) increased significantly (p0.05). The interaction between the sampling location in Korea and year was significant (p0.05).

Objective: To characterize the phenotype of spinocerebellar ataxia type 36 (SCA36), a novel dominant disorder (nicknamed “Asidan”) caused by a hexanucleotide GGCCTG repeat expansion in intron 1 of the NOP56 gene. Methods: We investigated the clinical, genetic, and neuropathologic characteristics of 18 patients with SCA36. We performed histologic evaluation of a muscle biopsy specimen from 1 patient with SCA36, and neuropathologic evaluation of an autopsied brain from another patient with SCA36. Results: The (GGCCTG)n expansion was found in 18 ataxic patients from 9 families. The age at onset of ataxia was 53.1 ± 3.4 years, with the most frequent symptoms being truncal ataxia (100% of patients), ataxic dysarthria (100%), limb ataxia (93%), and hyperreflexia (79%). Tongue fasciculation and subsequent atrophy were found in 71% of cases, particularly in those of long duration. Skeletal muscle fasciculation and atrophy of the limbs and trunk were found in 57% of cases. Lower motor involvement was confirmed by EMG and muscle biopsy. The neuropathologic study revealed significant cerebellar Purkinje cell degeneration with obvious loss of lower motor neurons. Immunohistochemical analysis showed that NOP56 was localized to the nuclei of various neurons. Cytoplasmic or intranuclear inclusion staining of NOP56, TDP-43, and ataxin-2 was not observed in the remaining neurons. Conclusions: This is the first description of the unique clinical features of SCA36, a relatively pure cerebellar ataxia with progressive motor neuron involvement. Thus, SCA36 is a disease that stands at the crossroads of SCA and motor neuron disease.

Moyamoya disease is an idiopathic vascular disorder of the intracranial arteries. Ring finger 213 (RNF213) was previously identified as the strongest susceptibility gene for moyamoya disease in East Asian people by a genome-wide linkage analysis and exome analysis. The coding variant p.R4810K in RNF213 was strongly associated with moyamoya disease in the Japanese (odds ratio: 338.94, p = 1.05 × 10(-100)) and Korean (odds ratio: 135.63, p = 7.59 × 10(-27)) populations, and much less strongly associated in the Chinese population (odds ratio: 14.70, p = 2.63 × 10(-5)). The present study investigated the distribution of variant p.R4810K in RNF213 in 2,508 participants from East and Southeast Asian countries using a TaqMan probe. p.R4810K was detected at an allele frequency of about 1.00% in 4 of 11 investigated locations in China. In contrast, p.R4810K was detected homogeneously at relatively high frequencies of 1.00-1.72% in all investigated locations in Korea and Japan, including Okinawa. p.R4810K was not detected in Southeast Asian populations. The population susceptible to moyamoya disease was estimated to be 16.16 million people in East Asian countries, including 11.41 million Chinese, 1.36 million Korean, and 3.39 million Japanese people. The number of patients with moyamoya disease, which was estimated at approximately one per 300 carriers of p.R4810K, was considered to be 53,800 in East Asian populations.

The fluorotelomer alcohols (FTOHs) have been detected in various environmental compartments, including indoor and outdoor air, in North America and Europe. In our previous studies, FTOHs were detected at a relative higher concentration in outdoor air in the Keihan (Kyoto–Osaka, one of the major industrial zones) area, Japan compared to reported data. The exposure level of FTOHs in indoor air in the Keihan area remains unclear. In the present study, indoor air FTOH concentrations were investigated using a passive air sampler containing activated carbon felts. The indoor air sampling was conducted in 49 households of the Keihan area, during winter and summer 2008. Most samples contained 6:2 FTOH, 8:2 FTOH, 10:2 FTOH and 8:2 FTOAc. The median concentration of 8:2 FTOH (5.84 ng m −3 ) was highest among fluorotelomers, followed by those of 10:2 FTOH (1.12 ng m −3 ), 6:2 FTOH (0.29 ng m −3 ), and others. Significant correlations among fluorotelomers were observed in collected samples. The association between housing conditions and 8:2 FTOH concentrations showed that samples collected from bed rooms have higher 8:2 FTOH concentrations than those collected from other locations. In addition, samples collected in winter showed lower levels of 8:2 FTOH than those collected in summer. These findings suggest that 8:2 FTOH is the predominant component among fluorotelomers in indoor air, and that there are emission sources of fluorotelomers in indoor environments of the Keihan area. Further investigations into the origins of fluorotelomers are needed to evaluate indoor contamination with fluorotelomers.

In this first 2012 issue of Environmental Health and Preventive Medicine (EHPM), I regrettably think back to the tragedy of the earthquake in north-east Japan and the Fukushima Daiichi Nuclear Power Plant accident. As president of our society, Dr. Hiroshi Sato issued a public statement representing the view of the board members that we had not been creative enough in science. Yes, creativity is important. But why public remark on ‘deficiency’ in creativity? I finally came up with my own answer: deficiency in generosity, curiosity and modesty. Deficiency in any of these three basic characters results in a vicious cycle that results in a deficiency in the other two characters. The authorities are not generous, curious and modest enough to listen to minority opinions because they are concerned about making concessions. I am now fully aware that many qualified scientists had warned of possible dangers prior to the earthquake. These warnings, however, were deliberately ignored by scientific authorities. EHPM is rapidly becoming one of the major journals in the area of public health in Asian countries. Not only established and influential scientists but also young, upcoming leaders in various scientific fields are becoming members of the EHPM editorial board. To further foster the global development of EHPM, the Editorial Board has decided to add a section entitled “International Forum” to each issue of the journal. The aim of the forum is to expose readers to a wide diversity of opinions. Each forum article will be written by one or more of the members of the International Editorial Board. The first forum article is contributed by myself, Dr. Kyungho Choi and other Asian colleagues. The article reports on vividly evident current environmental health problems in Asian countries and raises several important agendas for the next 30 years. Therefore, we strongly believe that it is a worthy first article for the “International Forum”.

OBJECTIVE: This study aims to estimate the ecological exposure of adult residents of Fukushima Prefecture to (134)cesium (Cs) and (137)Cs through ingestion and inhalation between July 2 and July 8, 2011. METHODS: Fifty-five sets of meals with tap water, each representing one person's daily intake, were purchased in local towns in Fukushima Prefecture. Locally produced cow's milk (21 samples) and vegetables (43 samples) were also purchased. In parallel, air sampling was conducted at 12 different sites using a high-volume sampler. Nineteen sets of control meals were collected in Kyoto in July 2011. (134)Cs and (137)Cs levels in the samples were measured using a germanium detector. RESULTS: Radioactivity was detected in 36 of the 55 sample meals from Fukushima, compared with one of 19 controls from Kyoto. The median estimated dose level (μSv/year) was 3.0, ranging from not detectable to 83.1. None of the cow's milk (21) or vegetable (49) samples showed levels of contamination above the current recommended limits (Bq/kg) of 200 for milk and 500 for vegetables. The total effective dose levels by inhalation were estimated to be, 福島県成人住民の放射性セシウムへの経口、吸入曝露の先行評価を実施. 京都大学プレスリリース. 2011-11-14.

This study focuses on the regional trends and possible sources of brominated organic contaminants accumulated in breast milk from mothers in southeastern (Okinawa) and northwestern (Hokkaido) areas of Japan. For persistent brominated flame retardants, polybrominated diphenyl ethers (PBDEs; major components, BDE-47 and BDE-153) were distributed at higher levels in mothers from Okinawa (mean, 2.1 ng/g lipid), while hexabromobenzene (HeBB) and its metabolite 1,2,4,5-tetrabromobenzene were more abundantly detected in mothers from Hokkaido (0.86 and 2.6 ng/g lipid), suggesting that there are regional differences in their exposure in Japan. We also detected naturally produced brominated compounds, one of which was identified as 2′-methoxy-2,3′,4,5′-tetrabromodiphenyl ether (2′-MeO-BDE68) at higher levels in mothers from Okinawa (0.39 ng/g lipid), while the other was identified as 3,3′,4,4′-tetrabromo-5,5′-dichloro-2,2′-dimethyl-1,1′-bipyrrole in mothers from Hokkaido (0.45 ng/g lipid). The regional variation may be caused by source differences, i.e. southern seafood for MeO-PBDEs and northern biota for halogenated bipyrroles in the Japanese coastal water.

Short-chain chlorinated paraffins (SCCPs) persist in the environment and bioaccumulate in biota and are under review by the Stockholm Convention on persistent organic pollutants. SCCP levels were measured semiquantitatively in pooled 24 h food composite samples from Chinese (n = 10), Korean (n = 10), and Japanese (n = 40) adults in the 1990 s and 2007-2009. In Japan, SCCPs were detected in 14 of 20 pooled samples in the 1990 s and 13 of 20 pooled samples in 2009. Between these two time points, the geometric mean (GM) of the dietary intake of total SCCPs per body weight was comparable in Japan (54 ng kg-bw(-1) day(-1) in the 1990 s and 54 ng kg-bw(-1) day(-1) in the 2000s). In Beijing, SCCP levels were elevated by 2 orders of magnitude from 1993 to 2009 (GM: 620 ng kg-bw(-1) day(-1) in 2009). The 95th percentile estimate of the dietary intake was 1200 ng kg-bw(-1) day(-1) (1% of tolerable daily intake). In Seoul, no samples in 1994 contained detectable SCCP levels and only one sample in 2007 showed trace levels of SCCPs. Preliminary evidence on the significant increase in SCCP exposure in Beijing in 2009 warrants urgent investigations to refine dietary intake estimates by targeting food types and source identification.

A Commentary on Identification of an autosomal dominant locus for intracranial aneurysm through a model-based family collection in a geographically limited area

In the North Central Province of Sri Lanka, chronic kidney disease of uncertain etiology (CKDue) has increased markedly over the past 15-20 years.From around 4,700 patients who were followed up, 106 affected patients who visited two local clinics in the endemic area for CKDue on August 10, 2009 and 10 pedigrees of 10 of these cases with familial clustering of CKDue participated in this study. Urine samples, collected from affected patients (n = 106), unaffected relative controls (n = 81), and Japanese controls (n = 50), were analyzed for two tubular markers: α1-microglobulin and N-acetyl-β-D: -glucosaminidase. Urine samples from patients with CKDue stages 1-4 (n = 101) and all the samples from unaffected relatives and Japanese controls were analyzed for urinary cadmium concentration.Urinary excretion of α1-microglobulin was elevated even in the earliest stage of CKDue compared with its levels in unaffected relative controls. Urinary excretion of N-acetyl-β-D: -glucosaminidase was elevated only in stage 5. In contrast, urinary cadmium excretion was similar in CKDue patients and in the unaffected relative controls, and levels in both these groups were significantly lower than the level in the Japanese controls. All levels were below the threshold level for renal toxicity, indicating the absence of any evidence of cadmium toxicity.The present study indicates that renal tubular damage occurs in the very early stage of CKDue and demonstrates the existence of familial clustering, suggesting that CKDue is likely to be the outcome of exposure to an unknown nephrotoxin in susceptible subjects in the endemic region.

Autosomal-dominant spinocerebellar ataxias (SCAs) are a heterogeneous group of neurodegenerative disorders. In this study, we performed genetic analysis of a unique form of SCA (SCA36) that is accompanied by motor neuron involvement. Genome-wide linkage analysis and subsequent fine mapping for three unrelated Japanese families in a cohort of SCA cases, in whom molecular diagnosis had never been performed, mapped the disease locus to the region of a 1.8 Mb stretch (LOD score of 4.60) on 20p13 (D20S906-D20S193) harboring 37 genes with definitive open reading frames. We sequenced 33 of these and observed a large expansion of an intronic GGCCTG hexanucleotide repeat in NOP56 and an unregistered missense variant (Phe265Leu) in C20orf194, but we found no mutations in PDYN and TGM6. The expansion showed complete segregation with the SCA phenotype in family studies, whereas Phe265Leu in C20orf194 did not. Screening of the expansions in the SCA cohort cases revealed four additional occurrences, but none were revealed in the cohort of 27 Alzheimer disease cases, 154 amyotrophic lateral sclerosis cases, or 300 controls. In total, nine unrelated cases were found in 251 cohort SCA patients (3.6%). A founder haplotype was confirmed in these cases. RNA foci formation was detected in lymphoblastoid cells from affected subjects by fluorescence insitu hybridization. Double staining and gel-shift assay showed that (GGCCUG)n binds the RNA-binding protein SRSF2 but that (CUG)(6) does not. In addition, transcription of MIR1292, a neighboring miRNA, was significantly decreased in lymphoblastoid cells of SCA patients. Our finding suggests that SCA36 is caused by hexanucleotide repeat expansions through RNA gain of function., 二つの難病（脊髄小脳変性症および筋萎縮性側索硬化症）の症状を示す新しい遺伝性神経変性疾患とその原因遺伝子を発見. 京都大学プレスリリース. 2011-06-17.

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.

DNA methylation is known to change with age in several mammalian species. Here we have examined the effect of dietary energy restriction on this age-associated change in liver DNA of C3H/SHN mice. The total 5-methyldeoxycytidine level in the genome decreased slightly soon after energy restriction started. The effect, however, diminished with time and no appreciable difference was detected at middle and old ages. The degree of methylation at the c-myc gene, on the other hand, was not affected by energy restriction at early periods, but the age-dependent alterations at later ages were repressed. This is a new finding to show that DNA methylation is one of the molecular indices of aging affected by energy restriction. It suggests an importance of DNA methylation in the aging process.

We explored the effects of O,O,S-trimethyl phosphorothioate (OOS-TMP) on body temperatures in Fischer 344 female rats. The 7-day LD50 p.o. for Fischer 344 female rats was found to be 11.8 mg/kg. OOS-TMP induced long-lasting (more than 48 h) and extensive hypothermia at dosesor = 14 mg/kg at a typical laboratory temperature (22 degrees C) while it produced typical symptoms at 10 mg/kg without hypothermia. In contrast, pair-fed (to 20 mg/kg rats) rats (n = 4) did not become hypothermic, negating any role of hypophagia in OOS-TMP associated hypothermia. We next investigated the effects of housing temperatures on toxicities at a LD50 dose (12 mg/kg). At 30 degrees C (n = 11) and 22 degrees C (n = 13), rats did not have hypothermic bouts but at 15 degrees C, eight out of ten rats had. Evidence that changes of housing temperatures neither modified clinical symptoms nor changed mortality rates discards a possibility of hypothermia being involved in delayed toxicity. A novel result of the present study suggests that thermoregulation may be heavily impaired by a special class of organophosphorus compounds.

In the present study, using a large automated health records database, we investigated the incidence of cardio-cerebrovascular events, diabetes new-onset events, and dialysis initiation events in hypertensive patients, and examined the effects of antihypertensive medications on these incidences. We conducted a search of an automated health records database that contained anonymous information from the health insurance claims and the results of laboratory tests at 15 medical facilities across Japan. The study cohort was defined as patients who were diagnosed with hypertension and who visited a medical institution in the registration period. Events were defined by diagnosis, medication history, and laboratory test results. We obtained a cohort of 20,686 patients diagnosed with hypertension. The mean (standard deviation, SD) age in the cohort was 67.9 (13.2) years, and the follow-up period was 2.56 (1.42) years. The total incidence rates per 1,000 person-years in the present study population showed good agreement with rates in reported cohort studies: 8.10 (5.6–11.1) for cerebrovascular events, 1.27 (0.5–7.4) for cerebral hemorrhage, 6.57 (4.6–8.9) for cerebral infarction, 0.46 (0.1–1.0) for subarachnoid hemorrhage, and 1.75 (1.6–4.4) for myocardial infarction. The standardized incidence rates of cardio-cerebrovascular events, diabetes new-onset events, and dialysis initiation events were 9.73, 20.94, and 5.99 events/1,000 person-years, respectively. In terms of the incidence of the investigated events in hypertensive patients, the study results suggested that the automated health records database data were as valid and reliable as data from other epidemiological studies.

A publicly available atmospheric transport model, the Weather Research and Forecasting Chemistry Model ( http://ruc.noaa.gov/wrf/WG11/ ), was used to simulate atmospheric perfluorooctanoic acid (PFOA) and perfluorooctanoate (PFO) emitted from a point source in the Osaka urban area (also known as Keihanshin), Japan. The time period of the simulation was from 1983 to 2008. The modeled air concentrations were highly correlated (r = 0.91) with the observed air concentrations. Intake levels by inhalation of simulated air concentrations and through the gastrointestinal tract as estimated by the food duplicate method were input to a pharmacokinetic model of the human body to simulate serum concentrations of PFOA and PFO (PFO(A)). For validation of the atmospheric model, simulated values were compared with those observed in serum samples. The simulated values generally agreed with those observed in serum samples from residents of the Keihanshin area (r = 0.93). It was confirmed that the atmospheric model was generally capable of projecting features of atmospheric PFO(A) as well as serum concentrations of PFO(A) in this case. The results indicated a dominant contribution of the atmospheric component to serum PFO(A) in humans near the point source in the Keihanshin area. In 2008, that contribution was about 70%.

A Commentary on genome-wide association study to identify genetic variants present in Japanese patients harboring intracranial aneurysms

Background and Purpose— Genetic factors are important determinants of intracranial aneurysm (IA). Recently, a multinational, genome-wide association study identified 3 loci associated with IA, located on 2q (rs700651), 8q (rs10958409), and 9p (rs1333040 and rs10757278). The aim of this study was to evaluate these associations. Methods— Familial and sporadic cases were investigated. Familial cases, consisting of 96 subjects with IA, and 46 subjects of unknown status from 31 pedigrees were analyzed with the transmission disequilibrium test and linkage analysis. Associations of single-nucleotide polymorphisms (SNPs) with IA were tested in 419 sporadic IA cases and in 408 control subjects. Sequencing of CDKN2A , CDKN2B , and CDKN2BAS revealed additional SNPs, and their associations with IA were also tested. Results— The transmission disequilibrium test revealed associations of 2 SNPs, rs700651 ( P =0.036) and rs1333040 ( P =0.002), with familial IA. Analysis of SNPs in sporadic cases revealed an allelic association of rs1333040 with IA (odds ratio=1.28; 95% CI, 1.04–1.57; P =0.02) but failed to show associations of rs10757278 and rs496892 with IA. We sequenced 3 candidate genes; CDKN2A , CDKN2B , and CDKN2BAS . All 6 index cases from IA families had the rs1333040-T allele and SNPs (rs10965215, rs10120688, and rs7341791) in CDKN2BAS . None of these SNPs had linkage disequilibrium with rs1333040 and was associated with IA. Conclusions— A region between introns 7 and 15 of CDKN2BAS carrying the rs1333040-T allele may confer risk for IA.

A novel mutation of insulin receptor gene (INSR gene) was identified in a three generation family with phenotypical variety. Proband was a 12-year-old Japanese girl with type A insulin resistance. She showed diabetes mellitus with severe acanthosis nigricans and hyperinsulinemia without obesity. Using direct sequencing, a heterozygous nonsense mutation causing premature termination at amino acid 331 in the alpha subunit of INSR gene (R331X) was identified. Her father, 40 years old, was not obese but showed impaired glucose tolerance. Her paternal grandmother, 66 years old, has been suffered from diabetes mellitus for 15 years. Interestingly, they had the same mutation. One case of leprechaunism bearing homozygous mutation at codon 331 was identified. These findings led to the hypothesis that R331X may contribute to the variation of DM in the general population in Japan. An extensive search was done in 272 participants in a group medical examination that included 92 healthy cases of normoglycemia and 180 cases already diagnosed type 2 DM or detected hyperglycemia. The search, however, failed to detect any R331X mutation in this local population. In addition, the proband showed low level C-peptide/insulin molar ratio, indicating that this ratio is considered to be a useful index for identifying patients with genetic insulin resistance. In conclusion, a nonsense mutation causing premature termination after amino acid 331 in the alpha subunit of the insulin receptor was identified in Japanese diabetes patients. Further investigations are called for to address the molecular mechanism.

We reviewed 57 pediatric patients admitted with acute liver failure to Kyoto University Hospital in Japan over a period of 15 years to compare the etiology and the long-term outcome of infants and children after living donor liver transplantation (LDLT). Patients were divided into two groups according to age at the time of liver transplantation, infants group (1 year, n = 20), and children group (1-18 years, n = 37). The overall survival rates were 73.6%, 69.5% and 67.2% at 1, 5, and 10 years after LDLT respectively. Age of recipients at the time of LDLT had a strong impact on their outcome, Children had significantly better outcome than infants (P = 0.001). Surgical complications were comparable between both groups. Infants had higher rates of acute cellular rejection (ACR), which was associated with features of hepatitis in many cases. Refractory ACR was the leading cause of death in eight out of 12 infants, while it resulted in loss of one child only. Cox's proportional hazard regression model was used to examine potential risk factors for graft loss and it shows that age1 year was associated with high risk of graft loss [hazard ratio (HR) = 11.393; CI = 1.961-76.1763] (P0.05). In conclusion, Infants had poorer prognosis than children and refractory ACR was the leading cause of death. Using additional immunosuppressant for cases with severe and atypical rejections is recommended.

Genetic dissection of diseases is one of the epoch-making achievements in modern medicine. Positional cloning is a key method to isolate disease-related genes. For positional cloning, there are two conventional methods: family-based studies and case-control studies. In this review, I would like to describe several family-based studies on single gene diseases which I had conducted including those of Akita diabetic mice, systemic carnitine deficiency and Hartnup disease. The study of systemic carnitine deficiency underscored a potential power of the "Carrier state." Furthermore, cultural and public health practices in Japan such as preservation of umbilical cords and mother and child passbooks enabled us to conduct linkage analysis even 20 years after the deaths of affected patients in Hartnup disease. For multifactorial diseases, I present three family-based studies: intracranial aneurysm, moyamoya and arteriovenous malformation. Finally, I discuss on theoretical issues concerning the relationship among odds ratio, phenocopy rate and penetrance by formulating a single-locus dominant association model. Analysis of the model predicted a notion that a large odds ratio facilitates familial clustering of multifactorial diseases and vice versa is the case. Furthermore, the analysis predicted that genetic markers for screening should have odds ratio >/= eight to maintain similar qualities commonly required for clinical tests. Collectively, the analysis predicted a two-stage study design composed of linkage analysis based on a family study and subsequent replication by a case-control association study is more rational than the currently used two-independent case-control design. This newly proposed method is expected to provide polymorphisms, which have large odds ratios, requiring only minimum research budgets.

Peroxisome proliferator-activated receptor-alpha (PPARalpha) has been suggested to protect against chemically induced hepatobiliary injuries in rodents. This function could mask the potential toxicities of perfluorooctanoic acid (PFOA) that is an emerging environmental contaminant and a weak ligand of PPARalpha. However its function has not been clarified. In this study, PFOA was found to elicit hepatocyte and bile duct injuries in Pparalpha-null mice after 4 wk treatment with PFOA ammonium salt (0, 12.5, 25, 50 micromol/kg/d, gavage). In wild-type mice, PFOA caused major hepatocellular damage dose-dependently and minor cholangiopathy observed only at 25 and 50 micromol/kg. In treated Pparalpha-null mice, PFOA produced marked fat accumulation, severe cholangiopathy, hepatocellular damage and apoptotic cells especially in bile ducts. Oxidative stress was also increased 4-fold at 50 micromol/kg and TNF-alpha mRNA was upregulated more than 3-fold at 25 micromol/kg in Pparalpha-null mice. Biliary bile acid/phospholipid ratios were higher in Pparalpha-null mice than in wild-type mice. Results from these studies suggest that PPARalpha is protective against PFOA and have a critical role in drug induced hepatobiliary injury.

Several studies have shown improved growth after liver transplantation, but long-term follow-up data have been lacking. This study was aimed at evaluating the ability of children to catch up in height after living donor liver transplantation (LDLT) and at clarifying factors affecting growth. Growth was assessed by serial height measurements performed during follow-up. Standardized height scores (z scores) were calculated for each patient preoperatively (at the baseline) and at 1, 2, 3, 5, 10, and 15 years after LDLT. The risk potential of several preoperative and postoperative variables was evaluated. A total of 237 patients, including 159 females (67.1%), met the inclusion criteria. The mean age at the time of transplant was 3.89 +/- 0.28 years. The mean z score was -1.70 +/- 0.09, whereas the baseline height deficit was -6.50 +/- 0.39 cm. After LDLT, the z score improved significantly and reached -0.64 +/- 0.14 by the end of the first year. The best height improvement was seen after 10 years (-0.33 +/- 0.16). However, significant growth retardation remained at 15 years (-0.47 +/- 0.17). Height showed 3 distinct phases after transplantation: a growth spurt, a plateau phase, and a late declining phase. Univariate and multivariate analyses showed that children under 2 years and those with the most growth retardation at the time of LDLT achieved the best height gain in the first year. Late growth retardation was related to the baseline z score, ABO-incompatible grafts, and graft dysfunction. In conclusion, children have the potential ability to catch up in growth to normal levels after LDLT; they can show impressive height gains in the first year followed by protracted improvement over 10 years and then late growth retardation. Young age is a determinant for early height gain, whereas ABO-incompatible grafts and graft dysfunction are determinants for late growth retardation. In contrast, the baseline z score is a determinant for both.

We monitored core body temperature by telemetry in energy-restricted (201 kJ/wk) and control (397 kJ/wk) C57BL/6 and SHN/C3H F1 mice to determine whether torpor may be involved in the suppression of mitotic activities resulting from energy restriction. The energy restriction regimen employed inhibited the development of cancer and greatly extended longevity in both these mouse strains. Male and female C57BL/6 mice subjected to energy restriction from 4 wk of age and tested at 3 mo of age became torporific (body temperature less than 31 degrees C) at ambient air temperatures of 20-22 degrees C, whereas control animals stayed euthermic (greater than 35 degrees C). Energy restriction also induced torpor in 3- and 13-mo-old SHN/C3H F1 female mice, whereas 3-, 13- and 24-mo-old control mice were euthermic. Energy restriction decreased mitotic activities to approximately 30% of control values in both jejunum and epidermis in 3-mo-old female C57BL/6 mice maintained at 20-22 degrees C. However, this suppression of mitotic activities was antagonized by housing the energy-restricted mice at 30 degrees C for 2 wk, indicating that torpor plays a substantial role in suppressing mitotic activities in energy-restricted mice.

KOIZUMI, A., TSUKADA, M. and KAMIYAMA, S. A Skin-Specific MMTV Provirus is Stably Transmitted by Horizontal Transmission. Tohoku J. Exp. Med., 1992, 167 (3), 171-180 -Mouse mammary tumor virus (MMTV) is a retrovirus with a life cycle that remains to be elucidated. In establishing Mtv-4 congenic mice, an extra copy of the MMTV provirus was found specifically in the skin of both sexes of SHN mice but not in C3H/He mice. This extra copy was not, however, found in other normal tissues such as brain, lung, liver, spleen, kidney, uterus, testis, and mammary glands from SHN or C3H/He mice. In the skin of SHN mice, an 11-kb DNA fragment, which approximates in size to the entire MMTV genome, was detected in undigested high-molecular weight DNA. This unique MMTV provirus was shown to be transmitted horizontally from SHN mice to exogenous-MMTV-free C3H/He mice through nursing. The virus is thus concluded to be exogenous. Clusters of MMTV type A particles were found in sebaceous glands by electronmicroscopic examination. The present findings indicate that exogenous MMTV may be Stably maintained in the skin as an unintegrated provirus during its life cycle.

Log-Normality of Air Contaminants and Its Hidden Characteristics Useful for Industrial Hygiene Technology: Akio KOIZUMI, et al. Department of Health and Environmental Sciences, School of Public Health, Kyoto University-It is well known that concentrations of contaminants in the air fluctuate log-normally, but there has been no mathematical model that can describe the stochastic nature of this. In the present study we constructed a stochastic model that can describe log-normal distribution of the air contaminantion in workplaces. The so-called Ito-process can describe the log-normality of the stochastic nature of the air contaminants by using two parameters: ventilation performance and random release of contaminants from the source. We also derived a theorem from the model. It indicates that splitting the source into small independent compartments decreases randomness and thereby reduces exposure intensity. The reduction of randomness of contaminant release is considered to be as effective as improvement of ventilation performance in reducing workers'' exposure.

The aim of this work is to investigate the association between transfusion history and cancer mortality in a prospective follow-up study. We conducted a prospective cohort study in four areas of Akita Prefecture, Japan, in 10,451 individuals (4,401 men and 6,050 women, aged 40–79 years) without history of cancer. The subjects were followed until 31 December 2003 and the number of deaths from cancer was recorded. After mean follow-up of 12.76 years (140,259 person–years), 520 individuals (333 men and 187 women) died of cancer. History of blood transfusion before 1990 was mildly but significantly associated with overall cancer mortality (hazard ratio = 1.75, 95% confidence interval: 1.32–2.18) and nonliver cancer mortality (HR = 1.68, 95% CI: 1.25–2.26). This significant association remained unchanged after excluding deaths that occurred within 5 years of baseline for overall cancer mortality (HR = 1.47, 95% CI: 1.04–2.09) and for nonliver cancer mortality (HR = 1.43, 95% CI: 1.00–2.04). The significant association for nonliver cancer mortality was confirmed in subjects with no smoking history and/or alcohol consumption (HR = 2.01, 95% CI: 1.35–3.00). Site-specific analysis showed a possible association between transfusion history and death from pancreatic cancer. History of blood transfusion before 1990 was found to be associated with increased risk for cancer mortality and was independent of liver diseases. The mechanism of the association between blood transfusion and cancer mortality warrants further research.

As a result of the patient efforts of ex-editor-in-chief Prof. H. Yuasa and the strong support of ex-president Prof. K. Morimoto, our journal Environmental Health and Preventive Medicine (EHPM) is currently being published by one of the more well-known international publishers, Springer Verlag. This decision to form a formal working relationship with Springer Verlag was undertaken with the aim of promoting the scientific values of our journal, EHPM, in the international scientific community. To improve our desire for internationalization, our Society, with the full support of the elected president of the Japanese Society for Hygiene, Prof. H. Sato, has opened a China office, which is chaired by Prof. Peiyu Wang, Peking University. EHPM is about to enter a new phase in its history. We, A. Koizumi (Kyoto University) and T. Otsuki (Kawasaki Medical School) will succeed to the position of editor-in-chief of EHPM and Nippon Eiseigaku Zasshi, respectively. Other new members of the Editorial Board are Prof. K. Aoki (Ryukyu University), Prof. M. Ichiba (Saga University), Prof. H. Inadera (Toyama University), Prof. K. Ogino (Okayama University), Dr. Y. Saijo (Asahikawa Medical College), Prof. T. Takeuchi (Kagoshima University), Prof. T. Nakayama (Kyoto University), Dr. Y. Nishiwaki (Keio University), Dr. K. Harada (Kyoto University), and Prof. C. Watanabe (University of Tokyo). We will do our best to develop as a team and to extend the group dynamics of our society by exploring new horizons for EHPM and Nippon Eiseigaku Zasshi. Japanese universities are increasingly being confronted with a heavy economic burden in their desire to purchase journals, particularly since the numbers of journals are increasing: PubMed central listed more than 5000 journals as of April 2009. Nature, Science, Cell, New England Journal of Medicine, Lancet and JAMA, which are very prestigious journals, often reject even excellent papers due to the limitation of space and scope of the journals. These journals have played key roles in reporting new findings to a broad readership. The growth of science has been accompanied by a growth in the number of journals. Journals compete with each other and are ranked by impact factors; scientists are evaluated by cumulative impact factors and cumulative numbers of cited works. To a large extent, the status of a journal is almost always a reflection of the quality of the papers it publishes and the impact factors. Consequently, the promotion of those studies published in a journal is in proportion to the reputation of that journal. There are, however, a number of caveats. Journals cost a lot of money to maintain. There is also always a page limitation as even a large, well-known journal, such as JBC, cannot exceed 40,000 pages annually. To cope with such limitations, a new online journal, PLoS (Public Library of Science), has been been created. This journal is published by NPO, which was founded by Dr. Harold Vermus and his colleagues. It has several innovative policies: (1) open access; (2) editorial performance at every level is transparent and accessible to the scientific community and public; (3) it addresses educational value; (4) it is committed to a fair, rigorous editorial process; (5) scientific quality and importance are the sole considerations in publication decisions without pragmatic considerations focusing on publishing high-impact research; (6) it intends to expand its scope as rapidly as practically possible to provide a vehicle for publication of other valuable scientific or scholarly articles. EHPM and Nippon Eiseigaku Zasshi may learn a number of core values from PLoS. Policies (3), (5) and (6) are of particular importance. In terms of policies (3) and (5), it may be generally difficult to evaluate the scientific quality and importance of an article and also to maintain a good balance between educational function and scientific quality. Policy (6) is also essential: good journals publish large-scale studies which are supported by large grants, with less attention given to cottage industry science. Cottage industry scientists are increasingly having the feeling that it is difficult to find the appropriate journals for publishing their studies, even when they are scientifically sound. Due to complex factors, including time trends, and cultural, social, and economic backgrounds, we cannot simply transplant PLoS policies into EHPM and Nippon Eiseigaku Zasshi. However, we may learn something from these principles. If a study has a sound scientific basis and the article is well written, it may stimulate other studies in a self-directive way. In modern societies such as ours, we should be sensitive to ethical aspects as professional scientists. Finally, since EHPM and Nippon Eiseigaku Zasshi is the official journal of the Japanese Society for Hygiene, it has to be a scientific vehicle for all members. The new editorial group, including those of the China office, would like to address the following core points for a new editorial policy: Scientific formalism: All submitted papers should be based on sound science. We address the scientific formalism of the paper as much as its quality and importance and more than its impact. Compliance of ethical guidelines for scientists and research: Authors should not follow the standard ethical codes for study participants but ensure that the study also conforms with research ethics. Educational function: The principle of “peer review” originally meant mutual learning. However, reviewers often tend to have the illusion of superiority or of being a “big cheese”. Such an illusion, if any, should be completely discarded, and the mutual learning principle of “peer review” should be recalled. This step will encourage young researchers to submit challenging works to EHPM and Nippon Eiseigaku Zasshi. EHPM and Nippon Eiseigaku Zasshi: Public health issues are often linked to the social system and their traditional backgrounds. EHPM is more oriented to international issues while Eiseigaku Zasshi is more oriented to domestic issues. In addition, since Nippon Eiseigaku Zasshi is written in Japanese, it is more reader friendly for Japanese scientists than EHPM. As such, Nippon Eiseigaku Zasshi may play a more important role in terms of educational function than EHPM. We, however, should never forget the value of Nippon Eiseigaku Zasshi, since it has been cited by PubMed for many years. Both journals will be promoted equally. We believe that those new policies will facilitate the rapid growth of EHPM and Nippon Eiseigaku Zasshi.

Exposure to polychlorinated biphenyls (PCBs) is considered to have culminated between 1950 and 1970 in Japan, and exposure through diet, the major exposure route, has decreased significantly over the last 10 years. The primary goal of the present study was to investigate the long-term trends and congener profiles of serum and dietary levels of PCBs using historical samples.Using banked samples collected in 1980, 1995, and 2003 surveys, we determined the daily intakes and serum concentrations of 13 PCB congeners (#74, #99, #118, #138, #146, #153, #156, #163, #164, #170, #180, #182, and #187) in women.The total daily PCB intake [ng/day, geometric mean (geometric standard deviation)] decreased significantly from 523 (2.5) in 1980 to 63 (3.2) in 2003. The serum total PCB level (ng/g lipid) in women40 years of age decreased significantly from 185 (1.8) in 1980 to 68 (1.8) in 2003. In contrast, the level in women50 years of age increased significantly from 125 (1.7) in 1980 to 242 (1.7) in 2003. Specifically, the serum concentrations of hexa (#138, #146, #153, #156, #163, and #164) and hepta (#170, #180, #182, and #187) congeners increased significantly. A comparison of the serum PCB levels of women born from 1940 to 1953 revealed that their serum total PCB level was significantly higher in the 2003 survey [242 (1.7), n = 9] than in the 1995 [128 (2.0), n = 17] surveys. This increase in the total PCB level was attributable to increases in the hepta congener groups.Present results suggest a decreased rate of elimination of hepta congeners with aging in females, rather than a birth-generation phenomenon.

Human breast milk samples collected in 2007–2008 from four countries, Vietnam (Hanoi), China (Beijing), Korea (Seoul) and Japan (Sendai, Kyoto and Takayama), were analyzed for persistent organic pollutants (POPs) such as dichlorodiphenyltrichloroethane and its metabolites (DDTs), chlordane-related compounds (CHLs), hexachlorocyclohexanes (HCHs), hexachlorobenzene (HCB), polychlorinated biphenyls (PCBs) and polybrominated diphenyl ethers (PBDEs). Comparing with previous surveys, the present study indicates that the DDTs in breast milk from China and Vietnam had gradually decreased during the last decade, but were still 5–10 times higher than those in other nations. The ratios of p,p′-DDE/p,p′-DDT and o,p′-DDT/p,p′-DDT were higher in Beijing than in the other countries, suggesting that there is less fresh intake of commercial DDT products and a possible exposure to dicofol in China. CHL and PCB levels were relatively higher in mothers from Japan, whereas β-HCH and HCB were more common in Chinese women. In Japan, it is suspected that mothers in the urban/coastal area (Sendai) were more continuously exposed to organochlorine pesticides (OCPs) than mothers in the rural/inland area (Takayama). In addition, OCP levels in primiparae were significantly higher than those in multiparae from Japan and Korea. These indicate that both parity and regional factors are major determinants of the levels of OCPs and PCBs in human milk. On the other hand, higher concentrations of PBDEs were observed in mothers' milk from Korea. The congener was dominated by BDE-47 (43–54%), followed by BDE-153 (23–33%) in all regions except for Beijing where BDE-28 (23%) was relatively abundant. In Japanese breast milk, regional and parity-dependent distributions were not observed for PBDEs. Among PBDE congeners, age-dependency was observed for BDE-153, which was negatively correlated (p

We investigated whether age-dependent reactivation of a repressed X-linked gene occurs. Subjects were female mice, carrying the X-autosomal translocation, T(X;16)16H (Searle's translocation). The mice were also heterozygous, for the X-linked gene coding for phosphoglycerate kinase (T16H pgk-1b/+ pgk-1a and pgk-1a was selectively repressed in these mice (McMahon and Monk, 1983). An electrophoretic method was applied to determine the PGK-1 allozyme patterns in blood, bone marrow, brain, gastrointestinal tract, liver, heart, spleen, and uterus (including tumor tissues when found). Samples were collected from mice of three different ages: 2 months (n = 4), 11 to 12 months (n = 10), or 18 to 21 months (n = 15). The lowest detection limit of the relative cellular population expressing the PGK-1A allozyme was found to be 2%, which was sensitive enough to detect the reported reactivation ratio (more than 10% of cells in a lower power microscopic field). We could not detect PGK-1A activity in any organ, including tumors in any age group, leading to the conclusion that reactivation of the repressed pgk-1a gene did not occur during aging.

Although it has long been known that energy restriction (ER) inhibits tumors and retards aging in rats and mice, its mode of action remains unknown. In rodents, ER alters the rate of age-related changes in physiological indices. Thus, it affects a broad array of age-sensitive parameters. However, present evidence does not indicate which parameters are primary contributors to the deceleration of aging. Compared to fasting or short-term underfeeding, little is known about the metabolic effects of long-term, life-prolonging ER. We thus investigated the effects of ER on hepatic enzyme activities, including drug-metabolizing enzymes and antioxidant enzymes such as superoxide dismutase and catalase. The catalase activity was found to be higher in ER mice than in control mice both at 12 and 24 months of age. In accord with the high catalase activity, lipid peroxidation in liver was much less in ER mice than in age matched control mice. beta-Naphthoflavone, known to induce P-450 related enzymes and xanthine oxidase, was given (ip) to increase lipid peroxidation. The ER was found to inhibit lipid peroxidation after beta-naphthoflavone treatment. It was, therefore, concluded that long-term life-prolonging ER increases antioxidant defence, supporting indirectly the free radical theory of aging. It is well known that ER delays puberty in rodents and has a profound influence on serum hormone levels, including those of prolactin (PRL) and thyroid hormones. However, it remains unknown how these effects are produced by ER. We therefore investigated the effects of ER on the islets of Langerhans in the pancreas and on the pituitary-ovarian axis. In the islets of Langerhans, ER was found to increase the density of alpha-cells significantly both in 11- and 67-week-old mice. In the pituitary gland in ER mice, the cellular density of PRL-producing cells diminished significantly while that of growth-hormone-producing cells did not. One of the modes of action of ER on the endocrine system is thus concluded to be mediated by changing cellular population. Since ER decreased PRL-producing cells and PRL plays a key role in mammary tumorigenesis, we investigated whether ER decreased the gene expression of mouse mammary tumor virus (MMTV) in SHN/C3H mice. The SHN strain, which was found to have a new MMTV provirus locus, mtv-4, was used in the study.(ABSTRACT TRUNCATED AT 400 WORDS)

Moyamoya disease (MMD) is characterized by progressive stenosis and occlusion of the terminal portion of the bilateral carotid arteries as well as arterial collateral vessels. The etiology of MMD, however, remains unknown. Several pieces of evidence suggest the involvement of genetic factors in MMD: over 10% of MMD patients have affected blood relatives; concordance in the affection status has been proven in 80% of identical twins; and there is an ethnic predisposition to MMD, the incidence of the disease being the highest in the Asian population. With regard to genetic factor (s), transmission of MMD does not follow the classic Mendelian law, i.e., skipping of a generation and discordance in identical twins, thereby indicating that genetic influence is likely to determine the susceptibility to MMD. This study aimed to overview the recent findings related to the genetic determinants in MMD and to provide research perspectives for the next decade. Pathophysiological investigations at molecular levels have uncovered the upregulation of various growth and stress response factors that are associated with angiogenesis in occlusive cerebral arteries.

Perfluorooctane sulfonate (PFOS) is found ubiquitously in the environment, and is known to cause developmental toxicity, including cleft plate (CP). The aim of the present study was to elucidate the mechanism of CP associated with in utero exposure to PFOS in mice. We first examined whether the concentration of PFOS in fetal serum was related to susceptibility to CP. We compared palatogenesis following the administration of various concentrations of PFOS to dams. We conducted histological examination on gestational day (GD) 15 and 18, and alizarin red/alcian blue staining of fetal heads on GD18. Finally, we cultured palatal shelves (PSs) of GD14 fetuses, which had not yet made contact with each other, for 48h, to examine whether the shelves maintained the ability to fuse. The incidence of CP increased from 7.3% with a fetal serum concentration of PFOS of 110.7+/-13.4microg/ml (13mg/kg) to 78.3% with 138.6+/-0.9microg/ml (20mg/kg). PFOS at 50mg/kg on GD11-15 caused CP at a rate of 6.1%, meanwhile PFOS at 20mg/kg on GD1-17 caused a CP rate of 89.3%. Failure of palatal shelf elevation was observed with 20mg/kg PFOS. PFOS at 20mg/kg on GD1-17 and 50mg/kg on GD11-15 inhibited mandibular growth to the same extent, even though the rate of CP was different. Explants exposed to PFOS 20mg/kg and Tween 20 showed 94% (34/36) and 100% (31/31) fusion, respectively. We demonstrated that increasing the oral dose of PFOS from 13 to 20mg/kg resulted in a significant increase in CP even though there was only a small increase in serum concentration of PFOS. PFOS prevented elevation of the PSs above the tongue because their growth/fusion potential was maintained. Mandibular hypoplasia did not seem to play a critical role in the pathogenesis of CP.

Daily dietary intake of perfluorinated chemicals (PFCs) in relation to serum levels was assessed by determination of nine PFCs including perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) in matched daily diet duplicates and serum samples. Diet and serum were collected in year 2004 from 20 women in Osaka and Miyagi, Japan. Only PFOS and PFOA were detected in the diet samples and no significant difference between cities was seen. After adjusted by water content, diet concentration of PFOA was significantly higher in Osaka. The median daily intake calculated using the measured diet concentrations was 1.47 ng PFOS/kg b.w. and 1.28 ng PFOA/kg b.w. for Osaka, and 1.08 ng PFOS/kg b.w. and 0.72 ng PFOA/kg b.w. for Miyagi. A significant difference between cities was seen for the serum concentrations with median of 31 ng/mL PFOS and PFOA in Osaka, compared to 14 ng/mL PFOS and 4.6 ng/mL PFOA in Miyagi. Carboxylates such as perfluorononanoic acid (PFNA) and perfluoroundecanoic acid (PFUnDA) were also detected in serum at median levels 6.9 ng/mL and 3.2 ng/mL (Osaka), and 2.8 ng/mL and 5.1 ng/mL (Miyagi). Based on one-compartment model under steady state, dietary intake of PFOS and PFOA accounted for only 22.4% and 23.7% of serum levels in Osaka females, and in contrast 92.5% and 110.6% in Miyagi females, respectively. Keywords: Perfluorinated chemicals, Food duplicate, Dietary intake, Japan

Perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) comprise a class of per- and poly-fluorinated compounds that have been detected in the environment as well as in humans. The aim of this review is to summarize several monitoring studies in Japan and characterize the toxicokinetics of these compounds. We found that the levels of contamination by these compounds had unique patterns in Japan. The levels of PFOA in serum from inhabitants of the Kansai region were higher than those of other regions. The PFOA levels in air and water samples from the Kansai region were also relatively high. The estimated intakes from these routes partly explain the differences in the serum levels. The toxicokinetics of these compounds have been investigated. Serum samples from male participants had significantly higher geometric means for PFOS and PFOA compared to samples from female participants. This sex-related difference was partly simulated by menstrual blood loss. There are large interspecies differences in the excretion pathways of these compounds. The serum clearances of PFOA via urine were 300–1,000-fold lower in humans than in Wistar rats and Japanese macaques. On the other hand, the biliary excretion of these compounds was comparable in rats and humans, and the long half-lives in humans may be attributable to the low levels of urinary excretion and high biliary reabsorption rates. These findings suggest that qualitative differences in the excretion routes exist between humans and other species. For risk assessment of these compounds, further information regarding sources of exposure and their toxicokinetics is needed.

O,O,S-Trimethylphosphorothioate (OOS-TMP) has been shown to induce hypophagia and hypopraxia. Recent studies suggest that OOS-TMP-induced anorexia is partly mediated by its effect on the central nervous system. In this study, we examined the profiles of N-acylethanolamines (NEAs), including five amide-linked compounds, in the gastrointestinal system in C57BL/6J (B6) mice. The present results shown an orexigenic profile of the levels of NEAs with downregulation of the anorectic lipid, N-stearoylethanolamine (SEA), upregulation of the orexigenic lipid, 2-arachidonoyl glycerol (2-AG), at 2 h and upregulation of 2-AG at 24 h albeit with significant anorexia. However, the data indicated that the high level of 2-AG may be responsible for the hypopraxia. We next explored whether OOS-TMP may affect two models of hyperphagia and hyperglycemia, ins2+/Akita B6 (Akita) and B6-leprdb/leprdb mice (db/db). We identified potential anorexigenic effects in B6, Akita and db/db mice. Moreover, OOS-TMP was found to reduce blood glucose in Akita mice but not in db/db mice. Collectively, these findings suggest that N-acylethanolamines are not involved in the hypophagia but rather hypopraxia, and may play multiple physiological roles in this process. OOS-TMP might be a promising candidate for anti-obesity and anti-diabetic drug development.

Diabetic Akita male mice are more hyperphagic because of downregulation of proopiomelanocortin (POMC) caused by hypoleptinemia. We investigated the role of estrogen receptor alpha (ERalpha) in the regulation of the hypothalamic POMC in females. ERaKOAkt mice consumed 30% greater food (g/3 weeks) than the Akita diabetic controls. Ovariectomized diabetic (AFO) and nondiabetic (B6FO) mice had significantly lower food intake and elevated serum leptin levels. ERaKOAkt and ERaKO mice also increased serum leptin concentrations, while hypoinsulinemia was observed in ERaKOAkt and hyperinsulinemia in ERaKO mice. RT-PCR showed a significant attenuation of POMC expression in both ERaKOAkt and ERaKO mice, irrespective of the elevated leptin serum levels or hyperinsulinemia, while elevated serum leptin levels in AFO and B6FO mice upregulated POMC gene expression. These results indicate that ERalpha plays an essential role in leptin- and insulin-stimulated upregulation of the POMC gene. This action of ERalpha is likely mediated in a ligand-independent manner.

Perfluorooctanoic acid, an environmental contaminant, is found in both wild animals and human beings. There are large species and sex differences in the renal excretion of perfluorooctanoic acid. In the present study, we aimed to characterize organic anion transporters 1-3 (OAT1-3) in human beings and rats to investigate whether the species differences in the elimination kinetics of perfluorooctanoic acid from the kidneys can be attributed to differences in the affinities of these transporters for perfluorooctanoic acid. We used human (h) and rat (r) OAT transient expression cell systems and measured the [(14)C] perfluorooctanoic acid transport activities. Both human and rat OAT1 and OAT3 mediated perfluorooctanoic acid transport to similar degrees. Specifically, the kinetic parameters, K(m), were 48.0 +/- 6.4 microM for h OAT1; 51.0 +/- 12.0 microM for rOAT1; 49.1 +/- 21.4 microM for hOAT3 and 80.2 +/- 17.8 microM for rOAT3, respectively. These data indicate that both human and rat OAT1 and OAT3 have high affinities for perfluorooctanoic acid and that the species differences in its renal elimination are not attributable to affinity differences in these OATs between human beings and rats. In contrast, neither hOAT2 nor rOAT2 transported perfluorooctanoic acid. In conclusion, OAT1 and OAT3 mediated perfluorooctanoic acid transport in vitro, suggesting that these transporters also transport perfluorooctanoic acid through the basolateral membrane of proximal tubular cells in vivo in both human beings and rats. Neither human nor rat OAT2 mediated perfluorooctanoic acid transport. Collectively, the difference between the perfluorooctanoic acid half-lives in human beings and rats is not likely to be attributable to differences in the affinities of these transporters for perfluorooctanoic acid.

Perfluorinated compounds (PFCs) have been employed as surface treatment agents in a variety of products. Perfluorooctanoic acid (PFOA), a PFC that is found globally in the environment and in human tissues, has been increasing significantly in serum levels over the past 50 years. Here, we demonstrated that PFOA inhibits feeding behavior as potently as the endogenous peroxisome proliferator-activated receptor (PPAR)-alpha ligand, oleoylethanolamide (OEA), via the activation of PPAR-alpha, the vagal nerve and hypothalamic neuropeptides. Peripherally administered PFOA decreased food intake as potently as OEA. PFOA decreased gastric emptying and increased the expression level of the gene encoding urocortin 1 in the hypothalamus and the immunoreaction for urocortin 1 in the paraventricular nucleus. Vagotomy attenuated the inhibitory effects of PFOA on feeding. The inhibition of food intake and body-weight gain by PFOA was completely mitigated in PPAR-alpha-/-mice. Our studies demonstrated that the ubiquitous environmental pollutant PFOA works as an imitator of OEA mimicking its action in the feeding regulatory system, providing a new mode of action as represented by environmental 'anorexigens'.

Perfluorinated compounds (PFCs) have been employed as surface treatment agents in a variety of products. Perfluorooctanoic acid (PFOA), a PFC that is found globally in the environment and in human tissues, has been increasing significantly in serum levels over the past 50 years. Here, we demonstrated that PFOA inhibits feeding behavior as potently as the endogenous peroxisome proliferator-activated receptor (PPAR)-alpha ligand, oleoylethanolamide (OEA), via the activation of PPAR-alpha, the vagal nerve and hypothalamic neuropeptides. Peripherally administered PFOA decreased food intake as potently as OEA. PFOA decreased gastric emptying and increased the expression level of the gene encoding urocortin 1 in the hypothalamus and the immunoreaction for urocortin 1 in the paraventricular nucleus. Vagotomy attenuated the inhibitory effects of PFOA on feeding. The inhibition of food intake and body-weight gain by PFOA was completely mitigated in PPAR-alpha-/-mice. Our studies demonstrated that the ubiquitous environmental pollutant PFOA works as an imitator of OEA mimicking its action in the feeding regulatory system, providing a new mode of action as represented by environmental 'anorexigens'.

Fluorotelomer alcohols (FTOHs), a class of per- and poly-fluorochemicals (PFCs), are environmental contaminants characterized by high volatility and are therefore prone to long-range atmospheric transport. In this study, airborne FTOHs were identified in Japan using a newly developed passive air sampler containing activated carbon felts. Air sampling was conducted at 33 sites all over Japan from April to June 2007. 8:2 FTOH, the dominant analyte, ranged from32 to 2466pgm(-3) (mean: 241pgm(-3)) followed by 6:2 FTOH from n.d. to 768pgm(-3) (mean: 51.6pgm(-3)), 10:2 FTOH from17 to 113pgm(-3) (mean: 26.8pgm(-3)) and 8:2 FTOAcryl from n.d. to 480pgm(-3) (mean: 25.8pgm(-3)). 8:2 FTOMethacryl was not detected above detection limit (3pgm(-3)) in all samples. The highest concentration of total investigated PFCs was found in Chiba (35 degrees 43'20'' N 140 degrees 39'46'' E) (2152-3181pgm(-3)). On the other hand, in three locations of Higashi-Yodogawa (34 degrees 44' N 135 degrees 32' E), high levels of 8:2 FTOAcryl (239pgm(-3), 109-480pgm(-3)) were detected, the levels of which were comparable to those of 8:2 FTOH (mean: 175pgm(-3), range: 115-292pgm(-3)). This study is the first national wide-scale survey of FTOHs and 8:2 FTOAcryl across Japan, showing that 8:2 FTOH was ubiquitous in the environment in Japan.

We investigated the effects of energy restriction on the pituitary-ovarian axis and on a hormone responsive gene, the mouse mammary tumor virus (MMTV). Female C3H/SHN F1-hybrid mice, known to display a high incidence of mammary tumors, ate an energy-restricted diet (48 kcal/wk) or a control diet (95 kcal/wk) beginning at the time of weaning. By 67 wk of age, 12 of 32 mice in the control group, but none of the 33 mice in the energy-restricted group, had developed mammary tumors. Six tumor-free mice from each group were studied in detail at 67 wk of age. All six tumor-free control mice, but none of the six energy-restricted mice, showed uterine endometrial hyperplasia at autopsy. Mice subjected to energy restriction did not display an estrous cycle. The average levels of MMTV mRNA in mammary glands and uteri were strongly reduced by energy restriction. MMTV mRNA levels in mammary glands from control mice were two orders of magnitude lower than those in mammary tumors. Energy restriction lowered the percentage of pituitary mammatropes and suppressed proliferation of mammatropes with advancing age. Energy restriction thus appeared to inhibit endometrial hyperplasia and to decrease MMTV production at the mRNA level in the mammary glands and in the uterus. These effects may be a consequence of hormonal changes originating at the pituitary-ovarian axis.

The structure-pulmonary toxicity relationship was investigated using three different O,O,S-trimethylphosphorothioate (OOS-TMP) analogues, which is known to induce a typical mortality pattern (delayed death) and pulmonary injury; O,O-dimethyl S-ethyl (OOS-DMEP), O,O-dimethyl S-propyl (OOS-DMPP), and O,O-dimethyl S-butyl (OOS-DMBP) phosphorothioates. The mortality pattern in rats dosed with OOS-DMEP was similar to the "delayed death" pattern: the LD50s in rats for OOS-DMEP decreased dramatically from more than 200 mg/kg within 24 hr to 41.1 (males) or 13.8 (females) mg/kg 7 days after treatment while the LD50s in rats for OOS-DMPP and OOS-DMBP did not. Histopathological examinations revealed that OOS-DMEP induced pulmonary oedema and bleeding at a dose of 1/2 LD50 by 72 hr after dosing while the other two compounds did not. Thus, it was concluded that OOS-DMEP induces pulmonary injury, thereby eliciting the "delayed death" mortality pattern.

Nitroarenes, potent rodent carcinogens, have attracted much attention as environmental contaminants. Recently, simultaneous exposure to dust have been found to potentiate tumorigenicity of nitroarenes in rats (14). We hypothesized that inhabitants of northwestern Japan may be exposed to such substances in winter. First, we sought to develop a sensitive yet time saving method to determine nitroarene concentrations in airborne particulate matter. The method developed used a high-performance liquid chromatograph equipped with a fluorescence detector. By on-column catalytic reduction, using silica coated with platinum and rhodium, 1-nitropyrene (1-NP), 1,3-dinitropyrene, 1,6-dinitropyrene, and 1,8-dinitropyrene were effectively reduced to their amino derivatives which were easily detectable with the fluorescence detector. The lowest detectable limits found using the present method were on the order of picograms for individual compounds. Subsequently, we sought to determine the concentrations of nitroarenes using the new method on airborne particulate matter during the period from November to April. Samples were collected at sampling sites on road with high-volume air samplers for 24 hours or 72 hours in four different rural areas in Iwate Prefecture. At one location in the city of Morioka, samples were collected at the height of 25 m from the ground. Among the four nitropyrenes mentioned above, only 1-NP was detected in the samples. The concentrations of 1-NP and airborne particulates changed significantly in all examined areas in parallel with the rise and fall of the frequencies of studded tire use. The highest 1-NP and airborne particulate concentrations were 1.210 pg/m3 and 1,291 micrograms/m3, respectively, being recorded at the busiest road in Kitakami. In contrast, the lowest values were 38 to 66 pg/m3 for 1-NP and 54 to 94 micrograms/m3 for airborne particulates in Morioka, being recorded in November and April, respectively. The mutagenicities of extracts of airborne particulate matter showed similar trends. Thus, exposure situations with high concentrations of dust and 1-NP were confirmed to occur in Iwate. Finally, the risk posed by 1-NP in airborne particulate matter was estimated using the equation proposed by Cuddihy and McClellan (37) and was found to range from 3.12 x 10(-2) to 24.2 x 10(-2) lung cancer/year/10(6) people. Although the estimated risk due to 1-NP is relatively small, further surveillance is highly recommended in those areas since heavy dust exposure also occurs there.

Previous linkage analyses of intracranial aneurysm (IA) have proposed several genetic susceptibility loci; however, some loci remain contradictory. The objective of this study was to confirm these loci in a Japanese population using allelic and haplotype association analyses. We set high-density single nucleotide polymorphism markers in previously suggested IA loci and conducted an association analysis in 29 cases and 35 controls from a small community in Akita, Japan. Genotyping was carried out using the GeneChip 10 K mapping array, and the association analysis was performed using GeneSpring GT2 software. The result was confirmed in a replication cohort consisting of 237 cases and 253 controls from all over Japan. Only one variant, rs767603, at chromosome 14q23, was significantly associated with IA, both in allelic analysis (p = 0.00017, Bonferroni-corrected p = 0.021) and haplotype analysis (p = 0.00178, Bonferroni-corrected p = 0.048). This association was confirmed in the replication cohort (p = 0.0046 for allelic association, p = 0.0060 for haplotype association). Our findings confirm 14q23 to be a susceptibility locus for intracranial aneurysm.