[Research on Occupational Poisoning and Biological Monitoring].

OBJECTIVE: A new system for the regulation of chemical substances was introduced in Japan in April 2023. Unlike delineated rules, the new system does not specify any specific measures for individual substances, but is rather based on the management of health through the voluntary implementation of measures in order to reduce exposure to all dangerous/hazardous chemical substances. Regarding specific methods, exposures will be mainly elucidated through the measurement of a work environment. However, the necessity of biological monitoring should also be considered. This study aimed to identify the importance of biological monitoring by reviewing changes made over time in biological monitoring methods used for occupational poisoning. METHOD: The study presents occupational poisoning by several compounds (methyl bromide, polycyclic aromatic hydrocarbons, and MOCA) and the corresponding biological monitoring methods utilized, as researched by the author. Changes in biological monitoring based on the history of the Study Group on Occupational Poisoning and Biological Monitoring of the Japan Society for Occupational Health are also introduced. RESULTS: The areas of occupational poisoning at different times and cases of occupational cancer caused by exposure to chemical substances were presented in lectures held by the Study Group on Occupational Poisoning and Biological Monitoring. These lectures showed that although biological monitoring was previously implemented primarily by measuring the urinary metabolites of the exposed substance, the monitoring methods used have changed as the nature of exposure has changed, leading to the development of new tools that detect trace, low concentration, and mixed exposures. CONCLUSION: The health management of workers handling chemical substances at occupational sites will be shifted to autonomous management. However, it should be noted that only biological monitoring can detect and prove true exposure. Particularly, risk assessment by biological monitoring is necessary for substances that are suspected to be absorbed through the skin, and the measurement methods used should continue to be developed and refined. The significance of biological monitoring will continue to increase.

Quantification of bromide ion in biological samples using headspace gas chromatography-mass spectrometry.

OBJECTIVES: In this study, we aimed to establish a method for quantifying bromide ions (Br- ) in blood and urine using gas chromatograph-mass spectrometer (GC-MS) equipped with a headspace sampler, for biological monitoring of workers exposed to methyl bromide. METHODS: Samples were mixed with dimethyl sulfate, and Br- ions were detected using GC-MS with a headspace sampler. The validity of the proposed method was evaluated based on most of the US FDA guidance. The values obtained were compared with reference values by analysis using SeronormTM Trace Elements Whole Blood L-1 RUO. RESULTS: The calibration curve showed good linearity in the Br- concentration range of 0.1-20.0 mg/L, and the coefficient of determination R2 value was >.999. Intraday and interday accuracy values were 99.3%-103.1% and 97.4%-101.8%, respectively. The measured and reference values of Seronorm were concordant. Herein, eight urine and serum samples of workers were analyzed; the samples' Br- concentrations were known. The correlation coefficients of urine and serum samples were 0.97 and 0.96, respectively, and results were consistent. CONCLUSIONS: This study established a simple and rapid method for the determination of Br- concentration in biological samples using GC-MS with a headspace sampler. Moreover, it can be used for biological monitoring of occupational exposure to methyl bromide and for the determination of Br- concentration in a wide range of biological samples.

Impact of dietary compositions and patterns on the prevalence of nonalcoholic fatty liver disease in Japanese men: a cross-sectional study.

BACKGROUND: This study aimed to examine the impact of dietary compositions and patterns on nonalcoholic fatty liver disease (NAFLD) morbidity in Japanese men. METHODS: We conducted a cross-sectional study of 281 individuals who underwent comprehensive medical examinations during health screening. Dietary intake was assessed using a semi-quantitative food frequency questionnaire, and factor analysis was performed to detect dietary patterns. NAFLD was diagnosed by the presence of fatty liver on abdominal ultrasonography in nondrinkers (< 30 g/day), and patients were categorized into control (n = 192) and NAFLD groups (n = 89). RESULTS: Compared with the control group, the NAFLD group consumed fewer mushrooms. Three dietary patterns were identified, namely, a healthy pattern, a western pattern, and a snack pattern. The score of healthy pattern was negatively correlated with the risk of NAFLD. Compared with the lowest tertile of the healthy pattern, the middle tertile was associated with a lower risk of NAFLD after adjusting for age, physical activity, and smoking (odds ratio: 0.47, 95% confidence interval: 0.25-0.91). After further adjustments for body mass index, the middle tertile was associated with a lower risk of NAFLD (odds ratio: 0.46, 95% confidence interval: 0.23-0.92). CONCLUSIONS: A healthy dietary pattern comprising frequent intake of seaweeds, vegetables, mushrooms, pulses, and potatoes and starches was associated with a lower risk of NAFLD in Japanese men. In our opinion, this healthy pattern closely resembles the Japanese Washoku diet, indicating that adherence to Washoku may help prevent NAFLD.

Zinc-containing Mohs' paste affects blood flow and angiogenesis suppression.

PURPOSE: Mohs' paste, which is composed of zinc chloride and zinc oxide starch, is used for hemostasis of superficial malignancy in the clinical setting. We investigated the concentration of intramuscular zinc in mice after Mohs' paste application and evaluated its relationship with angiogenesis from the perspective of blood flow levels within 24 h. METHODS: Male C57BL/6JJmsSlc mice were administered single dose of Mohs' paste at 25%, 50%, and 75% after unilateral hind limb surgery, and glycerin, a viscosity modifier, was administered to the control group (0%). Hind limb blood flow levels were measured with a laser Doppler perfusion imaging system (n = 6). The amounts of intramuscular zinc and vascular endothelial growth factor-A (VEGF-A) expression were analyzed using inductively coupled plasma mass spectrometry (ICP-MS) and western blotting, respectively (n = 5 or 3). RESULTS: Blood flow levels were significantly decreased in the 50% group after 8 h, and significantly decreased in the 25% and 50% groups after 24 h. Intramuscular zinc was significantly increased in the 50% and 75% groups after 8 h. Western blotting showed that VEGF-A levels were significantly increased in the 25% and 50% groups after 8 h. Based on analytical experiments and biological investigation, we predicated the pharmacological effect of Mohs' paste and found over 50% of it is critical in the blood flow and angiogenesis suppression after more than 8 h of its application. CONCLUSIONS: The results suggest that the mechanism of blood flow suppression is independent of VEGF-A levels and might suppress future angiogenesis. Our findings support that of previous studies, in which Mohs' paste was expected to induce hemostasis and suppress angiogenesis. It is an excellent ointment that facilitates hemostasis by suppressing blood flow regardless of angiogenesis, and may be apt for situations where hemostasis is required in the clinical setting.

STAT3 inhibitory activity of naphthoquinones isolated from Tabebuia avellanedae.

The extract of Tabebuia avellanedae has been used as a folk medicine, and the various biological activities of T. avellanedae have been extensively studied. However, few studies have reported which natural products play a role in their biological effects. In this study, we evaluated representative naphthoquinones isolated from T. avellanedae and found that furanonaphthoquinones were the key structures required to exhibit STAT3 phosphorylation inhibitory activities. Our SAR analysis indicated that removal of a hydroxyl group enhanced the STAT3 phosphorylation inhibitory activity. In addition, the combined results of a mobility shift assay, SH2 domain binding assay, and docking simulation by Autodock 4.2.6 suggested that (S)-5-hydroxy-2-(1-hydroxyethyl)naphtho[2,3-b]furan-4,9-dione (1) could directly bind to the hinge region of STAT3.

Metabolism of 3-[5'-deoxy-5'-(dimethylarsinoyl)-ß-ribofuranosyloxy]-2-hydroxypropylene glycol in an artificial digestive system.

Seaweeds contain large amounts of organoarsenic compounds, mostly arsenosugars (AsSug) and arsenolipids (AsLipid). AsSug is mainly metabolized into dimethylarsinic acid (DMA V ) in humans. However, this metabolic process is not well understood. We investigated the metabolism of an AsSug, 3-[5'-deoxy-5'-(dimethylarsinoyl)-ß-ribofuranosyloxy]-2-hydroxypropylene glycol (AsSug328), in the gastrointestinal tract using an in vitro artificial gastrointestinal digestion system. AsSug328 was incubated with gastric juice for 4 h, with bile-pancreatic juice for 0.5 h, and finally with enteric bacteria solution for 24 h. The conversion of arsenic compounds after artificial digestion was analyzed by HPLC-ICP-MS and HPLC-ESI-Q-TOF-MS. Our results show that artificial gastrointestinal digestion converted AsSug328 into thio-AsSug328. However, no formation of DMA V was detected. Under the artificial digestion system, the 5-deoxyribofuranose structure of AsSug was maintained. Therefore, AsSug should be absorbed in the intestinal tract after its sugar moiety is partially decomposed. They are then possibly metabolized to DMA V in the liver and subsequently excreted through urine.

Proteomic analysis of hippocampal proteins in acrylamide-exposed Wistar rats.

Acrylamide has been used industrially and also found in certain foods cooked at high temperatures. Previous reports described acrylamide-related human intoxication who presented with ataxia, memory impairment, and/or illusion. The aim of this study was to characterize the molecular mechanisms of neurotoxicity of acrylamide by analyzing the expression levels of various proteins in the hippocampus of rats exposed to acrylamide. Male Wistar rats were administered acrylamide by gavage at 0, 2, and 20 mg/kg for 1 week or 0, 0.2, 2, and 20 mg/kg for 5 weeks. At the end of the experiment, the hippocampus was dissected out and proteins were extracted for two-dimensional difference gel electrophoresis combined with matrix-assisted laser-desorption ionization time-of-flight/time-of-flight mass spectrometry (MALDI-TOF/TOF/MS). MALDI-TOF/TOF/MS identified significant changes in two proteins in the 1-week and 22 proteins in the 5-week exposure groups. These changes were up-regulation in 9 and down-regulation in 13 proteins in the hippocampus of rats exposed to acrylamide at 20 mg/kg for 5 weeks. PANTHER overrepresentation test based on the GO of biological process showed significant overrepresentation in proteins annotated to nicotinamide nucleotide metabolic process, coenzyme biosynthetic process, pyruvate metabolic process, and carbohydrate metabolic process. The test also showed significant overrepresentation in proteins annotated to creatinine kinase activity for the GO of molecular function as well as myelin sheath, cytoplasmic part, and cell body for the GO of cellular component. Comparison with a previous proteomic study on hippocampal proteins in rats exposed to 1-bromopropane identified triosephosphate isomerase, mitochondrial creatine kinase U-type, creatine kinase ß-type and proteasome subunit α type-1 as proteins affected by exposure to acrylamide and 1-bromopropane, suggesting a common mechanism of neurotoxicity for soft electrophiles.

Estimation of the effective reproduction number of influenza based on weekly reports in Miyazaki Prefecture.

In Japan, as part of surveillance for seasonal influenza, the number of patients per influenza sentinel site is counted on a weekly basis. Currently, reference values are set for the weekly reported number of influenza cases per sentinel, and pre-epidemic and epidemic warnings are issued based on these values. In this study, we examined the association between these reference values and the effective reproduction number (Rt) using surveillance data for Miyazaki Prefecture collected from 2010 to 2011. There are nine public health centre jurisdictions in this prefecture, and Rt exceeded 1.0 at the time when pre-epidemic warnings were issued in almost all the jurisdictions. Thus, it was indicated that the validity of the reference value was also high for influenza transmission. However, our results indicated the presence of secondary epidemic caused by infections originating both from other jurisdictions and inner jurisdictions, and it is occasionally not possible to evaluate the end of an epidemic in a jurisdiction using only the reference value of termination. It is necessary to establish new methods after considering the situation in the surrounding jurisdictions for more detailed epidemic predictions.

Assessing the chronic respiratory health risk associated with inhalation exposure to powdered toner for printing in actual working conditions: a cohort study on occupationally exposed workers over 10 years.

BACKGROUND: Little epidemiological evidence exists regarding the chronic respiratory effects of inhaled powdered toner exposure in humans, although several case reports have suggested the existence of lung disorders that might be related to exposure to toner dust. OBJECTIVE: We aimed to estimate the chronic health risk to humans associated with routine toner dust exposure in copier industry workers under current actual work conditions. DESIGN: A prospective observational cohort study of occupational population. METHODS: Changes in chest radiogram, spirometry measurements and serum and urine biomarkers of biomedical responses to extrinsic stress, as well as subjective symptoms were longitudinally observed for up to 10 years in Japanese copier industry workers responsible for the manufacturing, maintenance or recycling of powdered toner or toner-using machines. A total of 694 subjects who did not change their work category during the follow-up and were free from chronic respiratory diseases at the baseline survey provided reliable results on at least three survey occasions during 3 years or more of follow-up. RESULTS: Typical fibrosis findings associated with pneumoconiosis was not observed on chest radiograms. No significant differences associated with toner exposure were noted in the frequency of new incidence of either non-specific findings on chest radiogram or serum fibrosis biomarkers (sialylated carbohydrate antigen KL-6 and surfactant protein D). However, the exposed subjects tended to show increases in the frequency of respiratory symptoms and reduced spirometry results during the follow-up compared with the control group, although significant differences were only seen in chronic cough. CONCLUSIONS: Under the current reasonably controlled work environmental conditions, lung fibrotic changes caused by inhaled dust exposure, including powdered toner, appear to be relatively uncommon; however, non-specific temporal irritation causing subjective symptoms and inflammatory responses might exist.

Correction to: Association between NAT2, CYP1A1, and CYP1A2 genotypes, heterocyclic aromatic amines, and prostate cancer risk: a case control study in Japan.

Following publication of the original article [1], the authors reported a correction in the units in the methods section under "Subjects".

Association between NAT2, CYP1A1, and CYP1A2 genotypes, heterocyclic aromatic amines, and prostate cancer risk: a case control study in Japan.

BACKGROUND: Heterocyclic aromatic amines (HAAs) may confer prostate cancer risk; however, the evidence is inconclusive and the activity of HAA-metabolizing enzymes is modulated by gene variants. The purpose of our study was to determine whether there was evidence of an association between HAA intake, polymorphisms in NAT2, CYP1A1, and CYP1A2 and prostate cancer risk in Japanese men. METHODS: Secondary data analysis of an observational case control study was performed. Among 750 patients with prostate cancer and 870 healthy controls, 351 cases and 351 age-matched controls were enrolled for analysis. HAA intake was estimated using a food frequency questionnaire and genotypes were scored by TaqMan real-time PCR assay. Logistic regression analysis was conducted according to affected/control status. RESULTS: We found that high HAA intake was significantly associated with an increased risk of prostate cancer (odds ratio (OR), 1.90; 95% confidence interval (95% CI), 1.40-2.59). The increased risk of prostate cancer was observed among individuals with the NAT2 slow acetylator phenotype (OR, 1.65; 95% CI, 1.04-2.61), CYP1A1 GA + GG genotype (OR, 1.27; 95% CI, 1.02-1.59), and CYP1A2 CA + AA genotype (OR, 1.43; 95% CI, 1.03-2.00). In addition, CYP1A1 GA + GG genotypes were associated with increased cancer risk in low (OR, 2.05; 95% CI, 1.19-3.63), moderate (OR, 1.72; 95% CI, 1.07-2.76), and high (OR, 2.86; 95% CI, 1.83-4.47) HAA intake groups. CONCLUSIONS: Our results suggest that high HAA intake is a risk factor of prostate cancer, and genotypes related to HAA metabolic enzymes can modulate the degree of the risk.

Quantitative effects of a declaration of a state of emergency on foot-and-mouth disease.

OBJECTIVES: The law in Japan requires the declaration of a state of emergency and implementation of countermeasures for an epidemic of a new infectious disease. However, because a state of emergency has never been declared in Japan, its effects remain unknown. The required countermeasures are similar to those implemented in the foot-and-mouth disease epidemic in Miyazaki in 2010. This study aimed to quantitatively estimate the effect of the declaration in 2010 and investigate the nature of the epidemic based on the day on which the declaration took effect. METHODS: Only publicly available data were used. Data for farms in the most affected town were analyzed. A modified susceptible-infected-recovered model was used to estimate the effect and for the simulation. Another model was used to estimate the effective reproduction number. RESULTS: After the declaration, the intra-bovine transmission rate decreased by 18.1 %, and there were few days when the effective reproduction number was >1.0. A few weeks delay in the declaration significantly increased the possibility of epidemic, number of farms at peak, and final infection scale. CONCLUSIONS: Based on the substantial decrease in the transmission rate after the declaration of a state of emergency in 2010, a future declaration will have a similar effect for a new infectious disease even though a direct extrapolation is not valid. Although a declaration should be carefully considered owing to the potential socioeconomic effects, it is essential to prepare for the implementation given that a delay of only a few weeks should be acceptable.

A novel metabolic activation associated with glutathione in dimethylmonothioarsinic acid (DMMTA(V))-induced toxicity obtained from in vitro reaction of DMMTA(V) with glutathione.

The purpose of the present study was to elucidate the metabolic processing of dimethylmonothioarsinic acid (DMMTA(V)), which is a metabolite of inorganic arsenic and has received a great deal of attention recently due to its high toxicity. The metabolites produced from an in vitro reaction with GSH were analyzed by high performance liquid chromatography-time of flight mass spectrometer (HPLC-TOFMS), HPLC with a photodiode array detector (PDA), and also gas chromatography-mass spectrometry (GC-MS) and GC with a flame photometric detector (FPD). The reaction of dimethylarsinic acid (DMA(V)) with GSH did not generate DMA(V)-SG but did generate dimethylarsinous acid (DMA(III)) or DMA(III)-SG. On the contrary, we confirmed that the reaction of DMMTA(V) with GSH directly produced the stable complex of DMMTA(V)-SG without reduction through a trivalent dimethylated arsenic such as DMA(III) and DMA(III)-SG. Furthermore, the present study suggests the production of hydrogen sulfide (H2S) and dimethylmercaptoarsine (DMA(III)-SH), a trivalent dimethylated arsenic, as well as DMA(III) and DMA(III)-SG in the decomposition process of DMMTA(V)-SG. These results indicate that the toxicity of DMMTA(V) depends not only on the formation of DMA(III) but also on at least those of H2S and DMA(III)-SH.

Associations between estrogen receptor genetic polymorphisms, smoking status, and prostate cancer risk: a case-control study in Japanese men.

OBJECTIVE: Prostate cancer (PCa) is one of the major causes of death among men. Our study investigated the association of ESR1 and ESR2 genotypes with susceptibility to PCa in relation to smoking status in Japanese. METHOD: A case-control study was performed with 750 Japanese prostate cancer patients and 870 healthy controls. After age-matching in case-controls, 352 controls and 352 cases were enrolled in this study. By using logistic regression analysis, the different genotypes from ESR1 and ESR2 were analyzed according to case/control status. RESULT: ESR2 rs4986938 AG and AG + AA genotypes were associated with significantly decreased risk of PCa (AG: OR = 0.68, 95 % CI 0.47-0.97, P < 0.05 and AG + AA: OR = 0.67, 95 % CI 0.47-0.94, P < 0.05). However, there was no significant association between ESR1 rs2234693 and PCa risk. When patients were grouped according to smoking status, the ESR2 rs1256049 AA genotype (OR = 0.48, 95 % CI 0.25-0.95, P < 0.05) and ESR2 rs4986938 AG + AA genotype (OR = 0.64, 95 % CI 0.41-1.00, P < 0.05) showed significantly decreased PCa risk in the ever-smoker group. CONCLUSION: Our results suggest that the estrogen receptor ESR2 has a very important function to predict PCa and that different SNPs have different predictive values. Smoking may influence estrogenic activity and may influence PCa together with the estrogen receptor.

Possible production of arsenic hemoglobin adducts via exposure to arsine.

OBJECTIVES: Arsine is an arsenic compound generated as a by-product in metal refineries. Accidental poisoning occurs sporadically; however, the administrative level for workers has not been established. Thus, it is essential to identify a highly specific biomarker for risk management in the workplace. The aim of this study was to identify an arsenic adduct, a potential biomarker, in the plasma. METHODS: Preserved mouse blood was exposed to arsine in vitro, and the plasma was separated. The residual clot of the control sample was hemolyzed using ultrapure water, and the supernatant was collected. Plasma from mice exposed to arsine in vivo was also separated from blood. Immunoprecipitation assays were conducted using all samples after ultrafiltration, and three fractions were collected. The total arsenic concentration in each fraction was quantified using inductively coupled plasma mass spectrometry (ICP-MS). The three in vitro samples and the eluate fraction from immunoprecipitation were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). RESULTS: In the exposed samples, the arsenic concentration in the fraction containing immunocomplexes was higher when immunoprecipitation was conducted with an anti-globin antibody. Three peaks were specifically observed in arsine-exposed samples after MALDI-TOF-MS analysis. Two of them were around m/z 15,000, and the other was m/z 15,700. The latter peak was confirmed even after immunoprecipitation. CONCLUSIONS: Globin forms an adduct with arsenic after both in vitro and in vivo exposure to arsine. This adduct together with hemoglobinuria could be a candidate biomarker of acute arsine poisoning in plasma.

Association of arteriosclerosis index and oxidative stress markers in school children.

BACKGROUND: Recent years have seen increased numbers of children with conditions that contribute strongly to atherosclerotic disease, such as passive smoking, obesity, and dyslipidemia. In the present study, we evaluated the utility of non-invasive urinary markers in preventing lifestyle-related diseases by comparing lipid metabolism-related parameters with oxidative stress markers in school children. METHODS: Subjects were 85 first-grade students. The variables examined included the smoking in subjects' household; exercise habits; height and weight; blood pressure; and plasma total cholesterol, high-density lipoprotein cholesterol, triglyceride, leptin, blood sugar, urinary cotinine, 8-hydroxy-2'-deoxyguanosine (8-OHdG), and 8-isoprostaglandin F2α (IsoP). RESULTS: Of the subjects, 10.6% were obese (% overweight ≥ 20%), 3.5% had a high-risk arteriosclerosis index (AI; 3 ≤ AI < 5), and 29.4% were passive smokers. No significant differences were seen between boys and girls for any of the measurement parameters. Both urinary 8-OHdG (6.8-24.5 ng/mg creatinine) and IsoP (0.9-7.4 ng/mg creatinine) were detected in all subjects, and a significant positive correlation was seen between the two markers. On multiple regression analysis using AI as an objective variable and all non-invasive markers as explanatory variables, urinary IsoP correlated most strongly with AI (P ≤ 0.01). CONCLUSIONS: Risk factors for atherosclerosis in adults, such as obesity and hypercholesterolemia, are associated with oxidative stress and inflammation. The present findings of the strongest correlation between urinary IsoP and AI suggest that urinary IsoP may serve as a non-invasive and effective early marker in predicting risk in children of developing lifestyle-related diseases.

Arsine toxicity is induced by inhalation but not by percutaneous exposure in hairless mice.

Arsine (AsH3) is used in many industries, but there is insufficient knowledge about the potential for percutaneous absorption. In order to examine possible percutaneous absorption of arsine, we conducted inhalation studies. Arsine was generated by reducing arsenic trioxide with NaBH4. Male 5-week-old Hos:HR-1 hairless mice were subjected to a single percutaneous exposure or whole-body inhalation exposure of ca. 300 ppm arsine for 5 min. The examination was performed 0-6 hr after the exposure. Total arsenic in whole blood and hematocrit (Ht) values were measured. Generation of an arsenic-hemoglobin (As-Hb) adduct in the blood was detected using high-performance liquid chromatography with an inductively coupled plasma mass spectrometer (HPLC-ICP-MS). Ht values in the inhalation group significantly decreased after 3 hr, but those in the percutaneous exposure group did not. Total arsenic in the inhalation group was 9.0-14.2 mg/l, which was significantly higher than that in the percutaneous group. The As-Hb adduct was detected only in mice in the inhalation group. Histopathological changes were noted only in the inhalation group, with marked deposition of eosinophilic globules in the proximal convoluted tubules of the kidneys, the Kupffer cells of the liver, and the red pulp in the spleen, but not in the lungs. Immunohistochemically, these eosinophilic globules were stained positively by hemoglobin (Hb) antibody. In the present study, arsine-induced hemolysis and deposition of Hb occurred in the kidney via the inhalation route but not via percutaneous exposure. The presence of As-Hb adduct may be a useful indicator for confirming arsine poisoning.