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Cell-free DNA (cfDNA) is free DNA found in circulating blood that originates from apoptosis or necrosis, and elevated cfDNA concentrations have been reported in cancers and other diseases. In this study, the concentrations and fragment distributions of plasma cfDNA were investigated in elderly (n = 1) and pediatric (n = 1) patients with acute promyelocytic leukemia (APL) treated with arsenic trioxide (ATO). A slight increase in cfDNA concentrations was observed in the APL patients compared with healthy controls. In the elderly APL patient, the plasma cfDNA concentration increased with treatment duration, and significant positive correlations were observed between the plasma total arsenic concentration and cfDNA concentration during the first consolidation therapy. Ladder fragments (150–200bp, 300–400 bp, and 500–600 bp) were observed in a portion of plasma cfDNA from both APL patients. Moreover, APL-related gene mutations (FMS-like tyrosine kinase 3 internal tandem duplication, GATA-binding protein 2 gene, Janus-associated kinase 2 gene, Wilms’ tumor 1 gene, Kirsten-ras gene, and neuroblastoma-ras gene) were successfully genotyped from plasma cfDNA by using polymerase chain reaction-based methods. The findings of this study provide fundamental information on cfDNA from APL patients treated with ATO.

Extracellular matrix tenascin‑X (TNX) is the largest member of the tenascin family. Our previous study demonstrated that TNX was involved in hepatic dysfunction, including fibrosis, in mice that were administered a high‑fat and high‑cholesterol diet with high levels of phosphorus and calcium. The present study investigated whether overexpression of both the fibrinogen domain of TNX (TNX‑FG) and integrin α11, one of the TNX cell surface receptors, induces

Cell-free DNA (cfDNA) originates from apoptotic and/or necrotic cells. Few reports are available that examine cfDNA from postmortem samples. Therefore, this study investigated differences between postmortem and biogenic subjects in concentration and fragment distribution of serum cfDNA. We also clarified features of serum cfDNA in postmortem subjects. The results revealed that postmortem subjects had significantly higher cfDNA concentrations than healthy controls and patients with cardiac disease. Serum cfDNA concentrations increased slightly with postmortem interval in subjects who died of asphyxia, and they were slightly higher in subjects who died from internal vs. external causes. Microchip electrophoresis of serum cfDNA revealed a fragment larger than 10,000 bp in only two postmortem subjects; we speculate that the fragment may have originated from necrotic cells. A relatively high concentration of one 150–200 bp fragment was characteristic of postmortem samples. This fragment may have been derived from apoptosis or other processes. We also observed ladder fragments in some subjects who died from external causes. Although additional research is needed for verification, serum cfDNA concentrations and fragment patterns possibly be used as a tool to estimate postmortem intervals and cause of death.

Background Ne-Zake is the drinking of alcohol before sleeping for helping to fall asleep and sleep well, and Mukae-Zake is the drinking of alcohol in the morning for “calming down” or “curing hangovers”. Objective We sought to examine the relationship of insomnia with Ne-Zake and Mukae-Zake among healthy middle-aged Japanese farmers. Methods In a cross-sectional study of 746 participants (mean age, 59.5 years; women, 25.9%), Ne-Zake and Mukae-Zake were defined based on a self-administered questionnaire. Insomnia was defined as the Athens Insomnia Scale Japanese version ≥6 or usage of sleeping pills in the previous year. Logistic regression was used to calculate odds ratio (OR) of insomnia related to Ne-Zake and Mukae-Zake adjusting for sex, age, presence of sleep-related disorders, frequency of alcohol consumption, and quantity of alcohol consumed per one occasion. Results We observed insomnia, Ne-Zake, and Mukae-Zake in 174 (23.3%), 140 (18.8%), and 37 (5.0%) participants, respectively. After adjustment for demographic and confounding factors, participants with Ne-Zake had a significantly higher prevalence of insomnia (OR 2.00 [95% confidence interval, 1.27–3.16]), compared to those without Ne-Zake. Mukae-Zake was also independently associated with a higher prevalence of insomnia among men (OR 3.26 [1.55–6.87]). Participants with both Ne-Zake and Mukae-Zake had a highly significant association with insomnia (OR 4.77 [2.01–11.3]) than those with neither Ne-Zake nor Mukae-Zake. Additionally, for insomnia, the association of Mukae-Zake was more pronounced than that of Ne-Zake (OR 4.09, 95% CI 1.14–14.7, p = 0.031; and OR 1.81, 95% CI 1.08–3.06, p = 0.026, respectively). Conclusion Ne-Zake and Mukae-Zake were associated with insomnia independent of the quantity and frequency of alcohol consumption among Japanese farmers. This finding can be used for stratifying individuals with insomnia not only to improve sleep hygiene but also to prevent alcohol dependence by informing the general population that alcohol has a negative effect on sleep, contrary to popular beliefs.

Joint hypermobility syndrome (JHS) (also termed hypermobility type Ehlers-Danlos syndrome, hEDS) is a heritable connective tissue disorder that is characterized by generalized joint hypermobility, chronic pain, fatigue, and minor skin changes. Initially, it was reported that there is a small subset of patients with JHS/hEDS who have haploinsufficiency of tenascin-X (TNX). However, the relationship between TNXB and JHS/hEDS has not been reported at all afterwards. EDS was reclassified into thirteen types in 2017, and the causative gene of JHS/hEDS remained to be identified. Therefore, in this study in order to determine whether JHS/hEDS can be diagnosed by the concentrations of serum form of TNX (sTNX), we measured the concentrations of sTNX in 17 JHS/hEDS patients. The sTNX concentrations in half of the JHS/hEDS patients were significantly lower than those in healthy individuals. No mutations, insertions or deletions were detected in the TNX exon sequence of the JHS/hEDS patients except for one in patient. That patient has a heterozygous mutation. A correlation between sTNX concentration and mutation of the TNXB genomic sequence was not found in the JHS/hEDS patients. These results indicate that the decrease in sTNX concentration could be used as a risk factor for JHS/hEDS.

The human ABO blood group system is of great importance in blood transfusion and organ transplantation. ABO transcription is known to be regulated by a constitutive promoter in a CpG island and regions for regulation of cell-specific expression such as the downstream + 22.6-kb site for epithelial cells and a site in intron 1 for erythroid cells. Here we investigated whether the + 22.6-kb site might play a role in transcriptional regulation of the gene encoding odorant binding protein 2B (OBP2B), which is located on the centromere side 43.4 kb from the + 22.6-kb site. In the gastric cancer cell line KATOIII, quantitative PCR analysis demonstrated significantly reduced amounts of OBP2B and ABO transcripts in mutant cells with biallelic deletions of the site created using the CRISPR/Cas9 system, relative to those in the wild-type cells, and Western blotting demonstrated a corresponding reduction of OBP2B protein in the mutant cells. Moreover, single-molecule fluorescence in situ hybridization assays indicated that the amounts of both transcripts were correlated in individual cells. These findings suggest that OBP2B could be co-regulated by the + 22.6-kb site of ABO.

Although organophosphorus agents are used worldwide as pesticides, there have been many reports of pesticide poisoning. Nerve agents are organophosphorus agents that interfere with neurotransmission and have been used as chemical weapons in wars. These agents mainly irreversibly inhibit the action of acetylcholinesterase, an enzyme that breaks down acetylcholine, a neurotransmitter, and are believed to cause acute symptoms of poisoning. However, in recent years, the presence of subacute, delayed toxicity independent of acetylcholinesterase inhibition has been reported for some organophosphorus agents. We analyzed the subacute and delayed toxicity of bis(isopropylmethyl)phosphonate (BIMP), which has the same phosphonate group as sarin. BIMP rounded out the morphology of the cells and reduced the proportion of cells in the G1 phase of the cell cycle over time. No DNA damage was observed, suggesting that BIMP may affect cell division.

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Zinc homeostasis in cells depends on zinc transporters, which are divided into 2 families: ZnT (SLC30A) and ZIP (SLC39A). In this study, we examined the effect of 20 single nucleotide polymorphisms (SNPs) in 10 genes encoding zinc transporters on blood zinc concentration in Japanese subjects ( n = 102). Blood zinc levels were determined by microwave plasma-atomic emission spectrometry, and SNPs were analyzed by polymerase chain reaction followed by restriction fragment length polymorphism analysis. Among the 20 SNPs examined, 3 SNPs (SLC30A3 rs11126936, SLC39A8 rs233804, and SLC39A14 rs4872479) were significantly associated with blood zinc concentration. Individuals with genotype TT and TG in rs11126936 showed significantly higher blood zinc concentrations than those with GG. As for rs233804, individuals harboring the A allele had significantly higher blood zinc concentrations than those without this allele. Furthermore, the genotype TT and TG in rs4872479 had significantly higher blood zinc concentrations than those with GG. Among these three SNPs, combination of SLC30A3 rs11126936 and SLC39A8 rs233804 may strongly affect blood zinc levels. This study is the first comprehensive investigation of the effect of SNPs in genes encoding zinc transporters on blood zinc concentration. Adverse effects of zinc deficiency are reported and above 3 SNPs may be related to genetic susceptibility to zinc deficiency.

Background ABO and its paralogues, such as A3GALT2 and GGTA1, encoding α1,3-Gal(NAc) transferases, belong to the glycosyltransferase 6 (GT6) gene family. We have developed an alternative method for the identification of species based on sequence variations within the GT6 gene family, which is applicable to degraded DNA. Methods/materials DNA samples prepared from control mammalian species, together with an unknown sample, were polymerase chain reaction (PCR)-amplified using one universal primer pair targeting the sequences in the last coding exons of the GT6 gene family, yielding 141-bp products derived from those multiple loci. After cloning, sequence determination and Basic Local Alignment Search Tool analysis, phylogenetic trees were constructed. Results Comparison of the sequences obtained with those references showed good concordance with each of the starting species of mammals. This system was able to identify 'mouse' or 'rodent' as the origin of the unknown sample. Conclusion For the identification of species, genotyping of ABO and its homologues would be applicable for the analysis of degraded DNA samples. Although the method employed in this study is likely valid for mammals, it would not be suitable for birds, fish and reptiles. It may be possible to improve the present method for use with other species by employing an alternative universal primer set.

Recent studies of copy number variations (CNVs) associated with physical features, such as body mass index, body height or bone length, have suggested that such CNVs could serve as markers in forensic cases involving unidentified individuals. However, the process of cataloging CNVs has been slow because of the cumbersome nature and low reliability of the procedures involved. Here we describe a simple quantitative real-time PCR (Q-PCR) method for screening of medicolegally useful CNVs, which does not require reference DNA with known copy number. The first step is to prepare a chimeric plasmid vector including one copy each of the single-copy gene-specific sequence as the internal standard, and the target CNV-specific sequence. To assess the validity of this new method, we analyzed CNVs in the LTBP1 and ETV6 gene regions, both of which are candidate CNVs associated with body height. The PCR efficiencies for the single-copy (reference) gene and the target CNV were similar, indicating that quantitation was reliable. Furthermore, simulated analysis of the LTBP1 CNV using mock samples prepared by mixing vectors in varying proportions showed that this analytical method allowed correct determination of the LTBP1 copy number. These results demonstrated that our simple method has considerable potential for screening of trait-related CNVs that would be useful for forensic casework.

Problematic smartphone use among adolescents has become a social concern and is associated with poor sleep quality. The relationship between life habits, such as smartphone use and sleep duration, and levels of immunological and neuroendocrine biomarkers, including the stress hormone cortisol, in adolescents seems to be important to objectively comprehend their health and well-being in school life. However, such a relationship has not been well documented. We therefore studied rural junior high school students in Japan to elucidate the relationship between serum cortisol (SC) levels and their life habits. A total of 155 students in the seventh grade in 2016 were recruited as subjects. Of them, 140 students with eligible responses and blood samples (12-13 years; 80 boys, 60 girls) were finally included in the study (response rate 90.3%). A questionnaire survey concerning wake-up time, sleep duration, and the length of time using a smartphone per day was conducted. Blood samples were collected from peripheral veins of participants under fasting conditions between 8:30 and 11:00 a.m. The Spearman rank correlation coefficients were as follows: between SC and wake-up time, 0.199 (p = 0.018); between SC and sleep duration, 0.185 (p = 0.029); and between SC and time spent on smartphones, 0.172 (p = 0.042). The multiple regression analysis showed that high SC levels were significantly associated with late wake-up time and with short sleep duration. We therefore propose that measuring SC levels is useful for early detection of the change in the well-regulated daily life among junior high school students.

Circulating cell-free DNA (cfDNA) has been directly related to cancer, diabetes, stroke, systemic lupus erythematosus, trauma, rheumatoid arthritis, inflammation, infection, and myocardial infarction (MI). In this study, plasma cfDNA was extracted from the plasma of cardiac disease patients and the cfDNA fragment distribution as well as the relationships between cfDNA concentration and deoxyribonuclease I (DNase I) activity enzyme implicated in double-stranded DNA processing were examined. Results revealed that the cfDNA concentrations in patients with MI and cardiac angina were significantly higher than that in healthy control subjects. Microchip electrophoresis of plasma cfDNA revealed a single fragment (150–200 bp) in some healthy control subjects and three fragments (150–200 bp, 300–400 bp, and 500–600 bp) in all cardiac patient samples. Moreover, a cfDNA ratio of 150–200 bp/500–600 bp was significantly more prevalent in MI patients than in patients with other cardiac diseases (chest pain, cardiac angina, atrial fibrillation and cardiac failure). In addition, a positive correlation between DNase I activity and cfDNA concentration was observed. These results suggest that the plasma cfDNA in cardiac disease patients may originate from apoptosis and that the 150–200 bp/500–600 bp ratio for cfDNA may be a novel diagnostic indicator for MI.

Background Loss of blood group ABO antigens on red blood cells (RBCs) is well known in patients with leukemias, and such decreased ABO expression has been reported to be strongly associated with hypermethylation of the ABO promoter. We investigated the underlying mechanism responsible for A-antigen reduction on RBCs in a patient with myelodysplastic syndrome. Study design and methods Genetic analysis of ABO was performed by PCR and sequencing using peripheral blood. RT-PCR were carried out using cDNA prepared from total bone marrow (BM) cells. Bisulfite genomic sequencing was performed using genomic DNA from BM cells. Screening of somatic mutations was carried out using a targeted sequencing panel with genomic DNA from BM cells, followed by transient transfection assays. Results Genetic analysis of ABO did not reveal any mutation in coding regions, splice sites, or regulatory regions. RT-PCR demonstrated reduction of A-transcripts when the patient's RBCs were not agglutinated by anti-A antibody and did not indicate any significant increase of alternative splicing products in the patient relative to the control. DNA methylation of the ABO promoter was not obvious in erythroid cells. Targeted sequencing identified somatic mutations in ASXL1, EZH2, RUNX1, and WT1. Experiments involving transient transfection into K562 cells showed that the expression of ABO was decreased by expression of the mutated RUNX1. Conclusion Because the RUNX1 mutation encoded an abnormally elongated protein without a transactivation domain which could act as dominant negative inhibitor, this frame-shift mutation in RUNX1 may be a genetic candidate contributing to A-antigen loss on RBCs.

Deoxyribonucleases (DNases) might play a role in prevention of autoimmune conditions such as systemic lupus erythematosus through clearance of cell debris resulting from apoptosis and/or necrosis. Previous studies have suggested that variations in the in vivo activities of DNases I-like 3(1L3) and II have an impact on autoimmune-related conditions. The genes for these DNases are known to show copy number variations (CNVs) whereby copy loss leads to a reduction of the in vivo activities of the enzymes, thereby possibly affecting the pathophysiological background of autoimmune diseases. Using a simple newly developed quantitative real-time PCR method, we investigated the distributions of the CNVs for DNASE1L3 and DNASE2 in Japanese and German populations. It was found that only 2 diploid copy numbers for all of these DNASE CNVs was distributed in both of the study populations; no copy loss or gain was evident for any of the autoimmune-related DNase genes. Therefore, it was demonstrated that these human autoimmune-related DNase genes show low genetic diversity of CNVs resulting in alterations of the in vivo levels of DNase activity.

Each year in Japan from 1990 to 1997, approx. 21,000–24,000 individuals committed suicide. In 1998, the number of suicides increased to >30,000, and a trend of high suicide numbers then persisted for >10 years. Although Japan’s annual number of suicides has recently been decreasing, it remains among the highest worldwide. Herein, we assessed the annual suicide data (numbers and rates) related to three economic and life indicators: (1) the difference between actual income and consumer spending of one average month per year in one household, (2) the annual difference between exports and imports, and (3) the annual total debt determined by statistical analyses for both sexes/males/females during the 40-year period from 1979 to 2018 in Japan. Our findings indicated that [1] total debt may be associated with both the number of suicides and the suicide rate for both sexes, for males, and for females, and [2] the difference between actual income and consumer spending may be associated with both the number of suicides and the suicide rate only in females. These findings revealed factors that are clearly suicide-related, and it is necessary to design suicide prevention strategies based on the factors. Relevant public and private entities should become aware of the involvement of both debt and the difference between income and spending in suicide trends as they plan suicide prevention measures. Further analyses of suicide data should be performed in a wide range of fields including legal medicine, toward a greater understanding of suicide risk factors.

A foreign body impacted in the esophagus is not a rare incident among adults or children. In adults, a dental prosthesis is prone to become impacted in the esophagus. The diagnostic difficulty of this often causes a delay in its removal, which can lead to serious complications, including death. This report describes the autopsy case of a man who died of prolonged asphyxiation induced by the delayed removal of an impacted denture, which was misdiagnosed on his first visit notwithstanding that a part of the denture could be seen on X-rays. Cases in which an impacted denture led to death have rarely been reported in contrast to numerous papers about recovered cases.

A 45-year-old male, an alcohol addict with asthma, was found dead in his home, after several days of continued drinking. A forensic autopsy was performed 3days after the discovery of his death in order to specify the cause of death. A longitudinal perforation penetrating all layers of the esophagus measuring 1.8cm was present on the left wall approximately 2.0cm from the gastroesophageal junction. There were 1900mL of greenish to brownish turbid liquid in the left pleural cavity and 150mL of greenish viscous liquid in the stomach. Histopathologically, an infiltration of numerous neutrophils was evident in the submucosa layer, proper muscular layer, and serous membrane of the esophagus, corresponding to the esophageal laceration. The serum C-reactive protein (CRP) concentration was determined to be 3.1mg/dL. The alcohol concentrations were determined to be 1.49mg/g in the right cardiac blood, 1.31mg/g in the left cardiac blood, and 2.48mg/g in urine. Based upon the autopsy and histopathological findings, as well as the biochemical and toxicological analyses, we concluded that the cause of death was respiratory failure by pleural effusion, resulting from spontaneous esophageal perforation. This was the first report of a spontaneous esophageal perforation eventually causing respiratory failure.

This study examined the usefulness of statistical parametric mapping (SPM) for investigating postmortem changes on brain computed tomography (CT). This retrospective study included 128 patients (23 − 100 years old) without cerebral abnormalities who underwent unenhanced brain CT before and after death. The antemortem CT (AMCT) scans and postmortem CT (PMCT) scans were spatially normalized using our original brain CT template, and postmortem changes of CT values (in Hounsfield units; HU) were analysed by the SPM technique. Compared with AMCT scans, 58.6 % and 98.4 % of PMCT scans showed loss of the cerebral sulci and an unclear grey matter (GM)–white matter (WM) interface, respectively. SPM analysis revealed a significant decrease in cortical GM density within 70 min after death on PMCT scans, suggesting cytotoxic brain oedema. Furthermore, there was a significant increase in the density of the WM, lenticular nucleus and thalamus more than 120 min after death. The SPM technique demonstrated typical postmortem changes on brain CT scans, and revealed that the unclear GM–WM interface on early PMCT scans is caused by a rapid decrease in cortical GM density combined with a delayed increase in WM density. SPM may be useful for assessment of whole brain postmortem changes. • The original brain CT template achieved successful normalization of brain morphology. • Postmortem changes in the brain were independent of sex. • Cortical GM density decreased rapidly after death. • WM and deep GM densities increased following cortical GM density change. • SPM could be useful for assessment of whole brain postmortem changes.

The human ABO blood group system is of great importance in blood transfusion and organ transplantation. The ABO system is composed of complex carbohydrate structures that are biosynthesized by A- and B-transferases encoded by the ABO gene. However, the mechanisms regulating ABO gene expression in epithelial cells remain obscure. On the basis of DNase I-hypersensitive sites in and around ABO in epithelial cells, we prepared reporter plasmid constructs including these sites. Subsequent luciferase assays and histone modifications indicated a novel positive regulatory element, designated the +22.6-kb site, downstream from ABO, and this was shown to enhance ABO promoter activity in an epithelial cell-specific manner. Expression of ABO and B-antigen was reduced in gastric cancer KATOIII cells by biallelic deletion of the +22.6-kb site using the CRISPR/Cas9 system. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay demonstrated that the site bound to an epithelial cell-specific transcription factor, Elf5. Mutation of the Ets binding motifs to abrogate binding of this factor reduced the regulatory activity of the +22.6-kb site. Furthermore, ELF5 knockdown with shRNA reduced both endogenous transcription from ABO and B-antigen expression in KATOIII cells. Thus, Elf5 appeared to be involved in the enhancer potential of the +22.6-kb site. These results support the contention that ABO expression is dependent upon a downstream positive regulatory element functioning through a tissue-restricted transcription factor, Elf5, in epithelial cells.

Background Complete deficiency of an extracellular matrix tenascin-X (TNX) leads to a classical type of Ehlers-Danlos syndrome (EDS). TNX haploinsufficiency is a cause of hypermobility type of EDS. Human TNX is also present in a serum form (sTNX) with a molecular size of 140 kDa. In this study, we established a method for quantification of sTNX using nano-liquid chromatography tandem mass spectrometry (LC/MS/MS) with selected/multiple reaction monitoring. Methods Twelve abundant protein-depleted sera were reduced, alkylated, and digested with Lys-C and trypsin. Subsequently, the digests were fractionated by strong cation exchange chromatography. Optimal and validated transitions of precursor and product ions of the peptides from sTNX were developed on a triple quadrupole mass spectrometer. Results Serum concentrations of sTNX of healthy individuals were quantified as an average of 144 ng/ml. However, sTNX was not detected by this method in serum from a patient with a classical type of EDS in whom sTNX was not found by Western blot analysis. The limit of quantification (LOQ) of sTNX by nano-LC/MS/MS method was 2.8 pg whereas the detection sensitivity of sTNX by Western blot analysis was 19 pg. The nano-LC/MS/MS method is more sensitive than Western blot analysis. Conclusions The quantification method will be useful for diagnosis and risk stratification of EDS caused by TNX deficiency and haploinsufficiency.

Raman spectroscopy is commonly used in chemistry to identify molecular structure. This technique is a nondestructive analysis and needs no sample preparation. Recently, Raman spectroscopy has been shown to be effective as a multipurpose analytical method for forensic applications. In the present study, blood identification and discrimination between human and nonhuman blood were performed by a portable Raman spectrometer, which can be used at a crime scene. To identify the blood and to discriminate between human and nonhuman blood, Raman spectra of bloodstains from 11 species (human, rat, mouse, cow, horse, sheep, pig, rabbit, cat, dog, and chicken) were taken using a portable Raman spectrometer. Raman peaks for blood (742, 1001, 1123, 1247, 1341, 1368, 1446, 1576, and 1619 cm−1) could be observed by the portable Raman spectrometer in all 11 species, and the human bloodstain could be distinguished from the nonhuman ones by using a principal component analysis. This analysis can be performed on a bloodstain sample of at least 3 months old. The portable Raman spectrometer can be used at a crime scene, and this analysis is useful for forensic examination.

OBJECTIVE: To continue our previous investigations, we have extensively investigated the function of the 61, 41, and 35 non-synonymous single nucleotide polymorphisms (SNPs) in the human genes encoding DNASE1, DNASE1L3, and DNASE2, respectively, potentially relevant to autoimmune diseases. METHODS: The site-directed mutagenesis was employed to amino acid-substituted constructs corresponding to each SNP. The COS-7 cells were transfected with each vector and DNase activity was assayed by the single radial enzyme diffusion method. By using PolyPhen-2, changes in the DNase function of each non-synonymous SNP were predicted. Genotyping of all the non-synonymous SNPs was performed in 14 different populations including 3 ethnic groups using the polymerase chain reaction followed by the restriction fragment length polymorphism method. RESULTS: Expression analysis demonstrated these SNPs to be classified into four categories with regard to the effect on DNase activity: SNPs not affecting the activity level, ones reducing it, ones abolishing it, and ones elevating it. In particular, 9, 5, and 4 SNPs producing a loss-of-function variant of the enzymes in DNASE1, DNASE1L3, and DNASE2, respectively, were confirmed. SNPs producing DNase loss of function can be estimated by PolyPhen-2 to be "probably damaging" with a high accuracy of prediction. Almost all of these functional SNPs producing a loss of function or substantially low activity-harboring forms exhibited a mono-allelic distribution in all of the populations. CONCLUSION: A minor allele of functional SNPs, despite the remarkably low genetic heterogeneity of the SNPs, might be a genetic risk factor for autoimmune diseases.

The annual number of suicides in Japan totaled around 23,000 in 1997 and abruptly increased to around 31,000 in 1998. This figure has remained high since then. This abrupt increase in the number of suicides was primarily due to an increase in suicides occasioned by economic concerns. The association between various economic factors and suicide must be studied in detail and over the long term in order to ascertain the association between economic concerns and suicide. This study examined the relative poverty rate and the suicide rate in Japan over 30 years and discussed the association between those two rates. The results suggest that the relative poverty rate may be associated with the suicide rate for both sexes. This association is true for men in particular. The organizations and professionals involved in implementing suicide prevention measures should be cognizant of the current findings and consider formulating additional specific measures.

In Japan over the past few years, approximately 13,000 individuals were arrested for drug offenses each year. It is useful to know the trends in drug offenses, in order to devise the most effective countermeasures and addiction treatment programs. Herein, we have revealed the trends in drug offenses in the Tokyo Metropolitan Area. This report was researched the number of individuals arrested for drug offenses in Tokyo during the 3-year study period 2016-2018. The drugs are classified into the six categories: stimulants, narcotics, psychoactive drugs, opium, cannabis, and designated substances. We also calculated the percentages of individuals arrested for various drug offenses based on these six categories. Approximately 86% of the arrests for drug offenses in Tokyo during the 3-year period were for stimulants or cannabis. A higher percentage of individuals were arrested for stimulants, but the percentage of individuals arrested for cannabis increased each year. Given the percentage of individuals arrested for designated substances or narcotics, preventive measures for drug offenses involving stimulants and cannabis should be promptly implemented. Further campaigns to prevent drug offenses and public lectures are also needed. Public education must be provided to prevent drug offenses involving designated substances and narcotics.

Recently it has been recognized that a considerable number of copy number variations (CNVs) are associated with diseases and other complex human traits. In our previous study, we developed a simple quantitative real-time PCR (Q-PCR) method for analysis of CNV copy number, which had the advantage of obviating the need for reference DNA with a known copy number. Using DNA samples obtained from 231 Japanese individuals, we applied this method for analyzing the copy number of a candidate CNV associated with body height, located in the neural precursor cell expressed, developmentally down-regulated 4-like, E3 ubiquitin protein ligase (NEDD4L) gene. In addition, the appropriateness of the results was evaluated and confirmed by quantification of amplicons with an Agilent 2100 Bioanalyzer. The NEDD4L gene encodes a member of the Nedd4 family of HECT domain E3 ubiquitin ligases. The target CNV located in the intron has been found to be significantly associated with height variation in Chinese. However, it remains unknown whether such an association exists in other populations, including Japanese. Analysis of the correlations between copy number and body height using ANOVA revealed no statistically significant correlations in Japanese.

In the present study, we used micro-Raman spectroscopy with high-resolution analysis to discriminate between bloodstains from infants and bloodstains from adults. Raman peaks were detected at 674, 754, 976, 1002, 1105, 1127, 1176, 1248, 1340, 1368, 1390, 1560, and 1611 cm1; these peaks were derived from hemoglobin, albumin, and glucose. However, a peak was obtained at 1105 cm1, which was assigned to histidine; this peak was observed only for bloodstains from adults. Human adult hemoglobin (HbA) is composed of an a2b2 tetramer structure, whereas human fetal hemoglobin (HbF) is composed of an a2c2. Therefore, the lack of a Raman peak at 1105 cm1in bloodstains from infants indicates the possibility of two histidine substitutions (His116Ile and His143Ser) in the y chain of HbF. This study discriminates between bloodstains from infants and bloodstains from adults using micro-Raman spectroscopy, with beneficial implications in forensic science.

Iron is bound to mobile transferrin (TF) and ferritin in blood. TF receptors (TFRC and TFR2) regulate intracellular iron by delivering iron from TF into the cytoplasm. In this study, we examined the effects of 10 single nucleotide polymorphisms (SNPs) in each of the genes for TF and TF receptors on blood iron concentrations in Japanese subjects. Blood iron levels were determined by microwave plasma-atomic emission spectrometry and the SNPs were analyzed by polymerase chain reaction followed by restriction fragment length polymorphism analysis. Blood iron levels in males were significantly higher than those in females. Therefore, the analysis was performed only in males. Blood iron concentrations did not correlate with age and postmortem intervals in males. Among the 10 SNPs in TF, TFRC, and TFR2 genes, significant associations were observed between TF genotypes (rs12769) and male iron concentrations. Individuals with genotype GG in rs12769 had significantly higher blood iron concentrations than those with GA. Previous studies have shown the association between high tissue iron concentrations and disease, liver iron levels are higher in infants dying from sudden infant death syndrome and decreased blood iron concentrations were observed in critically ill children. Therefore, rs12769 in TF might be related to diseases and mortality risk.

There are numerous social issues that need to be quickly addressed in Japan [1-5]. Some of these issues concern children [6-8], and one such issue is refusal to attend school. Over the years from 1995-2006 the proportion of Japanese children who refused to attend school increased until 2001, but that proportion plateaued from 2002-2006 [9]. Recent trends in refusal to attend school need to be studied in detail, and specific measures to deal with refusal to attend school need to be devised.

Over the past few years, the number of traffic fatalities and the number of individuals injured in traffic accidents in Japan have decreased [1]. From 2006 to 2016, the number of fatalities within 30 days of a traffic accident decreased annually in Japan from 7,336 in 2006 to 4,838 in 2014. However, the number increased slightly to 4,885 in 2015, though it decreased again to 4,698 in 2016. From 2006 to 2016, the number of individuals injured in traffic accidents decreased annually in Japan from 1,098,564 in 2006 to 618,853 in 2016. We previously examined detailed trends in traffic fatalities by age group in Japan, Germany, and France [2]. In the study, individuals ages 25–64 accounted for a large proportion of traffic fatalities in Germany and France, but the elderly age 65 or older accounted for a large proportion of traffic fatalities in Japan. The elderly accounted for 20.2% of Japan’s population in 2005, 23.0% in 2010, and 26.7% in 2015 [3], so the elderly represent a growing proportion of the population.