Class III Alcohol Dehydrogenase Plays a Key Role in the Onset of Alcohol-Related/-Associated Liver Disease as an S-Nitrosoglutathione Reductase in Mice.

Lipid accumulation in the liver due to chronic alcohol consumption (CAC) is crucial in the development of alcohol liver disease (ALD). It is promoted by the NADH/NAD ratio increase via alcohol dehydrogenase (ADH)-dependent alcohol metabolism and lipogenesis increase via peroxisome proliferator-activated receptor γ (PPARγ) in the liver. The transcriptional activity of PPARγ on lipogenic genes is inhibited by S-nitrosylation but activated by denitrosylation via S-nitrosoglutathione reductase (GSNOR), an enzyme identical to ADH3. Besides ADH1, ADH3 also participates in alcohol metabolism. Therefore, we investigated the specific contribution of ADH3 to ALD onset. ADH3-knockout (Adh3-/-) and wild-type (WT) mice were administered a 10% ethanol solution for 12 months. Adh3-/- exhibited no significant pathological changes in the liver, whereas WT exhibited marked hepatic lipid accumulation (p < 0.005) with increased serum transaminase levels. Adh3-/- exhibited no death during CAC, whereas WT exhibited a 40% death. Liver ADH3 mRNA levels were elevated by CAC in WT (p < 0.01). The alcohol elimination rate measured after injecting 4 g/kg ethanol was not significantly different between two strains, although the rate was increased in both strains by CAC. Thus, ADH3 plays a key role in the ALD onset, likely by acting as GSNOR.

Development of prevention strategies against bath-related deaths based on epidemiological surveys of inquest records in Kagoshima Prefecture.

Sudden death in the bathroom (bath-related death) occurs more frequently in Japan than in other countries. To clarify the epidemiological characteristics of bath-related deaths, we reviewed inquest records of deaths in Kagoshima Prefecture from 2006 to 2019. We identified 2689 cases of bath-related death. Of these cases, 90% were among people aged ≥ 65 years. The majority occurred in a home bathtub between 16:00 and 20:00. Most deaths (52.0%) occurred in winter (December-February), and there were extremely strong negative correlations with the environmental temperatures (maximum, minimum, and mean) on the day of death. We identified the environmental temperature during cold winter months that bath-related deaths are likely to occur in Kagoshima, although further investigation concerning the effects of other confounding factors is required. Forensic autopsies have only been performed in 29 cases and the cause of death was not diagnosed correctly in the majority of cases. Although autopsies are essential to elucidate the pathogenesis of the deaths, it is difficult to increase the rate of autopsies under the current Japanese death investigation system. Therefore, we suggest that the best way to prevent bath-related death is establishing an "Alert system" based on our results, and to have people refrain from bathing on dangerous days.

Epidemiological analysis of intramuscular hemorrhage of respiratory and accessory respiratory muscles in fatal drowning cases.

The postmortem diagnosis of drowning death and understanding the mechanisms leading to drowning require a comprehensive judgment based on numerous morphological findings in order to determine the pathogenesis and epidemiological characteristics of the findings. Effortful breathing during the drowning process can result in intramuscular hemorrhage in respiratory and accessory respiratory muscles. However, the characteristics of this phenomenon have not been investigated. We analyzed the epidemiological characteristics of 145 cases diagnosed as drowning, in which hemorrhage, not due to trauma, was found in the respiratory muscles and accessory respiratory muscles. Hemorrhage was observed in 31.7% of these cases, and the incidence did not differ by gender or drowning location. The frequency of hemorrhage was significantly higher in months with a mean temperature below 20°C than in months above 20°C, suggesting a relationship between the occurrence of hemorrhage and low environmental temperature. Moreover, the frequency of hemorrhage was significantly higher in the elderly (aged ≥65 years) compared to those <65 years old. In the elderly, the weakening of muscles due to aging may contribute to the susceptibility for intramuscular hemorrhage. Moreover, these intramuscular hemorrhages do not need to be considered in cases of a potential bleeding tendency due to disease such as cirrhosis or medication such as anticoagulants. Our results indicate that intramuscular hemorrhage in respiratory and accessory respiratory muscles can serve as an additional criterion to differentiate between fatal drowning and other causes of death, as long as no cutaneous or subcutaneous hematomas above the muscles with hemorrhages are observed. In addition, the epidemiological features that such intramuscular hemorrhage is more common in cold environments and in the elderly may provide useful information for the differentiation.

Alcohol consumption induces murine osteoporosis by downregulation of natural killer T-like cell activity.

INTRODUCTION: Chronic alcohol consumption (CAC) can induce several deleterious effects on the body, including the promotion of osteoporosis; however, the immunological mechanism underlying alcohol-induced osteoporosis is still unclear. METHODS: We administered alcohol to mice for 4 weeks as the experimental CAC model and analyzed the bone and immune cells that are located in the vicinity of a bone. RESULTS: IL-4 is known to be a suppressive factor for osteoclastogenesis, and we found that natural killer T (NKT)-like cells, which showed NK1.1-positive, CD3-positive, and α-galactosylceramide-loaded CD1d tetramer-negative, produced IL-4 more effectively than CD4+ T and natural killer (NK) cells. The alcohol consumption facilitated a significant decrease of bone mineral density with the upregulation of nuclear factor of activated T cells 1 and receptor activator of NF-κB ligand expression. Meanwhile, we confirmed that alcohol consumption suppressed the activity of antigen-presenting cells (APCs) and NKT-like cells, leading to decreased IL-4 secretion. Moreover, these harmful effects of alcohol consumption were reduced by simultaneous treatment with a glycolipid antigen OCH. CONCLUSIONS: Our results indicate that the inactivation of innate immune cells, APCs, and NKT-like cells are likely to be crucial for alcohol-induced osteoporosis and provide a new therapeutic approach for preventing osteoporosis.

The Contribution of Alcohol Dehydrogenase 3 to the Development of Alcoholic Osteoporosis in Mice.

BACKGROUND: Alcohol dehydrogenase 3 (ADH3) plays major roles not only in alcohol metabolism but also in nitric oxide metabolism as S-nitrosoglutathione reductase (GSNOR). ADH3/GSNOR regulates both adipogenesis and osteogenesis through the denitrosylation of peroxisome proliferator-activated receptor γ. The current study investigated the contribution of ADH3 to the development of alcoholic osteoporosis in chronic alcohol consumption (CAC). METHODS: Nine-week-old male mice of different ADH genotypes [wild-type (WT) and Adh3-/-] were administered a 10% ethanol solution for 12 months. The femurs were evaluated by histochemical staining and computed tomography-based bone densitometry. The mRNA levels of ADH3 were evaluated in the WT mice by reverse transcription-quantitative polymerase chain reaction. RESULTS: The Adh3-/- control mice exhibited increased activities of both osteoblasts and osteoclasts and lower bone masses than the WT control mice. CAC exhibited no remarkable change in osteoblastic and osteoclastic activities, but decreased bone masses were observed in WT mice despite an increase in the mRNA levels of ADH3. Conversely, bone masses in the Adh3-/- control mice were not reduced after CAC. CONCLUSIONS: The Adh3-/- control mice exhibited a high turnover of osteoporosis since osteoclastogenesis dominated osteoblastogenesis; however, bone resorption was not enhanced after CAC. In comparison, CAC lead to alcoholic osteoporosis in WT mice, accompanied by increased mRNA levels of ADH3. Hence, ADH3 can prevent osteoporosis development in normal ADH genotypes with no alcohol ingestion. However, ADH3 contributes to the development of alcoholic osteoporosis under CAC by participating in alcohol metabolism, increasing metabolic toxicity, and lowering GSNO reducing activity.

Metabolic pharmacokinetics of early chronic alcohol consumption mediated by liver alcohol dehydrogenases 1 and 3 in mice.

BACKGROUND AND AIM: Alcohol dehydrogenases (ADHs) 1 and 3 are responsible for systemic alcohol metabolism. The current study investigated the contribution of liver ADH1 and ADH3 to the metabolic pharmacokinetics of chronic alcohol consumption (CAC). METHODS: The 9-week-old male mice of different ADH genotypes (wild-type [WT], Adh1-/- , and Adh3-/- ) were administered with 10% ethanol solution for 1 month, followed by acute ethanol administration (4.0 g/kg). The alcohol elimination rate (AER), area under the blood alcohol concentration curve (AUC), and the maximum blood alcohol concentration (Cmax ) were calculated. The liver content, activity, and mRNA levels of ADH were evaluated. RESULTS: Chronic alcohol consumption increased the AER and reduced the AUC in all ADH genotypes. The increased ADH1 content was correlated with AER in WT mice but not in the Adh3-/- mice. Similarly, the increased ADH3 content was also correlated with AER in both WT and Adh1-/- mice. The Cmax was significantly higher in Adh3-/- control mice than in WT control mice. It decreased in the Adh1-/- mice by CAC along with an increase in the ADH3 content. CONCLUSIONS: Alcohol dehydrogenases 1 and 3 would accomplish the pharmacokinetic adaptation to CAC in the early period. ADH1 contributes to the metabolic pharmacokinetics of CAC with a decrease in AUC in conjunction with an increase of AER by increasing the enzyme content in the presence of ADH3. ADH3 also contributes to a decrease in AUC in conjunction with not only an increase in AER but also a decrease in Cmax by increasing the enzyme content.

Association between posttraumatic stress disorder (PTSD) severity and ego structure of the Nanai people.

BACKGROUND: A man-made chemical disaster occurred in the Amur River, leading to posttraumatic stress disorder (PTSD) in the Nanai people indigenous to the river's surrounding area. PTSD severity measured by the total scores of Impact of Event Scale-Revised (IES-R) (Total-I) and Clinician-Administered PTSD Scale (CAPS) (Total-C) were not always identical in terms of demographic and ethnocultural characters. It is possible that the results derived using the Total-I and Total-C may differ for persons with different backgrounds and/or individual characteristics. In this study, the associations between PTSD severity and personal characteristics were evaluated. METHODS: The study was a field-type survey including 187 randomly selected participants (75 males and 112 females). In addition to Total-I/Total-C, scores for each IES-R/CAPS item, Intrusion, Avoidance, and Hyperarousal, and Ego Structure Test by Ammon (ISTA) score were examined to evaluate their personal characteristics. RESULTS: No specific trends in ISTA score were obvious among four groups defined according to Total-I/Total-C. The results of principal component analysis showed that all IES-R/CAPS items contributed positively to the 1st axis but to the 2nd axis in a different manner. ISTA items did not always show correlations to each other, but principal component analysis suggested that Construct contributed positively and Destruct and Deficient (with the exception of Destruct sexuality) contributed negatively. High IES-R scores were associated with Construct Aggression and Deficient Inner demarcation, but high CAPS score was less likely to exhibit Construct Narcissism. CONCLUSION: To avoid the misdiagnosis of PTSD, usage of both IES-R/CAPS may be required. Simultaneous application of personality/ego tests may be helpful, but appropriate numbers of their questions would be important.

Genistein and menaquinone-4 treatment-induced alterations in the expression of mRNAs and their products are beneficial to osteoblastic MC3T3-E1 cell functions.

The aim of the present study was to determine the molecular basis of the beneficial effects of genistein and/or menaquinone­4 (MK­4) on bone quality. Initially, 1 µM genistein was applied to MC3T3­E1 cells for 24 h and the upregulated mRNAs that were detected by microarray were selected for further examination by reverse transcription­quantitative­polymerase chain reaction. Among them, alterations were observed in the level of GATA­binding protein 6 (GATA6), Notch gene homolog 2 (NOTCH2), Wnt family member 5A (WNT5A), bone γ­carboxyglutamate protein (BGLAP), chondroadherin (CHAD), dipeptidyl peptidase 4 (DPP4), ectonucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2), alkaline phosphatase (ALP) 3 and ATPase phospholipid­transporting 11A (ATP11A) in response to treatment with 0.1 µM 17­ß­estradiol, 1 µM genistein, and/or 1 µM MK­4. GATA6, NOTCH2 and WNT5A are considered to be associated with osteoclast, but not osteoblast, function; however, increases in osteoblastic mRNAs, including BGLAP and CHAD, were observed in each of the treatment groups at 48 h. Immunocytochemical analysis confirmed an increase in CHAD and DPP4 proteins following the administration of genistein + MK­4. Furthermore, genistein + MK­4 led to alterations in cell morphology to spindle or oval shapes, and increased the intensity of ALP staining. Although the level of ALP mRNA was not consistently altered in response to the treatments, a marked increase in ALP activity was observed following 96 h treatment with genistein + MK­4. Therefore, the simultaneous intake of genistein and MK­4 appears to be beneficial for the maintenance of bone quality.

Effect of vitamin K2 on the development of stress-induced osteopenia in a growing senescence-accelerated mouse prone 6 strain.

Vitamin K2 (VK2) has been used as a therapeutic agent for osteoporosis, since it has been suggested to be able to reduce the frequency of fractures by improving bone quality; however, bone turnover is strictly regulated by various cytokines and hormones. In the present study, the effect of menaquinone-4 (MK-4) on bone turnover was investigated using the senescence-accelerated mouse prone 6 (SAMP6) strain. Since water-immersion restraint stress (WRS) causes a significant decrease in bone mineral density (BMD), WRS was used as the bone resorption model in the SAMP6 strain. Six-week-old SAMP6 male mice were divided into the following three groups: Control, WRS and WRS + MK-4. WRS was performed for 6 h per day, 5 times a week, for 4 weeks. Following WRS, MK-4 (30 mg/kg) was injected subcutaneously 3 times a week for 4 weeks. No growth retardation was observed in the WRS groups as compared with the control group. In the WRS groups, the BMD was significantly lower than that in the control group. The levels of bone formation and resorption markers were increased in the WRS groups, indicating that WRS reduced the BMD by promoting high bone turnover. A bone histomorphometrical examination showed that the trabecular (Tb) bone mass in the secondary spongiosa at the distal femur was significantly reduced in the WRS mice, and this reduction was abrogated by MK-4 treatment. Specifically, the Tb bone reduction was caused by the activation of osteoclasts (Ocs), and Oc activity was suppressed by MK-4. The number of osteoblasts and the mineral apposition rate were significantly increased in the WRS and WRS + MK-4 mice, suggesting that WRS triggered a significantly higher mineral apposition rate. These results indicate that MK-4 can induce recovery from the bone mineral loss caused by WRS treatment. Further studies are required to clarify the association between bone quality and MK-4.

Factors associated with tuberculosis cases in Semarang District, Indonesia: case-control study performed in the area where case detection rate was extremely low.

OBJECTIVES: Indonesia is ranked as the 4th highest contributor to tuberculosis (TB) in the world. Semarang District in Central Java displays extremely low case detection rate (CDR), possibly contributing to the local prevalence of TB. METHODS: A case-control study was performed to explore the factors that cause such low CDR. We recruited 129 TB cases and 83 controls that visited the same centers and were not diagnosed with TB. RESULTS: The cases had 7.5 ± 2.3 symptoms/person on average, indicating the delay in diagnosis because the controls only displayed 1.0 ± 1.7. The multiple logistic regression analysis comparing the cases/controls extracted following factors as a risk to have TB: farmer, close contact with TB patients, ignorance of whether Bacillus Calmette-Guérin (BCG) was accepted or no, smoking, low income, a lot of people living in the same room, irregular hand wash before meals, not wash hands after blow, soil floor, and no sunlight and no ventilation in the house. CONCLUSIONS: Neither the cases nor the controls knew the symptoms and how to avoid TB infection, which probably caused the delay in diagnosis. It is difficult to change the current living conditions. Thus, the amendment of the community-based education program of TB seems to be required.

Regulation of serine protease inhibitor Kazal type-5 (SPINK5) gene expression in the keratinocytes.

OBJECTIVES: Serine protease inhibitor Kazal type-5 (SPINK5) plays a crucial role in deciding the timing of desquamation of the skin. Its gene expression is limited at the very surface of the stratum granulosum (SG), whereas expression of kallikreins (KLKs) encoding proteases is usually found throughout the stratum spinosum and SG. METHODS: To explore the difference in expression regulation of these proteases/inhibitors, the function of SPINK5 promoter was examined using luciferase assay. RESULTS: Luciferase assay targeting the SPINK5 promoters (nucleotide -676/-532 and -318/-146 from the major transcription start site) showed high intensity in NHEK human keratinocyte. These two sites had neither common cis-elements nor GATA3 element but electrophoretic mobility shift assay showed similar retardation bands. Moreover, DNA footprinting did not display specific protected bands. Thus, we could not identify cis-element(s) that controlled these elements. Differentiation induced by high Ca(2+) medium failed to alter their luciferase activities. Transfection of GATA3 expressing vector significantly but slightly increased them and that of vector expressing its dominant negative form decreased. CONCLUSIONS: Although GATA3 is reportedly important for inhibition of proliferation and induction of differentiation of keratinocytes, its effect on SPINK5 expression was indirect and GATA3 alone was insufficient for final differentiation of keratinocytes where full SPINK5 expression was observed.

Posttraumatic stress disorders in the Nanai after pollution of the Amur River: ethnocultural analysis.

OBJECTIVES: Chemical pollution of the Amur River has seriously damaged traditions and caused posttraumatic stress disorder (PTSD) among the Nanai, the indigenous people living along this river. This study was performed to clarify the ethnographic characteristics of PTSD in this unique population. METHODS: The study group consisted of 75 male and 112 female randomly selected volunteers. PTSD severity measured using scores of the Impact of Event Scale--Revised (Total-I) and Clinical-Administered PTSD Scale (Total-C) was estimated according to demographic and ethnocultural backgrounds, clinical status, and ethnopsychological attitudes toward the Amur River. RESULTS: The differences in averages of Total-I and Total-C were not always the same in the groups divided by ethnographic information. Logistic regression analysis with a dependent variable, possibly without PTSD (Total-I <34 and Total-C <40)/possibly with PTSD (either Total-I ≥34 or Total-C ≥40), and categorical independent variables using ethnographic information extracted a low score when 'priority values' and 'the Amur River for me is' was "profession" and a high score when 'dominant role in spousal relationship' was "self," when 'predominant forms of response in stressful situations' was "try to organize," when 'preferred method of medical treatment' was specific for the Nanai, when "rely on something mystical" was manifested, and when the Amur River was believed to be "sacred". CONCLUSION: Those with a pragmatic attitude were less likely to have PTSD. However, those who were required to make decisions within close relationships and were intimate with the Nanai tradition and the Amur River had increased likelihood of PTSD.