Ryosuke FUJII

BACKGROUND: Current evidence suggested an increased risk for cancer mortality in those with low AHRR DNAm levels. Therefore, AHRR DNAm could identify a more "fragile" group at risk for cancer mortality than questionnaire-based evaluations. Given this, the aim was to identify "fragile" groups at risk of cancer mortality by integrating questionnaire-based smoking indices and leukocyte AHRR DNAm levels in the Japanese population. METHODS: The target population was 795 participants without a clinical history who underwent a health check-up in 1990. They were followed for up to 30 years for mortality. The AHRR DNAm levels in leukocytes were measured by the pyrosequencing method. Hazard ratios (HRs) for cancer mortality were calculated using a Cox proportional hazards model. RESULTS: Significantly higher HRs for all-cancer mortality were observed in low AHRR DNAm groups regardless of smoking intensity (Pack-years<20: 3.69 [1.46-9.37], Pack-years≥20: 2.13 [1.11-4.09]). Compared to current smokers, significantly lower HRs for all-cancer mortality were observed in the group with high AHRR DNAm regardless of years since quitting (YSQ) (YSQ≤10: 0.13 [0.02-0.96], YSQ>10: 0.28 [0.08-0.98]). Even for YSQ greater than 10, there was no significant mortality risk reduction in the low AHRR DNAm group. CONCLUSIONS: The population with low AHRR DNAm levels had a higher risk of cancer mortality even with low smoking exposure. Furthermore, no significant risk reduction was observed in former smokers with low AHRR DNAm levels. IMPACT: AHRR DNAm levels in leukocytes may help identify groups at risk for cancer mortality overlooked by questionnaire-based smoking indices.

AIMS/INTRODUCTION: Previous observational studies have suggested an increased risk of type 2 diabetes associated with both short and long sleep duration. However, there remains uncertainty, particularly regarding the adverse effects of long sleep duration. We investigated the association between self-reported questionnaire-based and objectively measured accelerometer-derived sleep duration and the risk of type 2 diabetes using data from the UK Biobank. MATERIALS AND METHODS: First, we performed conventional Cox regression analysis with restricted cubic splines to illustrate the potentially non-linear association between sleep duration and the risk of type 2 diabetes. Second, we performed non-linear Mendelian randomization (MR) analysis using the doubly-ranked method with 85 and 20 genetic variants associated with questionnaire-based and accelerometer-based sleep duration, respectively. Third, we performed two-sample MR analysis. RESULTS: The results of conventional analysis of accelerometer-derived sleep duration did not suggest a strong association between longer sleep duration and type 2 diabetes risk (hazard ratio [HR] of ≥10 h compared with 7-8 h, 1.08; 95% confidence interval [CI], 0.92-1.27). The results of non-linear MR showed no strong evidence for an increased risk of type 2 diabetes associated with questionnaire-based longer sleep duration (HR of 9 h compared with 7 h, 0.77; 95% CI, 0.52-1.15). This finding was consistent with non-linear MR of accelerometer-derived sleep duration (HR of 9 h compared with 7 h, 0.78; 95% CI, 0.29-2.06). CONCLUSIONS: Our findings suggest that longer sleep duration does not play a major role in the development of type 2 diabetes.

BACKGROUND: Although utility of composite trait-specific polygenic risk score (multi-trait PRS) has been examined among European ancestries, few studies investigated among East Asians and incorporated modifiable risk factors. We examined the associations of multi-trait PRS for cardiometabolic factors with cardiovascular disease mortality by integrating nongenetic determinants. METHODS: A total of 14 086 Japanese participants (mean age, 55±9; 55.8% women) of the J-MICC (Japan Multi-Institutional Collaborative Cohort) study were analyzed in this study. We calculated 6 PRSs for cardiometabolic traits (systolic blood pressure, body mass index, triglycerides, low-density lipoprotein cholesterol, estimated glomerular filtration rate, and hemoglobin A1c). Based on these PRSs, we developed multi-trait PRS and considered as a primary exposure. Three nongenetic factors (smoking, alcohol drinking, and educational attainment) from the self-reported questionnaire were also examined. RESULTS: During a median 12.1-year follow-up period, a total of 472 all-cause and 79 cardiovascular disease mortality cases were documented. Compared with 0% to 90% of multi-trait PRSs, an adjusted hazard ratio (HR) among the top 10% of multi-trait PRSs was 1.32 (95% CI, 1.00-1.73) for all-cause death and 2.63 (95% CI, 1.48-4.67) for cardiovascular disease death. Incorporation of educational attainment with multi-trait PRSs showed null associations in those who went beyond high school (HR, 2.07 [95% CI, 0.44-9.66]) even in the top 10% of multi-trait PRS. CONCLUSIONS: Our analysis combining both genetic and nongenetic determinants highlighted that lifestyle factors and educational attainment can slightly reduce an individual's composite genetic risk for cardiovascular disease death.

BACKGROUND: Mitochondria, which harbor their own genome (mtDNA), have attracted attention due to the potential of mtDNA copy number (mtDNA-CN) as an indicator of mitochondrial dysfunction. Although mtDNA-CN has been proposed as a simple and accessible biomarker for metabolic disorders such as metabolic dysfunction-associated steatotic liver disease, the underlying mechanisms and the causal relationship remain insufficiently elucidated. In this investigation, we combined longitudinal epidemiological data, animal studies, and in vitro assays to elucidate the potential causal relationship between reduced mtDNA-CN and the development of steatotic liver disease (SLD). METHODS: We conducted a longitudinal study using data from a health examination cohort initiated in 1981 in Yakumo, Hokkaido, Japan. Data from examinations performed in 2015 and 2022 were analyzed, focusing on 76 subjects without SLD at baseline (2015) to assess the association between baseline mtDNA-CN and subsequent risk of SLD development. In addition, 28-day-old SD rats were fed ad libitum on a 45% high-fat diet and dissected at 2 and 8 weeks of age. Blood and liver mtDNA-CN were measured and compared at each feeding period. Additionally, in vitro experiments were performed using HepG2 cells treated with mitochondrial function inhibitors to induce mtDNA-CN depletion and to examine its impact on intracellular lipid accumulation. RESULTS: Epidemiological analysis showed that the subjects with low mtDNA-CN had a significantly higher odds ratio for developing SLD compared to high (odds ratio [95% confidence interval]: 4.93 [1.08-22.50]). Analysis of the animal model showed that 8 weeks of high-fat diet led to the development of fatty liver and a significant decrease in mtDNA-CN. A further 2 weeks of high-fat diet consumption resulted in a significant decrease in hepatic mtDNA-CN, despite the absence of fatty liver development, and a similar trend was observed for blood. Complementary in vitro experiments revealed that pharmacologically induced mitochondrial dysfunction led to a significant reduction in mtDNA-CN and was associated with increases in intracellular lipid accumulation in HepG2 cells. CONCLUSIONS: Our findings suggest that reduced mtDNA-CN may contribute causally to SLD development and could serve as a convenient, noninvasive biomarker for early detection and risk assessment.

Chronic kidney disease (CKD) is a complex disease affecting >10% of the global population, with large between- and within-continent variability reflecting major environmental determinants. To identify molecular targets for treatment and prevention, genome-wide association study meta-analyses (GWAMAs) of CKD-defining traits have identified hundreds of genetic loci in aggregated population samples. However, while GWAMAs estimate the average allelic effect across studies, single population studies may be relevant to unravel specific mechanisms. To assess whether a study sample from a specific population could extend existing knowledge on kidney function genetics, we selected 147 kidney function relevant loci identified by a large GWAMA, assessing their association with the glomerular filtration rate estimated from serum creatinine (eGFRcrea) in 10,146 participants to the Cooperative Health Research In South Tyrol (CHRIS) study, conducted in an Alpine region where thyroid dysfunction is common. We identified associations with single nucleotide polymorphisms (SNPs) at 11 loci, showing up-to-5.4 times larger effect sizes than in the corresponding GWAMA, not explainable by allele frequency differences. Systematic mediation analysis across 70 quantitative traits identified serum magnesium and the activated partial thromboplastin time as partial mediators of the eGFRcrea associations at SHROOM3 and SLC34A1, respectively. Given that free triiodothyronine and thyroxine acted as effect modifiers across all loci, we conducted SNP-by-thyroid stimulating hormone (TSH) interaction analyses, identifying significant interactions at STC1: SNPs had larger effects on eGFRcrea at higher TSH levels, possibly reflecting stanniocalcin-1 autocrine and paracrine role. Individual population studies can help characterize genetic associations. The interplay between phenotypes at SHROOM3 and SLC34A1 and the role of thyroid function as a genetic effect modifier warrant further investigations.

BACKGROUND: Although serum carotene may contribute to dementia prevention, there is a lack of longitudinal evidence for early cognitive decline before dementia symptoms. The aim of this study was to examine whether serum carotene levels were associated with annually evaluated cognitive trajectories among the Japanese general population. METHODS: Among 581 baseline participants, 199 individuals (83 males; mean age [min, max], 62.7 [39, 90] years) who underwent cognitive assessments more than twice after baseline were analyzed. "Attention" levels were assessed using one- and three-target Digit Cancellation Tests (D-CAT1 and D-CAT3). "General cognitive ability" was assessed by the short version of Mini-Mental State Examination (SMMSE). Serum carotenes (α-carotene, β-carotene and lycopene) were measured by high-performance liquid chromatography. After the measurements, we calculated total carotene levels by summing up the levels of all measured carotene. Carotene levels were categorized into three groups for analysis (low: 0%-25%, middle: 25%-75%, and high: 75%-100%). A linear mixed model was used to estimate the slope of the D-CAT score trajectory and to compare it between three categories. RESULTS: Compared with the middle carotene group, decline of attention was faster in the D-CAT1 for low β-carotene (β = -3.48, p = 0.035), lycopene (β = -3.10, p = 0.062), and total carotene (β = -4.75, p = 0.003), but not for α-carotene (β = -2.60, p = 0.111). For the D-CAT3, decline of attention was faster in the group of low lycopene (β = -3.17, p = 0.002) and total carotene (β = -2.17, p = 0.037) compared with the middle carotene group, while no clear association for α-carotene (β = -0.67, p = 0.521) and β-carotene (β = -0.64, p = 0.639). There were no clear associations between serum carotene and the SMMSE score. CONCLUSIONS: These findings suggest low levels of serum lycopene are associated with a decline of attention in the setting of the general population.

BACKGROUND: Among the more than 300 mycotoxins that are known to have toxic effects on animals and humans, Fusarium toxins deoxynivalenol (DON), T-2 and HT-2 toxins (T2/HT2), and zearalenone (ZEN) are frequently detected in domestic agricultural products. This study aimed to assess DON, T2/HT2, and ZEN exposure in Japanese adults by measuring urinary mycotoxins, observing their distributions, and making comparisons with data from other countries. METHODS: A total of 201 individuals participated in the study. Twenty-four-hour urine samples were collected from young adults (34 men and 35 women) in the Tokai region (urban area) and spot urine samples were collected from middle-aged and elderly adults (64 men and 68 women) in the Donan area of Hokkaido Prefecture (rural area). Urinary DON, T2/HT2, and ZEN levels were measured using a validated liquid chromatography-tandem mass spectrometry method. RESULTS: For DON, T2/HT2, and ZEN, the detection frequencies above the limit of detection (LOD) level (0.15, 0.13, and 0.01 µg/L, respectively) in all the samples were 53%, 26%, and 71%, respectively. The median concentrations (95th percentile) of urinary DON, HT2, and ZEN were 0.19 (3.93), <LOD (0.55), and 0.02 (0.12) µg/L, respectively. Although at least one of the investigated mycotoxins was detected in the urine of 86% of study participants, the concentrations were similar to or lower than those found in other countries (from 19 reports within the past decade). Moreover, the probable daily intake (PDI) values in the present study were lower than the provisional maximum tolerable daily intake levels. The urinary mycotoxin levels did not significantly differ with respect to sex, age, or occupation. CONCLUSIONS: This study represents the first comprehensive exposure assessment for DON, T2/HT2, and ZEN in Japanese adults using human biomonitoring methods. These data provide valuable information for a better understanding of mycotoxin exposure in Japan.

BACKGROUND: Incidence of ischemic stroke increased after natural disasters. Therefore, it is important to establish a means of identifying high-risk populations for incident stroke. We performed a prospective cohort study to examine whether these three cardiovascular disease-related miRNAs (miR-126, miR-197, and miR-223) are associated with incident stroke among elderly survivors of the Great East Japan Earthquake. METHOD: This cohort study was conducted using the data of 1192 survivors of the Great East Japan Earthquake over 60-years old who underwent a health check-up in December 2011. We followed up participants to record stroke cases until the end of 2016. We measured serum miRNAs by quantitative real-time polymerase chain reaction. HRs for incident stroke were estimated by Cox proportional hazard regression analyses. RESULT: The serum miR-197 level was significantly associated with the incident stroke; the HR per one standard deviation change in the miR-197 level was 1.65 (95% confidence interval: 1.19 - 2.30). In contrast, the levels of miR-126 and miR-223 were not associated with the incident stroke. CONCLUSION: We found that a higher miR-197 level is associated with an increased risk of incident stroke; thus, miR-197 is expected to be useful as a predictive biomarker.

Although previous polygenic risk score (PRS) studies for cardiovascular disease (CVD) focused on incidence, few studies addressed CVD mortality and quantified risks by environmental exposures in different genetic liability groups. This prospective study aimed to examine the associations of blood pressure PRS with all-cause and CVD mortality and to quantify the attributable risk by modifiable lifestyles across different PRS strata. 9,296 participants in the Japan Multi-Institutional Collaborative Cohort Study without hypertension at baseline were analyzed in this analysis. PRS for systolic blood pressure and diastolic blood pressure (PRSSBP and PRSDBP) were developed using publicly available Biobank Japan GWAS summary statistics. CVD-related mortality was defined by the International Classification of Diseases 10th version (I00-I99). Cox-proportional hazard model was used to examine associations of PRSs and lifestyle variables (smoking, drinking, and dietary sodium intake) with mortality. During a median 12.6-year follow-up period, we observed 273 all-cause and 41 CVD mortality cases. Compared to the middle PRS group (20-80th percentile), adjusted hazard ratios for CVD mortality at the top PRS group ( > 90th percentile) were 3.67 for PRSSBP and 2.92 for PRSDBP. Attributable risks of CVD mortality by modifiable lifestyles were higher in the high PRS group ( > 80th percentile) compared with the low PRS group (0-80th percentile). In summary, blood pressure PRS is associated with CVD mortality in the general Japanese population. Our study implies that integrating PRS with lifestyle could contribute to identify target populations for lifestyle intervention even though improvement of discriminatory ability by PRS alone is limited.

INTRODUCTION: Previous Mendelian randomization (MR) studies for the coffee-kidney association have reported inconsistent relationships in European populations and never examined mediators of this association. We aimed to evaluate this causal relationship using two-sample MR among both East Asian and European ancestries and to explore underlying mechanisms using plasma caffeine levels. METHODS: Among East Asians, the largest genome-wide association study (GWAS) results for coffee intake, plasma caffeine levels, and kidney outcomes were obtained from 152,634; 8940; and 47,070 Japanese adults. Among Europeans, summary statistics were acquired from European GWAS with 428,860; 7719; and 564,470 adults for each trait. We applied different MR methods (inverse-variance weighted [IVW] with random effects, weighted median, weighted mode, and MR-Egger). RESULTS: After excluding possible pleiotropic variants, among East Asian ancestry, drinking an extra coffee intake per week showed a protective association on serum creatinine-based estimated glomerular filtration rate (eGFRcre) (β = 0.077; 95% confidence interval [CI] = 0.003 to 0.150). Analysis in European ancestry also showed a causal relationship between drinking an extra coffee intake per day and eGFRcre (β = 0.052; 95% CI = 0.027 to 0.078). These results were consistent across different MR methods accounting for invalid instruments. Higher plasma caffeine levels were associated with lower eGFRcre among both East Asian (β = -0.071; 95% CI = -0.137 to -0.006) and European ancestries (β = -0.048; 95% CI = -0.057 to -0.040). CONCLUSIONS: Our cross-ancestry MR study found beneficial effects of coffee intake on eGFRcre. However, given the possible adverse effects of plasma caffeine levels on eGFRcre, interpretation of the results should be carefully considered and further investigations on noncaffeine and biological pathways are needed.

Thioredoxin-interacting protein (TXNIP) plays an important role in glucose metabolism, and its expression is regulated by DNA methylation (DNAm). Although the association between TXNIP DNAm and type 2 diabetes mellitus has been demonstrated in studies with a cross-sectional design, prospective studies are needed. We therefore examined the association between TXNIP DNAm levels and longitudinal changes in glycemic traits by conducting a longitudinal study involving 169 subjects who underwent two health checkups in 2015 and 2019. We used a pyrosequencing assay to determine TXNIP DNAm levels in leukocytes (cg19693031). Logistic regression analyses were performed to assess the associations between dichotomized TXNIP DNAm levels and marked increases in glycemic traits. At four years, the TXNIP DNA hypomethylation group had a higher percentage of changes in fasting plasma glucose (FPG) and hemoglobin A1c (HbA1c) compared to those in the hypermethylation group. The adjusted odds ratios for FPG and HbA1c levels were significantly higher in the TXNIP DNA hypomethylation group than in the hypermethylation group. We found that TXNIP DNA hypomethylation at baseline was associated with a marked increase in glycemic traits. Leukocyte TXNIP DNAm status could potentially be used as an early biomarker for impaired glucose homeostasis.

The renal angina index (RAI) is a validated scoring tool for predicting acute kidney injury (AKI). We investigated the efficacy of the RAI in 2436 heterogeneous patients (mean age, 70 years) treated in cardiac intensive care units (CICUs). The RAI was calculated from creatinine and patient condition scores. AKI was diagnosed by the Kidney Disease: Improving Global Outcome criteria. The primary and secondary endpoints were the development of severe AKI and all-cause mortality, respectively. Four hundred thirty-three patients developed AKI, 87 of them severe. In multivariate analyses, the RAI was a significant independent predictor of severe AKI. During the 12-month follow-up period, 210 patients suffered all-cause death. Elevated RAI was independently associated with all-cause mortality, as was NT-proBNP (p < 0.001). The RAI is a potent predictor not only of severe AKI but also of adverse outcomes and substantially improved the 12-month risk stratification of patients hospitalized in CICUs.

OBJECTIVES: The mitochondrial DNA (mtDNA) is unique and circular with multiple copies of the genome. The lower mtDNA copy number (mtDNA-CN) in leukocytes is associated with the risk of all-cause mortality. However, its long-term association is unknown. Thus, the study examined the association between mtDNA-CN and the risk of all-cause mortality in a long-term follow-up study in the Japanese population. DESIGN: This longitudinal study included the study cohort from an annual, population-based health checkup in the town of Yakumo, Hokkaido, Japan. SETTING AND PARTICIPANTS: 814 participants (baseline age range: 38-80 years, mean: 56.3 years) were included in this study in 1990. They were followed-up regarding mortality for about 30 years (median: 28.1 years) till 2019. MEASURES: The genomic DNA was extracted from peripheral blood mononuclear cells and the mtDNA-CN was measured using real-time polymerase chain reaction. The level of the mtDNA-CN was divided into tertiles (low, middle, and high). The participants were categorized based on their age into middle-aged (<60 years old) or old-aged (≥60 years old). Survival analysis was performed for tertile of mtDNA-CN and compared using the log-rank test. Univariate and multivariable Cox proportional hazard regression analyses were performed to assess the association between mtDNA-CN and all-cause mortality. The model adjusted with age, sex, body mass index, systolic blood pressure, smoking habit, alcohol consumption, exercise habit, and education level. RESULTS: The low levels of mtDNA-CN resulted in a significant decrease in cumulative survival rate (P <  0.05). The risk of mortality was significantly higher in the middle-aged cohort when mtDNA-CN levels were low (hazard ratios [95% confidence intervals]: 1.98 [1.10-3.56]). CONCLUSION: This study demonstrated that leukocyte mtDNA-CN is associated with future mortality risk. Our study findings may lead to further research on the early prediction of mortality and its underlying mechanisms.

OBJECTIVE: The association between knee osteoarthritis (OA) and miRNAs has been widely reported. However, the utility of miRNAs as predictors of knee osteoarthritis (KOA) progression in longitudinal studies has not been reported. We aimed to identify circulating miRNAs (c-miRNAs) associated with KOA progression in the general population and to examine their potential use as predictors of KOA progression. METHODS: In 2012 and 2018, 66 participants (128 knees) took part in a resident health check-up in the Yakumo study. If the KL classification progressed two or more levels, the patient was classified as having progressive OA. Quantitative real-time polymerase chain reaction was used to screen 21 c-miRNAs. The expression levels of those c-miRNAs were compared between the progressive OA group and non-progressive OA group using student-t-test. Logistic analysis was performed in c-miRNAs less than p < 0.10 in univariate analysis. RESULTS: The progressive OA group consisted of 78 knees. The results of the comparison between the progressive OA group and the non-progressive OA group showed that six c-miRNAs as follows; let7d (p = 0.030), c-miRNA-122 (p < 0.001), 150 (p = 0.070), 199 (p = 0.078), 21 (p = 0.016) and 320 (p = 0.093) were extracted as factors related to the progression of knee OA. In addition, logistic regression analysis identified c-miRNA-122 as an independent factor involved in the progression of knee osteoarthritis (odds ratio: 1.510, 95% confidence interval: 1.060-2.140, p = 0.023). The ROC curve showed by c-miRNA-122 for the progression of OA risk had an area under the curve of 0.702 (95% CI: 0.609-0.795). The threshold of c-miRNA-122 was -4.609. CONCLUSION: The expression level of c-miRNA-122 was associated with the risk of KOA progression in community dwelling Japanese people.

Background: Carotenoids have been reported to exert protective effects against age-related diseases via changes in DNA methylation. Although lower thioredoxin-interacting protein (TXNIP) DNA methylation is associated with age-related diseases, only a few studies have investigated the factors influencing TXNIP DNA methylation. Carotenoids may be a factor linking TXNIP to specific pathophysiological functions. The aim of this study was to examine whether serum carotenoid levels are associated with TXNIP DNA methylation levels. Methods: We conducted a cross-sectional study using 376 health examination participants (169 men). DNA methylation levels were determined using a pyrosequencing assay. Serum carotenoid levels were determined by high-performance liquid chromatography. Multivariable regression analyses were performed to examine the associations between TXNIP DNA methylation levels and serum carotenoid levels with adjustment for age, BMI, HbA1c, CRP, smoking habits, alcohol consumption, exercise habits, and percentage of neutrophils. Results: Multiple linear regression analyses showed that TXNIP DNA methylation levels were positively associated with serum levels of zeaxanthin/lutein (β [95%CI]: 1.935 [0.184, 3.685]), β-cryptoxanthin (1.447 [0.324, 2.570]), α-carotene (1.061 [0.044, 2.077]), β-carotene (1.272 [0.319, 2.226]), total carotenes (1.255 [0.040, 2.469]), total xanthophylls (2.133 [0.315, 3.951]), provitamin A (1.460 [0.402, 2.519]), and total carotenoids (1.972 [0.261, 3.683]) in men (all p<0.05). Of these, provitamin A showed the stronger association (standardized β=0.216). No significant association of TXNIP DNA methylation and serum carotenoid was observed in women. Conclusions: The findings of this study suggest that carotenoid intake may protect against age-related diseases by altering TXNIP DNA methylation status in men.

BACKGROUND: Epigenetic studies have reported relationships between dietary nutrient intake and methylation levels. However, genetic variants that may affect DNA methylation (DNAm) pattern, called methylation quantitative loci (mQTL), are usually overlooked in these analyses. We investigated whether mQTL change the relationship between dietary nutrient intake and leukocyte DNAm levels with an example of estimated fatty acid intake and ATP-binding cassette transporter A1 (ABCA1). METHODS: A cross-sectional study on 231 participants (108 men, mean age: 62.7 y) without clinical history of cancer and no prescriptions for dyslipidemia. We measured leukocyte DNAm levels of 8 CpG sites within ABCA1 gene by pyrosequencing method and used mean methylation levels for statistical analysis. TaqMan assay was used for genotyping a genetic variant of ABCA1 (rs1800976). Dietary fatty acid intake was estimated with a validated food frequency questionnaire and adjusted for total energy intake by using residual methods. RESULTS: Mean ABCA1 DNAm levels were 5% lower with the number of minor alleles in rs1800976 (CC, 40.6%; CG, 35.9%; GG, 30.6%). Higher dietary n-3 PUFA intake was associated with lower ABCA1 DNAm levels (1st (ref) vs. 4th, β [95% CI]: -2.52 [-4.77, -0.28]). After controlling for rs180076, the association between dietary n-3 PUFA intake and ABCA1 DNAm levels was attenuated, but still showed an independent association (1st (ref) vs. 4th, β [95% CI]: -2.00 [-3.84, -0.18]). The interaction of mQTL and dietary n-3 PUFA intake on DNAm levels was not significant. CONCLUSIONS: This result suggested that dietary n-3 PUFA intake would be an independent predictor of DNAm levels in ABCA1 gene after adjusting for individual genetic background. Considering mQTL need to broaden into other genes and nutrients for deeper understanding of DNA methylation, which can contribute to personalized nutritional intervention.

Background: The increasing prevalence of non-alcoholic fatty liver disease (NAFLD) has become a global health problem. NAFLD has few initial symptoms and may be difficult to detect early, so there is need for a minimally invasive early detection marker. We hypothesized that miR-122 and miR-20a levels combined, as the miR-122/miR-20a ratio might detect NAFLD more sensitively. Methods: This study involved 167 participants with low alcohol intake. Those who had an increase in echogenicity of the liver parenchyma and hepato-renal contrast on ultrasonography were classified as the NAFLD group (n = 44), which was further classified into mild (n = 26) and severe (n = 18) groups based on echogenic intensity and hepatic vessel and diaphragm visualization. Participants without fatty liver were included in the normal group, except for those with an abnormal body mass index, glycated hemoglobin, and systolic blood pressure (n = 123) values. Serum miR-122 and miR-20a expression levels in participants were measured by real-time polymerase chain reaction, and the miR-122/miR-20a was calculated. Results: In the NAFLD group, miR-122 expression was significantly higher and the miR-20a was significantly lower than in the normal group, in agreement with previous studies. miR-122/miR-20a was also significantly higher in the NAFLD group. Receiver operating characteristic curve analysis was performed with miR-122/miR-20a as an NAFLD detection marker, and the area under the curve of miR-122/miR-20a was significantly larger than that of miR-122 or miR-20a alone. Conclusions: The miR-122/miR-20a ratio, combined with miR-122 and miR-20a levels, is a useful biomarker to detect NAFLD with high sensitivity.

OBJECTIVES: The adverse health effects of consuming sugar-sweetened beverages have been studied worldwide. However, no recent report on the actual sugar contents of Japanese sugar-sweetened beverages is available. Therefore, we analyzed the glucose, fructose, and sucrose contents of common Japanese beverages. METHODS: The glucose, fructose, and sucrose contents of 49 beverages (8 energy drinks, 11 sodas, 4 fruit juices, 7 probiotic drinks, 4 sports drinks, 5 coffee drinks, 6 green tea drinks, and 4 black tea drinks) were determined using enzymatic methods. RESULTS: Three zero calorie drinks, 2 sugarless coffee drinks, and 6 green tea drinks contained no sugar. Three coffee drinks contained only sucrose. The orders of median glucose, fructose, and sucrose contents in the categories of beverages containing sugars were as follows: for glucose, fruit juice > energy drink ≥ soda ≫ probiotic drink > black tea drink > sports drink; for fructose, probiotic drink ≥ energy drink > fruit juice > soda ≫ sports drink > black tea drink; and for sucrose, black tea drink > energy drink ≥ probiotic drink > fruit juice > soda > coffee drink ≫ sports drink. The total fructose as a percentage of the total sugar content in the 38 sugar-containing beverages was between 40% and 60%. The total sugar content analyzed was not always equivalent to the carbohydrate content indicated on the nutrition label. CONCLUSIONS: These results indicate that information on the actual sugar content of common Japanese beverages is necessary for the exact assessment of beverage-derived sugar intake.

BACKGROUND: Previous studies have proposed different formulas of estimating glomerular filtration rate (eGFR) among clinical patients. The comprehensive comparison of eGFR formulas is not well established in a Japanese population. We compared eGFR values and chronic kidney disease (CKD) classification of nine different eGFR in a Japanese general population sample. METHODS: We analyzed 469 Japanese community-dwelling adults (184 men) without any self-reported kidney disease. GFR estimated using the 4- and 6-parameter Modification of Diet in Renal Disease (MDRD) formulas (MDRD4 and MDRD6); the CKD-EPI formulas based on creatinine with (CKD-EPI-2009) and without race coefficient (CKD-EPI-2021), on cystatin C (CKD-EPI-Cys), on both (CKD-EPI-CreCys); the Japanese creatinine-based formula (JPN-Cre), cystatin C-based formula (JPN-Cys), and modified CKD-EPI formula (JPN-CKD-EPI). CKD stages were defined by KDIGO guidelines (eGFR < 60 ml/min/1.73 m2). RESULTS: eGFRJPN-Cre (mean = 71.2; SD = 14.3) were much lower than eGFRCKD-EPI-2021 (mean = 94.2; SD = 12.7), while eGFRJPN-Cys (mean = 102.8; SD = 24.2) was comparable to the MDRD and CKD-EPI formulas. The difference between eGFRCKD-EPI-2021 and eGFRJPN-Cre showed a V-shaped distribution across eGFR levels, indicating complex errors between these formulas. We observed very low agreement in CKD classification between eGFRJPN-Cre and the eGFRCKD-EPI-2021 (kappa = 0.13; 95% confidence interval: 0.06, 0.23). CONCLUSIONS: JPN-Cre was substantially different from the CKD-EPI formula without race term (CKD-EPI-2021), which means that it is impossible to recalibrate those with a simple coefficient. Although a comparison with measured GFR should be necessary, choice of the estimation method needs caution in clinical decision-making and academic research.

The consumption of high-fructose corn syrup (HFCS) has been increasing in recent decades, especially among children. Some reports suggest that children and adolescents are more sensitive to the adverse effects of fructose intake than adults. However, the underlying mechanism of the difference in vulnerability between adolescence and adulthood have not yet been elucidated. In this study, we attempted to elucidate the different effects of HFCS intake at different growth stages in rats: childhood and adolescence (postnatal day (PD) 21-60), young adulthood (PD60-100), and adulthood (PD100-140). Since alterations in hepatic glucocorticoid (GC) metabolism can cause diseases including insulin resistance, we focused on GC metabolizing enzymes such as 11 beta-hydroxysteroid dehydrogenase 1 and 2 (Hsd11b1 and Hsd11b2) and steroid 5 alpha-reductase 1 (Srd5a1). Western blotting showed an increase in Hsd11b1 expression and a decrease in Hsd11b2 expression in childhood and adolescence but not in adulthood. We also observed changes in Hsd11b1 and Hsd11b2 activities only in childhood and adolescence, consistent with the results of mRNA and protein expression analysis. The effect of high-fructose intake with regards to GC metabolism may therefore vary with developmental stage. This study provides insight into the adverse effects of fructose on GC metabolism in children in the context of increasing rates of HFCS consumption.

DNA methylation can be affected by numerous lifestyle factors, including diet. Tobacco smoking induces aryl hydrocarbon receptor repressor (AHRR) DNA hypomethylation, which increases the risk of lung and other cancers. However, no lifestyle habits that might increase or restore percentage of AHRR DNA methylation have been identified. We hypothesized that dietary intakes of vegetables/fruits and serum carotenoid concentrations are related to AHRR DNA methylation. A total of 813 individuals participated in this cross-sectional study. A food frequency questionnaire was used to assess dietary intake of vegetables and fruits. AHRR DNA methylation in peripheral blood mononuclear cells were measured using pyrosequencing method. In men, dietary fruit intake was significantly and positively associated with AHRR DNA methylation among current smokers (P for trend = .034). A significant positive association of serum provitamin A with AHRR DNA methylation was observed among current smokers (men: standardized β = 0.141 [0.045 to 0.237], women: standardized β = 0.570 [0.153 to 0.990]). However, compared with never smokers with low provitamin A concentrations, percentages of AHRR DNA methylation were much lower among current smokers, even those with high provitamin A concentrations (men: β = -19.1% [-33.8 to -19.8], women: β = -6.0% [-10.2 to -1.7]). Dietary intake of vegetables and fruits rich in provitamin A may increase percentage of AHRR DNA methylation in current smokers. However, although we found a beneficial effect of provitamin A on AHRR DNA methylation, this beneficial effect could not completely remove the effect of smoking on AHRR DNA demethylation.

Carotenoids are abundant pigments mainly contained in vegetables and fruits, and show antioxidant properties by quenching free radicals in human body. Few studies have investigated associations between serum carotenoid levels and premature mortality. The objective of this study was to investigate the association between serum carotenoid level and premature mortality in a Japanese population. This study included 446 Japanese adults (174 men, aged of 40-64) recruited as participants in the Japan Collaborative Cohort (JACC) Study. Serum carotenoid level was measured by high-performance liquid chromatography. Premature mortality was defined as death before 65 years old during the follow-up period. Premature mortality was ascertained in 60 men (34.5%) and 65 women (23.9%). In men, compared to the 1st tertile of serum β-cryptoxanthin and provitamin A, those who were in the 3rd tertile had lower risks of premature all-cause mortality (OR, 95% CI: 0.19, 0.07-0.47 for β-cryptoxanthin, and 0.24, 0.09-0.61 for provitamin A). In women, compared to the 1st tertile of serum β-cryptoxanthin, those who were in the 3rd tertile had higher risks of premature all-cause mortality (OR, 95% CI: 1.94, 1.00-4.03). These significant associations were observed in analyses for premature cancer mortality. We found significant associations between higher levels of serum β-cryptoxanthin and provitamin A and lower risks of premature mortality among Japanese men, while a different directional association was found in women. Although these findings suggest roles of serum carotenoids on premature mortality, further studies are needed to validate this association in other populations.

BACKGROUND: Although many polygenic risk scores (PRS) for cardiovascular traits have been developed in European populations, it is an urgent task to construct a PRS and to evaluate its ability in non-European populations. We developed a genome-wide PRS for blood pressure in a Japanese population and examined the associations between this PRS and hypertension prevalence. METHODS: We performed a cross-sectional study in 11 252 Japanese individuals who participated in the J-MICC (Japan Multi-Institutional Collaborative Cohort) study. Using publicly available GWAS summary statistics from Biobank Japan, we developed the PRS in the target data (n=7876). With >30 000 single nucleotide polymorphisms, we evaluated PRS performance in the test data (n=3376). Hypertension was defined as systolic blood pressure of 130 mm Hg or more, or diastolic blood pressure of 85 mm Hg or more, or taking an antihypertensive drug. RESULTS: Compared with the middle PRS quintile, the prevalence of hypertension at the top PRS quintile was higher independently from traditional risk factors (odds ratio, 1.73 [95% CI, 1.32-2.27]). The difference of mean systolic blood pressure and diastolic blood pressure between the middle and the top PRS quintile was 4.55 (95% CI, 2.26-6.85) and 2.32 (95% CI, 0.86-3.78) mm Hg, respectively. Subgroups reflecting combinations of Japanese PRS and modifiable lifestyles and factors (smoking, alcohol intake, sedentary time, and obesity) were associated with the prevalence of hypertension. A European-derived PRS was not associated with hypertension in our participants. CONCLUSIONS: A PRS for blood pressure was significantly associated with hypertension and BP traits in a general Japanese population. Our findings also highlighted the importance of a combination of PRS and risk factors for identifying high-risk subgroups.

BACKGROUND: Smoking is well known to be a major risk factor for cancer, and to decrease the levels of aryl hydrocarbon receptor repressor (AHRR) DNA methylation. AHRR is a key regulator for AHR signaling, which is involved in chemical metabolism and cancer development. Therefore, smoking-induced AHRR DNA hypomethylation may be associated with cancer development. However, it has not been reported that association between AHHR DNA methylation and cancer mortality in Asian population. Hence, we examined whether AHRR DNA methylation levels were associated with cancer mortality in a Japanese population. METHODS: This study was conducted with 812 participants (aged 38-80 years) who received a health check-up in 1990, and did not have a clinical histories. We followed up the participants until the end of 2019 (median: 27.8 years), and 100 participants died from cancer. The AHRR DNA methylation levels in peripheral blood mononuclear cells (PBMCs) were measured by the pyrosequencing method. We calculated the hazard ratio (HR) and 95% confidence interval (CI) for cancer mortality according to the baseline levels of AHRR DNA methylation. RESULTS: We found that AHRR DNA hypomethylation was associated with a higher risk of all cancer mortality, especially smoking related cancers and lung cancer. (all cancer: HR, 1.28, 95% CI, 1.09-1.51; smoking-related cancers: HR, 1.35, 95% CI, 1.12-1.62; lung cancer: HR, 1.68, 95% CI, 1.24-2.26). CONCLUSIONS: Smoking-induced AHRR DNA hypomethylation in PBMCs was associated with the risk of cancer mortality in Japanese population; therefore, hypomethylation of AHRR may be a useful biomarker of cancer mortality risk.

AIMS: This study aimed to analyze differences in sensitivity to hepatic lipid metabolism at different ages, through DNA methylation, using an experimental rat model of high-fructose corn syrup (HFCS) intake. MAIN METHODS: The experimental was divided into three periods: childhood and adolescence (postnatal day (PD) 21-60), young adulthood (PD61-100), and adulthood (PD101-140). Rats in the different age groups were assigned to receive either water (C: control group) or 20% HFCS solution (H: HFCS-fed group). We measured hepatic mRNA levels of peroxisome proliferator-activated receptor alpha (Ppara), carnitine palmitoyltransferase 1A (Cpt1a), fatty acid synthase (Fasn), and peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (Pgc1a) using real-time PCR. Additionally, we examined the DNA methylation levels of Ppara, Cpt1a, Fasn, and Pgc1a using pyrosequencing. KEY FINDINGS: Gene expressions of Cpt1a and Ppara in childhood and adolescence were significantly lower in the H group than in the C group. Conversely, Fasn and Pgc1a expressions were significantly higher in the H group than in the C group. Additionally, there was hypermethylation of Cpt1a and Ppara and hypomethylation of Fasn and Pgc1a in the H groups of childhood and adolescence. However, only one gene expression and methylation change was observed in young adulthood and adulthood groups. We found that HFCS intake in rats had stronger lipid metabolic effects in childhood and adolescence than in other generations, and that its mechanism involved epigenetic regulation. SIGNIFICANCE: We anticipate that these research findings will be a breakthrough for elucidating the varying effects of growth stage in the future.

BACKGROUND: Previous cross-sectional studies have shown that several circulating microRNA levels are associated with hypertension, but there are no prospective studies among general populations. OBJECTIVE: We evaluated the impact of circulating inflammatory- and oxidative stress-responsive microRNAs on changes in blood pressure and the development of hypertension in normotensive Japanese. METHOD: The study subjects were 84 normotensive participants (33 men and 51 women) who were given a health examination in both 2012 and 2017. In five years, 29 participants developed hypertension. Serum levels of miRNAs (miR-21, miR-27a, and miR-133a) were measured using qRT-PCR. Odds ratios (ORs) and 95% confidence intervals (CIs) for incident hypertension were estimated by logistic regression analysis. RESULTS: Serum miR-27a and -133a levels were lower in newly hypertensive subjects compared with normotensive subjects. With 1-unit lower serum miR-27a and -133a, the confounders adjusted ORs and 95% CI for incident hypertension were 0.84 (0.72-0.96) and 0.75 (0.58-0.91), respectively. The group with high levels of serum miR-27a and -133a had lower ORs than the group with low levels of these miRNAs (OR and 95% CI of miR-27a: 0.29, 0.08-0.91; miR-133a: 0.08, 0.01-0.37, respectively). CONCLUSIONS: Circulating miR-27a and -133a are potential biomarkers for the prediction and prevention of hypertension.

There are concerns about the negative effects of fructose intake during pregnancy on the next generation. We have previously reported that offspring from dams fed with fructose during gestation and lactation demonstrate abnormal lipid metabolism in the liver. In this study, we aimed to elucidate the molecular mechanism of the effects of maternal high-fructose corn syrup (HFCS) consumption on offspring. Pregnant Sprague-Dawley rats were fed with 20% HFCS water solution during gestation and lactation. Offspring were put on a normal diet after weaning, and the serum parameters and gene expression patterns were studied at predetermined intervals. Offsprings from pregnant rats fed with 20% HFCS (HFCS group) developed insulin resistance and hyperlipidemia at 60 d of age. RNA-seq analysis demonstrated that peroxisome proliferator-activated receptor α (PPARα) expression is downregulated by maternal HFCS intake. Hepatic Pparα expression in the HFCS group appeared to be suppressed by the enhanced DNA methylation of its promoter region. It is suggested that the development of insulin resistance and hyperlipidemia in the HFCS group may be attributable to aberrant Pparα methylation in the offspring liver. Pparα hypermethylation may be one of molecular mechanism underlying the toxicity of maternal fructose intake.

Background: Thioredoxin-interacting protein (TXNIP) controls the cellular redox balance by binding to and inhibiting the expression and function of thioredoxin. DNA methylation of the TXNIP gene is involved in the regulation of TXNIP mRNA expression. Changes in TXNIP DNA methylation levels are associated with the development of various diseases such as type 2 diabetes mellitus (T2DM). However, few studies have focused on the influence of lifestyle factors such as alcohol intake on TXNIP DNA methylation.Objectives: This research examines the association of drinking behaviors with TXNIP DNA methylation levels in the general Japanese population.Methods: We conducted a cross-sectional study of 404 subjects (176 males and 228 females) who were divided into non-, moderate and heavy drinkers based on self-reported drinking behaviors. TXNIP DNA methylation levels in leukocytes were determined using a pyrosequencing assay.Results: The mean TXNIP DNA methylation level in heavy drinkers (74.2%) was significantly lower than that in non- and moderate drinkers (non: 77.7%, p < .001; moderate: 76.6%, p = .011). Multivariable linear regression analysis showed that log-transformed values of daily (b = -1.34; p < .001) and cumulative (b = -1.06; p = .001) alcohol consumption were associated with decreased TXNIP DNA methylation levels.Conclusion: TXNIP DNA methylation levels in heavy drinkers was lower than in non- and- moderate drinkers. Decreased TXNIP DNA methylation level increases the expression of TXNIP and elevates the risk of developing of diseases such as T2DM. Therefore, decreasing alcohol use in heavy drinkers may lessen the likelihood of some alcohol-related illnesses moderated through TXNIP DNA methylation.

The increasing prevalence of nonalcoholic fatty liver disease (NAFLD) is a global health problem. In recent years, the inhibitory effect of brain-derived neurotrophic factor (BDNF) on diabetes mellitus and fatty liver has been clarified. The purpose of this study was to analyze the relationship between serum BDNF and NAFLD which caused by abnormal metabolism of glucose and lipids. This cross-sectional study involved 429 participants (mean age, 63.5 years: men, 38.5%) with low alcohol intake. Of the participants, those who had an increase in echogenicity of the liver parenchyma and hepato-renal contrast on ultrasonography were classified as the NAFLD group (n = 88), and the others were classified as the normal (n = 341) group. The NAFLD group was further classified into a mild group (n = 60) and a severe group (n = 28) based on the intensity of echogenicity and visualization of the hepatic vessels and diaphragm. Median BDNF levels were higher in the NAFLD group than the normal group (35.5 vs. 42.3 ng/mL, p < 0.01). Furthermore, BDNF levels tended to be associated with the severity of NAFLD (p < 0.01). In addition to the univariate analysis, in the sex- and age-adjusted model, there was a significant association between the BDNF levels and NAFLD severity (p < 0.01). The fully adjusted regression analysis also showed a positive association between the serum BDNF level and NAFLD (p < 0.01). These results suggest that NAFLD patients have a compensatory increase in circulating BDNF levels.

Metabolic syndrome (MetS) is cluster of metabolic diseases, including abdominal obesity, hyperglycemia, high blood pressure, and dyslipidemia, that directly escalate the risk of type 2 diabetes, heart disease, and stroke. Thioredoxin-interacting protein (TXNIP) is a binding protein for thioredoxin, a molecule that is a key inhibitor of cellular oxidation, and thus regulates the cellular redox state. Epigenetic alteration of the TXNIP-encoding locus has been associated with components of MetS. In the present study, we sought to determine whether the level of TXNIP methylation in blood is associated with MetS in the general Japanese population. DNA was extracted from the peripheral blood cells of 37 subjects with and 392 subjects without MetS. The level of TXNIP methylation at cg19693031 was assessed by the bisulfite-pyrosequencing method. We observed that TXNIP methylation levels were lower in MetS subjects (median 74.9%, range 71.7-78.4%) than in non-MetS subjects (median 77.7%, range 74.4-80.5%; p = 0.0024). Calculation of the confounding factor-adjusted odds ratios (ORs) and 95% confidence intervals (CIs) for hypomethylation revealed that subjects with MetS exhibited significantly higher ORs for hypomethylation than did those without MetS (OR, 2.92; 95% CI, 1.33-6.62; p = 0.009). Our findings indicated that lower levels of TXNIP methylation are associated with MetS in the general Japanese population. Altered levels of DNA methylation in TXNIP at cg19693031 might play an important role in the pathogenesis of MetS.

Consumption of fructose-containing beverages such as high-fructose corn syrup (HFCS) is increasing, raising concerns about the negative effects of excessive fructose intake. A recent report indicated that excess HFCS intake impairs hippocampal function. In this study, we focused on neurotrophic factors (NFs) in the hippocampus from the viewpoint of epigenetics to clarify the adverse effects of fructose. We analyzed the effects of HFCS intake on hippocampal function in three age categories: childhood and adolescence (postnatal day (PD) 21-60), young adulthood (PD60-100), and late adulthood (PD100-140). For the experiments, male Sprague-Dawley rats were divided into three age categories, the control group was received distilled water and the HFCS group was received 20% HFCS solution for 40 days in each period. We analyzed mRNA and protein levels for qPCR and western blotting, respectively, of a hippocampal NF, brain-derived neurotrophic factor (Bdnf). HFCS consumption reduced hippocampal Bdnf mRNA and protein expressions in childhood and adolescence. Moreover, pyrosequencing assays revealed increased DNA methylation at the Bdnf promoter in childhood and adolescence. This Bdnf levels reduction may be due to hypermethylation of the promoter regions. It should be noted that this phenomenon was observed only in childhood and adolescence fructose consumption. Our results indicate that the sensitivity of the hippocampus to fructose may vary with age. This study provides insight into the adverse effects of excessive HFCS consumption on the hippocampus in children.

Given that fructose consumption has increased by more than 10-fold in recent decades, it is possible that excess maternal fructose consumption causes harmful effects in the next generation. This study attempted to elucidate the mechanism of the harmful effects of excessive maternal fructose intake from the perspective of offspring liver function. Female rats during gestation and lactation were fed water containing fructose, and their offspring were fed normal water. We attempted to elucidate the mechanism of fructose-induced transgenerational toxicity by conducting a longitudinal study focusing on hepatic programming prior to disease onset. Impaired Insulin resistance and decreased high-density lipoprotein-cholesterol levels were observed at 160 days of age. However, metabolic disorders were not observed in 60-day-old offspring. Microarray analysis of 60-day-old offspring livers showed the reduction of hepatic insulin-like growth factor-1 (Igf1) mRNA expression. This reduction continued until the rats were aged 160 days and attenuated Igf1 signaling. Hepatic microRNA-29 (miR-29a) and miR-130a, which target Igf1 mRNA, were also found to be upregulated. Interestingly, these miRNAs were upregulated in the absence of metabolic disorder. In this study, we found that maternal fructose intake resulted in dysregulated expression of Igf1 and its target miRNAs in the offspring liver, and that these offspring were more likely to develop metabolic disorders. Abnormal hepatic programming induced by an imbalanced maternal nutritional environment is maintained throughout life, implying that it may contribute to metabolic disorders.

<title>Abstract</title><sec> <title>Background</title> Public perceptions and personal characteristics are heterogeneous between countries and subgroups, which may have different impacts on health-protective behaviors during the coronavirus disease 2019 (COVID-19) pandemic. To assess whether self-reported perceptions of COVID-19 and personal characteristics are associated with protective behaviors among general adults and to compare patterns in six different countries. </sec><sec> <title>Methods</title> This cross-sectional study uses the secondary data collected through an online survey between 15 and 23 April 2020 across six countries (China, Italy, Japan, Korea, the UK, and the USA). A total of 5945 adults aged 18 years or older were eligible for our analysis. A logistic regression model was used to estimate odds ratios (OR) and 95% confidence intervals (95%CI) of three recommended behaviors (wearing a mask, handwashing, and avoiding social gatherings). </sec><sec> <title>Results</title> In most countries except for China, the participants who perceived wearing a mask as being extremely effective to curtail the pandemic were more likely to wear a mask (OR, 95%CI: Italy: 4.14, 2.08–8.02; Japan: 3.59, 1.75–7.30; Korea: 7.89, 1.91–31.63: UK: 9.23, 5.14–17.31; USA: 4.81, 2.61–8.92). Those who perceived that handwashing was extremely effective had higher ORs of this preventive behavior (OR, 95%CI: Italy: 16.39, 3.56–70.18; Japan: 12.24, 4.03–37.35; Korea: 12.41, 2.02–76.39; UK: 18.04, 2.60–152.78; USA: 10.56, 2.21–44.32). The participants who perceived avoiding social gathering as being extremely effective to curtail the pandemic were more likely to take this type of preventive behavior (OR, 95%CI: China: 3.79, 1.28–10.23; Korea: 6.18, 1.77–20.60; UK: 4.45, 1.63–11.63; USA: 4.34, 1.84–9.95). The associations between personal characteristics, living environment, psychological status, and preventive behaviors varied across different countries. Individuals who changed their behavior because of recommendations from doctors/public health officials were more likely to take preventive behaviors in many countries. </sec><sec> <title>Conclusions</title> These findings suggest that higher perceived effectiveness may be a common factor to encourage preventive behaviors in response to the COVID-19 pandemic. These results may provide a better understanding of the homogeneity and heterogeneity of factors related to preventive behaviors and improve public health policies in various countries and groups. </sec>

DNA methylation (DNAm) is one of the most studied epigenetic modifications. DNAm has emerged as a key biological mechanism and biomarkers to test associations between environmental exposure and outcomes in epidemiological studies. Although previous studies have focused on associations between DNAm and either exposure/outcomes, it is useful to test for mediation of the association between exposure and outcome by DNAm. The purpose of this scoping review is to introduce the methodological essence of statistical mediation analysis and to examine emerging epidemiological research applying mediation analyses. We conducted this scoping review for published peer-reviewed journals on this topic using online databases (PubMed, Scopus, Cochrane, and CINAHL) ending in December 2020. We extracted a total of 219 articles by initial screening. After reviewing titles, abstracts, and full texts, a total of 69 articles were eligible for this review. The breakdown of studies assigned to each category was 13 for smoking (18.8%), 8 for dietary intake and famine (11.6%), 6 for other lifestyle factors (8.7%), 8 for clinical endpoints (11.6%), 22 for environmental chemical exposures (31.9%), 2 for socioeconomic status (SES) (2.9%), and 10 for genetic factors and race (14.5%). In this review, we provide an exposure-wide summary for the mediation analysis using DNAm levels. However, we found heterogenous methods and interpretations in mediation analysis with typical issues such as different cell compositions and tissue-specificity. Further accumulation of evidence with diverse exposures, populations and with rigorous methodology will be expected to provide further insight in the role of DNAm in disease susceptibility.

The ratio of human nonmercaptalbumin (HNA) and reduced albumin (HMA) may be a new marker for oxidative stress. Locomotive syndrome (LS) is reduced mobility due to impairment of locomotive organs. We investigated whether the HNA/HMA ratio could be a new biomarker of LS. This study included 306 subjects (mean age 64.24 ± 10.4 years) who underwent LS tests, grip strength, walking speed, and tests for HNA and HMA. Oxidative stress was measured by the ratio of HMA (f(HMA) = (HMA/(HMA + HNA) × 100)), and the subjects were divided into normal (N group; f[HMA] ≥ 70%) and low (L group; f[HMA] < 70%) groups. There were 124 non-elderly (<65 years) and 182 elderly subjects (≥65 years). There were no significant differences in LS, grip strength, and walking speed between the L and N groups in the non-elderly subjects. However, significant differences were found in the elderly subjects. In logistic regression analysis, there was an association between f(HMA) and the LS severity at older ages. LS in the elderly is associated with a decline in HMA and, thus, an increase in oxidative stress. Thus, f(HMA) is a new biomarker of LS.

OBJECTIVES: Although consumption of vegetable and 100% fruit juices are an acceptable alternative for vegetable and fruit intake, information about their actual effects on kidney function is sparse. The aim of this study was to determine the association between the consumption of vegetable and fruit juices and changes in kidney function in a Japanese population over a 5-y period. METHODS: In this prospective study, we analyzed 2755 Japanese (742 men and 2013 women) individuals who participated in both the baseline and follow-up surveys in the Daiko study (a study within the Japan Multi-Institutional Collaborative Cohort study). Estimated glomerular filtration rate (eGFR) was calculated by age, sex, and serum creatinine level. For each beverage, we categorized all participants into four groups-rare (rarely consumed), low (≤2 cups/wk), moderate (3-4 cups/wk), or frequent (≥5 cups/wk) consumers of the beverage-based on a food frequency questionnaire. RESULTS: The mean baseline and follow-up eGFR (SD) were 82.4 (14.6) and 72.2 (12.6), respectively. In fully adjusted regression analyses, moderate consumption of vegetable juice was associated a lower decline in eGFR compared with the rare consumption group (β = -1.30; P = 0.01). Moreover, stratified analyses revealed that this significant association remained in those who were young, female, non-obese, normotensive, smoked cigarettes, consumed alcohol, or exercised. However, no significant association was found in analyses for fruit juices. CONCLUSIONS: This 5-y prospective study suggested an association between self-reported moderate consumption of vegetable juice and changes (possibly smaller decline) in kidney function in a relatively healthy Japanese population.

Primary prevention of premature death is a public health concern worldwide. Circulating microRNAs (miRNAs) have been described as potential diagnostic biomarkers for diseases as cancer and cardiovascular disease (CVD). This case-cohort study aimed to investigate the potential relationship between circulating miRNAs and the risk of premature death. A total of 39,242 subjects provided baseline serum samples in 1988-1990. Of these, 345 subjects who died of intrinsic disease (< 65 years old) and for which measurable samples were available were included in this study. We randomly selected a sub-cohort of 879 subjects. Circulatring miR-21, miR-29a, and miR-126 were determined using qRT-PCR. Conditional logistic regression models were used to analyse the data with respect to stratified miRNA levels. Multivariable logistic regression revealed that subjects with high circulating miR-21 and miR-29a individual levels had a significantly higher risk of total death, cancer death, and CVD death than those with medium miR-21 and miR-29a individual levels. Conversely, subjects with low circulating miR-126 levels had a significantly higher risk of total death than those with medium levels. This suggests that circulating miRNAs are associated with the risk of premature death from cancer and CVD, identifying them as potential biomarkers for early detection of high-risk individuals.

BACKGROUND: Inflammation is thought to be a risk factor for kidney disease. However, discussion is controversial whether inflammatory status is either a cause or an outcome of chronic kidney disease. We aimed to investigate the causal relationship between high-sensitivity C-reactive protein (hs-CRP) and estimated glomerular filtration rate (eGFR) using mendelian randomization (MR) approaches. METHODS: A total of 10,521 participants of the Japan Multi-institutional Collaborative Cohort Study was analyzed in this study. We used two-sample MR approaches (the inverse-variance weighted (IVW), the weighted median (WM), and the MR-Egger method) to estimate the effect of genetically determined hs-CRP on kidney function. We selected four and three hs-CRP associated single nucleotide polymorphisms (SNPs) as two instrumental variables (IV): IVCRP and IVAsian, based on SNPs previously identified in European and Asian populations. IVCRP and IVAsian explained 3.4% and 3.9% of the variation in hs-CRP, respectively. RESULTS: Using the IVCRP, genetically determined hs-CRP was not significantly associated with eGFR in the IVW and the WM methods (estimate per 1 unit increase in ln(hs-CRP), 95%CI: 0.000, -0.019 to 0.020 and -0.003, -0.019 to 0.014). For IVAsian, we found similar results using the IVW and the WM methods (estimate, 95% CI: -0.005, -0.020 to 0.010 and -0.004, -0.020 to 0.012). The MR-Egger method also showed no causal relationships between hs-CRP and eGFR (IVCRP: -0.008, -0.058 to 0.042; IVAsian: 0.001, -0.036 to 0.036). CONCLUSIONS: Our two-sample MR analyses with different IVs did not support a causal effect of hs-CRP on eGFR.

OBJECTIVE: MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression and play essential roles in the pathogenesis of cardiovascular disease. Previous cross-sectional studies showed that the levels of several circulating miRNA are associated with hypertension, but there are no prospective longitudinal studies using a general population. The aim of this study is to evaluate the impact of circulating vascular-related miRNA (miR-126, miR-221, and miR-222) on changes in blood pressure and new-onset hypertension in a Japanese population. METHODS: We conducted a 5-year longitudinal study using 192 health examination participants (87 men and 105 women). Serum miRNAs were measured using quantitative reverse transcription-PCR. Information regarding lifestyle and health condition was obtained using a self-administered questionnaire. Logistic regression analyses were performed to calculate odds ratios and 95% confidence intervals for new-onset hypertension in the 5-year period between the low and high group of serum miRNAs. RESULTS: Serum levels of miR-126, miR-221, and miR-222 were significantly and negatively associated with changes in SBP and the rate of change of SBP. Serum miR-126, miR-221, and miR-222 levels were significantly lower in new-onset hypertensive patients compared with normotensive individuals. The confounding factors adjusted odds ratios of each 1 increment in serum miR-126, miR-221, and miR-222 levels were 0.82 (95% confidence interval: 0.69-0.98), 0.79 (0.68-0.91), and 0.61 (0.46-0.81) for new-onset hypertension, respectively. CONCLUSION: Low serum levels of miR-126, miR-221, and miR-222 were associated with increased blood pressure and new-onset of hypertension. These circulating miRNAs are potential candidate biomarkers for the prediction of hypertension.

Maternal fructose consumption affects the metabolic functions of offspring later in life. However, the molecular mechanism remains poorly understood. Differences of microRNA expression profile and DNA methylation status are a candidate mechanism to explain the developmental programming that contributes to the development of a metabolic disorder. This study examined the transgenerational effect of maternal fructose consumption from the perspective of epigenetic modification. To do this, we collected serum and liver tissues from male offspring rats that were exposed to maternal distilled water or 20% fructose water during gestation and lactation. A decreased serum high-density lipoprotein cholesterol (HDL-C) level was observed in the offspring of fructose-fed dams at postnatal day (PD) 160. Given research indicating a role of liver X receptor alpha (LXRA) in cholesterol metabolism, we analyzed Lxra expression. Real-time polymerase chain reaction analysis demonstrated that offspring that were delivered from fructose-fed dams exhibited decreased Lxra gene expression in their liver tissue. There is a well-established association between Lxra expression and the level of DNA methylation and miR-206 expression. Pyrosequencing assays revealed no differences in the level of DNA methylation in the Lxra promoter region, whereas miR-206 expression was increased in the liver at PD 60 and 160. Our data indicate that early-life exposure to maternal fructose results in changing of miR-206 expression level in the liver that suppresses the expression of Lxra. This phenomenon may be associated with the decreased serum HDL-C level in offspring.

Osteoporosis is a disease characterized by deterioration of bone tissue and mass, with an increasing global prevalence. Therefore, the discovery of biomarkers for osteoporosis would help to guide appropriate treatment. Circulating microRNAs (miRNAs) have become increasingly recognized as biomarkers for detecting diseases. However, few studies have investigated the association of circulating miRNA with osteoporosis in the general population. The aim of this study was to identify miRNA associated with osteoporosis in a general resident health check-up for potential use as an osteoporosis biomarker. We conducted a cross-sectional study as part of a health check-up program and recruited 352 volunteers (139 men, 213 women, mean age 64.1 ± 9.6 years). Osteoporosis was diagnosed according to the WHO classification. Twenty-two candidate microRNAs were screened through real-time quantitative PCR, and miRNAs associated with osteoporosis were analyzed using logistic regression analysis including other risk factors. In total, 95 females and 30 males were diagnosed with osteoporosis with bone mineral density tests (BMD: T-score < -2.5). We found that miR195 was significantly lower in females, while miR150 and miR222 were significantly higher in males. The results of the logistic regression analysis indicated that in females, higher age and lower miR195 (odds ratio: 0.45, 95% confidential interval: 0.03-0.98) were significant risk factors for lower BMD, while the presence of a smoking habit and lower miR150 (odds ratio: 1.35, 95% confidential interval: 1.02-1.79) were significant risk factors for osteoporosis. Serum levels of miR195 and miR150 are independently associated with low bone mineral density in females and males, respectively.