Metabolomics has developed as a powerful tool for investigating the complex pathophysiology underlying atherosclerosis and cardiovascular disease. Many epidemiological studies have applied this ...technique to accurately and comprehensively assess the effects of environmental factors on health outcomes, which used to be a perpetual challenge. Metabolites are defined as small molecules which are intermediate products of metabolic reactions catalyzed by numerous enzymes occurring within cells. Consequent to both genetic variation and environment, they allow us to explore the gene–environment interactions and to gain a better understanding of multifactorial diseases like cardiovascular disease. This review article highlights the findings of well-known prospective cohort studies around the world that have utilized metabolomics for a wide range of purposes, including biomarker discovery, improving cardiovascular risk prediction and early disease diagnosis, and exploring detailed mechanisms of disease onset and progression. However, technical challenges still exist in applying them clinically. One limitation is due to various analytical platforms that are used based on the judgment of each study; comparative assessments among different platforms need to be conducted in order to correctly interpret and validate each data externally. Secondly, metabolite levels obtained in most high-throughput metabolomics profiling studies are often semiquantitative rather than fully quantitative concentrations, which makes it difficult to compare and combine results among different studies and to determine the levels for practical use. In 2014, the Consortium of Metabolomics Studies was developed, which is expected to take the lead in overcoming these issues.
Background: Although self-reported questionnaires are widely employed in epidemiologic studies, their validity has not been sufficiently assessed. The aim of this study was to evaluate the validity ...of a self-reported questionnaire on medication use by comparison with health insurance claims and to identify individual determinants of discordance in the Tsuruoka Metabolomics Cohort Study.Methods: Participants were 2,472 community-dwellers aged 37 to 78 years from the Tsuruoka Metabolomics Cohort Study. Information on lifestyle and medications was collected through a questionnaire. Sensitivity and specificity were determined using health insurance claims from November 2014 to March 2016, which were used as a standard. Potential determinants of discordance were assessed using multivariable logistic regression.Results: The self-reported questionnaire on medication use showed high validity. Sensitivity and specificity were 0.95 (95% CI, 0.93–0.96) and 0.97 (95% CI, 0.96–0.98) for antihypertensive medications, 0.94 (95% CI, 0.91–0.97) and 0.98 (95% CI, 0.98–0.99) for diabetes medications, and 0.84 (95% CI, 0.82–0.87) and 0.98 (95% CI, 0.97–0.99) for dyslipidemia medications, respectively. Males without high education and those who currently smoke cigarettes were found to be associated with discordant reporting which affected sensitivity, especially those with medication use for dyslipidemia.Conclusions: In this population-based cohort study, we found that the self-reported questionnaire on medication use was a valid measure to capture regular medication users. Sensitivity for dyslipidemia medications was lower than those for the other medications. Type of medication, sex, education years, and smoking status influenced discordance, which affected sensitivity in self-reporting.
To assess the association of dietary saturated fatty acid (SFA) intake with the presence of early AMD in a Japanese population.
The population-based Tsuruoka Metabolomics Cohort Study enrolled ...general population individuals aged 35 to 74 years from among participants in annual health check-up programs that included fundus photographs in Tsuruoka, Japan. A total of 4010 individuals participated in the baseline survey. After excluding nonresponders to a dietary survey and participants with suboptimal fundus image quality, 3988 participants (median age, 62.4 years) were included in this cross-sectional analysis. Dietary intake was assessed by a validated food frequency questionnaire. Fatty acids intake was adjusted for total energy intake by the residuals method. The association between fatty acid intake and presence of early AMD was assessed by logistic regression models.
Median daily SFA intake was 11.3 g (interquartile range, 9.6, 13.0 g). After adjustments for potential confounding factors, participants in the highest quartile of SFA intake were less likely to have early AMD, compared with the lowest quartile (odds ratio, 0.71; 95% confidence interval: 0.52-0.96). A significant trend for decreased risk of early AMD with increasing SFA intake was noted (P = 0.011). There was no significant association between poly-unsaturated fatty acid (PUFA) including n3-PUFA intake and early AMD.
We found that increased SFA intake was associated with reduced risk of early AMD in a Japanese population with low SFA intake. Adequate fatty acid intake may be required to maintain retinal homeostasis and prevent AMD.
Currently, large-scale cohort studies for metabolome analysis have been launched globally. However, only a few studies have evaluated the reliability of urinary metabolome analysis. This study aimed ...to establish the reliability of urinary metabolomic profiling in cohort studies. In the Tsuruoka Metabolomics Cohort Study, 123 charged metabolites were identified and routinely quantified using capillary electrophoresis-mass spectrometry (CE-MS). We evaluated approximately 750 quality control (QC) samples and 6,720 participants' spot urine samples. We calculated inter- and intra-batch coefficients of variation in the QC and participant samples and technical intraclass correlation coefficients (ICC). A correlation of metabolite concentrations between spot and 24-h urine samples obtained from 32 sub-cohort participants was also evaluated. The coefficient of variation (CV) was less than 20% for 87 metabolites (70.7%) and 20-30% for 19 metabolites (15.4%) in the QC samples. There was less than 20% inter-batch CV for 106 metabolites (86.2%). Most urinary metabolites would have reliability for measurement. The 96 metabolites (78.0%) was above 0.75 for the estimated ICC, and those might be useful for epidemiological analysis. Among individuals, the Pearson correlation coefficient of 24-h and spot urine was more than 70% for 59 of the 99 metabolites. These results show that the profiling of charged metabolites using CE-MS in morning spot human urine is suitable for epidemiological metabolomics studies.
Food intake biomarkers can be critical tools that can be used to objectively assess dietary exposure for both epidemiological and clinical nutrition studies. While an accurate estimation of food ...intake is essential to unravel associations between the intake and specific health conditions, random and systematic errors affect self-reported assessments. This study aimed to clarify how habitual food intake influences the circulating plasma metabolome in a free-living Japanese regional population and to identify potential food intake biomarkers. To achieve this aim, we conducted a cross-sectional analysis as part of a large cohort study. From a baseline survey of the Tsuruoka Metabolome Cohort Study, 7,012 eligible male and female participants aged 40-69 years were chosen for this study. All data on patients' health status and dietary intake were assessed via a food frequency questionnaire, and plasma samples were obtained during an annual physical examination. Ninety-four charged plasma metabolites were measured using capillary electrophoresis mass spectrometry, by a non-targeted approach. Statistical analysis was performed using partial-least-square regression. A total of 21 plasma metabolites were likely to be associated with long-term food intake of nine food groups. In particular, the influential compounds in each food group were hydroxyproline for meat, trimethylamine-N-oxide for fish, choline for eggs, galactarate for dairy, cystine and betaine for soy products, threonate and galactarate for carotenoid-rich vegetables, proline betaine for fruits, quinate and trigonelline for coffee, and pipecolate for alcohol, and these were considered as prominent food intake markers in Japanese eating habits. A set of circulating plasma metabolites was identified as potential food intake biomarkers in the Japanese community-dwelling population. These results will open the way for the application of new reliable dietary assessment tools not by self-reported measurements but through objective quantification of biofluids.
The use of heated tobacco products (HTP) has increased exponentially in Japan since 2016; however, their effects on health remain a major concern.
Tsuruoka Metabolome Cohort Study participants (
= ...11,002) were grouped on the basis of their smoking habits as never smokers (NS), past smokers (PS), combustible tobacco smokers (CS), and HTP users for <2 years. Peripheral blood mononuclear cells were collected from 52 participants per group matched to HTP users using propensity scores, and DNA and RNA were purified from the samples. DNA methylation (DNAm) analysis of the 17 smoking-associated DNAm biomarker genes (such as
,
,
, and
), as well as whole transcriptome analysis, was performed.
Ten of the 17 genes were significantly hypomethylated in CS and HTP users compared with NS, among which
,
, and
showed intermediate characteristics between CS and NS; nonetheless,
expression was significantly higher in CS than in the other three groups. Conversely,
and
were more hypomethylated in HTP users than in NS, and
expression was markedly upregulated in all the groups when compared with that in NS.
HTP users (switched from CS <2 years) display abnormal DNAm and transcriptome profiles, albeit to a lesser extent than the CS. However, because the molecular genetic effects of long-term HTP use are still unknown, long-term molecular epidemiologic studies are needed.
This study provides new insights into the molecular genetic effects on DNAm and transcriptome profiles in HTP users who switched from CS.
Aims: The categories in the comprehensive lipid and risk management guidelines were proposed by the Japan Atherosclerosis Society (JAS Guidelines 2017), which adopted the estimated 10 year absolute ...risk of coronary artery disease (CAD) incidence in the Suita score. We examined whether those categories were concordant with the degree of arterial stiffness. Methods: In 2014, the cardio-ankle vascular index (CAVI), an arterial stiffness parameter, was measured in 1,972 Japanese participants aged 35–74 years in Tsuruoka City, Yamagata Prefecture, Japan. We examined the mean CAVI and the proportion and odds ratios (ORs) of CAVI ≥ 9.0 on the basis of the following three management classifications using the analysis of variance and logistic regression: “Category I (Low risk),” “Category II (Middle risk),” and “Category III (High risk).” Results: The mean CAVI and proportion of CAVI ≥ 9.0 were 8.6 and 34.8% among males and 8.1 and 18.3% among females, respectively. The mean CAVI and proportion of CAVI ≥ 9.0 were associated with an estimated 10 year absolute risk for CAD among males and females, excluding High risk for females. These results were similar to the management classification by the guideline: the multivariable-adjusted ORs (95% confidence intervals) of CAVI ≥ 9.0 among Category II and Category III compared with those among Category I were 2.96 (1.61–5.43) and 7.33 (4.03–13.3) for males and 3.99 (2.55–6.24) and 3.34 (2.16–5.16) for females, respectively. Conclusions: The risk stratification, which was proposed in the JAS Guidelines 2017, is concordant with the arterial stiffness parameter.
The purpose of this study was to evaluate the reproducibility and validity of a short food frequency questionnaire (FFQ) for food group intake in Japan, the reproducibility and partial validity of ...which were previously confirmed for nutrients.
A total of 288 middle-aged healthy volunteers from 11 different areas of Japan provided nonconsecutive 3-day weighed dietary records (DRs) at 3-month intervals over four seasons. We evaluated reproducibility based on the first (FFQ1) and second (FFQ2) questionnaires and their validity against the DRs by comparing the intake of 20 food groups. Spearman's rank correlation coefficients (SRs) were calculated between energy-adjusted intake from the FFQs and that from the DRs.
The intake of 20 food groups estimated from the two FFQs was mostly equivalent. The median energy-adjusted SRs between the FFQ1 and FFQ2 were 0.61 (range 0.38-0.86) for men and 0.66 (0.45-0.84) for women. For validity, the median de-attenuated SRs between DRs and the FFQ1 were 0.51 (0.17-0.76) for men and 0.47 (0.23-0.77) for women. Compared with the DRs, the proportion of cross-classification into exact plus adjacent quintiles with the FFQ1 ranged from 58 to 86% in men and from 57 to 86% in women. According to the robust Z scores and the Bland-Altman plot graphs, the underestimation errors in the FFQ1 tended to be greater in individuals with high mean levels of consumption for meat for men and for other vegetables for both men and women.
The FFQ demonstrated high reproducibility and reasonable validity for food group intake. This questionnaire is short and remains appropriate for identifying associations between diet and health/disease among adults in Japan.
Cohort studies with metabolomics data are becoming more widespread, however, large-scale studies involving 10,000s of participants are still limited, especially in Asian populations. Therefore, we ...started the Tsuruoka Metabolomics Cohort Study enrolling 11,002 community-dwelling adults in Japan, and using capillary electrophoresis-mass spectrometry (CE-MS) and liquid chromatography-mass spectrometry. The CE-MS method is highly amenable to absolute quantification of polar metabolites, however, its reliability for large-scale measurement is unclear. The aim of this study is to examine reproducibility and validity of large-scale CE-MS measurements. In addition, the study presents absolute concentrations of polar metabolites in human plasma, which can be used in future as reference ranges in a Japanese population.
Metabolomic profiling of 8,413 fasting plasma samples were completed using CE-MS, and 94 polar metabolites were structurally identified and quantified. Quality control (QC) samples were injected every ten samples and assessed throughout the analysis. Inter- and intra-batch coefficients of variation of QC and participant samples, and technical intraclass correlation coefficients were estimated. Passing-Bablok regression of plasma concentrations by CE-MS on serum concentrations by standard clinical chemistry assays was conducted for creatinine and uric acid.
In QC samples, coefficient of variation was less than 20% for 64 metabolites, and less than 30% for 80 metabolites out of the 94 metabolites. Inter-batch coefficient of variation was less than 20% for 81 metabolites. Estimated technical intraclass correlation coefficient was above 0.75 for 67 metabolites. The slope of Passing-Bablok regression was estimated as 0.97 (95% confidence interval: 0.95, 0.98) for creatinine and 0.95 (0.92, 0.96) for uric acid. Compared to published data from other large cohort measurement platforms, reproducibility of metabolites common to the platforms was similar to or better than in the other studies. These results show that our CE-MS platform is suitable for conducting large-scale epidemiological studies.
Metabolomics is a promising approach to the identification of biomarkers in plasma. Here, we performed a population-based, cross-sectional study to identify potential biomarkers of alcohol intake and ...alcohol-induced liver injury by metabolomic profiling using capillary electrophoresis-mass spectrometry (CE-MS).
Fasting plasma samples were collected from 896 Japanese men who participated in the baseline survey of the Tsuruoka Metabolomics Cohort Study, and 115 polar metabolites were identified and absolutely quantified by CE-MS. Information on daily ethanol intake was collected through a standardized, self-administered questionnaire. The associations between ethanol intake and plasma concentration of metabolites were examined. Relationships between metabolite concentrations or their ratios and serum liver enzyme levels in the highest ethanol intake group (>46.0 g/day) were then examined by linear regression analysis. Replication analysis was conducted in 193 samples collected from independent population of this cohort.
Nineteen metabolites were identified to have an association with daily alcohol consumption both in the original and replication population. Three of these metabolites (threonine, glutamine, and guanidinosuccinate) were found to associate well with elevated levels of serum liver enzymes in the highest ethanol intake group, but not in the non-drinker group. We also found that the glutamate/glutamine ratio had a much stronger relation to serum γ-glutamyltransferase, aspartate transaminase, and alanine transaminase than glutamate or glutamine alone (standardized beta = 0.678, 0.558, 0.498, respectively).
We found 19 metabolites associated with alcohol intake, and three biomarker candidates (threonine, guanidinosuccinate and glutamine) of alcohol-induced liver injury. Glutamate/glutamine ratio might also be good biomarker.
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