Chemical Exposomics in Human Plasma by Lipid Removal and Large-Volume Injection Gas Chromatography-High-Resolution Mass Spectrometry.

For comprehensive chemical exposomics in blood, analytical workflows are evolving through advances in sample preparation and instrumental methods. We hypothesized that gas chromatography-high-resolution mass spectrometry (GC-HRMS) workflows could be enhanced by minimizing lipid coextractives, thereby enabling larger injection volumes and lower matrix interference for improved target sensitivity and nontarget molecular discovery. A simple protocol was developed for small plasma volumes (100-200 µL) by using isohexane (H) to extract supernatants of acetonitrile-plasma (A-P). The HA-P method was quantitative for a wide range of hydrophobic multiclass target analytes (i.e., log Kow > 3.0), and the extracts were free of major lipids, thereby enabling robust large-volume injections (LVIs; 25 µL) in long sequences (60-70 h, 70-80 injections) to a GC-Orbitrap HRMS. Without lipid removal, LVI was counterproductive because method sensitivity suffered from the abundant matrix signal, resulting in low ion injection times to the Orbitrap. The median method quantification limit was 0.09 ng/mL (range 0.005-4.83 ng/mL), and good accuracy was shown for a certified reference serum. Applying the method to plasma from a Swedish cohort (n = 32; 100 µL), 51 of 103 target analytes were detected. Simultaneous nontarget analysis resulted in 112 structural annotations (12.8% annotation rate), and Level 1 identification was achieved for 7 of 8 substances in follow-up confirmations. The HA-P method is potentially scalable for application in cohort studies and is also compatible with many liquid-chromatography-based exposomics workflows.

Personalized mixture toxicity testing: A proof-of-principle in vitro study evaluating the steroidogenic effects of reconstructed contaminant mixtures measured in blood of individual adults.

Chemical risk assessments typically focus on single substances, often overlooking real-world co-exposures to chemical mixtures. Mixture toxicology studies using representative mixtures can reveal potential chemical interactions, but these do not account for the unique chemical profiles that occur in the blood of diverse individuals. Here we used the H295R steroidogenesis assay to screen personalized mixtures of 24 persistent organic pollutants (POPs) for cytotoxicity and endocrine disruption. Each mixture was reconstructed at a human exposure relevant concentration (1×), as well as at 10- and 100-fold higher concentration (10×, 100×) by acoustic liquid handling based on measured blood concentrations in a Swedish cohort. Among the twelve mixtures tested, nine mixtures decreased the cell viability by 4-18%, primarily at the highest concentration. While the median and maximum mixtures based on the whole study population induced no measurable effects on steroidogenesis at any concentration, the personalized mixture from an individual with the lowest total POPs concentration was the only mixture that affected estradiol synthesis (35% increase at the 100× concentration). Mixtures reconstructed from blood levels of three different individuals stimulated testosterone synthesis at the 1× (11-15%) and 10× concentrations (12-16%), but not at the 100× concentration. This proof-of-principle personalized toxicity study illustrates that population-based representative chemical mixtures may not adequately account for the toxicological risks posed to individuals. It highlights the importance of testing a range of real-world mixtures at relevant concentrations to explore potential interactions and non-monotonic effects. Further toxicological studies of personalized contaminant mixtures could improve chemical risk assessment and advance the understanding of human health, as chemical exposome data become increasingly available.

A regional comparison of children's blood cadmium, lead, and mercury in rural, urban and industrial areas of six European countries, and China, Ecuador, and Morocco.

OBJECTIVES: The authors aimed to evaluate whether blood cadmium (B-Cd), lead (B-Pb) and mercury (B-Hg) in children differ regionally in 9 countries, and to identify factors correlating with exposure. MATERIAL AND METHODS: The authors performed a cross-sectional study of children aged 7-14 years, living in 2007-2008 in urban, rural, or potentially polluted ("hot spot") areas (ca. 50 children from each area, in total 1363 children) in 6 European and 3 non-European countries. The authors analyzed Cd, Pb, and total Hg in blood and collected information on potential determinants of exposure through questionnaires. Regional differences in exposure levels were assessed within each country. RESULTS: Children living near industrial "hot-spots" had B-Cd 1.6 (95% CI: 1.4-1.9) times higher in the Czech Republic and 2.1 (95% CI:1.6-2.8) times higher in Poland, as compared to urban children in the same countries (geometric means [GM]: 0.13 µg/l and 0.15 µg/l, respectively). Correspondingly, B-Pb in the "hot spot" areas was 1.8 (95% CI: 1.6-2.1) times higher than in urban areas in Slovakia and 2.3 (95% CI: 1.9-2.7) times higher in Poland (urban GM: 19.4 µg/l and 16.3 µg/l, respectively). In China and Morocco, rural children had significantly lower B-Pb than urban ones (urban GM: 64 µg/l and 71 µg/l, respectively), suggesting urban exposure from leaded petrol, water pipes and/or coal-burning. Hg "hot spot" areas in China had B-Hg 3.1 (95% CI: 2.7-3.5) times higher, and Ecuador 1.5 (95% CI: 1.2-1.9) times higher, as compared to urban areas (urban GM: 2.45 µg/l and 3.23 µg/l, respectively). Besides industrial exposure, traffic correlated with B-Cd; male sex, environmental tobacco smoke, and offal consumption with B-Pb; and fish consumption and amalgam fillings with B-Hg. However, these correlations could only marginally explain regional differences. CONCLUSIONS: These mainly European results indicate that some children experience about doubled exposures to toxic elements just because of where they live. These exposures are unsafe, identifiable, and preventable and therefore call for preventive actions. Int J Occup Med Environ Health. 2023;36(3):349-64.

Adipokines are possible risk markers for aortic stenosis requiring surgery.

OBJECTIVES: Aortic stenosis (AS) is the most prevalent valvular heart disease among adults. The adipocyte-derived hormones, leptin and adiponectin, have profound metabolic actions. We examined whether these adipokines are independently associated with future aortic valve replacement (AVR). DESIGN: In this longitudinal case-control study, we identified 336 cases who had undergone AVR due to AS, and who had previously participated in population-based health surveys. Two referents were matched to each case and leptin and adiponectin concentrations were analysed from stored baseline survey samples. Uni- and multivariable logistic regression analyses were used to estimate the risk of future AVR. An additional cohort was identified for validation including 106 cases with AVR and 212 matched referents. RESULTS: Median age (interquartile range (IQR)) in years at survey was 59.9 (10.4) and at surgery 68.3 (12.7), and 48% were women. An elevated concentration of leptin was not associated with future AVR (odds ratio [95% confidence interval]) (1.10 [0.92-1.32]), although leptin was associated with a higher risk in patients with coronary artery disease (CAD) having more than 5 years between survey and AVR (1.41 [1.08-1.84]). Adiponectin was not associated with higher risk for future AVR (0.95 [0.82-1.11]), although after stratification for age, higher levels were associated with reduced risk for AVR in persons aged ≥60 years at surgery (0.79 [0.64-0.98]). In the validation study, leptin was associated with future AVR whereas adiponectin was not. None of the associations remained significant after adjustment for body mass index (BMI). CONCLUSIONS: The adipokine leptin may promote the development of AS.

Phospholipid Removal for Enhanced Chemical Exposomics in Human Plasma.

The challenge of chemical exposomics in human plasma is the 1000-fold concentration gap between endogenous substances and environmental pollutants. Phospholipids are the major endogenous small molecules in plasma, thus we validated a chemical exposomics protocol with an optimized phospholipid-removal step prior to targeted and non-targeted liquid chromatography high-resolution mass spectrometry. Increased injection volume with negligible matrix effect permitted sensitive multiclass targeted analysis of 77 priority analytes; median MLOQ = 0.05 ng/mL for 200 µL plasma. In non-targeted acquisition, mean total signal intensities of non-phospholipids were enhanced 6-fold in positive (max 28-fold) and 4-fold in negative mode (max 58-fold) compared to a control method without phospholipid removal. Moreover, 109 and 28% more non-phospholipid molecular features were detected by exposomics in positive and negative mode, respectively, allowing new substances to be annotated that were non-detectable without phospholipid removal. In individual adult plasma (100 µL, n = 34), 28 analytes were detected and quantified among 10 chemical classes, and quantitation of per- and polyfluoroalkyl substances (PFAS) was externally validated by independent targeted analysis. Retrospective discovery and semi-quantification of PFAS-precursors was demonstrated, and widespread fenuron exposure is reported in plasma for the first time. The new exposomics method is complementary to metabolomics protocols, relies on open science resources, and can be scaled to support large studies of the exposome.

Polybrominated diphenyl ethers in type 2 diabetes mellitus cases and controls: Repeated measurements prior to and after diagnosis.

BACKGROUND: Previous studies have reported associations between certain persistent organic pollutants (POPs) and type 2 diabetes mellitus (T2DM). Polybrominated diphenyl ethers (PBDEs) are a class of POPs that are found in increasing concentrations in humans. Although obesity is a known risk factor for T2DM and PBDEs are fat-soluble, very few studies have investigated associations between PBDEs and T2DM. No longitudinal studies have assessed associations between repeated measurements of PBDE and T2DM in the same individuals and compared time trends of PBDEs in T2DM cases and controls. OBJECTIVES: To investigate associations between pre- and post-diagnostic measurements of PBDEs and T2DM and to compare time trends of PBDEs in T2DM cases and controls. METHODS: Questionnaire data and serum samples from participants in the Tromsø Study were used to conduct a longitudinal nested case-control study among 116 T2DM cases and 139 controls. All included study participants had three pre-diagnostic blood samples (collected before T2DM diagnosis in cases), and up to two post-diagnostic samples after T2DM diagnosis. We used logistic regression models to investigate pre- and post-diagnostic associations between PBDEs and T2DM, and linear mixed-effect models to assess time trends of PBDEs in T2DM cases and controls. RESULTS: We observed no substantial pre- or post-diagnostic associations between any of the PBDEs and T2DM, except for BDE-154 at one of the post-diagnostic time-points (OR = 1.65, 95% CI: 1.00, 2.71). The overall time trends of PBDE concentrations were similar for cases and controls. DISCUSSION: The study did not support PBDEs increasing the odds of T2DM, prior to or after T2DM diagnosis. T2DM status did not influence the time trends of PBDE concentrations.

Lead-exposure associated miRNAs in humans and Alzheimer's disease as potential biomarkers of the disease and disease processes.

Alzheimer's disease (AD) is a neurodegenerative disease that eventually affects memory and behavior. The identification of biomarkers based on risk factors for AD provides insight into the disease since the exact cause of AD remains unknown. Several studies have proposed microRNAs (miRNAs) in blood as potential biomarkers for AD. Exposure to heavy metals is a potential risk factor for onset and development of AD. Blood cells of subjects that are exposed to lead detected in the circulatory system, potentially reflect molecular responses to this exposure that are similar to the response of neurons. In this study we analyzed blood cell-derived miRNAs derived from a general population as proxies of potentially AD-related mechanisms triggered by lead exposure. Subsequently, we analyzed these mechanisms in the brain tissue of AD subjects and controls. A total of four miRNAs were identified as lead exposure-associated with hsa-miR-3651, hsa-miR-150-5p and hsa-miR-664b-3p being negatively and hsa-miR-627 positively associated. In human brain derived from AD and AD control subjects all four miRNAs were detected. Moreover, two miRNAs (miR-3651, miR-664b-3p) showed significant differential expression in AD brains versus controls, in accordance with the change direction of lead exposure. The miRNAs' gene targets were validated for expression in the human brain and were found enriched in AD-relevant pathways such as axon guidance. Moreover, we identified several AD relevant transcription factors such as CREB1 associated with the identified miRNAs. These findings suggest that the identified miRNAs are involved in the development of AD and might be useful in the development of new, less invasive biomarkers for monitoring of novel therapies or of processes involved in AD development.

Longitudinal changes in concentrations of persistent organic pollutants (1986-2016) and their associations with type 2 diabetes mellitus.

BACKGROUND: Positive associations have been reported between persistent organic pollutants (POPs) and type 2 diabetes mellitus (T2DM); however, causality has not been established. Over the last decades, environmental exposure to legacy POPs has decreased, complicating epidemiological studies. In addition, physiological risk factors for T2DM may also influence POP concentrations, contributing to a complex network of factors that could impact associations with T2DM. Longitudinal studies on this topic are lacking, and few have assessed prospective and cross-sectional associations between repeated POP measurements and T2DM in the same individuals, which may shed light on causality. OBJECTIVES: To compare longitudinal trends in concentrations of polychlorinated biphenyls (PCBs) and organochlorine pesticides (OCPs) in T2DM cases and controls, and to examine prospective and cross-sectional associations between PCBs, OCPs and T2DM at different time-points before and after T2DM diagnosis in cases. METHODS: We conducted a longitudinal, nested case-control study (1986-2016) of 116 T2DM cases and 139 controls from the Tromsø Study. All participants had three blood samples collected before T2DM diagnosis in cases, and up to two samples thereafter. We used linear mixed-effect models to assess temporal changes of POPs within and between T2DM cases and controls, and logistic regression models to investigate the associations between different POPs and T2DM at different time-points. RESULTS: PCBs, trans-nonachlor, cis-nonachlor, oxychlordane, cis-heptachlor epoxide, p,p'-DDE, and p,p'-DDT declined more slowly in cases than controls, whereas ß-HCH and HCB declined similarly in both groups. Most POPs showed positive associations between both pre- and post-diagnostic concentrations and T2DM, though effect estimates were imprecise. These associations were most consistent for cis-heptachlor epoxide. DISCUSSION: The observed positive associations between certain POPs and T2DM may be because of higher POP concentrations within prospective T2DM cases, due to slower temporal declines as compared to controls.

Plasma concentrations of persistent organic pollutants and pancreatic cancer risk.

BACKGROUND: Findings and limitations of previous studies on persistent organic pollutants (POPs) and pancreatic cancer risk support conducting further research in prospective cohorts. METHODS: We conducted a prospective case-control study nested within the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort. Participants were 513 pancreatic cancer cases and 1020 matched controls. Concentrations of 22 POPs were measured in plasma collected at baseline. RESULTS: Some associations were observed at higher concentrations of p, p'-DDT, trans-nonachlor, ß-hexachlorocyclohexane and the sum of six organochlorine pesticides and of 16 POPs. The odds ratio (OR) for the upper quartile of trans-nonachlor was 1.55 (95% confidence interval 1.06-2.26; P for trend = 0.025). Associations were stronger in the groups predefined as most valid (participants having fasted >6 h, with microscopic diagnostic confirmation, normal weight, and never smokers), and as most relevant (follow-up ≥10 years). Among participants having fasted >6 h, the ORs were relevant for 10 of 11 exposures. Higher ORs were also observed among cases with microscopic confirmation than in cases with a clinical diagnosis, and among normal-weight participants than in the rest of participants. Among participants with a follow-up ≥10 years, estimates were higher than in participants with a shorter follow-up (for trans-nonachlor: OR = 2.14, 1.01 to 4.53, P for trend = 0.035). Overall, trans-nonachlor, three PCBs and the two sums of POPs were the exposures most clearly associated with pancreatic cancer risk. CONCLUSIONS: Individually or in combination, most of the 22 POPs analysed did not or only moderately increased the risk of pancreatic cancer.

Intraindividual Long-term Immune Marker Stability in Plasma Samples Collected in Median 9.4 Years Apart in 304 Adult Cancer-free Individuals.

BACKGROUND: Changes in immune marker levels in the blood could be used to improve the early detection of tumor-associated inflammatory processes. To increase predictiveness and utility in cancer detection, intraindividual long-term stability in cancer-free individuals is critical for biomarker candidates as to facilitate the detection of deviation from the norm. METHODS: We assessed intraindividual long-term stability for 19 immune markers (IL10, IL13, TNFα, CXCL13, MCP-3, MIP-1α, MIP-1ß, fractalkine, VEGF, FGF-2, TGFα, sIL2Rα, sIL6R, sVEGF-R2, sTNF-R1, sTNF-R2, sCD23, sCD27, and sCD30) in 304 cancer-free individuals. Repeated blood samples were collected up to 20 years apart. Intraindividual reproducibility was assessed by calculating intraclass correlation coefficients (ICC) using a linear mixed model. RESULTS: ICCs indicated fair to good reproducibility (ICCs ≥ 0.40 and < 0.75) for 17 of 19 investigated immune markers, including IL10, IL13, TNFα, CXCL13, MCP-3, MIP-1α, MIP-1ß, fractalkine, VEGF, FGF-2, TGFα, sIL2Rα, sIL6R, sTNF-R1, sTNF-R2, sCD27, and sCD30. Reproducibility was strong (ICC ≥ 0.75) for sCD23, while reproducibility was poor (ICC < 0.40) for sVEGF-R2. Using a more stringent criterion for reproducibility (ICC ≥ 0.55), we observed either acceptable or better reproducibility for IL10, IL13, CXCL13, MCP-3, MIP-1α, MIP-1ß, VEGF, FGF-2, sTNF-R1, sCD23, sCD27, and sCD30. CONCLUSIONS: IL10, IL13, CXCL13, MCP-3, MIP-1α, MIP-1ß, VEGF, FGF-2, sTNF-R1, sCD23, sCD27, and sCD30 displayed ICCs consistent with intraindividual long-term stability in cancer-free individuals. IMPACT: Our data support using these markers in prospective longitudinal studies seeking early cancer detection biomarkers.

MercuNorth - monitoring mercury in pregnant women from the Arctic as a baseline to assess the effectiveness of the Minamata Convention.

Exposure to mercury (Hg) is a global concern, particularly among Arctic populations that rely on the consumption of marine mammals and fish which are the main route of Hg exposure for Arctic populations.The MercuNorth project was created to establish baseline Hg levels across several Arctic regions during the period preceding the Minamata Convention. Blood samples were collected from 669 pregnant women, aged 18-44 years, between 2010 and 2016 from sites across the circumpolar Arctic including Alaska (USA), Nunavik (Canada), Greenland, Iceland, Norway, Sweden, Northern Lapland (Finland) and Murmansk Oblast (Russia). Descriptive statistics were calculated, multiple pairwise comparisons were made between regions, and unadjusted linear trend analyses were performed.Geometric mean concentrations of total Hg were highest in Nunavik (5.20 µg/L)  and Greenland (3.79 µg/L), followed by Alaska (2.13 µg/L), with much lower concentrations observed in the other regions (ranged between 0.48 and 1.29 µg/L). In Nunavik, Alaska and Greenland, blood Hg concentrations have decreased significantly since 1992, 2000 and 2010 respectively with % annual decreases of 4.7%, 7.5% and 2.7%, respectively.These circumpolar data combined with fish and marine mammal consumption data can be used for assessing long-term Hg trends and the effectiveness of the Minamata Convention.

Novel Biomarkers of Habitual Alcohol Intake and Associations With Risk of Pancreatic and Liver Cancers and Liver Disease Mortality.

BACKGROUND: Alcohol is an established risk factor for several cancers, but modest alcohol-cancer associations may be missed because of measurement error in self-reported assessments. Biomarkers of habitual alcohol intake may provide novel insight into the relationship between alcohol and cancer risk. METHODS: Untargeted metabolomics was used to identify metabolites correlated with self-reported habitual alcohol intake in a discovery dataset from the European Prospective Investigation into Cancer and Nutrition (EPIC; n = 454). Statistically significant correlations were tested in independent datasets of controls from case-control studies nested within EPIC (n = 280) and the Alpha-Tocopherol, Beta-Carotene Cancer Prevention (ATBC; n = 438) study. Conditional logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for associations of alcohol-associated metabolites and self-reported alcohol intake with risk of pancreatic cancer, hepatocellular carcinoma (HCC), liver cancer, and liver disease mortality in the contributing studies. RESULTS: Two metabolites displayed a dose-response association with self-reported alcohol intake: 2-hydroxy-3-methylbutyric acid and an unidentified compound. A 1-SD (log2) increase in levels of 2-hydroxy-3-methylbutyric acid was associated with risk of HCC (OR = 2.54, 95% CI = 1.51 to 4.27) and pancreatic cancer (OR = 1.43, 95% CI = 1.03 to 1.99) in EPIC and liver cancer (OR = 2.00, 95% CI = 1.44 to 2.77) and liver disease mortality (OR = 2.16, 95% CI = 1.63 to 2.86) in ATBC. Conversely, a 1-SD (log2) increase in questionnaire-derived alcohol intake was not associated with HCC or pancreatic cancer in EPIC or liver cancer in ATBC but was associated with liver disease mortality (OR = 2.19, 95% CI = 1.60 to 2.98) in ATBC. CONCLUSIONS: 2-hydroxy-3-methylbutyric acid is a candidate biomarker of habitual alcohol intake that may advance the study of alcohol and cancer risk in population-based studies.

Pre- and post-diagnostic blood profiles of chlorinated persistent organic pollutants and metabolic markers in type 2 diabetes mellitus cases and controls; a pilot study.

OBJECTIVE: Several risk factors for type 2 diabetes mellitus (T2DM) are also associated with blood concentrations of persistent organic pollutants (POPs), and factors related to the disease may affect POP concentrations, and subsequent associations between POPs and T2DM. The purpose of this pilot study was to investigate the change in concentrations of lipids, hormones and POPs pre- and post-diagnosis in T2DM cases compared to healthy controls and their associations with T2DM. METHODS: We measured POPs, lipids, and thyroid and steroid hormones in plasma from 44 female cases collected prior to (pre-diagnostic) and following (post-diagnostic) T2DM diagnosis, and in 44 healthy female age-matched controls. We compared cross-sectional differences and longitudinal changes within and between matched cases and controls with t-tests and multivariable linear regression models. Associations between POP concentrations and T2DM were investigated using conditional logistic regression. RESULTS: Between the pre- and post-diagnostic measurement, cases developed more favorable lipid profiles and the longitudinal changes in lipid-normalized concentrations of non-dioxin-like polychlorinated biphenyls (PCBs), dioxin-like PCBs, beta-hexachlorocyclohexane (HCH), HCB, and 1,1,1-trichloro-2,2-bis(4-chlorophenyl) ethane (p,p'-DDE) differed significantly between cases and controls. The longitudinal changes in POPs were mainly driven by changes in bodyweight, total lipids and T2DM status. Cases had significantly higher pre-diagnostic concentrations of POPs and triglycerides, and lower concentrations of high-density lipoprotein cholesterol and free thyroxin than controls. Pre-diagnostic POP concentrations were not significantly associated with incident T2DM, whereas several post-diagnostic POP concentrations were significantly positively associated with prevalent T2DM. CONCLUSIONS: This pilot study suggests that factors related to T2DM affect blood concentrations of POPs and may partly explain the positive associations between POPs and T2DM.

Plasma metabolites associated with exposure to perfluoroalkyl substances and risk of type 2 diabetes - A nested case-control study.

Perfluoroalkyl substances (PFAS) are widespread persistent environmental pollutants. There is evidence that PFAS induce metabolic perturbations in humans, but underlying mechanisms are still unknown. In this exploratory study, we investigated PFAS-related plasma metabolites for their associations with type 2 diabetes (T2D) to gain potential mechanistic insight in these perturbations. We used untargeted LC-MS metabolomics to find metabolites related to PFAS exposures in a case-control study on T2D (n = 187 matched pairs) nested within the Västerbotten Intervention Programme cohort. Following principal component analysis (PCA), six PFAS measured in plasma appeared in two groups: 1) perfluorononanoic acid, perfluorodecanoic acid and perfluoroundecanoic acid and 2) perfluorohexane sulfonic acid, perfluorooctane sulfonic acid and perfluorooctanoic acid. Using a random forest algorithm, we discovered metabolite features associated with individual PFAS and PFAS exposure groups which were subsequently investigated for associations with risk of T2D. PFAS levels correlated with 171 metabolite features (0.16 ≤ |r| ≤ 0.37, false discovery rate (FDR) adjusted p < 0.05). Out of these, 35 associated with T2D (p < 0.05), with 7 remaining after multiple testing adjustment (FDR < 0.05). PCA of the 35 PFAS- and T2D-related metabolite features revealed two patterns, dominated by glycerophospholipids and diacylglycerols, with opposite T2D associations. The glycerophospholipids correlated positively with PFAS and associated inversely with risk for T2D (Odds Ratio (OR) per 1 standard deviation (1-SD) increase in metabolite PCA pattern score = 0.2; 95% Confidence Interval (CI) = 0.1-0.4). The diacylglycerols also correlated positively with PFAS, but they associated with increased risk for T2D (OR per 1-SD = 1.9; 95% CI = 1.3-2.7). These results suggest that PFAS associate with two groups of lipid species with opposite relations to T2D risk.

Blood Transcriptome Response to Environmental Metal Exposure Reveals Potential Biological Processes Related to Alzheimer's Disease.

Alzheimer's disease (AD) is a neurodegenerative disease which is manifested by a progressive and irreversible decline of cognition, memory loss, a shortened attention span, and changes in personality. Aging and genetic pre-dispositions, particularly the presence of a specific form of apolipoprotein E (APOE), are main risk factors of sporadic AD; however, a large body of evidence has shown that multiple environmental factors, including exposure to toxic metals, increase the risk for late onset AD. Lead (Pb) and cadmium (Cd) are ubiquitous toxic metals with a wide range of applications resulting in global distribution in the environment and exposure of all living organisms on earth. In addition to being classified as carcinogenic (Cd) and possibly carcinogenic (Pb) to humans by the International Agency for Research on Cancer, both compounds disrupt metal homeostasis and can cause toxic responses at the cellular and organismal levels. Pb toxicity targets the central nervous system and evidence for that has emerged also for Cd. Recent epidemiological studies show that both metals possibly are etiological factors of multiple neurodegenerative diseases, including Alzheimer's disease (AD). To further explore the association between metal exposure and AD risk we applied whole transcriptome gene expression analysis in peripheral blood leukocytes (PBLs) from 632 subjects of the general population, taken from the EnviroGenomarkers project. We used linear mixed effect models to associate metal exposure to gene expression after adjustment for gender, age, BMI, smoking, and alcohol consumption. For Pb exposure only few associations were identified, including a downregulation of the human eukaryotic translation initiation factor 5 (eIF5). In contrast, Cd exposure, particularly in males, revealed a much stronger transcriptomic response, featuring multiple pathways related to pathomolecular mechanisms of AD, such as endocytosis, neutrophil degranulation, and Interleukin-7 signaling. A gender stratified analysis revealed that the Cd responses were male-specific and included a downregulation of the APOE gene in men. This exploratory study revealed novel hypothetical findings which might contribute to the understanding of the neurotoxic effects of chronic Pb and Cd exposure and possibly improve our knowledge on the molecular mechanisms linking metal exposure to AD risk.

Troponin T but not C reactive protein is associated with future surgery for aortic stenosis: a population-based nested case-referent study.

AIMS: High-sensitivity troponin T (hs-TnT) and high-sensitivity C reactive protein (hs-CRP) may convey prognostic information in patients with aortic stenosis (AS). This study evaluated if hs-TnT and hs-CRP associate with myocardial mass, and risk of future surgery for AS. METHODS: In total, 336 patients (48% women) with surgery for AS with previous participation in large population surveys were identified. Preoperatively, myocardial mass and the presence of coronary artery disease (CAD) were assessed. Two matched referents were allocated for each case, and hs-TnT and hs-CRP were determined in stored plasma from the baseline survey. Conditional logistic regression analysis was used to estimate the risk (OR (95% CI)) related to one (natural logarithm) SD increase in hs-TnT and hs-CRP. Kaplan-Mayer and Cox regression analyses were used to evaluate time to surgery. RESULTS: Median age (IQR) was 59.8 (10.3) years at survey, and median time between survey and surgery was 10.9 (9.3) years. Hs-TnT was independently associated with surgery for AS (1.24 (1.06-1.44)) irrespective of CAD, whereas Hs-CRP was not (1.05 (0.90-1.22)). Elevated hs-TnT levels at survey associated with shorter time to surgery (p<0.001), and with increased myocardial mass (p=0.002). Hs-CRP did not associate with time to surgery or with myocardial mass. CONCLUSIONS: Hs-TnT-but not hs-CRP-was associated with increased risk of-and shorter time to-future surgery for AS. Hs-TnT associated with myocardial mass at surgery which indicates that hs-TnT could be a potential biomarker for determining intervention.

High frequency of intermediary alleles in the HTT gene in Northern Sweden - The Swedish Huntingtin Alleles and Phenotype (SHAPE) study.

Trinucleotide (CAG) repeat expansions longer than 39 in the huntingtin (HTT) gene cause Huntington's disease (HD). The frequency of intermediate alleles (IA) with a length of 27-35 in the general population is not fully known, but studied in specific materials connected to the incidence of HD. The Swedish Huntingtin Alleles and Phenotype (SHAPE) study aims to assess the frequency of trinucleotide repeat expansions in the HTT gene in north Sweden. 8260 individuals unselected for HD from the counties of Norr- and Västerbotten in the north of Sweden were included. DNA samples were obtained and analysis of the HTT gene was performed, yielding data on HTT gene expansion length in 7379 individuals. A high frequency of intermediate alleles, 6.8%, was seen. Also, individuals with repeat numbers lower than ever previously reported (<5) were found. These results suggest a high frequency of HD in the norther parts of Sweden. Subsequent analyses may elucidate the influence of IA:s on traits other than HD.

Unexpected Acetylation of Endogenous Aliphatic Amines by Arylamine N-Acetyltransferase NAT2.

N-Acetyltransferases play critical roles in the deactivation and clearance of xenobiotics, including clinical drugs. NAT2 has been classified as an arylamine N-acetyltransferase that mainly converts aromatic amines, hydroxylamines, and hydrazines. Herein, we demonstrate that the human arylamine N-acetyltransferase NAT2 also acetylates aliphatic endogenous amines. Metabolomic analysis and chemical synthesis revealed increased intracellular concentrations of mono- and diacetylated spermidine in human cell lines expressing the rapid compared to the slow acetylator NAT2 phenotype. The regioselective N8 -acetylation of monoacetylated spermidine by NAT2 answers the long-standing question of the source of diacetylspermidine. We also identified selective acetylation of structurally diverse alkylamine-containing drugs by NAT2, which may contribute to variations in patient responses. The results demonstrate a previously unknown functionality and potential regulatory role for NAT2, and we suggest that this enzyme should be considered for re-classification.

Mild impairment of renal function (shrunken pore syndrome) is associated with increased risk for future surgery for aortic stenosis.

Recently, a new approach was proposed to detect mild impairment in renal function: a reduced ratio between estimated glomerular filtration rate (eGFR) calculated by cystatin C and eGFR calculated by creatinine. We aimed to evaluate if this ratio is associated with aortic stenosis (AS) requiring surgery. We identified 336 patients that first participated in population surveys and later underwent surgery for AS (median age [interquartile range] 59.8 [10.3] years at survey and 68.3 [12.7] at surgery, 48% females). For each patient, two matched referents were allocated. Cystatin C and creatinine were determined in stored plasma. eGFRcystatin C and eGFRcreatinine and their ratio were estimated. Conditional logistic regression analyses were used to estimate the risk (odds ratio (OR) with [95% confidence interval (CI)]) related to one (ln) standard deviation increase in the ratio between eGFRcystatin C and eGFRcreatinine. A high ratio was associated with lower risk for AS requiring surgery (OR [95% CI]) (OR 0.84 [0.73-0.97]), especially in women (0.74 [0.60-0.92] vs. 0.93 [0.76-1.13] in men). After further stratification for coronary artery disease (CAD), the association remained in women with CAD but not in women without CAD (0.60 [0.44-0.83] and 0.89 [0.65-1.23], respectively). In conclusion, a high ratio between eGFRcystatin C and eGFRcreatinine was associated with lower risk for surgery for AS, especially in women. Mild impairment of renal function is thus associated with future risk for AS requiring surgery.