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Biological age (BA) captures detrimental age-related changes. The best-known and most-used BA indicators include DNA methylation-based epigenetic clocks and telomere length (TL). The most common biological sample material for epidemiological aging studies, whole blood, is composed of different cell types. We aimed to compare differences in BAs between blood cell types and assessed the BA indicators' cell type-specific associations with chronological age (CA). An analysis of DNA methylation-based BA indicators, including TL, methylation level at cg16867657 in ELOVL2, as well as the Hannum, Horvath, DNAmPhenoAge, and DunedinPACE epigenetic clocks, was performed on 428 biological samples of 12 blood cell types. BA values were different in the majority of the pairwise comparisons between cell types, as well as in comparison to whole blood (p < 0.05). DNAmPhenoAge showed the largest cell type differences, up to 44.5 years and DNA methylation-based TL showed the lowest differences. T cells generally had the "youngest" BA values, with differences across subsets, whereas monocytes had the "oldest" values. All BA indicators, except DunedinPACE, strongly correlated with CA within a cell type. Some differences such as DNAmPhenoAge-difference between naïve CD4 + T cells and monocytes were constant regardless of the blood donor's CA (range 20-80 years), while for DunedinPACE they were not. In conclusion, DNA methylation-based indicators of BA exhibit cell type-specific characteristics. Our results have implications for understanding the molecular mechanisms underlying epigenetic clocks and underscore the importance of considering cell composition when utilizing them as indicators for the success of aging interventions., (© 2024. The Author(s).)

Eastern and Western Finns show a striking difference in coronary heart disease-related mortality; genetics is a known contributor for this discrepancy. Here, we discuss the potential role of DNA methylation in mediating the discrepancy in cardiometabolic disease-risk phenotypes between the sub-populations. We used data from the Young Finns Study ( n  = 969) to compare the genome-wide DNA methylation levels of East- and West-originating Finns. We identified 21 differentially methylated loci (FDR < 0.05; Δβ >2.5%) and 7 regions (smoothed FDR < 0.05; CpGs ≥ 5). Methylation at all loci and regions associates with genetic variants ( p  < 5 × 10 -8 ). Independently of genetics, methylation at 11 loci and 4 regions associates with transcript expression, including genes encoding zinc finger proteins. Similarly, methylation at 5 loci and 4 regions associates with cardiometabolic disease-risk phenotypes including triglycerides, glucose, cholesterol, as well as insulin treatment. This analysis was also performed in LURIC ( n  = 2371), a German cardiovascular patient cohort, and results replicated for the association of methylation at cg26740318 and DMR&#95;11p15 with diabetes-related phenotypes and methylation at DMR&#95;22q13 with triglyceride levels. Our results indicate that DNA methylation differences between East and West Finns may have a functional role in mediating the cardiometabolic disease discrepancy between the sub-populations.

Non-coding 886 ( nc886 , vtRNA2-1 ) is the only human polymorphically imprinted gene, in which the methylation status is not determined by genetics. Existing literature regarding the establishment, stability and consequences of the methylation pattern, as well as the nature and function of the nc886 RNAs transcribed from the locus, are contradictory. For example, the methylation status of the locus has been reported to be stable through life and across somatic tissues, but also susceptible to environmental effects. The nature of the produced nc886 RNA(s) has been redefined multiple times, and in carcinogenesis, these RNAs have been reported to have conflicting roles. In addition, due to the bimodal methylation pattern of the nc886 locus, traditional genome-wide methylation analyses can lead to false-positive results, especially in smaller datasets. Herein, we aim to summarize the existing literature regarding nc886 , discuss how the characteristics of nc886 give rise to contradictory results, as well as to reinterpret, reanalyse and, where possible, replicate the results presented in the current literature. We also introduce novel findings on how the distribution of the nc886 methylation pattern is associated with the geographical origins of the population and describe the methylation changes in a large variety of human tumours. Through the example of this one peculiar genetic locus and RNA, we aim to highlight issues in the analysis of DNA methylation and non-coding RNAs in general and offer our suggestions for what should be taken into consideration in future analyses.

Background: Studies have shown that cardiovascular health (CVH) is related to depression. We aimed to identify gene networks jointly associated with depressive symptoms and cardiovascular health metrics using the whole blood transcriptome., Materials and Methods: We analyzed human blood transcriptomic data to identify gene co-expression networks, termed gene modules, shared by Beck's depression inventory (BDI-II) scores and cardiovascular health (CVH) metrics as markers of depression and cardiovascular health, respectively. The BDI-II scores were derived from Beck's Depression Inventory, a 21-item self-report inventory that measures the characteristics and symptoms of depression. CVH metrics were defined according to the American Heart Association criteria using seven indices: smoking, diet, physical activity, body mass index (BMI), blood pressure, total cholesterol, and fasting glucose. Joint association of the modules, identified with weighted co-expression analysis, as well as the member genes of the modules with the markers of depression and CVH were tested with multivariate analysis of variance (MANOVA)., Results: We identified a gene module with 256 genes that were significantly correlated with both the BDI-II score and CVH metrics. Based on the MANOVA test results adjusted for age and sex, the module was associated with both depression and CVH markers. The three most significant member genes in the module were YOD1 , RBX1 , and LEPR . Genes in the module were enriched with biological pathways involved in brain diseases such as Alzheimer's, Parkinson's, and Huntington's., Conclusions: The identified gene module and its members can provide new joint biomarkers for depression and CVH., Competing Interests: BHM, NM, TL and PPM were affiliated with Fimlab Laboratories. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Mishra, Raitoharju, Mononen, Saarinen, Viikari, Juonala, Hutri-Kähönen, Kähönen, Raitakari, Lehtimäki and Mishra.)

Genome-wide association studies of human personality have been carried out, but transcription of the whole genome has not been studied in relation to personality in humans. We collected genome-wide expression profiles of adults to characterize the regulation of expression and function in genes related to human personality. We devised an innovative multi-omic approach to network analysis to identify the key control elements and interactions in multi-modular networks. We identified sets of transcribed genes that were co-expressed in specific brain regions with genes known to be associated with personality. Then we identified the minimum networks for the co-localized genes using bioinformatic resources. Subjects were 459 adults from the Young Finns Study who completed the Temperament and Character Inventory and provided peripheral blood for genomic and transcriptomic analysis. We identified an extrinsic network of 45 regulatory genes from seed genes in brain regions involved in self-regulation of emotional reactivity to extracellular stimuli (e.g., self-regulation of anxiety) and an intrinsic network of 43 regulatory genes from seed genes in brain regions involved in self-regulation of interpretations of meaning (e.g., production of concepts and language). We discovered that interactions between the two networks were coordinated by a control hub of 3 miRNAs and 3 protein-coding genes shared by both. Interactions of the control hub with proteins and ncRNAs identified more than 100 genes that overlap directly with known personality-related genes and more than another 4000 genes that interact indirectly. We conclude that the six-gene hub is the crux of an integrative network that orchestrates information-transfer throughout a multi-modular system of over 4000 genes enriched in liquid-liquid-phase-separation (LLPS)-related RNAs, diverse transcription factors, and hominid-specific miRNAs and lncRNAs. Gene expression networks associated with human personality regulate neuronal plasticity, epigenesis, and adaptive functioning by the interactions of salience and meaning in self-awareness., (© 2024. The Author(s).)

Schizophrenia is often regarded as a disorder of premature aging. We investigated (a) whether polygenic risk for schizophrenia (PRS sch ) relates to pace of epigenetic aging and (b) whether personal dispositions toward active and emotionally close relationships protect against accelerated epigenetic aging in individuals with high PRS sch . The sample came from the population-based Young Finns Study (n = 1348). Epigenetic aging was measured with DNA methylation aging algorithms such as AgeAccel Hannum , EEAA Hannum , IEAA Hannum , IEAA Horvath , AgeAccel Horvath , AgeAccel Pheno , AgeAccel Grim , and DunedinPACE. A PRS sch was calculated using summary statistics from the most comprehensive genome-wide association study of schizophrenia to date. Social dispositions were assessed in terms of extraversion, sociability, reward dependence, cooperativeness, and attachment security. We found that PRS sch did not have a statistically significant effect on any studied indicator of epigenetic aging. Instead, PRS sch had a significant interaction with reward dependence (p = 0.001-0.004), cooperation (p = 0.009-0.020), extraversion (p = 0.019-0.041), sociability (p = 0.003-0.016), and attachment security (p = 0.007-0.014) in predicting AgeAccel Hannum , EEAA Hannum , or IEAA Hannum . Specifically, participants with high PRS sch appeared to display accelerated epigenetic aging at higher (vs. lower) levels of extraversion, sociability, attachment security, reward dependence, and cooperativeness. A rather opposite pattern was evident for those with low PRS sch . No such interactions were evident when predicting the other indicators of epigenetic aging. In conclusion, against our hypothesis, frequent social interactions may relate to accelerated epigenetic aging in individuals at risk for psychosis. We speculate that this may be explained by social-cognitive impairments (perceiving social situations as overwhelming or excessively arousing) or ending up in less supportive or deviant social groups., (© 2023 The Authors. Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Advanced integrative analysis of DNA methylation and transcriptomics data may provide deeper insights into smoke-induced epigenetic alterations, their effects on gene expression and related biological processes, linking cigarette smoking and related diseases. We hypothesize that accumulation of DNA methylation changes in CpG sites across genomic locations of different genes might have biological significance. We tested the hypothesis by performing gene set based integrative analysis of blood DNA methylation and transcriptomics data to identify potential transcriptomic consequences of smoking via changes in DNA methylation in the Young Finns Study (YFS) participants ( n  = 1114, aged 34-49 years, women: 54%, men: 46%). First, we performed epigenome-wide association study (EWAS) of smoking. We then defined sets of genes based on DNA methylation status within their genomic regions, for example, sets of genes containing hyper- or hypomethylated CpG sites in their body or promoter regions. Gene set analysis was performed using transcriptomics data from the same participants. Two sets of genes, one containing 49 genes with hypomethylated CpG sites in their body region and the other containing 33 genes with hypomethylated CpG sites in their promoter region, were differentially expressed among the smokers. Genes in the two gene sets are involved in bone formation, metal ion transport, cell death, peptidyl-serine phosphorylation, and cerebral cortex development process, revealing epigenetic-transcriptomic pathways to smoking-related diseases such as osteoporosis, atherosclerosis, and cognitive impairment. These findings contribute to a deeper understanding of the pathophysiology of smoking-related diseases and may provide potential therapeutic targets.

Metabolic dysfunction, partly driven by altered liver function, predisposes to coronary artery disease (CAD), but the role of liver in vulnerable atherosclerotic plaque development remains unclear. Here we produced hepatocyte-like cells (HLCs) from 27 induced pluripotent stem cell (iPSC) lines derived from 15 study subjects with stable CAD (n = 5), acute CAD (n = 5) or healthy controls (n = 5). We performed a miRNA microarray screening throughout the differentiation, as well as compared iPSC-HLCs miRNA profiles of the patient groups to identify miRNAs involved in the development of CAD. MicroRNA profile changed during differentiation and started to resemble that of the primary human hepatocytes. In the microarray, 35 and 87 miRNAs were statistically significantly deregulated in the acute and stable CAD patients, respectively, compared to controls. Down-regulation of miR-149-5p, -92a-3p and -221-3p, and up-regulation of miR-122-5p was verified in the stable CAD patients when compared to other groups. The predicted targets of deregulated miRNAs were enriched in pathways connected to insulin signalling, inflammation and lipid metabolism. The iPSC-HLCs derived from stable CAD patients with extensive lesions had a distinct genetic miRNA profile possibly linked to metabolic dysfunction, potentially explaining the susceptibility to developing CAD. The iPSC-HLCs from acute CAD patients with only the acute rupture in otherwise healthy coronaries did not present a distinct miRNA profile, suggesting that hepatic miRNAs do not explain susceptibility to plaque rupture., (© 2023. The Author(s).)

Aim: We aimed at identifying the shared biological processes underlying atherosclerosis-osteoporosis co/multimorbidity., Methods: We performed gene set analysis (GSA) of whole-blood transcriptomic data to identify biological processes shared by the early markers of these two diseases. Early markers of diseases, carotid intima-media thickness (CIMT) for atherosclerosis and trabecular bone mineral density (BMD) from distal radius and tibia for osteoporosis, were used to categorize the study participants into cases and controls. Participants with high CIMT (>90th percentile) were defined as cases for subclinical atherosclerosis. Study population-based T-scores for BMD were calculated and T-score ≤ -1 was used for the definition of low BMD cases i.e., early indicator of osteoporosis., Results: We did not identify any gene sets jointly associated with early markers of atherosclerosis and osteoporosis. We identified three novel and replicated 234 gene sets significantly associated with high CIMT with false discovery rate (FDR) ≤ 0.01. Only two genes, both related to the immune system, were identified to be associated with high CIMT by traditional differential gene expression analysis. However, none of the studied gene sets or individual genes were significantly associated with tibial or radial BMD. The three novel CIMT associated gene sets contained genes involved in copper homeostasis, neural crest cell migration and nicotinate and nicotinamide metabolism. The 234 replicated gene sets in this study are related to the immune system, hypoxia and apoptosis, consistent with the existing literature on atherosclerosis., Conclusions: This study identified novel biological processes associated with high CIMT but not with reduced BMD., Competing Interests: Declaration of competing interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Aims & methods: The aim of this study was to characterize the methylation level of a polymorphically imprinted gene, VTRNA2-1 / nc886 , in human populations and somatic tissues.48 datasets, consisting of more than 30 tissues and >30,000 individuals, were used. Results: nc886 methylation status is associated with twin status and ethnic background, but the variation between populations is limited. Monozygotic twin pairs present concordant methylation, whereas ∼30% of dizygotic twin pairs present discordant methylation in the nc886 locus. The methylation levels of nc886 are uniform across somatic tissues, except in cerebellum and skeletal muscle. Conclusion: The nc886 imprint may be established in the oocyte, and, after implantation, the methylation status is stable, excluding a few specific tissues.

Mitochondria have a complex communication network with the surrounding cell and can alter nuclear DNA methylation (DNAm). Variation in the mitochondrial DNA (mtDNA) has also been linked to differential DNAm. Genome-wide association studies have identified numerous DNAm quantitative trait loci, but these studies have not examined the mitochondrial genome. Herein, we quantified nuclear DNAm from blood and conducted a mitochondrial genome-wide association study of DNAm, with an additional emphasis on sex- and prediabetes-specific heterogeneity. We used the Young Finns Study (n = 926) with sequenced mtDNA genotypes as a discovery sample and sought replication in the Ludwigshafen Risk and Cardiovascular Health study (n = 2317). We identified numerous significant associations in the discovery phase (P < 10-9), but they were not replicated when accounting for multiple testing. In total, 27 associations were nominally replicated with a P < 0.05. The replication analysis presented no evidence of sex- or prediabetes-specific heterogeneity. The 27 associations were included in a joint meta-analysis of the two cohorts, and 19 DNAm sites associated with mtDNA variants, while four other sites showed haplogroup associations. An expression quantitative trait methylation analysis was performed for the identified DNAm sites, pinpointing two statistically significant associations. This study provides evidence of a mitochondrial genetic control of nuclear DNAm with little evidence found for sex- and prediabetes-specific effects. The lack of a comparable mtDNA data set for replication is a limitation in our study and further studies are needed to validate our results., (© The Author(s) 2021. Published by Oxford University Press.)

Background: In humans, the nc886 locus is a polymorphically imprinted metastable epiallele. Periconceptional conditions have an effect on the methylation status of nc886, and further, this methylation status is associated with health outcomes in later life, in line with the Developmental Origins of Health and Disease (DOHaD) hypothesis. Animal models would offer opportunities to study the associations between periconceptional conditions, nc886 methylation status and metabolic phenotypes further. Thus, we set out to investigate the methylation pattern of the nc886 locus in non-human mammals., Data: We obtained DNA methylation data from the data repository GEO for mammals, whose nc886 gene included all three major parts of nc886 and had sequency similarity of over 80% with the human nc886. Our final sample set consisted of DNA methylation data from humans, chimpanzees, bonobos, gorillas, orangutangs, baboons, macaques, vervets, marmosets and guinea pigs., Results: In human data sets the methylation pattern of nc886 locus followed the expected bimodal distribution, indicative of polymorphic imprinting. In great apes, we identified a unimodal DNA methylation pattern with 50% methylation level in all individuals and in all subspecies. In Old World monkeys, the between individual variation was greater and methylation on average was close to 60%. In guinea pigs the region around the nc886 homologue was non-methylated. Results obtained from the sequence comparison of the CTCF binding sites flanking the nc886 gene support the results on the DNA methylation data., Conclusions: Our results indicate that unlike in humans, nc886 is not a polymorphically imprinted metastable epiallele in non-human primates or in guinea pigs, thus implying that animal models are not applicable for nc886 research. The obtained data suggests that the nc886 region may be classically imprinted in great apes, and potentially also in Old World monkeys, but not in guinea pigs., Competing Interests: The authors have declared that no competing interests exist.

Background: Women with a history of complications of pregnancy, including hypertensive disorders, gestational diabetes or an infant fetal growth restriction or preterm birth, are at higher risk for cardiovascular disease later in life. We aimed to examine differences in maternal DNA methylation following pregnancy complications., Methods: Data on women participating in the Young Finns study (n = 836) were linked to the national birth registry. DNA methylation in whole blood was assessed using the Infinium Methylation EPIC BeadChip. Epigenome-wide analysis was conducted on differential CpG methylation at 850 K sites. Reproductive history was also modeled as a predictor of four epigenetic age indices., Results: Fourteen significant differentially methylated sites were found associated with both history of pre-eclampsia and overall hypertensive disorders of pregnancy. No associations were found between reproductive history and any epigenetic age acceleration measure., Conclusions: Differences in epigenetic methylation profiles could represent pre-existing risk factors, or changes that occurred as a result of experiencing these complications., (© 2021. The Author(s).)

Phylogenetic, developmental, and brain-imaging studies suggest that human personality is the integrated expression of three major systems of learning and memory that regulate (1) associative conditioning, (2) intentionality, and (3) self-awareness. We have uncovered largely disjoint sets of genes regulating these dissociable learning processes in different clusters of people with (1) unregulated temperament profiles (i.e., associatively conditioned habits and emotional reactivity), (2) organized character profiles (i.e., intentional self-control of emotional conflicts and goals), and (3) creative character profiles (i.e., self-aware appraisal of values and theories), respectively. However, little is known about how these temperament and character components of personality are jointly organized and develop in an integrated manner. In three large independent genome-wide association studies from Finland, Germany, and Korea, we used a data-driven machine learning method to uncover joint phenotypic networks of temperament and character and also the genetic networks with which they are associated. We found three clusters of similar numbers of people with distinct combinations of temperament and character profiles. Their associated genetic and environmental networks were largely disjoint, and differentially related to distinct forms of learning and memory. Of the 972 genes that mapped to the three phenotypic networks, 72% were unique to a single network. The findings in the Finnish discovery sample were blindly and independently replicated in samples of Germans and Koreans. We conclude that temperament and character are integrated within three disjoint networks that regulate healthy longevity and dissociable systems of learning and memory by nearly disjoint sets of genetic and environmental influences., (© 2019. The Author(s).)

Background: Non-coding RNA 886 (nc886) is coded from a maternally inherited metastable epiallele. We set out to investigate the determinants and dynamics of the methylation pattern at the nc886 epiallele and how this methylation status associates with nc886 RNA expression. Furthermore, we investigated the associations between the nc886 methylation status or the levels of nc886 RNAs and metabolic traits in the YFS and KORA cohorts. The association between nc886 epiallele methylation and RNA expression was also validated in induced pluripotent stem cell (iPSC) lines., Results: We confirm that the methylation status of the nc886 epiallele is mostly binomial, with individuals displaying either a non- or hemi-methylated status, but we also describe intermediately and close to fully methylated individuals. We show that an individual's methylation status is associated with the mother's age and socioeconomic status, but not with the individual's own genetics. Once established, the methylation status of the nc886 epiallele remains stable for at least 25 years. This methylation status is strongly associated with the levels of nc886 non-coding RNAs in serum, blood, and iPSC lines. In addition, nc886 methylation status associates with glucose and insulin levels during adolescence but not with the indicators of glucose metabolism or the incidence of type 2 diabetes in adulthood. However, the nc886-3p RNA levels also associate with glucose metabolism in adulthood., Conclusions: These results indicate that nc886 metastable epiallele methylation is tuned by the periconceptional conditions and it associates with glucose metabolism through the expression of the ncRNAs coded in the epiallele region., (© 2021. The Author(s).)

MicroRNAs are important in prostate cancer development, progression and metastasis. The aim of this study was to test microRNA expression profile in prostate tissue obtained from prostate cancer patients for associations with various prostate cancer related factors and to pinpoint the predicted target pathways for these microRNAs. Prostate tissue samples were obtained at prostatectomy from patients participating in a trial evaluating impact of pre-operative atorvastatin on serum prostate specific antigen (PSA) and Ki-67 expression in prostate tissue. Prostate tissue microRNA expression profiles were analyzed using OpenArray ® MicroRNA Panel. Pathway enrichment analyses were conducted for predicted target genes of microRNAs that correlated significantly with studied factors. Eight microRNAs correlated significantly with studied factors of patients after Bonferroni multiple testing correction. MiR-485-3p correlated with serum HDL-cholesterol levels. In atorvastatin-treated subjects, miR-34c-5p correlated with a change in serum PSA and miR-138-3p with a change in total cholesterol. In the placebo arm, both miR-576-3p and miR-550-3p correlated with HDL-cholesterol and miR-627 with PSA. In pathway analysis, these eight microRNAs related significantly to several pathways relevant to prostate cancer. This study brings new evidence from the expression of prostate tissue microRNAs and related pathways that may link risk factors to prostate cancer and pinpoint new therapeutic possibilities.

Activation of the hypoxia-inducible factor (HIF) pathway reprograms energy metabolism. Hemoglobin (Hb) is the main carrier of oxygen. Using its normal variation as a surrogate measure for hypoxia, we explored whether lower Hb levels could lead to healthier metabolic profiles in mice and humans ( n = 7175) and used Mendelian randomization (MR) to evaluate potential causality ( n = 173,480). The results showed evidence for lower Hb levels being associated with lower body mass index, better glucose tolerance and other metabolic profiles, lower inflammatory load, and blood pressure. Expression of the key HIF target genes SLC2A4 and Slc2a1 in skeletal muscle and adipose tissue, respectively, associated with systolic blood pressure in MR analyses and body weight, liver weight, and adiposity in mice. Last, manipulation of murine Hb levels mediated changes to key metabolic parameters. In conclusion, low-end normal Hb levels may be favorable for metabolic health involving mild chronic activation of the HIF response., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution License 4.0 (CC BY).)

Preterm birth (PTB) is associated with increased risk of type 2 diabetes and neurocognitive impairment later in life. We analyzed for the first time the associations of PTB with blood miRNA levels in adulthood. We also investigated the relationship of PTB associated miRNAs and adulthood phenotypes previously linked with premature birth. Blood MicroRNA profiling, genome-wide gene expression analysis, computer-based cognitive testing battery (CANTAB) and serum NMR metabolomics were performed for Young Finns Study subjects (aged 34-49 years, full-term n = 682, preterm n = 84). Preterm birth (vs. full-term) was associated with adulthood levels of hsa-miR-29b-3p in a fully adjusted regression model (p = 1.90 × 10 -4 , FDR = 0.046). The levels of hsa-miR-29b-3p were down-regulated in subjects with PTB with appropriate birthweight for gestational age (p = 0.002, fold change [FC] = - 1.20) and specifically in PTB subjects with small birthweight for gestational age (p = 0.095, FC = - 1.39) in comparison to individuals born full term. Hsa-miR-29b-3p levels correlated with the expressions of its target-mRNAs BCL11A and CS and the gene set analysis results indicated a target-mRNA driven association between hsa-miR-29b-3p levels and Alzheimer's disease, Parkinson's disease, Insulin signaling and Regulation of Actin Cytoskeleton pathway expression. The level of hsa-miR-29b-3p was directly associated with visual processing and sustained attention in CANTAB test and inversely associated with serum levels of VLDL subclass component and triglyceride levels. In conlcusion, adult blood levels of hsa-miR-29b-3p were lower in subjects born preterm. Hsa-miR-29b-3p associated with cognitive function and may be linked with adulthood morbidities in subjects born preterm, possibly through regulation of gene sets related to neurodegenerative diseases and insulin signaling as well as VLDL and triglyceride metabolism.

We analysed whole blood genome-wide expression data to identify gene co-expression modules shared by early traits of osteoporosis and atherosclerosis. Gene expression was profiled for the Young Finns Study participants. Bone mineral density and content were measured as early traits of osteoporosis. Carotid and bulbus intima media thickness were measured as early traits of atherosclerosis. Joint association of the modules, identified with weighted co-expression analysis, with early traits of the diseases was tested with multivariate analysis. Among the six modules significantly correlated with early traits of both the diseases, two had significant (adjusted p-values (p.adj) < 0.05) and another two had suggestively significant (p.adj < 0.25) joint association with the two diseases after adjusting for age, sex, body mass index, smoking habit, alcohol consumption, and physical activity. The three most significant member genes from the significant modules were NOSIP, GXYLT2, and TRIM63 (p.adj ≤ 0.18). Genes in the modules were enriched with biological processes that have separately been found to be involved in either bone metabolism or atherosclerosis. The gene modules and their most significant member genes identified in this study support the osteoporosis-atherosclerosis comorbidity hypothesis and can provide new joint biomarkers for both diseases and their dual prevention.

Objective: This study aimed to investigate the role of cytokines as intermediates in the pathway from increased adiposity to disease., Methods: BMI and circulating levels of up to 41 cytokines were measured in individuals from three Finnish cohort studies (n = 8,293). Mendelian randomization (MR) was used to assess the impact of BMI on circulating cytokines and the impact of BMI-driven cytokines on risk of obesity-related diseases., Results: Observationally, BMI was associated with 19 cytokines. For every SD increase in BMI, causal effect estimates were strongest for hepatocyte growth factor, monocyte chemotactic protein-1 (MCP-1), and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and were as ratios of geometric means 1.13 (95% CI: 1.08-1.19), 1.08 (95% CI: 1.04-1.14), and 1.13 (95% CI: 1.04-1.21), respectively. TRAIL was associated with a small increase in the odds of coronary artery disease (odds ratio: 1.03; 95% CI: 1.00-1.06). There was inconsistent evidence for a protective role of MCP-1 against inflammatory bowel diseases., Conclusions: Observational and MR estimates of the effect of BMI on cytokine levels were generally concordant. There was little evidence for an effect of raised levels of BMI-driven cytokines on disease. These findings illustrate the challenges of MR when applied in the context of molecular mediation., (© 2021 The Authors. Obesity published by Wiley Periodicals LLC on behalf of The Obesity Society (TOS).)

High blood pressure (BP) is a major risk factor for many noncommunicable diseases. The effect of mitochondrial DNA single-nucleotide polymorphisms (mtSNPs) on BP is less known than that of nuclear SNPs. We investigated the mitochondrial genetic determinants of systolic, diastolic, and mean arterial BP. MtSNPs were determined from peripheral blood by sequencing or with genome-wide association study SNP arrays in two independent Finnish cohorts, the Young Finns Study and the Finnish Cardiovascular Study, respectively. In total, over 4200 individuals were included. The effects of individual common mtSNPs, with an additional focus on sex-specificity, and aggregates of rare mtSNPs grouped by mitochondrial genes were evaluated by meta-analysis of linear regression and a sequence kernel association test, respectively. We accounted for the predicted pathogenicity of the rare variants within protein-encoding and the tRNA regions. In the meta-analysis of 87 common mtSNPs, we did not observe significant associations with any of the BP traits. Sex-specific and rare-variant analyses did not pinpoint any significant associations either. Our results are in agreement with several previous studies suggesting that mtDNA variation does not have a significant role in the regulation of BP. Future studies might need to reconsider the mechanisms thought to link mtDNA with hypertension.

Experimental studies of learning suggest that human temperament may depend on the molecular mechanisms for associative conditioning, which are highly conserved in animals. The main genetic pathways for associative conditioning are known in experimental animals, but have not been identified in prior genome-wide association studies (GWAS) of human temperament. We used a data-driven machine learning method for GWAS to uncover the complex genotypic-phenotypic networks and environmental interactions related to human temperament. In a discovery sample of 2149 healthy Finns, we identified sets of single-nucleotide polymorphisms (SNPs) that cluster within particular individuals (i.e., SNP sets) regardless of phenotype. Second, we identified 3 clusters of people with distinct temperament profiles measured by the Temperament and Character Inventory regardless of genotype. Third, we found 51 SNP sets that identified 736 gene loci and were significantly associated with temperament. The identified genes were enriched in pathways activated by associative conditioning in animals, including the ERK, PI3K, and PKC pathways. 74% of the identified genes were unique to a specific temperament profile. Environmental influences measured in childhood and adulthood had small but significant effects. We confirmed the replicability of the 51 Finnish SNP sets in healthy Korean (90%) and German samples (89%), as well as their associations with temperament. The identified SNPs explained nearly all the heritability expected in each sample (37-53%) despite variable cultures and environments. We conclude that human temperament is strongly influenced by more than 700 genes that modulate associative conditioning by molecular processes for synaptic plasticity and long-term memory.

Human personality is 30-60% heritable according to twin and adoption studies. Hundreds of genetic variants are expected to influence its complex development, but few have been identified. We used a machine learning method for genome-wide association studies (GWAS) to uncover complex genotypic-phenotypic networks and environmental interactions. The Temperament and Character Inventory (TCI) measured the self-regulatory components of personality critical for health (i.e., the character traits of self-directedness, cooperativeness, and self-transcendence). In a discovery sample of 2149 healthy Finns, we identified sets of single-nucleotide polymorphisms (SNPs) that cluster within particular individuals (i.e., SNP sets) regardless of phenotype. Second, we identified five clusters of people with distinct profiles of character traits regardless of genotype. Third, we found 42 SNP sets that identified 727 gene loci and were significantly associated with one or more of the character profiles. Each character profile was related to different SNP sets with distinct molecular processes and neuronal functions. Environmental influences measured in childhood and adulthood had small but significant effects. We confirmed the replicability of 95% of the 42 SNP sets in healthy Korean and German samples, as well as their associations with character. The identified SNPs explained nearly all the heritability expected for character in each sample (50 to 58%). We conclude that self-regulatory personality traits are strongly influenced by organized interactions among more than 700 genes despite variable cultures and environments. These gene sets modulate specific molecular processes in brain for intentional goal-setting, self-reflection, empathy, and episodic learning and memory.

Smoking as a major risk factor for morbidity affects numerous regulatory systems of the human body including DNA methylation. Most of the previous studies with genome-wide methylation data are based on conventional association analysis and earliest threshold-based gene set analysis that lacks sensitivity to be able to reveal all the relevant effects of smoking. The aim of the present study was to investigate the impact of active smoking on DNA methylation at three biological levels: 5'-C-phosphate-G-3' (CpG) sites, genes and functionally related genes (gene sets). Gene set analysis was done with mGSZ, a modern threshold-free method previously developed by us that utilizes all the genes in the experiment and their differential methylation scores. Application of such method in DNA methylation study is novel. Epigenome-wide methylation levels were profiled from Young Finns Study (YFS) participants' whole blood from 2011 follow-up using Illumina Infinium HumanMethylation450 BeadChips. We identified three novel smoking related CpG sites and replicated 57 of the previously identified ones. We found that smoking is associated with hypomethylation in shore (genomic regions 0-2 kilobases from CpG island). We identified smoking related methylation changes in 13 gene sets with false discovery rate (FDR) ≤ 0.05, among which is olfactory receptor activity, the flagship novel finding of the present study. Overall, we extended the current knowledge by identifying: (i) three novel smoking related CpG sites, (ii) similar effects as aging on average methylation in shore, and (iii) a novel finding that olfactory receptor activity pathway responds to tobacco smoke and toxin exposure through epigenetic mechanisms., (© 2020 The Author(s).)

Markers in monocytes, precursors of macrophages, which are related to CAD, are largely unknown. Therefore, we aimed to identify genes in monocytes predictive of a new ischemic event in patients with CAD and/or discriminate between stable CAD and acute coronary syndrome. We included 66 patients with stable CAD, of which 24 developed a new ischemic event, and 19 patients with ACS. Circulating CD14+ monocytes were isolated with magnetic beads. RNA sequencing analysis in monocytes of patients with (n = 13) versus without (n = 11) ischemic event at follow-up and in patients with ACS (n = 12) was validated with qPCR (n = 85). MT-COI, STRN and COX10 predicted new ischemic events in CAD patients (power for separation at 1% error rate of 0.97, 0.90 and 0.77 respectively). Low MT-COI and high STRN were also related to shorter time between blood sampling and event. COX10 and ZNF484 together with MT-COI, STRN and WNK1 separated ACS completely from stable CAD patients. RNA expressions in monocytes of MT-COI, COX10, STRN, WNK1 and ZNF484 were independent of cholesterol lowering and antiplatelet treatment. They were independent of troponin T, a marker of myocardial injury. But, COX10 and ZNF484 in human plaques correlated to plaque markers of M1 macrophage polarization, reflecting vascular injury. Expression of MT-COI, COX10, STRN and WNK1, but not that of ZNF484, PBMCs paired with that in monocytes. The prospective study of relation of MT-COI, COX10, STRN, WNK1 and ZNF484 with unstable CAD is warranted., Competing Interests: We have the following interests: Bernward Klocke is employed by Intrexon Bioinformatics Germany which was hired by KU Leuven to deliver service in modeling. There are no patents, products in development or marketed products to declare. This does not alter our adherence to all the PLOS ONE policies on sharing data and materials.

Cytokines are essential regulatory components of the immune system, and their aberrant levels have been linked to many disease states. Despite increasing evidence that cytokines operate in concert, many of the physiological interactions between cytokines, and the shared genetic architecture that underlies them, remain unknown. Here, we aimed to identify and characterize genetic variants with pleiotropic effects on cytokines. Using three population-based cohorts (n = 9,263), we performed multivariate genome-wide association studies (GWAS) for a correlation network of 11 circulating cytokines, then combined our results in meta-analysis. We identified a total of eight loci significantly associated with the cytokine network, of which two (PDGFRB and ABO) had not been detected previously. In addition, conditional analyses revealed a further four secondary signals at three known cytokine loci. Integration, through the use of Bayesian colocalization analysis, of publicly available GWAS summary statistics with the cytokine network associations revealed shared causal variants between the eight cytokine loci and other traits; in particular, cytokine network variants at the ABO, SERPINE2, and ZFPM2 loci showed pleiotropic effects on the production of immune-related proteins, on metabolic traits such as lipoprotein and lipid levels, on blood-cell-related traits such as platelet count, and on disease traits such as coronary artery disease and type 2 diabetes., (Copyright © 2019 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

We analyzed the associations between whole blood microRNA profiles and the indices of glucose metabolism and impaired fasting glucose and examined whether the discovered microRNAs correlate with the expression of their mRNA targets. MicroRNA and gene expression profiling were performed for the Young Finns Study participants (n = 871). Glucose, insulin, and glycated hemoglobin (HbA1c) levels were measured, the insulin resistance index (HOMA2-IR) was calculated, and the glycemic status (normoglycemic [n = 534]/impaired fasting glucose [IFG] [n = 252]/type 2 diabetes [T2D] [n = 24]) determined. Levels of hsa-miR-144-5p, -122-5p, -148a-3p, -589-5p, and hsa-let-7a-5p associated with glycemic status. hsa-miR-144-5p and -148a-3p associated with glucose levels, while hsa-miR-144-5p, -122-5p, -184, and -339-3p associated with insulin levels and HOMA2-IR, and hsa-miR-148a-3p, -15b-3p, -93-3p, -146b-5p, -221-3p, -18a-3p, -642a-5p, and -181-2-3p associated with HbA1c levels. The targets of hsa-miR-146b-5p that correlated with its levels were enriched in inflammatory pathways, and the targets of hsa-miR-221-3p were enriched in insulin signaling and T2D pathways. These pathways showed indications of co-regulation by HbA1c-associated miRNAs. There were significant differences in the microRNA profiles associated with glucose, insulin, or HOMA-IR compared to those associated with HbA1c. The HbA1c-associated miRNAs also correlated with the expression of target mRNAs in pathways important to the development of T2D.

The effect of mitochondrial DNA (mtDNA) variation on peripheral blood transcriptomics in health and disease is not fully known. Sex-specific mitochondrially controlled gene expression patterns have been shown in Drosophila melanogaster but in humans, evidence is lacking. Functional variation in mtDNA may also have a role in the development of type 2 diabetes and its precursor state, i.e. prediabetes. We examined the associations between mitochondrial single-nucleotide polymorphisms (mtSNPs) and peripheral blood transcriptomics with a focus on sex- and prediabetes-specific effects. The genome-wide blood cell expression data of 19 637 probes, 199 deep-sequenced mtSNPs and nine haplogroups of 955 individuals from a population-based Young Finns Study cohort were used. Significant associations were identified with linear regression and analysis of covariance. The effects of sex and prediabetes on the associations between gene expression and mtSNPs were studied using random-effect meta-analysis. Our analysis identified 53 significant expression probe-mtSNP associations after Bonferroni correction, involving 7 genes and 31 mtSNPs. Eight probe-mtSNP signals remained independent after conditional analysis. In addition, five genes showed differential expression between haplogroups. The meta-analysis did not show any significant differences in linear model effect sizes between males and females but identified the association between the OASL gene and mtSNP C16294T to show prediabetes-specific effects. This study pinpoints new independent mtSNPs associated with peripheral blood transcriptomics and replicates six previously reported associations, providing further evidence of the mitochondrial genetic control of blood cell gene expression. In addition, we present evidence that prediabetes might lead to perturbations in mitochondrial control., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Correlation among traits is a fundamental feature of biological systems that remains difficult to study. To address this problem, we developed a flexible approach that allows us to identify factors associated with inter-individual variation in correlation. We use data from three human cohorts to study the effects of genetic and environmental variation on correlations among mRNA transcripts and among NMR metabolites. We first show that environmental exposures (infection and disease) lead to a systematic loss of correlation, which we define as 'decoherence'. Using longitudinal data, we show that decoherent metabolites are better predictors of whether someone will develop metabolic syndrome than metabolites commonly used as biomarkers of this disease. Finally, we demonstrate that correlation itself is under genetic control by mapping hundreds of 'correlation quantitative trait loci (QTLs)'. Together, this work furthers our understanding of how and why coordinated biological processes break down, and points to a potential role for decoherence in disease., Editorial Note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter)., Competing Interests: AL, MS, AK, TL, ER, MK, IS, NM, OR, MA, PP, NZ, JA No competing interests declared, (© 2019, Lea et al.)

Objective- Palmitoylethanolamide is an endogenous fatty acid mediator that is synthetized from membrane phospholipids by N-acyl phosphatidylethanolamine phospholipase D. Its biological actions are primarily mediated by PPAR-α (peroxisome proliferator-activated receptors α) and the orphan receptor GPR55. Palmitoylethanolamide exerts potent anti-inflammatory actions but its physiological role and promise as a therapeutic agent in chronic arterial inflammation, such as atherosclerosis remain unexplored. Approach and Results- First, the polarization of mouse primary macrophages towards a proinflammatory phenotype was found to reduce N-acyl phosphatidylethanolamine phospholipase D expression and palmitoylethanolamide bioavailability. N-acyl phosphatidylethanolamine phospholipase D expression was progressively downregulated in the aorta of apolipoprotein E deficient (ApoE -/- ) mice during atherogenesis. N-acyl phosphatidylethanolamine phospholipase D mRNA levels were also downregulated in unstable human plaques and they positively associated with smooth muscle cell markers and negatively with macrophage markers. Second, ApoE - /- mice were fed a high-fat diet for 4 or 16 weeks and treated with either vehicle or palmitoylethanolamide (3 mg/kg per day, 4 weeks) to study the effects of palmitoylethanolamide on early established and pre-established atherosclerosis. Palmitoylethanolamide treatment reduced plaque size in early atherosclerosis, whereas in pre-established atherosclerosis, palmitoylethanolamide promoted signs of plaque stability as evidenced by reduced macrophage accumulation and necrotic core size, increased collagen deposition and downregulation of M1-type macrophage markers. Mechanistically, we found that palmitoylethanolamide, by activating GPR55, increases the expression of the phagocytosis receptor MerTK (proto-oncogene tyrosine-protein kinase MER) and enhances macrophage efferocytosis, indicative of proresolving properties. Conclusions- The present study demonstrates that palmitoylethanolamide protects against atherosclerosis by promoting an anti-inflammatory and proresolving phenotype of lesional macrophages, representing a new therapeutic approach to resolve arterial inflammation.

α-melanocyte-stimulating hormone (α-MSH) is processed from pro-opiomelanocortin (POMC) and mediates anti-inflammatory actions in leukocytes. α-MSH also promotes macrophage reverse cholesterol transport by inducing ATP-binding cassette transporters ABCA1 and ABCG1. Here we investigated the regulation of POMC and α-MSH expression in atherosclerosis. First, transcript levels of POMC and its processing enzymes were analyzed in human arterial plaques (n = 68) and non-atherosclerotic controls (n = 24) as well as in whole blood samples from coronary artery disease patients (n = 55) and controls (n = 45) by microarray. POMC expression was increased in femoral plaques compared to control samples as well as in unstable advanced plaques. α-MSH-producing enzyme, carboxypeptidase E, was down-regulated, whereas prolylcarboxypeptidase, an enzyme inactivating α-MSH, was up-regulated in unstable plaques compared to stable plaques, suggesting a possible reduction in intraplaque α-MSH levels. Second, immunohistochemical analyses revealed the presence of α-MSH in atherosclerotic plaques and its localization in macrophages and other cell types. Lastly, supporting the role of α-MSH in reverse cholesterol transport, POMC expression correlated with ABCA1 and ABCG1 in human plaque and whole blood samples. In conclusion, α-MSH is expressed in atherosclerotic plaques and its processing enzymes associate with plaque stability, suggesting that measures to enhance the local bioavailability of α-MSH might protect against atherosclerosis.

Fatty liver (FL) disease is the most common type of chronic liver disease. We hypothesized that liver's response to the process where large droplets of triglyceride fat accumulate in liver cells is reflected also in gene pathway expression in blood. Peripheral blood genome wide gene expression analysis and ultrasonic imaging of liver were performed for 1,650 participants (316 individuals with FL and 1,334 controls) of the Young Finns Study. Gene set enrichment analysis (GSEA) was performed for the expression data. Fourteen gene sets were upregulated (false discovery rate, FDR < 0.05) in subjects with FL. These pathways related to extracellular matrix (ECM) turnover, immune response regulation, prothrombotic state and neural tissues. After adjustment for known risk factors and biomarkers of FL, we found i) integrin A4B1 signaling, ii) leukocyte transendothelial migration, iii) CD40/CD40L and iv) netrin-1 signaling pathways to be upregulated in individuals with FL (nominal p < 0.05). From these all but not ii) remained significantly upregulated when analyzing only subjects without history of heavy alcohol use. In conclusion, FL was associated with blood gene sets of ECM turnover, inflammatory response, immune system activation and prothrombotic state. These may form a systemic link between FL and the development of cardiovascular diseases.

Background: Prediabetes often occurs together with dyslipidaemia, which is paradoxically treated with statins predisposing to type 2 diabetes mellitus. We examined peripheral blood pathway profiles in prediabetic subjects with (PR D ) and without dyslipidaemia (PR 0 ) and compared these to nonprediabetic controls without dyslipidaemia (C 0 )., Methods: The participants were from the Cardiovascular Risk in Young Finns Study, including 1240 subjects aged 34 to 49 years. Genome-wide expression data of peripheral blood and gene set enrichment analysis were used to investigate the differentially expressed genes and enriched pathways between different subtypes of prediabetes., Results: Pathways for cholesterol synthesis, interleukin-12-mediated signalling events, and downstream signalling in naïve CD8+ T-cells were upregulated in the PR 0 group in comparison with controls (C 0 ). The upregulation of these pathways was independent of waist circumference, blood pressure, smoking status, and insulin. Adjustment for CRP left the CD8+ T-cell signalling and interleukin-12-mediated signalling event pathway upregulated. The cholesterol synthesis pathway was also upregulated when all prediabetic subjects (PR 0 and PR D ) were compared with the nonprediabetic control group. No pathways were upregulated or downregulated when the PR D group was compared with the C 0 group. Five genes in the PR 0 group and 1 in the PR D group were significantly differentially expressed in comparison with the C 0 group., Conclusions: Blood cell gene expression profiles differ significantly between prediabetic subjects with and without dyslipidaemia. Whether this classification may be used in detection of prediabetic individuals at a high risk of cardiovascular complications remains to be examined., (Copyright © 2017 John Wiley & Sons, Ltd.)

Ascending thoracic aortic aneurysm (ATAA) is a multifactorial disease with a strong inflammatory component. Surgery is often required to prevent aortic rupture and dissection. We performed gene expression analysis (Illumina HumanHT-12 version 3 Expression BeadChip) for 32 samples from ATAA (26 without/6 with dissection), and 28 left internal thoracic arteries (controls) collected in Tampere Vascular study. We compared expression profiles and conducted pathway analysis using Ingenuity Pathway Analysis (IPA) to reveal differences between ATAA and a healthy artery wall. Almost 5000 genes were differentially expressed in ATAA samples compared to controls. The most downregulated gene was homeobox (HOX) A5 (fold change, FC = -25.3) and upregulated cadherin-2 (FC = 12.6). Several other HOX genes were also found downregulated (FCs between -25.3 and -1.5, FDR < 0.05). 43, mostly inflammatory, canonical pathways in ATAA were found to be significantly (p < 0.05, FDR < 0.05) differentially expressed. The results remained essentially the same when the 6 dissected ATAA samples were excluded from the analysis. We show for the first time on genome level that ATAA is an inflammatory process, revealing a more detailed molecular pathway level pathogenesis. We propose HOX genes as potentially important players in maintaining aortic integrity, altered expression of which might be important in the pathobiology of ATAA.

Background: Immunometabolism plays a central role in many cardiometabolic diseases. However, a robust map of immune-related gene networks in circulating human cells, their interactions with metabolites, and their genetic control is still lacking. Here, we integrate blood transcriptomic, metabolomic, and genomic profiles from two population-based cohorts (total N = 2168), including a subset of individuals with matched multi-omic data at 7-year follow-up., Results: We identify topologically replicable gene networks enriched for diverse immune functions including cytotoxicity, viral response, B cell, platelet, neutrophil, and mast cell/basophil activity. These immune gene modules show complex patterns of association with 158 circulating metabolites, including lipoprotein subclasses, lipids, fatty acids, amino acids, small molecules, and CRP. Genome-wide scans for module expression quantitative trait loci (mQTLs) reveal five modules with mQTLs that have both cis and trans effects. The strongest mQTL is in ARHGEF3 (rs1354034) and affects a module enriched for platelet function, independent of platelet counts. Modules of mast cell/basophil and neutrophil function show temporally stable metabolite associations over 7-year follow-up, providing evidence that these modules and their constituent gene products may play central roles in metabolic inflammation. Furthermore, the strongest mQTL in ARHGEF3 also displays clear temporal stability, supporting widespread trans effects at this locus., Conclusions: This study provides a detailed map of natural variation at the blood immunometabolic interface and its genetic basis, and may facilitate subsequent studies to explain inter-individual variation in cardiometabolic disease.

Background: The melanocortin 1 receptor (MC1-R) is expressed by monocytes and macrophages, where it exerts anti-inflammatory actions on stimulation with its natural ligand α-melanocyte-stimulating hormone. The present study was designed to investigate the specific role of MC1-R in the context of atherosclerosis and possible regulatory pathways of MC1-R beyond anti-inflammation., Methods: Human and mouse atherosclerotic samples and primary mouse macrophages were used to study the regulatory functions of MC1-R. The impact of pharmacological MC1-R activation on atherosclerosis was assessed in apolipoprotein E-deficient mice., Results: Characterization of human and mouse atherosclerotic plaques revealed that MC1-R expression localizes in lesional macrophages and is significantly associated with the ATP-binding cassette transporters ABCA1 and ABCG1, which are responsible for initiating reverse cholesterol transport. Using bone marrow-derived macrophages, we observed that α-melanocyte-stimulating hormone and selective MC1-R agonists similarly promoted cholesterol efflux, which is a counterregulatory mechanism against foam cell formation. Mechanistically, MC1-R activation upregulated the levels of ABCA1 and ABCG1. These effects were accompanied by a reduction in cell surface CD36 expression and in cholesterol uptake, further protecting macrophages from excessive lipid accumulation. Conversely, macrophages deficient in functional MC1-R displayed a phenotype with impaired efflux and enhanced uptake of cholesterol. Pharmacological targeting of MC1-R in atherosclerotic apolipoprotein E-deficient mice reduced plasma cholesterol levels and aortic CD36 expression and increased plaque ABCG1 expression and signs of plaque stability., Conclusions: Our findings identify a novel role for MC1-R in macrophage cholesterol transport. Activation of MC1-R confers protection against macrophage foam cell formation through a dual mechanism: It prevents cholesterol uptake while concomitantly promoting ABCA1- and ABCG1-mediated reverse cholesterol transport., (© 2017 American Heart Association, Inc.)

Background: Cardiac hypertrophy increases the risk of developing heart failure and cardiovascular death. The neutrophil inflammatory protein, lipocalin-2 (LCN2/NGAL), is elevated in certain forms of cardiac hypertrophy and acute heart failure. However, a specific role for LCN2 in predisposition and etiology of hypertrophy and the relevant genetic determinants are unclear. Here, we defined the role of LCN2 in concentric cardiac hypertrophy in terms of pathophysiology, inflammatory expression networks, and genomic determinants., Methods and Results: We used 3 experimental models: a polygenic model of cardiac hypertrophy and heart failure, a model of intrauterine growth restriction and Lcn2 -knockout mouse; cultured cardiomyocytes; and 2 human cohorts: 114 type 2 diabetes mellitus patients and 2064 healthy subjects of the YFS (Young Finns Study). In hypertrophic heart rats, cardiac and circulating Lcn2 was significantly overexpressed before, during, and after development of cardiac hypertrophy and heart failure. Lcn2 expression was increased in hypertrophic hearts in a model of intrauterine growth restriction, whereas Lcn2 -knockout mice had smaller hearts. In cultured cardiomyocytes, Lcn2 activated molecular hypertrophic pathways and increased cell size, but reduced proliferation and cell numbers. Increased LCN2 was associated with cardiac hypertrophy and diastolic dysfunction in diabetes mellitus. In the YFS, LCN2 expression was associated with body mass index and cardiac mass and with levels of inflammatory markers. The single-nucleotide polymorphism, rs13297295, located near LCN2 defined a significant cis -eQTL for LCN2 expression., Conclusions: Direct effects of LCN2 on cardiomyocyte size and number and the consistent associations in experimental and human analyses reveal a central role for LCN2 in the ontogeny of cardiac hypertrophy and heart failure., (© 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.)

Cardiovascular diseases due to atherosclerosis are the leading cause of death globally. We aimed to investigate the potentially altered gene and pathway expression in advanced peripheral atherosclerotic plaques in comparison to healthy control arteries. Gene expression analysis was performed (Illumina HumanHT-12 version 3 Expression BeadChip) for 68 advanced atherosclerotic plaques (15 aortic, 29 carotid and 24 femoral plaques) and 28 controls (left internal thoracic artery (LITA)) from Tampere Vascular Study. Dysregulation of individual genes was compared to healthy controls and between plaques from different arterial beds and Ingenuity pathway analysis was conducted on genes with a fold change (FC) > ±1.5 and false discovery rate (FDR) < 0.05. 787 genes were significantly differentially expressed in atherosclerotic plaques. The most up-regulated genes were osteopontin and multiple MMPs, and the most down-regulated were cell death-inducing DFFA-like effector C and A (CIDEC, CIDEA) and apolipoprotein D (FC > 20). 156 pathways were differentially expressed in atherosclerotic plaques, mostly inflammation-related, especially related with leukocyte trafficking and signaling. In artery specific plaque analysis 50.4% of canonical pathways and 41.2% GO terms differentially expressed were in common for all three arterial beds. Our results confirm the inflammatory nature of advanced atherosclerosis and show novel pathway differences between different arterial beds., Competing Interests: The authors declare no competing financial interests.

Circulating cytokines and growth factors are regulators of inflammation and have been implicated in autoimmune and metabolic diseases. In this genome-wide association study (GWAS) of up to 8,293 Finns we identified 27 genome-widely significant loci (p < 1.2 × 10 -9 ) for one or more cytokines. Fifteen of the associated variants had expression quantitative trait loci in whole blood. We provide genetic instruments to clarify the causal roles of cytokine signaling and upstream inflammation in immune-related and other chronic diseases. We further link inflammatory markers with variants previously associated with autoimmune diseases such as Crohn disease, multiple sclerosis, and ulcerative colitis and hereby elucidate the molecular mechanisms underpinning these diseases and suggest potential drug targets., (Copyright © 2017 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

MicroRNAs are involved in disease development and may be utilized as biomarkers. We investigated the association of blood miRNA levels and a) fatty liver (FL), b) lipoprotein and lipid pathways involved in liver lipid accumulation and c) levels of predicted mRNA targets in general population based cohort. Blood microRNA profiling (TaqMan OpenArray), genome-wide gene expression arrays and nuclear magnetic resonance metabolomics were performed for Young Finns Study participants aged 34-49 years (n = 871). Liver fat status was assessed ultrasonographically. Levels of hsa-miR-122-5p and -885-5p were up-regulated in individuals with FL (fold change (FC) = 1.55, p = 1.36 * 10 -14 and FC = 1.25, p = 4.86 * 10 -4 , respectively). In regression model adjusted with age, sex and BMI, hsa-miR-122-5p and -885-5p predicted FL (OR = 2.07, p = 1.29 * 10 -8 and OR = 1.41, p = 0.002, respectively). Together hsa-miR-122-5p and -885-5p slightly improved the detection of FL beyond established risk factors. These miRNAs may be associated with FL formation through the regulation of lipoprotein metabolism as hsa-miR-122-5p levels associated with small VLDL, IDL, and large LDL lipoprotein subclass components, while hsa-miR-885-5p levels associated inversely with XL HDL cholesterol levels. Hsa-miR-885-5p levels correlated inversely with oxysterol-binding protein 2 (OSBPL2) expression (r = -0.143, p = 1.00 * 10 -4 ) and suppressing the expression of this lipid receptor and sterol transporter could link hsa-miR-885-5p with HDL cholesterol levels., Competing Interests: MAK, PS, and AJK are shareholders in Brainshake Ltd., a company offering NMR-based metabolite profiling.

Background and Aims: Focal adhesions (FA) play an important role in the tissue remodeling and in the maintenance of tissue integrity and homeostasis. Talin and vinculin proteins are among the major constituents of FAs contributing to cellular well-being and intercellular communication., Methods: Microarray analysis (MA) and qRT-PCR low-density array were implemented to analyze talin-1, talin-2, meta-vinculin and vinculin gene expression in circulating blood and arterial plaque., Results: All analyzed genes were significantly and consistently downregulated in plaques (carotid, abdominal aortic and femoral regions) compared to left internal thoracic artery (LITA) control. The use of LITA samples as controls for arterial plaque samples was validated using immunohistochemistry by comparing LITA samples with healthy arterial samples from a cadaver. Even though the differences in expression levels between stable and unstable plaques were not statistically significant, we observed further negative tendency in the expression in unstable atherosclerotic plaques. The confocal tissue imaging revealed gradient of talin-1 expression in plaque with reduction close to the vessel lumen. Similar gradient was observed for talin-2 expression in LITA controls but was not detected in plaques. This suggests that impaired tissue mechanostability affects the tissue remodeling and healing capabilities leading to development of unstable plaques., Conclusions: The central role of talin and vinculin in cell adhesions suggests that the disintegration of the tissue in atherosclerosis could be partially driven by downregulation of these genes, leading to loosening of cell-ECM interactions and remodeling of the tissue., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Red blood cell (RBC) traits are important heritable clinical biomarkers and modifiers of disease severity. To identify coding genetic variants associated with these traits, we conducted meta-analyses of seven RBC phenotypes in 130,273 multi-ethnic individuals from studies genotyped on an exome array. After conditional analyses and replication in 27,480 independent individuals, we identified 16 new RBC variants. We found low-frequency missense variants in MAP1A (rs55707100, minor allele frequency [MAF] = 3.3%, p = 2 × 10(-10) for hemoglobin [HGB]) and HNF4A (rs1800961, MAF = 2.4%, p < 3 × 10(-8) for hematocrit [HCT] and HGB). In African Americans, we identified a nonsense variant in CD36 associated with higher RBC distribution width (rs3211938, MAF = 8.7%, p = 7 × 10(-11)) and showed that it is associated with lower CD36 expression and strong allelic imbalance in ex vivo differentiated human erythroblasts. We also identified a rare missense variant in ALAS2 (rs201062903, MAF = 0.2%) associated with lower mean corpuscular volume and mean corpuscular hemoglobin (p < 8 × 10(-9)). Mendelian mutations in ALAS2 are a cause of sideroblastic anemia and erythropoietic protoporphyria. Gene-based testing highlighted three rare missense variants in PKLR, a gene mutated in Mendelian non-spherocytic hemolytic anemia, associated with HGB and HCT (SKAT p < 8 × 10(-7)). These rare, low-frequency, and common RBC variants showed pleiotropy, being also associated with platelet, white blood cell, and lipid traits. Our association results and functional annotation suggest the involvement of new genes in human erythropoiesis. We also confirm that rare and low-frequency variants play a role in the architecture of complex human traits, although their phenotypic effect is generally smaller than originally anticipated., (Copyright © 2016 American Society of Human Genetics. All rights reserved.)

Platelet production, maintenance, and clearance are tightly controlled processes indicative of platelets' important roles in hemostasis and thrombosis. Platelets are common targets for primary and secondary prevention of several conditions. They are monitored clinically by complete blood counts, specifically with measurements of platelet count (PLT) and mean platelet volume (MPV). Identifying genetic effects on PLT and MPV can provide mechanistic insights into platelet biology and their role in disease. Therefore, we formed the Blood Cell Consortium (BCX) to perform a large-scale meta-analysis of Exomechip association results for PLT and MPV in 157,293 and 57,617 individuals, respectively. Using the low-frequency/rare coding variant-enriched Exomechip genotyping array, we sought to identify genetic variants associated with PLT and MPV. In addition to confirming 47 known PLT and 20 known MPV associations, we identified 32 PLT and 18 MPV associations not previously observed in the literature across the allele frequency spectrum, including rare large effect (FCER1A), low-frequency (IQGAP2, MAP1A, LY75), and common (ZMIZ2, SMG6, PEAR1, ARFGAP3/PACSIN2) variants. Several variants associated with PLT/MPV (PEAR1, MRVI1, PTGES3) were also associated with platelet reactivity. In concurrent BCX analyses, there was overlap of platelet-associated variants with red (MAP1A, TMPRSS6, ZMIZ2) and white (PEAR1, ZMIZ2, LY75) blood cell traits, suggesting common regulatory pathways with shared genetic architecture among these hematopoietic lineages. Our large-scale Exomechip analyses identified previously undocumented associations with platelet traits and further indicate that several complex quantitative hematological, lipid, and cardiovascular traits share genetic factors., (Published by Elsevier Inc.)

Sleep loss and insufficient sleep are risk factors for cardiometabolic diseases, but data on how insufficient sleep contributes to these diseases are scarce. These questions were addressed using two approaches: an experimental, partial sleep restriction study (14 cases and 7 control subjects) with objective verification of sleep amount, and two independent epidemiological cohorts (altogether 2739 individuals) with questions of sleep insufficiency. In both approaches, blood transcriptome and serum metabolome were analysed. Sleep loss decreased the expression of genes encoding cholesterol transporters and increased expression in pathways involved in inflammatory responses in both paradigms. Metabolomic analyses revealed lower circulating large HDL in the population cohorts among subjects reporting insufficient sleep, while circulating LDL decreased in the experimental sleep restriction study. These findings suggest that prolonged sleep deprivation modifies inflammatory and cholesterol pathways at the level of gene expression and serum lipoproteins, inducing changes toward potentially higher risk for cardiometabolic diseases.

Many genetic variants influence complex traits by modulating gene expression, thus altering the abundance of one or multiple proteins. Here we introduce a powerful strategy that integrates gene expression measurements with summary association statistics from large-scale genome-wide association studies (GWAS) to identify genes whose cis-regulated expression is associated with complex traits. We leverage expression imputation from genetic data to perform a transcriptome-wide association study (TWAS) to identify significant expression-trait associations. We applied our approaches to expression data from blood and adipose tissue measured in ∼ 3,000 individuals overall. We imputed gene expression into GWAS data from over 900,000 phenotype measurements to identify 69 new genes significantly associated with obesity-related traits (BMI, lipids and height). Many of these genes are associated with relevant phenotypes in the Hybrid Mouse Diversity Panel. Our results showcase the power of integrating genotype, gene expression and phenotype to gain insights into the genetic basis of complex traits.

We used genome wide expression (GWE) data of circulating blood cells and pathway analysis to investigate the inflammatory and other molecular pathways that may be associated with long-standing depressive symptoms. Participants were 607 women and 316 men (mean age 42 years) from the Young Finns Study who participated in three consecutive study phases in 2001, 2007 and 2012. Using Gene-set enrichment analyses (GSEA) we focused our analyses to pathways (available in MSigDB database) that are likely to affect immunological and inflammatory processes. GSEA were performed for blood cell GWE data in 2012. Depressive symptoms were assessed using a modified 21-item Beck Depression Inventory in each of the three study phases. Participants who scored in the top quartile of depressive symptoms in each of the three measurement points (n = 191) differed from other participants (n = 732) in several gene-set pathways related to inflammatory processes or immune-inflammatory signaling including interleukin (IL-1) pathway, and pathways related to various immuno-inflammatory processes, such as toll-like, the NEF protein, the nuclear factor kB, the kinase AKT and the mature B cell antigen receptor pathway (false discovery rates, FDRs<0.12). The results provide novel genome wide molecular evidence that support the association between chronic depressive symptoms and altered immune-inflammatory regulation., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The biomarker glycoprotein acetylation (GlycA) has been shown to predict risk of cardiovascular disease and all-cause mortality. Here, we characterize biological processes associated with GlycA by leveraging population-based omics data and health records from >10,000 individuals. Our analyses show that GlycA levels are chronic within individuals for up to a decade. In apparently healthy individuals, elevated GlycA corresponded to elevation of myriad inflammatory cytokines, as well as a gene coexpression network indicative of increased neutrophil activity, suggesting that individuals with high GlycA may be in a state of chronic inflammatory response. Accordingly, analysis of infection-related hospitalization and death records showed that increased GlycA increased long-term risk of severe non-localized and respiratory infections, particularly septicaemia and pneumonia. In total, our work demonstrates that GlycA is a biomarker for chronic inflammation, neutrophil activity, and risk of future severe infection. It also illustrates the utility of leveraging multi-layered omics data and health records to elucidate the molecular and cellular processes associated with biomarkers., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: Kindlins (FERMT) are cytoplasmic proteins required for integrin (ITG) activation, leukocyte transmigration, platelet aggregation and thrombosis. Characterization of kindlins and their association with atherosclerotic plaques in human(s) is lacking., Methods and Results: Exploratory microarray (MA) was first performed followed by selective quantitative validation of robustly expressed genes with qRT-PCR low-density array (LDA). In LDA, ITGA1 (1.30-fold, p = 0.041) and ITGB3 (1.37-fold, p = 0.036) were upregulated in whole blood samples of patients with coronary artery disease (CAD) compared to healthy controls. In arterial plaques, both robustly expressed transcript variants of FERMT3 (MA: 5.90- and 3.4-fold; LDA: 3.99-fold, p < 0.0001 for all) and ITGB2 (MA: 4.81- and 4.92-fold; LDA: 5.29-fold, p < 0.0001 for all) were upregulated while FERMT2 was downregulated (MA: -1.61-fold; LDA: -2.88-fold, p < 0.0001 for both). The other integrins (ITGA1, ITGAV, ITGB3, ITGB5) were downregulated. All these results were replicated in at least one arterial bed. The latter FERMT3 transcript variant associated with unstable plaques (p = 0.0004). FERMT3 correlated with M2 macrophage markers and in hierarchical cluster analysis clustered with inflammatory and macrophage markers, while FERMT2 correlated with SMC-rich plaque markers and clustered with SMC markers. In confocal immunofluorescence analysis, FERMT3 protein colocalized with abundant CD68-positive cells of monocytic origin in the atherosclerotic plaques, while co-localization of FERMT3 with HHF35 indicative of smooth muscle cells was low., Conclusions: Kindlin-3 (FERMT3) is upregulated in atherosclerotic, especially unstable plaques, mainly in cells of monocytic origin and of M2 type. Simultaneous upregulation of ITGB2 suggests a synergistic effect on leukocyte adherence and transmigration into the vessel wall., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Proprotein convertase subtilisin/kexin (PCSK) enzymes cleave and convert their immature substrates into biologically active forms. Polymorphisms in the PCSK genes have been reported to associate with human diseases and phenotypes, including hypercholesterolemia and blood pressure (BP), and targeting PCSKs is considered a promising future form of drug therapy. PCSK processing is readily induced upon upregulation of the enzyme, but the genetic factors contributing to PCSK expression have not been thoroughly characterized. To gain a comprehensive understanding of the genetic regulation of PCSK expression, we performed, for the first time, a genome-wide expression quantitative trait loci (eQTL) analysis using mRNA expression in >1400 human peripheral blood samples from the Cardiovascular Risk in Young Finns Study and ca. ten million single-nucleotide polymorphisms (SNPs). The expression data showed clear expression for FURIN, PCSK5, PCSK7 and MBTPS1 (membrane-bound transcription factor peptidase, site 1) mRNAs in virtually all tested samples. A discovery analysis demonstrated a genome-wide significant (p < 5 × 10(-8)) association with the selected PCSK probes for 1024 variants, which were located at ten independent loci. Of these loci, 5/10 could be confirmed to regulate PCSK expression in two additional and independent sample sets. Finally, a phenotypic analysis demonstrated that a novel cis-eQTL SNP rs4702 for FURIN is strongly associated with both diastolic (p = 0.012) and systolic (p = 0.035) BP levels, as well as peripheral vascular resistance (p = 0.003). These findings indicate that the expression of the PCSK enzymes is regulated by genetic factors, which have biological roles in health and disease.

Background: Atherosclerosis is an inflammatory disease with possible contributions from bacterial antigens. We aimed to investigate the role of oral bacteria as inducers of inflammatory cascades in smooth muscle cells from carotid endarterectomy patients (AthSMCs) and healthy controls (HSMCs)., Findings: Inactivated Streptococcus mitis, S. sanguinis, S. gorgonii, Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis were used to stimulate inflammation in HSMCs and AthSMCs. Tumor necrosis factor-α (TNFα) was used as a positive control in all stimulations. Interleukin-6 (IL-6) levels were determined from cell culture supernatants and microRNA expression profiles from cells after 24 h of bacterial stimulation. Genome wide expression (GWE) analyses were performed after 5 h stimulation. All studied bacteria induced pro inflammatory IL-6 production in both SMCs. The most powerful inducer of IL-6 was A. actinomycetemcomitans (p < 0.001). Of the 84 studied miRNAs, expression of 9 miRNAs differed significantly (p ≤ 0.001) between HSMCs and AthSMCs stimulated with inactivated bacteria or TNFα. The data was divided into two groups: high IL-6 producers (A. actinomytectemcomititans and TNFα) and low IL-6 producers (streptococcal strains and P. gingivalis). The expression of 4 miRNAs (miR-181-5p, -186-5p, -28-5p and -155-5p) differed statistically significantly (p < 0.001) between healthy HSMCs and AthSMCs in the low IL-6 producer group. According to multidimensional scaling, two gene expression clusters were seen: one in HSMCs and one AthSMCs., Conclusions: Our results suggest that inactivated oral bacteria induce inflammation that is differently regulated in healthy and atherosclerotic SMCs.