Exploring the use of experimental small area estimates to examine the relationship between individual-level and area-level community belonging and self-rated health.

Background: Small area estimation refers to statistical modelling procedures that leverage information or "borrow strength" from other sources or variables. This is done to enhance the reliability of estimates of characteristics or outcomes for areas that do not contain sufficient sample sizes to provide disaggregated estimates of adequate precision and reliability. There is growing interest in secondary research applications for small area estimates (SAEs). However, it is crucial to assess the analytic value of these estimates when used as proxies for individual-level characteristics or as distinct measures that offer insights at the area level. This study assessed novel area-level community belonging measures derived using small area estimation and examined associations with individual-level measures of community belonging and self-rated health. Data and methods: SAEs of community belonging within census tracts produced from the 2016-2019 cycles of the Canadian Community Health Survey (CCHS) were merged with respondent data from the 2020 CCHS. Multinomial logistic regression models were run between area-level SAEs, individual-level sense of community belonging, and self-rated health on the study sample of people aged 18 years and older. Results: Area-level community belonging was associated with individual-level community belonging, even after adjusting for individual-level sociodemographic characteristics, despite limited agreement between individual- and area-level measures. Living in a neighbourhood with low community belonging was associated with higher odds of reporting being in fair or poor health, versus being in very good or excellent health (odds ratio: 1.53; 95% confidence interval: 1.22, 1.91), even after adjusting for other factors such as individual-level sense of community belonging, which was also associated with self-rated health. Interpretation: Area-level and individual-level sense of community belonging were independently associated with self-rated health. The novel SAEs of community belonging can be used as distinct measures of neighbourhood-level community belonging and should be understood as complementary to, rather than proxies for, individual-level measures of community belonging.

A national cohort study of community belonging and its influence on premature mortality.

BACKGROUND: Community belonging, an important constituent of subjective well-being, is an important target for improving population health. Ageing involves transitioning across different social conditions thus, community belonging on health may vary across the life course. Using a nationally representative cohort, this study estimates the life stage-specific impact of community belonging on premature mortality. METHODS: Six cycles of the Canadian Community Health Survey (2000-2012) were combined and linked to the Canadian Vital Statistics Database (2000-2017). Respondents were followed for up to 5 years. Multivariable-adjusted modified Poisson regression models were used to estimate the relative risk of premature mortality for three life stages: early adulthood (18-35 years), middle adulthood (36-55 years) and late adulthood (56-70 years). RESULTS: The final analytical sample included 477 100 respondents. Most reported a 'somewhat strong' sense of belonging (45.9%). Compared with their 'somewhat strong' counterparts, young adults reporting a 'somewhat weak' sense of belonging exhibited an increased relative risk (RR) of 1.76 (95% CI 1.27 to 2.43) for premature mortality, whereas middle-aged adults reporting the same exhibited a decreased RR of 0.82 (95% CI 0.69, 0.98). Among older adults, groups reporting a 'very strong' (RR 1.10, 95% CI 1.01, 1.21) or a 'very weak' sense (RR 1.14, 95% CI 1.01, 1.28) of belonging exhibited higher RRs for premature mortality. CONCLUSION: The results demonstrate how community belonging relates to premature mortality differs across age groups underscoring the importance of considering life stage-specific perspectives when researching and developing approaches to strengthen belonging.

Overcrowded housing during adolescence and future risk of premature mortality: a 28-year follow-up of 556,191 adolescents from Switzerland.

Background: Few large-scale studies have examined the health impacts of overcrowded housing in European countries. The aim of this study was to assess whether household crowding during adolescence increases the risk of all-cause and cause-specific mortality in Switzerland. Methods: Study participants were 556,191 adolescents aged 10-19 years at the 1990 census from the Swiss National Cohort. Household crowding at baseline was measured as the ratio between the number of persons living in the household and the number of available rooms, categorized as none (ratio ≤ 1), moderate (1 < ratio ≤ 1.5), and severe (ratio > 1.5). Participants were linked to administrative mortality records through 2018 and followed for premature mortality from all causes, cardiometabolic disease and self-harm or substance use. Cumulative risk differences between ages 10 and 45 were standardized by parental occupation, residential area, permit status and household type. Findings: Of the sample, 19% lived in moderately and 5% lived in severely crowded households. During an average follow-up of 23 years, 9766 participants died. Cumulative risk of death from all causes was 2359 (95% compatibility intervals: 2296-2415) per 100,000 persons when living in non-crowded households. Living in moderately crowded households led to 99 additional deaths (-63 to 256) per 100,000 persons and living in severely crowded households 258 additional deaths (-37 to 607) per 100,000 persons. The effect of crowding on mortality from cardiometabolic diseases, self-harm or substance use was negligible. Interpretation: Excess risk of premature mortality in adolescents living in overcrowded households appears to be small or negligible in Switzerland. Funding: University of Fribourg Scholarship Programme for foreign post-doctoral researchers.

An international comparative study of active living environments and hospitalization for Wales and Canada.

Rationale: Previous studies indicate active living environments (ALEs) are associated with higher physical activity levels across different geographic contexts, and could lead to reductions in hospital burden. Both Wales UK and Canada have advanced data infrastructure that allows record linkage between survey data and administrative health information. Objective: To assess the relationship between ALEs and hospitalization in Wales and Canada. Methods: We performed a population-based comparison using individual-level survey data from the Welsh Health Survey (N = 9968) linked to the Patient Episode Database for Wales, and the Canadian Community Health Survey (N = 40,335) linked to the Discharge Abstract Database. Using equivalent protocols and open-source data for street networks, destinations, and residential density, we derived 5-class measures of the ALE for Wales and Canada (classed 1 through 5, considered least favourable to most favourable for active living, respectively). We evaluated relationships of ALEs to health, behaviours and hospitalization using multivariate regression (reference group was the lowest ALE class 1, considered least favourable for active living). Results: For Canada, those living in the highest ALE class 5 had lower odds of all-cause hospitalization (OR 0.66, 95% CI 0.54 to 0.81; as compared to the lowest ALE class 1). In contrast, those living in the highest ALE class 5 in Wales had higher odds of all-cause hospitalization (OR 1.37, 95% CI 1.04 to 1.80). The relationship between ALEs and cardiometabolic hospitalization was inconclusive for Canada (OR 0.75, 95% CI 0.50 to 1.12), but we observed higher odds of cardiometabolic hospitalization for respondents living in higher ALE classes for Wales (OR 1.46, 95% CI 1.10 to 1.78; comparing ALE class 4 to ALE class 1). Conclusion: Canadian respondents living in high ALE neighbourhoods that are understood to be favourable for active living had lower odds of all-cause hospitalization, whereas Welsh respondents living in high ALEs that were deemed favourable for active living exhibited higher odds of all-cause hospitalization. Environments which promote physical activity in one geographic context may not do so in another. There remains a need to identify relevant context-specific factors that encourage active living.

Does living near hospital obscure the association between active living environments and hospitalization?

Hospitals tend to be among the destinations that make densely populated, well-connected neighbourhoods more conducive to active living. In this study, we determined whether living near a hospital distorts the association between living in favourable ALEs and hospitalization for physical inactivity-related cardiometabolic diseases. We used a record linkage of 442,345 respondents of the Canadian Community Health Survey and their hospitalization records for cardiometabolic disease. We then assessed respondents' neighbourhoods using the Canadian Active Living Environments measure (Can-ALE), a measure based on ≥3-way intersection density, residential density, and points of interest. We then calculated the distance in kilometers between the centroids of respondents' assigned dissemination areas and the nearest user-contributed location for hospitals from OpenStreetMap. We monitored changes in estimates for the association between ALEs and odds of cardiometabolic disease hospitalization using a series of logistic regressions with indicator variables for distances to hospital of 500 meters to 10 kilometers. We found that living between 500 meters and six kilometers of a hospital and was associated with modestly higher odds of cardiometabolic hospitalization (OR 1.10, 95% CI 1.02 to 1.18 for 500 meters; OR 1.05, 95% CI 1.01 to 1.09 for six kilometers). Living in more favourable ALEs was associated with lower odds of hospitalization (OR 0.79, 95% CI 0.68 to 0.91; comparing the most favourable to least favourable ALEs). Effect estimates between more favourable ALEs and lower odds of hospitalization were marginally strengthened when living within 2-6 kilometers to a hospital was accounted for. This study demonstrates the importance of disentangling interrelated geographic factors and underlines the potential for built environments to elicit reductions in health care.

Social Cohesion and Community Resilience During COVID-19 and Pandemics: A Rapid Scoping Review to Inform the United Nations Research Roadmap for COVID-19 Recovery.

Shock events uncover deficits in social cohesion and exacerbate existing social inequalities at the household, community, local, regional, and national levels. National and regional government recovery planning requires careful stakeholder engagement that centers on marginalized people, particularly women and marginalized community leaders. The aim of this rapid scoping review was to inform the United Nations Research Roadmap for the COVID-19 Recovery, based on Pillar 5 of the United Nations Framework for the Immediate Socioeconomic Response to COVID-19: Social Cohesion and Community Resilience. We present a summary of key concepts across the literature that helped situate this review. The results include a description of the state of the science and a review of themes identified as being crucial to sustainable and equitable recovery planning by the United Nations. The role of social cohesion during a disaster, particularly its importance for upstream planning and relationship building before a disaster occurs, is not well understood and is a promising area of future research. Understanding the applicability of social cohesion measurement methodologies and outcomes across different communities and geographies, as well as the development of new and relevant instruments and techniques, is urgently needed in the context of the global COVID-19 pandemic.

Active living environments, physical activity and premature cardiometabolic mortality in Canada: a nationwide cohort study.

OBJECTIVE: To evaluate sex-specific and age-specific associations of active living environments (ALEs) with premature cardiometabolic mortality. DESIGN: Population-based retrospective cohort study. SETTING: Residential neighbourhoods (1000-metre circular buffers from the centroids of dissemination areas) across Canada for which the Canadian ALE Measure was derived, based on intersection density, points of interest and dwelling density. PARTICIPANTS: 249 420 survey respondents from an individual-level record linkage between the Canadian Community Health Survey (2000-2010) and the Canadian Mortality Database until 2011, comprised of older women (65-85 years), older men (65-81 years), middle-aged women (45-64 years) and middle-aged men (45-64 years). PRIMARY OUTCOME MEASURES: Premature cardiometabolic mortality and average daily energy expenditure attributable to walking. Multivariable proportional hazards regression models were adjusted for age, educational attainment, dissemination area-level median income, smoking status, obesity, the presence of chronic conditions, season of survey response and survey cycle. RESULTS: Survey respondents contributed a total of 1 451 913 person-years. Greater walking was observed in more favourable ALEs. Walking was associated with lower cardiometabolic death in all groups except for middle-aged men. Favourable ALEs conferred a 22% reduction in death from cardiometabolic causes (HR 0.78, 95% CI 0.63 to 0.97) for older women. CONCLUSIONS: On average, people walk more in favourable ALEs, regardless of sex and age. With the exception of middle-aged men, walking is associated with lower premature cardiometabolic death. Older women living in neighbourhoods that favour active living live longer.

The early care environment and DNA methylome variation in childhood.

Prenatal adversity shapes child neurodevelopment and risk for later mental health problems. The quality of the early care environment can buffer some of the negative effects of prenatal adversity on child development. Retrospective studies, in adult samples, highlight epigenetic modifications as sentinel markers of the quality of the early care environment; however, comparable data from pediatric cohorts are lacking. Participants were drawn from the Maternal Adversity Vulnerability and Neurodevelopment (MAVAN) study, a longitudinal cohort with measures of infant attachment, infant development, and child mental health. Children provided buccal epithelial samples (mean age = 6.99, SD = 1.33 years, n = 226), which were used for analyses of genome-wide DNA methylation and genetic variation. We used a series of linear models to describe the association between infant attachment and (a) measures of child outcome and (b) DNA methylation across the genome. Paired genetic data was used to determine the genetic contribution to DNA methylation at attachment-associated sites. Infant attachment style was associated with infant cognitive development (Mental Development Index) and behavior (Behavior Rating Scale) assessed with the Bayley Scales of Infant Development at 36 months. Infant attachment style moderated the effects of prenatal adversity on Behavior Rating Scale scores at 36 months. Infant attachment was also significantly associated with a principal component that accounted for 11.9% of the variation in genome-wide DNA methylation. These effects were most apparent when comparing children with a secure versus a disorganized attachment style and most pronounced in females. The availability of paired genetic data revealed that DNA methylation at approximately half of all infant attachment-associated sites was best explained by considering both infant attachment and child genetic variation. This study provides further evidence that infant attachment can buffer some of the negative effects of early adversity on measures of infant behavior. We also highlight the interplay between infant attachment and child genotype in shaping variation in DNA methylation. Such findings provide preliminary evidence for a molecular signature of infant attachment and may help inform attachment-focused early intervention programs.

Worldwide patterns of human epigenetic variation.

DNA methylation is an epigenetic modification, influenced by both genetic and environmental variation, that plays a key role in transcriptional regulation and many organismal phenotypes. Although patterns of DNA methylation have been shown to differ between human populations, it remains to be determined how epigenetic diversity relates to the patterns of genetic and gene expression variation at a global scale. Here we measured DNA methylation at 485,000 CpG sites in five diverse human populations, and analysed these data together with genome-wide genotype and gene expression data. We found that population-specific DNA methylation mirrors genetic variation, and has greater local genetic control than mRNA levels. We estimated the rate of epigenetic divergence between populations, which indicates far greater evolutionary stability of DNA methylation in humans than has been observed in plants. This study provides a deeper understanding of worldwide patterns of human epigenetic diversity, as well as initial estimates of the rate of epigenetic divergence in recent human evolution.

Epigenetic correlates of neonatal contact in humans.

Animal models of early postnatal mother-infant interactions have highlighted the importance of tactile contact for biobehavioral outcomes via the modification of DNA methylation (DNAm). The role of normative variation in contact in early human development has yet to be explored. In an effort to translate the animal work on tactile contact to humans, we applied a naturalistic daily diary strategy to assess the link between maternal contact with infants and epigenetic signatures in children 4-5 years later, with respect to multiple levels of child-level factors, including genetic variation and infant distress. We first investigated DNAm at four candidate genes: the glucocorticoid receptor gene, nuclear receptor subfamily 3, group C, member 1 (NR3C1), µ-opioid receptor M1 (OPRM1) and oxytocin receptor (OXTR; related to the neurobiology of social bonds), and brain-derived neurotrophic factor (BDNF; involved in postnatal plasticity). Although no candidate gene DNAm sites significantly associated with early postnatal contact, when we next examined DNAm across the genome, differentially methylated regions were identified between high and low contact groups. Using a different application of epigenomic information, we also quantified epigenetic age, and report that for infants who received low contact from caregivers, greater infant distress was associated with younger epigenetic age. These results suggested that early postnatal contact has lasting associations with child biology.

Trends in DNA Methylation with Age Replicate Across Diverse Human Populations.

Aging is associated with widespread changes in genome-wide patterns of DNA methylation. Thousands of CpG sites whose tissue-specific methylation levels are strongly correlated with chronological age have been previously identified. However, the majority of these studies have focused primarily on cosmopolitan populations living in the developed world; it is not known if age-related patterns of DNA methylation at these loci are similar across a broad range of human genetic and ecological diversity. We investigated genome-wide methylation patterns using saliva- and whole blood-derived DNA from two traditionally hunting and gathering African populations: the Baka of the western Central African rain forest and the ≠Khomani San of the South African Kalahari Desert. We identified hundreds of CpG sites whose methylation levels are significantly associated with age, thousands that are significant in a meta-analysis, and replicate trends previously reported in populations of non-African descent. We confirmed that an age-associated site in the promoter of the gene ELOVL2 shows a remarkably congruent relationship with aging in humans, despite extensive genetic and environmental variation across populations. We also demonstrate that genotype state at methylation quantitative trait loci (meQTLs) can affect methylation trends at some age-associated CpG sites. Our study explores the relationship between CpG methylation and chronological age in populations of African hunter-gatherers, who rely on different diets across diverse ecologies. While many age-related CpG sites replicate across populations, we show that considering common genetic variation at meQTLs further improves our ability to detect previously identified age associations.

DNA methylation signature of human fetal alcohol spectrum disorder.

BACKGROUND: Prenatal alcohol exposure is the leading preventable cause of behavioral and cognitive deficits, which may affect between 2 and 5 % of children in North America. While the underlying mechanisms of alcohol's effects on development remain relatively unknown, emerging evidence implicates epigenetic mechanisms in mediating the range of symptoms observed in children with fetal alcohol spectrum disorder (FASD). Thus, we investigated the effects of prenatal alcohol exposure on genome-wide DNA methylation in the NeuroDevNet FASD cohort, the largest cohort of human FASD samples to date. METHODS: Genome-wide DNA methylation patterns of buccal epithelial cells (BECs) were analyzed using the Illumina HumanMethylation450 array in a Canadian cohort of 206 children (110 FASD and 96 controls). Genotyping was performed in parallel using the Infinium HumanOmni2.5-Quad v1.0 BeadChip. RESULTS: After correcting for the effects of genetic background, we found 658 significantly differentially methylated sites between FASD cases and controls, with 41 displaying differences in percent methylation change >5 %. Furthermore, 101 differentially methylated regions containing two or more CpGs were also identified, overlapping with 95 different genes. The majority of differentially methylated genes were highly expressed at the level of mRNA in brain samples from the Allen Brain Atlas, and independent DNA methylation data from cortical brain samples showed high correlations with BEC DNA methylation patterns. Finally, overrepresentation analysis of genes with up-methylated CpGs revealed a significant enrichment for neurodevelopmental processes and diseases, such as anxiety, epilepsy, and autism spectrum disorders. CONCLUSIONS: These findings suggested that prenatal alcohol exposure is associated with distinct DNA methylation patterns in children and adolescents, raising the possibility of an epigenetic biomarker of FASD.

DNA methylation profiling in human Huntington's disease brain.

Despite extensive progress in Huntington's disease (HD) research, very little is known about the association of epigenetic variation and HD pathogenesis in human brain tissues. Moreover, its contribution to the tissue-specific transcriptional regulation of the huntingtin gene (HTT), in which HTT expression levels are highest in brain and testes, is currently unknown. To investigate the role of DNA methylation in HD pathogenesis and tissue-specific expression of HTT, we utilized the Illumina HumanMethylation450K BeadChip array to measure DNA methylation in a cohort of age-matched HD and control human cortex and liver tissues. In cortex samples, we found minimal evidence of HD-associated DNA methylation at probed sites after correction for cell heterogeneity but did observe an association with the age of disease onset. In contrast, comparison of matched cortex and liver samples revealed tissue-specific DNA methylation of the HTT gene region at 38 sites (FDR < 0.05). Importantly, we identified a novel differentially methylated binding site in the HTT proximal promoter for the transcription factor CTCF. This CTCF site displayed increased occupancy in cortex, where HTT expression is higher, compared with the liver. Additionally, CTCF silencing reduced the activity of an HTT promoter-reporter construct, suggesting that CTCF plays a role in regulating HTT promoter function. Overall, although we were unable to detect HD-associated DNA methylation alterations at queried sites, we found that DNA methylation may be correlated to the age of disease onset in cortex tissues. Moreover, our data suggest that DNA methylation may, in part, contribute to tissue-specific HTT transcription through differential CTCF occupancy.

MicroRNA-223 dose levels fine tune proliferation and differentiation in human cord blood progenitors and acute myeloid leukemia.

A precise understanding of the role of miR-223 in human hematopoiesis and in the pathogenesis of acute myeloid leukemia (AML) is still lacking. By measuring miR-223 expression in blasts from 115 AML patients, we found significantly higher miR-223 levels in patients with favorable prognosis, whereas patients with low miR-223 expression levels were associated with worse outcome. Furthermore, miR-223 was hierarchically expressed in AML subpopulations, with lower expression in leukemic stem cell-containing fractions. Genetic depletion of miR-223 decreased the leukemia initiating cell (LIC) frequency in a myelomonocytic AML mouse model, but it was not mandatory for rapid-onset AML. To relate these observations to physiologic myeloid differentiation, we knocked down or ectopically expressed miR-223 in cord-blood CD34⁺ cells using lentiviral vectors. Although miR-223 knockdown delayed myeloerythroid precursor differentiation in vitro, it increased myeloid progenitors in vivo following serial xenotransplantation. Ectopic miR-223 expression increased erythropoiesis, T lymphopoiesis, and early B lymphopoiesis in vivo. These findings broaden the role of miR-223 as a regulator of the expansion/differentiation equilibrium in hematopoietic stem and progenitor cells where its impact is dose- and differentiation-stage-dependent. This also explains the complex yet minor role of miR-223 in AML, a heterogeneous disease with variable degree of myeloid differentiation.

A pooling-based approach to mapping genetic variants associated with DNA methylation.

DNA methylation is an epigenetic modification that plays a key role in gene regulation. Previous studies have investigated its genetic basis by mapping genetic variants that are associated with DNA methylation at specific sites, but these have been limited to microarrays that cover <2% of the genome and cannot account for allele-specific methylation (ASM). Other studies have performed whole-genome bisulfite sequencing on a few individuals, but these lack statistical power to identify variants associated with DNA methylation. We present a novel approach in which bisulfite-treated DNA from many individuals is sequenced together in a single pool, resulting in a truly genome-wide map of DNA methylation. Compared to methods that do not account for ASM, our approach increases statistical power to detect associations while sharply reducing cost, effort, and experimental variability. As a proof of concept, we generated deep sequencing data from a pool of 60 human cell lines; we evaluated almost twice as many CpGs as the largest microarray studies and identified more than 2000 genetic variants associated with DNA methylation. We found that these variants are highly enriched for associations with chromatin accessibility and CTCF binding but are less likely to be associated with traits indirectly linked to DNA, such as gene expression and disease phenotypes. In summary, our approach allows genome-wide mapping of genetic variants associated with DNA methylation in any tissue of any species, without the need for individual-level genotype or methylation data.

Prenatal antidepressant exposure associated with CYP2E1 DNA methylation change in neonates.

Some but not all neonates are affected by prenatal exposure to serotonin reuptake inhibitor antidepressants (SRI) and maternal mood disturbances. Distinguishing the impact of these 2 exposures is challenging and raises critical questions about whether pharmacological, genetic, or epigenetic factors can explain the spectrum of reported outcomes. Using unbiased DNA methylation array measurements followed by a detailed candidate gene approach, we examined whether prenatal SRI exposure was associated with neonatal DNA methylation changes and whether such changes were associated with differences in birth outcomes. Prenatal SRI exposure was first associated with increased DNA methylation status primarily at CYP2E1(ß(Non-exposed) = 0.06, ß(SRI-exposed) = 0.30, FDR = 0); however, this finding could not be distinguished from the potential impact of prenatal maternal depressed mood. Then, using pyrosequencing of CYP2E1 regulatory regions in an expanded cohort, higher DNA methylation status--both the mean across 16 CpG sites (P < 0.01) and at each specific CpG site (P < 0.05)--was associated with exposure to lower 3rd trimester maternal depressed mood symptoms only in the SRI-exposed neonates, indicating a maternal mood x SRI exposure interaction. In addition, higher DNA methylation levels at CpG2 (P = 0.04), CpG9 (P = 0.04) and CpG10 (P = 0.02), in the interrogated CYP2E1 region, were associated with increased birth weight independently of prenatal maternal mood, SRI drug exposure, or gestational age at birth. Prenatal SRI antidepressant exposure and maternal depressed mood were associated with altered neonatal CYP2E1 DNA methylation status, which, in turn, appeared to be associated with birth weight.

Brain-derived neurotrophic factor (BDNF) Val66Met polymorphism influences the association of the methylome with maternal anxiety and neonatal brain volumes.

Early life environments interact with genotype to determine stable phenotypic outcomes. Here we examined the influence of a variant in the brain-derived neurotropic factor (BDNF) gene (Val66Met), which underlies synaptic plasticity throughout the central nervous system, on the degree to which antenatal maternal anxiety associated with neonatal DNA methylation. We also examined the association between neonatal DNA methylation and brain substructure volume, as a function of BDNF genotype. Infant, but not maternal, BDNF genotype dramatically influences the association of antenatal anxiety on the epigenome at birth as well as that between the epigenome and neonatal brain structure. There was a greater impact of antenatal maternal anxiety on the DNA methylation of infants with the methionine (Met)/Met compared to both Met/valine (Val) and Val/Val genotypes. There were significantly more cytosine-phosphate-guanine sites where methylation levels covaried with right amygdala volume among Met/Met compared with both Met/Val and Val/Val carriers. In contrast, more cytosine-phosphate-guanine sites covaried with left hippocampus volume in Val/Val infants compared with infants of the Met/Val or Met/Met genotype. Thus, antenatal Maternal Anxiety × BDNF Val66Met Polymorphism interactions at the level of the epigenome are reflected differently in the structure of the amygdala and the hippocampus. These findings suggest that BDNF genotype regulates the sensitivity of the methylome to early environment and that differential susceptibility to specific environmental conditions may be both tissue and function specific.

Infinium monkeys: Infinium 450K array for the Cynomolgus macaque (Macaca fascicularis).

The Infinium Human Methylation450 BeadChip Array (Infinium 450K) is a robust and cost-efficient survey of genome-wide DNA methylation patterns. Macaca fascicularis (Cynomolgus macaque) is an important disease model; however, its genome sequence is only recently published, and few tools exist to interrogate the molecular state of Cynomolgus macaque tissues. Although the Infinium 450K is a hybridization array designed to the human genome, the relative conservation between the macaque and human genomes makes its use in macaques feasible. Here, we used the Infinium 450K array to assay DNA methylation in 11 macaque muscle biopsies. We showed that probe hybridization efficiency was related to the degree of sequence identity between the human probes and the macaque genome sequence. Approximately 61% of the Human Infinium 450K probes could be reliably mapped to the Cynomolgus macaque genome and contain a CpG site of interest. We also compared the Infinium 450K data to reduced representation bisulfite sequencing data generated on the same samples and found a high level of concordance between the two independent methodologies, which can be further improved by filtering for probe sequence identity and mismatch location. We conclude that the Infinium 450K array can be used to measure the DNA methylome of Cynomolgus macaque tissues using the provided filters. We also provide a pipeline for validation of the array in other species using a simple BLAST-based sequence identify filter.

The effect of genotype and in utero environment on interindividual variation in neonate DNA methylomes.

Integrating the genotype with epigenetic marks holds the promise of better understanding the biology that underlies the complex interactions of inherited and environmental components that define the developmental origins of a range of disorders. The quality of the in utero environment significantly influences health over the lifecourse. Epigenetics, and in particular DNA methylation marks, have been postulated as a mechanism for the enduring effects of the prenatal environment. Accordingly, neonate methylomes contain molecular memory of the individual in utero experience. However, interindividual variation in methylation can also be a consequence of DNA sequence polymorphisms that result in methylation quantitative trait loci (methQTLs) and, potentially, the interaction between fixed genetic variation and environmental influences. We surveyed the genotypes and DNA methylomes of 237 neonates and found 1423 punctuate regions of the methylome that were highly variable across individuals, termed variably methylated regions (VMRs), against a backdrop of homogeneity. MethQTLs were readily detected in neonatal methylomes, and genotype alone best explained ∼25% of the VMRs. We found that the best explanation for 75% of VMRs was the interaction of genotype with different in utero environments, including maternal smoking, maternal depression, maternal BMI, infant birth weight, gestational age, and birth order. Our study sheds new light on the complex relationship between biological inheritance as represented by genotype and individual prenatal experience and suggests the importance of considering both fixed genetic variation and environmental factors in interpreting epigenetic variation.

Comprehensive analysis of mammalian miRNA* species and their role in myeloid cells.

Processing of pre-miRNA through Dicer1 generates an miRNA duplex that consists of an miRNA and miRNA* strand. Despite the general view that miRNA*s have no functional role, we further investigated miRNA* species in 10 deep-sequencing libraries from mouse and human tissue. Comparisons of miRNA/miRNA* ratios across the miRNA sequence libraries revealed that 50% of the investigated miRNA duplexes exhibited a highly dominant strand. Conversely, 10% of miRNA duplexes showed a comparable expression of both strands, whereas the remaining 40% exhibited variable ratios across the examined libraries, as exemplified by miR-223/miR-223* in murine and human cell lines. Functional analyses revealed a regulatory role for miR-223* in myeloid progenitor cells, which implies an active role for both arms of the miR-223 duplex. This was further underscored by the demonstration that miR-223 and miR-223* targeted the insulin-like growth factor 1 receptor/phosphatidylinositol 3-kinase axis and that high miR-223* levels were associated with increased overall survival in patients with acute myeloid leukemia. Thus, we found a supporting role for miR-223* in differentiating myeloid cells in normal and leukemic cell states. The fact that the miR-223 duplex acts through both arms extends the complexity of miRNA-directed gene regulation of this myeloid key miRNA.