title: DNA methylation of cord blood cell types: Applications for mixed cell birth studies.
abstract: Epigenome-wide association studies of disease widely use DNA methylation measured in blood as a surrogate tissue. Cell proportions can vary between people and confound associations of exposure or outcome. An adequate reference panel for estimating cell proportions from adult whole blood for DNA methylation studies is available, but an analogous cord blood cell reference panel is not yet available. Cord blood has unique cell types and the epigenetic signatures of standard cell types may not be consistent throughout the life course. Using magnetic bead sorting, we isolated cord blood cell types (nucleated red blood cells, granulocytes, monocytes, natural killer cells, B cells, CD4(+)T cells, and CD8(+)T cells) from 17 live births at Johns Hopkins Hospital. We confirmed enrichment of the cell types using fluorescence assisted cell sorting and ran DNA from the separated cell types on the Illumina Infinium HumanMethylation450 BeadChip array. After filtering, the final analysis was on 104 samples at 429,794 probes. We compared cell type specific signatures in cord to each other and methylation at 49.2% of CpG sites on the array differed by cell type (F-test P < 10(-8)). Differences between nucleated red blood cells and the remainder of the cell types were most pronounced (36.9% of CpG sites at P < 10(-8)) and 99.5% of these sites were hypomethylated relative to the other cell types. We also compared the mean-centered sorted cord profiles to the available adult reference panel and observed high correlation between the overlapping cell types for granulocytes and monocytes (both r=0.74), and poor correlation for CD8(+)T cells and NK cells (both r=0.08). We further provide an algorithm for estimating cell proportions in cord blood using the newly developed cord reference panel, which estimates biologically plausible cell proportions in whole cord blood samples.
authors.affiliation: a Department of Epidemiology , Johns Hopkins University Bloomberg School of Public Health , Baltimore , Maryland , USA.|b Center for Epigenetics, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.|c Department of Epidemiology , University of Michigan School of Public Health , Ann Arbor , Michigan , USA.
authors.forename: Kelly M
authors.initials: KM
authors.lastname: Bakulski
authors.affiliation: b Center for Epigenetics, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.|d Department of Mental Health , Johns Hopkins University Bloomberg School of Public Health , Baltimore , Maryland , USA.
authors.forename: Jason I
authors.initials: JI
authors.lastname: Feinberg
authors.affiliation: a Department of Epidemiology , Johns Hopkins University Bloomberg School of Public Health , Baltimore , Maryland , USA.
authors.forename: Shan V
authors.initials: SV
authors.lastname: Andrews
authors.affiliation: e Division of Geriatric Medicine and Gerontology, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.
authors.forename: Jack
authors.initials: J
authors.lastname: Yang
authors.affiliation: a Department of Epidemiology , Johns Hopkins University Bloomberg School of Public Health , Baltimore , Maryland , USA.|f Wendy Klag Center for Autism and Developmental Disabilities, Johns Hopkins Bloomberg School of Public Health , Baltimore , Maryland , USA.
authors.forename: Shannon
authors.initials: S
authors.lastname: Brown
authors.affiliation: g Division of Neonatology, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.
authors.forename: Stephanie
authors.initials: S
authors.lastname: L McKenney
authors.affiliation: h Division of Gynecology and Obstetrics, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.|i Department of International Health , Johns Hopkins University Bloomberg School of Public Health , Baltimore , Maryland , USA.
authors.forename: Frank
authors.initials: F
authors.lastname: Witter
authors.affiliation: e Division of Geriatric Medicine and Gerontology, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.
authors.forename: Jeremy
authors.initials: J
authors.lastname: Walston
authors.affiliation: b Center for Epigenetics, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.|d Department of Mental Health , Johns Hopkins University Bloomberg School of Public Health , Baltimore , Maryland , USA.
authors.forename: Andrew P
authors.initials: AP
authors.lastname: Feinberg
authors.affiliation: b Center for Epigenetics, Johns Hopkins University School of Medicine , Baltimore , Maryland , USA.|d Department of Mental Health , Johns Hopkins University Bloomberg School of Public Health , Baltimore , Maryland , USA.|f Wendy Klag Center for Autism and Developmental Disabilities, Johns Hopkins Bloomberg School of Public Health , Baltimore , Maryland , USA.
authors.forename: M Daniele
authors.initials: MD
authors.lastname: Fallin
mesh_terms: D001402:B-Lymphocytes / Q000378:metabolism; D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism; D018414:CD8-Positive T-Lymphocytes / Q000378:metabolism; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation / Q000235:genetics*; D057890:Epigenomics; D004900:Erythroblasts / Q000378:metabolism; D005260:Female; D005312:Fetal Blood / Q000166:cytology / Q000378:metabolism*; D015894:Genome, Human; D006098:Granulocytes / Q000378:metabolism; D006801:Humans; D007231:Infant, Newborn; D007694:Killer Cells, Natural / Q000378:metabolism; D050498:Live Birth / Q000235:genetics; D008297:Male; D009000:Monocytes / Q000378:metabolism
keywords: Cellular heterogeneity; DNA methylation; cord blood; epidemiology; nucleated red blood cells