Air Pollution,Morphological Characteristics And Neonatal DNA Methylation

Objective The aim of this study was to screen the gene which has strong link in DNA methylation between peripheral blood in mothers and cord blood in newborn, and then explore the associations of exposures to air pollutants during pregnancy, DNA methylation level and morphological characters of newborn.Subjects and methods 1. Study populationA pilot study was conducted in Houzhai Center Hospital in Zhengzhou. Adult pregnant women, who came to the hospital for delivery between January 2010 and January 2012, were considered eligible for the study. And a total of 568 potentially eligible subjects were identified.2. Determination of morphological characteristicsThe birth weight and length of neonate were measured using horizontal height and weight measuring instrument, and the birth head circumference was measured by measuring tape. The characteristics of parents were measured using height and weight measuring instrument.3. Extraction of genomic DNAGenomic DNA samples were extracted from maternal peripheral blood and neonatal cord blood using the Bioteke extraction kit.4. Screening of methylated sites in genes which have strong link between peripheral blood in mothers and cord blood in newbornDNA methylated site which have strong link between peripheral blood in mothers and cord blood in newborn by genome-wide methylation analysis using Illumina Human Methylation 27 K Bead Chip.5. Determination of the methylation level in promoter region of target geneThe methylation level in promoter region of target gene were tested by Quantitative Methylation Specific Polymerase Chain Reaction（QMS-PCR）.6. Statistical analysisAll the data was entered by the software of Epidata, version 3.1. The data visualization was processed using the R ggplot2 package. All of data was further analyzed by the software of SPSS（version 21.0）.Results 1. Screening and verification of target gene.G Protein-Coupled Receptor（GPR61）gene was screened out from total 27578 Cp G sites as the target gene. The determination coefficient of methylation levels of promoter region of GPR61 in mothers and cord blood was 0.291.2. Associations of morphological characters of neonates with air pollution during pregnancy and morphological characters of neonates.In male neonates, the birth weight was correlated with exposed to SO₂ and NO₂ during pregnancy（P<0.05 respectively）, and the birth height was correlated with exposed to SO₂, NO₂ and PM10 respectively during pregnancy. During first trimester, the birth weight and height were negatively correlated with SO₂ exposure（P<0.05 respectively）, and birth height was positively correlated with NO₂ exposure（β=0.386, P<0.001）. There was a significant correlation between birth weight, as well as birth height, and exposed to SO₂ and NO₂ during second trimester（P<0.05 respectively）. In addition, birth weight and height was negatively correlated with exposed to PM10 during third trimester（P<0.05 respectively）.In female neonates, the birth weight and height were significantly correlated with exposed to SO₂, NO₂ during pregnancy（P<0.05 respectively）. There was a negative correlation between birth height and exposures of SO₂ during first trimester（β=-0.278, P=0.014）. During first trimester, the birth height was positively correlated with NO₂ exposure（β=0.245, P=0.025）, while the birth height was negatively correlated with the exposure of PM10（β=-0.193, P=0.012）.3. Methylation level of GPR61, maternal morphological characters and the morphological characters of neonates.There was a positive correlation between male neonatal birth weight and maternal weight gain during pregnancy（r=0.260, P<0.001）, and a significant negative correlation between male neonatal birth height and maternal weight gain during pregnancy（r=-0.129, P=0.046）. Besides, the female neonatal birth heightwas positively correlated with the maternal weight gain during pregnancy（r=0.189, P=0.009）.4. Methylation level of GPR61 in promoter region and the exposures of air pollutant during pregnancy.When we adjusted confounding factors of GPR61 methylation levels of neonates（namely, the GPR61 methylation levels of mothers）, we found that the exposures of SO₂, NO₂ and PM10 during pregnancy was significantly correlated with neonatal GPR61 methylation levels（P<0.05 respectively）. During first trimester, the exposure of SO₂ was negatively correlated with the GPR61 methylation levels of neonates（β=-0.258, P<0.001）. Besides, there was a significant correlation between the neonatal GPR61 methylation level and the exposures of SO₂ and NO₂ during second trimester（P<0.05 respectively）. In addition, the exposures of PM10 during third trimester was negatively correlated with the GPR61 methylation level of neonates（β=-0.145, P<0.001）.Conclusions1. The methylation level of GPR61 promoter region in cord blood is affected by the exposures of SO₂, NO₂ and PM10 during pregnancy.2. The methylation level in promoter region of GPR61 in neonate is affected by the methylation level in this site of mother.3. The morphological characters of neonate are correlated with the exposures of SO₂, NO₂ and PM10, and mother’s weight gain during pregnancy.