DNA methylation profiling in peripheral lung tissues of smokers and patients with COPD

Background: Epigenetics changes have been shown to be affected by cigarette smoking. Cigarette smoke (CS)-mediated DNA methylation can potentially affect several cellular and pathophysiological processes, acute exacerbations, and comorbidity in the lungs of patients with chronic obstructive pulmonary disease (COPD). We sought to determine whether genome-wide lung DNA methylation profiles of smokers and patients with COPD were significantly different from non-smokers. We isolated DNA from parenchymal lung tissues of patients including eight lifelong non-smokers, eight current smokers, and eight patients with COPD and analyzed the samples using Illumina's Infinium HumanMethylation450 BeadChip. Results: Our data revealed that the differentially methylated genes were related to top canonical pathways (e.g., G beta gamma signaling, mechanisms of cancer, and nNOS signaling in neurons), disease and disorders (organismal injury and abnormalities, cancer, and respiratory disease), and molecular and cellular functions (cell death and survival, cellular assembly and organization, cellular function and maintenance) in patients with COPD. The genome-wide DNA methylation analysis identified suggestive genes, such as NOS1AP, TNFAIP2, BID, GABRB1, ATXN7, and THOC7 with DNA methylation changes in COPD lung tissues that were further validated by pyrosequencing. Pyrosequencing validation confirmed hyper-methylation in smokers and patients with COPD as compared to non-smokers. However, we did not detect significant differences in DNA methylation for TNFAIP2, ATXN7, and THOC7 genes in smokers and COPD groups despite the changes observed in the genome-wide analysis. Conclusions: Our study suggests that DNA methylation in suggestive genes, such as NOS1AP, BID, and GABRB1 may be used as epigenetic signatures in smokers and patients with COPD if the same is validated in a larger cohort. Future studies are required to correlate DNA methylation status with transcriptomics of selective genes identified in this study and elucidate their role and involvement in the progression of COPD and its exacerbations.Peer reviewe

O USO DE ÍNDICES DE AVALIAÇÃO COMO ARGUMENTO DE QUALIDADE DAS INSTITUIÇÕES DE EDUCAÇÃO SUPERIOR

CpGs differentially methylated in lung DNA in relation to COPD vs. non-smokers (P < 0.01). (CSV 273 kb

The bisphenol A experience: a primer for the analysis of environmental effects on mammalian reproduction

It is increasingly evident that environmental factors are a veritable Pandora's box from which new concerns and complications continue to emerge. Although previously considered the domain of toxicologists, it is now clear that an understanding of the effects of the environment on reproduction requires a far broader range of expertise and that, at least for endocrine-disrupting chemicals, many of the tenets of classical toxicology need to be revisited. Indeed, because of the wide range of reproductive effects induced by these chemicals, interest among reproductive biologists has grown rapidly: in 2000, the program for the annual Society for the Study of Reproduction meeting included a single minisymposium on the fetal origins of adult disease, one platform session on endocrine disruption, and 23 toxicology poster presentations. In contrast, environmental factors featured prominently at the 2009 meeting, with strong representation in the plenary, minisymposia, platform, and poster sessions. Clearly, a lot has happened in a decade, and environmental issues have become an increasingly important research focus for reproductive biologists. In this review, we summarize some of the inherent difficulties in assessing environmental effects on reproductive performance, focusing on the endocrine disruptor bisphenol A (BPA) to illustrate important emerging concerns. In addition, because the BPA experience serves as a prototype for scientific activism, public education, and advocacy, these issues are also discussed

Bisphenol A Exposure In Utero Disrupts Early Oogenesis in the Mouse

Estrogen plays an essential role in the growth and maturation of the mammalian oocyte, and recent studies suggest that it also influences follicle formation in the neonatal ovary. In the course of studies designed to assess the effect of the estrogenic chemical bisphenol A (BPA) on mammalian oogenesis, we uncovered an estrogenic effect at an even earlier stage of oocyte development—at the onset of meiosis in the fetal ovary. Pregnant mice were treated with low, environmentally relevant doses of BPA during mid-gestation to assess the effect of BPA on the developing ovary. Oocytes from exposed female fetuses displayed gross aberrations in meiotic prophase, including synaptic defects and increased levels of recombination. In the mature female, these aberrations were translated into an increase in aneuploid eggs and embryos. Surprisingly, we observed the same constellation of meiotic defects in fetal ovaries of mice homozygous for a targeted disruption of ERb, one of the two known estrogen receptors. This, coupled with the finding that BPA exposure elicited no additional effects in ERb null females, suggests that BPA exerts its effect on the early oocyte by interfering with the actions of ERb. Together, our results show that BPA can influence early meiotic events and, importantly, indicate that the oocyte itself may be directly responsive to estrogen during early oogenesis. This raises concern that brief exposures during fetal development to substances that mimic or antagonize the effects of estrogen may adversely influence oocyte development in the exposed female fetus

Bisphenol A Exposure Disrupts Genomic Imprinting in the Mouse

<div><p>Exposure to endocrine disruptors is associated with developmental defects. One compound of concern, to which humans are widely exposed, is bisphenol A (BPA). In model organisms, BPA exposure is linked to metabolic disorders, infertility, cancer, and behavior anomalies. Recently, BPA exposure has been linked to DNA methylation changes, indicating that epigenetic mechanisms may be relevant. We investigated effects of exposure on genomic imprinting in the mouse as imprinted genes are regulated by differential DNA methylation and aberrant imprinting disrupts fetal, placental, and postnatal development. Through allele-specific and quantitative real-time PCR analysis, we demonstrated that maternal BPA exposure during late stages of oocyte development and early stages of embryonic development significantly disrupted imprinted gene expression in embryonic day (E) 9.5 and 12.5 embryos and placentas. The affected genes included <i>Snrpn, Ube3a, Igf2, Kcnq1ot1, Cdkn1c</i>, and <i>Ascl2</i>; mutations and aberrant regulation of these genes are associated with imprinting disorders in humans. Furthermore, the majority of affected genes were expressed abnormally in the placenta. DNA methylation studies showed that BPA exposure significantly altered the methylation levels of differentially methylated regions (DMRs) including the <i>Snrpn</i> imprinting control region (ICR) and <i>Igf2</i> DMR1. Moreover, exposure significantly reduced genome-wide methylation levels in the placenta, but not the embryo. Histological and immunohistochemical examinations revealed that these epigenetic defects were associated with abnormal placental development. In contrast to this early exposure paradigm, exposure outside of the epigenetic reprogramming window did not cause significant imprinting perturbations. Our data suggest that early exposure to common environmental compounds has the potential to disrupt fetal and postnatal health through epigenetic changes in the embryo and abnormal development of the placenta.</p> </div

BPA exposure altered DNA methylation at the <i>Igf2</i> DMR1 and <i>H19/Igf2</i> ICR in the embryos.

<p>A. Pyrosequencing assays tested methylation of 4 and 6 CpGs (highlighted in bold) in 457 bp and 221 bp genomic regions at the <i>Igf2</i> DMR1 and <i>H19/Igf2</i> ICR, respectively. Methylation at the (B) <i>Igf2</i> DMR1 and (C) <i>H19/Igf2</i> ICR was analyzed in embryos from controls, lower dose and upper dose exposure groups. Analysis in the upper dose exposure group included embryos that showed monoallelic (black circles) or biallelic (red circles) expression of the <i>Igf2</i> gene. Sample sizes analyzed in each exposure group are indicated with a = P<0.05 and b = P<0.01. Black horizontal line indicates average methylation level in each exposure group.</p

Representative LC-SRM/MS chromatograms demonstrating significantly higher BPA level in serum from treated mice.

<p>Top panels represent the chromatograms for unlabeled BPA and lower panels labeled BPA ([¹³C₁₂]-BPA) spiked prior to sample extraction to quantify BPA levels in serum from representative control (B) and upper dose treated mice (C). Blank sample used as control for storage and extraction is indicated in (A). Y-axis represents relative abundance of signal intensity and X-axis retention time in minute. Differences in BPA concentrations were determined based on peak heights (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003401#s4" target="_blank">Materials and Methods</a>). NL = normalized levels of intensity; <i>m/z = </i>mass to charge ratio.</p

Map of mouse chromosome 7 showing imprinted domains analyzed in the current work is illustrated.

<p>Black = paternally expressed genes; dark grey = maternally expressed genes; and light grey = differentially methylated regions (DMRs). Arrows indicated direction of gene transcription. C = centromere and T = telomere. Not drawn to scale.</p

Impact of BPA exposure is shown as number of genes exhibiting imprinting perturbations.

<p>Effects of BPA exposure were more significant in the (B) placentas compared to (A) embryos. For each exposure group (control [blue], lower dose [pink] or upper dose [red]), percentage of tissues showing LOI (Y-axis) of at least 1 gene (≥1), 2 genes (≥2) or 3 genes (≥3) is indicated; a = P<0.05; b = P<0.01; c = P<0.001.</p