DNA methylation heatmaps of CpG sites in PBMCs and AECs from healthy, atopic and asthmatic pediatric subjects.

<p>AECs and PBMCs were analyzed for 1027 CpG loci in 671 genes from healthy (A), atopic (B), and asthmatic (C) subjects. Heatmaps of z-scores for AECs and PBMCs are shown with individuals (columns) and differential CpG sites (rows). Increased methylation is shown in red and decreased methylation in blue. D. Venn diagram showing overlap of differentially methylated sites between healthy, atopic and asthmatic subjects. Numbers in black indicate total number of CpG sites while numbers in red/blue indicate more/less methylated in AECs (compared to PBMCs). E. The molecular and cellular functions in the 47 genes classified by IPA. The x-axis shows functions while the y-axis shows –log(p-value).</p

Patient demographics in DNA methylation cohort prior to surgery.

<p>Subjects are identified as healthy, atopic, atopic asthmatic, or non-atopic asthmatic.</p

DNA methylation profile of AECs compared to PBMCs.

<p>DNA methylation for 1027 CpG sites was assessed in AECs compared to PBMCs from all subjects. A. Volcano plot of CpG sites interrogated with red and blue points indicating significantly over- and under-methylated sites. Q-values are shown on the y-axis (−log₁₀) and z-score difference on the x-axis (log₂). Dashed lines indicate cut-offs for significance. B. Heatmap illustrating z-scores of 80 differentially methylated loci in AECs compared to PBMCs. Columns represent subjects and rows represent CpG sites while red/blue indicates more/less methylated. C. The molecular and cellular functions of the 67 genes classified by IPA. The x-axis shows functions while the y-axis shows –log(p-value).</p

Patient demographics in gene expression cohort prior to surgery.

<p>Subjects are identified as healthy, atopic, or atopic asthmatic.</p

Differentially Methylated CpGs in Atopic Compared to Asthmatic Derived AECs. Z-score difference is presented as atopic relative to asthmatic derived AECs.

<p>Differentially Methylated CpGs in Atopic Compared to Asthmatic Derived AECs. Z-score difference is presented as atopic relative to asthmatic derived AECs.</p

DNA methylation analysis of atopic asthmatics compared to non-atopic asthmatics individuals in AECs and PBMCs.

<p>Volcano plots of airway epithelial cells (AECs) (A) and PBMCs (B), analyzed for 1027 CpG loci in 671 genes, from atopic and non-atopic asthmatic subjects. Y-axis represents the q-values (−log₁₀) for all of the CpG sites analyzed and the x-axis is the z-score difference (log₂). Dashed lines indicate cut-offs for significance. These results show no differences between atopic and non-atopic asthma.</p

STAT5A and CRIP1 Gene Expression and DNA Methylation Status in AECs.

<p>AECs from atopic, healthy and asthmatic individuals were analyzed for STAT5A (A) and CRIP1 (C) mRNA expression using RT-PCR. Results are expressed as gene expression normalized to PPIA (y-axis). DNA methylation status is shown as M-values for STAT5A_E42_F (B) and CRIP1_P874_R (D) for the three phenotypes. * indicates differential methylation as detailed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044213#pone-0044213-t003" target="_blank">Table 3</a>.</p

Differential methylation between disease phenotypes in AECs or PBMCs.

<p>Volcano plots of CpG sites interrogated with red and blue points indicating significantly over- and under-methylated sites. Q-values are shown on the y-axis (−log₁₀) and z-score difference on the x-axis (log₂). Dashed lines indicate cut-offs for significance. Within AECs, differences in DNA methylation were assessed in healthy subjects compared to atopics (A) and asthmatics (B) as well as atopic subjects compared to asthmatics (C). The same comparisons were performed in PBMCs (D, E, and F).</p