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A varying T cell subtype explains apparent tobacco smoking induced single CpG hypomethylation in whole blood.

Bauer M, Linsel G, Fink B, Offenberg K, Hahn AM, Sack U, Knaack H, Eszlinger M, Herberth G - Clin Epigenetics (2015)

Bottom Line: Within two independent cohorts, we confirmed the differentially expression of the GPR15 gene when smokers and non-smokers subjects are compared.Treatment of peripheral blood mononuclear cell (PBMC) cultures with aqueous cigarette smoke extract did not induce a higher proportion of this T cell subtype.Our results underline that DNA hypomethylation at cg19859270 site, observed in WBCs of smokers, did not arise by direct effect of tobacco smoking compounds on methylation of DNA but rather by the enrichment of a tobacco-smoking-induced lymphocyte population in the peripheral blood.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Immunology, Helmholtz Centre for Environmental Research-UFZ, Leipzig, 04318 Germany.

ABSTRACT

Background: Many recent epigenetic studies report that cigarette smoking reduces DNA methylation in whole blood at the single CpG site cg19859270 within the GPR15 gene.

Results: Within two independent cohorts, we confirmed the differentially expression of the GPR15 gene when smokers and non-smokers subjects are compared. By validating the GPR15 protein expression at the cellular level, we found that the observed decreased methylation at this site in white blood cells (WBC) of smokers is mainly caused by the high proportion of CD3+GPR15+ expressing T cells in peripheral blood. In current smokers, the percentage of GPR15+ cells among CD3+ T cells in peripheral blood is significantly higher (15.5 ± 7.2 %, mean ± standard deviation) compared to non-smokers (3.7 ± 1.6 %). Treatment of peripheral blood mononuclear cell (PBMC) cultures with aqueous cigarette smoke extract did not induce a higher proportion of this T cell subtype.

Conclusions: Our results underline that DNA hypomethylation at cg19859270 site, observed in WBCs of smokers, did not arise by direct effect of tobacco smoking compounds on methylation of DNA but rather by the enrichment of a tobacco-smoking-induced lymphocyte population in the peripheral blood.

No MeSH data available.


Percentage of GPR15 protein expressing lymphocytes in blood specimens of the Replication Cohort (91 non-smokers vs. 32 smokers in total). a Representative dot plots gated on lymphocytes show GPR15 expression in CD3+ T cells and CD19+ B cells in smoker and non-smoker. Percentages represent the frequency of these cells in the lymphocyte gate. b Cumulative data of the GPR15 expressing cells as percentage of CD3+ or CD19+ lymphocytes. P values from Mann-Whitney U Test
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Fig2: Percentage of GPR15 protein expressing lymphocytes in blood specimens of the Replication Cohort (91 non-smokers vs. 32 smokers in total). a Representative dot plots gated on lymphocytes show GPR15 expression in CD3+ T cells and CD19+ B cells in smoker and non-smoker. Percentages represent the frequency of these cells in the lymphocyte gate. b Cumulative data of the GPR15 expressing cells as percentage of CD3+ or CD19+ lymphocytes. P values from Mann-Whitney U Test

Mentions: In order to identify the blood cell type expressing GPR15, flow cytometric analyses were performed using whole blood samples from the Replication Cohort as well as peripheral blood mononuclear cells (PBMCs) from buffy coats. The main population expressing GPR15 was CD3+ T cells (Fig. 2a, see Additional file 3). Smokers had a significantly (p = 1.8 × 10−10) increased proportion of GPR15+ cells among CD3+ T cells (15.5 ± 7.2 %, mean ± standard deviation) compared to non-smoker (3.7 ± 1.6 %, Fig. 2b). By setting a cutoff of 9 % GPR15 expressing cells among CD3+ T cells in blood, the flow cytometric analysis could distinguish smoker from non-smoker with high sensitivity (0.88 %) and high specificity (0.99 %). In addition to T cells, a low proportion of B cells expressed GPR15 (Fig. 2a). The proportion of CD19+GPR15+ B cells was significantly (p = 1.5 × 10−5) higher in smoker (7.98 ± 7.54 %) compared to non-smoker (3.81 ± 2.89 %, Fig. 2b).Fig. 2


A varying T cell subtype explains apparent tobacco smoking induced single CpG hypomethylation in whole blood.

Bauer M, Linsel G, Fink B, Offenberg K, Hahn AM, Sack U, Knaack H, Eszlinger M, Herberth G - Clin Epigenetics (2015)

Percentage of GPR15 protein expressing lymphocytes in blood specimens of the Replication Cohort (91 non-smokers vs. 32 smokers in total). a Representative dot plots gated on lymphocytes show GPR15 expression in CD3+ T cells and CD19+ B cells in smoker and non-smoker. Percentages represent the frequency of these cells in the lymphocyte gate. b Cumulative data of the GPR15 expressing cells as percentage of CD3+ or CD19+ lymphocytes. P values from Mann-Whitney U Test
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4526203&req=5

Fig2: Percentage of GPR15 protein expressing lymphocytes in blood specimens of the Replication Cohort (91 non-smokers vs. 32 smokers in total). a Representative dot plots gated on lymphocytes show GPR15 expression in CD3+ T cells and CD19+ B cells in smoker and non-smoker. Percentages represent the frequency of these cells in the lymphocyte gate. b Cumulative data of the GPR15 expressing cells as percentage of CD3+ or CD19+ lymphocytes. P values from Mann-Whitney U Test
Mentions: In order to identify the blood cell type expressing GPR15, flow cytometric analyses were performed using whole blood samples from the Replication Cohort as well as peripheral blood mononuclear cells (PBMCs) from buffy coats. The main population expressing GPR15 was CD3+ T cells (Fig. 2a, see Additional file 3). Smokers had a significantly (p = 1.8 × 10−10) increased proportion of GPR15+ cells among CD3+ T cells (15.5 ± 7.2 %, mean ± standard deviation) compared to non-smoker (3.7 ± 1.6 %, Fig. 2b). By setting a cutoff of 9 % GPR15 expressing cells among CD3+ T cells in blood, the flow cytometric analysis could distinguish smoker from non-smoker with high sensitivity (0.88 %) and high specificity (0.99 %). In addition to T cells, a low proportion of B cells expressed GPR15 (Fig. 2a). The proportion of CD19+GPR15+ B cells was significantly (p = 1.5 × 10−5) higher in smoker (7.98 ± 7.54 %) compared to non-smoker (3.81 ± 2.89 %, Fig. 2b).Fig. 2

Bottom Line: Within two independent cohorts, we confirmed the differentially expression of the GPR15 gene when smokers and non-smokers subjects are compared.Treatment of peripheral blood mononuclear cell (PBMC) cultures with aqueous cigarette smoke extract did not induce a higher proportion of this T cell subtype.Our results underline that DNA hypomethylation at cg19859270 site, observed in WBCs of smokers, did not arise by direct effect of tobacco smoking compounds on methylation of DNA but rather by the enrichment of a tobacco-smoking-induced lymphocyte population in the peripheral blood.

View Article: PubMed Central - PubMed

Affiliation: Department of Environmental Immunology, Helmholtz Centre for Environmental Research-UFZ, Leipzig, 04318 Germany.

ABSTRACT

Background: Many recent epigenetic studies report that cigarette smoking reduces DNA methylation in whole blood at the single CpG site cg19859270 within the GPR15 gene.

Results: Within two independent cohorts, we confirmed the differentially expression of the GPR15 gene when smokers and non-smokers subjects are compared. By validating the GPR15 protein expression at the cellular level, we found that the observed decreased methylation at this site in white blood cells (WBC) of smokers is mainly caused by the high proportion of CD3+GPR15+ expressing T cells in peripheral blood. In current smokers, the percentage of GPR15+ cells among CD3+ T cells in peripheral blood is significantly higher (15.5 ± 7.2 %, mean ± standard deviation) compared to non-smokers (3.7 ± 1.6 %). Treatment of peripheral blood mononuclear cell (PBMC) cultures with aqueous cigarette smoke extract did not induce a higher proportion of this T cell subtype.

Conclusions: Our results underline that DNA hypomethylation at cg19859270 site, observed in WBCs of smokers, did not arise by direct effect of tobacco smoking compounds on methylation of DNA but rather by the enrichment of a tobacco-smoking-induced lymphocyte population in the peripheral blood.

No MeSH data available.