Particulate matter-induced hypomethylation of Alu and LINE1 in normal human bronchial epithelial cells and epidermal keratinocytes

Abstract

Background: Airborne particulate matter (PM), a complex mixture of organic and inorganic compounds, is a major public health concern due to its adverse health effects. Understanding the biological action of PM is of particular importance in the improvement of public health. Differential methylation of repetitive elements (RE) by PM might have severe consequences for the structural integrity of the genome and on transcriptional activity, thereby affecting human health. This study aimed to evaluate the effect of inhaled and non-inhaled PM (PM_2.5, PM₁₀, and PM₁₀-PAH) exposure on DNA methylation. We quantitatively measured the methylation content of Alu and LINE1 in PM-treated normal human bronchial epithelial cells (NHBE) and normal human epidermal keratinocytes (NHEK) by using whole-genome bisulfite sequencing and pyrosequencing.

Results: All PMs exposure significantly lowered Alu and LINE1 methylation in both cells than in mock-treated controls. Hypomethylation was more prominent in PM₁₀-PAH exposed-NHBE and PM₁₀ exposed-NHEK. Alu and LINE1 methylation change exhibited different sensitivity according to the subfamily evolutionary ages, with stronger effects on the oldest L1-M and Alu J in NHBE, and oldest L1-M and youngest Alu S in NHEK.

Conclusions: These results demonstrate that the differential susceptibility of PM-induced hypomethylation of Alu and LINE1 depends upon RE evolutionary age and PM type.

Keywords: Alu; DNA methylation; LINE1; Particulate matter; Subfamily; WGBS.

Fig. 1

Alu and LINE1 methylation change in NHBE and NHEK after PM exposure. The hypomethylation ratio was calculated as a percentage of CpGs with low methylation to analyzed total CpG sites and the means ± SE of three independent WGBS experiments are shown. The mean methylation ratios of PM-treated cells were compared to mock-treated cells using one-way ANOVA, and Bonferroni’s correction p-value was used for post-hoc comparison between two groups where the ANOVA was significant

Fig. 2

Differential hypomethylation of Alu and LINE1 evolutionary subfamilies in NHBE and NHEK following PM treatment. (A) Alu change in NHBE. (B) LINE1 change in NHBE. (C) Alu change in NHEK. (D) LINE1 change in NHEK. The means ± SE of three independent WGBS experiments are shown. Bonferroni-adjusted p-value was the result of comparing between the two groups

Fig. 3

Distribution of Alu and LINE1 methylation level in PM-exposed NHBE (A) and NHEK (B). Methylation level was expressed as a percentage of 5-methylcytosine divided by the sum of methylated and unmethylated cytosines. The mean ± 95% confidence interval of three independent pyrosequencing is shown. The comparisons of mean methylation levels were evaluated using ANOVA. Bonferroni-adjusted p-value was the result of comparing between the two groups. Cells maintained in culture medium with vehicle were used as control group (CTL^a, 0.1% DMSO; CTL^b, 1% PBS)