Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2018 Jun 15:290:63-72.
doi: 10.1016/j.toxlet.2018.03.021. Epub 2018 Mar 20.

Effects of developmental lead exposure on the hippocampal methylome: Influences of sex and timing and level of exposure

G Singh  1 , V Singh  2 , Zi-Xuan Wang  2 , G Voisin  3 , F Lefebvre  4 , J-M Navenot  2 , B Evans  2 , M Verma  2 , D W Anderson  2 , J S Schneider  2
Affiliations

Effects of developmental lead exposure on the hippocampal methylome: Influences of sex and timing and level of exposure

G Singh et al. Toxicol Lett. .

Abstract

Developmental lead (Pb) exposure results in persistent cognitive/behavioral impairments as well as an elevated risk for developing a variety of diseases in later life. Environmental exposures during development can result in a variety of epigenetic changes, including alterations in DNA methylation, that can influence gene expression patterns and affect the function and development of the nervous system. The present promoter-based methylation microarray profiling study explored the extent to which developmental Pb exposure may modify the methylome of a brain region, hippocampus, known to be sensitive to the effects of Pb exposure. Male and female Long Evans rats were exposed to 0 ppm, 150 ppm, 375 ppm, or 750 ppm Pb through perinatal exposures (gestation through lactation), early postnatal exposures (birth through weaning), or long-term postnatal exposures (birth through postnatal day 55). Results showed a significant contribution of sex to the hippocampal methylome and effects of Pb exposure level, with non-linear dose response effects on methylation. Surprisingly, the developmental period of exposure contributed only a small amount of variance to the overall data and gene ontology (GO) analysis revealed the largest number of overrepresented GO terms in the groups with the lowest level of exposure. The highest number of significant differentially methylated regions was found in females exposed to Pb at the lowest exposure level. Our data reinforce the significant effect that low level Pb exposure may have on gene-specific DNA methylation patterns in brain and that this occurs in a sex-dependent manner.

Keywords: Brain; DNA methylation; Developmental exposure; Epigenetics; Heavy metal; Hippocampus; Lead toxicity; Sex.

PubMed Disclaimer

Figures

Figure 1
Figure 1. Experimental design
Animals were exposed to 150 ppm, 375 ppm and 750 ppm lead (Pb) in food during different exposure periods: PERI (perinatal: dams exposed to Pb 10 days before breeding and continued through weaning (PND21)), EPN (early postnatal: dams were exposed to Pb at the time of parturition (PND0) and continued through weaning (PND21)), and LPN (long-term postnatal: Pb exposure started at the time of parturition (PND0) and continued through PND55). All pups were euthanized at PND55.
Figure 2
Figure 2. RDA analysis
(A–D) RDA plots for the covariable sex (A), Pb exposure level (B) and developmental period of exposure (C) are shown depicting a global view of the relationships between the methylation profiles and experimental conditions. Each colored dot represents a probe. RDA analysis showed that compared to exposure level and developmental period of exposure, sex was the major driver of the observed methylation changes in the hippocampus. The sex covariable is strongly associated with the RDA1 axis but exposure levels and exposure periods are not clearly associated with the RDA2 axis. In the RDA plot (D), each red dot represents a probe and the blue arrows represent covariables. This plot shows that the sex effect is associated with the RDA1 axis (sex effect) and exposure levels (150ppm and 375ppm) are associated with the RDA2 axis (exposure level effect). The two exposure levels were found to be anti-correlated (i.e., effects in opposite directions). The length of the arrow reflects the size of the effect of the covariable.
Figure 3
Figure 3. Representative probe heat maps for LPN 150 ppm males and females
The heat maps show the methylation log2 ratios for the 50 top-most significant (by p-value) probes in females (left) and males (right) in the long-term postnatal (LPN), 150 ppm exposure groups (compared to untreated controls). Levels 0 to −3 indicate hypomethylation and levels 0 to 3 indicate hypermethylation. The effects in males and females are for the most part in opposite directions.
Figure 4
Figure 4. Gene Ontology (GO) enrichment analysis
The most significantly enriched biological processes categories within hypermethylated genes (black bars) and hypomethylated genes (grey bars) are shown for comparison groups: LPN Female vs Control Female, 150 ppm Pb (A); PERI Female vs Control Female, 375 ppm Pb (B); LPN Male vs Control Male, 150 ppm (C); LPN Male vs Control Male, 750 ppm (D). In these groups significantly overrepresented biological functions composed of minimum of 5 genes at p<0.05 were found.
Figure 5
Figure 5. Relative expression of genes exhibiting hypermethylation and hypomethylation at Pb exposure levels 150 ppm and 375 ppm in LPN Females and Males
(A) The list of genes selected based on significant methylation change for expression level analysis. (B) The mRNA expression levels were normalized to Gapdh and were expressed as relative to controls (no Pb). Values represent mean ± SEM from N of 6–8; asterisks denote statistically significant change compared to control (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001).
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. ACCLPP. Low level lead exposure harms children: A renewed call for primary prevention 2012
    1. Adriaens ME, Jaillard M, Eijssen LM, Mayer CD, Evelo CT. An evaluation of two-channel ChIP-on-chip and DNA methylation microarray normalization strategies. BMC Genomics. 2012;13:42. - PMC - PubMed
    1. Anderson DW, Mettil W, Schneider JS. Effects of low level lead exposure on associative learning and memory in the rat: Influences of sex and developmental timing of exposure. Toxicol Lett. 2016;246:57–64. - PMC - PubMed
    1. Bock C, Beerman I, Lien WH, Smith ZD, Gu H, Boyle P, Gnirke A, Fuchs E, Rossi DJ, Meissner A. DNA methylation dynamics during in vivo differentiation of blood and skin stem cells. Mol Cell. 2012;47(4):633–47. - PMC - PubMed
    1. Boellmann F, Zhang L, Clewell HJ, Schroth GP, Kenyon EM, Andersen ME, Thomas RS. Genome-wide analysis of DNA methylation and gene expression changes in the mouse lung following subchronic arsenate exposure. Toxicol Sci. 2010;117(2):404–17. - PubMed