Sex and Exposure to Postnatal Chlorpyrifos Influence the Epigenetics of Feeding-Related Genes in a Transgenic APOE Mouse Model: Long-Term Implications on Body Weight after a High-Fat Diet

Abstract

Developmental exposure to toxicants and diet can interact with an individual's genetics and produce long-lasting metabolic adaptations. The different isoforms of the apolipoprotein E (APOE) are an important source of variability in metabolic disorders and influence the response to the pesticide chlorpyrifos (CPF). We aimed to study the epigenetic regulation on feeding control genes and the influence of postnatal CPF exposure, APOE genotype, and sex, and how these modifications impact on the metabolic response to a high-fat diet (HFD). Both male and female apoE3- and apoE4-TR mice were exposed to CPF on postnatal days 10-15. The DNA methylation pattern of proopiomelanocortin, neuropeptide Y, leptin receptor, and insulin-like growth factor 2 was studied in the hypothalamus. At adulthood, the mice were given a HFD for eight weeks. The results highlight the importance of sex in the epigenetic regulation and the implication of CPF treatment and APOE genotype. The body weight progression exhibited sex-dimorphic differences, apoE4-TR males being the most susceptible to the effects induced by CPF and HFD. Overall, these results underscore the pivotal role of sex, APOE genotype, and developmental exposure to CPF on subsequent metabolic disturbances later in life and show that sex is a key variable in epigenetic regulation.

Keywords: APOE; chlorpyrifos; epigenetics; feeding control; high-fat diet.

Figure 1

Experimental timeline including the treatment period from postnatal days (PND) 10 to 15 and sacrifice to obtain biological samples at PND 45. At postnatal month (PNM) 3, the animals were subjected to the high-fat diet (HFD) until PNM 5. Abbreviations: CPF: chlorpyrifos.

Figure 2

Pomc gene methylation at the CpG positions studied in the target zone in the promoter and overall methylation levels, which are sharing the same y axis. Below, there is the graphical representation of the CpG positions studied and the exact sequence of the target region (A). Methylation at the specific position −134 (B) and gene expression of the Pomc gene (C). All groups included 6 animals except for the APOE3-CNT that included 5. An asterisk (*) indicates significant differences between groups at p < 0.05. Abbreviations: CNT, control; CPF, chlorpyrifos-treated; ttm, treatment. Data is contained within the Supplementary File S1.

Figure 3

Npy gene methylation at the CpG positions studied in the target zone in the promoter and overall methylation levels, which are sharing the same y axis. Below, there is the graphical representation of the CpG positions studied and the exact sequence of the target region (A). Gene expression of the Npy gene (B). All groups included 6 animals except for the APOE3-CNT that included 5. An asterisk (*) indicates significant differences between groups at p < 0.05. Abbreviations: CNT, control; CPF, chlorpyrifos-treated. Data is contained within the Supplementary File S1.

Figure 4

Lepr gene methylation at the CpG positions studied in the target zone in the promoter and overall methylation levels, which are sharing the same y axis. Below, there is the graphical representation of the CpG positions studied and the exact sequence of the target region (A). Methylation at the specific position −40 (B) and gene expression of the Lepr gene (C). All groups included 6 animals except for the APOE3-CNT that included 5. An asterisk (*) indicates significant differences between groups at p < 0.05. Abbreviations: CNT, control; CPF, chlorpyrifos-treated. Data is contained within the Supplementary File S1.

Figure 5

Igf2 gene methylation at the CpG positions studied in the target zone in the DMR2 and overall methylation levels, which are sharing the same y axis. Below, there is the graphical representation of the CpG positions studied and the exact sequence of the target region (A). Methylation at the specific position +12395 (B) and gene expression of the Igf2 gene (C). All groups included 6 animals except for the APOE3-CNT that included 5. An asterisk (*) indicates significant differences between groups at p < 0.05. Different letters (a, b) represent significant differences at p < 0.05. Abbreviations: CNT, control; CPF, chlorpyrifos-treated; ttm, treatment. Data is contained within the Supplementary File S1.

Figure 6

Body weight progression during the postnatal period and the high-fat diet for APOE3 males (A), APOE3 females (B), APOE4 males (C), and APOE4 females (D), with the BW of week 0, week 4 and week 8 described in the corresponding table. The symbol $ indicates a significant effect of the diet at p < 0.05, while the symbol # represents a significant effect of the treatment at p < 0.05. Different letters (a, b) represent significant differences at p < 0.05. Abbreviations: CNT, control; CPF, chlorpyrifos-treated; HFD, high-fat diet. Data is contained within the Supplementary File S1.