Table9_Effects of Prenatal Exposure to Titanium Dioxide Nanoparticles on DNA Methylation and Gene Expression Profile in the Mouse Brain.pdf

Background and Objectives: Titanium dioxide nanoparticles (TiO2-NP) are important materials used in commercial practice. Reportedly, TiO2-NP exposure during pregnancy can affect the development of the central nervous system in mouse offspring; however, the underlying mechanism remains unknown. In the present study, we investigated the impact of prenatal TiO2-NP exposure on global DNA methylation and mRNA expression patterns in the brains of neonatal mice.
Materials and Methods: Pregnant C57BL/6J mice were intratracheally administered a TiO2-NP suspension (100 μg/mouse) on gestational day 10.5, and brains were collected from male and female offspring at day 1 postpartum. After extraction of methylated DNA by immunoprecipitation, the DNA methylation profile was analyzed using a mouse CpG island microarray. Total RNA was obtained, and mRNA expression profiles were comprehensively assessed using microarray analysis.
Results: Among genes in the CpG island microarray, DNA methylation was increased in 614 and 2,924 genes and decreased in 6,220 and 6,477 genes in male and female offspring, respectively. Combined with mRNA microarray analysis, 88 and 89 genes were upregulated (≥1.5-fold) accompanied by demethylation of CpG islands, whereas 13 and 33 genes were downregulated (≤0.67-fold) accompanied by methylation of CpG islands in male and female offspring mice, respectively. Gene Set Enrichment Analysis (GSEA) revealed that these genes were enriched in gene ontology terms related to the regulation of transcription factors, cell proliferation, and organism development. Additionally, MeSH terms related to stem cells and morphogenesis were enriched.
Conclusion: Prenatal TiO2-NP exposure induced genome-wide alterations in DNA methylation and mRNA expression in the brains of male and female offspring. Based on GSEA findings, it can be speculated that prenatal TiO2-NP exposure causes adverse effects on brain functions by altering the DNA methylation state of the fetal brain, especially neural stem cells, resulting in the subsequent abnormal regulation of transcription factors that modulate development and differentiation.

背景与目的：二氧化钛纳米颗粒（Titanium dioxide nanoparticles, TiO₂-NP）是商业化应用的重要材料。已有研究表明，妊娠期暴露于TiO₂-NP可影响小鼠子代的中枢神经系统发育，但其潜在分子机制尚未明确。本研究旨在探讨产前暴露于TiO₂-NP对新生小鼠大脑全基因组DNA甲基化及mRNA表达谱的影响。
材料与方法：于妊娠第10.5天，对妊娠C57BL/6J小鼠经气管内给予TiO₂-NP混悬液（剂量为100 μg/只）；并于产后第1天收集雌雄子代的脑组织。通过免疫沉淀法提取甲基化DNA后，采用小鼠CpG岛芯片分析DNA甲基化谱。提取总RNA后，利用芯片技术全面评估mRNA表达谱。
结果：在CpG岛芯片检测的基因中，雄性子代分别有614个基因的DNA甲基化水平升高、2924个基因的DNA甲基化水平降低，雌性子代则分别有6220个基因的DNA甲基化水平升高、6477个基因的DNA甲基化水平降低。结合mRNA芯片分析结果，雄性子代中共鉴定出88个伴随CpG岛去甲基化的上调基因（表达量≥1.5倍），雌性子代则有89个；而伴随CpG岛甲基化的下调基因（表达量≤0.67倍）在雄性子代中为13个，雌性子代中为33个。基因集富集分析（Gene Set Enrichment Analysis, GSEA）显示，上述基因富集于与转录因子调控、细胞增殖及机体发育相关的基因本体术语。此外，还富集到与干细胞及形态发生相关的医学主题词表（MeSH）术语。
结论：产前暴露于TiO₂-NP可诱导雌雄子代大脑发生全基因组范围的DNA甲基化与mRNA表达改变。基于GSEA分析结果，可推测产前TiO₂-NP暴露通过改变胎脑（尤其是神经干细胞）的DNA甲基化状态，进而异常调控参与发育与分化的转录因子，最终对脑功能产生不良影响。