Nanoscale titanium dioxide (TiO2) is manufactured in wide scale, with a range of applications in consumer products. Significant toxicity of TiO2 nanoparticles has, however, been recognized, suggesting considerable risk to human health. To evaluate fully their toxicity, assessment of the epigenetic action of these nanoparticles is critical. However, only few studies are available examining capability of nanoparticles to alter epigenetic integrity. In the present study, the effect of TiO2 nanoparticles exposure on DNA methylation, a major epigenetic mechanism, was investigated in in vitro cellular model systems. A panel of cells relevant to portals of human exposure (Caco-2 (colorectal), HepG2 (liver), NL20 (lung), and A-431 (skin)) was exposed to TiO2 nanoparticles to assess effects on global methylation, gene-specific methylation, and expression levels of DNA methyltransferases, MBD2, and UHRF1. Global methylation was determined by enzyme-linked immunosorbent assay-based immunochemical analysis. Degree of promoter methylation across a defined panel of genes was evaluated using EpiTect Methyl II Signature PCR System Array technology. Expression of DNMT1, DNMT3a, DNMT3b, MBD2, and URHF1 was quantified by qRT-PCR. Decrease in global DNA methylation in cell lines Caco-2, HepG2, and A-431 exposed to TiO2 nanoparticles was shown. Across four cell lines, eight genes (CDKN1A, DNAJC15, GADD45A, GDF15, INSIG1, SCARA3, TP53, and BNIP3) were identified in which promotors were methylated after exposure. Altered expression of these genes is associated with disease etiology. The results also revealed aberrant expression of epigenetic regulatory genes involved in DNA methylation (DNMT1, DNMT3a, DNMT3b, MBD2, and UHRF1) in TiO2 exposed cells, which was cell type dependent. Findings from this study clearly demonstrate the impact of TiO2 nanoparticles exposure on DNA methylation in multiple cell types, supporting potential involvement of this epigenetic mechanism in the toxicity of TiO2 nanoparticles. Hence for complete assessment of potential risk from nanoparticle exposure, epigenetic studies are critical.

中文翻译：

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二氧化钛纳米粒子对多种人类细胞系DNA甲基化的影响。

纳米级二氧化钛（TiO2）被大规模生产，在消费产品中具有一系列应用。然而，已经认识到TiO 2纳米颗粒的显着毒性，表明对人体健康具有相当大的风险。为了充分评估其毒性，评估这些纳米颗粒的表观遗传作用至关重要。然而，只有很少的研究可用于检查纳米粒子改变表观遗传完整性的能力。在本研究中，在体外细胞模型系统中研究了TiO2纳米颗粒暴露对DNA甲基化（一种主要的表观遗传机制）的影响。将一组与人类暴露的门相关的细胞（Caco-2（结肠直肠），HepG2（肝脏），NL20（肺）和A-431（皮肤））暴露于TiO2纳米颗粒，以评估其对总体甲基化的影响，特定的甲基化 DNA甲基转移酶，MBD2和UHRF1的表达水平。通过基于酶联免疫吸附测定的免疫化学分析确定总体甲基化。使用EpiTect Methyl II Signature PCR System Array技术评估了定义的一组基因中启动子的甲基化程度。通过qRT-PCR定量DNMT1，DNMT3a，DNMT3b，MBD2和URHF1的表达。显示了暴露于TiO2纳米粒子的细胞系Caco-2，HepG2和A-431中总体DNA甲基化的降低。在四个细胞系中，鉴定了八个基因（CDKN1A，DNAJC15，GADD45A，GDF15，INSIG1，SCARA3，TP53和BNIP3），其中启动子在暴露后被甲基化。这些基因的表达改变与疾病病因有关。结果还表明，与DNA甲基化有关的表观遗传调控基因（DNMT1，暴露于TiO2的细胞中的DNMT3a，DNMT3b，MBD2和UHRF1），这取决于细胞类型。这项研究的发现清楚地证明了TiO2纳米颗粒暴露对多种细胞类型的DNA甲基化的影响，支持了这种表观遗传机制潜在参与TiO2纳米颗粒的毒性。因此，对于完整评估纳米颗粒暴露的潜在风险，表观遗传学研究至关重要。