DNA methylation analysis in rat kidney epithelial cells exposed to 3-MCPD and glycidol

3-Monochloropropane-1,2-diol (3-MCPD) is a well-known food processing contaminant that has been regarded as a rat carcinogen, which is known to induce Leydig-cell and mammary gland tumors in males, as well as kidney tumors in both genders. 3-MCPD is highly suspected to be a non-genotoxic carcinogen. 2,3-Epoxy-1-propanol (glycidol) can be formed via dehalogenation from 3-MCPD. We aimed to investigate the cytotoxic effects of 3-MCPD and glycidol, then to demonstrate the possible epigenetic mechanisms with global and gene-specific DNA methylation in rat kidney epithelial cells (NRK-52E). IC50 value of 3-MCPD was determined as 48mM and 41.39 mM, whereas IC50 value of glycidol was 1.67mM and 1.13mM by MTT and NRU test, respectively. Decreased global DNA methylation at the concentrations of 100 mM and 1000 mu M for 3-MCPD and 100 mu M and 500 mu M for glycidol were observed after 48 h exposure by using 5-methylcytosine (5-mC) ELISA kit. Methylation changes were detected in promoter regions of c-myc and Rassf1a in 3-MCPD and glycidol treated NRK-52E cells by using methylation-specific PCR (MSP), whereas changes on gene expression of c-myc and Rassf1a were observed by using real-time PCR. However, e-cadherin, p16, VHL and p15 genes were unmethylated in their CpG promoter regions in response to treatment with 3-MCPD and glycidol. Alterations in DNA methylation might be key events in the toxicity of 3-MCPD and glycidol. Alterations in DNA methylation might be key events in the toxicity of 3-MCPD and glycidol

Investigation of the genotoxic and cytotoxic effects of widely used neonicotinoid insecticides in HepG2 and SH-SY5Y cells

Neonicotinoids are a relatively new type of insecticide to control a variety of pests. Although they are generally considered to be safe, they can lead to harmful effects on human and environmental health. We aimed to investigate possible effects of common neonicotinoid insecticides (acetamiprid, clothianidin, imidacloprid, thiacloprid, and thiamethoxam) on cytotoxicity and DNA damage in human neuroblastoma (SH-SY5Y) and human hepatocellular carcinoma (HepG2) cells. Our results indicated that 50% of inhibitory concentration values of neonicotinoids are in the range of 0.96 to >4 mM in SH-SY5Y cells and 0.53 to >4 mM in HepG2 cells by the methyl tetrazolium and neutral red uptake tests after 24 and 48 h exposure. We observed significant DNA damage at 500 mu M of five neonicotinoids in SHSY-5Y cells, while only imidacloprid, thiametoxam, and thiacloprid showed some alterations in HepG2 cells after 24 h exposure using the alkaline comet assay. In conclusion, neonicotinoid insecticides may induce cytotoxicity and DNA damage in cell cultures; therefore, further studies are needed to better understand the toxicity of neonicotinoids

Alteration on global and gene-spesific DNA methylation and global histone modifications in HepG2 cells in response to BPA

Bisphenol A (BPA), as synthetic monomer used in industry in the production of polycarbonate plastic and epoxy resins, has endocrine disruptor properties and high risk on human health. Continuous release of free BPA into food, beverages, and the environment has resulted in a widespread human exposure to this chemical. Recent studies have showed the role of endocrine effects of environmental chemicals on the changes in gene expression may be associated with epigenetic mechanisms such as DNA methylation and histone modifications. The aim of the study was to investigate dose-related effects of BPA (0, 0.1 mu M, 1 and 10 mM for 48 and 96 h) on global and gene-spesific (p16, cyclin D2 and Rassf1 genes) DNA methylation in human hepatocarcinoma (HepG2) cells. We also investigated global histone modifications such as H3K9 trimethylation (H3K9me3), H3K9 acetylation (H3K9ac), H3K27 trimethylation (H3K27me3). The 50% inhibitory concentration (IC50) value of BPA was determined as 134 and 180 mu M in HepG2 cells for 24 h by MTT and neutral red uptake (NRU) tests, respectively. We observed decrease on the global levels of 5-methylcytosine and 5-hydroxymethylcytosine at 1 and 10 mM after 96 h BPA exposure. There is no significant alterations on the promoter-region of methylation and expression of p16, cyclin D2 and Rassf1 genes. Global levels of H3K9me3 decreased after 0.1 and 1 mu M concentration of BPA exposure for 48 h, while increased after 96 h 0.1 and 1 mu M of BPA exposure for. Hovewer, it has not been observed significantly changes for the global levels of H3K27me3 and H3K9ac. In this study we suggest that BPA may disrupt epigenetic events by altering global and gene-spesific DNA methylation and histone modifications in HepG2 cells

Effects of BPA on global DNA methylation and global histone 3 lysine modifications in SH-SY5Y cells: An epigenetic mechanism linking the regulation of chromatin modifiying genes

Bisphenol A (BPA), an estrogenic endocrine disruptor, is widely used in the production of polycarbonate plastic and epoxy resins, resulting in high risk on human health. In present study we aimed to investigate the effects of BPA on global and gene specific DNA methylation, global histone modifications and regulation of chromatin modifiying enzymes in human neuroblastoma cells (SH-SY5Y). Cells were treated with BPA at 0.1, 1 and 10 mu M concentrations for 48 and 96h. IC50 value of BPA was determined as 183 and 129 mu M in SH-SY5Y cells after 24h by MTT and NRU tests, respectively. We observed significant alterations on the 5-mC% levels (1.3 fold) and 5-hmC% levels (1.67 fold) after 10 mu M of BPA for 96h. Significant decrease was identified in H3K9me3 and H3K9ac after 10 mu M of BPA for 96h while decrease was observed in H3K4me3 at 10 mu M of BPA for 48h. Alterations were observed in chromatin modifiying genes including G9a, EZH2, SETD8, SETD1A, HAT1, SIRT1, DNMT1, RIZ1 and Suv39h1 after 96h of BPA exposure. Taken together, this study suggests that BPA might modulate the epigenetic regulators which would be key molecular events in the toxicity of endocrine disrupting chemicals