So Yeon Yu

Genome-wide profiling in melatonin-exposed human breast cancer cell lines identifies differentially methylated genes involved in the anticancer effect of melatonin

Epigenetic alterations have emerged as an important mechanism involved in tumorigenesis. The epigenetic impact of DNA methylation in various types of human cancer is not completely understood. Previously, we observed melatonin‐induced differential expression of miRNA and miRNA‐related genes in human breast cancer cell lines that indicated an anticancer effect of melatonin. In this report, we further characterized epigenetic changes in melatonin‐exposed MCF‐7 cells through the analysis of DNA methylation profiles in breast cancer cells to provide new insights into the potential mechanisms of the anticancer effect of melatonin. Microarray‐based DNA methylation and gene expression profiling were carried out using human breast cancer cell lines. We further identified a number of mRNAs whose expression levels show an inverse correlation with DNA methylation levels. The mRNA expression levels and methylation status of candidate genes in melatonin‐exposed cells were confirmed by real‐time quantitative PCR and bisulfite PCR. This approach led to the detection of cancer‐related genes, which were oncogenic genes, including EGR3 and POU4F2/Brn‐3b were down‐regulated, while the tumor suppressor gene, GPC3, was up‐regulated by 1 nm melatonin‐treated MCF‐7 cells. Our results provide detailed insights into the DNA methylation patterns induced by melatonin and suggest a potential mechanism of the anticancer effect of aberrant DNA methylation in melatonin‐treated breast cancer cells.

Gene expression profiling of HepG2 cells treated with endocrine disrupting chemicals using the HazChem human array V3

The endocrine system is a system of glands that secrete hormones to regulate the body. This system is disrupted by endocrine disrupting chemicals (EDCs), which are similar to sex hormones. These chemicals function as androgen antagonists or estrogen agonists. To determine the effect of EDCs on the gene expression profile in human cells, we treated HepG2 cells with 3 chemicals (bisphenol A, 17β-estradiol, vinclozolin), and analyzed common gene expression using a custom-made HazChem human array V3. The Haz-Chem human array V3 included a total of 1136 genes, all of which were differentially expressed by exposure to VOCs, PAHs, POPs, and LTCs. Of these genes, 24 genes were commonly expressed after exposure to all 3 chemicals, based on the SAM (Significant Analysis of Microarray) method (q-value < 0.5%). These genes were analyzed further and found to be involved in coagulation, hemostasis, wound healing, angiogenesis, homeostasis, cell redox homeostasis, and cell proliferation based on GO function and pathway networks.

Environmental risk assessment of toxicity exposure: High-throughput expression profiling

Recently, volatile organic compounds (VOCs) in outdoor air pollution have led to major public health problems and the identification of specific minimally invasive biomarkers for assessing environmental toxicant exposure has become increasingly important. However, research into the human health effects of inhalation exposure to VOCs remains insufficient. Using a microarray based approach, we identified and validated characteristic mRNA expression profiles in the human whole blood of workers exposed to VOCs (toluene, ethylbenzene, and xylene), which were then compared with genomic level expressions in workers not exposed to the toxicants. We surveyed 141 workers working in a chemical production factory, of which 66 were not exposed to VOCs. We identified 4384 characteristic discernible exposure indicator mRNAs for toluene, 1296 for ethylbenzene, and 5821 for xylene. Using these, we were able to discern those subjects from the control group to a higher accuracy, sensitivity, and specificity than when using urinary biomarkers. The results showed that altered levels of mRNA can be a reliable, novel, and minimally invasive biological indicator of occupational exposure to VOCs. Future research directions should consider the adverse effects of exposure to VOCs on epigenetic regulation.

Integrative analyses of differential gene expression and DNA methylation of ethylbenzene-exposed workers

Humans can absorb volatile organic compounds (VOCs), such as ethylbenzene (EB), via inhalation resulting in various effects such as hepatotoxicity and even carcinogenicity. Because occupational EB exposure occurs frequently in the synthetic rubber and plastic industry, its toxicity has been previously reported by various in vitro as well as in vivo animal studies. EB has also been classified as a group 2B possible carcinogen by the IARC. However, these studies investigated the toxicity of EB used a much higher dose than expected occupational exposure. Thus the results are likely to be far away from to be the basis for the validation of the toxicity of the EB exposure. Because the previous studies examine only phenotypes, it is still unclear what and how EB affects physiological pathways in the human body. Using microarray platform, we investigated the gene expression profiles and genomic methylation patterns of 66 human blood samples from employees of local plants that use EB. The affected genes were functionally analyzed using the DAVID Tool and IPA. The methylation array demonstrated that, compared to the control group, a total of 1446 genes were hypermethylated and 60 genes were hypomethylated. Among the 378 differentially expressed genes, 12 had proper epigenetic correlation with the altered methylation patterns. Our study demonstrated how EB exposure affects the human genome by investigating the gene expression profiles and DNA methylation profiles of blood cells. We expect that our results will help establish the genomic basis of the effect of EB exposure on humans.

Innate immune response gene expression profiles in specific pathogen-free chickens infected with avian influenza virus subtype H9N2

In poultry species, H9N2 low pathogenic avian influenza (LPAI) virus infections frequently lead to low to moderate mortality rates and morbidity characterized by depression, respiratory disease symptoms, and decreases in egg production. Since current knowledge on the avian immune response to H9N2 infection is limited, we used microarray analysis to examine global changes in immune-related gene expression induced by H9N2 LPAI infection in specific pathogen-free chickens. The expression profiles of approximately 800 genes, including those that influence cell differentiation, transcription, transport, immune responses, and signal transduction, were altered by H9N2 infection. A complete chicken genome microarray analysis identified a strong innate antiviral host response after infection in spleens. In particular, a significant number of immune response genes, including interferon genes and related immune response genes, were upregulated following H9N2 infection. The increased transcription of 2′-5′-oligoadenylate synthetase and myxovirus resistance genes was confirmed by real-time RT-PCR. These results suggest that the host response generated against H9N2 infection may contribute to protective effects by manipulating innate immune responses.

Application of the emerging technologies in toxicogenomics: An overview

Toxicogenomics is a rapidly developing area of research which combines the understanding of molecular and cellular regulation in response to interaction of chemical stressors. Moreover, the emergence of comprehensive research method covering genomic, proteomic, and metabonomics analysis including bioinformatics has further substantiated the field of toxicogenomics. Thus, evaluation of such integrated analysis, conversing from variety of experimental techniques, along with the resulting database can decipher the importantance of such techniques in toxicogenomics. In addition, recent integration of epigenetic study (DNA Methylation, histone modification, small and noncoding RNAs) with toxicogenomics can pave the path to elucidate, highly reliable research results. The objective of the present review is to expand these understandings of toxicogenomics approach and validate the contribution of the developing research methods (genome, proteome, metabolites) to emphasize its importance in future toxicogenomics research.

Analysis of toxicity of tetrabutyltin: comparing with EDC chemicals

Exposure to environmental chemicals has been implicated in a number of adverse health effects, including immunotoxic, neurotoxic, reproductive toxic effects, and, above all, cancers. In the present study, we investigated toxicity of three environmental chemicals-bisphenol A (BPA), nonylphenol (NP), and tetrabutyltin (TTBT) in a human breast cancer cell line by HazChem Human array. We then found toxicity markers of TTBT through comparison with BPA and NP. Fifty three genes were differentially expressed and seventeen genes showed similar expression patterns. Ontology analysis showed differential expression of genes related to aging and apoptosis and genes having similarities associated with pregnancy and insemination. In this paper, we revealed potential TTBT toxic markers and endotoxicity of TTBT. This study could be a foundation for additional studies of TTBT toxicity.

Identification of time-dependent biomarkers and effects of exposure to volatile organic compounds using high-throughput analysis.

Volatile organic compounds (VOCs) can be easily taken up by humans, leading to various diseases, such as respiratory system and central nervous system disorders. Environmental risk assessment is generally conducted using traditional tests, which may be time‐consuming and technically challenging. Therefore, analysis of the effects of VOCs, such as toluene, ethylbenzene, and xylene, may be improved by use of novel, high‐throughput methods, such as microarray analysis. In this study, we examined the effects of VOCs exposure in humans on gene expression and methylation using microarray analysis. We recruited participants who had short‐term exposure, long‐term exposure, or no exposure. We then analyzed changes in gene expression in blood samples from these participants. A total of 866 genes were upregulated, while 366 genes were downregulated in the short‐term exposure group. Similarly, in the long‐term exposure group, a total of 852 and 480 genes were up‐ or downregulated, respectively. Hierarchical clustering analysis was used to divide the clustered genes into nine clusters to investigate the expression of variations in accordance with the exposure period. And the methylation microarray was performed at the same time to see whether this expression variation is related to the epigenetic study. Finally, we have 5 genes that were upregulated and 12 genes that were downregulated, gradually and respectively, so these genes are expected to function as biomarkers of the duration of exposure to VOCs. Further research is required to determine the time‐dependent effects of VOCs on epigenetic regulation of gene expression.

Identification of potential biomarkers for xylene exposure by microarray analyses of gene expression and methylation

Xylene is volatile organic compound that has been reported to increase the incidence of cancer and various diseases related to the immune system, cardiovascular systems, respiratory and reproductive organs. However, there are currently few biomarkers in human cases. Using microarray, we analysed 10 participants for the xylene-exposure group and 10 controls that were not exposed to xylene. The two groups were compared in terms of expression levels and methylation patterns. We identified 6 genes that were down-regulated and hyper-methylated, and 132 that were up-regulated and hypo-methylated in the xylene- exposure group compared to control. We sorted out and 28 biomarker candidates were chosen using DAVID. And then, we used IPA to select the significant potential biomarkers in them. We used network analysis and selected 6 significant genes, and these 6 genes showed altered expression and methylation in xylene-exposure group, suggesting that they are suitable potential biomarkers for xylene exposure.

Association analysis of toluene exposure time with high-throughput mRNA expressions and methylation patterns using in vivo samples.

The emission of volatile organic compounds (VOCs) resulting from outdoor air pollution can contribute to major public health problems. However, there has been limited research on the health effects in humans from the inhalation of VOCs. Therefore, this study conducted an in vivo analysis of the effects of toluene, one of the most commonly used chemicals in many industries, on gene expression and methylation over time using the high-throughput technique of microarray analysis. We separated participants into three groups (control, short-term exposure, and long-term exposure) to investigate the influence of toluene exposure time on gene expression. We then comprehensively analyzed and investigated the correlation between variations in gene expression and the occurrence of methylation. Twenty-six genes were upregulated and hypomethylated, while 32 genes were downregulated and hypermethylated. The pathways of these genes were confirmed to be associated with cell survival and the immune system. Based on our findings, these genes can help predict the effects of time-dependent exposure to toluene on human health. Thus, observations from our data may have implications for the identification of biomarkers of toluene exposure.