(W1302) EXPOSURE TO 2,3,7,8-TETRACHLOROBENZO-P-DIOXIN ALTERED THE DNA METHYLATION PATTERNS OF GERMLINE DEVELOPMENTAL GENES IN HUMAN EMBRYONIC STEM CELLS
The University of Hong Kong The University of Hong Kong, Hong Kong
Abstract: 2,3,7,8-tetrachlorobenzo-p-dioxin (TCDD) is an environmental toxin reported to be associated with many systematic diseases including cancers, dementia and reproductive defects. Previous studies revealed that TCDD exposure compromised ovarian function in adult mice. More importantly, TCDD exposure exhibited transgenerational effect on fetal sperm development in rats. Whether early TCDD exposure affected embryonic primordial germ cells (PGC) specification in humans is unclear. In this study, we hypothesized that TCDD exposure induced aberration of DNA methylation in early embryo, leading to dysregulated human PGC development. Human embryonic stem cell (hESC) is biologically equivalent to inner cell mass (ICM), the origin of human PGC. Hence, hESC line VAL3 was cultured with physiological doses of TCDD (10 and 100 pM), followed by genome-wide DNA methylome profiling by reduced representation bisulfite sequencing. The results showed that nearly 4000 CpG sites were differentially methylated in VAL3 upon TCDD exposure. Interestingly, Gene ontology analysis of the hypermethylated genes demonstrated enrichments of biological processes including “gamete generation”, “spermatid differentiation”, “germ cell development”, “multicellular organism reproduction” and “reproductive process in a multicellular organism”. Among them, Desert Hedgehog (DHH) and Cadherin EGF LAG Seven-Pass G-Type Receptor 2 (CELSR2), were selected for further validation because they have been reported to be important for male gamete development. Gene-specific bisulfite sequencing demonstrated that the average methylation rates of DHH and CELSR2 were significantly increased in VAL3 upon TCDD exposure. The results suggested a potential mechanism that TCDD might pose on germ cell specification during early embryonic development. The established TCDD-treated hESC lines in this study provided a powerful model to validate and study the relationship between TCDD exposure and germ cell specification.
