ESPU Congress 2018 - Abstract Book

36 29 th CONGRESS OF THE ESPU 15:38–15:41 S3-2 (PP) THE EFFECT OF XENOESTROGENS ON FORESKIN FIBROBLASTS AND THE ETIOLOGY OF HYPOSPADIAS Karen AITKEN  1 , Jia-Xin JIANG  2 , Shreya BATRA  2 , Sevan HOPYAN  2 and Darius BAGLI  2 1) PGCRL room 159420TUV, DSCB, Sickkids Research Institute, Toronto, CANADA - 2) Hospital for Sick Children, Developmental and Stem Cell Biology, Research Institute, Toronto, CANADA PURPOSE Hypospadias is a developmental defect in males where the development of the urethra foreskin and ventricle surface of the penis is altered. Although the etiology of hypospadias is still debated, it has been suggested that hypospadias is linked to environmentally altered gene expression. Previous studies have found that in utero exposure to xenoestrogens (XE) underlying many congenital anom- alies also alters the development of the genital tubercle (GT) through developmentally important genes wnt5a and hoxa13. We hypothesize that expression of hoxa13 and wnt5a are rescued in the presence of a methylase inhibitors indicating that XE mediation of GT is under epigenetic regulation. MATERIAL AND METHODS Human foreskin fibroblast cell line (BJ cells) were maintained in EMEM, 10 % heat-treated charcoal- stripped (hormone-free) fetal calf serum in 37°C, 5 % CO2 incubator. For exposure to XE, cells were starved of serum for 24–48 hours, and exposed to Diethylstilbestrol for 0–120 hours. EZH (methylase) inhibitor (UNC 1999) or DNAmethylation inhibitor (decitibine) were introduced to the BJ cells for 48–120 hours. Cells were harvested and real time PCR was performed using SYBR-Green Master Mix. Relative levels of hoxa13 and wnt5a was determined with GAPDH used to normalize the findings. p<0.05 was considered to be significant. RESULTS Exposure to XE decreased hoxa13 and wnt5a expression in human foreskin fibroblasts. After the introduction of UNC1999, the expression of hoxa13 at day 6 was rescued, p<0.05. Wnt5a expres- sion was partially rescued by DNA methylation inhibition, p<0.05. CONCLUSIONS XE have the potential to affect genital tubercle development through epigenetic mechanisms, though the mechanisms may differ depending on the genes. Further work is underway to uncover the regulation of wnt5 and hoxa13 by histone and DNA methylation profiles using ChIP-PCR and pyrosequencing.