Effects of triclosan exposure on placental extravillous trophoblast motility, relevant IGF2/H19 signaling and DNA methylation-related enzymes of HTR-8/SVneo cell line

Abstract

Triclosan (TCS) is an antimicrobial agent widely used in personal care products and a potential endocrine disruptor chemical (EDC). TCS can pass through the placental barrier. Any influence of EDCs on epigenetic changes of placenta and embryo may bring profound impact on later health. This study aimed to investigate the effects of TCS exposure on cell proliferation and migration, and the expression of imprinted genes IGF2/H19 and DNA methylation-related enzymes in human placental extravillous trophoblast cell line HTR-8/SVneo. After exposure to TCS levels of 0 (DMSO Control), 10−11, 10−10, 10−9, 10−8, 10−7, 10−6, 10−5, 3 × 10−5, 6 × 10−5, 10−4 M and incubated for up to 36 h, cell proliferation and migration were examined by CCK-8, EdU incorporation assay and wound healing assay; the mRNA levels of IGF2, H19, DNA methyltransferases (Dnmt3a, Dnmt3b and Dnmt1), ten-eleven translocation enzymes (Tet1, Tet2 and Tet3) and IGF2 Receptor (IGF2R) were analyzed by qRT-PCR. The protein levels of IGF2 were measured by Western blot and ELISA. The cell viability turned to decline at TCS treatment of 3 × 10−5 M and above (all p < 0.05, compared to the DMSO Control). The cell migration decreased at TCS 10−5 and 3 × 10−5 M treatment (p < 0.05), but consistently unchanged at low dose of TCS from 10−9 to 10−7 M (p > 0.05). TCS treatments below cytotoxicity doses (< 10−5 M) did not significantly alter the mRNA levels of IGF2, Dnmt1, Dnmt3a, Dnmt3b, Tet1, Tet2 and Tet3, compared to DMSO Control treatment (all p > 0.05). The transcription level of H19 was up-regulated by TCS at 3 × 10−5 M. TCS at 10−7 and 6 × 10−5 M increased the protein level of IGF2 in cell supernatant. Our data suggest that high TCS exposure may suppress HTR-8/SVneo cells viability and migration, increase H19 gene expressions and IGF2 protein secretion. The exact mechanism of TCS action in human trophoblast needs further studies.