Studies have shown that microRNA-148a (miR-148a) was proved to be silenced while DNA methyltransferase 1 (DNMT1) was over-expressed in gastric cancer. But the mechanism of aberrant expression of miR-148a and DNMT1 and their relationships in gastric cancer are still unknown. The aims of this study were to investigate the expression profile of miR-148a and DNMT1 and reveal whether they have any relationships. We used reverse-transcriptase quantitative real-time PCR, methylation-specific PCR and Western blot to measure the level of miR-148a expression, DNA methylation level and DNMT1 expression, respectively. Gastric cancer cells were transfected with plasmid or siRNA or treated with 5-aza-2'-deoxycytidine. Cell proliferation and apoptosis were detected by cell counting and flow cytometric analysis. In this study, we demonstrated that gastric cancer tissues and cell lines displayed a consistent down-regulation of miR-148a and hypermethylation of promoter region. DNMT1 was over-expressed in primary tumors and cell lines, while knockdown of DNMT1 using siRNA could decrease methylation level of miR-148a promoter and restore its expression. Furthermore, ectopic over-expression of miR-148a in cancer cell lines caused reduction in DNMT1 expression and inhibited cell proliferation, but no obvious change was found in apoptosis rate. These results suggest that miR-148a is inactivated by DNA hypermethylation of promoter region in gastric cancer, which is mediated through DNMT1 over-expression. Additionally, the silence of miR-148a reduces its suppression to DNMT1 in gastric cancer, and this may in turn result in over-expression of DNMT1 and promote DNA hypermethylation.

Abstract-Perfluorooctane sulfonate (PFOS) is a widespread and persistent chemical in the environment. We investigated the endocrine-disrupting effects of PFOS using a combination of in vitro and in vivo assays. Reporter gene assays were used to detect receptor-mediated (anti-)estrogenic, (anti-)androgenic, and (anti-)thyroid hormone activities. The effect of PFOS on steroidogenesis was assessed both at hormone levels in the supernatant and at expression levels of hormone-induced genes in the H295R cell. A zebrafishbased short-term screening method was developed to detect the effect of PFOS on endocrine function in vivo. The results indicate that PFOS can act as an estrogen receptor agonist and thyroid hormone receptor antagonist. Exposure to PFOS decreased supernatant testosterone (T), increased estradiol (E2) concentrations in H295R cell medium and altered the expression of several genes involved in steroidogenesis. In addition, PFOS increased early thyroid development gene (hhex and pax8) expression in a concentration-dependent manner, decreased steroidogenic enzyme gene (CYP17, CYP19a, CYP19b) expression, and changed the expression pattern of estrogen receptor production genes (esr1, esr2b) after 500 mg/L PFOS treatment in zebrafish embryos. These results indicate that PFOS has the ability to act as an endocrine disruptor both in vitro and in vivo by disrupting the function of nuclear hormone receptors, interfering with steroidogenesis, and altering the expression of endocrine-related genes in zebrafish embryo. Environ. Toxicol. Chem. 2013;32:353-360. # 2012 SETAC

Triclosan (TCS) and triclocarban (TCC), as broad spectrum antibacterial agents, are distributed widely in the environment and humans. Most studies have focused on their distribution and biodegradation, but the endocrine-disrupting effects of these chemicals, especially their estrogenic effects, are still unclear. In the present study, we investigated the estrogenic effects of TCS and TCC using a series of in vitro assays, including the ER reporter gene assay in the CV-1 cells, E-screen assay and evaluation of estrogen-responsive genes in the MCF-7 cells. The tested concentrations of TCS and TCC were both from 1 × 10(-9) to 1 × 10(-6)  M. Results showed that TCS and TCC exerted estrogenic activities by inducing luciferase activities in an ER reporter gene assay, promoting the proliferation of the MCF-7 cells, up-regulating the expression of pS2 and down-regulating ERα expression at both the mRNA and protein levels in the MCF-7 cells. We further found that TCS and TCC could alter the expression of multiple microRNAs (mir-22, mir-206 and mir-193b) in the MCF-7 cells, which would help understand the mechanisms of their estrogenic effects on regulating the expression of ERα. In brief, our results demonstrated the potential estrogenic effects and profiled in vitro data for further risk assessment of TCS and TCC.

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