407033

MIR30D

OBJECTIVE: MicroRNAs (miRs) are essential regulators of gene expression by suppressing translation or causing degradation of target mRNA. Growing evidence sheds light on the crucial roles of miR dysregulation in cancer development and progression. In this study, we focused on the role of miR-30d in transforming growth factor beta1 (TGF-beta1)-initiated epithelial-mesenchymal transition (EMT) in ovarian cancer cells. METHODS: Transforming growth factor beta1 (10 ng/mL) was used to initiate EMT in SKOV3 and 3AO cells. The expression of miR-30 family members was determined by quantitative real-time polymerase chain reaction. Messenger RNA and protein levels of E-cadherin, N-cadherin, vimentin, and Snail were detected by quantitative real-time polymerase chain reaction and Western blot, respectively. Cell migration and invasion capacities were evaluated by Transwell chamber assay. Luciferase activity assay was performed to verify the direct inhibition of Snail by miR-30d. RESULTS: MiR-30b, MiR-30c, and MiR-30d were down-regulated during TGF-beta1-induced EMT in SKOV3 and 3AO ovarian cancer cells. Restoration of miR-30d by miR-30d mimic reversed TGF-beta1-induced EMT phenotypes including the morphological changes, expression pattern of molecular markers (E-cadherin, N-cadherin), and migratory and invasive capabilities in ovarian cancer cells. Furthermore, Snail was identified as the direct target of miR-30d. CONCLUSIONS: Our results revealed that miR-30d functioned as a suppressor of ovarian cancer progression by decreasing Snail expression and thus blocking TGF-beta1-induced EMT process, suggesting the potentiality of miR-30d analogs to be used as therapeutics for ovarian cancer.

BACKGROUND: Abnormal DNA methylation/demethylation is recognized as a hallmark of cancer. TET (ten-eleven translocation) family members are novel DNA demethylation related proteins that dysregulate in multiple malignances. However, their effects on ovarian cancer remain to be elucidated. METHODS: The changes of TET family members during TGF-beta1-induced epithelial-mesenchymal transition (EMT) in SKOV3 and 3AO ovarian cancer cells were detected. TET3 was ectopically expressed in TGF-beta1-treated ovarian cancer cells to examine its effect on TGF-beta1-induced EMT phenotype. The downstream target of TET3 was further identified. Finally, the relationships of TET3 expression to clinic-pathological parameters of ovarian cancer were investigated with a tissue microarray using immunohistochemistry. RESULTS: TET3 was downregulated during TGF-beta1-initiatd epithelial-mesenchymal transition (EMT) in SKOV3 and 3AO ovarian cancer cells. Overexpression of TET3 reversed TGF-beta1-induced EMT phenotypes including the expression pattern of molecular markers (E-cadherin, Vimentin, N-cadherin, Snail) and migratory and invasive capabilities of ovarian cancer cells. miR-30d was identified as a downstream target of TET3, and TET3 overexpression resumed the demethylation status in the promoter region of miR-30d precursor gene, resulting in restoration of miR-30d (an EMT suppressor of ovarian cancer cells proven in our previous study) level in TGF-beta1-induced EMT. We further found that TET3 expression was decreased in ovarian cancer tissues, especially in serous ovarian cancers. The overall positivity of TET3 was inversely correlated with the grade of differentiation status of ovarian cancer. CONCLUSION: Our results revealed that TET3 acted as a suppressor of ovarian cancer by demethylating miR-30d precursor gene promoter to block TGF-beta1-induced EMT.