ABSTRACT
Increasing evidence has suggested that dysregulation of microRNAs (miRNAs) could contribute to tumor progression. The miR-34 family is directly transactivated by tumor suppressor p53 which is frequently mutated in various cancers; however, the effect of miR-34a on the ovarian cancer cells remains unclear. The aim of the paper was to study the expression of miR-34a in ovarian cancer and miR-34a's relation to the cell proliferation and metastasis in ovarian cancer in vitro. miR-34a expression was determined by quantitative RT-PCR in a panel of 60 human ovarian cancer samples. Functional characterization of miR-34a was accomplished by reconstitution of miR-34a expression in ovarian cancer cells by determining changes in proliferation, migration, and invasion. Our results showed that miR-34a is downregulated in ovarian cancer tissues compared with the corresponding adjacent non-neoplastic tissues, and the expression level of miR-34a was significantly lower in ovarian cancer cell lines in comparison with normal human fallopian tube epithelial cell line. The 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-diphenytetrazoliumromide (MTT) assay revealed significant cell proliferation inhibition in miR-34a transfectant compared with the control from HO8910 and SKOV3 cells, which displayed lowest expressions of miR-34a. Furthermore, the transwell assay also showed significant cell migration inhibition in miR-34a transfectant, compared with cell lines transfected with NC. Overexpression of miR-34a led to the inhibition of AXL expression, indicating that AXL is a target gene for miR-34a. Our data suggest that miR-34a may function as a tumor suppressor through repression of oncogenic AXL in ovarian cancer.
Subject(s)
MicroRNAs/genetics , Ovarian Neoplasms/genetics , Proto-Oncogene Proteins/genetics , Receptor Protein-Tyrosine Kinases/genetics , Tumor Suppressor Protein p53/genetics , Apoptosis/genetics , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Fallopian Tubes/cytology , Fallopian Tubes/metabolism , Female , Gene Expression Regulation, Neoplastic , Humans , MicroRNAs/biosynthesis , Neoplasm Invasiveness/genetics , Neoplasm Metastasis , Ovarian Neoplasms/pathology , Proto-Oncogene Proteins/antagonists & inhibitors , Proto-Oncogene Proteins/biosynthesis , Receptor Protein-Tyrosine Kinases/antagonists & inhibitors , Receptor Protein-Tyrosine Kinases/biosynthesis , Axl Receptor Tyrosine KinaseABSTRACT
OBJECTIVE: The effect of miR-449 and miR-34 on the growth, cell cycle and target gene expressions of ovarian cancer cell line SKOV3 and SKOV3-ipl was discussed. METHOD: Real-time quantitative reverse transcription PCR was employed to detect the expressions of miR-449a/b and miR-34b, c in SKOV3 and SKOV3-ipl cells. The two miRNAs were successfully expressed in SKOV3-ipl cells by transfection. The variations in cell growth rate and cell cycle were determined by MTS assay and flow cytometry, respectively. The expressions of cell cycle-related proteins were detected by Western Blot. RESULTS: miR-449b and miR-34c induced the decline of the adhesiveness of SKOV3-ipl cells by 20%-30%. The number of cells arrested in G1-phase increased and the number of cells arrested in S-phase decreased significantly. The cell cycle-related proteins CDK6 and CDC254 were downregulated. miR-449b caused the expression of CDK6 and CDC25A to decrease. After the co-transfection with miR-449b and miR-34c, the relevant proteins were downregulated more significantly. The expressions of CDK6, CDC25A and cyclin A were decreased significantly. CONCLUSION: miR-449b and miR-34c can induce cell cycle arrest in SKOV3-ipl cells and the downregulation of CDK6, CDC25A and cyclin A.
