The Role of Abnormal Methylation of Wnt5a Gene Promoter Regions in Human Epithelial Ovarian Cancer: A Clinical and Experimental Study

Abstract EN

Objective.

In the current study, the role of abnormal methylation of Wnt5a gene promoter regions in human epithelial ovarian cancer was investigated.

Methods.

Wnt5a expressions were examined by immunohistochemistry in epithelial ovarian tissues (30 normal and 79 human EOC tissues).

SKOV3 cells were treated with different concentrations of 5-Aza-CdR (0.5, 5, and 50 μmol/L).

The methylation status of the Wnt5a promoter was analyzed using a methylation-specific polymerase chain reaction (MSP), and the expression level of Wnt5a mRNA was detected using quantitative real-time polymerase chain reaction (qRT-PCR).

Cell proliferation was measured by MTT assay, and apoptosis was analyzed using flow cytometry.

Results.

(1) Compared with normal tissues, Wnt5a expressions were reduced or lost in EOC (P<0.05).

Wnt5a expression had a close relationship with histological grade, FIGO stage, and lymph node metastasis (P=0.005, P=0.022, and P=0.037, resp.).

(2) Wnt5a abnormal methylation status existed in ovarian cancer tissues and was higher than that of normal ovarian tissue (P<0.01).

(3) Before treatment with 5-Aza-CdR, the promoter of the Wnt5a gene was methylated in SKOV3 cells; accordingly, Wnt5a mRNA levels were low to absent in SKOV3 cells.

(4) Following 5-Aza-CdR treatment, MSP analysis revealed complete demethylation of the Wnt5a promoter in the SKOV3 cell line, particularly at 5 μmol/L 5-Aza-CdR.

Wnt5a expression increased in SKOV3 cells following treatment with a demethylating agent (P≤0.001).

(5) The growth rate of the cells was inhibited in a dose-dependent manner by treatment with 5-Aza-CdR.

(6) The cell apoptosis rate increased gradually after treatment with 0.5, 5, and 50 μmol/L 5-Aza-CdR.

The apoptosis rate exists in a dose-dependent relationship with 5-Aza-CdR concentration (F=779.73, P<0.01).

Conclusions.

Wnt5a gene region promoter aberrant methylation existed in epithelial ovarian cancer, and abnormal methylation of Wnt5a gene promoter regions may be a new target for the treatment of epithelial ovarian cancer.