Molecular Mechanism of Mir-148A regulating cell adhesion molecule1 gene methylation by acting on DNA methyltransferase 1 in ovarian cancer
10.3760/cma.j.cn.115807-20200902-00266
- VernacularTitle:卵巢癌中miR-148a通过作用DNA甲基化转移酶1调控细胞黏附分子1基因甲基化的分子机制研究
- Author:
Gaoming ZHENG
1
;
Xiaoru SU
;
Lan LI
Author Information
1. 杭州师范大学附属医院检验科 310015
- Keywords:
Ovarian cancer;
miRNA-148a;
Cell adhesion molecule 1;
DNA methyltransferase 1
- From:
Chinese Journal of Endocrine Surgery
2021;15(5):509-513
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effects of cell adhesion molecule 1 (CADM1) promoter methylation in ovarian cancer on gene transcription and protein expression levels, and the regulation mechanism of mirNA-148A on CADM1 methylation levels.Methods:A total of 86 patients with ovarian cancer who received surgical treatment in the Affiliated Hospital of Hangzhou Normal University from Jun. 2018 to Jun. 2020 were selected as study subjects. The methylation level of CADM1 gene CpG island in ovarian cancer tissues and adjacent tissues was quantitatively detected. Quantitative real-time polymerase chain reaction was used to detect the mRNA and mirNA-148a expressions of CADM1 gene. The CADM1 gene and DNMT1 protein levels were detected by Western blot. Human ovarian cancer SKOV3 cells were treated with different concentrations of methyltransferase inhibitors (5-Azacytidine, 5-aza) , and CADM1 mRNA expression was detected 72 h later. Human ovarian cancer cell lines SKOV3 were transfected with mir-335-5p mimic, inhibitor and negative control respectively. Then mir-335-5p expression level and CADM1 gene methylation level were detected after transfection.Results:The methylation level of CADM1-1 island in ovarian cancer tissues was 2.89%±0.82%, significantly higher than that of paracancerous normal tissues 1.86%±0.68% ( t=4.936, P<0.001) , and that of CADM1-2 island in ovarian cancer tissues was 3.12%±0.93%, significantly higher than that of paracancerous normal tissues (2.27%±0.69%, t=5.114, P<0.001) . Pearson correlation analysis showed that the methylation level of CADM1-1 island and CADM1-2 island in ovarian cancer tissues was significantly negatively correlated with the relative mRNA expression (r was -0.615 and -0.582, respectively, and both P<0.001) , and with the protein expression level of CADM1 (r was -0.521 and -0.612, respectively, and both P<0.001) . The relative expression level of mirNA-148a in ovarian cancer tissues was 1.53±0.42, significantly lower than that in paracancer tissues (2.59±0.73, t=6.113, P<0.001) . After treatment with different concentrations of 5-AZA, mRNA expression levels of CADM1 gene in SKOV3 cells were significantly higher in the low concentration group and the high concentration group than in the control group (both P<0.05) , and mRNA expression levels in the high concentration group were significantly higher than in the low concentration group ( P<0.05) . After mirNA-148A transfected SKOV3 cells, the relative expression levels of mirNA-148a in the mimic group were significantly increased, while those of inhibitor group were significantly decreased ( P<0.001) . The DNMT1 expression level and CADM1 gene methylation level of mimic group were significantly decreased, while those of inhibitor group were significantly increased (P<0.001) . Conclusion:In ovarian cancer, miRNA-148a can regulate the DNA methylation level of CADM1 gene by acting on the downstream target protein DNMT1, thus affecting the mRNA and protein expression levels of CDM1 gene and participating in the pathogenesis of ovarian cancer.
