Xianqi Feng, Ling Zhang, Shumin Nie, Zhan Su, Xue Shi, Yan Gao, Xiangyun Chen, Wenyuan Niu, Zhongguang Cui, Hongguo Zhao, Fanjun Meng and Chunting Zhao Pages 31 - 35 ( 5 )
Objective: To investigate the effects of the DNA methyltransferase inhibitor 5-Aza-2'- Deoxycytidine (5-Aza-Dc) on DLC-1 gene expression, methylation level, and the expression of downstream signaling molecules Cdc42 in the human multiple myeloma cell line RPMI8226 cells.
Methods: The RPMI8226 cells were treated with 5-Aza-Dc, the methylation status of CpG island of DLC-1 gene was detected by bisulfate sequencing PCR(BSP) in RPMI8226 cells. The expression of DLC-1 and Cdc42 mRNA was determined by reverse transcriptase polymerase chain reaction (RT-PCR). Enzyme-linked immunoabsorbent assay (ELISA) was used to detect the expression of Cdc42 protein.
Results: The methylation of DLC-1 gene was detected in the RPMI8226 cells without 5-Aza-Dc pretreament. DLC-1 gene methylation status was decreased after the 5-Aza-Dc treatment for 72h, and DLC-1 gene didn't display DNA methylation on the highest concentration of 5-Aza-Dc. DLC-1 mRNA was weakly expressed in the control group, and the expression was gradually increased in 5-Aza-Dc treatment group. The Cdc42 mRNA and protein expression of experimental group were significantly decreased and were dose-dependent compared with the control group (P <0.05).
Conclusion: The results of this research indicated that 5-Aza-Dc can effectively inhibit the methylation status of DLC-1 gene, reversal DLC-1 gene expression, and significantly decrease the expression of downstream signaling molecules Cdc42 mRNA and protein in RPMI-8226 cell.
5-aza-2`-deoxycytidine, Cdc42, DLC-1, methylation, multiple myeloma.
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