Effect Of DNA Methylation Inhibitor And Histone Deacetylase Inhibitor On RASSF1A, SOCS-1 Gene Expression In Multiple Myeloma Cells

Background:Multiple myeloma (MM) is malignant plasma cells disease, characterized by osteolytic lesions, osteocopic pain, pathological fractures, hypercalcemia and anemia. The pathogenesis of MM is not understood fully yet. According to molecular biological study of cancers, a series of genetic or epigenetic alteration, especially the activation of oncogenes and the inhibition of tumor-suppressor genes have major contributions to the development of many human malignant tumors. Epigenetic events are defined as alterations in gene expression without changes in the DNA coding sequence that are heritable through cell division. As the most important part of the emerging epigenetics and epigenomics, methylation of CpG island resulting in the inhibited expression of cancer suppressor genes, as well as its impact on the pathogenesis and progression of tumors has become a hot issue. The reversible characteristics of epigenetic modification hints that it can be used as treatment targets for tumor patients.Objective:To investigate the effect of DNA methylation and histone deacetylase inhibitor on transcription regulation of RASSF1A and SOCS-1 tumor suppressor genes and the molecular biological behaviors in MM cells. Method:The cells were treated with different doses of 5-Aza-CdR and VPA either alone or combined. MTT was used to detect the proliferation of cells. The apoptosis and cell cycle were analyzed by flow cytometry. Methylation-specific polymerase chain reaction (MSP) was used to detect CpG island methylation in RASSF1A and SOCS-1 genes promoter. Quantitative real-time reverse transcription polymerase chain reaction (RQ-PCR) was used to examine the expression of RASSFIA and SOCS-1 genes in cells.Results:Increased growth inhibition and apoptosis effect were shown in combining treatment of 5-Aza-CdR and VPA than 5-Aza-CdR or VPA used alone (P<0.05). After treatment with 5-Aza-CdR or VPA alone for 48 h, cells were arrested in G(0)/G(1) phase in contrast to the control group(P<0.05), more cells were arrested in G0/G1 phases by combining of 5-Aza-CdR and VPA than they were used alone(P<0.05). The methylation of RASSFIA and SOCS-1 genes promoter was detected in U266 cells, and the methylation of SOCS-1 gene promoter was detected in RPMI8226 cells. There was few gene expressing in the control group. The demethylation effect could be detected in the groups treated with 5-Aza-CdR alone or both drugs, whereas no demethylation effect happened in the VPA group. The expression level of genes was induced by 5-Aza-CdR in a concentration-dependent manner. VPA was not able to induce the expression of RASSFIA or SOCS-1. The expression level was increased significantly by combination of VPA and 5-Aza-CdR. The RASSFIA gene was non-methylationed in RPMI8226 cells, and it was the same state in 5-Aza-CdR and VPA treatment groups.Conclusion:DNA methylation and histone deacetylase inhibitor could synergistically induce demethylation of the RASSFIA and SOCS-1 genes in U266 cells, the SOCS-1 gene in RPMI8226 cells, re-express genes silenced in MM cells, inhibit the proliferation of MM cells and induce cells apoptosis markedly.Keywords:gene,RASSF1A; DNA methyltransferase; histone deacetylase; Ras Association Domain Family 1A; multiple myeloma...