[Expression of Insulin-Like Growth Factor Binding Protein 7 Gene in Multiple Myeloma Cell Line and Methylation Regulation].

OBJECTIVE

To investigate the expression of insulin-like growth factor binding protein 7 (IGFBP7 ) gene in multiple myeloma cell line U266, and study the effect of 5-aza-2'-deoxycytidine (5-aza-dc) on proliferation of U266 cells.

METHODS

multiple myeloma cell line U266 was cultured in vitro. The bone marrow mononuclear cells from healthy persons(N-BMMNC) were collected and used as normal controls. The IGFBP7 mRNA expression of U266 cells and N-BMMNC were detected by real-time fluorescence quantitative PCR, the DNA methylation status of the IGFBP7 CpG island was measured by using methylation-specific PCR(MSP). The different concentrations of 5-aza-dc (5 µmol/L, 10 µmol/L, 20 µmol/L) were used to treat U266 cells for 48 hours, the RT-PCR and Western blot were used to detect the effect of IGFBP7 mRNA and protein expressions, the cell growth curve and Annexin V/PI were analyzed by flow cytometry.

RESULTS

As compared with normal BMMNC, the lower expression of IGFBP7 gene was found in U266 cells, the obvious hypermethylation of the CpG island in the IGFBP7 promoter was observed. After treatment of U266 treating with different concentrations of 5-aza-dc, the IGFBP7 mRNA expression was up-regulated dose-dependently(P<0.05), the U266 cells grew slowly and apoptosis rates were enhanced dose-dependently.

CONCLUSION

As the hypermethylalion of CpG island in IGFBP7 promoter is a frequent event in lower expression of IGFBP7 gene in U226 cells, the 5-aza-dc can up-regulate the expression of IGFBP7 , and can inhibit cell proliferation through induction of cell apoptosis and arrest of cell cycle.