Negative regulation of microRNA - 9 by ecotropic viral integration site1 involved in the pathogenesis of acute myeloid leukemia / 中华实用儿科临床杂志

ABSTRACT

Objective To investigate the regulation mechanism of microRNA - 9(miR - 9)by ecotropic viral integration site1(EVI1)its impact on proliferation of AML cells and its role in the pathogenesis of myelogenous leuke-mia. Methods EVI1 was forced to express in Uocm1 cell lines by murine stem cell virus - EVI1(MSCV - EVI1) plasmid infection. EVI1 overexpressed Uocm1 cells were then treated with 0. 1 μmol/ L 5 - aza - 2′ - deoxycytidine (5 - AZA)dissolved in dimethyl sulfoxide (DMSO). The methylation level of miR - 9 promoter was tested by DNA bi-sulfite sequencing technology. The cell cycle was observed by flow cytometry (FCM). The proliferation ability of the cells was detected by the colony forming assay in semi - solid Methylcellulose medium culture. Results EVI1 level was dramatically increased after being infected by MSCV - EVI1 plasmid. Forced expression of EVI1 in Uocm1 signifi-cantly downregulated miR - 9 by inducing hypermethylation of miR - 9 promoter. Relative expression level of miR - 9 was lower in EVI1 overexpressed group(0. 004 ± 0. 000)than that of the control group(0. 006 ± 0. 001)(t = 4. 09,P <0. 05). When EVI1 was overexpressed in Uocm1,the rate of G0 / G1 cells decreased markedly(P < 0. 05),while rates of S phage and G2 phage increased significantly(all P < 0. 05). Seven days after 500 cells plated in semi - solid medium, EVI1 overexpressed Uocm1 cells gave rise to more colony (122. 3 ± 7. 8)than Uocm1 cells infected with vector (45. 7 ± 6. 1)(t = - 13. 44,P < 0. 01). 0. 1 μmol/ L 5 - AZA recovered miR - 9 expression(P < 0. 01)by decreasing EVI1 induced hypermethylation of miR - 9 promoter. G0 / G1 phase cell proportion was(48. 25 ± 2. 19)％ in control group,while (65. 90 ± 2. 90)％ in 5 - AZA group (t = - 6. 85,P < 0. 05). 5 - AZA group formed less colony (51. 00 ± 10. 01)than the control group (123. 40 ± 8. 12)(t = 9. 59,P < 0. 01),which indicated that 5 - AZA inhibi-ted cell proliferation by G0 / G1 cell cycle retardation in EVI1 overexpressed uocm1 cells. Conclusions EVI1 may en-hance proliferation ability of myeloid leukemia cells by downregulating miR - 9 through inducing hypermethylation of miR - 9 promotor,which plays a crucial role in the pathogenesis of AML. 5 - AZA may be an effective hypomethylating agent in the therapy of EVI1 high acute myeloid leukemia.