RÉSUMÉ
Objective@#To investigate the effect and mechanism of all-trans retinoic acid (ATRA) on leukemic cell line U937 cells with NPM1 mutation.@*Methods@#Human acute myeloid leukemia cell line U937 was explored, NPM1 mutated (A type) plasmids were transfected into U937 to form stable clones A1 and A2, which were identified by Western blot and Co-immunoprecipitation. The cell proliferation was measured by methylthiazolyl tetrazolium bromide (MTT) ; cell cycle and cell apoptosis were explored by flow cytometric; cell colony formation was measured by microscope count, the molecular pathways related to cell proliferation were measured by Western blot.@*Results@#①The cell proliferations of mutant A1 and A2 were inhibited significantly by 52.6% and 35.8% (P<0.05) , respectively under ATRA exposure. ②The percentages of G0/G1 stage of mutant A1 and A2 increased by 20.1% and 35.8%, respectively under ATRA exposure. ③All the U937 leukemic cells were inhibited under ATRA exposure; the decreased percentages of vector, wild-type and mutant NPM1 cells were 32.7%, 57.9% and 90.9% respectively. ④p-ERK decreased obviously after ATRA exposure in NPM1 mutated leukemic cells. ⑤More mutant NPM1 cells inclined to apoptosis under the exposure of ATRA and cytotoxic drugs than cytotoxic drugs alone, meanwhile more cells apoptosis occurred when ATRA was administrated after cytotoxic drugs exposure.@*Conclusions@#ATRA could inhibit cell proliferation and colony formation, blocked the cell cycle in the G0/G1 stage accompanied by the significant reduction of p-ERK in U937 leukemic cells with NPM1 mutation. Besides, ATRA could synergize with drugs to suppress the leukemic cells survival more effectively when ATRA was administered after the cytotoxic drugs exposure in U937 leukemic cells with NPM1 mutation.
RÉSUMÉ
Objective@#To investigate the impact and mechanism of NPM1 gene expression on acute myeloid leukemia (AML) cell lines.@*Methods@#Human AML cell line U937 and HL-60 cells were transfected with NPM1 plasmid to establish stable clones, and the high NPM1 protein expression (NPM1hi) clones were screened by Western blot. The cell proliferation was assayed by methylthiazolyl tetrazolium bromide (MTT) , cell cycle and cell apoptosis by flow cytometric, cell colony formation by microscope count, the molecular pathways related to cell cycle by Western blot. The expression of NPM1 gene in primary AML bone marrow mononuclear cells (BMMC) was investigated by RQ-PCR.@*Results@#①The proliferation of NPM1hi U937 and HL-60 cells was similar with that of control cells (4.68±1.28 vs 3.89±0.81, 3.34±0.37 vs 2.68±0.29, P>0.05) . ②The percentage of S phase in NPM1hi U937 and HL-60 cells was higher than that in control cells[ (50.22±3.42) % vs (39.78±3.80) %, (59.01±3.27) % vs (43.94±2.08) %, P<0.05]. ③The anti-apoptosis capacity and colony formation abilities of NPM1hi U937 cells increased than that of control cells[ (68.8±10.2) % vs (48.7±3.22) %, and (772.7±24.0) vs (652.3±16.5) , P<0.05], but the above abilities of NPM1hi HL60 cells were similar with that of control cells. ④ The expressions of CDK4, Cyclin D1, Cyclin D2 and Cyclin E in NPM1hi leukemia cells were higher than that of control cells, but the expression of Cyclin D3 was lower. ⑤The NPM1 expression levels in AML patients with favorable cytogenetic prognosis were lower than that of patients with intermediate prognosis.@*Conclusions@#NPM1 protein could promote more cells to enter S phase, enhance the ability of antiapoptosis and colony formation in AML cell lines. The quantitative level of NPM1 may predict the cytogenetic risk of AML patients.