| [objective] To explore the relationships between the percentage of CD34+/CD123+ leukemia stem cells in acute myeloid leukemia’s bone marrow mononuclear cells and outcomes in newly diagnosed, refractory /relapse and CR AML(no M3) patients.Then explore the relationships between the ID4, CDH13 gene methylations in CD34+/CD123+ leukemia stem cells with AML’s outcomes, in order to provide the theoretical basis to methylation target therapy in clinical maintenance therapy.[methods] 1.collected 20 cases of AML (no M3) patients’s bone marrow specimens and the related clinical datas (classification, staging and other clinical informations);Using immunomagnetic heads separating specimens of bone marrow in patients with AML, obtain CD34+/CD123+leukemia stem cells;Using flow cytometry to assess the effect of immunomagnetic heads separating method;explore the relationship between the percentage of CD34+/CD123+ leukemia stem cells in acute myeloid leukemia and the outcomes.2.extract the DNA from the CD34+/CD123+leukemia stem cells, using the methylation specific PCR method to detection the ID4, CDH13 gene methylation in CD34+/CD123+leukemia stem cells, and explore the ID4, CDH13 methylations in leukemia stem cells correlation with the prognosis of AML.[results] 1. immunomagnetic heads separating results show that: after CD34 positive immunomagnetic heads separated, purity of CD34+ cells reached (74.87±10.49%) cells recovery rate was (21.77±8.26%), after CD 123 positive immunomagnetic heads separated, purity for CD34+/CD123+cells reached (68.29±12.99%), cells recovery rate was (16.35±3.51%).2. CD34 and CD 123 antigen are expressed in newly diagnosed, refractory/relapse and CR AML patients, almost no expression in the control groups. The highest proportion of CD34+ cells existed in refractory/relapse groups, the mean was 62.72%;the lowest existed in CR groups, the mean was 4.73%, two groups to compare P<0.05,the difference was statistically significant.CD34+/CD123+cells in refractory/relapse groups was the highest, the mean was 23.41%, the lowest was the CR groups, the mean was 2.91%,two groups to compare P< 0.05, the difference was statistically significant. Newly diagnosed groups compared with refractory/relapse groups and CR groups, P> 0.05, there were no statistically significant differences.3.MSP results show that:3.1 ID4 gene methylation in refractory/relapse groups was highest, reached 100%, CR groups was 60%, newly diagnosed was 77.78%, not methylation in normal control groups, and comparing every two groups, P< 0.05, the difference was statistically significant.3.2 CDH13 gene methylation in refractory/relapse groups was highest, reached 66.67%, the lowest was CR groups, reached 20%, newly diagnosed group was 55.56%, the normal control group did not detect the methylation, comparing every two groups, P< 0.05, the difference was statistically significant.3.3 ID4, CDH13 all methylation, refractory/relapse groups was highest, reached 66.67%, newly diagnosed groups was 44.44%, the CR groups was 0, comparing every two groups, P<0.05, the difference was statistically significant.ID4, CDH13 gene methylation have obvious relationship with AML disease states.[conclusion] 1. immunomagnetic heads separating method can effective sorting out of CD34+/CD123+ leukemia stem cells.2.CD34+/CD123+ leukemia stem cells proportion in the refractory recurrent groups is highest, the lowest in the CR group, its associated with prognosis of AML, the higher the ratio, the worse prognosis,and it can be used as one of the targets inAML,and MRD monitoring.3.ID4, CDH13 gene in AML patient’sCD34/CD123+ leukemia stem cells exist in different degree of methylation, degree of abnormal methylation occurs widely associated with prognosis of AML, the more the worse prognosis, it can be used as one of AML prognosis judgement index.Gene methylation can be used as one of index of the AML prognosis. In the treatment of AML, especially, in the consolidation and maintenance therapy, demethylation therapy has a certain value. |
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