| Objective: Differential expression of microRNAs(miRNAs) has been implicated in leukemogenesis. Based on the prediction of bioinformatics technology, we investigated the expressions of several mi RNAs closely related to initial pediatric acute myeloid leukemia, analyzed its clinical significances in diagnosis and prognosis, and studied its potential functions.Methods: Using bioinformatics technology, we predicted three miRNAs closely related to the diagnosis of pediatric AML. 83 AML children were included, and 29 non-malignant disease matched children were set up as controls. Real-time quantitative polymerase chain response(qRT-PCR) was applied to validate the expressions of these miRNAs in bone marrow samples. We detected the expressions(the results were expressed in 2-△△Ct) and analyzed its clinical significances. Furtherly, we studied its functions on leukemia cell lines and explored the underlying mechanism.Results:1. miR-196 b, miR-155 and miR-25 were predicted as candidate miRNAs closely related to initial AML in children.Based on improved POMA model, miR-196 b, miR-155 and miR-25 were revealed to be responsiable for the occurrence of pediatric AML, due to its larger NOD and TFP(Wilcoxon test, p<0.05; NOD=30, 28, and 32, TFP=0.289, 0.242 and 0.191). In addition, IGFBP7 gene was predicted as potential target of miR-196 b.2. Verifications in clinical specimens:The expressions of miR-155 in initial AML children were significantly higher than those in controls(p<0.0001). Differential expression of miR-196 b was significantly associated with specific subgroups, although no significant differential expression was found between AML full corhort and controls. Overexpressions of miR-196 b existed in M4/5 group compared to controls(p=0.032) and non-M4/5 group(p<0.0001).3. The correlations between the expressions of miRNAs and prognosis of AML Children.A significant association was observed between high expression of miR-196 b or miR-155 and inferior overall survival(OS) of pediatric AML(Log Rank p<0.0001, =0.011, respectively), with higher early mortality. No obvious association existed between miR-25 expression and OS of AML patients(Log Rank p>0.05)。Among prognosis groups according to NCCN 2015 gudline and WHO classification, the lowest level of miR-196 b or miR-155 was observed in favorable prognosis group(p<0.001, =0.0002). miR-196 b expression was significantly higher in non-remission group as compared to remission group after the first induction remission therapy(p=0.018), but no result similar to this for miR-155 or miR-25(p>0.05). Among various molecular mutation subgroups, the highest expression was observed in FLT3-ITD for miR-196b(p=0.037), but in C-Kit for miR-155 and mi R-25(p=0.004, 0.047). Among abnormal cytogenetic subgroups, expression level of miR-196 b was the lowest in t(8;21) and the highest in MLL-rearrangement compared with other cytogenetic subgroups(p=0.0002, 0.0002, respectively). The lowest level of miR-155 was in t(15;17) group(p=0.0004)。4. The correlations between the expressions of miRNAs and other clinical data.We observed higher miR-196 b expression in WBC≥100?109/L and age≤12 months group at presentation(p=0.004, 0.029, respectively). Using Spearman’s correlation analysis, we found that miR-196 b expression was positively associated with platelet(PLT) count(Spearman’s r=0.31, p=0.004). No significant correlation between the expressions of miR-155 or miR-25 and the counts of WBC or PLT(p>0.05).5. The expression of IGFBP7 in initial bone marrows of AML children was not only higher than that in controls(p<0.0001), but also related to favorable prognosis of patients(Log Rank p=0.045)6. Exogenous miR-196 b overexpressed in leukemia cell lines can promote itsproliferation and inhibit the apoptosis induced by VP16 in MV4-11 cells, as well as decreased differentiation of NB4 cells induced by retinoic acid.7. On the levels of mRNA and protein, negative regulation roles in IGFBP7 expression were observed after transient transfection for miR-196b-mimics/inhibitors in 293 T cells.Conclusions:1. Differential expressions of mi R-155 and miR-196 b not only exist in initial pediatric AML, but also have correlations to the outcome of patients. miR-155 and miR-196 b potentially can be promising biomarkers for diagnosis and prognosis in pediatric AML.2. Overexpressed mi R-196 b can promote proliferation, inhibit apoptosis, and reduce differentiation in leukemia cell lines. It is expected to be a new target for treatment in pediatric AML.3. IGFBP7 could be a target of miR-196 b. The regulatory relationships between them promote the occurrence and development of AML. It is necessary for further validation. |
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