| ObjectivemiRNAs play important roles in the regulation of cell proliferation, differentiationand apoptosis. The deregulation of miRNAs expression contributes to tumorigenesisby modulating oncogenic and tumor suppressor signaling pathways. Oncogenictranscription factor Myc can control expression of a large set of microRNAs(miRNAs). Previous studies have shown that the expression of miR-17-92cluster hasclose relationship with the expression of Myc. In current study, silencing Myc inmultiple myeloma (MM) cells induced cell death and growth inhibition, anddownregulated expression of miR-17-92cluster. Overexpression of miR-17ormiR-18a could partly abrogated Myc-knockdown-induced MM cell apoptosis. One ofthe mechanism of Myc inhibiting MM cell apoptosis is through Myc activatingmiR-17-92cluster and subsequently down-modulating proapoptotic protein Bim.This research aimed to investigate the role of miRNA-17-92cluster in multiplemyeloma.MethodsWe used lentiviral transduction to silence Myc expression and evaluated theeffects of Myc-shRNA on cell growth, survival and apoptosis in KMS28PE,OCI-My5and XG-1myeloma cell lines. miR-17-92cluster miRNAs expression wereexamined in KMS28PE, XG1and OCI-My5Myc silencing cell lines compared toKMS28PE, XG1and OCI-My5scramble cell lines by quantitative RT-PCR. We alsoused luciferase reporter assays to demonstrate the target gene of miR-17andmiR-18a.ResultsGrowth analysis indicated that silencing Myc in these myeloma cell lines didnot inhibit cell proliferation until day3after transfection; whereas, on day7a dramatic decrease of cell viability could be observed in Myc-shRNA-expressing cellscompared to the cells expressing the scramble control shRNA (40–52%vs88–92%, p<0.05). miR-17, miR-18a, miR-19a, miR-20a and miR-92-1all decreased in theKMS28PE, XG1and OCI-My5Myc silencing cell lines (p <0.05). OverexpressingmiR-17and miR-18a could enhance cell proliferation in KMS28PE and OCI-My5-Myc-knockdown cell lines, and the cell viabilities at day6increased from56.9%±0.42%to80.2%±3.39%(overexpression miR-17, p=0.003) or78.75%±1.06%(overexpression miR-18, p=0.003) in KMS28PE-Myc-knockdown cell line and from49.05%±5.16%to81.45%±2.19%(overexpression miR-17, p=0.002) or72.9%±0.28%(overexpression miR-18, p=0.006) in OCI-My5Myc-knockdown cell line.ConclusionsMyc silence induced myeloma cell apoptosis. Myc positively modulatedmiR-17-92cluster, and Bim is the direct downstream target of miR-17and miR-18a.Overexpression of miR-17or miR-18a could partly abrogatedMyc-knockdown-induced MM cell apoptosis. ObjectiveMultiple myeloma (MM) is a clonal plasma cell disorder characterized by clonalproliferation of malignant plasma cells in bone marrow, monoclonal protein in bloodor urine, and associated organ dysfunction. It accounts for approximately10%of allhematologic cancers. The conventional therapy results in lower response and MM isalso incurable today. The clinical heterogeneity of MM is decided by multivariatefactors. The most important genetic abnormality with specific prognostic value wasthe loss of chromosome13[2-5]. Loss of chromosome13, observed in almost half ofpatients, negatively affects prognosis. miR-15a, miR-16-1located on13q14，andmiR-17-92cluster, located on13q31.3, play important roles in the regulation of cellproliferation, differentiation and apoptosis. We measured the expression of miR-15a,miR-16-1and miR-17-92cluster in85newly diagnosed MM patients by quantitativereal-time PCR analyses, to investigate the relationship between expression ofmiR-15a, miR-16-1and miR-17-92cluster with chromosome13deletions (del(13))and prognosis.MethodWe measured the expression of miR-15a, miR-16-1and miR-17-92cluster in85newly diagnosed MM patients by quantitative real-time PCR analyses, to analyze ifthese miRNAs expressions are correlated with age, disease stage, bone marrowplasma cell percentage, β2-MG, serum albumin, LDH, serum creatinine andC-reactive protein (CRP). We also assessed whether a direct correlation existsbetween different microRNA expression and PFS in MM patients. ResultsWe studied85newly diagnosed MM patients, including54males and31femaleswith a median age of60years (range33-82years). According to Durie and Salmonstaging,7patients had stage I,12patients had stage II, and66patients had stage III.According to International Staging System,13patients had stage I,20patients hadstage II, and52patients had stage III. The paraprotein was IgG in46patients, IgA in12patients, kappa light chain in16patients and lambda light chain in11patients. Weidentified del(13q14) in53/85(62.4%) patients for miR-15a, miR-16-1andmiR-17-92cluster using the I-FISH, and non-del(13q14) in32/85(37.6%). Themedian levels of miR-15a, miR-16-1, miR-17, miR-18a, miR-19a, miR-20a andmiR-92-1in del(13q14) MM patients were33.13±6.68,105.11±4.32,31.78±2.62,391.45±8.41,25.94±1.48,60.70±3.27and31.70±4.89, respectively; while the medianlevels of miR-15a, miR-16-1, miR-17, miR-18a, miR-19a, miR-20a and miR-92-1inpatients not harboring this deletion were25.25±3.17,95.67±4.66,28.64±4.95,134.54±8.17,23.91±2.88,29.95±3.83and25.90±2.09respectively. The resultsshowed that miR-15a, miR-16-1, miR-17, miR-18a, miR-19a, miR-20a and miR-92-1expression levels didn’t decrease in patients with the deletion of13q14. In85MMpatients, the expression levels of miR-15a, miR-16-1, miR-17, miR-18a, miR-19a,miR-20a and miR-92-1are not associated with age, disease stage, bone marrowplasma cell percentage, β2-MG, serum albumin, LDH, serum creatinine andC-reactive protein (CRP)(p>0.05). We also assessed whether a direct correlationexists between different microRNA expression and PFS in MM cases. The medianduration of PFS of patients highly expressing miR-15a, miR-16-1, miR-17, miR-20aand miR-92-1were11,13,13,12.5and16months, respectively. However, themedian duration of PFS of low expression patients were22.5,18,26,23.5and20.5months, respectively. These results indicated that high expression of miR-15a,miR-16-1, miR-17, miR-20a and miR-92-1were associated with shorter PFS(p=0.025,0.046,0.027,0.032,0.048, respectively).ConclusionsmiR-15a, miR-16-1, miR-17-92cluster expression levels didn’t decrease in patients with the deletion of13q14, which suggested that miR-15a, miR-16-1,miR-17-92cluster expression levels are independent of the deletion of chromosome13. High expression of miR-15a, miR-16-1, miR-17, miR-20a and miR-92-1wereassociated with shorter PFS compared with low expression, which suggested thathigh expression is a poor prognosis factor. |
PDF Full Text Request