Evaluation of miR-15a, miR-16-1, ZAP-70, Ang-2, and Bcl-2 as potential prognostic biomarkers in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) is a blood cancer characterized by the accumulation of clonal B-lymphocytes. This study evaluated the mRNA gene expression of miR-15a, miR-16- 1, ZAP-70, and Ang-2 by qPCR, as well as the plasma levels of Bcl-2 by Elisa immunoassay, in CLL patients and healthy controls. Significant differences were observed when comparing patients and controls regarding miR-15a (p < 0.001), miR-16-1 (p < 0.001) mRNA, Ang-2 gene expression, and Bcl-2 plasma levels (p < 0.001). When stratified by risk, differences were maintained with a significantly reduced expression in high-risk patients. A positive correlation was observed between miR-15a and platelets (R2 = 0.340; p = 0.009) as well as between Bcl-2 and leukocytes (R2 = 0.310; p = 0.019). Conversely, negative correlations were observed between ZAP-70 and platelets (R2 = - 0.334; p = 0.011), between miR-15a and lymphocytes (R2 = - 0.376; p = 0.004), as well as between miR-16-and lymphocytes (R2 = - 0.515; p = 0.00004). The data suggest that a reduction in miR-15a and miR-16-1 expressions, in addition to an overexpression of Bcl-2, are associated with the reduction in apoptosis and, consequently, to a longer survival of lymphocytes, thus contributing to lymphocyte accumulation and aggravation of the disease. By contrast, Ang-2 expression was significantly higher in A than in B + C Binet groups. This context leads to the speculation that this biomarker should be investigated in more robust studies within populations with a still relevantly indolent form of the disease in an attempt to identify those patients with a greater potential for an aggravation of the disease

MiR-15a and miR-16-1 enhance sensitivity of Raji cells to Ara-C / 第二军医大学学报

Objective To study whether miR-15a and miR-16-1 can enhance the sensitivity of Raji cells to cytarabine (Ara-C). Methods MiR-15a and miR-16-1 oligonucleotides were transfected into Raji cells with Lipofectamine™ 2000, and then the cells were treated with Ara-C. The IC50 values of Ara-C was detected by CCK8 assay. The growth of Raji cells was measured by trypan blue dye exclusion method. The apoptotic cells were observed by Hoechst dyeing; AnnexinV/PI double dyeing and glow cytometry(FCM) were used to examine the cell apoptotic rate. Results After transfection of miR-15a or miR-16-1 into Raji cells, the IC50 values of Ara-C were 10. 41 and 10. 86, respectively, which were significantly lower than that of the untransfected group(15.43)and scrambled oligonucleotides (SODN)transfection group(14. 92, P<0.05). Trypan blue dye exclusion assay showed that miR-15a/miR-16-1 transfection group had obviously decreased the cell growth compared to miR-15a, miR-16-1 group, untransfected group and SODN transfected group; Hoechst dyeing demonstrated plenty of apoptotic cells. AnnexinV/PI double dyeing assays by FCM indicated that the cell apoptotic rates in earlier period and late period were 20. 93% and 25. 27% in the miR-15a+Ara-C group, and 20. 69% and 23. 13% in the miR-16-1 + Ara-C group, which were obviously higher than those in miR-15a group (6. 99%, 10. 08%), miR-16-1 group(4. 73%, 10. 64%), Ara-C group (10. 88%, 11. 83%) and control group (14. 39%, 11. 93%). Conclusion MiR-15a and miR-16-1 oligonucleotides can enhance the sensitivity of Raji cells to Ara-C.