Mir-15a and Mir-16-1 Induce Apoptosis of Raji Cell Line

MicroRNAs are endogenous small noncoding RNAs that regulate gene expression negatively at posttranscriptional level. Mir-15a and mir-16-1 are deleted or downregulated in the majority of chronic lymphomatic leukemia(CLL), which is the most common human leukemia and overexpresses the antiapoptotic B cell lymphoma2 (Bcl-2) protein. Here we investigated the effect of mir-15a and mir-16-1 on human lymhpomatic cell line Raji in induction of apoptosis. We transfected chemically synthesized mir-15a and mir-16-1 oligonucleotide into Raji cells using Lipofectamine 2000 reagent. At 24,48,72 hours after transfection, the growth inhibitory effect of Raji cells was measured by trypan blue dye exclusion method and CCK8 assay. Apoptosis was determined by morphological observation and flow cytomertry analysis after AnnexinV/PI double labeling. The expression levels of Bcl-2 protein were detected by immunofluorescence assay. Trypan blue dye exclusion method and CCK8 assay showed that transfection of mir-15a or mir-16-1 decreasesd the cell growth at 24, 48 and 72 h, which were significantly lower than those cells transfected with control oligonucleotide and untransfected cells, respectively (P<0.05). Using Hoechst staining, cells treated with mir-15a or mir-16-1 oligonucleotide displayed changes of apoptosis at 48,72h after transfection. AnnexinV/PI double dyeing assays showed early and late apoptotic cells at 48 and 72 h post-transfection. At 48,72h after transfection with mir-15a or mir-16-1 oligonucleotide, early and late apoptotic cell rate were obviously higher than untransfected cells and control miRNA group. Indirect immunofluorescence assay demonstrated that the expression of Bcl-2 was degraded at post-transfection, suggesting that Bcl-2 is a direct target of mir-15a and mir-16-1. These results show that mir-15a and mir-16-1 can induce apoptosis of human lymhpomatic cell line Raji. Our results suggest that mir-15a and mir-16-1 could be used for therapy of Bcl-2 overexpression tumors.