Effect of Chlorambucil on the Apoptotic Signaling Pathway in Lymphoma Cells / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To study whether chlorambucil has apoptotic effect on the B cell lymphoma A20 cells and its exact mechanisms in apoptotic signaling pathway.</p><p><b>METHODS</b>The experimental cells were treated with 20 μmol/L chlorambucil, the control cells were treated with PBS. Annexin V-FITC Cell Apoptosis Detection Kit was used to examine cell apoptosis. Western blot was used to detect the expressions of active caspase-3, Survivin, NF-B and pAKT. Real-time fluorescent quantitative PCR was performed to examine the mRNA expression of Survivin.</p><p><b>RESULTS</b>Compared with the control group, the proportion of FITC/PI apoptotic cells and the expression of active caspase-3 (t=7.384, P=0.000) in the chlorambucil treatment group was significantly elevated. However, the expression of Survivin mRNA (t=4.384, P=0.000), protein expressions of survivin (t=12.360, P=0.000), NF-B (t=5.462, P=0.000) and pAKT (t=7.183, P=0.000) in the chlorambucil-treated group all significantly decreased.</p><p><b>CONCLUSION</b>The chlorambucil can induce the apoptosis of lymphoma cells, its mechanism may related with inhibition of PI3K/AKT signaling pathway, and expression of NF-B and survivin.</p>

Action Mechanism of Chlorambucil against Mantle Cell Lymphoma Cell Line Jeko-1 / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To explore the action mechanism of chlorambucil against mantle cell lymphoma cell line Jeko-1.</p><p><b>METHODS</b>The effect of chlorambucil on Jeko-1 cell proliferation was measured by MTT method. The effect of chlorambucil on the apoptosis of Jeko-1 cell was detected by Hoechst staining and Annexin V-FITC dual staining. The activation of PI3K/AKT signaling pathway and the expression of BAX, BCL-2, procaspase 3, procaspase 8 and procaspase 9 were detected by Western blot.</p><p><b>RESULTS</b>0, 5, 10, 20 µmol/L chlorambucil could inhibit Jeko-1 cell proliferation at 24, 48, 72 h in a time- and dose-dependent manner. Chlorambucil of 0, 5, 10, 20 µmol/L increased the apoptotic rate of Jeko-1 cells, upregulated the expression of BAX, procaspase 3, procaspase 8, procaspase 9 and PI3K, increased the phosphorylation of AKT and down-regulated the expression of BCL-2.</p><p><b>CONCLUSION</b>The chlorambucil can induce the apoptosis of mantle cell lymphoma Jeko-1 cells via blocking PI3K/AKT signaling pathway.</p>

Expression and Promoter CpG Island Methylation Status of miR-34b in Leukemia Cell Lines and Their Clinical Significance / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To explore the expression and promoter CpG island methylation status of miR-34b in leukemia cell lines and their clinical significance.</p><p><b>METHODS</b>A total of 10 cases of non-hematologic diseases were selected as control group, and the bone marrow cells of control group and HL-60, K562 cells were selected; the relative expression of miR-34b was detected in bone marrow cells, HL-60 and K562 cell lines by fluorescence quantitative PCR, and the MiR-34b methylation status was detected by methylation-specific PCR, the HL-60 and K562 cell lines were treated with decitabine, and the expression levels and methylation status of miR-34b in the 2 cell lines were detected by the same method. Has-miR-34b was transfected into K562 cells, which were divided into non-transfection group, negative control group and Has-miR-34b transfection group; if the transfection was successful, the cell proliferation should be recorded at different time points of culture, and the proliferation inhibition rate should be calculated.</p><p><b>RESULTS</b>The relative expression level of miR-34b in the control group was (5.23 ± 0.75), in HL-60 was (0.05 ± 0.01) and in K562 was (0.04 ± 0.01). The difference between 3 groups was statistically significant (F = 44.812, P < 0.01). The promoter regions of CpG island in HL-60 and K562 cell lines were methylated, while the bone marrow cells were not methylated in 10 cases of non hematologic diseases children.Through miR-34b expression levels of HL-60 and K562 cell lines significantly increased by decitabine treatment (P < 0.05), and the methylation of leukemia cell line promoter region CpG island was found before and after decitabine treatment, but after administration of decitabine the methylation significantly decreased, suggesting that decitabine has an inhibitory effect on methylation of promoter region CpG island. After being cultured for 48, 72, 96 and 120 hrs, the cell proliferation in Has-miR-34b transfection group reached to 24.8%, 46.7%, 33.6% and 4.7%, repectively, and significantly lower than that in non transfection group (P < 0.05).</p><p><b>CONCLUSION</b>CpG island methylation of miR-34b promoter region in leukemia cell lines can decrease the expression levels of miR-34b, which is also the reason why miR-34b can reduce the inhibition of cell proliferation, thus miR-34b might be a tumor suppressor gene involved in the regulation of leukemia.</p>