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Objective:To detect the expression of microRNA-218 (miR-218) in human acute lymphocyte leukemia (ALL) T lymphocytes ( CCRF-CEM) ,explore its effects on the biological features of CCRF-CEM cells and the expression of its target gene c-kit, so as to provide new insights for leukemia treatment.Methods: Using the quantitative real-time polymerase chain reaction ( qRT-PCR) ,we detected the expression of miR-218 in the normal peripheral blood T lymphocytes and CCRF-CEM cells.Forty-eight hours after the miR-218 mimic was transfected into the CCRF-CEM cells,the expression of miR-218 in the CCRF-CEM cells was detected by qRT-PCR.The effect of miR-218 on the CCRF-CEM cell viability was detected using MTT.The effect of miR-218 on the proliferation and apoptosis of CCRF-CEM cell was analyzed using flow cytometry.c-kit gene was identified to be a target gene of miR-218 by luciferase reporter enzyme system,and the effect of miR-218 on the expression of KIT protein in cells were determined using Western blot.Results:As shown by qRT-PCR,compared with that in the normal peripheral blood T lymphocytes,the expressions of miR-218 in ALL T lymphocytes cell lines were significantly decreased ( P<0.01 ) .Compared with the control group, the expression of miR-218 increase significantly in CCRF-CEM cells transfected with miR-218 mimic for 48 hours ( P<0.01).MTT showed that the cell viability decreased significantly after the over-expression of miR-218 in the CCRF-CEM cells ( P<0.05 ) .Flow cytometry showed that the S-phase fraction significantly declined after the over-expression of miR-218 ( P<0.01 ) , and meanwhile the apoptosis of cells also significantly increased (P<0.01).Luciferase reporter gene assay showed that,compared with the control group,the relative luciferase activity significantly declined in the miR-218 mimic transfection group (P<0.01).Compared with the control group,the expression of KIT protein in the CCRF-CEM cells transfected with miR-218 mimic for 48 hours significantly decreased ( P<0.01).Conclusion:The expression of miR-218 decreases in ALL T lymphocytes cell lines.MiR-218 can negatively regulate the expression of KIT protein,inhibit the proliferation and increase the apoptosis of CCRF-CEM cells.Treatment based on the enhanced expression of miR-218 may be a promising strategy for leukemia.
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Objective:To investigate the expression and significance of B cell activating factor (BAFF) and a proliferation-inducing ligand ( APRIL) in children with acute lymphoblastic leukemia ( ALL).Methods:The mRNA and protein expressions in ALL.
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Objective To investigate the effects of a proliferation-inducing ligand(APRIL)gene silencing by small interfering RNA(siRNA)on cell cycle and proliferation of colon carcinoma SW480 cells.Methods The siRNA plasmid vector targeting APRIL gene,named as siRNA-APRIL,was transfected into SW480 cells,transfected with scrambled vector as a nontargeting control and nontransfected group as another control.APRIL mRNA and protein expression were examined by real-time PCR and Western blot,respectively.Cell proliferation activity was analyzed by cell counting kit-8(CK-8),cell cycle was detected by flow cytometry,and p21 together with p27,two important regulatory genes in cell cycle,were measured by RTPCR.Results Compared with nontargeting control and nontransfected control,APRIL expression was inhibited significantly at both mRNA and protein level by siRNA-APRIL being transfected in SW480 cells(P ＜0.05).Cell proliferation ability was drastically repressed after siRNA-APRIL being transfected at 48 h,72 h and 96 h(P ＜ 0.05).After transfected 48 h,the percent of Go/G1 phase cell was significantly increased,S and G2/M phase cell were significantly decreased,the number of cell in apoptosis was increased and the expression of p21 and p27 mRNA were up-regulated(P ＜ 0.05).There was no significant difference when compared the two control groups each other(P ＞ 0.05).Conclusion siRNA-APRIL can effectively knockdown the expression of APRIL gene in SW480 cells,moreover,it can inhibit the cell proliferation and induce G0/G1 phase cell cycle arrest,which occurrence may involve in upregulation the mRNA expression of p21 and p27.
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Objective To investigate the regulation of B-lymphocyte stimulator(BLyS) levels in response to IFN-γand IL-6. Methods Flow cytometry, quantitative polymerase chain reaction, ELISA and Western blot were applied to examine the expression level of BLyS in response to IFN-γ and IL-6 . Results IFN-γand IL-6 induced BLyS expression in KM3 cells. After treated with BAY11-7082, an IkB-α phospho- rylation inhibitor, the up regulation of BI,yS induced by IFN-γ was completely inhibited. Inhibiting the nu-clear faetor-kB (NF-kB) and mitogen activated protein kinase(MAPK) activation in KM3 cells reduced BLyS protein and gene expression. Conclusion MAPK and NF-kB pathways are involved in the regulation of BLyS expression, which suggests that MAPK and NF-kB might be used for the treatment of multiple mye- loma.
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Objective:To investigate the expression of APRIL in NSCLC and to analyze the relationship between expression of APRIL and clinicalpathological characteristics of NSCLC.Methods:RTQ-PCR and Western blot were used to detect the expression of APRIL mRNA and protein in 68 pair of tumor tissues and corresponding adjacent normal tissues of NSCLC.50 Formalin-fixed,paraffin-embedded NSCLC samples were tested for APRIL protein location and expression by IHC,with results compared to the clinical and pathological parameters of NSCLC to determine significance.Results:The results showed that the expression level of APRIL mRNA and protein in tumor tissues was significantly higher than in normal tissues(both P