ABSTRACT
Objective:To investigate the effect of miR-21 on the proliferation and apoptosis of leukemia cell line K562 and the regulation of PI3K/AKT signaling pathway.Methods:K562 cells were divided into control group,miR-21 NC group and miR-21 interference group,the first group without treatment,the two groups after using cationic liposome transfection of LipofectamineTM2000 miR-21 inhibitor and miR-21 negative control.After transfection 48 h,the expression of miR-21 mRNA in cells was detected by Real-time fluorescence quantitative (qRT-PCR),MTT assay was used to test the effect of miR-21 on cell proliferation,flow cytometry was used to detected the effect of miR-21 on cell cycle apoptosis rate,the protein expression of PI3K,AKT and p-AKT in PI3K/AKT signaling pathway after miR-21 transfection were detected by protein immunoblotting (Western blot).Results:The expression level of miR-21 mRNA and the survival rate of cells in the miR-21 interference group were significantly lower than those in the control group and the miR-21 NC group;compared with control group and miR-21 group NC,flow cytometry detected the cells accounted for proportion of G0/G1 phase in miR-21 interference group increased significantly,the proportion of cells at S phase significantly decreased,and apoptosis rate increased significantly.Western blot test results show that the expression levels of p-AKT in miR-21 inter-ference group were significantly lower than those in the control group and miR-21 NC group,but the expression levels of PI3K and AKT protein were not changed significantly.Conclusion:Down-regulation of miR-21 can inhibit the proliferation and promote apoptosis of leukemia K562 cells,and its mechanism may be related to the inhibition of PI3K/AKT signaling pathway.
ABSTRACT
<p><b>OBJECTIVES</b>To explore the feasibility of DC being in vitro induced from AML cells with cytokine cocktails and their biological properties.</p><p><b>METHODS</b>AML cells were cultured in either presence or absence of cytokine cocktails. DC were studied for morphology, and cytochemical and immunofluorescent staining. Functions of DC were examined by MLC, FITC-conjugated dextran uptake test, and LDH release assay. RT-PCR and FISH were used to analyze the specific fusion genes of culture-derived DC.</p><p><b>RESULTS</b>Classical DC morphological changes occurred in all 15 cultured AML cells. DC-associated surface molecules such as CD(1a), CD(80), CD(86), CD(106), CD(83) and HLA-DR were upregulated (P < 0.05). The allostimulatory abilities of culture-derived DC were significantly higher than those of AML cells uncultured or cultured in the absence of cytokines (P < 0.05). Culture-derived DC only in the presence of GM-CSF + IL-4 have phagocytotic activities. CTL assay was performed in 5 of the 15 samples. At effector/target ratio of 20:1, auto-T lymphocytes primed with the culture-derived DC exhibited no more killing activity to auto-AML cells than those stimulated by IL-2 or uncultured AML cells. Culture-derived DC presenced the native AML-specific aberrant karyotype and related fusion gene.</p><p><b>CONCLUSIONS</b>Cytokine cocktails could in vitro induce AML cells into DC with classical morphology, immunophenotype and function. DC maturity induced by different cytokine cocktails could be variable. Culture-derived DC were originated from the native AML cells. AML cells could make the auto-T lymphocyte anergy.</p>