ABSTRACT
AIM:To study the growth-inhibiting and proapoptotic effects of Pim-1 kinase inhibitor SMI-4a on human acute myeloid leukemia cell line U937.METHODS:The effect of SMI-4a on U937 cell viability was measured by CCK-8 assay.The apoptotic rate was assessed by flow cytometry with Annexin V-PI staining and by fluorescence microscopy with Hoechst 33342 staining.Methylcellulose was used to assess colony formation ability of the cells.The expression of β-catenin in the cell cytosol and nucleus was detected by Western blot,and the expression of apoptosis-related proteins in the U937 cells was also examined.Intracellular distribution of β-catenin was detected by the method of immunofluorescence.RESULTS:SMI-4a inhibited the viability of U937 cells.Annexin V-PI staining showed that SMI-4a induced apoptosis in dose-and time-dependent manners.Hoechst 33342 staining also verified the apoptosis.SMI-4a significantly inhibited the colony formation capacity of the U937 cells.The results of Western blot demonstrated that SMI-4a upregulated the expression of PARP and Bax,downregulated the expression of Bcl-2 and change the distribution of β-catenin in intracellular compartment.Immunofluorescence observation found that SMI-4a decreased the expression level of β-catenin in the U937 cells.CONCLUSION:SMI-4a induces U937 cell apoptosis through regulating the expression of apoptosis-related genes.
ABSTRACT
AIM:To study the growth-inhibiting and proapoptotic effects of Pim-1 kinase inhibitor SMI-4a on human acute myeloid leukemia cell line U937.METHODS:The effect of SMI-4a on U937 cell viability was measured by CCK-8 assay.The apoptotic rate was assessed by flow cytometry with Annexin V-PI staining and by fluorescence microscopy with Hoechst 33342 staining.Methylcellulose was used to assess colony formation ability of the cells.The expression of β-catenin in the cell cytosol and nucleus was detected by Western blot,and the expression of apoptosis-related proteins in the U937 cells was also examined.Intracellular distribution of β-catenin was detected by the method of immunofluorescence.RESULTS:SMI-4a inhibited the viability of U937 cells.Annexin V-PI staining showed that SMI-4a induced apoptosis in dose-and time-dependent manners.Hoechst 33342 staining also verified the apoptosis.SMI-4a significantly inhibited the colony formation capacity of the U937 cells.The results of Western blot demonstrated that SMI-4a upregulated the expression of PARP and Bax,downregulated the expression of Bcl-2 and change the distribution of β-catenin in intracellular compartment.Immunofluorescence observation found that SMI-4a decreased the expression level of β-catenin in the U937 cells.CONCLUSION:SMI-4a induces U937 cell apoptosis through regulating the expression of apoptosis-related genes.