| Objectives:Acute myelogenous leukemia(acute myeloid leukemia,AML)is a common malignant tumor of hematopoietic system,and is the most common kind of leukemia in adults.the main performance for myeloid progenitor cells to differentiate different levels of maturity disorder and malignant bone marrow cells infinite proliferation and apoptosis.In recent years,as the incidence and mortality of leukemia have increased gradually,the need for research and treatment of leukemia drugs and methods has become more urgent.Although along with the continuous improvement of chemotherapeutic drugs and molecular target therapy of progress,and gradually development of hematopoietic stem cell transplantation and improvement the survival rate of leukaemia,the bone marrow hematopoietic stem cell transplantation due to the strict,more expensive and complications and risk are restricted by some disadvantages such as wide.Therefore,it is very important to explore new anticancer drugs for the clinical treatment of leukemia and the survival quality of patients.Study abroad in recent years,bromo domain structure(bromo domain inhibitors,BRD4)inhibitors JQ1 have stronger inhibitory effect of leukemia cells,it may inhibit MYC transcription in the role of inhibition of cell proliferation.However,the proliferation of cells involves the activation of various models,especially the jak-stat signal transduction pathway.Therefore,this research with different concentrations of JQ1 role in U937-AML cell lines,through the cell inhibition rate and STAT5 signal transduction pathways related important molecular FAK the expression of PAK1 detection,discusses JQ1 in leukemia cells,the mechanism of inhibition and indirect understanding at the same time affect the AML STAT5 signal path in the process of cell proliferation,in order to Provide a new perspective for AML therapy.Methods:1.4 methyl azazole blue(MTT)method to detect the proliferation inhibition rate of JQ1 in U937 cell lines;2.Flow cytometry tests the apoptosis rate of U937 cells;3.Real-time fluorescence quantitative polymerase chain reaction(rt-pcr)method to detect the expression of FAKand PAK1 mRNA;4.Western Blot detects stat-5 expression.Results:1.Determined by MTT test results:U937 cell lines and different concentrations of JQ1 jointly develop after 24 h,48h,72 h,different concentration of JQ1 has obvious inhibitory effect to the U937 cells proliferation,and cell proliferation inhibition rate of the experimental group with the increase of drug concentration and prolonged effect,rendering time-dose dependent.2.Flow cytometry test result shows: different drug concentrations of JQ1 intervention U937 cells after 24 h,48h,results show that the 24 h time point control early apoptosis rate was(0.44±0.07)%,while the observation group JQ1 in 0.2,1.0,4.0 mu mol/L concentration early apoptosis rate(2.72±0.53)%,respectively(5.29±0.66)%,(8.83± 0.28)%.The early apoptosis rate of 48 h control group was(0.91±0.29)%,and the different concentration groups of the experimental group JQ1were(5.79 ± 0.27)%,(12.03 ± 0.80)%,(15.89 ± 0.79)%.Through the single factor analysis of variance of statistics,according to the results of the same point in time there were significant differences between groups,the concentration and the concentration of the same set of apoptosis rate with the extension of time increased significantly,compared with the control group was statistically significant(P < 0.05),characterized by time-dose dependent.3.Real-time fluorescence quantitative PCR test results showed that after 48 h of different drug concentrations,rt-pcr detected that the dissolution curves of FAK and PAK1 related genes showed a single amplification curve,with goodspecificity.Results show that the observation group of FAK(PAK1 quantity than the control group dropped significantly,gene expression and gene expression quantity change trend with the increase of drug concentration significantly(F value were454.651,2434.610,P values < 0.001),there are statistically significant differences between group(P < 0.05).4.Western Blot test showed that different drug concentrations of JQ1 U937 cells after 48 h,compared with the control group,experimental group of STAT-5 protein expression level with the increase of drug concentration protein relative content gradually reduced,while the control group the STAT-5 protein expression level increased significantly.During the same period,the stat-5 protein,which was phosphorylated,showed a decrease in the expression level of the non-phosphorylated stat-5 protein.(F value were 263.135,20.658,P values < 0.001),there are statistically significant differences between group(P < 0.05).Conclusion:1.JQ1 can significantly inhibit the proliferation of U937 cells and show time and dose dependence.2.JQ1 can effectively induce apoptosis of U937 cells,and its apoptosis effect is also time-dependent and dose-dependent.3.JQ1 can significantly inhibit the FAK gene and PAK1 gene in U937 cells,and promote cell apoptosis by inhibiting the above gene regulation of STAT-5 signal.4.Western Blot test showed that JQ1 can significantly inhibit the expression level of STAT-5 in U937 cells,thus achieving the effect of the progress of the leukemia cells.5.JQ1 may in time dose dependent inhibition of proliferation of U937 cells and promote its apoptosis,the mechanism may be JQ1 of FAK(,the influence of PAK1 important molecules to inhibit the STAT-5 ways to one of the ways to promote apoptosis of U937 cells. |
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