首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献，搜索用时 31 毫秒
1.
  相似文献   

2.
AIM:To investigate the effects of sodium valproate (VPA) on the cell cycle and apoptosis of chronic myeloid leukemia cell line K562, and to explore the possible mechanisms. METHODS:K562 cells were treated with VPA. Cell cycle and apoptosis were analyzed by flow cytometry. The expression of p21WAF1 mRNA was detected by RT-PCR. RESULTS:After treatment with VPA, cell cycle was arrested obviously at G0/G1 phase ［(82.30±9.41)% vs (40.13±2.12)%, P<0.05］. The apoptotic rate was significantly higher in the cells treated with VPA than that in untreated cells ［(11.47%±0.25%) vs (4.77%±0.40%), P<0.05］. The level of p21WAF1 mRNA was increased ［(1.65±0.91) vs (0.25±0.04), P<0.05］. CONCLUSION:VPA induces elevated expression of p21WAF1 mRNA in K562 cells, resulting in G0/G1 phase arrest and apoptosis in vitro.  相似文献   

3.
AIM: To investigate the characteristic of T-cell acute lymphocytic leukemia 1 (TAL1) gene expression in acute myeloid leukemia (AML) cell lines and in primary AML cells from de novo AML patients with different subtypes. METHODS: Real-time PCR was used to determine the expression of TAL1 mRNA in acute leukemia cell lines (Jurkat, CCRF-CEM, HL-60 and NB4 cell lines) and peripheral blood mononuclear cells from 47 newly diagnosed AML patients. Twelve healthy individuals were served as healthy control group. RESULTS: A significantly increased level in TAL1 mRNA was found in AML cell lines (HL-60 and NB4), T-cell acute lymphacytic leukemia (T-ALL) cell lines (Jurkat, CCRF-CEM) and primary AML cells compared with the healthy controls. Over-expression of TAL1 was found in all detected AML subtypes, the highest level of TAL-1 mRNA was found in AML-M1 and AML-M5 subtype (P<0.05). CONCLUSION: High expression of TAL1 in AML might influence the differentiation and proliferation of myeloid cells, further investigation needs to confirm whether it would be as a biomarker for pathogenesis of AML.  相似文献   

4.
AIM: To investigate the effect of siRNA-induced knockdown of v-ral simian leukemia viral oncogene homolog A(RALA) on proliferation and apoptosis of chronic myelogenous leukemia(CML) K562 cells. METHODS: The chemically synthesized siRNA targeting to RALA gene was transfected into K562 cells using LipofectamineTM 2000. The proliferation and viability of K562 cells were detected by MTT assay and trypan blue dye exclusion. The expression levels of RALA mRNA and protein were determined by quantitative real-time PCR and Western blotting,respectively. The cell apoptosis was analyzed using flow cytometry by double staining with annexin V and propidium iodide, and the apoptotic morphological changes were detected by Hoechst 33258 staining. RESULTS: RALA siRNA significantly down-regulated RALA mRNA and protein expression in K562 cells(P<0.05). The proliferation of K562 cells in RALA siRNA group was inhibited compared with control group(P<0.05). The apoptotic rate was much higher in RALA siRNA group than that in negative control group(P<0.05). The apoptotic morphological changes were observed in the nuclei of K562 cells transfected with RALA siRNA. CONCLUSION: The siRNA-mediated knockdown of RALA results in inhibition of proliferation and induction of apoptosis in K562 cells, indicating that RALA might be used as a potential therapeutic target in chronic myelogenous leukemia.  相似文献   

5.
AIM: The effects of selenium dioxide (SeO2) on proliferation, apoptosis, intracellular reactive oxygen species (ROS) and Ca2+ levels in three leukemia cell lines NB4, K562 and HL-60 were investigated. METHODS: Three leukemia cell lines were treated with 3-30 μmol/L SeO2. Flow cytometry was used to detect apoptosis rate, and analyze the changes of ROS and Ca2+ level within cells. RESULTS: SeO2 at 10 and 30 μmol/L inhibited proliferation in three leukemia cell lines. Treatment with 30 μmol/L SeO2 for 48 h induced 54.0%, 46.5%, 49.6% apoptosis in NB4, K562, and HL-60 cells, respectively, and also markedly decreased ROS and Ca2+ levels among three cell lines. The rate of ROS positive cells in NB4 and HL-60 decreased with the increase in SeO2 concentrations. ROS was clearly reduced with 30 μmol/L SeO2 in K562. Ca2+ levels were tardily declined with 10, 30 μmol/L SeO2 in NB4 and HL-60 cells. Ca2+ levels were clearly reduced with 30 μmol/L SeO2 in K562. CONCLUSION: SeO2 induces apoptosis in three leukemia cells. The declines of intracellular ROS and Ca2+ levels are involved in apoptosis induced by SeO2.  相似文献   

6.
AIM: To transfer 4 full-length WT1 isoforms cDNA into the leukemia cell line NB4 so as to provide a cell model for studying the WT-1 gene function. METHODS: The eukaryotic expression recombinant vectors for WT1 isoforms (pCB6+/WT1) were introduced into the leukemia cell line NB4 by electroporation. The positive cell clones were screened by G418 culture. The integration of WT1 gene isoforms in NB4 cells as confirmed by PCR. The mRNA and protein of WT1 were detected by RT-PCR and Western blotting. RESULTS: WT1 gene isoforms were successfully transferred into NB4 cells. WT1 mRNA and protein expression in the G418-selected cells increased remarkably compared with the control. CONCLUSION: WT1 gene isoforms were effectively transferred into NB4 cells by electroporation and stably expressed in the transfected cells.  相似文献   

7.
AIM: To investigate the relationship between promoter hypermethylation of secreted frizzled-related protein (SFRP) genes and acute leukemia (AL)，and to study the mechanism how 5-aza-2-deoxycytidine (5-Aza-CdR) reverses the hypermethylation of SFRP genes in human AL cell lines. METHODS：Methylation-specific PCR (MSP) was used to detect the methylation levels of SFRP1, SFRP2, SFRP4 and SFRP5 genes in different human AL cell lines (Molt-4, Jurkat, HL-60 and NB4). The methylation levels of these genes in Jurkat cell line before and after 5-Aza-CdR treatment were also analyzed by MSP. The expression of SFRP1, SFRP2, SFRP4 and SFRP5 mRNA was detected by real-time fluorescence quantitative RT-PCR. The mRNA levels of DNA methyltransferase (DNMT) 1, DNMT3A and DNMT3B were analyzed by semiquantitative RT-PCR. RESULTS：None of normal human blood or bone marrow mononuclear cells showed methylation of SFRP genes. Hypermethylation in the promoter regions of SFRP1, SFRP2 and SFRP5 genes was observed in all of the four AL cell lines. SFRP4 gene was totally methylated in NB4, Molt-4 and Jurkat cell lines but partially methylated in HL60 cell line. Treatment with 5-Aza-CdR for 72 h successfully reversed the hypermethylation of SFRP genes, and significantly increased the mRNA expression of SFRP. Moreover, the mRNA expression of DNMT1, DNMT3A and DNMT3B was down-regulated by 5-Aza-CdR in a concentration-dependent manner. CONCLUSION：Methylated SFRP genes may serve as potential independent biomarkers for early detection of AL. 5-Aza-CdR activates SFRP gene expression by demethylation of SFRP genes and down-regulation of DNMT1, DNMT3A and DNMT3B mRNA expression.  相似文献   

8.
  相似文献   

9.
AIM: To evaluate the effect of interfering TGF-β receptor Ⅱ (TβRⅡ) expression on the viability and differentiation of human acute promyelocytic leukemia NB4 cells induced by all-trans retinoic acid (ATRA) and their apoptosis induced by arsenic trioxide (ATO). METHODS: The technique of lentivirus-mediated RNA interference was used to obtain stable NB4 cells with TβRⅡ knockdown, named TβRⅡ-shRNA NB4 cells. CCK-8 assay was used to detect the viability of TβRⅡ-shRNA NB4 cells. The expression level of CD11b was analyzed by flow cytometry, and Wright-Giemsa staining was used to detect the effects of ATRA on the differentiation of TβRⅡ-shRNA NB4 cells. Double staining (Annexin V-FITC/PI) and AO/EB staining were used to detect the effects of ATO on the apoptosis of TβRⅡ-shRNA NB4 cells. RESULTS: The viability of TβRⅡ-shRNA NB4 cells was significantly higher than that of NB4 parental cells. The differentiation was induced in TβRⅡ-shRNA NB4 cells and NB4 parent cells by treatment with ATRA at different concentration (0.01, 0.02, 0.04, 0.08, 0.1 μmol/L) for 96 h. The differentiation rate of TβRⅡ-shRNA NB4 cells was lower than that of NB4 parental cells in a dose-dependent manner. ATO induced apoptosis of TβRⅡ-shRNA NB4 cells and NB4 parent cells at different concentrations (2, 4 and 8 μmol/L) for 24 h. The apoptotic rate of TβRⅡ-shRNA NB4 cells was lower than that of NB4 parental cells dose-dependently. At the concentration of 8 μmol/L for 24 h, the apoptotic rates in TβRⅡ-shRNA NB4 cells and NB4 cells were (49.15±2.05)% and (66.85±2.41)%, respectively (P<0.01). CONCLUSION: Down-regulation of TβRⅡ increases the viability of NB4 cells, inhibits NB4 cell differentiation induced by ATRA, and also inhibits apoptosis induced by ATO.  相似文献   

10.
AIM: To study whether inhibition of forkhead box protein M1(FoxM1) sensitizes leukemia K562 cells to homoharringtonine (HHT). METHODS: K562 cells were incubated with HHT at different concentrations (0μmol/L, 0.015μmol/L, 0.030μmol/L and 0.045μmol/L) for different time (0 h, 24 h, 48 h and 72 h). The mRNA and protein levels of FoxM1 were detected by real-time PCR and Western blot. FoxM1 siRNA was transfected into K562 cells with 0.015μmol/L HHT after 6 h. After 72 h incubation, the cell proliferation was detected by cell counting and soft agar assay, and the proportion of apoptotic K562 cells was determined by flow cytometry. The expression of c-Myc and Sp1 were detected by real-time PCR and Western blot. RESULTS: FoxM1 expression was reduced time-dependently and dose-dependently, suggesting that HHT mediated the downregulation of FoxM1 in K562 cells. In K562 cells, treatment with FoxM1 siRNA and HHT inhibited the cell proliferation and promoted the apoptosis significantly. Therefore, inhibition of FoxM1 sensitized leukemia K562 cells to HHT. The expression of c-Myc and Sp1 was positively regulated by FoxM1. CONCLUSION: HHT inhibits Forkhead box protein M1 expression in K562 cells. Inhibition of FoxM1 sensitizes K562 cells to HHT.  相似文献   

11.
AIM: To investigate whether glucosylceramide synthase (GCS) regulates apoptosis-related gene bcl-2 expression via MEK/ERK signaling pathway, thus enhancing drug resistance of K562/A02 human leukemia multidrug resistant cell line. METHODS: siRNA targeting GCS was transfected into K562/A02 cells. Bcl-2, p-ERK and total ERK expression at mRNA and protein levels after GCS knockdown were detected by real-time PCR and Western blotting. After exposed to MEK-ERK pathway inhibitor U0126, the expression of Bcl-2 at mRNA and protein levels also was analyzed by real-time PCR and Western blotting, respectively. The viability of the cells was evaluated by CCK-8 assay. RESULTS: The expression of GCS and Bcl-2, as well as MEK/ERK signaling were significantly inhibited in K562/A02 cells by GCS siRNA transfection compared with negative control group. Inactivation of MEK/ERK signaling due to U0126 treatment decreased Bcl-2 mRNA and protein levels in a concentration-dependent manner, and sensitized K562/A02 cells to adriamycin. CONCLUSION: GCS may affect the expression of apoptosis-related gene bcl-2 by MEK/ERK signaling pathway, thus regulating multidrug resistance of human leukemia K562/A02 cells.  相似文献   

12.
AIM: To study the effects of phorbol 12-myristate 13-acetate (PMA) on the mRNA expression of BCR/ABL and Fyn in K562 cells, and to explore their relationship. METHODS: The K562 cells were stimulated by PMA at a series of concentrations (1~250 μg/L) for 24 h, and the mRNA expression levels of BCR/ABL and Fyn in K562 cells were detected by real-time fluorescence quantitative PCR. The relative changes of both mRNA expression were measured using 2-ΔΔCt formula. RESULTS: PMA significantly inhibits the mRNA levels of BCR/ABL and Fyn in a dose-dependent manner, and the correlation of these inhibitory effects were significant. Compared with gene Molt-4 cells, the inhibition by PMA was specific for K562 cells. The K562 cells were induced to differentiate to be pseudopodium-like cells. CONCLUSION: The PMA downregulates the mRNA level of Fyn by inhibiting BCR/ABL fusion gene.  相似文献   

13.
AIM: Open reading frame(ORF) of death associated protein kinase1(DAPK1) gene was cloned for studying on tumor forming and metastasis.METHODS: Based on nucleotide sequence of DAPK1 gene from GenBank, a pair of primers was designed. DAPK1 gene ORF was transfected into Raji cells in expression vector pcDNA3.1(+) with lipofectamine reagent. Morphologic assessment of apoptosis was performed with fluorescence microscope cytotoxicity and cell viability was assayed by MTT. RESULTS: DAPK1 gene ORF was amprified from K562 cells by RT-PCR. It was cloned into plasmid pMD18-T and sequenced. There were seven mutation in 4 300 bp nucleotide sequence relatively to DAPK1 nucleotide sequence from GenBank, but six was synonymous mutation and one was single nucleotide polymorphism. 4 300 bp nucleotide of DAPK1 gene ORF was transfected into Raji cells. DAPK1 gene expression was detected in 48 h after it was transfected into Raji cells. Then Raji cells showed apoptosis. CONCLUSION: Large fragment gene was cloned by RT-PCR and transfected into Raji cells successfully. Over-expression of DAPK1 gene induced Raji cells apoptosis.  相似文献   

14.
AIM:To study the expression of telomerase inhibitor Pinx1 in acute leukemia cells and during the differentiation of acute promyelocytic leukemia cells,and to realize its effect on telomerase activity.METHODS:Realtime quantitative PCR with fluorescence probe hybridization was used to measure the expression of Pinx1 and hTERT mRNA in acute leukemia cells and during differentiation of NB4 cells induced by ATRA.The correlations between Pinx1 and hTERT expression were also analyzed.RESULTS:Pinx1 mRNA expression in acute leukemia samples (0.00312,5.42×10-4-0.024) was significantly higher than that in normal bone marrow mononuclear cells (7.89×10-4,0-0.00863,P<0.01).The expression of Pinx1 mRNA had significant positive correlation with hTERT mRNA expression (r=0.296,P<0.05).Pinx1 mRNA expression decreased during differentiation,its expression was positive correlated with hTERT mRNA expression (r=0.900,P<0.05).CONCLUSIONS:As an inhibitor of telomerase,however,Pinx1 also had the same direction of regulation with telomerase activity in acute leukemia cells,suggesting its expression variation may be a subsequent reaction induced by that of hTERT to stabilize telomerase activity.The exact mechanisms remained to be verified.  相似文献   

15.
AIM:To study the effect of p19ARF on the biological behavior of human leukemia cells. METHODS:p19ARF was cloned in eukaryotic expression vector pcDNA3.1 and transferred into INK4a/ARF locus-deficient leukemia cells HEL and K562. The changes in biological characteristics of the two p19ARF-transfected cells were observed.RESULTS:The growth of the p19ARF-transfected HEL cells was significantly inhibited compared with the vector-transfected cells; Cell cycle analysis showed that the expression of foreign p19ARF gene resulted in G1 phase cell cycle arrest and apoptosis cell death in some of HEL cells. However, p19ARF had no marked effects on the growth of K562 cells with p53 gene mutation and did not induce apoptosis in K562 cells.CONCLUSION:p19ARF suppressed the growth of leukemia cells by p53-dependent pathway.  相似文献   

16.
AIM: To study the changes of mitochondrial membrane potential (△ψm) and mitochondrial mass in apoptosis of Jurkat cells induced by camptothecin (CPT). METHODS: Jurkat cells were treated with CPT. Annexin V-FITC/propidium iodine (PI) double stainig was used to detected early stage of apoptosis and PI staining for analyzing the cell cycle. Jurkat cells were stained by annexin V-PE/DiOC6(3) to detect changes of △ψm. The mitochondrial mass was measured by cytometry with NAO staining. RESULTS: 6 h after treated with 10 μmol/L CPT, the rate of early apoptotic cells (22.59±1.04)% had significantly difference compared with control group (3.93±0.73)% (P<0.01). The necrotic rate (2.48±0.53)% had no significant difference to that in control group (2.78±0.63)% (P>0.05). Apoptotic peak appeared obviously after treated with CPT, the percentage of late apoptotic cells (13.58±0.97)% had distinctly difference compared with control group (3.18±0.51)% (P<0.01). The cells in G0/G1 phase (48.14±0.96)% were much higher than that in control group (44.09±0.43)% (P<0.01). Mitochondrial depolarization was very obviously in CPT group. The percentage of annexin V+DiOC6 (3)- cells was (19.47±0.69)%, while in control group, was (4.21±0.40)% (P<0.01). Mitochondrial mass in CPT group was significantly lower than that in control group, the percentage of NAO+ cells (74.77±1.66) % had significantly difference compared with control group (92.24±1.41)% (P<0.01). CONCLUSION: During the process of CPT-induced apoptosis in Jurkat cells, mitochondrial depolarization was very obviously and mitochondrial mass decreased, indicating that the process of apoptosis is nearly related to the mitochondrial pathway.  相似文献   

17.
AIM:To study the effect of bcr- abl gene antisense phosphorothioate oligonucleotides(Aspo) on K562 cell line and explore its significance in chrenic myelogeneous leukemia (CML) gene therapy.METHODS:Cells were exposed to oligomeis, observed by inverted microscope.Cells inhibitory rate were determined by 0.4 trypan blue exclusion . CFU-K562 were cultured in 0.8 % methylcellulose . P210 was measured by flow cytomety RESULTS: K562 cells were treated with Aspo, they still grew in clone state and show antisense sequence specific and dose dependent. When the concentration of Aspo was more than Spznol/L, the growth of cells was inhibited and P210 was down regulated or completely suppressed, and the greatest growth inhibition was at 120h . There was signifi-cant inhibition of cell proliferation in a rang‘cells number from 1×104/mL to 5×104/mL after treatment with 10unol/L Aspo. b2a2 Aspo was also effect on K562 cells which expressing b3a2 mRNA.CONCLUSION: bcr-abl Aspo has a specific growth inhibition effect on K562 cells, and worths further study in CML gene therapy.  相似文献   

18.
AIM To investigate the expression relevance of GATA binding protein-1 (GATA-1) and microR?NA-451a (miR-451a) in erythroid differentiation of human chronic myeloid leukemia K562 cells under hypoxia. METHODS The K562 cells were divided into 2 groups: normoxia group and hypoxia (1% O2) group, and 40 μmol/L hemin chloride was used to induce K562 cell differentiation for 48 and 72 h. The mRNA expression of γ-globin was detected by RT-qPCR, hemoglobin production was observed by benzidine staining, and flow cytometry was used to detect CD235a expression for verifying erythroid differentiation model. The protein expression of GATA-1 during K562 cell differentiation under normoxia and hypoxia was determined by Western blot. RT-qPCR was used to detect the mRNA expression of GATA-1 and the expression level of miR-451a, and their correlation was analysis. The K562 cells were infected by lentivirus for over-expression or knock-down of GATA-1. Meanwhile, the morphological changes of the cells in the above groups were analyzed by Wright-Giemsa staining method to clarify the erythroid differentiation of K562 cells. The expression miR-451a was detected by RT-qPCR after GATA-1 over-expression or knock-down. REULTS: Under normoxia and hypoxia conditions, the expression levels of γ?-globin and CD235a and the positive rate of benzidine staining at 48 and 72 h were significantly higher than those at 0 h (P<0.05).At 72 h, the expression levels of γ?-globin and CD235a and the benzidine staining positive rate in hypoxia group were significantly higher than normoxia group (P<0.05). The expression of GATA-1 mRNA and miR-451a under hypoxia showed an upward trend during the erythroid differentiation of K562 cells, and was significantly higher than that in normoxia group at 72 h (P<0.05). Correlation analysis showed that the mRNA expression of GATA-1 was positively correlated with miR-451a expression under hypoxia (P<0.01). After over-expression of GATA-1 under hypoxia, the expression of γ-globin and CD235a, the positive rate of benzidine staining, and the cell counts of size augmentation, nuclear deflection and nuclear shrinkage at 72 h were significantly higher than those in negative control group (P<0.05). After knock-down of GATA-1 under hypoxia, the expression of γ-globin and CD235a, the benzidine staining positive rate, and the cell counts of size augmentation, nuclear deflection and nuclear shrinkage at 72 h were significantly lower than those in negative control group (P<0.05). Compared with negative control group under hypoxia, the expression of miR-451a was significantly increased after GATA-1 over-expression (P<0.05), while the expression of miR-451a was significantly decreased after GATA-1 knock-down (P<0.05). CONCLUSION Hypoxia increases the expression of GATA-1 and then up-regulates miR-451a to promote erythroid differentiation of K562 cells.  相似文献   

19.
AIM: To investigate the role of Forkhead box M1 (FoxM1) and B-cell leukemia/lymphoma-2 (Bcl-2) in the pathogenesis of acute myeloid leukemia (AML). METHODS: RT-qPCR and immunofluorescence analysis were used to determine the expression of FoxM1 at mRNA and protein levels in AML-de novo patients, AML-complete remission (CR) patients, AML-refractoriness and relapse (RR) patients and healthy controls. HL60 cells and K562 cells were transfected with FoxM1 siRNA. The cell proliferation was detected by cell proliferation assay and colony formation assay on soft agar, and the cell apoptosis was determined by flow cytometry. The expression of FoxM1 and Bcl-2 at mRNA and protein levels was detected by RT-qPCR and Western blotting. The activity of bcl-2 promoter was examined by luciferase reporter assay with FoxM1 targetting. RESULTS: FoxM1 expression level in the AML-de novo patients was significantly higher than that in the healthy controls. As compared with the AML-de novo patients, FoxM1 expression in the AML-CR patients was reduced, and the FoxM1 expression level was the highest in the AML-RR patients. FoxM1 expression was inhibited in the HL60 cells and K562 cells transfected with FoxM1 siRNA. Transfection with FoxM1 siRNA in the HL60 cells and K562 cells inhibited the proliferation as compared with NC siRNA transfection, and impaired the colony formation ability. On the contrary, transfection with FoxM1 siRNA promoted the cell apoptosis. FoxM1 regulated bcl-2 expression positively. CONCLUSION: FoxM1 promotes the development of AML by regulating bcl-2 expression. Silencing of FoxM1 expression suppresses cell proliferation and promotes cell apoptosis. FoxM1 is a potential target for AML treatment.  相似文献   

20.
AIM：To investigate the effect of c-Myc inhibitor 10058-F4 on human chronic myeloid leukemia (CML) K562 cells and imatinib-resistant K562/G cells. METHODS：The protein expression of c-Myc was detected by Western blotting. Cell proliferation was evaluated by MTT assay and colony formation assay. PI staining was used to determine the cell cycle distribution. Annexin V-PI staining was applied for apoptosis detection. RESULTS：Imatinib-resistant K562/G cells displayed lower sensitivity to imatinib than K562 cells with high expression of c-Myc. Treatment with specific c-Myc inhibitor 10058-F4 inhibited the cell proliferation in a dose- and time-dependent manner, and K562/G displayed more sensitivity to 10058-F4 than K562 cells. 10058-F4 also induced cell cycle arrest in G0/G1 phase and induced apoptotic cell death in the 2 cells. Importantly, 10058-F4 suppressed the colony formation ability in K562 and K562/G cells. CONCLUSION：c-Myc is a novel target to overcome imatinib-induced drug resistance, and c-Myc inhibitor provides a new approach in CML therapy.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号