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Objective To investigate the effect and the mechanism of different G-CSF-priming protocols on leukemia cell lines (HL-60 and U937) in vitro and provide the clinical guidance to clinical treatment of acute leukemia.Methods The leukemia cell lines HL-60 and U937 were used as model to detect the effects of three drugs alone and combined two drugs (HA) or three drugs (HAG) respectively.Cell viability and cell growth inhibition were performed by cell count kit-8 (CCK-8) assay.Apoptotic marker AnnexinV/PI,cell membrane surface antigen CD11b,cell cycle,mitochondrial membrane potential (JC-1) and Caspase-3 were determined by flow cytometry.Results After using of HAG for 48 h,HL-60 and U937 cells counts were decreased significantly and the apoptotic marker Annexin V was significantly increased. To compare the single drug group with two drug combination group,the result was significantly different (P ＜0.05),and the apoptosis of U937 cells was higher than HL-60 cell line.CD11b expression among the three groups did not change (P ＞ 0.05).Using of CAG and MAG,the mitochondrial nembrane potential of HL-60 and U937cells was increased,the three-drug combination group was significantly higher than single-drug group and control group (P ＜0.05); Caspase-3 was activated,the fluorescence intensities of Caspase-3 of the three-drug combination group and single drug group were significantly higher (P ＜0.05) comparing with the control group.Conclusion HAG regimen could induce leukemia cells to apoptosis through the reduction of mitochondrial membrane potential and the activation of Caspase-3 to induce apoptosis of leukemia cells.
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Objective To investigate the effect of arsenic trioxide on expression VEGF of lymphoma cells. Methods The VEGF mRNA expression was analysed by by Real-time PCR, and VEGF protein expression in Raji and Jurkat lymphoma cell lines by ELISA. Results ATO can inhibit lymphoma cells by inducing apoptosis. ATO induced lymphoma cell apoptosis was due to time.With the period of ATO effecting on cells goes, the expression of VEGF mRNA and the protein were down-regulated significantly (after 24, 48, 72 h). There were, the VEGF mRNA △△Ct data of treated with ATO, at 12 h, for Raji: 0.75±0.15, 72 h, Jurkat: 1.67±0.13. After 72 h, Raji: 8.95±0.38; Jurkat: 9.09±0.16 (f =3.54, P <0.01; t =3.65, P <0.01). And about the VEGF protein, at 12 h, Raji: 198.38±4.37; Jurkat: 563.11±3.81. After 72 h, Raji: 23.55±2.06; Jurkat: 57.11 ±3.88 (t =2.48, P <0.05; t =2.59, P <0.05). Conclusion ATO can inhibit the proliferation of lymphoma cells by down-regulating the expression of VEGF mRNA and its protein.
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Objective To evaluate the growth inhibition and apoptosis of human monocytic leukemia THP-1 cell line by using 5,8-dimethyl-2-β-hydroxyisovalerylshikonin (SK36) and explore its preliminary mechanism. Methods CCK colorimetric assay and cell counting was used to examine the growth inhibition of shikonin on THP-1 cells. The apoptosis of THP-1 cells was detected by Annexin V/PI double labeling. The activation of Caspase-3 apoptosis pathway was determined by FCM. The apoptosis and the necrosis of THP-1 cells were detected by the laser scanning confocal microscopy. Results When the THP-1 cells were treated with SK36 at 1.02 μg/ml for 24 h and 48 h, the growth inhibition was dose-dependent. The cell apoptotic rate of THP-1 cells treated with 1.02 μg/ml evaluated by FCM with Annexin V/PI double labeling staining were (40.61 ±2.13) % and (67.40±9.15) % at 24 h and 48 h after treatment, respectively, which were significantly higher than that of the control group [(16.97±0.61) %] ([ = 18.444, t = 9.528, P <0.01). SK36 could induce THP-1 cells apoptosis involving the activation of Caspase-3 (F= 323.61, P<0.01). Conclusion SK36 can induce human THP-1 cells to undergo apoptosis, and its primary mechanism was to activate the Caspase-3.
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Background and purpose:Recombinant human granulocyte colony-stimulating factor(rhG-CSF)is a kind of polypeptide.It can stimulate the haemopoietic stem cell and granule progenitor cell to differentiate and proliferate.The neutrophil can be accelerated to release and promote its function of phagocytosis,chemotaxis and oxidative burst by rhG-CSF.In this study,we aimed to explore the effect of rhG-CSF on morphology、phenotype and function of neutrophil in patients with malignant lymphoma after treatment of chemotherapy.Methods:White blood cell(WBC) was counted by the method of COULTER.The morphology of neutrophil was observed on Wright's-stained peripheral blood smear.The phagocytotic function and chemotaxis of neutrophil were assessed by either measuring the amount of hydrogen peroxide or by the agarose method.Oxidative burst and phenotype of neutrophil was analysed by flow cytometry using immunofluorescence technique.Results:①There was Hypogranular in some neutrophilic cytoplasm after chemotherapy,the number of "toxic" granulation,vacuoles and Dohle bodies increased from 5.4%,3.2%,0.5% to 55.0%,11.8%,4.1% in neutrophils from patients with malignant lymphoma after rhG-CSF administration,.②Compared to normal control,5.76?mol/LH_(2)O_(2).10~(-6)PMN,1.85,76.4,the neutrophil functions of phagocytosis,chemotaxis,oxidative burst were impaired after chemotherapy 2.73?mol/LH_(2)O_(2).10~(-6)PMN,0.97,51.8%,rhG-CSF increased the function,7.02?mol/LH_(2)O_(2).10~(-6)PMN,1.89,78.4%,the neutrophil function from these patients had returned to normal or were even exceeded.③ A mild increase of CD64 expression from 5.4%,3.2%,0.5% and a significant decrease of CD62L expression from 7.47% to 3.95% were found in patients after rhG-CSF treatment.No changes of CD16,CD32,CD14 and CD11b expression were detected in these patients before or after G-CSF administration.Conclusions:rhG-CSF administration can modify the morphology,phenotypeand partially recover the impaired function of neutrophil from the patients with Malignant Lymphoma after chemotherapy,andit may improve anti-infection ability of the human body.