Objective To explore the method for differentiation induction of leukemia cells into dendritic cells(DC) by A23187 in vitro. Methods Chronic myeloid leukemia K562 cells were cultured with A23187 or cytokine to induce differentiation and form DC. The morphologic features of cells were observed under inverted microscope, the changes of DC surface marks were determined by flow cytometry and RT-PCR, the ability to stimulate lymphocyte proliferation was tested by MTT colorimetry. Results Under the condition of the does (385 ng/ml) of A23187 for four days, some of K562 cells were found in typical dendritic appearance.The expression of DC markers CD1a,CD83 ,HLA-DR,CD86 and CD80 was 6.65 ±2.70,7.37 ±2.40,6.24 ±4.29, 21.60 ± 3.84, 18.52 ± 4.48 repectively, and increased obviously compared with the negative control group(2.80 ±0.52,1.85 ±0.56,2.25 ±0.47,6.69 ±0.83,9.96 ±3.53). The differences had statistical significance (P < 0.05). K562 cells derived from DC acquired the ability to stimulate lymphocyte proliferation.Conclusion A23187 can induce the leukemia cells differenntiation into activated DC-like cells rapidly.

Background and purpose:The expression of ERβin triple negative breast cancer(TNBC) might be associated with good prognosis in TNBC patients. ERβand ERαhave considerable homology. FOXA1 plays an important role in ERαexpression and function. The aim of this study was to analyze the expression of FOXA1 and ERβin TNBC and the relationship between them and the clinicopathologic characteristics and prognosis. Methods:The breast cancer samples in Peking Union Medical College Hospital were collected from Nov. in 2011 to Dec. in 2013, and TNBC were screened out based on the expression of ERα, PR and HER-2. Thirty ERβ-negative samples and 30 ERβ-positive samples were selected randomly according to the ERβexpression. We used immunohistochemical method to detect the expression of FOXA1. Finally, 48 TNBC samples were obtained to analyze the results. Results:The total positive rate of FOXA1 was 35.4%(17/48). In the ERβ-positive group, the positive rate of FOXA1 was 35.7%(10/28),and in the ERβ-negative group, the positive rate of FOXA1 was 35% (7/20). The expression of FOXA1 in these 2 groups had no signiifcant difference (P=0.83), which indicated that there was no relation between ERβand FOXA1. The FOXA1 positive group and FOXA1 negative group also showed no signiifcant difference in age, tumor size, and lymphatic metastasis number in axilla, tumor grade, tumor stage, NPI and DFS. However, Ki-67 showed negative correlation with FOXA1 expression (P<0.01). Conclusion:FOXA1 expression had no relationship with ERβexpression in TNBC. Ki-67 showed negative correlation with FOXA1 expression, which might hint that the proliferation of tumor cell was lower in FOXA1 positive TNBC.

Objective To investigate the effect of TGF-β1/SMAD signaling pathway on K562 cells growth inhibition caused by HMBA. Methods After establishing the in vitro differentiation model with HMBA on K562 cells, the MTT assay was used to detect the proliferation of K562 cells, the cell cycle profile was detected by flow cytometry, and the mRNA expression of TGF-β1, SMAD3, SMAD4 and EVI1 was measured by RT-PCR assay. Results HMBA could inhibit the proliferation and promote the differentiation of K562 cells obviously, which was time and concentration-dependent, and the 72 h corresponding IC50, was about 2 mmol/L. Within 72 h, flow cytometry assay indicated that the ration of G0-G1 phase cells was up-regulated, and the results of RT-PCR showed that relative mRNA expression of TGF-β1, SMAD3 and SMAD4 at mRNA level was increased gradually while that of EVI1 was decreased gradually. Conclusion HMBA can inhibit K562 cells proliferation through TGF-β1/SMAD signaling pathway.

Objective To detect the synergetic effect and mechanism of arsenic trioxide(As2O3)and Trichostatin A(TSA)during inducing apoptosis of HL-60 cells.Methods MTT method was used to test the proliferation of HL-60 cells.Cell cycle and apoptosis were detected by FCM.Semi-quantitative RT-PCR was used to detect the mRNA expression of Bax and Bcl-2 in the cells treated by As2O3 and(or)TSA.Results As2O3 combined with TSA could inhibit proliferation and induce cell cycle arrest at G0 and G1.The percent of apoptosis induced by combination of As2O3 and TSA was obviously higher than that of either As2O3 or TSA.Bax gene expression was increased,while Bcl-2 gene expression was decreased,Bax/Bel-2 ratio was up-regulated.Conclusion Synergetic effect by As2O3 and TSA is remarkable in inducing apoptosis of HL-60 cells.Cell cycte arrest and Bax/Bcl-2 ratio play an important role in apoptosis of HL-60 cells induced by As2O3,TSA or their combination.

Objective To investigate the reversal effect of the monomer of traditional Chinese medicine on muhidrug resistance(MDR) and its possible mechanism in K562/ADM cell line in vitro. Methods With different concentrations of baicalin, geniposide administered to K562/ADM cells, the proliferation of K562/ ADM cells was detected by the MTY assay. Expression of mdr-1 mRNA, Topo Ⅱ mRNA was measured by semi-quantitive RT-PCR. Results Thatbaicalin and geniposide could increase the sensitivity of K562/ADM cells to adriamycin, multiples of reversion were 1.95 times and 1.46 times. The proliferation of K562/ADM cells was in-hibited obviously by baicalin and geniposide, the level of mdr-1 mRNA expression was down-regulated and the Topo Ⅱ mRNA was up-regulated(P<0.01 ). Conclusion Baicalin and geniposide may reverse the multi-drag-resistance of K562/ADM cells, which was related to the down-regulation of mdr-1 expression and up-reg-ulation of Topo Ⅱ beta expression.

Objective To explore molecular mechanism of expression of vascular endothelial growth factor (VEGF) mRNA and secretion of VEGF protein in HL-60 cells induced by all-trans refinoic acid (ATRA). Methods MTr method was used to detect the proliferation of HL-60 cells induced by ATRA,cell cycle and CD11b expression in HL-60 cells were detected by flow cytometry. Expression of VEGF, c-myc, by-poxia-inducible factor(HIF)-lα, matrix metalloproteinase (MMP)-9 and MMP-2 mRNA were detected by semi-quantitative RT-PCR. VEGF protein in HL-60 cells culture supernatant was measured by ELISA before and after being induced by ATRA. Results After treatment with ATRA,the proliferation of HL-60 cells was obviously inhibited, CD11b expression increased, trend of granulocyte directional differentiation emerged, and differentiation degree was increasd(P <0. 05) ;expression level of c-my and VEGF mRNA was down-regulated (P < 0. 05), but expression level of HIF-1α mRNA was up-regulated (P < 0. 05). VEGF protein level in HL-60 cells culture supernatant was decreased by blocking the expression of MMP-9 or MMP-2(P <0. 05).Conclusion VEGF expression has positive correlation with c-myc expression,but has negative correlation with HIF-1α expression. MMP-9 and MMP-2 may be the main factors regulating VEGF secretion in HL-60 cells.

Objective To study the molecular mechanism of different sensitivities to apoptosis induced by low concentration of As2O3 in PML-RARα negative HL-60 cells and PML-RARα positive NB4 cells. Meth-ods NB4 and HL-60 cells were cultured with As2O3 for 1 to 4 days; cell proliferation were detected by MTT method; the apoptosis was detected by flow cytometry,Bcl-2,Bax and Fas mRNA were determined by RT-PCR. Results The proliferation of NB4 cells was inhibited obviously by As2O3(1.0 μmol/L)with the induction of apoptosis( P <0.05) ,which was accompanied by the down-regulation of Bcl-2 mRNA expression( P <0.05)and the ratio of Bcl-2/Bax(P <0.05), but there was no obvious variation of Bax and Fas expression( P >0.05). Inhibition of proliferation and apoptosis were not obvious in PML-RARα negative HL-60 cells induced by low concentration As2O3 ( P >0.05), and there was no obvious variation of Bcl-2, Bax, Fas mRNA expres-sion or Bcl/Bax ratio( P >0.05). Conclusion The ratio of Bcl-2/Bax is contributed to the different sensitiv-ities of PML-RARα negative HL-60 cells and positive NB4 cells induced by low concentration of As2O3.

Objective To evaluate the positive effects of cisplatin on sensitivity of human glioma U251 to tumor necrosis factor-related apoptosis inducing ligand and to investigate the potential mechanism. Methods The expression of green fluorescent protein (GFP) in U251 which was transfected with pAdxsi-GFP-TRAIL was observed by inverted fluorescent microscope ×400) and to ascertain the MOI. The proliferation inhibition was studied by MTT method. Morphological change was detected through inverted florescent microscope and the Hoechst33342 staining assay was used to verify whether cell apoptosis could be induced or not. The cell apoptosis was also analyzed by flow cvtometry with propidium iodide staining. Semi-quantitative RT-PCR was introduced to detect the mRNA expression of apoptosis related gene.Results The expression of TRAIL mRN A was significantly upregulated after transfection. Compared with treatment group of cisplatin and TRAIL alone, the proliferation of U2S1 was significantly inhibited in the cisplatin sensitizing TRAIL group (P < 0.05 ). Nuclear shrinkage and pyknosis fragmentation were observed by Hoechst 33342 staining assay; Apoptotic peak was detected from the results of flow cvtometry and there were significant differences between the sensitizing group and the other two groups ( P < 0.05 ) ; Moreover, the relatively high expression of TRAIL, DR5, caspaseS and down - regulated survivin genes were also observed. There was no significant changes in DR4 expression. Conclusion Cisplatin could extremely enhance the sensitivity of U251 cells to TRAIL And the potential mechanism may related to the increase of TRAIL, DRS, caspaseS genes while the reduction of surivivin gene.

Objective To study effect and mechanism of fungus polysaccharide PSM-a of Polyporus sp.M05 on S180 bearing mice. Methods MTT method was used to detect the inhibiting role of PSM-a on S180 cells proliferation in vitro. S180 mice model was established,and was administered by gavage. Tumor volume was detected, and the ratio of tumor to mile weight and inhibiting tumor rate. The activity of NK and LAK cells on the target cells was analyzed by MTT colorimetric assay ;HE stain was used to detect the necrosis of tumor cells. Results PSM-a could inhibit S180 cells grouth in vitro. PSM-a could decrease the tumor weight and increase the ratio of tumor volume and mice weight; Tumor inhibiting rate reached 80% and above when treated with 250 μg/nml PSM-a. PSM-a could increase the activity of NK and LAK cells, and necrosis happened. Conclusion PSM-a could significantly inhibit the growth of S180 cells, and the mechanism bnay be related with the increased killing activity of immunne cells to tumor cells.

Objective To detect the proliferation inhibition and apoptosis of HL-60 cell induced by APS-1 and APS-2 isolated from Polyporus sp. M05 and to investigate its mechanism. Methods The proliferation inhibition was detected by living cells count method,and chosed proper concentration.Flow cytometry with propidium iodide staining was used to detect cell apoptosis. Semi-quantitative RT-PCR was used to detect the expression of apoptosis related gene. Results APS-1 and APS-2 could significantly inhibit the proliferation of HL-60 cells on a time and dose dependent manner. Apoptosis ratio increased to 47. 9% and 26. 8% after HL-60 cells were exposed to APS-1 and APS-2 respectively for 48 h,and the differences had statistical significance (P <0.01). After being induced by APS-1,mRNA of Bax,Fas,Caspase-3 was upregulated. And after being induced by APS-2,mRNA of Bax,Caspase-3 was upregulated,while Fas mRNA did not change. Conclusion APS-1 and APS-2 can inhibit the proliferation and induce apoptosis of HL-60 cells. Mechanism of HL-60 cell apoptosis induced by APS-1 is related to both mitochondrial pathway and Fas signaling pathway,while apoptosis induced by APS-2 is only related to mitochondrial pathway.