The effect of MPO inhibitor on the gene expression profiles in HL-60 cell lines exposed to 1, 2, 4,-benzenetriol
ABSTRACT: To reveal the gene expression profile changes after exposure to BT, one of benzene metabolites, with or without ABAH, a myeloperoxidase (MPO) inhibitor, and to elucidate how MPO is involved in myelotoxicity of BT, we performed microarray analysis. HL-60 cells suspended in RPMI 1640 containing with 10% of heat-inactivated FBS at 4×10^5/ mL were incubated with or without BT (50 μM) at 37℃ for one or 4 hours. For MPO inhibition experiment, HL-60 (4×10^5 cells/ mL) were pretreatment with 100 μM of ABAH in RPMI 1640/10% of heat-inactivated FBS at 37℃ for 24 hours. Media was then replaced with new media containing the regent plus BT (50 μM) and incubate at 37℃ for one or 4 hours. Unexposed HL-60 cells were used as controls. To validate the microarray results, we selected 22 genes and conducted a real-time PCR. Gene expression alteration induced by 50 μM of BT in HL-60 cells, with or without ABAH, was measured at 1 and 4 hours after exposure to it. Three independent experiments were performed at each time (1 or 4 hours).
Project description:We aim to characterize to effects of the absence of CD40L on neutrophil transcriptome and the effect of soluble CD40L on HL60 cells. For this purpose Total RNA of isolated neutrophils from three CD40L-deficient patients and three healthy controls as well as HL-60 cells from ATCC (HL-60 (ATCC CCL-240) were analyzed by RNAseq. Before RNA obtention, neutrophils were incubated for 2 hours in the presence or absence of 100 U/ml rhIFN- (Immukine, Boehringer Ingelheim), and HL-60 cells cultured for 6 days in the presence or absence of 500 ng/mL sCD40L and/or 1,0 % dimethyl sulfoxide (DMSO).
Project description:GATA2 mutants were discovered in families predisposed to MDS/AML and in sporadic cases of CML-blast crisis. Promyelocytic HL-60 cells were transduced with lentiviral vectors that express GATA2 WT or T354M, 355delT or L359V mutants upon addition of 4-hydroxy tamoxifen (4HT). Microarrays were performed to identify GATA2 WT signatures and differences caused by these mutations. HL-60 cells transduced with GATA2 (WT, T354M, 355delT and L359V) lentiviruses were treated with 4HT for 24 hours and RNA isolated and gene expression measured on Affymetrix microarrays.
Project description:Transcriptional profiling of human leukemia HL-60 cells comparing ATRA treated HL-60 cells with ATRA plus ATO. Goal was to determine the effects of ATO on ATRA induced differentiation of HL-60 cells. Two-condition experiment, ATRA vs. ATRA plus ATO treated HL-60 cells.
Project description:Daunorubicin (DNR) and Cytarabin (Ara-C) are the main chemotherapeutic drugs used for Acute Myeloid Leukemias (AML) treatment. However, their mode of action is not well understood. To decipher if these drug would induce rapid transcriptional reprogramming, we treated HL-60 AML cell line with DNR or Ara-C for 3 hours and carried out a transcriptomic analyses. In addition, we had shown that Reactive Oxigen Species (ROS) produced by NADPH oxidases (NOX) are involved in DNR-induced gene expression. We therefore also performed a transcriptomic analyses in HL-60 cells treated with DNR and VAS2870, an NADPH oxidase inhibitor. HL-60 cells were treated or not for 3 hours with 2 µM Ara-C, 1 µM DNR or 1 µM DNR + 20µM VAS 2870. RNAs were purified from 3 independent experiments and used to probe Affymetrix Human Gene 2.0 ST Genechips
Project description:Curcumin has antileukemic potential that is mediated by dfifferent pathways. Because we could not confirm that the Arylhydrocarbonreceptor, a ligand activated transcription factor, is involved in curcumins antileukemic effects, we performed microarray experiments to have an overall screening of the pathways affected by curcumin in HL-60 cells. We identified an involvement of the vitamin D receptor in the effects of curcumin on HL-60 cells by comparing our results with other experiments. Experiment Overall Design: HL-60 cells were treated for 24h with 100 µM curcumin, 10 nM 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or 0.5 % DMSO as control.
Project description:We investigate the functional complexity of the Plutella xylostella transcriptome in defending against a Bt toxin using Illumina sequencing technology. Over 2,900 differentially expressed unigenes were obtained in resistant P. xylostella comparison to their susceptible counterpart. All the P. xylostella were maintained on cabbage.The susceptible strain (MM) was cultured without exposure to any Bt toxins.Before the sample collected, Cry1Ac-resistant P. xylostella were treated with 750μg/mL Bt toxin Cry1Ac to eliminate the heterozygous individuals. Then the survivors were collected after 48 hours and designed as the resistant sample (MK and GK). Then fourth-instars larvae midgut tissues of MK,GK and MM were collected, respectively, The RNA was extracted and sequenced using Illunima HiSeq 2000.
Project description:All-trans retinoic acid (ATRA)-based differentiation therapy has achieved success with the treatment of acute promyelocytic leukemia (APL), a unique subtype of acute myeloid leukemia (AML). However, other subtypes of AML display resistance to ATRA-based treatment. Here, we demonstrate that a novel natural vibsane-type diterpenoid vibsanine A promotes the differentiation of myeloid leukemia cell lines and primary AML blasts. To reveal how vibsanine A function on promoting myeloid leukemia cell differentiation, we analyzed and compared the gene expression profiles in myeloid leukemia HL-60 cells treated with vibsanine A, PMA, and ATRA. HL-60 cells were treated with vibsanine A, PMA and ATRA for 6 hours or longer up to 24 hours. Gene expression profiling was conducted
Project description:The purpose of the present study was to elucidate in more details the molecular mechanisms of neutrophil-mediated inflammation. We therefore investigated the time-dependent stimulatory potential of LPS from Escherichia coli on cytokine response in neutrophil-like HL-60 cells. <br><br>To get a general overview on the total mRNA changes in DMSO-differentiated HL-60 cells, we performed whole-transcript analysis of LPS-stimulated dHL-60 cells after 2h and 6h of LPS treatment. The samples were collected from three independent experiments from three different passages in cell culture. Non-stimulated dHL-60 cells served as control. We identified the differentially expressed cytokine genes implicated in the human inflammatory response and prominent for their role in neutrophil-mediated inflammatory processes.
Project description:Multiple-condition experiment was desinged to be any number of conditions in an experiment without replicate observations for microarray and used to identify genes differentially expressed between different pairs of conditions (treatments).<br> In this study we used breast cancer stable cell lines for overexpressing and silencing annexin A1 (ANXA1), which belongs to a family of -dependent phospholipid binding proteins and are preferentially located on the cytosolic face of the plasma membrane. Cell lines overexpressing ANXA1 (MDA_MB-453/cDNA) were generated by introducing retroviral vectors containing ANXA1 cDNA (pBabe/ANXA1 cDNA) into breast cancer cell line MDA-MB-453 (a low expressor of ANXA1). Breast cancer cell line BT-474, a high expressor of ANXA1, was infected with ANXA1 siRNA-plasmid viruses to knockdown ANAXAI expressor (BT-474/siRNA) where nucleotides corresponding to siRNA were synthesized and ligated into the pLNCX retroviral vector [35,36]. We also used a pLNCX/U6 empty vector to infect BT-474 and obtained an empty vector expressor. Therefore, 5 breast cancer cell lines (MDA_MB-453, MDA_MB-453/cDNA, BT-474, BT-474/siRNA, and BT-474/U6) are attributed to two genotypes: MDA_MB-453 and BT-474. MCE was performed for microarray analysis with these 5 breast cancer cell lines, that is, only one sample was drawn from each breast cancer cell line.
Project description:We used an in vivo short hairpin RNA (shRNA) screening approach to identify genes that are essential for MLL-AF9 acute myeloid leukemia (AML). We found that Integrin Beta 3 (Itgb3) is essential for murine leukemia cells in vivo, and for human leukemia cells in xenotransplantation studies. In leukemia cells, Itgb3 knockdown impaired homing, downregulated LSC transcriptional programs, and induced differentiation via the intracellular kinase, Syk. In contrast, loss of Itgb3 in normal HSPCs did not affect engraftment, reconstitution, or differentiation. Finally, we confirmed that Itgb3 is dispensable for normal hematopoiesis and required for leukemogenesis using an Itgb3 knockout mouse model. Our results establish the significance of the Itgb3 signaling pathway as a potential therapeutic target in AML. R940406 (R406, the active metabolite of fostamatinib) was supplied by Rigel Pharmaceuticals, Inc., South San Francisco, CA, and AstraZeneca Pharmaceuticals, Wilmington, DE, USA. R406 was resuspended in dimethyl sulfoxide (DMSO) (Sigma-Aldrich) and stored at −80°C. . HL-60, U937 and KG-1 cell lines were purchased from the American Type Culture Collection. MOLM-14 cell lines were provided by Dr. Scott Amstrong (Dana-Farber Cancer Institute, Boston MA, USA.) All cell lines were maintained in RPMI 1640 (Cellgro) supplemented with 1% penicillin-streptomycin and 10% fetal bovine serum (FBS, Sigma-Aldrich) at 37 °C with 5% CO2. MOLM-14, U937, HL-60 and KG-1 cells were grown in 4mM R406 for 24 hours