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European Review For Medical and... Aug 2019To explore the regulatory role of PLZF in the malignant phenotype of non-APL acute myeloid leukemia (AML) and its underlying mechanism.
OBJECTIVE
To explore the regulatory role of PLZF in the malignant phenotype of non-APL acute myeloid leukemia (AML) and its underlying mechanism.
MATERIALS AND METHODS
The expression level of PLZF in AML cell lines KG-1a, HL-60, OCI-AML3, THP-1 and K562 was detected by quantitative Polymerase Chain Reaction (qPCR) and Western blot, respectively. Subsequently, THP-1 cells were divided into mock group (no treatment), scramble group (transfection with scramble shRNA) and shPLZF group (transfection with shPLZF). THP-1 cell line stably expressing shPLZF was constructed, followed by determination of its transfection efficiency by qPCR and Western blot, respectively. The proliferation and colony formation of THP-1 cells were accessed using CCK-8 (cell counting kit-8) assay and colony formation assay, respectively. The apoptotic rate in THP-1 cells was determined using flow cytometry. Protein levels of apoptosis-related genes in THP-1 cells were detected by Western blot. Finally, protein levels of AKT, Foxo3a, pAKT and pFoxo3a were detected by Western blot as well.
RESULTS
Both mRNA and protein levels of PLZF were relatively high in THP-1 cells, and were selected for the following experiments. After construction of THP-1 cell line stably expressing shPLZF, proliferative rate and colony formation abilities increased in the shPLZF group compared with the mock group and the scramble group. We found a decreased apoptotic rate, downregulated Bax and upregulated Bcl-2 in the shPLZF group than those of the mock group and scramble group. Phosphorylation levels of AKT and Foxo3a increased after interference with PLZF, whereas no significant changes in total levels of AKT and Foxo3a were observed.
CONCLUSIONS
PLZF inhibits the malignant phenotype of AML by regulating the AKT/Foxo3a pathway.
Topics: Apoptosis; Cell Proliferation; Forkhead Box Protein O3; HL-60 Cells; Humans; K562 Cells; Leukemia, Myeloid; Promyelocytic Leukemia Zinc Finger Protein; Proto-Oncogene Proteins c-akt; Signal Transduction
PubMed: 31378879
DOI: 10.26355/eurrev_201908_18522 -
Fitoterapia Jun 2021A phytochemical investigation of a medicinal plant Artemisia atrovirens was carried out, resulting in the characterization of a novel bis-nor seco-guaianolide,...
A phytochemical investigation of a medicinal plant Artemisia atrovirens was carried out, resulting in the characterization of a novel bis-nor seco-guaianolide, seco-atrovirenolide A (1), a new 1,10-seco-guaianolide derivative, seco-atrovirenoic acid A (2), and a new artifact 10-methanoyloxy-seco-atrovirenoic acid A (3), together with eight known guaianolide and seco-guaianolide derivatives (4-11). The structures of new compounds were fully established by extensive analysis of MS, 1D and 2D NMR spectroscopic data. The absolute configurations of the isolated compounds were confirmed by TDDFT ECD calculation, Mosher's method, and X-ray crystal diffraction experiment. All the compounds were tested in vitro for their cytotoxicity against HL-60 and A549 cell lines. Some of them showed moderate inhibitory activity against HL-60 cell lines with IC values ranging from 5.99 to 11.74 μM.
Topics: A549 Cells; Antineoplastic Agents, Phytogenic; Artemisia; China; HL-60 Cells; Humans; Limonins; Molecular Structure; Phytochemicals; Plants, Medicinal
PubMed: 33781859
DOI: 10.1016/j.fitote.2021.104900 -
Biomeditsinskaia Khimiia Dec 2023Plasma membrane proteins with extracellular-exposed domains are responsible for transduction of extracellular signals into intracellular responses, and their...
Plasma membrane proteins with extracellular-exposed domains are responsible for transduction of extracellular signals into intracellular responses, and their accessibility to therapeutic molecules makes them attractive targets for drug development. In this work, using omics technologies and immunochemical methods, we have studied changes in the content of markers of clusters of differentiation (CD markers) of neutrophils (CD33, CD97, CD54, CD38, CD18, CD11b, CD44, and CD71) at the level of transcripts and proteins in NB4, HL-60 and K562 cell lines, induced by the treatment with all-trans-retinoic acid (ATRA). Transcriptomic analysis revealed the induction of CD38, CD54, CD11b, and CD18 markers as early as 3 h after the addition of the inducer in the ATRA-responsive cell lines HL-60 and NB4. After 24 h, a line-specific expression pattern of CD markers could be observed in all cell lines. Studies of changes in the content of CD antigens by means of flow cytometry and targeted mass spectrometry (MS) gave similar results. The proteomic profile of the surface markers (CD38, CD54, CD11b, and CD18), characteristic of the NB4 and HL-60 lines, reflects different molecular pathways for the implementation of ATRA-induced differentiation of leukemic cells into mature neutrophils.
Topics: Humans; Leukemia, Promyelocytic, Acute; Proteomics; Tretinoin; HL-60 Cells; Cell Differentiation
PubMed: 38153053
DOI: 10.18097/PBMC20236906383 -
Biomolecules Oct 2022Autophagy is a fundamental catabolic process of cellular survival. The role of autophagy in cancer is highly complex: in the early stages of neoplastic transformation,...
Autophagy is a fundamental catabolic process of cellular survival. The role of autophagy in cancer is highly complex: in the early stages of neoplastic transformation, it can act as a tumor suppressor avoiding the accumulation of proteins, damaged organelles, and reactive oxygen species (ROS), while during the advanced stages of cancer, autophagy is exploited by cancer cells to survive under starvation. 6-(Methylsulfonyl) hexyl isothiocyanate (6-MITC) is the most interesting compound in the rizhome. Recently, we proved its ability to induce cytotoxic, cytostatic, and cell differentiation effects on leukemic cell lines and its antimutagenic activity on TK6 cells. In the current study, to further define its chemopreventive profile, Jurkat and HL-60 cells were treated with 6-MITC for 24 h. The modulation of the autophagic process and the involvement of ROS levels as a possible trigger mechanisms were analyzed by flow cytometry. We found that 6-MITC induced autophagy in Jurkat and HL-60 cells at the highest concentration tested and increased ROS intracellular levels in a dose-dependent manner. Our results implement available data to support 6-MITC as an attractive potential chemopreventive agent.
Topics: Humans; Reactive Oxygen Species; Cytostatic Agents; Isothiocyanates; Leukemia; Autophagy; HL-60 Cells; Apoptosis; Cell Line, Tumor
PubMed: 36291694
DOI: 10.3390/biom12101485 -
Experimental Hematology May 2020Mds1-Evi1 (also known as Prdm3) and Prdm16 are two highly related zinc finger transcription factors that, within the hematopoietic system, are both expressed primarily...
Mds1-Evi1 (also known as Prdm3) and Prdm16 are two highly related zinc finger transcription factors that, within the hematopoietic system, are both expressed primarily in hematopoietic stem cells (HSCs). Our laboratory previously found that constitutive Mds1-Evi1 knockout mice are viable, but their HSCs are unable to withstand myeloablative chemotherapy or effectively transplant irradiated recipient mice. A similar phenotype has been observed for Prdm16, except that the Prdm16 constitutive knockout is lethal. Here, we created a novel double-knockout model of Mds1-Evi1 and Prdm16 in the bone marrow, in which double knockout occurs only in cells that endogenously express Mds1-Evi1 and only upon induction with tamoxifen. We show that combined Mds1-Evi1/Prdm16 deficiency causes bone marrow failure within 15 days, with rapid loss in all progenitor compartments, while the peripheral blood exhibits progressive reductions in peripheral monocytes and granulocytes. We found that surviving hematopoietic stem cells and granulocytic progenitors had elevated apoptosis and cell division, and were unable to form colonies in vitro; adding back wild-type Mds1-Evi1 or Prdm16 to double-knockout bone marrow restores colony formation, and for MDS1-EVI1, this activity depends on a functional PR domain. All of these phenotypic effects were exhibited at milder levels in Mds1-Evi1 and Prdm16 single-knockout controls. Overall, these results illustrate that Mds1-Evi1 and Prdm16 play additive roles in maintaining normal hematopoietic stem cell survival.
Topics: Animals; Apoptosis; Cell Line; Cell Survival; DNA-Binding Proteins; Granulocyte Precursor Cells; Hematopoiesis; MDS1 and EVI1 Complex Locus Protein; Mice; Mice, Knockout; Models, Biological; Transcription Factors
PubMed: 32437910
DOI: 10.1016/j.exphem.2020.04.010 -
Nutrition and Cancer 2022Acute myeloid leukemia is characterized by abnormal differentiation of hematopoietic stem cells, leading to the accumulation of immature myeloid cells. Differentiation...
Acute myeloid leukemia is characterized by abnormal differentiation of hematopoietic stem cells, leading to the accumulation of immature myeloid cells. Differentiation therapy has been a successful treatment option for acute promyelocytic leukemia but suffers from adverse effects. Therefore, search for novel differentiation-inducing agents with minimal side effects is desirable. Securinine, a naturally-occurring alkaloid, induces differentiation in various leukemic cells and apoptosis in other types of cancers. However, the underlying molecular mechanism(s) remain elusive. Our study aimed to elucidate the possible molecular mechanism(s) and signaling events involved in securinine-induced differentiation of HL-60 cells. Securinine inhibited proliferation in a time- and dose-dependent manner and triggered differentiation. A higher CD14+ population indicated maturation toward monocytic lineage. Securinine caused cell cycle arrest at the G0/G1 phase and enhanced ROS generation. Quantitative gene expression analysis showed significant down-regulation of , , , and and up-regulation of gene. The expression of distinct protein kinases Lyn, Chk-2, Yes, FAK, c-Jun, and JNK were enhanced. Use of specific inhibitors of crucial intracellular signaling proteins indicated that JNK and ERK blockade resulted in a significant decline in differentiation. These data thus confirm that securinine induces differentiation through the activation of the JNK-ERK signaling pathway in HL-60 cells.
Topics: Azepines; Cell Differentiation; HL-60 Cells; Heterocyclic Compounds, Bridged-Ring; Humans; Lactones; MAP Kinase Signaling System; Piperidines
PubMed: 33998358
DOI: 10.1080/01635581.2021.1925710 -
Molecules (Basel, Switzerland) May 2021A new series of mollugin-1,2,3-triazole derivatives were synthesized using a copper(I)-catalyzed Huisgen 1,3-dipolar cycloaddition reaction of corresponding...
A new series of mollugin-1,2,3-triazole derivatives were synthesized using a copper(I)-catalyzed Huisgen 1,3-dipolar cycloaddition reaction of corresponding -propargylated mollugin with aryl azides. All the compounds were evaluated for their cytotoxicity on five human cancer cell lines (HL-60, A549, SMMC-7721, SW480, and MCF-7) using MTS assays. Among the synthesized series, most of them showed cytotoxicity and most of all, compounds and exhibited significant cytotoxicity of all five cancer cell lines.
Topics: A549 Cells; Antineoplastic Agents; Azides; Cell Line, Tumor; Drug Screening Assays, Antitumor; HL-60 Cells; Humans; Inhibitory Concentration 50; MCF-7 Cells; Magnetic Resonance Spectroscopy; Molecular Structure; Pyrans; Structure-Activity Relationship; Triazoles
PubMed: 34071319
DOI: 10.3390/molecules26113249 -
American Journal of Clinical Pathology Sep 2021Myelodysplastic/myeloproliferative neoplasms (MDS/MPN) are a group of rare and heterogeneous hematopoietic disorders that frequently present a diagnostic challenge. Here...
OBJECTIVES
Myelodysplastic/myeloproliferative neoplasms (MDS/MPN) are a group of rare and heterogeneous hematopoietic disorders that frequently present a diagnostic challenge. Here we present our institutional experience with next-generation sequencing (NGS), together with morphologic, flow cytometric, and cytogenetic evaluation, in the diagnosis of MDS/MPN, with particular emphasis on MDS/MPN unclassifiable (MPN-U).
METHODS
We evaluated the morphologic, flow cytometric, cytogenetic, and molecular characteristics of all MDS/MPN cases that underwent NGS at our institution between April 2016 and February 2019.
RESULTS
Thirty-seven cases of MDS/MPN were identified, including 14 cases of MDS/MPN-U. Ninety-seven percent harbored mutations and immunophenotypic aberrancies (36/37), while only 38% had cytogenetic abnormalities (12/32). The MDS/MPN-U group had the highest rate of myeloblast phenotypic abnormalities and had a high mutation rate of approximately 2.7 mutated genes per case, most commonly in JAK2, SRSF2, and ASXL1.
CONCLUSIONS
No single ancillary study was abnormal in every case, but all cases had at least one abnormal finding, demonstrating the usefulness of a multiparameter approach to the diagnosis of MDS/MPN. Although a few specific mutations were found exclusively in MDS/MPN-U and JAK2 mutations were most prevalent, larger studies are needed to determine whether MDS/MPN-U has a mutational "fingerprint," which may aid in diagnosis and targeted therapy.
Topics: Adult; Aged; Aged, 80 and over; Cytogenetics; Female; Flow Cytometry; Granulocyte Precursor Cells; High-Throughput Nucleotide Sequencing; Humans; Immunophenotyping; Male; Middle Aged; Mutation; Mutation Rate; Myelodysplastic-Myeloproliferative Diseases; Myeloproliferative Disorders; Sequence Analysis, DNA; Young Adult
PubMed: 33877292
DOI: 10.1093/ajcp/aqaa281 -
The Journal of Gene Medicine Oct 2021Pre-existing immunities hamper the application of human adenovirus (HAdV) vectors in gene therapy or vaccine development. Fowl adenovirus (FAdV)-based vector might...
BACKGROUND
Pre-existing immunities hamper the application of human adenovirus (HAdV) vectors in gene therapy or vaccine development. Fowl adenovirus (FAdV)-based vector might represent an alternative.
METHODS
An intermediate plasmid containing FAdV-4 fiber genes, pMD-FAV4Fs, was separated from FAdV-4 adenoviral plasmid pKFAV4GFP. An overlap extension polymerase chain reaction (PCR) was employed for fiber modification in pMD-FAV4Fs, and the modified fibers were restored to generate new adenoviral plasmids through restriction-assembly. FAdV-4 vectors were rescued and amplified in chicken LMH cells. Fluorescence microscopy and flow cytometry were used to evaluate the gene transfer efficiency. The amount of viruses binding to cells was determined by a real-time PCR. A plaque-forming assay and one-step growth curve were used to evaluate virus growth.
RESULTS
Four sites in the CD-, DE-, HI- and IJ-loop of fiber1 knob could tolerate the insertion of exogenous peptide. The insertion of RGD4C peptide in the fiber1 knob significantly promoted FAdV-4 transduction to human adherent cells such as 293, A549 and HEp-2, and the insertion to the IJ-loop demonstrated the best performance. The replacement of the fiber2 knob of FAdV-4 with that of HAdV-35 improved the gene transfer to human suspension cells such as Jurkat, K562 and U937. Fiber-modified FAdV-4 vectors could transduce approximately 80% human cells at an acceptable multiplicity of infection. Enhanced gene transfer mainly resulted from increased virus binding. Fiber modifications did not significantly influence the growth of recombinant FAdV-4 in packaging cells.
CONCLUSIONS
As a proof of principle, it was feasible to enhance gene transduction of FAdV-4 vectors to human cells by modifying the fibers.
Topics: A549 Cells; Adenoviruses, Human; Cell Line; Cell Line, Tumor; Genetic Therapy; Genetic Vectors; HEK293 Cells; HL-60 Cells; Humans; Jurkat Cells; Plasmids; Transduction, Genetic; U937 Cells; Vaccine Development
PubMed: 34050587
DOI: 10.1002/jgm.3368 -
G3 (Bethesda, Md.) Aug 2019Tumor necrosis factor alpha (TNF-α) is a potent cytokine involved in systemic inflammation and immune modulation. Signaling responses that involve TNF-α are context...
Tumor necrosis factor alpha (TNF-α) is a potent cytokine involved in systemic inflammation and immune modulation. Signaling responses that involve TNF-α are context dependent and capable of stimulating pathways promoting both cell death and survival. TNF-α treatment has been investigated as part of a combined therapy for acute myeloid leukemia due to its modifying effects on all-trans retinoic acid (ATRA) mediated differentiation into granulocytes. To investigate the interaction between cellular differentiation and TNF-α, we performed RNA-sequencing on two forms of the human HL-60/S4 promyelocytic leukemia cell line treated with TNF-α. The ATRA-differentiated granulocytic form of HL-60/S4 cells had an enhanced transcriptional response to TNF-α treatment compared to the undifferentiated promyelocytes. The observed TNF-α responses included differential expression of cell cycle gene sets, which were generally upregulated in TNF-α treated promyelocytes, and downregulated in TNF-α treated granulocytes. This is consistent with TNF-α induced cell cycle repression in granulocytes and cell cycle progression in promyelocytes. Moreover, we found evidence that TNF-α treatment of granulocytes shifts the transcriptome toward that of a macrophage. We conclude that TNF-α treatment promotes a divergent transcriptional program in promyelocytes and granulocytes. TNF-α promotes cell cycle associated gene expression in promyelocytes. In contrast, TNF-α stimulated granulocytes have reduced cell cycle gene expression, and a macrophage-like transcriptional program.
Topics: Biomarkers; Databases, Genetic; Gene Expression Profiling; Gene Expression Regulation, Leukemic; Genes, cdc; HL-60 Cells; Humans; Leukemia, Promyelocytic, Acute; Transcriptome; Tumor Necrosis Factor-alpha
PubMed: 31263060
DOI: 10.1534/g3.119.400361