Dose- and Time-Dependent Response of Human Leukemia (HL-60 Cells to Arsenic Trioxide Treatment

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Paul B. Tchounwou

2006-06-01

Full Text Available The treatment of acute promyelocytic leukemia (APL has been based on the administration of all-trans retinoic acid plus anthracycline chemotherapy, which is very effective as first line therapy; however 25 to 30% of patients will relapse with their disease becoming refractory to conventional therapy. Recently, studies have shown arsenic trioxide to be effective in the treatment of acute promyelocytic leukemia. In this study, we used the human leukemia (HL-60 cell line as a model to evaluate the cytoxicity of arsenic trioxide based on the MTT assay. Data obtained from this assay indicated that arsenic trioxide significantly reduced the viability of HL-60 cells, showing LD50 values of 14.26 + 0.5μg/mL, 12.54 + 0.3μg/mL, and 6.4 + 0.6μg/mL upon 6, 12, and 24 hours of exposure, respectively; indicating a dose- and time-dependent response relationship. Findings from the present study indicate that arsenic trioxide is highly cytotoxic to human leukemia (HL-60 cells, supporting its use as an effective therapeutic agent in the management of acute promyelocytic leukemia.

Effect of citrus flavonoids on HL-60 cell differentiation.

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Kawaii, S; Tomono, Y; Katase, E; Ogawa, K; Yano, M

1999-01-01

Twenty-seven Citrus flavonoids were examined for their activity of induction of terminal differentiation of human promyelocytic leukemia cells (HL-60) by nitro blue tetrazolium (NBT) reducing, nonspecific esterase, specific esterase, and phagocytic activities. 10 flavonoids were judged to be active (percentage of NBT reducing cells was more than 40% at a concentration of 40 microM), and the rank order of potency was natsudaidain, luteolin, tangeretin, quercetin, apigenin, 3, 3, '4, '5, 6, 7, 8-heptamethoxyflavone, nobiletin, acacetin, eriodictyol, and taxifolin. These flavonoids exerted their activity in a dose-dependent manner. HL-60 cells treated with these flavonoids differentiated into mature monocyte/macrophage. The structure-activity relationship established from comparison between flavones and flavanones revealed that ortho-catechol moiety in ring B and C2-C3 double bond had an important role for induction of differentiation of HL-60. In polymethoxylated flavones, hydroxyl group at C3 and methoxyl group at C8 enhanced the differentiation-inducing activity.

Antiproliferative activity of pristimerin isolated from Maytenus ilicifolia (Celastraceae) in human HL-60 cells.

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Costa, Patricia Marçal da; Ferreira, Paulo Michel Pinheiro; Bolzani, Vanderlan da Silva; Furlan, Maysa; de Freitas Formenton Macedo Dos Santos, Vânia Aparecida; Corsino, Joaquim; de Moraes, Manoel Odorico; Costa-Lotufo, Letícia Veras; Montenegro, Raquel Carvalho; Pessoa, Cláudia

2008-06-01

Pristimerin has been shown to be cytotoxic to several cancer cell lines. In the present work, the cytotoxicity of pristimerin was evaluated in human tumor cell lines and in human peripheral blood mononuclear cells (PBMC). This work also examined the effects of pristimerin (0.4; 0.8 and 1.7 microM) in HL-60 cells, after 6, 12 and 24h of exposure. Pristimerin reduced the number of viable cells and increased number of non-viable cells in a concentration-dependent manner by tripan blue test showing morphological changes consistent with apoptosis. Nevertheless, pristimerin was not selective to cancer cells, since it inhibited PBMC proliferation with an IC50 of 0.88 microM. DNA synthesis inhibition assessed by 5-bromo-2'-deoxyuridine (BrdU) incorporation in HL-60 cells was 70% and 83% for the concentrations of 0.4 and 0.8 microM, respectively. Pristimerin (10 and 20 microM) was not able to inhibit topoisomerase I. In AO/EB (acridine orange/ethidium bromide) staining, all tested concentrations reduced the number of HL-60 viable cells, with the occurrence of necrosis and apoptosis in a concentration-dependent manner, results in agreement with trypan blue exclusion findings. The analysis of membrane integrity and internucleosomal DNA fragmentation by flow cytometry in the presence of pristimerin indicated that treated cells underwent apoptosis. The present data point to the importance of pristimerin as representative of an emerging class of potential anticancer chemicals, exhibiting an antiproliferative effect by inhibiting DNA synthesis and triggering apoptosis.

Utilization of the human cell line HL-60 for chemiluminescence based detection of microorganisms and related substances

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Timm, Michael; Hansen, Erik W; Moesby, Lise

2006-01-01

species (ROS) when challenged with pyrogenic substances. In a luminol enhanced chemilumimetric assay the responsiveness of differentiated HL-60 cells is tested towards Salmonella typhimurium, Bacillus subtilis, Saccharomyces cerevisiae, Candida albicans, lipopolysaccharide (LPS) and lipoteichoic acid (LTA......). The results show a poor sensitivity to S. typhimurium but displays good sensitivity towards B. subtilis, LTA and LPS. Furthermore, the sensitivity towards the yeasts C. albicans and S. cerevisiae is considerably better than obtained in other in vitro cell systems. Overall these results indicate that the HL-60...... cell assay possibly could be evolved to a supplementary assay for the known pyrogenic detection assays. Furthermore, the utilization of the assay for pyrogenic examination of recombinant drugs derived from yeast expression systems would be relevant to examine....

Down-regulation of procaspase-8 expression by focal adhesion kinase protects HL-60 cells from TRAIL-induced apoptosis

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Tamagiku, Yuji; Sonoda, Yoshiko; Kunisawa, Mari; Ichikawa, Daiju; Murakami, Yayoi; Aizu-Yokota, Eriko; Kasahara, Tadashi

2004-01-01

We have demonstrated that focal adhesion kinase (FAK)-overexpressed (HL-60/FAK) cells have marked resistance against various apoptotic stimuli such as hydrogen peroxide, etoposide, and ionizing radiation compared with the vector-transfected (HL-60/Vect) cells. HL-60/FAK cells are highly resistant to TRAIL-induced apoptosis, while original HL-60 or HL-60/Vect cells were sensitive. TRAIL at 500 ng/ml induced significant DNA fragmentation, activation of caspase-8 and 3, the processing of a proapoptotic BID, and mitochondrial release of cytochrome c in HL-60/Vect cells, whereas no such events were observed in the HL-60/FAK cells. In particular, the expression of procaspase-8 gene and subsequent cleavage of caspase-8 were markedly reduced in HL-60/FAK cells, while expression of TRAIL-receptor 2 and 3, TRADD, and FADD was equivalent in both types of cells. In HL-60/FAK cells, the phosphoinositide 3 (PI3)-kinase/Akt survival pathway was constitutively activated, accompanied by significant induction of inhibitor-of-apoptosis proteins, XIAP, RIP, and Bcl-XL. The introduction of FAK siRNA in HL-60/FAK cells sensitized them against TRAIL-induced apoptosis, confirming that overexpressed FAK downregulates procaspase-8 expression, which subsequently inhibits downstream apoptosis pathway in the HL-60/FAK cells

The synthesis of phosphatidylalcohols in HL-60 cells

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Tettenborn, C.S.

1988-01-01

This study focuses upon the phenomenon that 12-O-tetradecanoyl-phorbol-13-acetate (TPA) stimulates a pathway for the synthesis of phosphatidylalcohols in HL-60 cells during the differentiation of these cells to macrophages. Using exogenous ethanol as a substrate for this pathway, the synthesis of phosphatidylethanol is induced by TPA well before full expression of differentiation. Experiments done in whole cell cultures demonstrated that this pathway could utilize a wide range of other alcohols as substrates, including glycerol, a potential endogenous substrate. Using radiolabeling and iodine-staining as criteria, this TPA-inducible pathway was shown to result in a dramatic alteration of phospholipid composition, depending on the availability of the alcohol headgroup. A cell-free system was developed to explore the enzymatic reactions involved in phosphatidylalcohol synthesis, a main goal of these studies. For this purpose, the lipid pools of HL-60 cells were prelabeled with [ 3 H]arachidonic acid in the absence of ethanol and total cell homogenates were prepared by sonication. The ability of the cell lysates to synthesize phosphatidylethanol was assayed after addition of ethanol to the system

FUSION OF SENDAI VIRUS WITH HUMAN HL-60 AND CEM CELLS - DIFFERENT KINETICS OF FUSION FOR 2 ISOLATES

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DELIMA, MCP; NIR, S; FLASHER, D; KLAPPE, K; HOEKSTRA, D; DUZGUNES, N

1991-01-01

The kinetics of fusion of Sendai virus (Z strain) with the human promyelocytic leukemia cell line HL-60, and the human T lymphocytic leukemia cell line CEM was investigated. Fusion was monitored by fluorescence dequenching of octadecylrhodamine (R-18) incorporated in the viral membrane. For one

Effects of highly ripened cheeses on HL-60 human leukemia cells: antiproliferative activity and induction of apoptotic DNA damage.

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Yasuda, S; Ohkura, N; Suzuki, K; Yamasaki, M; Nishiyama, K; Kobayashi, H; Hoshi, Y; Kadooka, Y; Igoshi, K

2010-04-01

To establish cheese as a dairy product with health benefits, we examined the multifunctional role of cheeses. In this report, we clarify whether different types of commercial cheeses may possess antiproliferative activity using HL-60 human promyelocytic leukemia cell lines as a cancer model. Among 12 cheese extracts tested, 6 (Montagnard, Pont-l'Eveque, Brie, Camembert, Danablue, and Blue) revealed strong growth inhibition activity and induction of DNA fragmentation in HL-60 cells. Based on the quantification of nitrogen contents in different cheese samples, a positive correlation between the ripeness of various cheeses and their antiproliferative activity tested in HL-60 cells was displayed. Four varieties of Blue cheese ripened for 0, 1, 2, or 3 mo demonstrated that the Blue cheese ripened for a long term was capable of causing the strong suppression of the cell growth and the induction of apoptotic DNA damage as well as nucleic morphological change in HL-60 cells. Collectively, these results obtained suggest a potential role of highly ripened cheeses in the prevention of leukemic cell proliferation. Copyright (c) 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Uric acid disrupts hypochlorous acid production and the bactericidal activity of HL-60 cells.

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Carvalho, Larissa A C; Lopes, João P P B; Kaihami, Gilberto H; Silva, Railmara P; Bruni-Cardoso, Alexandre; Baldini, Regina L; Meotti, Flavia C

2018-06-01

Uric acid is the end product of purine metabolism in humans and is an alternative physiological substrate for myeloperoxidase. Oxidation of uric acid by this enzyme generates uric acid free radical and urate hydroperoxide, a strong oxidant and potentially bactericide agent. In this study, we investigated whether the oxidation of uric acid and production of urate hydroperoxide would affect the killing activity of HL-60 cells differentiated into neutrophil-like cells (dHL-60) against a highly virulent strain (PA14) of the opportunistic pathogen Pseudomonas aeruginosa. While bacterial cell counts decrease due to dHL-60 killing, incubation with uric acid inhibits this activity, also decreasing the release of the inflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF- α). In a myeloperoxidase/Cl - /H 2 O 2 cell-free system, uric acid inhibited the production of HOCl and bacterial killing. Fluorescence microscopy showed that uric acid also decreased the levels of HOCl produced by dHL-60 cells, while significantly increased superoxide production. Uric acid did not alter the overall oxidative status of dHL-60 cells as measured by the ratio of reduced (GSH) and oxidized (GSSG) glutathione. Our data show that uric acid impairs the killing activity of dHL-60 cells likely by competing with chloride by myeloperoxidase catalysis, decreasing HOCl production. Despite diminishing HOCl, uric acid probably stimulates the formation of other oxidants, maintaining the overall oxidative status of the cells. Altogether, our results demonstrated that HOCl is, indeed, the main relevant oxidant against bacteria and deviation of myeloperoxidase activity to produce other oxidants hampers dHL-60 killing activity. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Cloning of a glutathione S-transferase decreasing during differentiation of HL60 cell line

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Kim, Jae Chul; Park, In Kyu; Lee, Kyu Bo; Sohn, Sang Kyun; Kim, Moo Kyu; Kim, Jung Chul [College of Medicine, Kyungpook National Univ., Taegu (Korea, Republic of)

1999-06-01

By sequencing the Expressed Sequence Tags of human dermal papilla cDNA library, we identified a clone named K872 of which the expression decreased during differentiation of HL60 cell line. K872 plasmid DNA was isolated according to QIA plasmid extraction kit (Qiagen GmbH, Germany). The nucleotide sequencing was performed by Sanger's method with K872 plasmid DNA. The most updated GenBank EMBL necleic acid banks were searched through the internet by using BLAST (Basic Local Alignment Search Tools) program. Northern bots were performed using RNA isolated from various human tissues and cancer cell lines. The gene expression of the fusion protein was achieved by His-Patch Thiofusion expression system and the protein product was identified on SDS-PAGE. K872 clone is 1006 nucleotides long, and has a coding region of 675 nucleotides and a 3' non-coding region of 280 nucleotides. The presumed open reading frame starting at the 5' terminus of K872 encodes 226 amino acids, including the initiation methionine residue. The amino acid sequence deduced from the open reading frame of K872 shares 70% identity with that of rat glutathione S-transferase kappa 1 (rGSTK1). The transcripts were expressed inh a variety of human tissues and cancer cells. The levels of transcript were relatively high in those tissues such as heart, skeletal muscle, and peripheral blood leukocyte. It is noteworthy that K872 was found to be abundantly expressed in colorectal cancer and melanoma cell lines. Homology search result suggests that K872 clone is the human homolog of the rGSTK1 which is known to be involved in the resistance of cytotoxic therapy. We propose that meticulous functional analysis should be followed to confirm that.

Cytotoxicity, Antiproliferative Effects, and Apoptosis Induction of Methanolic Extract of Cynometra cauliflora Linn. Whole Fruit on Human Promyelocytic Leukemia HL-60 Cells

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T-Johari S. A. Tajudin

2012-01-01

Full Text Available Methanolic extract of Cynometra cauliflora whole fruit was assayed for cytotoxicity against the human promyelocytic leukemia HL-60 and the normal mouse fibroblast NIH/3T3 cell lines by using the MTT assay. The CD50 of the extract for 72 hours was 0.9 μg/mL whereas the value for the cytotoxic drug vincristine was 0.2 μg/mL. The viability of the NIH/3T3 cells was at 80.0% when treated at 15.0 μg/mL. The extract inhibited HL-60 cell proliferation with dose dependence. AO/PI staining of HL-60 cells treated with the extract revealed that majority of cells were in the apoptotic cell death mode. Flow cytometry analysis of HL-60 cells treated at CD50 of the extract showed that the early apoptotic cells were 31.0, 26.3 and 19.9% at 24, 48, and 72 hours treatment, respectively. The percentage of late apoptotic cells was increased from 62.0 at 24 hours to 64.1 and 70.2 at 48 and 72 hours, respectively. Meanwhile, percent of necrotic cells were 4.9, 6.6, and 8.5 at 24, 48, and 72 hours, respectively. This study has shown that the methanolic extract of C. cauliflora whole fruit was cytotoxic towards HL-60 cells and induced the cells into apoptotic cell death mode, but less cytotoxic towards NIH/3T3 cells.

Rare Coumarins Induce Apoptosis, G1 Cell Block and Reduce RNA Content in HL60 Cells

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Widelski Jarosław

2017-02-01

Full Text Available The rare coumarins stenocarpin, stenocarpin isobutyrate, oficinalin, oficinalin isobutyrate, 8-methoxypeucedanin and the known xanthotoxin, isoimperatorin, bergapten, peucedanin and 8–methoxyisoimperatorin were isolated from Peucedanum luxurians Tamamsch. (Apiaceae and identified by means of spectral data (1D and 2D NMR. Their immunomodulating activity was evaluated by flow cytometry and their influence on HL60 cells as well as on PHA-stimulated PBLs was tested. All tested coumarins induce apoptosis (maximal in the 48 h culture and decrease cell proliferation in a time- and dose-dependent manner, especially in HL60 cells. They also induce partial G1 block, but only in HL60 cells (at 100 µM concentrations. Dose-dependent reduction of RNA content was also found in G1 cells treated by the coumarins. All of the tested coumarins also possessed immunomodulatory activities. Bergapten and xanthotoxin were found to be the best candidates for further evaluation as anti-cancer drugs.

The Critical Role of Redox Homeostasis in Shikonin-Induced HL-60 Cell Differentiation via Unique Modulation of the Nrf2/ARE Pathway

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Bo Zhang

2012-01-01

Full Text Available Among various cancer cell lines, the leukemia cell line HL-60 was most sensitive to Shikonin, with evidence showing both the prooxidative activities and proapoptotic effects of micromolar concentrations of Shikonin. However, the mechanism involved in the cytotoxicity of Shikonin in the submicromolar range has not been fully characterized. Using biochemical and free radical biological experiments in vitro, we identified the prodifferentiated profiles of Shikonin and evaluated the redox homeostasis during HL-60 differentiation. The data showed a strong dose-response relationship between Shikonin exposure and the characteristics of HL-60 differentiation in terms of morphology changes, nitroblue tetrazolium (NBT reductive activity, and the expression level of surface antigens CD11b/CD14. During drug exposure, intercellular redox homeostasis changes towards oxidation are necessary to support Shikonin-induced differentiation, which was proven by additional enzymatic and non-enzymatic redox modulators. A statistically significant and dose-dependent increase (P<0.05 was recorded with regard to the unique expression levels of the Nrf2/ARE downstream target genes in HL-60 cells undergoing late differentiation, which were restored with further antioxidants employed with the Shikonin treatment. Our research demonstrated that Shikonin is a differentiation-inducing agent, and its mechanisms involve the Nrf2/ARE pathway to modulate the intercellular redox homeostasis, thus facilitating differentiation.

Relationship between structure and antiproliferative activity of polymethoxyflavones towards HL60 cells.

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Kawaii, Satoru; Ikuina, Tomoyasu; Hikima, Takeshi; Tokiwano, Tetsuo; Yoshizawa, Yuko

2012-12-01

As part of our continuing investigation of polymethoxyflavone (PMF) derivatives as potential anticancer substances, a series of PMF derivatives was synthesized. The synthesized compounds were evaluated for cytotoxicity against the promyelocytic leukemic HL60 cell line, and structure-activity relationship correlations were investigated along with previously isolated PMFs from the peel of king orange (Citrus nobilis). 7,3'-Dimethoxyflavone demonstrated the most potent activity among the synthetic PMFs. Consideration of correlation between the methoxylation pattern and antiproliferative activity revealed the importance of the 3'-methoxyl group and the higher degree of methoxylation on the A-ring moiety of PMFs.

A study on apoptotic signaling pathway in HL-60 cells induced by radiation

International Nuclear Information System (INIS)

Kim, Hye Jung; Moon, Sung Keun; Lee, Jae Hoon; Moon, Sun Rock

2001-01-01

The mechanical insights of death at cancer cells by ionizing radiation are not yet clearly defined. Recent evidences have demonstrated that radiation therapy may induce cell death via activation of signaling pathway for apoptosis in target cells. This study is designed whether ionizing radiation may activate the signaling cascades of apoptosis including caspase family cysteine proteases, Bcl2/Bax, cytochrome c and Fas/Fas-L in target cells. HL-60 cells were irradiated in vitro with 6 MV X-ray at dose ranges from 2 Gy to 32 Gy. The cell viability was tested by MTT assay and the extent of apoptosis was determined using agarose gel electrophoresis. The activities of caspase proteases were measured by proteolytic cleavages of substrates. Western blot analysis was used to monitor PARP, caspase-3, Cytochrome-c, BcI-2, Bax, Fas and Fas-L. Ionizing radiation decreases the viability of HL -60 cells in a time and dose dependent manner. Ionizing radiation-induced death in HL- 60 cells is an apoptotic death which is revealed as characteristic ladder-pattern fragmentation at genomic DNA over 16 Gy at 4 hours. Ionizing radiation induces the activation of caspase-2, 3, 6, 8 and 9 of HL --60 cells in a time-dependent manner. The activation of caspase- 3 protease is also evidenced by the digestion of poly (ADP-ribose) polymerase and procaspase 3 with 16Gy ionizing irradiation. Anti-apoptotic Bcl2 expression is decreased but apoptotic Bax expression is increased with mitochondrial cytochrome c release in a time- dependent manner. In addition, expression of Fas and Fas-L is also increased in a time dependent manner. These data suggest that ionizing radiation-induced apoptosis is mediated by the activation of various signaling pathways including caspase family cysteine proteases, BcI 2 /Bax, Fas and Fas-L in a time and dose dependent manner

Induced apoptosis by mild hyperthermia occurs via telomerase inhibition on the three human myeloid leukemia cell lines: TF-1, K562, and HL-60.

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Deezagi, Abdolkhaleg; Manteghi, Sanaz; Khosravani, Pardis; Vaseli-Hagh, Neda; Soheili, Zahra-Soheila

2009-09-01

The purpose of this research was to understand the effect of hyperthermia on the telomerase activity in human leukemic cell lines (HL-60, K562, and TF-1). The cells were treated by hyperthermia at the range of 41-44 degrees C for 120 min and incubated for 96 h. Then telomerase activity, cell proliferation, and apoptosis were assessed. The results indicated that hyperthermia significantly induced apoptosis on the cells. The cells exhibited pre-apoptotic pattern at 41 and 42 degrees C at 60-120 min and apoptotic pattern at 43 and 44 degrees C over 30 min after hyperthermia. Telomerase activity (that was assayed immediately after hyperthermia) was stable at 41-42 degrees C for 60 min but decreased to 35-40% at 120 min. However, at severe hyperthermia (43-44 degrees C) telomerase activity was decreased in a time- and dose-dependent manner. Following hyperthermia (41-44 degrees C up to 120 min), the cells were incubated for 96 h. In these conditions, the telomerase activity was decreased by about 60-80% in comparison with that untreated control cells.

Discovery of novel inducers of cellular differentiation using HL-60 promyelocytic cells.

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Mata-Greenwood, E; Ito, A; Westenburg, H; Cui, B; Mehta, R G; Kinghorn, A D; Pezzuto, J M

2001-01-01

Non-physiological inducers of terminal differentiation have been used as novel therapies for the prevention and therapy of cancer. We have used cultured HL-60 promyelocytic cells to monitor differentiation, proliferation and cell death events as induced by a large set of extracts derived from plants. Screening of more than 1400 extracts led to the discovery of 34 with potent activity (ED50 Petiveria alliacea, and desmethylrocaglamide from Aglaia ponapensis. Zapotin demonstrated the most favorable biological profile in that induction of differentiation correlated with proliferation arrest, and a lack of cytotoxicity. We conclude that the HL-60 cell model is a useful system for the discovery of novel pharmacophores with potential to suppress the process of carcinogenesis, and that flavonoids may be especially useful in this capacity.

Differentiation-inducing effects of small fruit juices on HL-60 leukemic cells.

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Yoshizawa, Y; Kawaii, S; Urashima, M; Fukase, T; Sato, T; Murofushi, N; Nishimura, H

2000-08-01

Epidemiological studies indicate that high intakes of fruits and vegetables are associated with a reduced risk of cancer, and several plant-derived drugs have been developed in medical oncology. Since only a small part of the flora has been tested for any kind of bioactivity, we chose small fruits as sources of differentiation-inducing activity against HL-60 leukemic cells. We have prepared juices from various small fruits that grow mainly in the northern part of Japan. Screening of 43 samples indicated that juices of Actinidia polygama Maxim., Rosa rugosa Thunb., Vaccinium smallii A. Gray, and Sorbus sambucifolia Roem. strongly induced differentiation of HL-60 cells to monocyte/macrophage characteristics in a concentration-dependent manner as indicated by histochemical and biochemical examinations.

Berberine Induces Apoptotic Cell Death via Activation of Caspase-3 and -8 in HL-60 Human Leukemia Cells: Nuclear Localization and Structure-Activity Relationships.

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Okubo, Shinya; Uto, Takuhiro; Goto, Aya; Tanaka, Hiroyuki; Nishioku, Tsuyoshi; Yamada, Katsushi; Shoyama, Yukihiro

2017-01-01

Berberine (BBR), an isoquinoline alkaloid, is a well-known bioactive compound contained in medicinal plants used in traditional and folk medicines. In this study, we investigated the subcellular localization and the apoptotic mechanisms of BBR were elucidated. First, we confirmed the incorporation of BBR into the cell visually. BBR showed antiproliferative activity and promptly localized to the nucleus from 5[Formula: see text]min to 15[Formula: see text]min after BBR treatment in HL-60 human promyelocytic leukemia cells. Next, we examined the antiproliferative activity of BBR (1) and its biosynthetically related compounds (2-7) in HL-60 cells. BBR exerted strongest antiproliferative activity among 1-7 and the results of structures and activity relation suggested that a methylenedioxyl group in ring A, an [Formula: see text]-alkyl group at C-9 position, and the frame of isoquinoline may be necessary for antiproliferative activity. Moreover, BBR showed the most potent antiproliferative activity in HL-60 cells among human cancer and normal cell lines tested. Next, we examined the effect of BBR on molecular events known as apoptosis induction. In HL-60 cells, BBR induced chromatin condensation and DNA fragmentation, and triggered the activation of PARP, caspase-3 and caspase-8 without the activation of caspase-9. BBR-induced DNA fragmentation was abolished by pretreatment with inhibitors against caspase-3 and caspase-8, but not against caspase-9. ERK and p38 were promptly phosphorylated after 15 min of BBR treatment, and this was correlated with time of localization to the nucleus of BBR. These results demonstrated that BBR translocated into nucleus immediately after treatments and induced apoptotic cell death by activation of caspase-3 and caspase-8.

Involvement of the histamine H4 receptor in clozapine-induced hematopoietic toxicity: Vulnerability under granulocytic differentiation of HL-60 cells

International Nuclear Information System (INIS)

Goto, Aya; Mouri, Akihiro; Nagai, Tomoko; Yoshimi, Akira; Ukigai, Mako; Tsubai, Tomomi; Hida, Hirotake; Ozaki, Norio; Noda, Yukihiro

2016-01-01

Clozapine is an effective antipsychotic for treatment-resistant schizophrenia, but can cause fatal hematopoietic toxicity as agranulocytosis. To elucidate the mechanism of hematopoietic toxicity induced by clozapine, we developed an in vitro assay system using HL-60 cells, and investigated the effect on hematopoiesis. HL-60 cells were differentiated by all-trans retinoic acid (ATRA) into three states according to the following hematopoietic process: undifferentiated HL-60 cells, those undergoing granulocytic ATRA-differentiation, and ATRA-differentiated granulocytic cells. Hematopoietic toxicity was evaluated by analyzing cell survival, cell proliferation, granulocytic differentiation, apoptosis, and necrosis. In undifferentiated HL-60 cells and ATRA-differentiated granulocytic cells, both clozapine (50 and 100 μM) and doxorubicin (0.2 µM) decreased the cell survival rate, but olanzapine (1–100 µM) did not. Under granulocytic differentiation for 5 days, clozapine, even at a concentration of 25 μM, decreased survival without affecting granulocytic differentiation, increased caspase activity, and caused apoptosis rather than necrosis. Histamine H 4 receptor mRNA was expressed in HL-60 cells, whereas the expression decreased under granulocytic ATRA-differentiation little by little. Both thioperamide, a histamine H 4 receptor antagonist, and DEVD-FMK, a caspase-3 inhibitor, exerted protection against clozapine-induced survival rate reduction, but not of live cell counts. 4-Methylhistamine, a histamine H 4 receptor agonist, decreased the survival rate and live cell counts, as did clozapine. HL-60 cells under granulocytic differentiation are vulnerable under in vitro assay conditions to hematopoietic toxicity induced by clozapine. Histamine H 4 receptor is involved in the development of clozapine-induced hematopoietic toxicity through apoptosis, and may be a potential target for preventing its occurrence through granulocytic differentiation. - Highlights: • HL-60

Relationship Between Structure and Antiproliferative Activity of Novel 5-amino-4-cyanopyrazole-1-formaldehydehydrazono Derivatives on HL-60RG Human Leukemia Cells.

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Nagahara, Yukitoshi; Nagahara, Katsuhiko

2017-11-01

Pyrazole derivatives have been reported to have potent antimicrobial and anticancer activity. We recently synthesized and determined the effects of analogs, benzamidoxime derivatives, on mammalian cells and discovered that benzamidoximes had an antiproliferative effect. Here we synthesized and determined the anticancer effects of hydrazonopyrazole derivatives on a mammalian cancer cell line. We synthesized 12 hydrazonopyrazole derivatives with several constant alkyl chain length or branched chains at the side chain to investigate their anticancer cell activity, using the human myelogenous leukemia cell line HL-60RG. Among all hydrazonopyrazole derivatives we synthesized, the hydrazonopyrazole derivative with a branched chain at the side chain rather than a constant alkyl chain significantly inhibited cell viability. The strongest hydrazonopyrazole derivative, 5-amino-4-cyanopyrazole-1-formaldehydehydrazono-3'-pentanal, tended to damage cells dose-dependently. This cell growth attenuation was a result of apoptosis, activating caspase-3 and fragmented DNA. Hydrazonopyrazole derivatives induced apoptosis of HL-60RG leukemia cells. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

[Ca2+]i in exterior of cells effected on apoptosis of HL-60 cells induced by irradiation

International Nuclear Information System (INIS)

He Ziyi; Meng Qingyong

2005-01-01

Objective: To investigate of the different [Ca 2+ ]i in exterior of cells promotion function on apoptosis of HL-60 cells induced by irradiation. Methods: To put ration dose 32 P and different [Ca 2+ ]i into culture of HL-60 and measure the apoptosis rate with FCM after 24 and 48 hours. Result: Apoptosis rate increased with the increase of [Ca 2+ ]i which shows an obvious function to promote apoptosis, r 24 =0.9001 (P=0.0145); r48=0.9343 (P=0.0063). Conclusion: [Ca 2+ ]i in exterior of cells has a obvious function in promoteing apoptosis induced by irradiation. (authors)

Involvement of the histamine H{sub 4} receptor in clozapine-induced hematopoietic toxicity: Vulnerability under granulocytic differentiation of HL-60 cells

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Goto, Aya; Mouri, Akihiro; Nagai, Tomoko; Yoshimi, Akira; Ukigai, Mako; Tsubai, Tomomi; Hida, Hirotake [Division of Clinical Sciences and Neuropsychopharmacology, Faculty and Graduate School of Pharmacy, Meijo University, 150 Yagotoyama, Tempaku-ku, Nagoya 468-8503 (Japan); Ozaki, Norio [Department of Psychiatry, Graduate School of Medicine, Nagoya University, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Noda, Yukihiro, E-mail: ynoda@meijo-u.ac.jp [Division of Clinical Sciences and Neuropsychopharmacology, Faculty and Graduate School of Pharmacy, Meijo University, 150 Yagotoyama, Tempaku-ku, Nagoya 468-8503 (Japan)

2016-09-01

Clozapine is an effective antipsychotic for treatment-resistant schizophrenia, but can cause fatal hematopoietic toxicity as agranulocytosis. To elucidate the mechanism of hematopoietic toxicity induced by clozapine, we developed an in vitro assay system using HL-60 cells, and investigated the effect on hematopoiesis. HL-60 cells were differentiated by all-trans retinoic acid (ATRA) into three states according to the following hematopoietic process: undifferentiated HL-60 cells, those undergoing granulocytic ATRA-differentiation, and ATRA-differentiated granulocytic cells. Hematopoietic toxicity was evaluated by analyzing cell survival, cell proliferation, granulocytic differentiation, apoptosis, and necrosis. In undifferentiated HL-60 cells and ATRA-differentiated granulocytic cells, both clozapine (50 and 100 μM) and doxorubicin (0.2 µM) decreased the cell survival rate, but olanzapine (1–100 µM) did not. Under granulocytic differentiation for 5 days, clozapine, even at a concentration of 25 μM, decreased survival without affecting granulocytic differentiation, increased caspase activity, and caused apoptosis rather than necrosis. Histamine H{sub 4} receptor mRNA was expressed in HL-60 cells, whereas the expression decreased under granulocytic ATRA-differentiation little by little. Both thioperamide, a histamine H{sub 4} receptor antagonist, and DEVD-FMK, a caspase-3 inhibitor, exerted protection against clozapine-induced survival rate reduction, but not of live cell counts. 4-Methylhistamine, a histamine H{sub 4} receptor agonist, decreased the survival rate and live cell counts, as did clozapine. HL-60 cells under granulocytic differentiation are vulnerable under in vitro assay conditions to hematopoietic toxicity induced by clozapine. Histamine H{sub 4} receptor is involved in the development of clozapine-induced hematopoietic toxicity through apoptosis, and may be a potential target for preventing its occurrence through granulocytic differentiation

Inward rectifier potassium channels in the HL-1 cardiomyocyte-derived cell line.

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Goldoni, Dana; Zhao, YouYou; Green, Brian D; McDermott, Barbara J; Collins, Anthony

2010-11-01

HL-1 is a line of immortalized cells of cardiomyocyte origin that are a useful complement to native cardiomyocytes in studies of cardiac gene regulation. Several types of ion channel have been identified in these cells, but not the physiologically important inward rectifier K(+) channels. Our aim was to identify and characterize inward rectifier K(+) channels in HL-1 cells. External Ba(2+) (100 µM) inhibited 44 ± 0.05% (mean ± s.e.m., n = 11) of inward current in whole-cell patch-clamp recordings. The reversal potential of the Ba(2+)-sensitive current shifted with external [K(+)] as expected for K(+)-selective channels. The slope conductance of the inward Ba(2+)-sensitive current increased with external [K(+)]. The apparent Kd for Ba(2+) was voltage dependent, ranging from 15 µM at -150  mV to 148 µM at -75  mV in 120  mM external K(+). This current was insensitive to 10 µM glybenclamide. A component of whole-cell current was sensitive to 150 µM 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), although it did not correspond to the Ba(2+)-sensitive component. The effect of external 1 mM Cs(+) was similar to that of Ba(2+). Polymerase chain reaction using HL-1 cDNA as template and primers specific for the cardiac inward rectifier K(ir)2.1 produced a fragment of the expected size that was confirmed to be K(ir)2.1 by DNA sequencing. In conclusion, HL-1 cells express a current that is characteristic of cardiac inward rectifier K(+) channels, and express K(ir)2.1 mRNA. This cell line may have use as a system for studying inward rectifier gene regulation in a cardiomyocyte phenotype. © 2010 Wiley-Liss, Inc.

Gene expression profiling for nitric oxide prodrug JS-K to kill HL-60 myeloid leukemia cells.

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Liu, Jie; Malavya, Swati; Wang, Xueqian; Saavedra, Joseph E; Keefer, Larry K; Tokar, Erik; Qu, Wei; Waalkes, Michael P; Shami, Paul J

2009-07-01

The nitric oxide (NO) prodrug JS-K is shown to have anticancer activity. To profile the molecular events associated with the anticancer effects of JS-K, HL-60 leukemia cells were treated with JS-K and subjected to microarray and real-time RT-PCR analysis. JS-K induced concentration- and time-dependent gene expression changes in HL-60 cells corresponding to the cytolethality effects. The apoptotic genes (caspases, Bax, and TNF-alpha) were induced, and differentiation-related genes (CD14, ITGAM, and VIM) were increased. For acute phase protein genes, some were increased (TP53, JUN) while others were suppressed (c-myc, cyclin E). The expression of anti-angiogenesis genes THBS1 and CD36 and genes involved in tumor cell migration such as tissue inhibitors of metalloproteinases, were also increased by JS-K. Confocal analysis confirmed key gene changes at the protein levels. Thus, multiple molecular events are associated with JS-K effects in killing HL-60, which could be molecular targets for this novel anticancer NO prodrug.

Effects of nicotine on cellular proliferation, cell cycle phase distribution, and macromolecular synthesis in human promyelocytic HL-60 leukaemia cells

International Nuclear Information System (INIS)

Konno, S.; Wu, J.M.; Chiao, J.W.

1986-01-01

Addition of nicotine causes a dose- and time-dependent inhibition of cell growth in the human promyelocytic HL-60 leukemia cells, with 4 mM nicotine resulting in a 50% inhibition of cellular proliferation after 48-50h. Accompanying the anticellular effect of nicotine is a significant change in the cell cycle distribution of HL-60 cells. For example, treatment with 4 mM nicotine for 20h causes an increase in the proportion of G1-phase cells (from 49% to 57%) and a significant decrease in the proportion of S-phase cells (from 41% to 32%). These results suggest that nicotine causes partial cell arrest in the G-1 phase which may in part account for its effects on cell growth. To determine whether nicotine changes the cellular uptake/transport to macromolecular precursors, HL-60 cells were treated with 216 mM nicotine for 30h, at the end of which time cells were labelled with ( 3 H)thymidine, ( 3 H)uridine, ( 14 C)lysine and( 35 S)methionine, the trichloroacetic acid soluble and insoluble radioactivities from each of the labelling conditions were determined. These studies show that nicotine mainly affects the ''de novo synthesis'' of proteins. (author)

Cdc6 is a rate-limiting factor for proliferative capacity during HL60 cell differentiation

International Nuclear Information System (INIS)

Barkley, Laura R.; Hong, Hye Kyung; Kingsbury, Sarah R.; James, Michelle; Stoeber, Kai; Williams, Gareth H.

2007-01-01

The DNA replication (or origin) licensing pathway represents a critical step in cell proliferation control downstream of growth signalling pathways. Repression of origin licensing through down-regulation of the MCM licensing factors (Mcm2-7) is emerging as a ubiquitous route for lowering proliferative capacity as metazoan cells exit the cell division cycle into quiescent, terminally differentiated and senescent 'out-of-cycle' states. Using the HL60 monocyte/macrophage differentiation model system and a cell-free DNA replication assay, we have undertaken direct biochemical investigations of the coupling of origin licensing to the differentiation process. Our data show that down-regulation of the MCM loading factor Cdc6 acts as a molecular switch that triggers loss of proliferative capacity during early engagement of the somatic differentiation programme. Consequently, addition of recombinant Cdc6 protein to in vitro replication reactions restores DNA replication competence in nuclei prepared from differentiating cells. Differentiating HL60 cells over-expressing either wild-type Cdc6 or a CDK phosphorylation-resistant Cdc6 mutant protein (Cdc6A4) exhibit an extended period of cell proliferation compared to mock-infected cells. Notably, differentiating HL60 cells over-expressing the Cdc6A4 mutant fail to down-regulate Cdc6 protein levels, suggesting that CDK phosphorylation of Cdc6 is linked to its down-regulation during differentiation and the concomitant decrease in cell proliferation. In this experimental model, Cdc6 therefore plays a key role in the sequential molecular events leading to repression of origin licensing and loss of proliferative capacity during execution of the differentiation programme

Differentiation-promoting activity of pomegranate (Punica granatum) fruit extracts in HL-60 human promyelocytic leukemia cells.

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Kawaii, Satoru; Lansky, Ephraim P

2004-01-01

Differentiation refers to the ability of cancer cells to revert to their normal counterparts, and its induction represents an important noncytotoxic therapy for leukemia, and also breast, prostate, and other solid malignancies. Flavonoids are a group of differentiation-inducing chemicals with a potentially lower toxicology profile than retinoids. Flavonoid-rich polyphenol fractions from the pomegranate (Punica granatum) fruit exert anti-proliferative, anti-invasive, anti-eicosanoid, and pro-apoptotic actions in breast and prostate cancer cells and anti-angiogenic activities in vitro and in vivo. Here we tested flavonoid-rich fractions from fresh (J) and fermented (W) pomegranate juice and from an aqueous extraction of pomegranate pericarps (P) as potential differentiation-promoting agents of human HL-60 promyelocytic leukemia cells. Four assays were used to assess differentiation: nitro blue tetrazolium reducing activity, nonspecific esterase activity, specific esterase activity, and phagocytic activity. In addition, the effect of these extracts on HL-60 proliferation was evaluated. Extracts W and P were strong promoters of differentiation in all settings, with extract J showing only a relatively mild differentiation-promoting effect. The extracts had proportional inhibitory effects on HL-60 cell proliferation. The results highlight an important, previously unknown, mechanism of the cancer preventive and suppressive potential of pomegranate fermented juice and pericarp extracts.

Induction of apoptosis by Citrus paradisi essential oil in human leukemic (HL-60) cells.

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Hata, Tomona; Sakaguchi, Ikuyo; Mori, Masahiro; Ikeda, Norikazu; Kato, Yoshiko; Minamino, Miki; Watabe, Kazuhito

2003-01-01

Limonene is a primary component of citrus essential oils (EOs) and has been reported to induce apoptosis on tumor cells. Little is known about induction of apoptosis by citrus EOs. In this study, we examined induction of apoptosis by Citrus aurantium var. dulcis (sweet orange) EO, Citrus paradisi (grapefruit) EO and Citrus limon (lemon) EO. These EOs induced apoptosis in HL-60 cells and the apoptosis activities were related to the limonene content of the EOs. Moreover, sweet orange EO and grapefruit EO may contain components besides limonene that have apoptotic activity. To identify the components with apoptotic activity, grapefruit EO was fractionated using silica gel columns, and the components were analyzed by GC-MS. The n-hexane fraction contained limonene, and the dichloromethane fraction (DF) contained aldehyde compounds and nootkatone. Decanal, octanal and citral in the DF showed strong apoptotic activity, suggesting that the aldehyde compounds induced apoptosis strongly in HL-60 cells.

Genes encoded within 8q24 on the amplicon of a large extrachromosomal element are selectively repressed during the terminal differentiation of HL-60 cells.

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Hirano, Tetsuo; Ike, Fumio; Murata, Takehide; Obata, Yuichi; Utiyama, Hiroyasu; Yokoyama, Kazunari K

2008-04-02

Human acute myeloblastic leukemia HL-60 cells become resistant to differentiation during long-term cultivation. After 150 passages, double minute chromosomes (dmins) found in early-passaged cells are replaced by large extrachromosomal elements (LEEs). In a DNA library derived from a purified fraction of LEEs, 12.6% (23/183) of clones were assigned to 8q24 and 9.2% (17/183) were assigned to 14q11 in the human genome. Fluorescence in situ hybridization (FISH) revealed a small aberrant chromosome, which had not been found in early-passaged cells, in addition to the purified LEEs. We determined that each LEE consisted of six discontinuous segments in a region that extended for 4.4Mb over the 8q24 locus. Five genes, namely, Myc (a proto-oncogene), NSMCE2 (for a SUMO ligase), CCDC26 (for a retinoic acid-dependent modulator of myeloid differentiation), TRIB1 (for a regulator of MAPK kinase) and LOC389637 (for a protein of unknown function), were encoded by the amplicon. Breaks in the chromosomal DNA within the amplicon were found in the NSMCE2 and CCDC26 genes. The discontinuous structure of the amplicon unit of the LEEs was identical with that of dmins in HL-60 early-passaged cells. The difference between them seemed, predominantly, to be the number (10-15 copies per LEE versus 2 or 3 copies per dmin) of constituent units. Expression of the Myc, NSMCE2, CCDC26 and LOC389637 and TRIB1 genes was constitutive in all lines of HL-60 cells and that of the first four genes was repressed during the terminal differentiation of early-passaged HL-60 cells. We also detected abnormal transcripts of CCDC26. Our results suggest that these genes were selected during the development of amplicons. They might be amplified and, sometimes, truncated to contribute to the maintenance of HL-60 cells in an undifferentiated state.

In Vitro Antioxidant and Antiproliferative Activities of Novel Orange Peel Extract and It's Fractions on Leukemia HL-60 Cells.

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Diab, Kawthar A E; Shafik, Reham Ezzat; Yasuda, Shin

2015-01-01

In the present work, novel orange peel was extracted with 100%EtOH (ethanol) and fractionated into four fractions namely F1, F2, F3, F4 which were eluted from paper chromatographs using 100%EtOH, 80%EtOH, 50%EtOH and pure water respectively. The crude extract and its four fractions were evaluated for their total polyphenol content (TPC), total flavonoid content (TFC) and radical scavenging activity using DPPH (1,1-diphenyl-2-picrylhydrazyl) assay. Their cytotoxic activity using WST assay and DNA damage by agarose gel electrophoresis were also evaluated in a human leukemia HL-60 cell line. The findings revealed that F4 had the highest TPC followed by crude extract, F2, F3 and F1. However, the crude extract had the highest TFC followed by F4, F3, F2, and F1. Depending on the values of EC50 and trolox equivalent antioxidant capacity, F4 possessed the strongest antioxidant activity while F1 and F2 displayed weak antioxidant activity. Further, incubation HL-60 cells with extract/fractions for 24h caused an inhibition of cell viability in a concentration- dependent manner. F3 and F4 exhibited a high antiproliferative activity with a narrow range of IC50 values (45.9 - 48.9 μg/ml). Crude extract exhibited the weakest antiproliferative activity with an IC50 value of 314.89 μg/ml. Analysis of DNA fragmentation displayed DNA degradation in the form of a smear-type pattern upon agarose gel after incubation of HL-60 cells with F3 and F4 for 6 h. Overall, F3 and F4 appear to be good sources of phytochemicals with antioxidant and potential anticancer activities.

Beta-mangostin from Cratoxylum arborescens activates the intrinsic apoptosis pathway through reactive oxygen species with downregulation of the HSP70 gene in the HL60 cells associated with a G0/G1 cell-cycle arrest.

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Omer, Fatima Abdelmutaal Ahmed; Hashim, Najihah Binti Mohd; Ibrahim, Mohamed Yousif; Dehghan, Firouzeh; Yahayu, Maizatulakmal; Karimian, Hamed; Salim, Landa Zeenelabdin Ali; Mohan, Syam

2017-11-01

Xanthones are phytochemical compounds found in a number of fruits and vegetables. Characteristically, they are noted to be made of diverse properties based on their biological, biochemical, and pharmacological actions. Accordingly, the apoptosis mechanisms induced by beta-mangostin, a xanthone compound isolated from Cratoxylum arborescens in the human promyelocytic leukemia cell line (HL60) in vitro, were examined in this study. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was done to estimate the cytotoxicity effect of β-mangostin on the HL60 cell line. Acridine orange/propidium iodide and Hoechst 33342 dyes and Annexin V tests were conducted to detect the apoptosis features. Caspase-3 and caspase-9 activities; reactive oxygen species; real-time polymerase chain reaction for Bcl-2, Bax, caspase-3, and caspase-9 Hsp70 genes; and western blot for p53, cytochrome c, and pro- and cleavage-caspase-3 and caspase-9 were assessed to examine the apoptosis mechanism. Cell-cycle analysis conducted revealed that β-mangostin inhibited the growth of HL60 at 58 µM in 24 h. The administration of β-mangostin with HL60 caused cell morphological changes related to apoptosis which increased the number of early and late apoptotic cells. The β-mangostin-catalyzed apoptosis action through caspase-3, caspase-7, and caspase-9 activation overproduced reactive oxygen species which downregulated the expression of antiapoptotic genes Bcl-2 and HSP70. Conversely, the expression of the apoptotic genes Bax, caspase-3, and caspase-9 were upregulated. Meanwhile, at the protein level, β-mangostin activated the formation of cleaved caspase-3 and caspase-9 and also upregulated the p53. β-mangostin arrested the cell cycle at the G 0 /G 1 phase. Overall, the results for β-mangostin showed an antiproliferative effect in HL60 via stopping the cell cycle at the G 0 /G 1 phase and prompted the intrinsic apoptosis pathway.

[Expression of ICAT and Wnt signaling-related proteins in the monocytic differentiation of HL-60 cells induced by a new steroidal drug NSC67657].

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Wang, J S; Wang, W J; Wang, T; Zhang, Y

2016-04-01

To investigate the expression of mRNA and proteins of β-catenin, TCF-4 (ICAT) and Wnt signaling pathway-related genes in the monocytic differentiation of acute myeloid leukemia HL-60 cells induced by a new steroidal drug NSC67657. Wright's staining and α-NBE staining were used to observe the differentiation of HL-60 cells after 5 days of 10 μmol/L NSC67657 treatment. Flow cytometry (FCM) was used to detect the differentiation and cell cycles. The expressions of mRNA and proteins of ICAT and Wnt signaling pathway-related factors, including β-catenin, TCF-4, c-myc, cyclin D1 and TCF-1 before and after differentiation, were detected by RT-PCR and Western blot. Morphological observation showed that NSC67657 induced monocytic differentiation of HL-60 cells. At 5 days after 10 μmol/L NSC67657 treatment, the number of CD14(+) HL-60 cells was (94.37±2.84)%, significantly higher than the (1.31±0.09)% in control group (Pcells were of (18.76±0.98)%, significantly lower than that of (34.38±2.61) % in the control group (Pprotein, and down-regulated the expression of β-catenin mRNA and protin (Pprotein and nuclear protein in the HL-60 cells (P>0.05 for all). The target genes of Wnt signaling pathway, including c-myc, cyclinD1 and TCF-1 mRNA and proteins in the HL-60 cells were significantly down-regulated after NSC67657 treatment (Pcells, and down-regulates the expression of β-catenin and target genes of Wnt signaling pathway. These results indicate that Wnt signaling pathway may be directly or indirectly involved in the monocytic differentiation process of HL-60 cells.

SKP2 siRNA inhibits the degradation of P27kip1 and down-regulates the expression of MRP in HL-60/A cells.

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Xiao, Jie; Yin, Songmei; Li, Yiqing; Xie, Shuangfeng; Nie, Danian; Ma, Liping; Wang, Xiuju; Wu, Yudan; Feng, Jianhong

2009-08-01

S-phase kinase-associated protein 2 (SKP2) gene is a tumor suppressor gene, and is involved in the ubiquitin-mediated degradation of P27kip1. SKP2 and P27kip1 affect the proceeding and prognosis of leukemia through regulating the proliferation, apoptosis and differentiation of leukemia cells. In this study, we explored the mechanism of reversing of HL-60/A drug resistance through SKP2 down-regulation. HL-60/A cells were nucleofected by Amaxa Nucleofector System with SKP2 siRNA. The gene and protein expression levels of Skp2, P27kip1, and multi-drug resistance associated protein (MRP) were determined by reverse transcription-polymerase chain reaction and western blot analysis, respectively. The cell cycle was analyzed by flow cytometry. The 50% inhibitory concentration value was calculated using cytotoxic analysis according to the death rate of these two kinds of cells under different concentrations of chemotherapeutics to compare the sensitivity of the cells. HL-60/A cells showed multi-drug resistance phenotype characteristic by cross-resistance to adriamycin, daunorubicin, and arabinosylcytosine, due to the expression of MRP. We found that the expression of SKP2 was higher in HL-60/A cells than in HL-60 cells, but the expression of P27kip1 was lower. The expression of SKP2 in HL-60/A cells nucleofected by SKP2 siRNA was down-regulated whereas the protein level of P27kip1 was up-regulated. Compared with the MRP expression level in the control group (nucleofected by control siRNA), the mRNA and protein expression levels of MRP in HL-60/A cells nucleofected by SKP2 siRNA were lower, and the latter cells were more sensitive to adriamycin, daunorubicin, and arabinosylcytosine. Down-regulating the SKP2 expression and arresting cells in the G0/G1 phase improve drug sensitivity of leukemia cells with down-regulated MRP expression.

Antileukemic Potential of Momordica charantia Seed Extracts on Human Myeloid Leukemic HL60 Cells

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Ramani Soundararajan

2012-01-01

Full Text Available Momordica charantia (bitter gourd has been used in the traditional system of medicine for the treatment of various diseases. Anticancer activity of M. charantia extracts has been demonstrated by numerous in vitro and in vivo studies. In the present study, we investigated the differentiation inducing potential of fractionated M. charantia seed extracts in human myeloid HL60 cells. We found that the HL60 cells treated with the fractionated seed extracts differentiated into granulocytic lineage as characterized by NBT staining, CD11b expression, and specific esterase activity. The differentiation inducing principle was found to be heat-stable, and organic in nature. The differentiation was accompanied by a downregulation of c-myc transcript, indicating the involvement of c-myc pathway, at least in part, in differentiation. Taken together these results indicate that fractionated extracts of M. charantia seeds possess differentiation inducing activity and therefore can be evaluated for their potential use in differentiation therapy for leukemia in combination with other inducers of differentiation.

Antileukemic Potential of Momordica charantia Seed Extracts on Human Myeloid Leukemic HL60 Cells

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Soundararajan, Ramani; Prabha, Punit; Rai, Umesh; Dixit, Aparna

2012-01-01

Momordica charantia (bitter gourd) has been used in the traditional system of medicine for the treatment of various diseases. Anticancer activity of M. charantia extracts has been demonstrated by numerous in vitro and in vivo studies. In the present study, we investigated the differentiation inducing potential of fractionated M. charantia seed extracts in human myeloid HL60 cells. We found that the HL60 cells treated with the fractionated seed extracts differentiated into granulocytic lineage as characterized by NBT staining, CD11b expression, and specific esterase activity. The differentiation inducing principle was found to be heat-stable, and organic in nature. The differentiation was accompanied by a downregulation of c-myc transcript, indicating the involvement of c-myc pathway, at least in part, in differentiation. Taken together these results indicate that fractionated extracts of M. charantia seeds possess differentiation inducing activity and therefore can be evaluated for their potential use in differentiation therapy for leukemia in combination with other inducers of differentiation. PMID:22654956

HA117 endows HL60 cells with a stem-like signature by inhibiting the degradation of DNMT1 via its ability to down-regulate expression of the GGL domain of RGS6.

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Shuangshuang Li

Full Text Available All-trans retinoic acid (ATRA induces complete remission in almost all patients with acute promyelocytic leukemia (APL via its ability to induce the in vivo differentiation of APL blasts. However, prolonged ATRA treatment can result in drug resistance. In previous studies, we generated a multi-drug-resistant HL60/ATRA cell line and found it to contain a new drug resistance-related gene segment, HA117. In this study, we demonstrate that ATRA induces multi-drug-resistant subpopulations of HL60 cells with a putative stem-like signature by up-regulating the expression of the new gene segment HA117. Western blot analysis and quantitative real-time PCR demonstrated that HA117 causes alternative splicing of regulator of G-protein signaling 6 (RGS6 and down-regulation of the expression of the GGL domain of RGS6, which plays an important role in DNA methyltransferase 1 (DNMT1 degradation. Moreover, DNMT1 expression was increased in multi-drug resistance HL60/ATRA cells. Knockdown of HA117 restored expression of the GGL domain and blocked DNMT1 expression. Moreover, resistant cells displayed a putative stem-like signature with increased expression of cancer steam cell markers CD133 and CD123. The stem cell marker, Nanog, was significantly up-regulated. In conclusion, our study shows that HA117 potentially promotes the stem-like signature of the HL60/ATRA cell line by inhibiting by the ubiquitination and degradation of DNMT1 and by down-regulating the expression of the GGL domain of RGS6. These results throw light on the cellular events associated with the ATRA-induced multi-drug resistance phenotype in acute leukemia.

Immunomodulatory effects of testosterone evaluated in all-trans retinoic acid differentiated HL-60 cells, granulocytes, and monocytes

DEFF Research Database (Denmark)

Boje, Alex; Moesby, Lise; Timm, Michael

2012-01-01

The sex hormones are known to affect innate immunity in humans. In this study we evaluated the immunomodulatory effects of testosterone in a model system comprising of all-trans retinoic acid differentiated HL-60 cells, and confirmed the results in human granulocytes and monocytes. Results showed...... that testosterone at pharmacological doses reduced the production of interleukin-8 and reactive oxygen species from differentiated HL-60 cells in a concentration dependent manner without affecting phagocytosis. The cells were stimulated with zymosan, lipopolysaccharide, or Bacillus subtilis. At the highest...... concentration of testosterone (120 µM), interleukin-8 secretion was reduced 42-80%, and production of reactive oxygen species was reduced 32-46%. Flutamide, an antagonist of the classical intracellular androgen receptor, was unable to antagonize the immunosuppressive effect of testosterone. We further...

1,25 dihydroxyvitamin D3 (1,25) regulation of c-myc mRNA in HL-60 leukemia cells

International Nuclear Information System (INIS)

Simpson, R.U.; Bresnick, E.H.; Begley, D.A.

1986-01-01

Recently, 1,25 was shown to induce differentiation and decrease c-myc levels in HL-60 cells. The authors have confirmed these observations by RNA dot blot analysis. Cells treated with 50 nM 1,25 for 4, 24 and 48 hr showed c-myc mRNA levels of 26, 17 and 15% of control respectively. β-Actin mRNA levels were not altered. To ascertain whether 1, 25 regulated c-myc transcriptionally, an HL-60 nuclear RNA runoff assay was developed. Assay of total nuclei transcriptional activity revealed that 50% of RNA elongation was α-amanitin (0.8 μg/ml) sensitive and was linear with nuclei concentration (0.1-1 x 10 7 nuclei). 1,25 (50 nM) treated (45-96 hr) cells had decreased (approx.40%) total transcription rate relative to control. Decreased total RNA synthesis occurred concomitant with NBT reducing activity. 32 P-RNA runoff transcripts from HL-60 nuclei were hybridized to excess (5 μg DNA was excess) Pst I linearized c-myc and β-actin clones (in pBR322) immobilized on nitrocellulose filter. 32 P-RNA input from 2 x 10 6 to 2 x 10 7 cpm yielded linear hybridization signal. Analysis of blot dot intensity revealed no difference in transcription of c-myc in nuclei from 1,25 dosed or control cells. (myc/actin ratios: 1,25 (50 nM, 72 hr) =1.1 +/- 0.3 and control (72 hr) = 1.0, N=3 or 2 or 3 dots ea). These preliminary data suggest 1,25 does not affect c-myc transcription in HL-60 nuclei and may regulate c-myc mRNA post-transcriptionally

Structure-activity relationship studies of 5,7-dihydroxyflavones as naturally occurring inhibitors of cell proliferation in human leukemia HL-60 cells.

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Ninomiya, Masayuki; Nishida, Kyohei; Tanaka, Kaori; Watanabe, Kunitomo; Koketsu, Mamoru

2013-07-01

Flavonoids are widely occurring polyphenols that are found in plants. The aim of this study was to investigate the structure-activity relationships of 5,7-dihydroxyflavones, with a focus on the effect of B ring structure substitution on the antiproliferative effects of the compounds in human leukemia HL-60 cells. We prepared a series of 5,7-dihydroxyflavones and evaluated their ability to inhibit the proliferation of HL-60 cells by using the MTT assay. The apoptosis- and cell differentiation-inducing ability of the most potent flavones were investigated using staining and morphological analyses. This study explored the antileukemic and chemopreventive potency of 5,7-dihydroxyflavones, particularly diosmetin and chrysoeriol, which have both hydroxy and methoxy groups on the B ring.

Biosynthesis of platelet activating factor (PAF) via alternate pathways: subcellular distribution of products in HL-60 cells

International Nuclear Information System (INIS)

Record, M.; Snyder, F.

1986-01-01

Final steps in the biosynthesis of PAF can be catalyzed by two different routes: CDP-choline:1-alkyl-2-acetyl-Gro cholinephosphotransferase [dithiothrietol (DTT)-insensitive] or acetyl-CoA:1-alkyl-2-lyso-GroPCho acetyltransferase. The authors have investigated the conversion of tritium-labeled 1-alkyl-2-acetyl-Gro and 1-alkyl-2-lyso-GroPCho (lyso-PAF) to PAF and other lipid products in HL-60 cells and in subcellular organelles isolated by centrifugation in a Percoll gradient. When cells are incubated with the labeled precursors (2 μM) the total amount of labeled PAF and 1-alkyl-2-acyl-GroPCho formed was similar from both precursors (60 pmol from 1-alkyl-2-acetyl-Gro and 50 pmol from lyso-PAF). However, PAF formed from 1-alkyl-2-acetyl-Gro represented 70% of the total products, whereas with lyso-PAF the major labeled product was 1-alkyl-2-acyl-GroPCho. Formation of PAF from 1-[ 3 H]alkyl-2-acetyl-Gro was linear to at least 30 min at 20 0 C. After a 15-min incubation of this neutral lipid with HL-60 cells, the labeled PAF produced was located exclusively in the plasma membrane fraction as opposed to the label in the 1-alkyl-2-acyl-GroPCho, which was found only in the endoplasmic reticulum; none of the labeled PAF product was released to the media. The authors results suggest PAF might be synthesized by the DTT-insensitive cholinephosphotransferase at the site of the plasma membrane in HL-60 cells

The content of DNA and RNA in microparticles released by Jurkat and HL-60 cells undergoing in vitro apoptosis

International Nuclear Information System (INIS)

Reich, Charles F.; Pisetsky, David S.

2009-01-01

Microparticles are small membrane-bound vesicles that are released from apoptotic cells during blebbing. These particles contain DNA and RNA and display important functional activities, including immune system activation. Furthermore, nucleic acids inside the particle can be analyzed as biomarkers in a variety of disease states. To elucidate the nature of microparticle nucleic acids, DNA and RNA released in microparticles from the Jurkat T and HL-60 promyelocytic cell lines undergoing apoptosis in vitro were studied. Microparticles were isolated from culture media by differential centrifugation and characterized by flow cytometry and molecular approaches. In these particles, DNA showed laddering by gel electrophoresis and was present in a form that allowed direct binding by a monoclonal anti-DNA antibody, suggesting antigen accessibility even without fixation. Analysis of RNA by gel electrophoresis showed intact 18s and 28s ribosomal RNA bands, although lower molecular bands consistent with 28s ribosomal RNA degradation products were also present. Particles also contained messenger RNA as shown by RT-PCR amplification of sequences for β-actin and GAPDH. In addition, gel electrophoresis showed the presence of low molecular weight RNA in the size range of microRNA. Together, these results indicate that microparticles from apoptotic Jurkat and HL-60 cells contain diverse nucleic acid species, indicating translocation of both nuclear and cytoplasmic DNA and RNA as particle release occurs during death

Compound K, a metabolite of ginseng saponin, induces apoptosis via caspase-8-dependent pathway in HL-60 human leukemia cells

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Cho, Sung-Hee; Chung, Kyung-Sook; Choi, Jung-Hye; Kim, Dong-Hyun; Lee, Kyung-Tae

2009-01-01

Compound K [20-O-β-(D-glucopyranosyl)-20(S)-protopanaxadiol], a metabolite of the protopanaxadiol-type saponins of Panax ginseng C.A. Meyer, has been reported to possess anti-tumor properties to inhibit angiogenesis and to induce tumor apoptosis. In the present study, we investigated the effect of Compound K on apoptosis and explored the underlying mechanisms involved in HL-60 human leukemia cells. We examined the effect of Compound K on the viabilities of various cancer cell lines using MTT assays. DAPI assay, Annexin V and PI double staining, Western blot assay and immunoprecipitation were used to determine the effect of Compound K on the induction of apoptosis. Compound K was found to inhibit the viability of HL-60 cells in a dose- and time-dependent manner with an IC 50 of 14 μM. Moreover, this cell death had typical features of apoptosis, that is, DNA fragmentation, DNA ladder formation, and the externalization of Annexin V targeted phosphatidylserine residues in HL-60 cells. In addition, compound-K induced a series of intracellular events associated with both the mitochondrial- and death receptor-dependent apoptotic pathways, namely, (1) the activation of caspases-3, -8, and -9; (2) the loss of mitochondrial membrane potential; (3) the release of cytochrome c and Smac/DIABLO to the cytosol; (4) the translocation of Bid and Bax to mitochondria; and (5) the downregulations of Bcl-2 and Bcl-xL. Furthermore, a caspase-8 inhibitor completely abolished caspase-3 activation, Bid cleavage, and subsequent DNA fragmentation by Compound K. Interestingly, the activation of caspase-3 and -8 and DNA fragmentation were significantly prevented in the presence of cycloheximide, suggesting that Compound K-induced apoptosis is dependent on de novo protein synthesis. The results indicate that caspase-8 plays a key role in Compound K-stimulated apoptosis via the activation of caspase-3 directly or indirectly through Bid cleavage, cytochrome c release, and caspase-9 activation

Compound K, a metabolite of ginseng saponin, induces apoptosis via caspase-8-dependent pathway in HL-60 human leukemia cells

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Choi Jung-Hye

2009-12-01

Full Text Available Abstract Background Compound K [20-O-β-(D-glucopyranosyl-20(S-protopanaxadiol], a metabolite of the protopanaxadiol-type saponins of Panax ginseng C.A. Meyer, has been reported to possess anti-tumor properties to inhibit angiogenesis and to induce tumor apoptosis. In the present study, we investigated the effect of Compound K on apoptosis and explored the underlying mechanisms involved in HL-60 human leukemia cells. Methods We examined the effect of Compound K on the viabilities of various cancer cell lines using MTT assays. DAPI assay, Annexin V and PI double staining, Western blot assay and immunoprecipitation were used to determine the effect of Compound K on the induction of apoptosis. Results Compound K was found to inhibit the viability of HL-60 cells in a dose- and time-dependent manner with an IC50 of 14 μM. Moreover, this cell death had typical features of apoptosis, that is, DNA fragmentation, DNA ladder formation, and the externalization of Annexin V targeted phosphatidylserine residues in HL-60 cells. In addition, compound-K induced a series of intracellular events associated with both the mitochondrial- and death receptor-dependent apoptotic pathways, namely, (1 the activation of caspases-3, -8, and -9; (2 the loss of mitochondrial membrane potential; (3 the release of cytochrome c and Smac/DIABLO to the cytosol; (4 the translocation of Bid and Bax to mitochondria; and (5 the downregulations of Bcl-2 and Bcl-xL. Furthermore, a caspase-8 inhibitor completely abolished caspase-3 activation, Bid cleavage, and subsequent DNA fragmentation by Compound K. Interestingly, the activation of caspase-3 and -8 and DNA fragmentation were significantly prevented in the presence of cycloheximide, suggesting that Compound K-induced apoptosis is dependent on de novo protein synthesis. Conclusions The results indicate that caspase-8 plays a key role in Compound K-stimulated apoptosis via the activation of caspase-3 directly or indirectly through

Altered binding of human histone gene transcription factors during the shutdown of proliferation and onset of differentiation in HL-60 cells

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Stein, G.; Lian, J.; Stein, J.; Shalhoub, V.; Wright, K.; Pauli, U.; Van Wijnen, A.; Briggs, R.

1989-01-01

Two sites of protein-DNA interaction have been identified in vivo and in vitro in the proximal promoter regions of an H4 and an H3 human histone gene. In proliferating cells, these genes are transcribed throughout the cell cycle, and both the more distal site I and the proximal site II are occupied by promoter-binding factors. In this report the authors demonstrate that during the shutdown of proliferation and onset of differentiation of the human promyelocytic leukemia cell line HL-60 into cells that exhibit phenotypic properties of monocytes, histone gene expression is down-regulated at the level of transcription. In vivo occupancy of site I by promoter factors persists in the differentiated HL-60 cells, but protein-DNA interactions at site II are selectively lost. Furthermore, in vitro binding activity of the site II promoter factor HiNF-D is lost in differentiated cells, and nuclear extracts from differentiated cells do not support in vitro transcription of these histone genes. The results suggest that the interaction of HiNF-D with proximal promoter site II sequences plays a primary role in rendering cell growth-regulated histone genes transcribable in proliferating cells. It appears that while cell-cycle control of histone gene expression is mediated by both transcription and mRNA stability, with the shutdown of proliferation and onset of differentiation, histone gene expression is regulated at the transcriptional level

Alterations of energy metabolism and glutathione levels of HL-60 cells induced by methacrylates present in composite resins.

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Nocca, G; De Palma, F; Minucci, A; De Sole, P; Martorana, G E; Callà, C; Morlacchi, C; Gozzo, M L; Gambarini, G; Chimenti, C; Giardina, B; Lupi, A

2007-03-01

Methacrylic compounds such as 2-hydroxyethyl methacrylate (HEMA), triethylene glycol dimethacrylate (TEGDMA) and bisphenol A glycerolate (1 glycerol/phenol) dimethacrylate (Bis-GMA) are largely present in auto- or photopolymerizable composite resins. Since the polymerization reaction is never complete, these molecules are released into the oral cavity tissues and biological fluids where they could cause local adverse effects. The aim of this work was to verify the hypothesis that the biological effects of HEMA, TEGDMA and Bis-GMA - at a non-cytotoxic concentration - depend on the interaction with mitochondria and exert consequent alterations of energy metabolism, GSH levels and the related pathways in human promyelocytic cell line (HL-60). The biological effects of methacrylic monomers were determined by analyzing the following parameters: GSH concentration, glucose-6-phosphate dehydrogenase (G6PDH) and glutathione reductase (GR) activity, oxygen and glucose consumption and lactate production along with cell differentiation and proliferation. All monomers induced both cellular differentiation and decrease in oxygen consumption. Cells treated with TEGDMA and Bis-GMA showed a significant enhancement of glucose consumption and lactate production. TEGDMA and HEMA induced GSH depletion stimulating G6PDH and GR activity. All the monomers under study affect the metabolism of HL-60 cells and show differentiating activity. Since alterations in cellular metabolism occurred at compound concentrations well below cytotoxic levels, the changes in energy metabolism and glutathione redox balance could be considered as potential mechanisms for inducing clinical and sub-clinical adverse effects and thus providing useful parameters when testing biocompatibility of dental materials.

Phospholipase D catalyzes phospholipid metabolism in chemotactic peptide-stimulated HL-60 granulocytes

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Pai, J.K.; Siegel, M.I.; Egan, R.W.; Billah, M.M.

1988-01-01

There exists circumstantial evidence for activation of phospholipase D (PLD) in intact cells. However, because of the complexity of phospholipid remodeling processes, it is essential to distinguish PLD clearly from other phospholipases and phospholipid remodeling enzymes. Therefore, to establish unequivocally PLD activity in dimethyl sulfoxide-differentiated HL-60 granulocytes, to demonstrate the relative contribution of PLD to phospholipid turnover, and to validate the hypothesis that the formation of phosphatidylethanol is an expression of PLD-catalyzed transphosphatidylation, we have developed methodologies to label HL-60 granulocytes in 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine (alkyl-PC) with 32P without labeling cellular ATP. These methodologies involve (a) synthesis of alkyl-lysoPC containing 32P by a combination of enzymatic and chemical procedures and (b) incubation of HL-60 granulocytes with this alkyl-[32P] lysoPC which enters the cell and becomes acylated into membrane-associated alkyl-[32P]PC. Upon stimulation of these 32P-labeled cells with the chemotactic peptide, N-formyl-Met-Leu-Phe (fMLP), alkyl-[32P]phosphatidic acid (alkyl-[32P]PA) is formed rapidly. Because, under these conditions, cellular ATP has not been labeled with 32P, alkyl-[32P]PA must be formed via PLD-catalyzed hydrolysis of alkyl-[32P]PC at the terminal phosphodiester bond. This result conclusively demonstrates fMLP-induced activation of PLD in HL-60 granulocytes. These 32P-labeled HL-60 granulocytes have also been stimulated in the presence of ethanol to produce alkyl-[32P]phosphatidylethanol (alkyl-[32P]PEt). Formation of alkyl-[32P]PEt parallels that of alkyl-[32P]PA with respect to time course, fMLP concentration, inhibition by a specific fMLP antagonist (t-butoxycarbonyl-Met-Leu-Phe), and Ca2+ concentration

Cytotoxicity of Vitex agnus-castus fruit extract and its major component, casticin, correlates with differentiation status in leukemia cell lines.

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Kikuchi, Hidetomo; Yuan, Bo; Nishimura, Yoshio; Imai, Masahiko; Furutani, Ryota; Kamoi, Saki; Seno, Misako; Fukushima, Shin; Hazama, Shingo; Hirobe, Chieko; Ohyama, Kunio; Hu, Xiao-Mei; Takagi, Norio; Hirano, Toshihiko; Toyoda, Hiroo

2013-12-01

We have demonstrated that an extract from the ripe fruit of Vitex agnus-castus (Vitex) exhibits cytotoxic activities against various types of solid tumor cells, whereas its effects on leukemia cells has not been evaluated to date. In this study, the effects of Vitex and its major component, casticin, on leukemia cell lines, HL-60 and U-937, were investigated by focusing on proliferation, induction of apoptosis and differentiation. Identification and quantitation by NMR spectroscopy showed that casticin accounted for approximate 1% weight of Vitex. Dose-dependent cytotoxicity of Vitex and casticin was observed in both cell lines, and HL-60 cells were more sensitive to the cytotoxicity of Vitex/casticin compared to U-937 cells. Furthermore, compared to unstimulated HL-60 cells, phorbol 12-myristate 13-acetate (PMA)- and 1,25-dihydroxyvitamin D₃ (VD₃)-differentiated HL-60 cells acquired resistance to Vitex/casticin based on the results from cell viability and apoptosis induction analysis. Since the HL-60 cell line is more immature than the U-937 cell line, these results suggested that the levels of cytotoxicity of Vitex/casticin were largely attributed to the degree of differentiation of leukemia cells; that is, cell lines with less differentiated phenotype were more susceptible than the differentiated ones. RT-PCR analysis demonstrated that PMA upregulated the expression of intercellular adhesion molecule-1 (ICAM-1) in HL-60 cells, and that anti-ICAM-1 monoclonal antibody not only abrogated PMA-induced aggregation and adhesion of the cells but also restored its sensitivity to Vitex. These results suggested that ICAM-1 plays a crucial role in the acquired resistance in PMA-differentiated HL-60 cells by contributing to cell adhesion. These findings provide fundamental insights into the clinical application of Vitex/casticin for hematopoietic malignancy.

Generation of Adducts of 4-Hydroxy-2-nonenal with Heat Shock 60 kDa Protein 1 in Human Promyelocytic HL-60 and Monocytic THP-1 Cell Lines

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Alessia Arcaro

2015-01-01

Full Text Available Heat shock 60 kDa protein 1 (HSP60 is a chaperone and stress response protein responsible for protein folding and delivery of endogenous peptides to antigen-presenting cells and also a target of autoimmunity implicated in the pathogenesis of atherosclerosis. By two-dimensional electrophoresis and mass spectrometry, we found that exposure of human promyelocytic HL-60 cells to a nontoxic concentration (10 μM of 4-hydroxy-2-nonenal (HNE yielded a HSP60 modified with HNE. We also detected adducts of HNE with putative uncharacterized protein CXorf49, the product of an open reading frame identified in various cell and tissue proteomes. Moreover, exposure of human monocytic THP-1 cells differentiated with phorbol 12-myristate 13-acetate to 10 μM HNE, and to light density lipoprotein modified with HNE (HNE-LDL or by copper-catalyzed oxidation (oxLDL, but not to native LDL, stimulated the formation of HNE adducts with HSP60, as detected by immunoprecipitation and western blot, well over basal levels. The identification of HNE-HSP60 adducts outlines a framework of mutually reinforcing interactions between endothelial cell stressors, like oxLDL and HSP60, whose possible outcomes, such as the amplification of endothelial dysfunction, the spreading of lipoxidative damage to other proteins, such as CXorf49, the activation of antigen-presenting cells, and the breaking of tolerance to HSP60 are discussed.

Regulation of C/EBPβ isoforms by MAPK pathways in HL60 cells induced to differentiate by 1,25-dihydroxyvitamin D3

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Marcinkowska, Ewa; Garay, Edward; Gocek, Elzbieta; Chrobak, Agnieszka; Wang, Xuening; Studzinski, George P.

2006-01-01

C/EBPβ is known to be important for monocytic differentiation and macrophage function. Here, we found that expression of all three C/EBPβ isoforms induced in HL60 cells by 1,25-dihydroxyvitamin D 3 (1,25D) was upregulated in a sustained manner that correlates with the appearance of monocytic phenotype and with the G1 phase cell cycle arrest. In 1,25D-resistant HL60-40AF cells, isoforms β-1 and β-3 were expressed at levels comparable to 1,25D-sensitive HL60-G cells, but isoform β-2 was difficult to detect. Treatment of sensitive HL60 cells with 1,25D resulted in predominantly nuclear localization of C/EBP isoforms β-2 and β-3, while a large proportion of C/EBPβ-1 remained in the cytoplasm. Attenuation of the MEK-ERK MAPK pathway by the inhibitor PD98059 markedly reduced the expression, 1,25D-induced phosphorylation and nuclear localization of C/EBPβ-2 and C/EBPβ-3. Interestingly, only the lower molecular mass isoforms of C/EBPβ phosphorylated on Thr235 were found in the nuclei, while C/EBPβ-1 was constitutively phosphorylated and was detected principally in the cytoplasmic fraction. Although the role of C/EBPβ isoforms in 1,25D-induced differentiation is complex, our results taken together strongly suggest that the phosphorylation of C/EBPβ isoforms on Thr235 takes place mainly via the MEK-ERK pathway and that C/EBPβ-2 is the principal transcription factor in this cell system

Proteomic analysis of cell proliferation in a human hepatic cell line (HL-7702) induced by perfluorooctane sulfonate using iTRAQ

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Cui, Ruina; Zhang, Hongxia [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Guo, Xuejiang [State Key Laboratory of Reproductive Medicine, Department of Histology and Embryology, Nanjing Medical University, Nanjing 210029 (China); Cui, Qianqian; Wang, Jianshe [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Dai, Jiayin, E-mail: daijy@ioz.ac.cn [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China)

2015-12-15

Highlights: • PFOS stimulates cell proliferation of human liver cell line (HL-7702). • Differential expressed proteins are identified by iTRAQ. • Most of differential proteins caused by PFOS are related to cell proliferation. • Up-regulation of cyclin/cdk by PFOS plays a role in driving cells into cell cycle. - Abstract: Perfluorooctane sulfonate (PFOS) is a commonly used and widely distributed perfluorinated compound proven to cause adverse health outcomes. However, how PFOS affects liver cell proliferation is not well understood. In this experiment, we exposed a human liver cell line (HL-7702) to 50 μM PFOS for 48 h and 96 h. We identified 52 differentially expressed proteins using a quantitative proteomic approach. Among them, 27 were associated with cell proliferation, including hepatoma-derived growth factor (Hdgf) and proliferation biomarkers Mk167 (Ki67) and Top2α. Results from MTT, cell counting, and cell cycle analysis showed low-dose PFOS (<200 μM) stimulated HL-7702 cell viability at 48 h and 96 h, reduced the G0/G1 percentage, and increased the S + G2/M percentage. Moreover, levels of Cyclin D1, Cyclin E2, Cyclin A2, Cyclin B1 and their partner Cdks were elevated, and the expression of regulating proteins like c-Myc, p53, p21 waf/cip1 and Myt1, as well as the phosphorylation levels of p-Wee1(S642), p-Chk1(S345) and p-Chk2(T68), were disturbed. We hypothesized that low-dose PFOS stimulated HL-7702 proliferation by driving cells into G1 through elevating cyclins/cdks expression, and by promoting cell cycle progression through altering other regulating proteins. This research will shed light on the mechanisms behind PFOS-mediated human hepatotoxicity.

Proteomic analysis of cell proliferation in a human hepatic cell line (HL-7702) induced by perfluorooctane sulfonate using iTRAQ

International Nuclear Information System (INIS)

Cui, Ruina; Zhang, Hongxia; Guo, Xuejiang; Cui, Qianqian; Wang, Jianshe; Dai, Jiayin

2015-01-01

Highlights: • PFOS stimulates cell proliferation of human liver cell line (HL-7702). • Differential expressed proteins are identified by iTRAQ. • Most of differential proteins caused by PFOS are related to cell proliferation. • Up-regulation of cyclin/cdk by PFOS plays a role in driving cells into cell cycle. - Abstract: Perfluorooctane sulfonate (PFOS) is a commonly used and widely distributed perfluorinated compound proven to cause adverse health outcomes. However, how PFOS affects liver cell proliferation is not well understood. In this experiment, we exposed a human liver cell line (HL-7702) to 50 μM PFOS for 48 h and 96 h. We identified 52 differentially expressed proteins using a quantitative proteomic approach. Among them, 27 were associated with cell proliferation, including hepatoma-derived growth factor (Hdgf) and proliferation biomarkers Mk167 (Ki67) and Top2α. Results from MTT, cell counting, and cell cycle analysis showed low-dose PFOS (<200 μM) stimulated HL-7702 cell viability at 48 h and 96 h, reduced the G0/G1 percentage, and increased the S + G2/M percentage. Moreover, levels of Cyclin D1, Cyclin E2, Cyclin A2, Cyclin B1 and their partner Cdks were elevated, and the expression of regulating proteins like c-Myc, p53, p21 waf/cip1 and Myt1, as well as the phosphorylation levels of p-Wee1(S642), p-Chk1(S345) and p-Chk2(T68), were disturbed. We hypothesized that low-dose PFOS stimulated HL-7702 proliferation by driving cells into G1 through elevating cyclins/cdks expression, and by promoting cell cycle progression through altering other regulating proteins. This research will shed light on the mechanisms behind PFOS-mediated human hepatotoxicity.

Loss of mitochondrial transmembrane potential and caspase-9 activation during apoptosis induced by the novel styryl-lactone goniothalamin in HL-60 leukemia cells.

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Inayat-Hussain, S H; Annuar, B O; Din, L B; Ali, A M; Ross, D

2003-08-01

Styryl-lactones such as goniothalamin represent a new class of compounds with potential anti-cancer properties. In this study, we investigated the mechanisms of goniothalamin (GTN), a plant styryl-lactone induced apoptosis in human promyelocytic leukemia HL-60 cells. This plant extract resulted in apoptosis in HL-60 cells as assessed by the externalisation of phosphatidylserine. Using the mitochondrial membrane dye (DIOC(6)) in conjunction with flow cytometry, we found that GTN treated HL-60 cells demonstrated a loss of mitochondrial transmembrane potential (Deltapsi(m)). Further immunoblotting on these cells showed activation of initiator caspase-9 and the executioner caspases-3 and -7. Pretreatment with the pharmacological caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp fluoromethyl ketone (Z-VAD.FMK) abrogated apoptosis as assessed by all of the apoptotic features in this study. In summary, our results demonstrate that goniothalamin-induced apoptosis occurs via the mitochondrial pathway in a caspase dependent manner.

Purification and Characterization of Glutaminase Free Asparaginase from Enterobacter cloacae: In-Vitro Evaluation of Cytotoxic Potential against Human Myeloid Leukemia HL-60 Cells.

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Islam Husain

Full Text Available Asparaginase is an important antileukemic agent extensively used worldwide but the intrinsic glutaminase activity of this enzymatic drug is responsible for serious life threatening side effects. Hence, glutaminase free asparaginase is much needed for upgradation of therapeutic index of asparaginase therapy. In the present study, glutaminase free asparaginase produced from Enterobacter cloacae was purified to apparent homogeneity. The purified enzyme was found to be homodimer of approximately 106 kDa with monomeric size of approximately 52 kDa and pI 4.5. Purified enzyme showed optimum activity between pH 7-8 and temperature 35-40°C, which is close to the internal environment of human body. Monovalent cations such as Na+ and K+ enhanced asparaginase activity whereas divalent and trivalent cations, Ca2+, Mg2+, Zn2+, Mn2+, and Fe3+ inhibited the enzyme activity. Kinetic parameters Km, Vmax and Kcat of purified enzyme were found to be 1.58×10-3 M, 2.22 IU μg-1 and 5.3 × 104 S-1, respectively. Purified enzyme showed prolonged in vitro serum (T1/2 = ~ 39 h and trypsin (T1/2 = ~ 32 min half life, which is therapeutically remarkable feature. The cytotoxic activity of enzyme was examined against a panel of human cancer cell lines, HL-60, MOLT-4, MDA-MB-231 and T47D, and highest cytotoxicity observed against HL-60 cells (IC50 ~ 3.1 IU ml-1, which was comparable to commercial asparaginase. Cell and nuclear morphological studies of HL-60 cells showed that on treatment with purified asparaginase symptoms of apoptosis were increased in dose dependent manner. Cell cycle progression analysis indicates that enzyme induces apoptosis by cell cycle arrest in G0/G1 phase. Mitochondrial membrane potential loss showed that enzyme also triggers the mitochondrial pathway of apoptosis. Furthermore, the enzyme was found to be nontoxic for human noncancerous cells FR-2 and nonhemolytic for human erythrocytes.

Benzoquinone activates the ERK/MAPK signaling pathway via ROS production in HL-60 cells

International Nuclear Information System (INIS)

Ruiz-Ramos, Ruben; Cebrian, Mariano E.; Garrido, Efrain

2005-01-01

Benzene (BZ) is a class I carcinogen and its oxidation to reactive intermediates is a prerequisite of hematoxicity and myelotoxicity. The generated metabolites include hydroquinone, which is further oxidized to the highly reactive 1,4-benzoquinone (BQ) in bone marrow. Therefore, we explored the mechanisms underlying BQ-induced HL-60 cell proliferation by studying the role of BQ-induced reactive oxygen species (ROS) in the activation of the ERK-MAPK signaling pathway. BQ treatment (0.01-30 μM) showed that doses below 10 μM did not significantly reduce viability. ROS production after 3 μM BQ treatment increased threefold; however, catalase addition reduced ROS generation to basal levels. FACS analysis showed that BQ induced a fivefold increase in the proportion of cells in S-phase. We also observed a high proportion of Bromodeoxyuridine (BrdU) stained cells, indicating a higher DNA synthesis rate. BQ also produced rapid and prolonged phosphorylation of ERK1/2 proteins. Simultaneous treatment with catalase or PD98059, a potent MEK protein inhibitor, reduced cell recruitment into the S-phase and also abolished the ERK1/2 protein phosphorylation induced by BQ, suggesting that MEK/ERK is an important pathway involved in BQ-induced ROS mediated proliferation. The prolonged activation of ERK1/2 contributes to explain the increased S-phase cell recruitment and to understand the leukemogenic processes associated with exposure to benzene metabolites. Thus, the possible mechanism by which BQ induce HL-60 cells to enter the cell cycle and proliferate is linked to ROS production and its growth promoting effects by specific activation of regulating genes known to be activated by redox mechanisms

Synthesis and Biological Activity of Diastereomeric and Geometric Analogs of Calcipotriol, PRI-2202 and PRI-2205, Against Human HL-60 Leukemia and MCF-7 Breast Cancer Cells.

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Milczarek, Magdalena; Chodyński, Michał; Filip-Psurska, Beata; Martowicz, Agnieszka; Krupa, Małgorzata; Krajewski, Krzysztof; Kutner, Andrzej; Wietrzyk, Joanna

2013-10-31

Diastereomeric and geometric analogs of calcipotriol, PRI-2202 and PRI-2205, were synthesized as advanced intermediates from vitamin D C-22 benzothiazoyl sulfones and side-chain aldehydes using our convergent strategy. Calcitriol, calcipotriol (PRI-2201) and tacalcitol (PRI-2191) were used as the reference compounds. Among a series of tested analogs the diastereomeric analog PRI-2202 showed the strongest antiproliferative activity on the human breast cancer cell line MCF-7, whereas the geometric analog PRI-2205 was the weakest. Both analogs were less potent in antiproliferative activity against HL-60 cells compared to the reference compounds. The ability to potentiate antiproliferative effect of cisplatin or doxorubicin against HL-60 cells or that of tamoxifen against the MCF-7 cell line was observed at higher doses of PRI-2202 or PRI-2205 than those of the reference compounds. The proapoptotic activity of tamoxifen, expressed as the diminished mitochondrial membrane potential, as well as the increased phosphatidylserine expression, was partially attenuated by calcitriol, PRI-2191, PRI-2201 and PRI-2205. The treatment of the MCF-7 cells with tamoxifen alone resulted in an increase in VDR expression. Moreover, a further increase in VDR expression was observed when the analogs PRI-2201 or PRI-2205, but not PRI-2191, were used in combination with tamoxifen. This observation could partially explain the potentiation of the antiproliferative effect of tamoxifen by vitamin D analogs.

Elimination of clonogenic tumor cells from HL-60, Daudi, and U-937 cell lines by laser photoradiation therapy: implications for autologous bone marrow purging

International Nuclear Information System (INIS)

Gulliya, K.S.; Pervaiz, S.

1989-01-01

Laser photoradiation therapy was tested in an in vitro model for its efficacy in the elimination of non-Hodgkin's lymphoma cells. Results show that at 31.2 J/cm2 of laser light in the presence of 20 micrograms/mL of merocyanine 540 (MC540) there was greater than 5 log reduction in Burkitt's lymphoma (Daudi) cells. Similar tumor cell kill was obtained for leukemia (HL-60) cells at a laser light dose of 93.6 J/cm2. However, to obtain the same efficiency of killing for histiocytic lymphoma (U-937) cells, a higher dose of MC540 (25 micrograms/mL) was required. Clonogenic tumor stem cell colony formation was reduced by greater than 5 logs after laser photoradiation therapy. Under identical conditions for each cell line the percent survival for granulocyte-macrophage colony-forming units (CFU-GM, 45.9%, 40%, 17.5%), granulocyte/erythroid/macrophage/megakaryocyte (GEMM, 40.1%, 20.1%, 11.5%), colony-forming units (CFU-C, 16.2%, 9.1%, 1.8%), and erythroid burst-forming units (BFU-E, 33.4%, 17.8%, 3.9%) was significantly higher than the tumor cells. Mixing of gamma ray-irradiated normal marrow cells with tumor cells (1:1 and 10:1 ratio) did not interfere with the elimination of tumor cells. The effect of highly purified recombinant interferon alpha (rIFN) on laser photoradiation therapy of tumor cells was also investigated. In the presence of rIFN (30 to 3,000 U/mL), the viability of leukemic cells was observed to increase from 0% to 1.5% with a concurrent decrease in membrane polarization, suggesting an increase in fluidity of cell membrane in response to rIFN. However, at higher doses of rIFN (6,000 to 12,000 U/mL) this phenomenon was not observed. The viability of lymphoma cells remained unaffected at all doses of rIFN tested

C60 Fullerene Effects on Diphenyl-N-(trichloroacetyl)-amidophosphate Interaction with DNA In Silico and Its Cytotoxic Activity Against Human Leukemic Cell Line In Vitro

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Grebinyk, A.; Prylutska, S.; Grynyuk, I.; Kolp, B.; Hurmach, V.; Sliva, T.; Amirkhanov, V.; Trush, V.; Matyshevska, O.; Slobodyanik, M.; Prylutskyy, Yu.; Frohme, M.; Ritter, U.

2018-03-01

New representative of carbacylamidophosphates - diphenyl-N-(trichloroacetyl)-amidophosphate (HL), which contains two phenoxy substituents near the phosphoryl group, was synthesized, identified by elemental analysis and IR and NMR spectroscopy, and tested as a cytotoxic agent itself and in combination with C60 fullerene. According to molecular simulation results, C60 fullerene and HL could interact with DNA and form a rigid complex stabilized by stacking interactions of HL phenyl groups with C60 fullerene and DNA G nucleotide, as well as by interactions of HL CCl3 group by ion-π bonds with C60 molecule and by electrostatic bonds with DNA G nucleotide. With the use of MTT test, the cytotoxic activity of HL against human leukemic CCRF-CM cells with IC50 value detected at 10 μM concentration at 72 h of cells treatment was shown. Under combined action of 16 μM C60 fullerene and HL, the value of IC50 was detected at lower 5 μM HL concentration and at earlier 48 h period of incubation, besides the cytotoxic effect of HL was observed at a low 2.5 μM concentration at which HL by itself had no influence on cell viability. Binding of C60 fullerene and HL with minor DNA groove with formation of a stable complex is assumed to be one of the possible reasons of their synergistic inhibition of CCRF-CEM cells proliferation. Application of C60 fullerene in combination with 2.5 μM HL was shown to have no harmful effect on structural stability of blood erythrocytes membrane. Thus, combined action of C60 fullerene and HL in a low concentration potentiated HL cytotoxic effect against human leukemic cells and was not followed by hemolytic effect.

Taraxinic acid, a hydrolysate of sesquiterpene lactone glycoside from the Taraxacum coreanum NAKAI, induces the differentiation of human acute promyelocytic leukemia HL-60 cells.

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Choi, Jung-Hye; Shin, Kyung-Min; Kim, Na-Young; Hong, Jung-Pyo; Lee, Yong Sup; Kim, Hyoung Ja; Park, Hee-Juhn; Lee, Kyung-Tae

2002-11-01

The present work was performed to elucidate the active moiety of a sesquiterpene lactone, taraxinic acid-1'-O-beta-D-glucopyranoside (1). from Taraxacum coreanum NAKAI on the cytotoxicity of various cancer cells. Based on enzymatic hydrolysis and MTT assay, the active moiety should be attributed to the aglycone taraxinic acid (1a). rather than the glycoside (1). Taraxinic acid exhibited potent antiproliferative activity against human leukemia-derived HL-60. In addition, this compound was found to be a potent inducer of HL-60 cell differentiation as assessed by a nitroblue tetrazolium reduction test, esterase activity assay, phagocytic activity assay, morphology change, and expression of CD 14 and CD 66 b surface antigens. These results suggest that taraxinic acid induces the differentiation of human leukemia cells to monocyte/macrophage lineage. Moreover, the expression level of c-myc was down-regulated during taraxinic acid-dependent HL-60 cell differentiation, whereas p21(CIP1) and p27(KIP1) were up-regulated. Taken together, our results suggest that taraxinic acid may have potential as a therapeutic agent in human leukemia.

Synthesis and Biological Activity of Diastereomeric and Geometric Analogs of Calcipotriol, PRI-2202 and PRI-2205, Against Human HL-60 Leukemia and MCF-7 Breast Cancer Cells

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Andrzej Kutner

2013-10-01

Full Text Available Diastereomeric and geometric analogs of calcipotriol, PRI-2202 and PRI-2205, were synthesized as advanced intermediates from vitamin D C-22 benzothiazoyl sulfones and side-chain aldehydes using our convergent strategy. Calcitriol, calcipotriol (PRI-2201 and tacalcitol (PRI-2191 were used as the reference compounds. Among a series of tested analogs the diastereomeric analog PRI-2202 showed the strongest antiproliferative activity on the human breast cancer cell line MCF-7, whereas the geometric analog PRI-2205 was the weakest. Both analogs were less potent in antiproliferative activity against HL-60 cells compared to the reference compounds. The ability to potentiate antiproliferative effect of cisplatin or doxorubicin against HL-60 cells or that of tamoxifen against the MCF-7 cell line was observed at higher doses of PRI-2202 or PRI-2205 than those of the reference compounds. The proapoptotic activity of tamoxifen, expressed as the diminished mitochondrial membrane potential, as well as the increased phosphatidylserine expression, was partially attenuated by calcitriol, PRI-2191, PRI-2201 and PRI-2205. The treatment of the MCF-7 cells with tamoxifen alone resulted in an increase in VDR expression. Moreover, a further increase in VDR expression was observed when the analogs PRI-2201 or PRI-2205, but not PRI-2191, were used in combination with tamoxifen. This observation could partially explain the potentiation of the antiproliferative effect of tamoxifen by vitamin D analogs.

Cell cycle sensitivity of HL-60 cells to the differentiation-inducing effects of 1-alpha,25-dihydroxyvitamin D3

International Nuclear Information System (INIS)

Studzinski, G.P.; Bhandal, A.K.; Brelvi, Z.S.

1985-01-01

A recently described system for monocyte-like differentiation of HL-60 cells was utilized to determine if the initiation of this pathway can be linked to a set of replicative cellular events. The standard induction system consisted of a 4-h exposure to 100 nM 1-alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3] followed by determination of nonspecific esterase and phagocytic activity 24 h later. The cell cycle status was ascertained by the incorporation of [ 3 H]thymidine and autoradiography. Studies in which cell cycle block in the G1/S phase boundary region was produced by a partial inhibition of DNA synthesis with thymidine, or sodium butyrate, showed that the exposure of such semisynchronous cultures to 1,25(OH)2D3 resulted in an increased proportion of differentiated cells. Conversely, blocking the cell cycle with vinblastine (G2/M block) or theobromine (mid-G1 block) inhibited the initiation of differentiation by 1,25(OH)2D3. Experiments in which the differentiated cells were examined for the cell cycle position at the time of the exposure to 1,25(OH)2D3 by [ 3 H]thymidine labeling and autoradiography confirmed that the late G1 and early S phase cells are those which predominate in the differentiated fraction of 1,25(OH)2D3-treated HL-60 cultures. These results link pre- and early replicative cellular events to the induction of monocytic differentiation by 1,25(OH)2D3

Kaempferol induces DNA damage and inhibits DNA repair associated protein expressions in human promyelocytic leukemia HL-60 cells.

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Wu, Lung-Yuan; Lu, Hsu-Feng; Chou, Yu-Cheng; Shih, Yung-Luen; Bau, Da-Tian; Chen, Jaw-Chyun; Hsu, Shu-Chun; Chung, Jing-Gung

2015-01-01

Numerous evidences have shown that plant flavonoids (naturally occurring substances) have been reported to have chemopreventive activities and protect against experimental carcinogenesis. Kaempferol, one of the flavonoids, is widely distributed in fruits and vegetables, and may have cancer chemopreventive properties. However, the precise underlying mechanism regarding induced DNA damage and suppressed DNA repair system are poorly understood. In this study, we investigated whether kaempferol induced DNA damage and affected DNA repair associated protein expression in human leukemia HL-60 cells in vitro. Percentages of viable cells were measured via a flow cytometry assay. DNA damage was examined by Comet assay and DAPI staining. DNA fragmentation (ladder) was examined by DNA gel electrophoresis. The changes of protein levels associated with DNA repair were examined by Western blotting. Results showed that kaempferol dose-dependently decreased the viable cells. Comet assay indicated that kaempferol induced DNA damage (Comet tail) in a dose-dependent manner and DAPI staining also showed increased doses of kaempferol which led to increased DNA condensation, these effects are all of dose-dependent manners. Western blotting indicated that kaempferol-decreased protein expression associated with DNA repair system, such as phosphate-ataxia-telangiectasia mutated (p-ATM), phosphate-ataxia-telangiectasia and Rad3-related (p-ATR), 14-3-3 proteins sigma (14-3-3σ), DNA-dependent serine/threonine protein kinase (DNA-PK), O(6)-methylguanine-DNA methyltransferase (MGMT), p53 and MDC1 protein expressions, but increased the protein expression of p-p53 and p-H2AX. Protein translocation was examined by confocal laser microscopy, and we found that kaempferol increased the levels of p-H2AX and p-p53 in HL-60 cells. Taken together, in the present study, we found that kaempferol induced DNA damage and suppressed DNA repair and inhibited DNA repair associated protein expression in HL-60

Cyclic AMP-dependent protein kinase interferes with GTP γS stimulated IP3 formation in differentiated HL-60 cell membranes

International Nuclear Information System (INIS)

Misaki, Naoyuki; Imaizumi, Taro; Watanabe, Yashuiro

1989-01-01

The effects of addition of activated cyclic AMP-dependent protein kinase (PKA) on the function of islet-activating protein (IAP)-sensitive GTP-binding (G) protein were studied in the plasma membranes of 3 H-inositol-labeled differentiated human leukemic (HL-60) cells. Pretreatment of the membranes with activated PKA in the presence of MgATP for 15 min. at 37 degree C decreased GTP γS-stimulated inositol trisphosphate (IP 3 ) formation by about 30%, but had no influence on Ca 2+ -stimulated IP 3 formation. And autoradiography in the phosphorylation experiments of solubilized HL-60 cell membranes by PKA showed some 32 P incorporated bands, and among them one of the major bands showed the migration at 40 kDa supporting that the G protein coupling with PI response was phosphorylated by PKA. These results showed that pretreatment with activated PKA inhibited the mediating function of the G protein between the fMLP receptor and phospholipase C by its phosphorylation

Isolation of furocoumarins from bergamot fruits as HL-60 differentiation-inducing compounds.

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Kawaii, S; Tomono, Y; Katase, E; Ogawa, K; Yano, M

1999-10-01

The HL-60 differentiation-inducing compounds in bergamot fruits were isolated with column chromatography and identified as bergamottin, bergapten, and citropten by (1)H and (13)C NMR. Their HL-60 differentiation-inducing activity was measured by examining nitro blue tetrazolium (NBT) reducing, nonspecific acid esterase (NSE), specific esterase (SE), and phagocytic activities, and bergamottin showed the strongest activity among the coumarins isolated from bergamot fruits. The structure-activity relationship obtained from HL-60 differentiation assay suggests that hydrophobicity of furocoumarins is correlated with their activity.

Nano-hole induction by nanodiamond and nanoplatinum liquid, DPV576, reverses multidrug resistance in human myeloid leukemia (HL60/AR

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Ghoneum A

2013-07-01

Full Text Available Alia Ghoneum,1,2 Shivani Sharma,1,3 James Gimzewsk1,3 1Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, 2Department of Otalaryngology, Drew University of Medicine and Science, Los Angeles, 3California Nanosystems Institute (CNSI at University of California, Los Angeles, Los Angeles, CA, USA Abstract: Recently nanoparticles have been extensively studied and have proven to be a promising candidate for cancer treatment and diagnosis. In the current study, we examined the chemo-sensitizing activity of a mixture of nanodiamond (ND and nanoplatinum (NP solution known as DPV576, against multidrug-resistant (MDR human myeloid leukemia (HL60/AR and MDR-sensitive cells (HL60. Cancer cells were cultured with different concentrations of daunorubicin (DNR (1 × 10-9–1 × 10-6 M in the presence of selected concentrations of DPV576 (2.5%–10% v/v. Cancer cell survival was determined by MTT assay, drug accumulation by flow cytometry and confocal laser scanning microscopy (CLSM, and holes and structural changes by atomic force microscopy (AFM. Co-treatment of HL60/AR cells with DNR plus DPV576 resulted in the reduction of the IC50 to 1/4th. This was associated with increased incidences of holes inside the cells as compared with control untreated cells. On the other hand, HL60 cells did not show changes in their drug accumulation post-treatment with DPV576 and DNR. We conclude that DPV576 is an effective chemo-sensitizer as indicated by the reversal of HL60/AR cells to DNR and may represent a potential novel adjuvant for the treatment of chemo-resistant human myeloid leukemia. Keywords: nanodiamond, nanoplatinum, daunorubicin, flow cytometry, AFM

DNA fingerprinting of the NCI-60 cell line panel.

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Lorenzi, Philip L; Reinhold, William C; Varma, Sudhir; Hutchinson, Amy A; Pommier, Yves; Chanock, Stephen J; Weinstein, John N

2009-04-01

The National Cancer Institute's NCI-60 cell line panel, the most extensively characterized set of cells in existence and a public resource, is frequently used as a screening tool for drug discovery. Because many laboratories around the world rely on data from the NCI-60 cells, confirmation of their genetic identities represents an essential step in validating results from them. Given the consequences of cell line contamination or misidentification, quality control measures should routinely include DNA fingerprinting. We have, therefore, used standard DNA microsatellite short tandem repeats to profile the NCI-60, and the resulting DNA fingerprints are provided here as a reference. Consistent with previous reports, the fingerprints suggest that several NCI-60 lines have common origins: the melanoma lines MDA-MB-435, MDA-N, and M14; the central nervous system lines U251 and SNB-19; the ovarian lines OVCAR-8 and OVCAR-8/ADR (also called NCI/ADR); and the prostate lines DU-145, DU-145 (ATCC), and RC0.1. Those lines also show that the ability to connect two fingerprints to the same origin is not affected by stable transfection or by the development of multidrug resistance. As expected, DNA fingerprints were not able to distinguish different tissues-of-origin. The fingerprints serve principally as a barcodes.

Downregulation of telomerase activity in human promyelocytic cell line using RNA interference.

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Miri-Moghaddam, E; Deezagi, A; Soheili, Z S

2009-12-01

Telomerase is a ribonucleoprotein complex. It consists of two main components, human telomerase reverse transcriptase (hTERT) and human telomerase RNA. High telomerase activity is present in most malignant cells, but it is barely detectable in majority of somatic cells. The direct correlation between telomerase reactivation and carcinogens has made hTERT a key target for anticancer therapeutic studies. In this study, for the first time, we evaluated the ability of the new generation of short interfering RNA (siRNA) to regulate telomerase activity in the human promyelocytic leukemia cell line (HL-60). Transient transfection cell line by hTERT siRNAs resulted in statistically significant suppression of hTERT messenger RNAs which were detected by quantitative real-time polymerase chain reaction, while the expressed hTERT protein levels were measured by flow cytometry. The results of telomeric repeat amplification protocol showed that telomerase activity was significantly reduced upon transfection of the HL-60 cell line with hTERT siRNAs. The results of this study showed that telomerase activity and cell proliferation were efficiently inhibited in the hTERT siRNA-treated leukemic cell line.

Retinoic acid-induced granulocytic differentiation of HL60 human promyelocytic leukemia cells is preceded by downregulation of autonomous generation of inositol lipid-derived second messengers

International Nuclear Information System (INIS)

Porfiri, E.; Hoffbrand, A.V.; Wickremasinghe, R.G.

1991-01-01

Inositol phosphates (InsPs) and diacyglycerol (DAG) are second messengers derived via the breakdown of inositol phospholipids, and which play important signalling roles in the regulation of proliferation of some cell types. The authors have studied the operation of this pathway during the early stages of retionic acid (RA)-induced granulocytic differentiation of HL60 myeloid leukemia cells. The autonomous breakdown of inositol lipids that occurred in HL60 cells labeled with [3H] inositol was completely abolished following 48 hours of RA treatment. The rate of influx of 45Ca2+ was also significantly decreased at 48 hours, consistent with the role of inositol lipid-derived second messengers in regulating Ca2+ entry into cells. The downregulation of inositol lipid metabolism clearly preceded the onset of reduced proliferation induced by RA treatment, and was therefore not a consequence of decreased cell growth. The generation of InsPs in RA-treated cells was reactivated by the fluoroaluminate ion, a direct activator of guanine nucleotide-binding protein(s) (G proteins) that regulate the inositol lipid signalling pathway. Subtle alterations to a regulatory mechanism may therefore mediate the RA-induced downregulation of this pathway. The data are consistent with the hypothesis that the autonomous generation of inositol lipid-derived second messengers may contribute to the continuous proliferation of HL60 cells, and that the RA-induced downregulation of this pathway may, in turn, play a role in signalling the cessation of proliferation that preceedes granulocytic differentiation

An Effective Model of the Retinoic Acid Induced HL-60 Differentiation Program.

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Tasseff, Ryan; Jensen, Holly A; Congleton, Johanna; Dai, David; Rogers, Katharine V; Sagar, Adithya; Bunaciu, Rodica P; Yen, Andrew; Varner, Jeffrey D

2017-10-30

In this study, we present an effective model All-Trans Retinoic Acid (ATRA)-induced differentiation of HL-60 cells. The model describes reinforcing feedback between an ATRA-inducible signalsome complex involving many proteins including Vav1, a guanine nucleotide exchange factor, and the activation of the mitogen activated protein kinase (MAPK) cascade. We decomposed the effective model into three modules; a signal initiation module that sensed and transformed an ATRA signal into program activation signals; a signal integration module that controlled the expression of upstream transcription factors; and a phenotype module which encoded the expression of functional differentiation markers from the ATRA-inducible transcription factors. We identified an ensemble of effective model parameters using measurements taken from ATRA-induced HL-60 cells. Using these parameters, model analysis predicted that MAPK activation was bistable as a function of ATRA exposure. Conformational experiments supported ATRA-induced bistability. Additionally, the model captured intermediate and phenotypic gene expression data. Knockout analysis suggested Gfi-1 and PPARg were critical to the ATRAinduced differentiation program. These findings, combined with other literature evidence, suggested that reinforcing feedback is central to hyperactive signaling in a diversity of cell fate programs.

Anticancer Potential of Aqueous Ethanol Seed Extract of Ziziphus mauritiana against Cancer Cell Lines and Ehrlich Ascites Carcinoma

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Tulika Mishra

2011-01-01

Full Text Available Ziziphus mauritiana (Lamk. is a fruit tree that has folkloric implications against many ailments and diseases. In the present study, anticancer potential of seed extract of Ziziphus mauritiana in vitro against different cell lines (HL-60, Molt-4, HeLa, and normal cell line HGF by MTT assay as well as in vivo against Ehrich ascites carcinoma bearing Swiss albino mice was investigated. The extract was found to markedly inhibit the proliferation of HL-60 cells. Annexin and PI binding of treated HL-60 cells indicated apoptosis induction by extract in a dose-dependent manner. The cell cycle analysis revealed a prominent increase in sub Go population at concentration of 20 μg/ml and above. Agarose gel electrophoresis confirmed DNA fragmentation in HL-60 cells after 3 h incubation with extract. The extract also exhibited potent anticancer potential in vivo. Treatment of Ehrlich ascites carcinoma bearing Swiss albino mice with varied doses (100–800 mg/kg b.wt. of plant extract significantly reduced tumor volume and viable tumor cell count and improved haemoglobin content, RBC count, mean survival time, tumor inhibition, and percentage life span. The enhanced antioxidant status in extract-treated animals was evident from decline in levels of lipid peroxidation and increased levels of glutathione, catalase, and superoxide dismutase.

Cellular uptake of 99mTcN-NOET in human leukaemic HL-60 cells is related to calcium channel activation and cell proliferation

International Nuclear Information System (INIS)

Guillermet, Stephanie; Vuillez, Jean-Philippe; Caravel, Jean-Pierre; Marti-Batlle, Daniele; Fagret, Daniel; Fontaine, Eric; Pasqualini, Roberto

2006-01-01

A major goal of nuclear oncology is the development of new radiolabelled tracers as proliferation markers. Intracellular calcium waves play a fundamental role in the course of the cell cycle. These waves occur in non-excitable tumour cells via store-operated calcium channels (SOCCs). Bis(N-ethoxy, N-ethyldithiocarbamato) nitrido technetium (V)-99m ( 99m TcN-NOET) has been shown to interact with L-type voltage-operated calcium channels (VOCCs) in cultured cardiomyocytes. Considering the analogy between VOCCs and SOCCs, we sought to determine whether 99m TcN-NOET also binds to activated SOCCs in tumour cells in order to clarify the potential value of this tracer as a proliferation marker. Uptake kinetics of 99m TcN-NOET were measured in human leukaemic HL-60 cells over 60 min and the effect of several calcium channel modulators on 1-min tracer uptake was studied. The uptake kinetics of 99m TcN-NOET were compared both with the variations of cytosolic free calcium concentration measured by indo-1/AM and with the variations in the SG 2 M cellular proliferation index. All calcium channel inhibitors significantly decreased the cellular uptake of 99m TcN-NOET whereas the activator thapsigargin induced a significant 10% increase. In parallel, SOCC activation by thapsigargin, as measured using the indo-1/AM probe, was inhibited by nicardipine. These results indicate that the uptake of 99m TcN-NOET is related to the activation of SOCCs. Finally, a correlation was observed between the tracer uptake and variations in the proliferation index SG 2 M. The uptake of 99m TcN-NOET seems to be related to SOCC activation and to cell proliferation in HL-60 cells. These results indicate that 99m TcN-NOET might be a marker of cell proliferation. (orig.)

The Comparative PDT Experiment of the Inactivation of HL60 on Modified TiO2 Nanoparticles

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Kaiqi Lu

2015-01-01

Full Text Available Four samples of modified titanium dioxide (TiO2, Fe/TiO2 (2 wt%, Fe/TiO2 (5 wt%, and 5-ALA/TiO2, were experimented in photodynamic therapy (PDT on leukemia cells HL60, performing promising photocatalytic inactivation effect. Fe/TiO2 and 5-ALA/TiO2 were synthesized in methods of precipitation and ultrasonic methods, respectively. X-ray diffraction spectra and UV-Vis spectra were studied for the samples’ crystalline phase and redshift of absorption peak. Further, FTIR spectra and Raman spectra were obtained to examine the combination of 5-aminolevulinic (5-ALA and TiO2 nanoparticles. The toxicity of these four kinds of nanoparticles was studied through darkroom experiments. And based on the concentration which caused the same toxic effect (90% on HL60, PDT experiments of TiO2, Fe/TiO2 (2%, Fe/TiO2 (5%, and ALA/TiO2 were done, resulting in the fact that the photokilling efficiency was 69.7%, 71.6%, 72%, and 80.6%, respectively. Scanning electron microscope (SEM images of the samples were also taken to study the morphology of HL60 cells before and after PDT, resulting in the fact the activation of the modified TiO2 from PDT was the main cause of cell apoptosis.

19-nor vitamin-D analogs: a new class of potent inhibitors of proliferation and inducers of differentiation of human myeloid leukemia cell lines.

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Asou, H; Koike, M; Elstner, E; Cambell, M; Le, J; Uskokovic, M R; Kamada, N; Koeffler, H P

1998-10-01

We have studied the in vitro biological activities and mechanisms of action of 1,25-dihydroxyvitamin D3 (1,25D3) and nine potent 1,25D3 analogs on proliferation and differentiation of myeloid leukemia cell lines (HL-60, retinoic acid-resistant HL-60 [RA-res HL-60], NB4 and Kasumi-1). The common novel structural motiff for almost all the analogs included removal of C-19 (19-nor); each also had unsaturation of the side chain. All the compounds were potent; for example, the concentration of analogs producing a 50% clonal inhibition (ED50) ranged between 1 x 10(-9) to 4 x 10(-11) mol/L when using the HL-60 cell line. The most active compound [1, 25(OH)2-16,23E-diene-26-trifluoro-19-nor-cholecalciferol (Ro 25-9716)] had an ED50 of 4 x 10(-11) mol/L; in contrast, the 1,25D3 produced an ED50 of 10(-9) mol/L with the HL-60 target cells. Ro 25-9716 (10(-9) mol/L, 3 days) was a strong inducer of myeloid differentiation because it caused 92% of the HL-60 cells to express CD11b and 75% of these cells to reduce nitroblue tetrazolium (NBT). This compound (10(-8) mol/L, 4 days) also caused HL-60 cells to arrest in the G1 phase of the cell cycle (88% cells in G1 v 48% of the untreated control cells). The p27(kip-1), a cyclin-dependent kinase inhibitor which is important in blocking the cell cycle, was induced more quickly and potently by Ro 25-9716 (10(-7) mol/L, 0 to 5 days) than by 1,25D3, suggesting a possible mechanism by which these analogs inhibit proliferation of leukemic growth. The NB4 promyelocytic leukemia cells cultured with the Ro 25-9716 were also inhibited in their clonal proliferation (ED50, 5 x 10(-11) mol/L) and their expression of CD11b was enhanced (80% positive [10(-9) mol/L, 4 days] v 27% untreated NB4 cells). Moreover, the combination of Ro 25-9716 (10(-9) mol/L) and all-trans retinoic acid (ATRA, 10(-7) mol/L) induced 92% of the NB4 cells to reduce NBT, whereas only 26% of the cells became NBT positive after a similar exposure to the combination of 1,25D3

Andrographolide interferes with binding of nuclear factor-κB to DNA in HL-60-derived neutrophilic cells

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Hidalgo, María A; Romero, Alex; Figueroa, Jaime; Cortés, Patricia; Concha, Ilona I; Hancke, Juan L; Burgos, Rafael A

2005-01-01

Andrographolide, the major active component from Andrographis paniculata, has shown to possess anti-inflammatory activity. Andrographolide inhibits the expression of several proinflammatory proteins that exhibit a nuclear factor kappa B (NF-κB) binding site in their gene. In the present study, we analyzed the effect of andrographolide on the activation of NF-κB induced by platelet-activating factor (PAF) and N-formyl-methionyl-leucyl-phenylalanine (fMLP) in HL-60 cells differentiated to neutrophils. PAF (100 nM) and fMLP (100 nM) induced activation of NF-κB as determined by degradation of inhibitory factor B α (IκBα) using Western blotting in cytosolic extracts and by binding to DNA using electrophoretic mobility shift assay (EMSA) in nuclear extracts. Andrographolide (5 and 50 μM) inhibited the NF-κB-luciferase activity induced by PAF. However, andrographolide did not reduce phosphorylation of p38 MAPK or ERK1/2 and did not change IκBα degradation induced by PAF and fMLP. Andrographolide reduced the DNA binding of NF-κB in whole cells and in nuclear extracts induced by PAF and fMLP. Andrographolide reduced cyclooxygenase-2 (COX-2) expression induced by PAF and fMLP in HL-60/neutrophils. It is concluded that andrographolide exerts its anti-inflammatory effects by inhibiting NF-κB binding to DNA, and thus reducing the expression of proinflammatory proteins, such as COX-2. PMID:15678086

Effect of a 2.45-GHz radiofrequency electromagnetic field on neutrophil chemotaxis and phagocytosis in differentiated human HL-60 cells.

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Koyama, Shin; Narita, Eijiro; Suzuki, Yoshihisa; Taki, Masao; Shinohara, Naoki; Miyakoshi, Junji

2015-01-01

The potential public health risks of radiofrequency (RF) fields have been discussed at length, especially with the use of mobile phones spreading extensively throughout the world. In order to investigate the properties of RF fields, we examined the effect of 2.45-GHz RF fields at the specific absorption rate (SAR) of 2 and 10 W/kg for 4 and 24 h on neutrophil chemotaxis and phagocytosis in differentiated human HL-60 cells. Neutrophil chemotaxis was not affected by RF-field exposure, and subsequent phagocytosis was not affected either compared with that under sham exposure conditions. These studies demonstrated an initial immune response in the human body exposed to 2.45-GHz RF fields at the SAR of 2 W/kg, which is the maximum value recommended by the International Commission for Non-Ionizing Radiation Protection (ICNIRP) guidelines. The results of our experiments for RF-field exposure at an SAR under 10 W/kg showed very little or no effects on either chemotaxis or phagocytosis in neutrophil-like human HL-60 cells. © The Author 2014. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.

Biochemical studies of the differentiation of HL-60 cells into monocytes by either IFN, VIT, D3 or TPA

International Nuclear Information System (INIS)

Miyamoto, S.; Whyzmuzis, C.; Oronsky, B.; Wu, J.M.

1987-01-01

The authors have studied the differentiation process of the human promyelocytic cell line, HL-60, by treatment of these cells with either gamma interferon, 1, 25 dihydroxyvitamin D 3 or a phorbol ester, TPA. The cells were grown in RPMI 1640, 10% FCS with each respective agent, then pulsed labeled with 35 S-Met, harvested, lysed and subfractionated by centrifugation into post-ribosomal and ribosomal salt was fractions (RSW). These fractions were examined by SDS gel electrophoresis. The culture supernatant from the treated cells was dialyzed and passed over a heparin agarose affinity column. The absorbed material was eluted from the column by a step-wise salt gradient and analyzed by SDS gel electrophoresis. They have also observed that in a rabbit reticulocyte lysate assay, the RSW from control cells show inhibition of protein synthesis. The RSW from cells treated with either high concentrations (200-1000 units/ml) of gamma interferon, Vit D 3 or TPA did not show this inhibition. Some possible explanations for this phenomenon are the loss or inactivation of a component necessary for protein synthesis which is triggered by differentiation, or the differentiation-related modulation of translational inhibitor(s). They have used FPLC to further analyze the RSW, but because the factor(s) are present in such small quantities further analytical and more sensitive procedures need to be pursued

Isolation of HL-60 cancer cells from the human serum sample using MnO2-PEI/Ni/Au/aptamer as a novel nanomotor and electrochemical determination of thereof by aptamer/gold nanoparticles-poly(3,4-ethylene dioxythiophene) modified GC electrode.

Science.gov (United States)

Amouzadeh Tabrizi, Mahmoud; Shamsipur, Mojtaba; Saber, Reza; Sarkar, Saeed

2018-07-01

Herein, aptamer-modified self-propelled nanomotors were used for transportation of human promyelocytic leukemia cells (HL-60) from a human serum sample. For this purpose, the fabricated manganese oxide nanosheets-polyethyleneimine decorated with nickel/gold nanoparticles (MnO 2 -PEI/Ni/Au) as nanomotors were added to a vial containing thiolated aptamer KH1C12 solution as a capture aptamer to attach to the gold nanoparticles on the surface of nanomotors covalently. The aptamer-modified self-propelled nanomotors (aptamer KH1C12 /nanomotors) were then separated by placing the vial in a magnetic stand. The aptamer-modified self-propelled nanomotors were rinsed three times with water to remove the non-attached aptamers. Then, the resulting aptamer KH1C12 /nanomotors were applied for the on-the-fly" transporting of HL-60 cancer cell from a human serum sample. To release of the captured HL-60 cancer cells, the complementary nucleotide sequences of KH1C12 aptamer solution (releasing aptamer) that has a with capture aptamer was added to phosphate buffer solution (1 M, pH 7.4) containing HL-60/aptamer KH1C12 /nanomotors. Because of the high affinity of capture aptamer to complementary nucleotide sequences of aptamer KH1C12 , the HL-60 cancer cells released on the surface of aptamer KH1C12 /nanomotors into the solution. The second goal of the present work was determining the concentration of HL-60 cancer cell in the human serum samples. The electrochemical impedance spectroscopy technique (EIS) was used for the determination of HL-60 cancer cell. The concentration of separated cancer cell was determined by aptamer/gold nanoparticles-poly(3,4-ethylene dioxythiophene) modified GC electrode (GC/PEDOT-Au nano /aptamer KH1C12 ). The proposed aptasensor exhibited a good response to the concentration of HL-60 cancer cells in the range of 2.5 × 10 1 to 5 × 10 5 cells mL -1 with a low limit of detection of 250 cells mL -1 . Copyright © 2018 Elsevier B.V. All rights reserved.

Apoptotic and antiproliferative properties of 3β-hydroxy-Δ5-steroidal congeners from a partially purified column fraction of Dendronephthya gigantea against HL-60 and MCF-7 cancer cells.

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Fernando, I P Shanura; Sanjeewa, K K Asanka; Kim, Hyun-Soo; Wang, Lei; Lee, Won Woo; Jeon, You-Jin

2018-04-01

Organisms belonging to the genus Dendronephthya are among a group of marine invertebrates that produce a variety of terpenoids with biofunctional properties. Many of these terpenoids have been proven effective as anticancer drugs. Here, we report the antiproliferative effect of 3β-hydroxy-Δ5-steroidal congeners against the proliferation of HL-60 human leukemia cells and MCF-7 human breast cancer cells. The sterol-rich fraction (DGEHF2-1) inhibited the growth of HL-60 and MCF-7 cells with IC 50 values of 13.59 ± 1.40 and 29.41 ± 0.87 μg ml -1 respectively. Treatment with DGEHF2-1 caused a dose-dependent increase in apoptotic body formation, DNA damage and the sub-G 1 apoptotic cell population. Moreover, DGEHF2-1 downregulated the expression of Bcl-xL while upregulating Bax, caspase-9, and PARP cleavage in both HL-60 and MCF-7 cells. The steroid fraction was found to act via the mitochondria-mediated apoptosis pathway. Identification of the sterols was performed via gas chromatography-tandem mass spectrometry analysis. Studying the mechanism of the anticancer effect caused by these sterol derivatives could lead to the identification of other natural products with anticancer properties. Copyright © 2017 John Wiley & Sons, Ltd.

The studies of effects of 60Co γ-rays on DNA damage and repair in tumor cells

International Nuclear Information System (INIS)

Su Liaoyuan; Huang Hanxian; Cen Jiannong

1997-06-01

The effects of 60 Co γ-rays and its combination with hyperthermia on DNA strand breaks and their repair in L 5178y cells were studied using alkaline elution technique. When the cells were heated at 43 degree C for 30 min, there was obvious inhibition of repair of DNA damage caused by γ-irradiation. The hyperthermia before irradiation produced better inhibiting effect than that after irradiation. The effects of radiation on DNA strand breaks and its repair in HL-60 cells and HL-60 (VCR) cells were analyzed using technique of hydroxyapatite chromatography and the results sowed that the extent of DNA strand breaks in HL-60 cells and HL-60 (VCR) cells induced by irradiation was not markedly different, but the power of repair of strand breaks and the radioresistance of function of DNA synthesis in HL-60 (VCR) cells were higher than those in HL-60 cells. The difference was obvious. The results suggest that the heterogeneity of radiosensitivity of leukemia cells is correlated with its drug resistance. (10 refs., 3 tabs.)

Radiation response of haematopoietic cell lines of human origin

International Nuclear Information System (INIS)

Lehnert, S.; Rybka, W.B.; Suissa, S.; Giambattisto, D.

1986-01-01

Six human haematopoietic cell lines, five of leukaemic origin, including cells with myeloid, lymphoid and undifferentiated phenotype have been studied with respect to radiation response. The intrinsic radio-sensitivity of the cells varied widely, the D 0 s ranging from 0.53 to 1.39 Gy. Five of the cell lines showed some capacity to accumulate sublethal damage; in three of these, enhanced survival was demonstrated in split-dose experiments. One cell line (HL-60) was anomalous in that although little accumulation of sublethal damage was demonstrable, survival was enhanced by fractionation of the dose. Five of the six cell lines studied were of leukaemic origin. The results support the belief that, in contrast to the almost constant radiosensitivity of normal haematopoietic cell progenitors, leukaemic cell progenitors may show a wide range of radiosensitivities. (author)

BCL-x{sub L}/MCL-1 inhibition and RARγ antagonism work cooperatively in human HL60 leukemia cells

Energy Technology Data Exchange (ETDEWEB)

Perri, Mariarita; Yap, Jeremy L.; Yu, Jianshi [Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, 20 N Pine Street, Baltimore, MD 21201 (United States); Cione, Erika [Department of Pharmacy, Health and Nutritional Sciences, Ed. Polifunzionale, University of Calabria, 87036 Rende, CS (Italy); Fletcher, Steven [Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, 20 N Pine Street, Baltimore, MD 21201 (United States); Kane, Maureen A., E-mail: mkane@rx.umaryland.edu [Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, 20 N Pine Street, Baltimore, MD 21201 (United States)

2014-10-01

The acute promyelocytic leukemia (APL) subtype of acute myeloid leukemia (AML) is characterized by chromosomal translocations that result in fusion proteins, including the promyelocytic leukemia–retinoic acid receptor, alpha fusion protein (PML–RARα). All-trans retinoic acid (atRA) treatment is the standard drug treatment for APL yielding cure rates >80% by activating transcription and proteasomal degradation of retinoic acid receptor, alpha (RARα). Whereas combination therapy with As{sub 2}O{sub 3} has increased survival further, patients that experience relapse and are refractory to atRA and/or As{sub 2}O{sub 3} is a clinically significant problem. BCL-2 family proteins regulate apoptosis and over-expression of anti-apoptotic B-cell leukemia/lymphoma 2 (BCL-2) family proteins has been associated with chemotherapeutic resistance in APL including impairment of the ability of atRA to induce growth arrest and differentiation. Here we investigated the novel BH3 domain mimetic, JY-1-106, which antagonizes the anti-apoptotic BCL-2 family members B-cell lymphoma-extra large (BCL-x{sub L}) and myeloid cell leukemia-1 (MCL-1) alone and in combination with retinoids including atRA, AM580 (RARα agonist), and SR11253 (RARγ antagonist). JY-1-106 reduced cell viability in HL-60 cells alone and in combination with retinoids. The combination of JY-1-106 and SR11253 had the greatest impact on cell viability by stimulating apoptosis. These studies indicate that dual BCL-x{sub L}/MCL-1 inhibitors and retinoids could work cooperatively in leukemia treatment. - Highlights: • Novel Bcl-x{sub L}/Mcl-1 inhibitor JY-1-106 reduces HL60 cell viability. • JY-1-106 is investigated in combination with retinoic acid, AM580, and SR11253. • AM580 is an RARα agonist; SR11253 is an RARγ antagonist. • Combined use of JY-1-106/SR11253 exhibited the greatest cell viability reduction. • JY-1-106 alone or in combination with retinoids induces apoptosis.

Mechanism of the protective effects of long chain n-alkyl glucopyranosides against ultrasound-induced cytolysis of HL-60 cells

OpenAIRE

Cheng, Jason Y.; Riesz, Peter

2007-01-01

Recently it has been shown that long chain (C5 to C8) n-alkyl glucopyranosides completely inhibit ultrasound-induced cytolysis [1]. This protective effect has possible applications in HIFU (high intensity focused ultrasound) for tumor treatment, and in ultrasound assisted drug delivery and gene therapy. n-Alkyl glucopyranosides with hexyl (5mM), heptyl (3mM), octyl (2mM) n-alkyl chains protected 100% of HL-60 cells in-vitro from 1.057 MHz ultrasound induced cytolysis under a range of conditio...

Cellular uptake of {sup 99m}TcN-NOET in human leukaemic HL-60 cells is related to calcium channel activation and cell proliferation

Energy Technology Data Exchange (ETDEWEB)

Guillermet, Stephanie; Vuillez, Jean-Philippe; Caravel, Jean-Pierre; Marti-Batlle, Daniele; Fagret, Daniel [Universite de Grenoble, Radiopharmaceutiques Biocliniques, La Tronche (France); Fontaine, Eric [Universite de Grenoble, Laboratoire de Bioenergetique Fondamentale et Appliquee, Grenoble (France); Pasqualini, Roberto [Cis Bio International Schering SA, Gif-sur-Yvette (France)

2006-01-01

A major goal of nuclear oncology is the development of new radiolabelled tracers as proliferation markers. Intracellular calcium waves play a fundamental role in the course of the cell cycle. These waves occur in non-excitable tumour cells via store-operated calcium channels (SOCCs). Bis(N-ethoxy, N-ethyldithiocarbamato) nitrido technetium (V)-99m ({sup 99m}TcN-NOET) has been shown to interact with L-type voltage-operated calcium channels (VOCCs) in cultured cardiomyocytes. Considering the analogy between VOCCs and SOCCs, we sought to determine whether {sup 99m}TcN-NOET also binds to activated SOCCs in tumour cells in order to clarify the potential value of this tracer as a proliferation marker. Uptake kinetics of {sup 99m}TcN-NOET were measured in human leukaemic HL-60 cells over 60 min and the effect of several calcium channel modulators on 1-min tracer uptake was studied. The uptake kinetics of {sup 99m}TcN-NOET were compared both with the variations of cytosolic free calcium concentration measured by indo-1/AM and with the variations in the SG{sub 2}M cellular proliferation index. All calcium channel inhibitors significantly decreased the cellular uptake of {sup 99m}TcN-NOET whereas the activator thapsigargin induced a significant 10% increase. In parallel, SOCC activation by thapsigargin, as measured using the indo-1/AM probe, was inhibited by nicardipine. These results indicate that the uptake of {sup 99m}TcN-NOET is related to the activation of SOCCs. Finally, a correlation was observed between the tracer uptake and variations in the proliferation index SG{sub 2}M. The uptake of {sup 99m}TcN-NOET seems to be related to SOCC activation and to cell proliferation in HL-60 cells. These results indicate that {sup 99m}TcN-NOET might be a marker of cell proliferation. (orig.)

Anthocyanins from roselle extract arrest cell cycle G2/M phase transition via ATM/Chk pathway in p53-deficient leukemia HL-60 cells.

Science.gov (United States)

Tsai, Tsung-Chang; Huang, Hui-Pei; Chang, Kai-Ting; Wang, Chau-Jong; Chang, Yun-Ching

2017-04-01

Cell cycle regulation is an important issue in cancer therapy. Delphinidin and cyanidin are two major anthocyanins of the roselle plant (Hibiscus sabdariffa). In the present study, we investigated the effect of Hibiscus anthocyanins (HAs) on cell cycle arrest in human leukemia cell line HL-60 and the analyzed the underlying molecular mechanisms. HAs extracted from roselle calyces (purity 90%) markedly induced G2/M arrest evaluated with flow cytometry analysis. Western blot analyses revealed that HAs (0.1-0.7 mg mL -1 ) induced G2/M arrest via increasing Tyr15 phosphorylation of Cdc2, and inducing Cdk inhibitors p27 and p21. HAs also induced phosphorylation of upstream signals related to G2/M arrest such as phosphorylation of Cdc25C tyrosine phosphatase at Ser216, increasing the binding of pCdc25C with 14-3-3 protein. HAs-induced phosphorylation of Cdc25C could be activated by ATM checkpoint kinases, Chk1, and Chk2. We first time confirmed that ATM-Chk1/2-Cdc25C pathway as a critical mechanism for G2/M arrest in HAs-induced leukemia cell cycle arrest, indicating that this compound could be a promising anticancer candidate or chemopreventive agents for further investigation. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1290-1304, 2017. © 2016 Wiley Periodicals, Inc.

Measurement of tyrosine kinase activity in non-denaturing polyacrylamide gels and its induction during differentiation of human leukemia HL-60 cells

International Nuclear Information System (INIS)

Glazer, R.I.; Chapekar, M.S.; Knode, M.C.

1986-01-01

The authors have developed a PAGE assay to measure tyrosine kinase (PK-T) activity in cell extracts. HL-60 cells were extracted sequentially with 0.1% and 1.0% Triton X-100 buffer to obtain soluble and membrane-bound proteins, respectively. Extracts were separated by PAGE at 4 0 and gels were incubated in the presence of the substrate Glu:Tyr (4:1) and an assay mixture containing Mn ++ , Mg ++ and [γ 32 P]ATP. Gels were washed in TCA:PPi, dried and autoradiographed. 0.1% Triton extracts of untreated cells in log phase displayed several PK-T activities of which the major species was ∼75 kD; 1% Triton extracts exhibited only residual 75 kD PK-T. In contrast, induction of differentiation in HL-60 cells with optimal concentrations of DMSO, retinoic acid, 1.25(OH) 2 vitamin D 3 , phorbol ester, immune interferon (IFN-γ) or tumor necrosis factor (TNF) induced the appearance of a ∼170 kD PK-T in the 1% Triton extracts and a reduction of the 75 kD PK-T in the 0.1% Triton extracts. The 170 kD PK-T could also utilize Glu:Tyr (6:3:1), histone H 1 , pp60/sup src/ substrate Lys-14-Gly and vinculin as substrates but not Glu:Tyr (1:1). The 170 kD PK-T appeared within 2 days after treatment with 2500 μ/ml of IFN-γ or 100 μ/ml of TNF and its appearance coincided with the induction of the monocyte phenotype. These results suggest that the appearance of the 170 kD PK-T is related to the mature granulocyte or monocyte phenotype. Studies are in progress with radiolabeled IFN-γ to determine if the 170 kD PK-T is related to the IFN-γ receptor

Protodioscin, Isolated from the Rhizome of Dioscorea tokoro Collected in Northern Japan is the Major Antiproliferative Compound to HL-60 
Leukemic Cells.

Science.gov (United States)

Oyama, Manami; Tokiwano, Tetsuo; Kawaii, Satoru; Yoshida, Yasunori; Mizuno, Kouichi; Oh, Keimei; Yoshizawa, Yuko

2017-06-01

The rhizome of Oni-dokoro (a wild yam, Dioscorea tokoro) has extremely bitter taste and is not generally regarded edible;, however, in northern part of Japan, such as Iwate and a part of Aomori, it is used as health promoting food. To clarify the reason, we examined the biologically active compounds in the rhizome collected at Iwate and compared them from the other area in literature. The acetonitrile extract from northern part of Japan was purified by bioassay-guided separation using antiproliferative activity to human leukemia HL-60 cell, and protodioscin (PD) was isolated and identified by instrumental analyses as the major active compound. PD known as a saponin with four sugar moieties, an inhibitor for platelet aggregation, and a low density lipoprotein (LPL) lowering agent, displayed strong growth inhibitory effect to HL-60. The literature search suggested that the rhizome from other area contained dioscin and other saponins with three sugar moieties as their major component. We assume that the edible and health promoting effect of the rhizome in the particular area is partially derived from these different components. We were interested in the differences of utilization in the rhizome of wild yam Dioscorea tokoro, and examined the chemical composition in the rhizome to find protodioscin as antiproliferative compound to HL-60. In the report from other area, the rhizome exhibited dioscin as the major compound. Our study indicated that the protodioscin/dioscin composition varied regionally, although the reason is still needs to be investigated.

Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

Directory of Open Access Journals (Sweden)

Q. Sun

2012-10-01

Full Text Available The distal cytoplasmic motifs of leukemia inhibitory factor receptor α-chain (LIFRα-CT3 can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML cells by gene transfection; however, there are significant limitations in the potential clinical use of these motifs due to liposome-derived genetic modifications. To produce a potentially therapeutic LIFRα-CT3 with cell-permeable activity, we constructed a eukaryotic expression pcDNA3.0-TAT-CT3-cMyc plasmid with a signal peptide (ss inserted into the N-terminal that codes for an ss-TAT-CT3-cMyc fusion protein. The stable transfection of Chinese hamster ovary (CHO cells via this vector and subsequent selection by Geneticin resulted in cell lines that express and secrete TAT-CT3-cMyc. The spent medium of pcDNA3.0-TAT-CT3-cMyc-transfected CHO cells could be purified using a cMyc-epitope-tag agarose affinity chromatography column and could be detected via SDS-PAGE, with antibodies against cMyc-tag. The direct administration of TAT-CT3-cMyc to HL-60 cell culture media caused the enrichment of CT3-cMyc in the cytoplasm and nucleus within 30 min and led to a significant reduction of viable cells (P < 0.05 8 h after exposure. The advantages of using this mammalian expression system include the ease of generating TAT fusion proteins that are adequately transcripted and the potential for a sustained production of such proteins in vitro for future AML therapy.

Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

International Nuclear Information System (INIS)

Sun, Q.; Xiong, J.; Lu, J.; Xu, S.; Li, Y.; Zhong, X.P.; Gao, G.K.; Liu, H.Q.

2012-01-01

The distal cytoplasmic motifs of leukemia inhibitory factor receptor α-chain (LIFRα-CT3) can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML) cells by gene transfection; however, there are significant limitations in the potential clinical use of these motifs due to liposome-derived genetic modifications. To produce a potentially therapeutic LIFRα-CT3 with cell-permeable activity, we constructed a eukaryotic expression pcDNA3.0-TAT-CT3-cMyc plasmid with a signal peptide (ss) inserted into the N-terminal that codes for an ss-TAT-CT3-cMyc fusion protein. The stable transfection of Chinese hamster ovary (CHO) cells via this vector and subsequent selection by Geneticin resulted in cell lines that express and secrete TAT-CT3-cMyc. The spent medium of pcDNA3.0-TAT-CT3-cMyc-transfected CHO cells could be purified using a cMyc-epitope-tag agarose affinity chromatography column and could be detected via SDS-PAGE, with antibodies against cMyc-tag. The direct administration of TAT-CT3-cMyc to HL-60 cell culture media caused the enrichment of CT3-cMyc in the cytoplasm and nucleus within 30 min and led to a significant reduction of viable cells (P < 0.05) 8 h after exposure. The advantages of using this mammalian expression system include the ease of generating TAT fusion proteins that are adequately transcripted and the potential for a sustained production of such proteins in vitro for future AML therapy

Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

Energy Technology Data Exchange (ETDEWEB)

Sun, Q. [Department of Hyperbaric Medicine, No. 401 Hospital of PLA, Qingdao (China); Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China); Xiong, J. [Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China); Lu, J. [Office of Medical Education, Training Department, Second Military Medical University, Shanghai (China); Xu, S. [Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China); Li, Y. [State Food and Drug Administration of China,Huangdao Branch, Qingdao (China); Zhong, X.P.; Gao, G.K. [Department of Hyperbaric Medicine, No. 401 Hospital of PLA, Qingdao (China); Liu, H.Q. [2Department of Histology and Embryology, Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai (China)

2012-06-22

The distal cytoplasmic motifs of leukemia inhibitory factor receptor α-chain (LIFRα-CT3) can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML) cells by gene transfection; however, there are significant limitations in the potential clinical use of these motifs due to liposome-derived genetic modifications. To produce a potentially therapeutic LIFRα-CT3 with cell-permeable activity, we constructed a eukaryotic expression pcDNA3.0-TAT-CT3-cMyc plasmid with a signal peptide (ss) inserted into the N-terminal that codes for an ss-TAT-CT3-cMyc fusion protein. The stable transfection of Chinese hamster ovary (CHO) cells via this vector and subsequent selection by Geneticin resulted in cell lines that express and secrete TAT-CT3-cMyc. The spent medium of pcDNA3.0-TAT-CT3-cMyc-transfected CHO cells could be purified using a cMyc-epitope-tag agarose affinity chromatography column and could be detected via SDS-PAGE, with antibodies against cMyc-tag. The direct administration of TAT-CT3-cMyc to HL-60 cell culture media caused the enrichment of CT3-cMyc in the cytoplasm and nucleus within 30 min and led to a significant reduction of viable cells (P < 0.05) 8 h after exposure. The advantages of using this mammalian expression system include the ease of generating TAT fusion proteins that are adequately transcripted and the potential for a sustained production of such proteins in vitro for future AML therapy.

The Putative Role of the Non-Gastric H+/K+-ATPase ATP12A (ATP1AL1 as Anti-Apoptotic Ion Transporter: Effect of the H+/K+ ATPase Inhibitor SCH28080 on Butyrate-Stimulated Myelomonocytic HL-60 Cells

Directory of Open Access Journals (Sweden)

Martin Jakab

2014-10-01

Full Text Available Background/Aims: The ATP12A gene codes for a non-gastric H+/K+ ATPase, which is expressed in a wide variety of tissues. The aim of this study was to test for the molecular and functional expression of the non-gastric H+/K+ ATPase ATP12A/ATP1AL1 in unstimulated and butyrate-stimulated (1 and 10 mM human myelomonocytic HL-60 cells, to unravel its potential role as putative apoptosis-counteracting ion transporter as well as to test for the effect of the H+/K+ ATPase inhibitor SCH28080 in apoptosis. Methods: Real-time reverse-transcription PCR (qRT-PCR was used for amplification and cloning of ATP12A transcripts and to assess transcriptional regulation. BCECF microfluorimetry was used to assess changes of intracellular pH (pHi after acute intracellular acid load (NH4Cl prepulsing. Mean cell volumes (MCV and MCV-recovery after osmotic cell shrinkage (Regulatory Volume Increase, RVI were assessed by Coulter counting. Flow-cytometry was used to measure MCV (Coulter principle, to assess apoptosis (phosphatidylserine exposure to the outer leaflet of the cell membrane, caspase activity, 7AAD staining and differentiation (CD86 expression. Results: We found by RT-PCR, intracellular pH measurements, MCV measurements and flow cytometry that ATP12A is expressed in human myelomonocytic HL-60 cells. Treatment of HL-60 cells with 1 mM butyrate leads to monocyte-directed differentiation whereas higher concentrations (10 mM induce apoptosis as assessed by flow-cytometric determination of CD86 expression, caspase activity, phosphatidylserine exposure on the outer leaflet of the cell membrane and MCV measurements. Transcriptional up-regulation of ATP12A and CD86 is evident in 1 mM butyrate-treated HL-60 cells. The H+/K+ ATPase inhibitor SCH28080 (100 µM diminishes K+-dependent pHi recovery after intracellular acid load and blocks RVI after osmotic cell shrinkage. After seeding, HL-60 cells increase their MCV within the first 24 h in culture, and subsequently

Evidence for Transfer of Membranes from Mesenchymal Stem Cells to HL-1 Cardiac Cells.

Science.gov (United States)

Boomsma, Robert A; Geenen, David L

2014-01-01

This study examined the interaction of mouse bone marrow mesenchymal stem cells (MSC) with cardiac HL-1 cells during coculture by fluorescent dye labeling and then flow cytometry. MSC were layered onto confluent HL-1 cell cultures in a 1 : 4 ratio. MSC gained gap junction permeant calcein from HL-1 cells after 4 hours which was partially reduced by oleamide. After 20 hours, 99% MSC gained calcein, unaffected by oleamide. Double-labeling HL-1 cells with calcein and the membrane dye DiO resulted in transfer of both calcein and DiO to MSC. When HL-1 cells were labeled with calcein and MSC with DiO, MSC gained calcein while HL-1 cells gained DiO. Very little fusion was observed since more than 90% Sca-1 positive MSC gained DiO from HL-1 cells while less than 9% gained gap junction impermeant CMFDA after 20 hours with no Sca-1 transfer to HL-1 cells. Time dependent transfer of membrane DiD was observed from HL-1 cells to MSC (100%) and vice versa (50%) after 20 hours with more limited transfer of CMFDA. These results demonstrate that MSC and HL-1 cells exchange membrane components which may account for some of the beneficial effect of MSC in the heart after myocardial infarction.

JS-K, a nitric oxide prodrug, induces cytochrome c release and caspase activation in HL-60 myeloid leukemia cells.

Science.gov (United States)

Udupi, Vidya; Yu, Margaret; Malaviya, Swati; Saavedra, Joseph E; Shami, Paul J

2006-10-01

Nitric oxide (NO) induces differentiation and apoptosis in acute myelogenous leukemia (AML) cells. The NO prodrug O2-(2,4-dinitrophenyl)1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate, or JS-K, has potent antileukemic activity. JS-K induces apoptosis in HL-60 cells by a caspase-dependent mechanism. The purpose of this study was to determine the pathway through which JS-K induces apoptosis. We show that JS-K alters mitochondrial membrane potential (DeltaPsim) and induces cytochrome c release from mitochondria into the cytoplasm. Treatment with JS-K resulted in activation of Caspase (Casp) 9, Casp 3 and Casp 8. JS-K constitutes a promising lead for a new class of anti-leukemic agents.

Vitamin K2 and cotylenin A synergistically induce monocytic differentiation and growth arrest along with the suppression of c-MYC expression and induction of cyclin G2 expression in human leukemia HL-60 cells.

Science.gov (United States)

Maniwa, Yasuhisa; Kasukabe, Takashi; Kumakura, Shunichi

2015-08-01

Although all-trans retinoic acid (ATRA) is a standard and effective drug used for differentiation therapy in acute promyelocytic leukemia, ATRA-resistant leukemia cells ultimately emerge during this treatment. Therefore, the development of new drugs or effective combination therapy is urgently needed. We demonstrate that the combined treatment of vitamin K2 and cotylenin A synergistically induced monocytic differentiation in HL-60 cells. This combined treatment also synergistically induced NBT-reducing activity and non-specific esterase-positive cells as well as morphological changes to monocyte/macrophage-like cells. Vitamin K2 and cotylenin A cooperatively inhibited the proliferation of HL-60 cells in short-term and long-term cultures. This treatment also induced growth arrest at the G1 phase. Although 5 µg/ml cotylenin A or 5 µM vitamin K2 alone reduced c-MYC gene expression in HL-60 cells to approximately 45% or 80% that of control cells, respectively, the combined treatment almost completely suppressed c-MYC gene expression. We also demonstrated that the combined treatment of vitamin K2 and cotylenin A synergistically induced the expression of cyclin G2, which had a positive effect on the promotion and maintenance of cell cycle arrest. These results suggest that the combination of vitamin K2 and cotylenin A has therapeutic value in the treatment of acute myeloid leukemia.

Synthesis of minoxidil conjugates and their evaluation as HL-60 differentiation agents.

Science.gov (United States)

Stoica, Sonia; Magoulas, George E; Antoniou, Antonia I; Suleiman, Sherif; Cassar, Analisse; Gatt, Lucienne; Papaioannou, Dionissios; Athanassopoulos, Constantinos M; Schembri-Wismayer, Pierre

2016-02-15

Activation of minoxidil (MNX) with N,N'-carbonyldiimidazole and coupling with natural polyamines (PAs) and commercially available aliphatic or aromatic amines provided a series of new conjugates which were evaluated for their ability to induce differentiation to HL-60 acute myeloid leukemia cancer cells, using a modified NBTZ reduction test. Although neither MNX nor 4,4'-methylenedianiline (MDA) or 2,7-diaminofluorene (DAF), alone or in combination, had any effect, the MNX-spermine (SPM) conjugate (11) and the conjugates 7 and 8 of MNX with MDA and DAF exhibited a differentiation-inducing effect at a concentration of 10 μM without being toxic on proliferating human peripheral blood mononuclear cells. Copyright © 2016 Elsevier Ltd. All rights reserved.

Induction of apoptosis by hydrolyzable tannins from Eugenia jambos L. on human leukemia cells.

Science.gov (United States)

Yang, L L; Lee, C Y; Yen, K Y

2000-08-31

Eugenia jambos L. (Myrtaceae) is an antipyretic and anti-inflammatory herb of Asian folk medicine. A 70% acetone extract exerted the strongest cytotoxic effects on human leukemia cells (HL-60) from a preliminary screening of 15 plants. The cytotoxic principles were separated by bio-assay-guided fractionation to HL-60 cells; two hydrolyzable tannins (1-O-galloyl castalagin and casuarinin) were isolated from the 70% acetone extract. All significantly inhibited human promyelocytic leukemia cell line HL-60 and showed less cytotoxicity to human adenocarcinoma cell line SK-HEP-1 and normal cell lines of human lymphocytes and Chang liver cells. Thus, these compounds were exhibited the dose-dependent manner in HL-60 cells and the IC(50) were 10.8 and 12.5 microM, respectively. Flow cytometric analysis demonstrated the presence of apoptotic cells with low DNA content, a decrease of cell population at G(2)/M phase, and a concomitant increase of cell population at G(1) phase. The apoptosis induced by these two compounds was also demonstrated by DNA fragmentation assay and microscopic observation. These results suggest that the cytotoxic mechanism of both antitumor principle constituents might be the induction of apoptosis in HL-60 cells.

Apoptosis and pro-death autophagy induced by a spirostanol saponin isolated from Rohdea chinensis (Baker) N. Tanaka (synonym Tupistra chinensis Baker) on HL-60 cells.

Science.gov (United States)

Yi, Xiaomin; Xiang, Limin; Huang, Yuying; Wang, Yihai; He, Xiangjiu

2018-03-15

Our previous study has revealed that the spirostanol saponins isolated from the rhizomes of Rohdea chinensis (Baker) N. Tanaka (synonym Tupistra chinensis Baker) (Convallariaceae) (a reputed folk medicine) exhibited potent antiproliferative activity. However, the underlying mechanism of purified saponins remains unclear. More studies are necessary to assess the apoptosis and autophagy activities of the saponins from R. chinensis and clarify their antiproliferative mechanisms. The present study certificated the potential antiproliferative activity and mechanism of 5β-spirost-25(27)-en-1β,3β-diol-1-O-α-L-rhamnopyranosyl-(1→2)- β-D-xylopyranosyl-3-O-α-L-rhamnopyranoside (SPD), a spirostanol saponin from R. chinensis, against human acute promyelocytic leukemia cells (HL-60). The antiproliferative activity of SPD in vitro was evaluated by MTT assay compared with cis-dichlorodiammineplatinum (II). The autophagic activity was assessed using MDC staining and western blot, cell apoptosis inspection was detected by Annexin V-FITC/PI double staining and the mitochondrial membrane potential was detected by JC-1 fluorescence dye combined with flow cytometry. The potential mechanisms for protein levels of apoptosis and autophagy were evaluated by western blot. Treatment of HL-60 cells with SPD resulted in growth inhibition (IC 50 value of 2.0 ± 0.2 µM, after 48 h treatment) and induction of apoptosis and autophagy. Results from Annexin V-FITC/PI double-staining assay and mitochondrial membrane potential detection showed that apoptosis was happened after SPD treatment. The regulation of caspase-3, Bax, Bcl-2, PARP following SPD treatment contributed to the induction of mitochondria-dependent apoptosis. Meanwhile, SPD induced autophagy related with Akt/mTOR/p70S6K signaling and activated of AMPK signaling pathway. Furthermore, blocking autophagy with bafilomycin A1 reduced the cytotoxicity of SPD in HL-60 cells. The antiproliferative, apoptosis and pro

Vildagliptin and its metabolite M20.7 induce the expression of S100A8 and S100A9 in human hepatoma HepG2 and leukemia HL-60 cells.

Science.gov (United States)

Asakura, Mitsutoshi; Karaki, Fumika; Fujii, Hideaki; Atsuda, Koichiro; Itoh, Tomoo; Fujiwara, Ryoichi

2016-10-19

Vildagliptin is a potent, orally active inhibitor of dipeptidyl peptidase-4 (DPP-4) for the treatment of type 2 diabetes mellitus. It has been reported that vildagliptin can cause hepatic dysfunction in patients. However, the molecular-mechanism of vildagliptin-induced liver dysfunction has not been elucidated. In this study, we employed an expression microarray to determine hepatic genes that were highly regulated by vildagliptin in mice. We found that pro-inflammatory S100 calcium-binding protein (S100) a8 and S100a9 were induced more than 5-fold by vildagliptin in the mouse liver. We further examined the effects of vildagliptin and its major metabolite M20.7 on the mRNA expression levels of S100A8 and S100A9 in human hepatoma HepG2 and leukemia HL-60 cells. In HepG2 cells, vildagliptin, M20.7, and sitagliptin - another DPP-4 inhibitor - induced S100A9 mRNA. In HL-60 cells, in contrast, S100A8 and S100A9 mRNAs were significantly induced by vildagliptin and M20.7, but not by sitagliptin. The release of S100A8/A9 complex in the cell culturing medium was observed in the HL-60 cells treated with vildagliptin and M20.7. Therefore, the parental vildagliptin- and M20.7-induced release of S100A8/A9 complex from immune cells, such as neutrophils, might be a contributing factor of vildagliptin-associated liver dysfunction in humans.

Cytotoxic and DNA-damaging effects of methyl tert-butyl ether and its metabolites on HL-60 cells in vitro

Energy Technology Data Exchange (ETDEWEB)

Tang, G.H. [Xian Medical Univ. (China); Shen, Y.; Shen, H.M. [National Univ. of Singapore (Singapore)] [and others

1996-12-31

Methyl tert-butyl ether (MTBE) is a widely used oxygenate in unleaded gasoline; however, few studies have been conducted on the toxicity of this compound. This study evaluates the cytotoxic and DNA-damaging effects of MTBE and its metabolites in a human haemopoietic cell line, HL-60. The metabolites of MTBE studied include tertiary butyl alcohol (TBA), {alpha}-hydroxyisobutyric acid (HIBA), and formaldehyde. Comet assay is used to assess DNA damage, and the cytotoxicity is investigated by lactate dehydrogenease (LDH) release. The results show no significant cytotoxic effects of MTBE, TBA, and HIBA over a concentration ranging from 1 to 30 mM. Formaldehyde, in contrast, causes a substantial LDH release at a concentration of 5 {mu}M. Hydrogen peroxide, a known oxidative agent, at concentrations ranging from 10 to 100 {mu}M, produces a significant dose-related increase in DNA damage, whereas a much higher concentration of MTBE (1 to 30 mM) is required to produce a similar observation. The genotoxic effects of TBA and HIBA appear to be identical to that of MTBE. Conversely, DNA damage is observed for formaldehyde at a relatively low concentration range (5 to 100 {mu}M). These findings suggest that MTBE and its metabolites, except formaldehyde, have relatively low cytotoxic and genotoxic effects. 16 refs., 4 figs.

An antisense oligodeoxynucleotide targeted against the type IIβ regulatory subunit mRNA of protein kinase inhibits cAMP-induced differentiation in HL-60 leukemia cells without affecting phorbol ester effects

International Nuclear Information System (INIS)

Tortora, G.; Clair, T.; Cho-Chung, Y.S.

1990-01-01

The type II β regulatory subunit of cAMP-dependent protein kinase (RII β ) has been hypothesized to play an important role in the growth inhibition and differentiation induced by site-selective cAMP analogs in human cancer cells, but direct proof of this function has been lacking. To address this tissue, HL-60 human promyelocytic leukemia cells were exposed to RII β antisense synthetic oligodeoxynucleotide, and the effects on cAMP-induced growth regulation were examined. Exposure of these cells to RII β antisense oligodeoxynucleotide resulted in a decrease in cAMP analog-induced growth inhibition and differentiation without apparent effect on differentiation induced by phorbol esters. This loss in cAMP growth regulatory function correlated with a decrease in basal and induced levels of RII β protein. Exposure to RII β sense, RI α and RII α antisense, or irrelevant oligodeoxynucleotides had no such effect. These results show that the RII β regulatory subunit of protein kinase plays a critical role in the cAMP-induced growth regulation of HL-60 leukemia cells

Genes encoded within 8q24 on the amplicon of a large extrachromosomal element are selectively repressed during the terminal differentiation of HL-60 cells

OpenAIRE

Hirano, Tetsuo; Ike, Fumio; Murata, Takehide; Obata, Yuichi; Utiyama, Hiroyasu; Yokoyama, Kazunari K.

2008-01-01

Human acute myeloblastic leukemia HL-60 cells become resistant to differentiation during long­term cultivation. After 150 passages, double minute chromosomes (dmins) found in early­-passaged cells are replaced by large extrachromosomal elements (LEEs). In a DNA library derived from a purified fraction of LEEs, 12.6% (23/183) of clones were assigned to 8q24 and 9.2% (17/183) were assigned to 14q11 in the human genome. Fluorescence in situ hybridization (FISH) revealed a small aberrant chromos...

Enhancement of the incorporation of 5-fluorodeoxyuridylate into DNA of HL-60 cells by metabolic modulations

International Nuclear Information System (INIS)

Tanaka, M.; Kimura, K.; Yoshida, S.

1983-01-01

The exposure of HL-60 human promyelocytic leukemia cells to 0.5 microM 5-fluoro-2'-[ 3 H]deoxyuridine (FdUrd) for 16 hr resulted in the incorporation of 5.14 +/- 0.31 (S.D.) X 10(-7) mol FdUrd into DNA per mol of DNA nucleotide, which corresponds to 0.146 +/- 0.082 pmol FdUrd per 10(7) cells. Pretreatment with 50 microM deoxythymidine for 24 hr led to a 2.7-fold increase in the incorporation of this analogue into newly synthesized DNA during the ensuing 16-hr exposure to 0.5 microM [ 3 H]FdUrd. Pretreatment with 0.5 microM methotrexate for 3 hr also increased the [ 3 H]FdUrd incorporation into newly synthesized DNA approximately 5-fold. The coexistence of deoxythymidine or methotrexate with [ 3 H]FdUrd, however, led to decreased incorporation of FdUrd into DNA. More than 50% of the radioactivity in DNA separated by Cs2SO4 equilibrium density gradient centrifugation was proven to be fluorodeoxyuridylate by means of its binding to Lactobacillus casei deoxythymidine monophosphate synthetase

MicroRNA expression changes during human leukemic HL-60 cell differentiation induced by 4-hydroxynonenal, a product of lipid peroxidation.

Science.gov (United States)

Pizzimenti, Stefania; Ferracin, Manuela; Sabbioni, Silvia; Toaldo, Cristina; Pettazzoni, Piergiorgio; Dianzani, Mario Umberto; Negrini, Massimo; Barrera, Giuseppina

2009-01-15

4-Hydroxynonenal (HNE) is one of several lipid oxidation products that may have an impact on human pathophysiology. It is an important second messenger involved in the regulation of various cellular processes and exhibits antiproliferative and differentiative properties in various tumor cell lines. The mechanisms by which HNE affects cell growth and differentiation are only partially clarified. Because microRNAs (miRNAs) have the ability to regulate several cellular processes, we hypothesized that HNE, in addition to other mechanisms, could affect miRNA expression. Here, we present the results of a genome-wide miRNA expression profiling of HNE-treated HL-60 leukemic cells. Among 470 human miRNAs, 10 were found to be differentially expressed between control and HNE-treated cells (at p<0.05). Six miRNAs were down-regulated (miR-181a*, miR-199b, miR-202, miR-378, miR-454-3p, miR-575) and 4 were up-regulated (miR-125a, miR-339, miR-663, miR-660). Three of these regulated miRNAs (miR-202, miR-339, miR-378) were further assayed and validated by quantitative real-time RT-PCR. Moreover, consistent with the down-regulation of miR-378, HNE also induced the expression of the SUFU protein, a tumor suppressor recently identified as a target of miR-378. The finding that HNE could regulate the expression of miRNAs and their targets opens new perspectives on the understanding of HNE-controlled pathways. A functional analysis of 191 putative gene targets of miRNAs modulated by HNE is discussed.

Cytotoxic activity of vitamins K1, K2 and K3 against human oral tumor cell lines.

Science.gov (United States)

Okayasu, H; Ishihara, M; Satoh, K; Sakagami, H

2001-01-01

Vitamin K1, K2 and K3 were compared for their cytotoxic activity, radical generation and O2- scavenging activity. Among these compounds, vitamin K3 showed the highest cytotoxic activity against human oral tumor cell lines (HSC-2, HSG), human promyelocytic leukemic cell line (HL-60) and human gingival fibroblast (HGF). Vitamin K3 induced internucleosomal DNA fragmentation in HL-60 cells, but not in HSC-2 or HSG cells. The cytotoxic activity of vitamins K2 and K1 was one and two orders lower, respectively, than K3. Vitamin K2, but not vitamin K3, showed tumor-specific cytotoxic action. ESR spectroscopy showed that only vitamin K3 produced radical(s) under alkaline condition and most potently enhanced the radical intensity of sodium ascorbate and scavenged O2- (generated by hypoxanthine-xanthine oxidase reaction system); vitamin K2 was much less active whereas vitamin K1 was inactive. These data suggest that the cytotoxic activity of vitamin K3 is generated by radical-mediated oxidation mechanism and that this vitamin has two opposing actions (that is, antioxidant and prooxidant), depending on the experimental conditions.

Expression of human kinase suppressor of Ras 2 (hKSR-2) gene in HL60 leukemia cells is directly upregulated by 1,25-dihydroxyvitamin D3 and is required for optimal cell differentiation

International Nuclear Information System (INIS)

Wang Xuening; Wang, T.-T.; White, John H.; Studzinski, George P.

2007-01-01

Induction of terminal differentiation of neoplastic cells offers potential for a novel approach to cancer therapy. One of the agents being investigated for this purpose in preclinical studies is 1,25-dihydroxyvitamin D 3 (1,25D), which can convert myeloid leukemia cells into normal monocyte-like cells, but the molecular mechanisms underlying this process are not fully understood. Here, we report that 1,25D upregulates the expression of hKSR-2, a new member of a small family of proteins that exhibit evolutionarily conserved function of potentiating ras signaling. The upregulation of hKSR-2 is direct, as it occurs in the presence of cycloheximide, and occurs primarily at the transcriptional level, via activation of vitamin D receptor, which acts as a ligand-activated transcription factor. Two VDRE-type motifs identified in the hKSR-2 gene bind VDR-RXR alpha heterodimers present in nuclear extracts of 1,25D-treated HL60 cells, and chromatin immunoprecipitation assays show that these VDRE motifs bind VDR in 1,25D-dependent manner in intact cells, coincident with the recruitment of RNA polymerase II to these motifs. Treatment of the cells with siRNA to hKSR-2 reduced the proportion of the most highly differentiated cells in 1,25D-treated cultures. These results demonstrate that hKSR-2 is a direct target of 1,25D in HL60 cells, and is required for optimal monocytic differentiation

Antimicrobial Activity of Hippurate Nano composite and Its Cytotoxicity Effect in Combination with Cytarabine against HL-60

International Nuclear Information System (INIS)

Al Ali, S.H.H.; Al-Qubaisi, M.; Ismail, M.; El Zowalaty, M.; Hussein, M.Z.; Ismail, M.

2013-01-01

Hippuric acid (HA) was intercalated into a zinc-layered hydroxide (ZLH) by direct reaction of an aqueous suspension of zinc oxide with an aqueous solution of hippuric acid to obtain hippurate nano composite (HAN). Various concentrations of hippuric acid (0.05, 0.2, and 0.4 molar) were used for the synthesis of the nano composite. The as-synthesized HAN using 0.2 molar was found to give a well-ordered layered nano composite material with an increase in the basal spacing to 21.3 Å which indicated the insertion of hippurate organic moiety into the ZLH interlayers. The cytotoxicity of HAN in combination with cytarabine against human promyelocytic leukemia cells (HL-60) was tested using MTT cell viability assay and trypan blue dye exclusion assay. The combination of cytarabine with HAN showed higher tumor suppression efficiency as compared to that of cytarabine alone. The IC 50 values of HAN/cytarabine combination and cytarabine alone were μg/mL and μg/mL, respectively. DNA fragmentation was also studied, and the exposure of HL-60 cells to cytarabine produced % DNA fragmentation compared to % when cells were exposed to combination of cytarabine with HAN. The antimicrobial activity of hippuric acid and HAN nano composite was carried out against Gram-positive bacteria, Gram-negative bacteria, and yeasts. It was found that Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus were more sensitive to HAN compared to Bacillus subtilis and Salmonella choleraesuis

Berberine and a Berberis lycium extract inactivate Cdc25A and induce α-tubulin acetylation that correlate with HL-60 cell cycle inhibition and apoptosis

International Nuclear Information System (INIS)

Khan, Musa; Giessrigl, Benedikt; Vonach, Caroline; Madlener, Sibylle; Prinz, Sonja; Herbaceck, Irene; Hoelzl, Christine; Bauer, Sabine; Viola, Katharina; Mikulits, Wolfgang; Quereshi, Rizwana Aleem; Knasmueller, Siegfried; Grusch, Michael; Kopp, Brigitte; Krupitza, Georg

2010-01-01

Berberis lycium Royle (Berberidacea) from Pakistan and its alkaloids berberine and palmatine have been reported to possess beneficial pharmacological properties. In the present study, the anti-neoplastic activities of different B. lycium root extracts and the major constituting alkaloids, berberine and palmatine were investigated in p53-deficient HL-60 cells. The strongest growth inhibitory and pro-apoptotic effects were found in the n-butanol (BuOH) extract followed by the ethyl acetate (EtOAc)-, and the water (H 2 O) extract. The chemical composition of the BuOH extract was analyzed by TLC and quantified by HPLC. 11.1 μg BuOH extract (that was gained from 1 mg dried root) contained 2.0 μg berberine and 0.3 μg/ml palmatine. 1.2 μg/ml berberine inhibited cell proliferation significantly, while 0.5 μg/ml palmatine had no effect. Berberine and the BuOH extract caused accumulation of HL-60 cells in S-phase. This was preceded by a strong activation of Chk2, phosphorylation and degradation of Cdc25A, and the subsequent inactivation of Cdc2 (CDK1). Furthermore, berberine and the extract inhibited the expression of the proto-oncogene cyclin D1. Berberine and the BuOH extract induced the acetylation of α-tubulin and this correlated with the induction of apoptosis. The data demonstrate that berberine is a potent anti-neoplastic compound that acts via anti-proliferative and pro-apoptotic mechanisms independent of genotoxicity.

Mitochondrial DNA damage and oxidative damage in HL-60 cells exposed to 900 MHz radiofrequency fields

Energy Technology Data Exchange (ETDEWEB)

Sun, Yulong; Zong, Lin; Gao, Zhen [School of Public Health, Soochow University, Suzhou, Jiangsu Province (China); Zhu, Shunxing [Laboratory Animal Center, Nantong University, Nantong, Jiangsu Province (China); Tong, Jian [School of Public Health, Soochow University, Suzhou, Jiangsu Province (China); Cao, Yi, E-mail: yicao@suda.edu.cn [School of Public Health, Soochow University, Suzhou, Jiangsu Province (China)

2017-03-15

Highlights: • Increased reactive oxygen species. • Decreased mitochondrial transcription Factor A and polymerase gamma. • Decreased mitochondrial transcripts (ND1 and 16S) and mtDNA copy number. • Increased 8-hydroxy-2′deoxyguanosine. • Decreased adenosine triphosphate. - Abstract: HL-60 cells, derived from human promyelocytic leukemia, were exposed to continuous wave 900 MHz radiofrequency fields (RF) at 120 μW/cm{sup 2} power intensity for 4 h/day for 5 consecutive days to examine whether such exposure is capable damaging the mitochondrial DNA (mtDNA) mediated through the production of reactive oxygen species (ROS). In addition, the effect of RF exposure was examined on 8-hydroxy-2′-dexoyguanosine (8-OHdG) which is a biomarker for oxidative damage and on the mitochondrial synthesis of adenosine triphosphate (ATP) which is the energy required for cellular functions. The results indicated a significant increase in ROS and significant decreases in mitochondrial transcription factor A, mtDNA polymerase gamma, mtDNA transcripts and mtDNA copy number in RF-exposed cells compared with those in sham-exposed control cells. In addition, there was a significant increase in 8-OHdG and a significant decrease in ATP in RF-exposed cells. The response in positive control cells exposed to gamma radiation (GR, which is also known to induce ROS) was similar to those in RF-exposed cells. Thus, the overall data indicated that RF exposure was capable of inducing mtDNA damage mediated through ROS pathway which also induced oxidative damage. Prior-treatment of RF- and GR-exposed the cells with melatonin, a well-known free radical scavenger, reversed the effects observed in RF-exposed cells.

Expression of MIF and CD74 in leukemic cell lines: correlation to DR expression destiny.

Science.gov (United States)

Georgouli, Mirella; Papadimitriou, Lina; Glymenaki, Maria; Patsaki, Valia; Athanassakis, Irene

2016-06-01

Invariant chain (Ii) or CD74 is a non-polymorphic glycoprotein, which apart from its role as a chaperone dedicated to MHCII molecules, is known to be a high-affinity receptor for macrophage migration inhibitory factor (MIF). The present study aimed to define the roles of CD74 and MIF in the immune surveillance escape process. Towards this direction, the cell lines HL-60, Raji, K562 and primary pre-B leukemic cells were examined for expression and secretion of MIF. Flow cytometry analysis detected high levels of MIF and intracellular/membrane CD74 expression in all leukemic cells tested, while MIF secretion was shown to be inversely proportional to intracellular HLA-DR (DR) expression. In the MHCII-negative cells, IFN-γ increased MIF expression and induced its secretion in HL-60 and K562 cells, respectively. In K562 cells, CD74 (Iip33Iip35) was shown to co-precipitate with HLA-DOβ (DOβ), inhibiting thus MIF or DR binding. Induced expression of DOα in K562 (DOα-DOβ+) cells in different transfection combinations decreased MIF expression and secretion, while increasing surface DR expression. Thus, MIF could indeed be part of the antigen presentation process.

Global Proteome Analysis of the NCI-60 Cell Line Panel

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Amin Moghaddas Gholami

2013-08-01

Full Text Available The NCI-60 cell line collection is a very widely used panel for the study of cellular mechanisms of cancer in general and in vitro drug action in particular. It is a model system for the tissue types and genetic diversity of human cancers and has been extensively molecularly characterized. Here, we present a quantitative proteome and kinome profile of the NCI-60 panel covering, in total, 10,350 proteins (including 375 protein kinases and including a core cancer proteome of 5,578 proteins that were consistently quantified across all tissue types. Bioinformatic analysis revealed strong cell line clusters according to tissue type and disclosed hundreds of differentially regulated proteins representing potential biomarkers for numerous tumor properties. Integration with public transcriptome data showed considerable similarity between mRNA and protein expression. Modeling of proteome and drug-response profiles for 108 FDA-approved drugs identified known and potential protein markers for drug sensitivity and resistance. To enable community access to this unique resource, we incorporated it into a public database for comparative and integrative analysis (http://wzw.tum.de/proteomics/nci60.

Prostaglandin E2 blocks menadione-induced apoptosis through the Ras/Raf/Erk signaling pathway in promonocytic leukemia cell lines.

Science.gov (United States)

Yeo, Hyun-Seok; Shehzad, Adeeb; Lee, Young Sup

2012-04-01

Altered oxidative stress has long been observed in cancer cells, and this biochemical property of cancer cells represents a specific vulnerability that can be exploited for therapeutic benefit. The major role of an elevated oxidative stress for the efficacy of molecular targeted drugs is under investigation. Menadione is considered an attractive model for the study of oxidative stress, which can induce apoptosis in human leukemia HL-60 cell lines. Prostaglandin E(2) (PGE(2)) via its receptors not only promotes cell survival but also reverses apoptosis and promotes cancer progression. Here, we present evidence for the biological role of PGE(2) as a protective agent of oxidative stress-induced apoptosis in monocytic cells. Pretreatment of HL-60 cells with PGE(2) markedly ameliorated the menadione-induced apoptosis and inhibited the degradation of PARP and lamin B. The EP(2) receptor antagonist AH6809 abrogated the inhibitory effect of PGE(2), suggesting the role of the EP(2)/cAMP system. The PKA inhibitor H89 also reversed apoptosis and decreased the PKA activity that was elevated 10-fold by PGE(2). The treatment of HL-60 cells with NAC or zinc chloride showed a similar protective effect as with PGE(2) on menadione-treated cells. Furthermore, PGE(2) activated the Ras/Raf/MEK pathway, which in turn initiated ERK activation, and ultimately protected menadione-induced apoptosis. These results imply that PGE(2) via cell survival pathways may protect oxidative stress-induced apoptosis in monocytic cells. This study warrants further pre-clinical investigation as well as application towards leukemia clinics.

Hematopoietic Cancer Cell Lines Can Support Replication of Sabin Poliovirus Type 1

Science.gov (United States)

van Eikenhorst, Gerco; de Gruijl, Tanja D.; van der Pol, Leo A.; Bakker, Wilfried A. M.

2015-01-01

Viral vaccines can be produced in adherent or in suspension cells. The objective of this work was to screen human suspension cell lines for the capacity to support viral replication. As the first step, it was investigated whether poliovirus can replicate in such cell lines. Sabin poliovirus type 1 was serially passaged on five human cell lines, HL60, K562, KG1, THP-1, and U937. Sabin type 1 was capable of efficiently replicating in three cell lines (K562, KG1, and U937), yielding high viral titers after replication. Expression of CD155, the poliovirus receptor, did not explain susceptibility to replication, since all cell lines expressed CD155. Furthermore, we showed that passaged virus replicated more efficiently than parental virus in KG1 cells, yielding higher virus titers in the supernatant early after infection. Infection of cell lines at an MOI of 0.01 resulted in high viral titers in the supernatant at day 4. Infection of K562 with passaged Sabin type 1 in a bioreactor system yielded high viral titers in the supernatant. Altogether, these data suggest that K562, KG1, and U937 cell lines are useful for propagation of poliovirus. PMID:25815312

Importance of glutamine metabolism in leukemia cells by energy production through TCA cycle and by redox homeostasis.

Science.gov (United States)

Goto, Mineaki; Miwa, Hiroshi; Shikami, Masato; Tsunekawa-Imai, Norikazu; Suganuma, Kazuto; Mizuno, Shohei; Takahashi, Miyuki; Mizutani, Motonori; Hanamura, Ichiro; Nitta, Masakazu

2014-07-01

Some cancer cells depend on glutamine despite of pronounced glycolysis. We examined the glutamine metabolism in leukemia cells, and found that HL-60 cells most depended on glutamine in the 4 acute myelogenous leukemia (AML) cell lines examined: growth of HL-60 cells was most suppressed by glutamine deprivation and by inhibition of glutaminolysis, which was rescued by tricarboxylic acid (TCA) cycle intermediate, oxaloacetic acid. Glutamine is also involved in antioxidant defense function by increasing glutathione. Glutamine deprivation suppressed the glutathione content and elevated reactive oxygen species most evidently in HL-60 cells. Glutamine metabolism might be a therapeutic target in some leukemia.

Effect of С(60 fullerene on metabolic and proliferative activity of PKE cell line

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I. V. Belochkina

2014-04-01

Full Text Available The effect of С60 fullerene aqueous colloid solution (C60FAS on activity of redox and proliferative processes in PKE (transplantable cell line of pig kidney embryo cells has been studied. In particular, it was established that the presence of С60 fullerene (127 μМ in culturing medium of PKE cells during 48 h did not change their ability to reduce non-toxic АlamarBlue redox indicator and proliferative acti­vity.

Stimulation of granulocytic cell iodination by pine cone antitumor substances

International Nuclear Information System (INIS)

Unten, S.; Sakagami, H.; Konno, K.

1989-01-01

Antitumor substances (Fractions VI and VII) prepared from the NaOH extract of pine cone significantly stimulated the iodination (incorporation of radioactive iodine into an acid-insoluble fraction) of human peripheral blood adherent mononuclear cells, polymorphonuclear cells (PMN), and human promyelocytic leukemic HL-60 cells. In contrast, these fractions did not significantly increase the iodination of nonadherent mononuclear cells, red blood cells, other human leukemic cell lines (U-937, THP-1, K-562), human diploid fibroblast (UT20Lu), or mouse cell lines (L-929, J774.1). Iodination of HL-60 cells, which were induced to differentiate by treatment with either retinoic acid or tumor necrosis factor, were stimulated less than untreated cells. The stimulation of iodination of both PMN and HL-60 cells required the continuous presence of these fractions and was almost completely abolished by the presence of myeloperoxidase inhibitors. The stimulation activity of these fractions was generally higher than that of various other immunopotentiators. Possible mechanisms of extract stimulation of myeloperoxidase-containing cell iodination are discussed

Expression and function of β-adrenergic receptors in human hematopoietic cell lines

International Nuclear Information System (INIS)

Maeki, T.; Andersson, L.C.; Kontula, K.K.

1992-01-01

We investigated the expression and functional characteristics of β-adrenoceptors in a panel of 10 phenotypically different human hematopoietic cell lines. A binding assay with [ 125 I]iodocyanopindolol as the ligand revealed that cell lines of myelomonocytic or histiocytic derivation (HL-60, ML-2, RC-2A, U-937) expressed high numbers of β-adrenoceptors. An intermediate density of receptors was found in a non-T, non-B cell leukemia line (Nall-1), whereas T-cell (JM, CCRF-CEM), B-cell (Raji) or erythroleukemic cell lines (K-562, HEL) displayed minimala or undetectable binding of the radioligand. Isoprenaline-stimulated cAMP production by the cells correlated to their extent of β-adrenoceptor expression. Southern blot hybridization analysis of genomic DNA from the cell lines with a 32 P-labelled β 2 -adrenoceptor cDNA probe revealed no evidence for major rearrangement or amplification of the receptor gene. Incubation with isoprenaline in vitro suppressed the proliferation of the receptor-rich RC-2A cells but did not affect the growth rate of the receptor-deficient K-562 cells. Treatment with propranolol slightly enhanced the proliferation of the RC-2A cells but did not markedly alter the growth rate of two other cell lines, regardless of their β-adrenoceptor status. These findings indicate a regulatory influence by the sympathoadrenergic system on selected cells of the myelomonocytic lineage. (au)

Culture of human cell lines by a pathogen-inactivated human platelet lysate.

Science.gov (United States)

Fazzina, R; Iudicone, P; Mariotti, A; Fioravanti, D; Procoli, A; Cicchetti, E; Scambia, G; Bonanno, G; Pierelli, L

2016-08-01

Alternatives to the use of fetal bovine serum (FBS) have been investigated to ensure xeno-free growth condition. In this study we evaluated the efficacy of human platelet lysate (PL) as a substitute of FBS for the in vitro culture of some human cell lines. PL was obtained by pools of pathogen inactivated human donor platelet (PLT) concentrates. Human leukemia cell lines (KG-1, K562, JURKAT, HL-60) and epithelial tumor cell lines (HeLa and MCF-7) were cultured with either FBS or PL. Changes in cell proliferation, viability, morphology, surface markers and cell cycle were evaluated for each cell line. Functional characteristics were analysed by drug sensitivity test and cytotoxicity assay. Our results demonstrated that PL can support growth and expansion of all cell lines, although the cells cultured in presence of PL experienced a less massive proliferation compared to those grown with FBS. We found a comparable percentage of viable specific marker-expressing cells in both conditions, confirming lineage fidelity in all cultures. Functionality assays showed that cells in both FBS- and PL-supported cultures maintained their normal responsiveness to adriamycin and NK cell-mediated lysis. Our findings indicate that PL is a feasible serum substitute for supporting growth and propagation of haematopoietic and epithelial cell lines with many advantages from a perspective of process standardization, ethicality and product safety.

Combined Treatment with Low Concentrations of Decitabine and SAHA Causes Cell Death in Leukemic Cell Lines but Not in Normal Peripheral Blood Lymphocytes

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Barbora Brodská

2013-01-01

Full Text Available Epigenetic therapy reverting aberrant acetylation or methylation offers the possibility to target preferentially tumor cells and to preserve normal cells. Combination epigenetic therapy may further improve the effect of individual drugs. We investigated combined action of demethylating agent decitabine and histone deacetylase inhibitor SAHA (Vorinostat on different leukemic cell lines in comparison with peripheral blood lymphocytes. Large decrease of viability, as well as huge p21WAF1 induction, reactive oxygen species formation, and apoptotic features due to combined decitabine and SAHA action were detected in leukemic cell lines irrespective of their p53 status, while essentially no effect was observed in response to the combined drug action in normal peripheral blood lymphocytes of healthy donors. p53-dependent apoptotic pathway was demonstrated to participate in the wtp53 CML-T1 leukemic cell line response, while significant influence of reactive oxygen species on viability decrease has been detected in p53-null HL-60 cell line.

Improved viability and activity of neutrophils differentiated from HL-60 cells by co-culture with adipose tissue-derived mesenchymal stem cells

International Nuclear Information System (INIS)

Park, Yoon Shin; Lim, Goh-Woon; Cho, Kyung-Ah; Woo, So-Youn; Shin, Meeyoung; Yoo, Eun-Sun; Chan Ra, Jeong; Ryu, Kyung-Ha

2012-01-01

Highlights: ► Neutropenia is a principal complication of cancer treatment. ► Co-culture of neutrophils with AD-MSC retained cell survival and proliferation and inhibited neutrophil apoptosis under serum starved conditions. ► AD-MSC increased functions of neutrophil. ► AD-MSC promoted the viability of neutrophils by enhancing respiratory burst through the expression of IFN-α, G-CSF, and TGF-β. ► AD-MSC can be used to improve immunity for neutropenia treatment. -- Abstract: Neutropenia is a principal complication of cancer treatment. We investigated the supportive effect of adipose tissue-derived mesenchymal stem cells (AD-MSCs) on the viability and function of neutrophils. Neutrophils were derived from HL-60 cells by dimethylformamide stimulation and cultured with or without AD-MSCs under serum-starved conditions to evaluate neutrophil survival, proliferation, and function. Serum starvation resulted in the apoptosis of neutrophils and decreased cell survival. The co-culture of neutrophils and AD-MSCs resulted in cell survival and inhibited neutrophil apoptosis under serum-starved conditions. The survival rate of neutrophils was prolonged up to 72 h, and the expression levels of interferon (IFN)-α, granulocyte colony-stimulating factor (G-CSF), granulocyte–macrophage colony-stimulating factor, and transforming growth factor (TGF)-β in AD-MSCs were increased after co-culture with neutrophils. AD-MSCs promoted the viability of neutrophils by inhibiting apoptosis as well as enhancing respiratory burst, which could potentially be mediated by the increased expression of IFN-α, G-CSF, and TGF-β. Thus, we conclude that the use of AD-MSCs may be a promising cell-based therapy for increasing immunity by accelerating neutrophil function.

Low dose perfluorooctanoate exposure promotes cell proliferation in a human non-tumor liver cell line

Energy Technology Data Exchange (ETDEWEB)

Zhang, Hongxia; Cui, Ruina [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Guo, Xuejiang [State Key Laboratory of Reproductive Medicine, Nanjing Medical University, Nanjing 210029 (China); Hu, Jiayue [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Dai, Jiayin, E-mail: daijy@ioz.ac.cn [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China)

2016-08-05

Highlights: • Differential expression of proteins induced by PFOA in HL-7702 was identified. • Most of the differentially expressed proteins are related to cell proliferation. • A low dose of PFOA stimulates HL-7702 cell proliferation. • A high dose of PFOA inhibits HL-7702 cell proliferation. - Abstract: Perfluorooctanoate (PFOA) is a well-known persistent organic pollutant widely found in the environment, wildlife and humans. Medical surveillance and experimental studies have investigated the potential effects of PFOA on human livers, but the hepatotoxicity of PFOA on humans and its underlying mechanism remain to be clarified. We exposed a human liver cell line (HL-7702) to 50 μM PFOA for 48 h and 96 h, and identified 111 significantly differentially expressed proteins by iTRAQ analysis. A total of 46 proteins were related to cell proliferation and apoptosis. Through further analysis of the cell cycle, apoptosis and their related proteins, we found that low doses of PFOA (50–100 μM) promoted cell proliferation and numbers by promoting cells from the G1 to S phases, whereas high doses of PFOA (200–400 μM) led to reduced HL-7702 cell numbers compared with that of the control mainly due to cell cycle arrest in the G0/G1 phase. To our knowledge, this is the first report on the promotion of cell cycle progression in human cells following PFOA exposure.

Low dose perfluorooctanoate exposure promotes cell proliferation in a human non-tumor liver cell line

International Nuclear Information System (INIS)

Zhang, Hongxia; Cui, Ruina; Guo, Xuejiang; Hu, Jiayue; Dai, Jiayin

2016-01-01

Highlights: • Differential expression of proteins induced by PFOA in HL-7702 was identified. • Most of the differentially expressed proteins are related to cell proliferation. • A low dose of PFOA stimulates HL-7702 cell proliferation. • A high dose of PFOA inhibits HL-7702 cell proliferation. - Abstract: Perfluorooctanoate (PFOA) is a well-known persistent organic pollutant widely found in the environment, wildlife and humans. Medical surveillance and experimental studies have investigated the potential effects of PFOA on human livers, but the hepatotoxicity of PFOA on humans and its underlying mechanism remain to be clarified. We exposed a human liver cell line (HL-7702) to 50 μM PFOA for 48 h and 96 h, and identified 111 significantly differentially expressed proteins by iTRAQ analysis. A total of 46 proteins were related to cell proliferation and apoptosis. Through further analysis of the cell cycle, apoptosis and their related proteins, we found that low doses of PFOA (50–100 μM) promoted cell proliferation and numbers by promoting cells from the G1 to S phases, whereas high doses of PFOA (200–400 μM) led to reduced HL-7702 cell numbers compared with that of the control mainly due to cell cycle arrest in the G0/G1 phase. To our knowledge, this is the first report on the promotion of cell cycle progression in human cells following PFOA exposure.

Irradiated KHYG-1 retains cytotoxicity: potential for adoptive immunotherapy with a natural killer cell line.

Science.gov (United States)

Suck, G; Branch, D R; Keating, A

2006-05-01

To evaluate gamma-irradiation on KHYG-1, a highly cytotoxic natural killer (NK) cell line and potential candidate for cancer immunotherapy. The NK cell line KHYG-1 was irradiated at 1 gray (Gy) to 50 Gy with gamma-irradiation, and evaluated for cell proliferation, cell survival, and cytotoxicity against tumor targets. We showed that a dose of at least 10 Gy was sufficient to inhibit proliferation of KHYG-1 within the first day but not its cytolytic activity. While 50 Gy had an apoptotic effect in the first hours after irradiation, the killing of K562 and HL60 targets was not different from non-irradiated cells but was reduced for the Ph + myeloid leukemia lines, EM-2 and EM-3. gamma-irradiation (at least 10 Gy) of KHYG-1 inhibits cell proliferation but does not diminish its enhanced cytolytic activity against several tumor targets. This study suggests that KHYG-1 may be a feasible immunotherapeutic agent in the treatment of cancers.

Dysfunctional p53 deletion mutants in cell lines derived from Hodgkin's lymphoma

DEFF Research Database (Denmark)

Feuerborn, Alexander; Moritz, Constanze; von Bonin, Frederike

2006-01-01

Classical Hodgkin's lymphoma (cHL) is a distinct malignancy of the immune system. Despite the progress made in the understanding of the pathology of cHL, the transforming events remain to be elucidated. It has been proposed that mutations in the TP53 gene in biopsy material as well as cell lines ...

Ethyl acetate extract of Chinese medicinal herb Sarcandra glabra induces growth inhibition on human leukemic HL-60 cells, associated with cell cycle arrest and up-regulation of pro-apoptotic Bax/Bcl-2 ratio.

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Li, W Y; Chiu, Lawrence C M; Lam, W S; Wong, W Y; Chan, Y T; Ho, Y P; Wong, Elaine Y L; Wong, Y S; Ooi, Vincent E C

2007-02-01

Sarcandra glabra (Thunb.) Nakai, colloquially known as Caoshanhu, is a Chinese medicinal herb with reported anti-tumor, anti-inflammatory, anti-viral and non-specific immunoenhancing properties. Although the plant has been clinically used for treating a variety of diseases, its bioactive ingredients are largely unknown and its mode of action has never been investigated. In this study, the anti-tumor property of ethyl acetate (EA) extract of S. glabra was investigated by determining its in vitro growth-inhibitory effects on a panel of human cancer cell lines of different histotypes. Growth inhibition of the EA extract on the cancer cells seemed to be selective, and the leukemic HL-60 was found to be the most responsive after 48 h of treatment (IC50=58 microg/ml). Flow cytometric studies further illustrated that the extract might interfere with DNA replication and thus arrested the cell cycle at S phase in the leukemic cells, followed by DNA fragmentation and loss of phospholipid asymmetry in the plasma membrane after 72 h of treatment. Concurrently, the pro-apoptotic Bax/Bcl-2 ratio was also up-regulated by more than 178% of the control level. All these findings suggested that the extract had initiated apoptosis to kill the leukemic cells. Results from this pioneer study help to establish a scientific foundation for future research and development of the bioactive ingredients in EA extract of S. glabra as efficacious anti-cancer agents.

Improved viability and activity of neutrophils differentiated from HL-60 cells by co-culture with adipose tissue-derived mesenchymal stem cells

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Park, Yoon Shin; Lim, Goh-Woon [Department of Pediatrics, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of); Cho, Kyung-Ah; Woo, So-Youn; Shin, Meeyoung [Department of Microbiology, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of); Yoo, Eun-Sun [Department of Pediatrics, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of); Chan Ra, Jeong [Stem Cell Research Center, RNL BIO, Seoul 153-768 (Korea, Republic of); Ryu, Kyung-Ha, E-mail: ykh@ewha.ac.kr [Department of Pediatrics, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of)

2012-06-22

Highlights: Black-Right-Pointing-Pointer Neutropenia is a principal complication of cancer treatment. Black-Right-Pointing-Pointer Co-culture of neutrophils with AD-MSC retained cell survival and proliferation and inhibited neutrophil apoptosis under serum starved conditions. Black-Right-Pointing-Pointer AD-MSC increased functions of neutrophil. Black-Right-Pointing-Pointer AD-MSC promoted the viability of neutrophils by enhancing respiratory burst through the expression of IFN-{alpha}, G-CSF, and TGF-{beta}. Black-Right-Pointing-Pointer AD-MSC can be used to improve immunity for neutropenia treatment. -- Abstract: Neutropenia is a principal complication of cancer treatment. We investigated the supportive effect of adipose tissue-derived mesenchymal stem cells (AD-MSCs) on the viability and function of neutrophils. Neutrophils were derived from HL-60 cells by dimethylformamide stimulation and cultured with or without AD-MSCs under serum-starved conditions to evaluate neutrophil survival, proliferation, and function. Serum starvation resulted in the apoptosis of neutrophils and decreased cell survival. The co-culture of neutrophils and AD-MSCs resulted in cell survival and inhibited neutrophil apoptosis under serum-starved conditions. The survival rate of neutrophils was prolonged up to 72 h, and the expression levels of interferon (IFN)-{alpha}, granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor, and transforming growth factor (TGF)-{beta} in AD-MSCs were increased after co-culture with neutrophils. AD-MSCs promoted the viability of neutrophils by inhibiting apoptosis as well as enhancing respiratory burst, which could potentially be mediated by the increased expression of IFN-{alpha}, G-CSF, and TGF-{beta}. Thus, we conclude that the use of AD-MSCs may be a promising cell-based therapy for increasing immunity by accelerating neutrophil function.

1,1-Diphenyl-2-picrylhydrazyl radical-scavenging compounds from soybean miso and antiproliferative activity of isoflavones from soybean miso toward the cancer cell lines.

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Hirota, A; Taki, S; Kawaii, S; Yano, M; Abe, N

2000-05-01

Guided by their DPPH radical-scavenging activity, nine compounds were isolated from soybean miso. Of these, 8-hydroxydaidzein, 8-hydroxygenistein and syringic acid had as high DPPH radical-scavenging activity as that of alpha-tocopherol. The antiproliferative activity of four of the isolated isoflavones toward three cancer cell lines was examined. 8-Hydroxygenistein showed the highest activity (IC50=5.2 microM) toward human promyelocytic leukemia cells (HL-60).

Radotinib Induces Apoptosis of CD11b+ Cells Differentiated from Acute Myeloid Leukemia Cells.

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Sook-Kyoung Heo

Full Text Available Radotinib, developed as a BCR/ABL tyrosine kinase inhibitor (TKI, is approved for the second-line treatment of chronic myeloid leukemia (CML in South Korea. However, therapeutic effects of radotinib in acute myeloid leukemia (AML are unknown. In the present study, we demonstrate that radotinib significantly decreases the viability of AML cells in a dose-dependent manner. Kasumi-1 cells were more sensitive to radotinib than NB4, HL60, or THP-1 cell lines. Furthermore, radotinib induced CD11b expression in NB4, THP-1, and Kasumi-1 cells either in presence or absence of all trans-retinoic acid (ATRA. We found that radotinib promoted differentiation and induced CD11b expression in AML cells by downregulating LYN. However, CD11b expression induced by ATRA in HL60 cells was decreased by radotinib through upregulation of LYN. Furthermore, radotinib mainly induced apoptosis of CD11b+ cells in the total population of AML cells. Radotinib also increased apoptosis of CD11b+ HL60 cells when they were differentiated by ATRA/dasatinib treatment. We show that radotinib induced apoptosis via caspase-3 activation and the loss of mitochondrial membrane potential (ΔΨm in CD11b+ cells differentiated from AML cells. Our results suggest that radotinib may be used as a candidate drug in AML or a chemosensitizer for treatment of AML by other therapeutics.

An essential role of syntaxin 3 protein for granule exocytosis and secretion of IL-1α, IL-1β, IL-12b, and CCL4 from differentiated HL-60 cells.

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Naegelen, Isabelle; Plançon, Sébastien; Nicot, Nathalie; Kaoma, Tony; Muller, Arnaud; Vallar, Laurent; Tschirhart, Eric J; Bréchard, Sabrina

2015-03-01

Besides their roles in the killing of pathogens, neutrophils have the capacity to package a variety of cytokines into cytoplasmic granules for subsequent release upon inflammatory conditions. Because the rapid secretion of cytokines orchestrates the action of other immune cells at the infection site and thus, can contribute to the development and chronicity of inflammatory diseases, we aimed to determine the intracellular SNARE machinery responsible for the regulation of cytokine secretion and degranulation. From a constructed gene-expression network, we first selected relevant cytokines for functional validation by the CBA approach. We established a cytokine-secretion profile for human neutrophils and dHL-60 cells, underlining their similar ability to secrete a broad variety of cytokines within proinflammatory conditions mimicked by LPS stimulation. Secondly, after screening of SNARE genes by microarray experiments, we selected STX3 for further functional studies. With the use of a siRNA strategy, we show that STX3 is clearly required for the maximal release of IL-1α, IL-1β, IL-12b, and CCL4 without alteration of other cytokine secretion in dHL-60 cells. In addition, we demonstrate that STX3 is involved in MMP-9 exocytosis from gelatinase granules, where STX3 is partly localized. Our results suggest that the secretion of IL-1α, IL-1β, IL-12b, and CCL4 occurs during gelatinase degranulation, a process controlled by STX3. In summary, these findings provide first evidence that STX3 has an essential role in trafficking pathways of cytokines in neutrophil granulocytes. © Society for Leukocyte Biology.

Modulation of butyrate anticancer activity by solid lipid nanoparticle delivery: an in vitro investigation on human breast cancer and leukemia cell lines.

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Foglietta, Federica; Serpe, Loredana; Canaparo, Roberto; Vivenza, Nicoletta; Riccio, Giovanna; Imbalzano, Erica; Gasco, Paolo; Zara, Gian Paolo

2014-01-01

Histone modification has emerged as a promising approach to cancer therapy. The short-chain fatty acid, butyric acid, a histone deacetylase (HD) inhibitor, has shown anticancer activity. Butyrate transcriptional activation is indeed able to withdraw cancer cells from the cell cycle, leading to programmed cell death. Since butyrate's clinical use is hampered by unfavorable pharmacokinetic and pharmacodynamic properties, delivery systems, such as solid lipid nanoparticles (SLN), have been developed to overcome these constraints. In order to outline the influence of butyrate delivery on its anticancer activity, the effects of butyrate as a free (sodium butyrate, NB) or nanoparticle (cholesteryl butyrate solid lipid nanoparticles, CBSLN) formulation on the growth of different human cancer cell lines, such as the promyelocytic leukemia, HL-60, and the breast cancer, MCF-7 was investigated. A detailed investigation into the mechanism of the induced cytotoxicity was also carried out, with a special focus on the modulation of HD and cyclin-dependent kinase (CDK) mRNA gene expression by real time PCR analysis. In HL-60 cells, CBSLN induced a higher and prolonged expression level of the butyrate target genes at lower concentrations than NB. This led to a significant decrease in cell proliferation, along with considerable apoptosis, cell cycle block in the G0/G1 phase, significant inhibition of total HD activity and overexpression of the p21 protein. Conversely, in MCF-7 cells, CBSLN did not enhance the level of expression of the butyrate target genes, leading to the same anticancer activity as that of NB. Solid lipid nanoparticles were able to improve butyrate anticancer activity in HL-60, but not in MCF-7 cells. This is consistent with difference in properties of the cells under study, such as expression of the TP53 tumor suppressor, or the transporter for short-chain fatty acids, SLC5A8.

Hypoxia-induced alteration of tracer accumulation in cultured cancer cells and xenografts in mice: implications for pre-therapeutic prediction of treatment outcomes with 99mTc-sestamibi, 201Tl chloride and 99mTc-HL91

International Nuclear Information System (INIS)

Kinuya, Seigo; Yokoyama, Kunihiko; Li, Xiao-Feng; Bai, Jingming; Michigishi, Takatoshi; Tonami, Norihisa; Watanabe, Naoto; Shuke, Noriyuki; Takayama, Teruhiko; Bunko, Hisashi

2002-01-01

Weak visualization of tumours in pre-therapeutic scintigrams with technetium-99m sestamibi (MIBI) is likely a predictive sign of unfavourable tumour response to radiotherapy and chemotherapy. However, factors relating to this scintigraphic finding are not well understood. The presence of hypoxic tumour cells is one of the major reasons for therapeutic failure; consequently, we attempted to determine whether oxygenation status affects 99m Tc-MIBI accumulation in tumour cells. LS180 human colon cancer and T24 human bladder cancer cells were incubated in air or N 2 gas at 37 C. Cellular uptake of 99m Tc-MIBI was subsequently determined at 15, 60 and 120 min. Uptake of thallium-201 chloride was also assessed. Uptake of 99m Tc-HL91 was assessed as a hypoxic marker. Accumulation of the tracers in LS180 xenografts was observed in mice treated with 5 mg/kg hydralazine and compared with that in untreated mice. pO 2 in the medium and tumours was measured with O 2 microelectrodes. N 2 gas flow gradually reduced pO 2 in the cell suspension to 1-2 mmHg in 60 min. Cellular uptake of 99m Tc-MIBI in LS180 cells decreased by approximately 30% in N 2 gas in comparison to that in air throughout the study. Hypoxia had a more prominent influence on 201 Tl uptake, which displayed a reduction of approximately 60% in N 2 gas at 120 min, than on 99m Tc-MIBI uptake. On the other hand, N 2 gas induced an increase of 170% in 99m Tc-HL91 uptake at 120 min, indicating the hypoxic condition of cells. The results of in vitro assays employing the T24 cell line were similar to those obtained with the LS180 cell line. Hydralazine treatment markedly reduced 99m Tc-MIBI and 201 Tl accumulation in LS180 xenografts; moreover, intratumoural pO 2 decreased from 14.5±6.6 mmHg to 7.6±6.2 mmHg. 99m Tc-HL91 accumulation in xenografts was markedly increased by hydralazine. In conclusion, hypoxia reduced accumulation of 99m Tc-MIBI and 201 Tl in tumour cells. Accordingly, hypoxia may be an important factor in

HL-60 differentiating activity and flavonoid content of the readily extractable fraction prepared from citrus juices.

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Kawaii, S; Tomono, Y; Katase, E; Ogawa, K; Yano, M

1999-01-01

Citrus plants are rich sources of various bioactive flavonoids. To eliminate masking effects caused by hesperidin, naringin, and neoeriocitrin, the abundant flavonoid glycosides which make up 90% of the conventionally prepared sample, the readily extractable fraction from Citrus juice was prepared by adsorbing on HP-20 resin and eluting with EtOH and acetone from the resin and was subjected to HL-60 differentiation assay and quantitative analysis of major flavonoids. Screening of 34 Citrus juices indicated that King (C. nobilis) had a potent activity for inducing differentiation of HL-60, and the active principles were isolated and identified as four polymethoxylated flavonoids, namely, nobiletin, 3,3',4',5,6,7, 8-heptamethoxyflavone, natsudaidain, and tangeretin. HPLC analysis of the readily extractable fraction also indicated that King contained high amounts of these polymethoxylated flavonoids among the Citrus juices examined. Principal component and cluster analyses of the readily extractable flavonoids indicated peculiarities of King and Bergamot.

Analysis of myelomonocytic leukemic differentiation by a cell surface marker panel including a fucose-binding lectin from Lotus tetragonolobus.

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Elias, L; Van Epps, D E

1984-06-01

The fucose-binding lectin from Lotus tetragonolobus ( FBL -L) has been previously shown to bind specifically to normal cells of the myeloid and monocytic lineages. The purpose of this study was to explore the utility of fluoresceinated FBL -L as a leukemia differentiation marker in conjunction with a panel of other frequently used surface markers (Fc receptor, HLA-DR, OKM1, and antimonocyte antibody). FBL -L reacted with leukemic cells in 8/9 cases of clinically recognized acute myeloid leukemia, including myeloid blast crisis of chronic granulocytic leukemia, 3/3 cases of chronic phase chronic myelogenous leukemia, and in 2/7 cases of clinically undifferentiated acute leukemia. Correlations were noted between reactivity with FBL -L, and DR and Fc receptor expression. Among continuous cell lines, FBL -L bound with high intensity to a majority of HL-60 and U937 cells. The less well differentiated myeloblast cell lines, KG-1, KG1a , and HL-60 blast II, exhibited less FBL -L binding than HL-60 and U937. A moderate proportion of K562 cells exhibited low level binding of FBL -L. Several lymphoblastic cell lines exhibited a pattern of low intensity binding that was distinguishable from the high intensity binding pattern of the myeloblastic lines. FBL -L reactivity of U937 was enhanced by induction of differentiation with leukocyte conditioned medium, but not dimethylsulfoxide. Such treatments induced contrasting patterns of change of HL-60 and U937 when labeled with OKM1, alpha-Mono, and HLA-DR. These studies demonstrate the application of FBL -L to analysis and quantitation of myelomonocytic leukemic differentiation.

2,2',4,4'-Tetrachlorobiphenyl upregulates cyclooxygenase-2 in HL-60 cells via p38 mitogen-activated protein kinase and NF-κB

International Nuclear Information System (INIS)

Bezdecny, Steven A.; Karmaus, Peer; Roth, Robert A.; Ganey, Patricia E.

2007-01-01

Polychlorinated biphenyls (PCBs) are ubiquitous, persistent environmental contaminants that affect a number of cellular systems, including neutrophils. Among the effects caused by the noncoplanar PCB 2,2',4,4'-tetrachlorobiphenyl (2244-TCB) in granulocytic HL-60 cells are increases in superoxide anion production, activation of phospholipase A 2 with subsequent release of arachidonic acid (AA) and upregulation of the inflammatory gene cyclooxygenase-2 (COX-2). The objective of this study was to determine the signal transduction pathways involved in the upregulation of COX-2 by 2244-TCB. Treatment of HL-60 cells with 2244-TCB led to increased expression of COX-2 mRNA. This increase was prevented by the transcriptional inhibitor actinomycin D in cells pretreated with 2244-TCB for 10 min. The increase in COX-2 mRNA was associated with release of 3 H-AA, phosphorylation of p38 and extracellular signal-regulated kinase (ERK) mitogen-activated protein (MAP) kinases, increased levels of nuclear NF-κB and increased superoxide anion production. Bromoenol lactone, an inhibitor of the calcium-independent phospholipase A 2 , reduced 3 H-AA release but had no effect on COX-2 mRNA, protein or activity. Pretreatment with SB-202190 or SB-203580, inhibitors of the p38 MAP kinase pathway, prevented the 2244-TCB-mediated induction of COX-2 and phosphorylation of p38 and ERK MAP kinases. These inhibitors did not alter 3 H-AA release. Treatment with PD 98059 or U 0126, inhibitors of the MAP/ERK (MEK) pathway, prevented the 2244-TCB-mediated activation of ERK but had no effect on COX-2 induction or p38 phosphorylation. 2244-TCB treatment did not affect c-Jun N-terminal kinase (JNK) phosphorylation. 2244-TCB exposure increased the amount of nuclear NF-κB. This increase was prevented by pretreatment with p38 MAP kinase inhibitors, but not by pretreatment with MEK inhibitors. Pretreatment with inhibitors of NF-κB prevented the 2244-TCB-mediated induction of COX-2 mRNA. 2244-TCB

Human monoclonal antibodies reactive with human myelomonocytic leukemia cells.

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Posner, M R; Santos, D J; Elboim, H S; Tumber, M B; Frackelton, A R

1989-04-01

Peripheral blood mononuclear cells from a patient with chronic myelogenous leukemia (CML), in remission, were depleted of CD8-positive T-cells and cultured with Epstein-Barr virus. Four of 20 cultures (20%) secreted human IgG antibodies selectively reactive with the cell surfaces of certain human leukemia cell lines. Three polyclonal, Epstein-Barr virus-transformed, B-cell lines were expanded and fused with the human-mouse myeloma analogue HMMA2.11TG/O. Antibody from secreting clones HL 1.2 (IgG1), HL 2.1 (IgG3), and HL 3.1 (IgG1) have been characterized. All three react with HL-60 (promyelocytic), RWLeu4 (CML promyelocytic), and U937 (monocytic), but not with KG-1 (myeloblastic) or K562 (CML erythroid). There is no reactivity with T-cell lines, Burkitt's cell lines, pre-B-leukemia cell lines, or an undifferentiated CML cell line, BV173. Leukemic cells from two of seven patients with acute myelogenous leukemia and one of five with acute lymphocytic leukemia react with all three antibodies. Normal lymphocytes, monocytes, polymorphonuclear cells, red blood cells, bone marrow cells, and platelets do not react. Samples from patients with other diverse hematopoietic malignancies showed no reactivity. Immunoprecipitations suggest that the reactive antigen(s) is a lactoperoxidase iodinatable series of cell surface proteins with molecular weights of 42,000-54,000 and a noniodinatable protein with a molecular weight of 82,000. Based on these data these human monoclonal antibodies appear to react with myelomonocytic leukemic cells and may detect a leukemia-specific antigen or a highly restricted differentiation antigen.

CellMiner: a relational database and query tool for the NCI-60 cancer cell lines

Directory of Open Access Journals (Sweden)

Reinhold William C

2009-06-01

Full Text Available Abstract Background Advances in the high-throughput omic technologies have made it possible to profile cells in a large number of ways at the DNA, RNA, protein, chromosomal, functional, and pharmacological levels. A persistent problem is that some classes of molecular data are labeled with gene identifiers, others with transcript or protein identifiers, and still others with chromosomal locations. What has lagged behind is the ability to integrate the resulting data to uncover complex relationships and patterns. Those issues are reflected in full form by molecular profile data on the panel of 60 diverse human cancer cell lines (the NCI-60 used since 1990 by the U.S. National Cancer Institute to screen compounds for anticancer activity. To our knowledge, CellMiner is the first online database resource for integration of the diverse molecular types of NCI-60 and related meta data. Description CellMiner enables scientists to perform advanced querying of molecular information on NCI-60 (and additional types through a single web interface. CellMiner is a freely available tool that organizes and stores raw and normalized data that represent multiple types of molecular characterizations at the DNA, RNA, protein, and pharmacological levels. Annotations for each project, along with associated metadata on the samples and datasets, are stored in a MySQL database and linked to the molecular profile data. Data can be queried and downloaded along with comprehensive information on experimental and analytic methods for each data set. A Data Intersection tool allows selection of a list of genes (proteins in common between two or more data sets and outputs the data for those genes (proteins in the respective sets. In addition to its role as an integrative resource for the NCI-60, the CellMiner package also serves as a shell for incorporation of molecular profile data on other cell or tissue sample types. Conclusion CellMiner is a relational database tool for

Involvement of histone H3 phosphorylation via the activation of p38 MAPK pathway and intracellular redox status in cytotoxicity of HL-60 cells induced by Vitex agnus-castus fruit extract.

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Kikuchi, Hidetomo; Yuan, Bo; Yuhara, Eisuke; Imai, Masahiko; Furutani, Ryota; Fukushima, Shin; Hazama, Shingo; Hirobe, Chieko; Ohyama, Kunio; Takagi, Norio; Toyoda, Hiroo

2014-08-01

We have demonstrated that an extract from the ripe fruit of Vitex angus-castus (Vitex), might be a promising anticancer candidate. In order to further provide a molecular rationale for clinical development in anticancer therapy, a detailed mechanism underlying the efficacy of Vitex against HL-60 cells was investigated. Vitex induced a dose- and time-dependent decrease in cell viability associated with induction of apoptosis and G(2)/M cell cycle arrest, both of which were suppressed by the addition of SB203580, an inhibitor for p38 MAPK. Furthermore, SB203580 significantly suppressed Vitex-induced phosphorylation of histone H3, a downstream molecule of p38 MAPK known to be involved in apoptosis induction in tumor cells. Notably, Vitex induced upregulation of intracellular ATP, known to bind its binding pocket inside activated p38 MAPK and to be required for the activation of p38 MAPK pathway. These results, thus, suggest that upregulation of intracellular ATP and phosphorylation of histone H3 are closely associated with the activation of p38 MAPK pathway, consequently contributing to Vitex-mediated cytotoxicity. Intriguingly, a significant decrease of intracellular ROS levels and downregulation of expression level of gp91(phox), an important component of NADPH oxidase, were observed in Vitex-treated cells. A greater decline in ROS levels along with enhanced apoptosis was observed after treatment with Vitex in combination with SnPP, an inhibitor specific for HO-1. Since NADPH oxidase and HO-1 are closely correlated to redox status associated with intracellular ROS levels, the two enzymes are suggested to be implicated in Vitex-mediated cytotoxicity in HL-60 cells by regulating ROS generation. We also suggest that activation of the p38 MAPK pathway may be dependent on the alterations of intracellular ATP levels, rather than that of intracellular ROS levels. These results may have important implications for appropriate clinical uses of Vitex and provide novel insights

Is P-glycoprotein (ABCB1) a phase 0 or a phase 3 colchicine transporter depending on colchicine exposure conditions?

International Nuclear Information System (INIS)

Decleves, Xavier.; Niel, Elisabeth; Debray, Marcel; Scherrmann, Jean-Michel

2006-01-01

This study investigates the P-glycoprotein (Pgp)-mediated transport of its substrates in accumulation or efflux modes under steady-state conditions. The kinetics of colchicine uptake and efflux, a substrate of both Pgp and intracellular tubulin, were studied in HL60 and HL60/DNR cells; HL60/DNR cells contain 25 times more Pgp than do HL60 cells. HL60/DNR cells in a medium containing 6.25 nM colchicine, which mimics therapeutic conditions, reached steady-state twice as rapidly as did HL60 cells, and accumulated 24-times less colchicine than did HL60 cells. The Pgp inhibitor GF120918, increased colchicine uptake by HL60 cells 1.2-fold and that of HL60/DNR cells 17-fold, while it had no effect on colchicine efflux from either cell line that had been incubated with colchicine for 24 h. Colchicine kinetics fitted well a two closed-compartment model, showing that the low intracellular accumulation of colchicine in HL60/DNR cells resulted from a 11-fold decrease in colchicine uptake and a 2.3-fold increase in colchicine efflux, that could be attributed to Pgp-mediated efflux activity in HL60/DNR cells. Intracellular colchicine was mainly and similarly distributed in the cytosol in both cell lines. These data demonstrate that the kinetics of the intracellular colchicine accumulation depend on the density of Pgp and that Pgp is more a phase 0 (preventing cellular uptake) than a phase 3 (effluxing intracellular substrate) transporter under steady-state conditions, although the situation is reversed after a short incubation time (30 min), when intracellular free colchicine concentration is probably high enough for it to be removed from the cell by Pgp

Permissivity of the NCI-60 cancer cell lines to oncolytic Vaccinia Virus GLV-1h68

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Ascierto, Maria Libera; Bedognetti, Davide; Uccellini, Lorenzo; Rossano, Fabio; Ascierto, Paolo A; Stroncek, David F; Restifo, Nicholas P; Wang, Ena; Szalay, Aladar A; Marincola, Francesco M; Worschech, Andrea; Yu, Zhiya; Adams, Sharon; Reinboth, Jennifer; Chen, Nanhai G; Pos, Zoltan; Roychoudhuri, Rahul; Di Pasquale, Giovanni

2011-01-01

Oncolytic viral therapy represents an alternative therapeutic strategy for the treatment of cancer. We previously described GLV-1h68, a modified Vaccinia Virus with exclusive tropism for tumor cells, and we observed a cell line-specific relationship between the ability of GLV-1h68 to replicate in vitro and its ability to colonize and eliminate tumor in vivo. In the current study we surveyed the in vitro permissivity to GLV-1h68 replication of the NCI-60 panel of cell lines. Selected cell lines were also tested for permissivity to another Vaccinia Virus and a vesicular stomatitis virus (VSV) strain. In order to identify correlates of permissity to viral infection, we measured transcriptional profiles of the cell lines prior infection. We observed highly heterogeneous permissivity to VACV infection amongst the cell lines. The heterogeneity of permissivity was independent of tissue with the exception of B cell derivation. Cell lines were also tested for permissivity to another Vaccinia Virus and a vesicular stomatitis virus (VSV) strain and a significant correlation was found suggesting a common permissive phenotype. While no clear transcriptional pattern could be identified as predictor of permissivity to infection, some associations were observed suggesting multifactorial basis permissivity to viral infection. Our findings have implications for the design of oncolytic therapies for cancer and offer insights into the nature of permissivity of tumor cells to viral infection

Keeping HL-LHC accountable

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2015-01-01

This week saw the cost and schedule of the High Luminosity LHC (HL-LHC) and LHC Injectors Upgrade (LIU) projects come under close scrutiny from the external review committee set up for the purpose.    HL-LHC, whose implementation requires an upgrade to the CERN injector complex, responds directly to one of the key recommendations of the updated European Strategy for Particle Physics, which urges CERN to prepare for a ‘major luminosity upgrade’, a recommendation that is also perfectly in line with the P5 report on the US strategy for the field. Responding to this recommendation, CERN set up the HL-LHC project in 2013, partially supported by FP7 funding through the HiLumi LHC Design Study (2011-2015), and coordinated with the American LARP project, which oversees the US contribution to the upgrade. A key element of HL-LHC planning is a mechanism for receiving independent expert advice on all aspects of the project.  To this end, several technical reviews h...

Initial analysis of the HL-2A tokamak visible light measurements

International Nuclear Information System (INIS)

Shi Le; Zheng Yinjia

2005-01-01

Measurement of H α radial distribution is an important part in a plasma experiment. During the HL-2A experiment, CCD photos recorded the plasma radiation information using a plasma imaging system, particularly the H α line emission information (with H α filter in front of the camera). Since the line of light radiation measurement involves a line integral of the radiating region, data handling and analysis are necessary. The H α radial distribution along an HL-2A tokamak radial radius can be obtained by using an Abel inversion transformation with plasma imaging on the HL-2A tokamak. This paper provides a reliable measurement method on the HL-2A tokamak. (authors)

Overexpression of the Large-Conductance, Ca2+-Activated K+ (BK) Channel Shortens Action Potential Duration in HL-1 Cardiomyocytes.

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Stimers, Joseph R; Song, Li; Rusch, Nancy J; Rhee, Sung W

2015-01-01

Long QT syndrome is characterized by a prolongation of the interval between the Q wave and the T wave on the electrocardiogram. This abnormality reflects a prolongation of the ventricular action potential caused by a number of genetic mutations or a variety of drugs. Since effective treatments are unavailable, we explored the possibility of using cardiac expression of the large-conductance, Ca2+-activated K+ (BK) channel to shorten action potential duration (APD). We hypothesized that expression of the pore-forming α subunit of human BK channels (hBKα) in HL-1 cells would shorten action potential duration in this mouse atrial cell line. Expression of hBKα had minimal effects on expression levels of other ion channels with the exception of a small but significant reduction in Kv11.1. Patch-clamped hBKα expressing HL-1 cells exhibited an outward voltage- and Ca2+-sensitive K+ current, which was inhibited by the BK channel blocker iberiotoxin (100 nM). This BK current phenotype was not detected in untransfected HL-1 cells or in HL-1 null cells sham-transfected with an empty vector. Importantly, APD in hBKα-expressing HL-1 cells averaged 14.3 ± 2.8 ms (n = 10), which represented a 53% reduction in APD compared to HL-1 null cells lacking BKα expression. APD in the latter cells averaged 31.0 ± 5.1 ms (n = 13). The shortened APD in hBKα-expressing cells was restored to normal duration by 100 nM iberiotoxin, suggesting that a repolarizing K+ current attributed to BK channels accounted for action potential shortening. These findings provide initial proof-of-concept that the introduction of hBKα channels into a cardiac cell line can shorten APD, and raise the possibility that gene-based interventions to increase hBKα channels in cardiac cells may hold promise as a therapeutic strategy for long QT syndrome.

Mechanism of the protective effects of long chain n-alkyl glucopyranosides against ultrasound-induced cytolysis of HL-60 cells.

Science.gov (United States)

Cheng, Jason Y; Riesz, Peter

2007-07-01

Recently it has been shown that long chain (C5-C8) n-alkyl glucopyranosides completely inhibit ultrasound-induced cytolysis [J.Z. Sostaric, N. Miyoshi, P. Riesz, W.G. DeGraff, and J.B. Mitchell, Free Radical Biol. Med., 39 (2005) 1539]. This protective effect has possible applications in HIFU (high intensity focused ultrasound) for tumor treatment, and in ultrasound assisted drug delivery and gene therapy. n-Alkyl glucopyranosides with hexyl (5mM), heptyl (3mM), octyl (2mM) n-alkyl chains protected 100% of HL-60 cells in vitro from 1.057 MHz ultrasound-induced cytolysis under a range of conditions that resulted in 35-100% cytolysis in the absence of glucopyranosides. However the hydrophilic methyl-beta-d-glucopyranoside did not protect cells. The surface active n-alkyl glucopyranosides accumulate at the gas-liquid interface of cavitation bubbles. The OH radicals and H atoms formed in collapsing cavitation bubbles react by H-atom abstraction from either the n-alkyl chain or the glucose moiety of the n-alkyl glucopyranosides. Owing to the high concentration of the long chain surfactants at the gas-liquid interface of cavitation bubbles, the initially formed carbon radicals on the alkyl chains are transferred to the glucose moieties to yield radicals which react with oxygen leading to the formation of hydrogen peroxide. In this work, we find that the sonochemically produced hydrogen peroxide yields from oxygen-saturated solutions of long chain (hexyl, octyl) n-alkyl glucopyranosides at 614 kHz and 1.057 MHz ultrasound increase with increasing n-alkyl glucopyranoside concentration but are independent of concentration for methyl-beta-D-glucopyranoside. These results are consistent with the previously proposed mechanism of sonoprotection [J.Z. Sostaric, N. Miyoshi, P. Riesz, W.G. DeGraff, and J.B. Mitchell, Free Radical Biol. Med., 39 (2005) 1539]. This sequence of events prevents sonodynamic cell killing by initiation of lipid peroxidation chain reactions in cellular

Investigation of the Antiproliferative Properties of Natural Sesquiterpenes from Artemisia asiatica and Onopordum acanthium on HL-60 Cells in Vitro

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Judit Molnár

2016-02-01

Full Text Available Plants and plant extracts play a crucial role in the research into novel antineoplastic agents. Four sesquiterpene lactones, artecanin (1, 3β-chloro-4α,10α-dihydroxy-1α,2α-epoxy-5α,7αH-guaia-11(13-en-12,6α-olide (2, iso-seco-tanapartholide 3-O-methyl ether (3 and 4β,15-dihydro-3-dehydrozaluzanin C (4, were isolated from two traditionally used Asteraceae species (Onopordum acanthium and Artemisia asiatica. When tested for antiproliferative action on HL-60 leukemia cells, these compounds exhibited reasonable IC50 values in the range 3.6–13.5 μM. Treatment with the tested compounds resulted in a cell cycle disturbance characterized by increases in the G1 and G2/M populations, while there was a decrease in the S phase. Additionally, 1–3 elicited increases in the hypodiploid (subG1 population. The compounds elicited concentration-dependent chromatin condensation and disruption of the membrane integrity, as revealed by Hoechst 33258–propidium staining. Treatment for 24 h resulted in significant increases in activity of caspases-3 and -9, indicating that the tested sesquiterpenes induced the mitochondrial pathway of apoptosis. The proapoptotic properties of the sesquiterpene lactones were additionally demonstrated withannexin V staining. Compounds 1 and 2 increased the Bax/Bcl-2 expression and decreased the expressions of CDK1 and cyclin B2, as determined at the mRNA level by means of RT-PCR. These experimental results indicate that sesquiterpene lactones may be regarded as potential starting structures for the development of novel anticancer agents.

1,25-Dihydroxyvitamin D3 induces biphasic NF-κB responses during HL-60 leukemia cells differentiation through protein induction and PI3K/Akt-dependent phosphorylation/degradation of IκB

International Nuclear Information System (INIS)

Tse, A.K.-W.; Wan, C.-K.; Shen, X.-L.; Zhu, G.-Y.; Cheung, H.-Y.; Yang, M.; Fong, W.-F.

2007-01-01

1,25-Dihydroxyvitamin D 3 (VD 3 ) induces differentiation in a number of leukemia cell lines and under various conditions is able to either stimulate or inhibit nuclear factor kappa B (NF-κB) activity. Here we report a time-dependent biphasic regulation of NF-κB in VD 3 -treated HL-60 leukemia cells. After VD 3 treatment there was an early ∼ 4 h suppression and a late 8-72 h prolonged reactivation of NF-κB. The reactivation of NF-κB was concomitant with increased IKK activities, IKK-mediated IκBα phosphorylation, p65 phosphorylation at residues S276 and S536, p65 nuclear translocation and p65 recruitment to the NF-κB/vitamin D responsive element promoters. In parallel with NF-κB stimulation, there was an up-regulation of NF-κB controlled inflammatory and anti-apoptotic genes such as TNFα, IL-1β and Bcl-xL. VD 3 -triggered reactivation of NF-κB was associated with PI3K/Akt phosphorylation. PI3K/Akt antagonists suppressed VD 3 -stimulated IκBα phosphorylation as well as NF-κB-controlled gene expression. The early ∼ 4 h VD 3 -mediated NF-κB suppression coincided with a prolonged increase of IκBα protein which require de novo protein synthesis, lasted for as least 72 h and was insensitive to MAPK, IKK or PI3K/Akt inhibitors. Our data suggest a novel biphasic regulation of NF-κB in VD 3 -treated leukemia cells and our results may have provided the first molecular explanation for the contradictory observations reported on VD 3 -mediated immune-regulation

Whole genome transcription profiling of Anaplasma phagocytophilum in human and tick host cells by tiling array analysis

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Chavez Adela

2008-07-01

Full Text Available Abstract Background Anaplasma phagocytophilum (Ap is an obligate intracellular bacterium and the agent of human granulocytic anaplasmosis, an emerging tick-borne disease. Ap alternately infects ticks and mammals and a variety of cell types within each. Understanding the biology behind such versatile cellular parasitism may be derived through the use of tiling microarrays to establish high resolution, genome-wide transcription profiles of the organism as it infects cell lines representative of its life cycle (tick; ISE6 and pathogenesis (human; HL-60 and HMEC-1. Results Detailed, host cell specific transcriptional behavior was revealed. There was extensive differential Ap gene transcription between the tick (ISE6 and the human (HL-60 and HMEC-1 cell lines, with far fewer differentially transcribed genes between the human cell lines, and all disproportionately represented by membrane or surface proteins. There were Ap genes exclusively transcribed in each cell line, apparent human- and tick-specific operons and paralogs, and anti-sense transcripts that suggest novel expression regulation processes. Seven virB2 paralogs (of the bacterial type IV secretion system showed human or tick cell dependent transcription. Previously unrecognized genes and coding sequences were identified, as were the expressed p44/msp2 (major surface proteins paralogs (of 114 total, through elevated signal produced to the unique hypervariable region of each – 2/114 in HL-60, 3/114 in HMEC-1, and none in ISE6. Conclusion Using these methods, whole genome transcription profiles can likely be generated for Ap, as well as other obligate intracellular organisms, in any host cells and for all stages of the cell infection process. Visual representation of comprehensive transcription data alongside an annotated map of the genome renders complex transcription into discernable patterns.

Enhanced Missing Proteins Detection in NCI60 Cell Lines Using an Integrative Search Engine Approach.

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Guruceaga, Elizabeth; Garin-Muga, Alba; Prieto, Gorka; Bejarano, Bartolomé; Marcilla, Miguel; Marín-Vicente, Consuelo; Perez-Riverol, Yasset; Casal, J Ignacio; Vizcaíno, Juan Antonio; Corrales, Fernando J; Segura, Victor

2017-12-01

The Human Proteome Project (HPP) aims deciphering the complete map of the human proteome. In the past few years, significant efforts of the HPP teams have been dedicated to the experimental detection of the missing proteins, which lack reliable mass spectrometry evidence of their existence. In this endeavor, an in depth analysis of shotgun experiments might represent a valuable resource to select a biological matrix in design validation experiments. In this work, we used all the proteomic experiments from the NCI60 cell lines and applied an integrative approach based on the results obtained from Comet, Mascot, OMSSA, and X!Tandem. This workflow benefits from the complementarity of these search engines to increase the proteome coverage. Five missing proteins C-HPP guidelines compliant were identified, although further validation is needed. Moreover, 165 missing proteins were detected with only one unique peptide, and their functional analysis supported their participation in cellular pathways as was also proposed in other studies. Finally, we performed a combined analysis of the gene expression levels and the proteomic identifications from the common cell lines between the NCI60 and the CCLE project to suggest alternatives for further validation of missing protein observations.

Hibiscus anthocyanins rich extract-induced apoptotic cell death in human promyelocytic leukemia cells

International Nuclear Information System (INIS)

Chang, Y.-C.; Huang, H.-P.; Hsu, J.-D.; Yang, S.-F.; Wang, C.-J.

2005-01-01

Hibiscus sabdariffa Linne (Malvaceae), an attractive plant believed to be native to Africa, is cultivated in the Sudan and Eastern Taiwan. Anthocyanins exist widely in many vegetables and fruits. Some reports demonstrated that anthocyanins extracted from H. sabdariffa L., Hibiscus anthocyanins (HAs) (which are a group of natural pigments existing in the dried calyx of H. sabdariffa L.) exhibited antioxidant activity and liver protection. Therefore, in this study, we explored the effect of HAs on human cancer cells. The result showed that HAs could cause cancer cell apoptosis, especially in HL-60 cells. Using flow cytometry, we found that HAs treatment (0-4 mg/ml) markedly induced apoptosis in HL-60 cells in a dose- and time-dependent manner. The result also revealed increased phosphorylation in p38 and c-Jun, cytochrome c release, and expression of tBid, Fas, and FasL in the HAs-treated HL-60 cells. We further used SB203580 (p38 inhibitor), PD98059 (MEK inhibitor), SP600125 (JNK inhibitor), and wortmannin (phosphatidylinositol 3-kinase; PI-3K inhibitor) to evaluate their effect on the HAs-induced HL-60 death. The data showed that only SB203580 had strong potential in inhibiting HL-60 cell apoptosis and related protein expression and phosphorylation. Therefore, we suggested that HAs mediated HL-60 apoptosis via the p38-FasL and Bid pathway. According to these results, HAs could be developed as chemopreventive agents. However, further investigations into the specificity and mechanism(s) of HAs are needed

Effects of Ligusticum porteri (Osha) Root Extract on Human Promyelocytic Leukemia Cells.

Science.gov (United States)

Nguyen, Khanh; Sparks, Jean; Omoruyi, Felix

2017-01-01

Ligusticum porteri roots have been traditionally used in folk medicine, but the scientific basis is unclear. To investigate the cytotoxicity, antioxidant, and immunomodulatory effects of L. porteri root extract on human promyelocytic leukemia (HL-60) cells and H 2 O 2 -induced oxidative damaged HL-60 cells. HL-60 cells were incubated with different concentrations of root extract, and cells were harvested for viability assays on day 3 and 7. Cytokine levels (interferon-gamma [IFN-γ], interleukin-2 [IL-2], and interleukin-10 [IL-10]) and antioxidant indexes (malondialdehyde [MDA], reduced glutathione [GSH], superoxide dismutase [SOD], and catalase [CAT]) in H 2 O 2 -induced-stressed HL-60 were measured after 2 days. The viability of HL-60 challenged with H 2 O 2 declined by 42% compared to unstressed cells. After 7 days of incubation with 200 or 400 μg/mL L. porteri , the viability of HL-60 cells was two-fold higher than the control. Stressed HL-60 cells treated with 100, 200, and 400 μg/mL L. porteri reduced the lipid peroxidation by 12%-13%. We noted an increase in GSH levels, SOD and CAT activities in stressed HL-60 supplemented with 400 μg/mL root extract. Treatment with 400 μg/mL L. porteri significantly ( P effect against the oxidation of reduced glutathione (GSH)Treatment with L. porteri root extract may be effective in preventing oxidative damage through increasing the activities of antioxidant enzymes (superoxide dismutase [SOD] and catalase [CAT]) in acute promyelocytic leukemia cells.

Von Hippel-Lindau disease (vHL)

DEFF Research Database (Denmark)

Binderup, Marie Louise Mølgaard; Bisgaard, Søs Marie Luise; Harbud, Vibeke

2013-01-01

These clinical guidelines outline the criteria and recommendations for diagnostic and genetic work-up of families suspected of von Hippel-Lindau disease (vHL), as well as recommendations for prophylactic surveillance for vHL patients. The guideline has been composed by the Danish Coordination Group...... for vHL which is comprised of Danish doctors and specialists interested in vHL. The recommendations are based on longstanding clinical experience, Danish original research, and extensive review of the international literature. vHL is a hereditary multi-tumour disease caused by germline mutations...... of the disease, and/or in individuals with a vHL-associated manifestation; i.e. a hemangioblastoma in the retina or the central nervous system, familial or bilateral pheochromocytomas, familial, multiple, or early onset renal cell carcinomas, and in individuals with an endolymphatic sac tumour in the inner ear...

Isolation and purification of a novel anticancer 60 K daltons protein from the Persian Gulf sea hare, Aplysia Dactylomela

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Keyvan Zandi

2004-02-01

Full Text Available Sea hares have attracted the interest of many workers investigating chemical defense substances. Most of these substances are low molecular weight compounds derived from algal diets. Anticancer effects of a novel protein isolated from purple fluid of A. dactylomela are reported. The purification procedure consisted basically of ammonium sulphate fractionation, ion exchange and ultrafiltration techniques. For cytotoxicity effects, L929, K562, HL60 and NB4 cell lines and MTT assay were used. A protein of 60000 Da of the purple fluid of A. dactylomela had antiproliferative effects on the cell lines it was maximally active at 0.5-1.5 microgram/ml on NB4 cell line. Therefore, the purple fluid of A. dactylomela has a novel antiproliferative agent.

Evaluation of Antiradical and Anti-Inflammatory Activities of Ethyl Acetate and Butanolic Subfractions of Agelanthus dodoneifolius (DC. Polhill & Wiens (Loranthaceae Using Equine Myeloperoxidase and Both PMA-Activated Neutrophils and HL-60 Cells

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Rainatou Boly

2015-01-01

Full Text Available The ethyl acetate and n-butanolic subfractions of Agelanthus dodoneifolius were investigated for their antioxidant and antimyeloperoxidase (MPO activities. The reactive oxygen species (ROS generation was assessed by lucigenin-enhanced chemiluminescence (CL and dichlorofluorescein- (DCF- induced fluorescence techniques from phorbol myristate acetate- (PMA- stimulated equine neutrophils and human myeloid cell line HL-60, respectively. In parallel, the effects of the tested subfractions were evaluated on the total MPO release by stimulated neutrophils and on the specific MPO activity by means of immunological assays. The results showed the potent activity of the butanolic subfraction, at least in respect of the chemiluminescence test (IC50 = 0.3±0.1 µg/mL and the ELISA and SIEFED assays (IC50 = 2.8±1.2 µg/mL and 1.3±1.0 µg/mL, respectively. However, the ethyl acetate subfraction was found to be the most potent in the DCF assay as at the highest concentration, DCF fluorescence intensity decreases of about 50%. Moreover, we demonstrated that the ethyl acetate subfraction was rich in catechin (16.51% while it was not easy to identify the main compounds in the butanolic subfraction using the UPLC-MS/MS technique. Nevertheless, taken together, our results provide evidence that Agelanthus dodoneifolius subfractions may represent potential sources of natural antioxidants and of antimyeloperoxidase compounds.

In vitro response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells exposed to 60Co at single fraction

International Nuclear Information System (INIS)

Andrade, Lidia Maria; Campos, Tarcisio Passos Ribeiro de; Leite, M.F.; Goes, A.M.

2005-01-01

Radiotherapy using gamma rays is a common modality of breast cancer treatment. The aim of this research is to investigate the biological response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells (PBMC) exposed in vitro to 60 Co irradiation at a single fraction of 10 Gy, 25 Gy and 50 Gy doses at 136,4 cGy.min -1 rate. Cells were irradiated at room temperature by the Theratron 80 radiotherapy system. Biological response was evaluated through cellular viability using MTT assay and nucleus damages visualized by Propidium Iodide assay and electrophoresis agarose gel after gamma irradiation. Nucleus damages induced by 60 Co irradiation were compared to damage caused by cell exposure to 10% methanol. The 50 Gy dose of irradiation did not stimulate nucleus damages at the same level as that affected by 10% methanol induction in the MDAMB-231. Further studies are necessary to understand these mechanisms in the MDAMB-231 human breast carcinoma cell line.(author)

Mitochondrial genome-knockout cells demonstrate a dual mechanism of action for the electron transport complex I inhibitor mycothiazole.

Science.gov (United States)

Meyer, Kirsten J; Singh, A Jonathan; Cameron, Alanna; Tan, An S; Leahy, Dora C; O'Sullivan, David; Joshi, Praneta; La Flamme, Anne C; Northcote, Peter T; Berridge, Michael V; Miller, John H

2012-04-01

Mycothiazole, a polyketide metabolite isolated from the marine sponge Cacospongia mycofijiensis, is a potent inhibitor of metabolic activity and mitochondrial electron transport chain complex I in sensitive cells, but other cells are relatively insensitive to the drug. Sensitive cell lines (IC(50) 0.36-13.8 nM) include HeLa, P815, RAW 264.7, MDCK, HeLa S3, 143B, 4T1, B16, and CD4/CD8 T cells. Insensitive cell lines (IC(50) 12.2-26.5 μM) include HL-60, LN18, and Jurkat. Thus, there is a 34,000-fold difference in sensitivity between HeLa and HL-60 cells. Some sensitive cell lines show a biphasic response, suggesting more than one mechanism of action. Mitochondrial genome-knockout ρ(0) cell lines are insensitive to mycothiazole, supporting a conditional mitochondrial site of action. Mycothiazole is cytostatic rather than cytotoxic in sensitive cells, has a long lag period of about 12 h, and unlike the complex I inhibitor, rotenone, does not cause G(2)/M cell cycle arrest. Mycothiazole decreases, rather than increases the levels of reactive oxygen species after 24 h. It is concluded that the cytostatic inhibitory effects of mycothiazole on mitochondrial electron transport function in sensitive cell lines may depend on a pre-activation step that is absent in insensitive cell lines with intact mitochondria, and that a second lower-affinity cytotoxic target may also be involved in the metabolic and growth inhibition of cells.

Rhein triggers apoptosis via induction of endoplasmic reticulum stress, caspase-4 and intracellular calcium in primary human hepatic HL-7702 cells

Energy Technology Data Exchange (ETDEWEB)

KoraMagazi, Arouna [Department of Clinical Pharmacy, China Pharmaceutical University, Nanjing, Jiangsu (China); Wang, Dandan [Department of Pharmacology, China Pharmaceutical University, Nanjing, Jiangsu (China); Yousef, Bashir; Guerram, Mounia [Jiangsu Key Laboratory of Drug Screening, China Pharmaceutical University, Nanjing, Jiangsu (China); Yu, Feng, E-mail: yufengcpu14@yahoo.com [Department of Clinical Pharmacy, China Pharmaceutical University, Nanjing, Jiangsu (China); Department of Pharmacology, China Pharmaceutical University, Nanjing, Jiangsu (China); Key Laboratory of Drug Quality Control and Pharmacovigilance, China Pharmaceutical University, Nanjing, Jiangsu (China)

2016-04-22

Rhein is an active component of rhubarb; a traditional Chinese medicine reported to induce apoptosis and cause liver toxicity. However, rhein's apoptotic-inducing effects, as well as its molecular mechanisms of action on hepatic cells need to be further explored. In the present study, rhein was found to trigger apoptosis in primary human hepatic HL-7702 cells as showed by annexin V/PI double staining assay and nuclear morphological changes demonstrated by Hoechst 33258 staining. Moreover, it was observed that the mechanism implicated in rhein-induced apoptosis was caspase-dependent, presumably via ER-stress associated pathways, as illustrated by up-regulation of glucose-regulated protein 78 (GRP 78), PKR-like ER kinase (PERK), C-Jun N-terminal kinase (JNK) and CCAAT/enhancer-binding protein homologous protein (CHOP). Meanwhile, caspase-4 as a hallmark of ER-stress, was also showed to be activated following by caspase-3 activation. Furthermore, rhein also promoted intracellular elevation of calcium that contributed in apoptosis induction. Interestingly, pre-treatment with calpain inhibitor I reduced the effects of rhein on apoptosis induction and JNK activation. These data suggested that rhein-induced apoptosis through ER-stress and elevated intracellular calcium level in HL-7702 cells. - Highlights: • Rhein triggers apoptotic cell death on primary human hepatic HL-7702 cells. • Rhein leads to caspase-4 activation in HL-7702 cells. • Rhein induces endoplasmic reticulum stress pathways in HL-7702 cells. • Rhein causes elevation of intracellular calcium concentrations in HL-7702 cells.

Effects of Ligusticum porteri (Osha) Root Extract on Human Promyelocytic Leukemia Cells

OpenAIRE

Nguyen, Khanh; Sparks, Jean; Omoruyi, Felix

2017-01-01

Background: Ligusticum porteri roots have been traditionally used in folk medicine, but the scientific basis is unclear. Objective: To investigate the cytotoxicity, antioxidant, and immunomodulatory effects of L. porteri root extract on human promyelocytic leukemia (HL-60) cells and H2O2-induced oxidative damaged HL-60 cells. Materials and Methods: HL-60 cells were incubated with different concentrations of root extract, and cells were harvested for viability assays on day 3 and 7. Cytokine l...

Synthetically derived bat influenza A-like viruses reveal a cell type- but not species-specific tropism.

Science.gov (United States)

Moreira, Étori Aguiar; Locher, Samira; Kolesnikova, Larissa; Bolte, Hardin; Aydillo, Teresa; García-Sastre, Adolfo; Schwemmle, Martin; Zimmer, Gert

2016-10-24

Two novel influenza A-like viral genome sequences have recently been identified in Central and South American fruit bats and provisionally designated "HL17NL10" and "HL18NL11." All efforts to isolate infectious virus from bats or to generate these viruses by reverse genetics have failed to date. Recombinant vesicular stomatitis virus (VSV) encoding the hemagglutinin-like envelope glycoproteins HL17 or HL18 in place of the VSV glycoprotein were generated to identify cell lines that are susceptible to bat influenza A-like virus entry. More than 30 cell lines derived from various species were screened but only a few cell lines were found to be susceptible, including Madin-Darby canine kidney type II (MDCK II) cells. The identification of cell lines susceptible to VSV chimeras allowed us to recover recombinant HL17NL10 and HL18NL11 viruses from synthetic DNA. Both influenza A-like viruses established a productive infection in MDCK II cells; however, HL18NL11 replicated more efficiently than HL17NL10 in this cell line. Unlike conventional influenza A viruses, bat influenza A-like viruses started the infection preferentially at the basolateral membrane of polarized MDCK II cells; however, similar to conventional influenza A viruses, bat influenza A-like viruses were released primarily from the apical site. The ability of HL18NL11 or HL17NL10 viruses to infect canine and human cells might reflect a zoonotic potential of these recently identified bat viruses.

Fludarabine-mediated circumvention of cytarabine resistance is associated with fludarabine triphosphate accumulation in cytarabine-resistant leukemic cells.

Science.gov (United States)

Yamamoto, Shuji; Yamauchi, Takahiro; Kawai, Yasukazu; Takemura, Haruyuki; Kishi, Shinji; Yoshida, Akira; Urasaki, Yoshimasa; Iwasaki, Hiromichi; Ueda, Takanori

2007-02-01

The combination of cytarabine (ara-C) with fludarabine is a common approach to treating resistant acute myeloid leukemia. Success depends on a fludarabine triphosphate (F-ara-ATP)-mediated increase in the active intracellular metabolite of ara-C, ara-C 5'-triphosphate (ara-CTP). Therapy-resistant leukemia may exhibit ara-C resistance, the mechanisms of which might induce cross-resistance to fludarabine with reduced F-ara-ATP formation. The present study evaluated the effect of combining ara-C and fludarabine on ara-C-resistant leukemic cells in vitro. Two variant cell lines (R1 and R2) were 8-fold and 10-fold more ara-C resistant, respectively, than the parental HL-60 cells. Reduced deoxycytidine kinase activity was demonstrated in R1 and R2 cells, and R2 cells also showed an increase in cytosolic 5'-nucleotidase II activity. Compared with HL-60 cells, R1 and R2 cells produced smaller amounts of ara-CTP. Both variants accumulated less F-ara-ATP than HL-60 cells and showed cross-resistance to fludarabine nucleoside (F-ara-A). R2 cells, however, accumulated much smaller amounts of F-ara-ATP and were more F-ara-A resistant than R1 cells. In HL-60 and R1 cells, F-ara-A pretreatment followed by ara-C incubation produced F-ara-ATP concentrations sufficient for augmenting ara-CTP production, thereby enhancing ara-C cytotoxicity. No potentiation was observed in R2 cells. Nucleotidase might preferentially degrade F-ara-A monophosphate over ara-C monophosphate, leading to reduced F-ara-ATP production and thereby compromising the F-ara-A-mediated potentiation of ara-C cytotoxicity in R2 cells. Thus, F-ara-A-mediated enhancement of ara-C cytotoxicity depended on F-ara-ATP accumulation in ara-C-resistant leukemic cells but ultimately was associated with the mechanism of ara-C resistance.

Evaluation of anticancer properties of a new α-methylene-δ-lactone DL-249 on two cancer cell lines

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Pomorska Dorota K.

2017-06-01

Full Text Available The anticancer activity of a new synthetic α-methylene-δ-lactone DL-249 was reported in leukemia HL-60 and breast cancer MCF-7 cells and compared with the activity of a natural α-methylene-γ-lactone from Tanacetum parthenium, parthenolide.

Anti-leukemic effect of a synthetic compound, (±) trans-dihydronarciclasine (HYU-01) via cell-cycle arrest and apoptosis in acute myeloid leukemia.

Science.gov (United States)

Kim, Seo Ju; Park, Hyun Ki; Kim, Ju Young; Yoon, Jin Sun; Kim, Eun Shil; Cho, Cheon-Gyu; Kim, Byoung Kook; Park, Byeong Bae; Lee, Young Yiul

2012-10-01

(±) trans-Dihydronarciclasine, isolated from Chinese medicinal plant Zephyranthes candida, has been shown to possess quite potent anti-tumoral effect against selected human cancer cell lines. However, little is known about the anti-tumoral effect of (±) trans-dihydronarciclasine in acute myeloid leukemia (AML). This study was performed to investigate the effect of a novel synthetic (±) trans-dihydronarciclasine (code name; HYU-01) in AML. The HYU-01 inhibited the proliferation of various AML cell lines including HL-60 as well as primary leukemic blasts in a dose-dependent manner. To investigate the mechanism of the anti-proliferative effect of HYU-01, cell-cycle analysis was attempted in HL-60 cells, resulting in G1 arrest. The expression levels of CDK2, CDK4, CDK6, cyclin E, and cyclin A were decreased in a time-dependent manner. In addition, HYU-01 up-regulated the expression of the p27, and markedly enhanced the binding of p27 with CDK2, 4, and 6, ultimately resulting in the decrease of their kinase activities. Furthermore, HYU-01 induced the apoptosis through the induction of proapoptotic molecules and reduction of antiapoptotic molecules in association with the activation of caspase-3, -8, and -9. These results suggest that HYU-01 may inhibit the proliferation of HL-60 cells, via apoptosis, as well as G1 block in association with the induction of p27. © 2012 The Authors APMIS © 2012 APMIS.

Probiotic assessment of Enterococcus durans 6HL and Lactococcus lactis 2HL isolated from vaginal microflora.

Science.gov (United States)

Nami, Yousef; Abdullah, Norhafizah; Haghshenas, Babak; Radiah, Dayang; Rosli, Rozita; Khosroushahi, Ahmad Yari

2014-08-01

Forty-five lactic acid bacteria (LAB) were isolated from the vaginal specimens of healthy fertile women, and the identities of the bacteria were confirmed by sequencing of their 16S rDNA genes. Among these bacteria, only four isolates were able to resist and survive in low pH, bile salts and simulated in vitro digestion conditions. Lactococcus lactis 2HL, Enterococcus durans 6HL, Lactobacillus acidophilus 36YL and Lactobacillus plantarum 5BL showed the best resistance to these conditions. These strains were evaluated further to assess their ability to adhere to human intestinal Caco-2 cells. Lactococcus lactis 2HL and E. durans 6HL were the most adherent strains. In vitro tests under neutralized pH proved the antimicrobial activity of both strains. Results revealed that the growth of Escherichia coli O26, Staphylococcus aureus and Shigella flexneri was suppressed by both LAB strains. The antibiotic susceptibility tests showed that these strains were sensitive to all nine antibiotics: vancomycin, tetracycline, ampicillin, penicillin, gentamicin, erythromycin, clindamycin, sulfamethoxazole and chloramphenicol. These data suggest that E. durans 6HL and Lactococcus lactis 2HL could be examined further for their useful properties and could be developed as new probiotics. © 2014 The Authors.

In vitro response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells exposed to {sup 60}Co at single fraction

Energy Technology Data Exchange (ETDEWEB)

Andrade, Lidia Maria; Campos, Tarcisio Passos Ribeiro de [Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil). Dept. de Engenharia Nuclear]. E-mail: lidia.andrade@unifenas.br; Leite, M.F. [Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil). Dept. de Fisiologia e Biofisica; Goes, A.M. [Universidade Federal de Minas Gerais, Belo Horizonte, MG (Brazil). Dept. de Bioquimica e Imunologia

2005-10-15

Radiotherapy using gamma rays is a common modality of breast cancer treatment. The aim of this research is to investigate the biological response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells (PBMC) exposed in vitro to {sup 60} Co irradiation at a single fraction of 10 Gy, 25 Gy and 50 Gy doses at 136,4 cGy.min{sup -1} rate. Cells were irradiated at room temperature by the Theratron 80 radiotherapy system. Biological response was evaluated through cellular viability using MTT assay and nucleus damages visualized by Propidium Iodide assay and electrophoresis agarose gel after gamma irradiation. Nucleus damages induced by {sup 60} Co irradiation were compared to damage caused by cell exposure to 10% methanol. The 50 Gy dose of irradiation did not stimulate nucleus damages at the same level as that affected by 10% methanol induction in the MDAMB-231. Further studies are necessary to understand these mechanisms in the MDAMB-231 human breast carcinoma cell line.(author)

In Vitro Effects of Bromoalkyl Phenytoin Derivatives on Regulated Death, Cell Cycle and Ultrastructure of Leukemia Cells.

Science.gov (United States)

Śladowska, Katarzyna; Opydo-Chanek, Małgorzata; Król, Teodora; Trybus, Wojciech; Trybus, Ewa; Kopacz-Bednarska, Anna; Handzlik, Jadwiga; Kieć-Kononowicz, Katarzyna; Mazur, Lidia

2017-11-01

To search for new antileukemic agents, the chemical structure of phenytoin was modified. A possible cytotoxic activity of three bromoalkyl phenytoin analogs, methyl 2-(1-(3-bromopropyl)-2,4-dioxo-5,5-diphenylimidazolidin-3-yl) propanoate (PH2), 1-(3-bromopropyl)-3-methyl-5,5-diphenylimidazolidine-2,4-dione (PH3) and 1-(4-bromobutyl)-3-methyl-5,5-diphenylimidazolidine-2,4-dione (PH4) on regulated cell death, the cell cycle and cell ultrastructure was assessed. The experiments were performed in vitro on HL-60 and U937 cells, using flow cytometry and electron microscopy methods. Application of PH2, PH3, and PH4 resulted in cell surface exposure of phosphatidylserine and plasma membrane impairment, caspase-8, -9, and -3/7 activation, dissipation of mitochondrial membrane potential, DNA breakage, cell-cycle disturbance and cell ultrastructural changes. In general, PH3 appeared to be the most active against the leukemia cells, and all bromoalkyl hydantoins, PH2-PH4, were more active in HL-60 cells than in U937 cells. The antileukemic activity of the bromoalkyl phenytoin analogs depended on the combination of N-hydantoin substituents and the human cell line used. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Bezafibrate induces a mitochondrial derangement in human cell lines: a PPAR-independent mechanism for a peroxisome proliferator.

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Scatena, R; Bottoni, P; Vincenzoni, F; Messana, I; Martorana, G E; Nocca, G; De Sole, P; Maggiano, N; Castagnola, M; Giardina, B

2003-11-01

Bezafibrate is a hypolipidemic drug that belongs to the group of peroxisome proliferators because it binds to peroxisome proliferator-activated receptors type alpha (PPARs). Peroxisome proliferators produce a myriad of extraperoxisomal effects, which are not necessarily dependent on their interaction with PPARs. An investigation on the peculiar activities of bezafibrate could clarify some of the molecular events and the relationship with the biochemical and pharmacological properties of this class of compounds. In this view, the human acute promyelocytic leukemia HL-60 cell line and human rabdomiosarcoma TE-671 cell line were cultured in media containing bezafibrate and a number of observations such as spectrophotometric analysis of mitochondrial respiratory chain enzymes, NMR metabolite determinations, phosphofructokinase enzymatic analysis, and differentiation assays were carried on. Bezafibrate induced a derangement of NADH cytochrome c reductase activity accompanied by metabolic alterations, mainly a shift to anaerobic glycolysis and an increase of fatty acid oxidation, as shown by NMR analysis of culture supernatants where acetate, lactate, and alanine levels increased. On the whole, the present results suggest a biochemical profile and a therapeutic role of this class of PPARs ligands more complex than those previously proposed.

Autologous stem cell transplantation for patients aged 60 years or older with refractory or relapsed classical Hodgkin's lymphoma: a retrospective analysis from the French Society of Bone Marrow Transplantation and Cell Therapies (SFGM-TC).

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Stamatoullas, A; Brice, P; Gueye, M S; Mareschal, S; Chevallier, P; Bouabdallah, R; Nguyenquoc, S; Francois, S; Turlure, P; Ceballos, P; Monjanel, H; Bourhis, J-H; Guillerm, G; Mohty, M; Biron, P; Cornillon, J; Belhadj, K; Bonmati, C; Dilhuydy, M-S; Huynh, A; Bernard, M; Chrétien, M-L; Peffault de Latour, R; Tilly, H

2016-07-01

This report retrospectively analyzed the outcome of 91 patients aged 60 years or older with refractory/relapsed (R/R) classical Hodgkin's lymphoma (cHL) who underwent autologous stem cell transplantation (ASCT) between 1992 and 2013 and were reported to the French Society of Bone Marrow Transplantation and Cell Therapies registry. The median age at transplant was 63 years. The majority of patients exhibited disease chemosensitivity to salvage treatment (57 complete responses, 30 partial responses, 1 progressive disease and 3 unknown). The most frequent conditioning regimen consisted of BCNU, cytarabine, etoposide, melphalan (BEAM) chemotherapy (93%). With a median follow-up of 54 months, 5-year estimates of overall survival (OS) and progression free survival (PFS) for the entire group were 67 and 54%, respectively. Despite the missing data, in univariate analysis, the number of salvage chemotherapy lines (1-2 versus ⩾3) significantly influenced the OS, unlike the other prognostic factors (stage III-IV at relapse, disease status before ASCT and negative positron emission tomography (PET) scan) encountered in younger patients. In spite of its limitations, this retrospective study with a long-term follow-up suggests that ASCT is a valid treatment option for chemosensitive R/R cHL in selected elderly patients, with an acceptable rate of toxicity.

CD25 targeted therapy of chemotherapy resistant leukemic stem cells using DR5 specific TRAIL peptide

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Jayaprakasam Madhumathi

2017-03-01

Full Text Available Chemotherapy resistant leukemic stem cells (LSCs are being targeted as a modern therapeutic approach to prevent disease relapse. LSCs isolated from methotrexate resistant side population (SP of leukemic cell lines HL60 and MOLT4 exhibited high levels of CD25 and TRAIL R2/DR5 which are potential targets. Recombinant immunotoxin conjugating IL2α with TRAIL peptide mimetic was constructed for DR5 receptor specific targeting of LSCs and were tested in total cell population and LSCs. IL2-TRAIL peptide induced apoptosis in drug resistant SP cells from cell lines and showed potent cytotoxicity in PBMCs derived from leukemic patients with an efficacy of 81.25% in AML and 100% in CML, ALL and CLL. IL2-TRAIL peptide showed cytotoxicity in relapsed patient samples and was more effective than TRAIL or IL2-TRAIL proteins. Additionally, DR5 specific IL2-TRAIL peptide was effective in targeting and killing LSCs purified from cell lines [IC50: 952 nM in HL60, 714 nM in MOLT4] and relapsed patient blood samples with higher efficacy (85% than IL2-TRAIL protein (46%. Hence, CD25 and DR5 specific targeting by IL2-TRAIL peptide may be an effective strategy for targeting drug resistant leukemic cells and LSCs.

Radiosensitivity of mesothelioma cell lines

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Haekkinen, A.M.; Laasonen, A.; Linnainmaa, K.; Mattson, K.; Pyrhoenen, S.

1996-01-01

The present study was carried out in order to examine the radiosensitivity of malignant pleural mesothelioma cell lines. Cell kinetics, radiation-induced delay of the cell cycle and DNA ploidy of the cell lines were also determined. For comparison an HeLa and a human foetal fibroblast cell line were simultaneously explored. Six previously cytogenetically and histologically characterized mesothelioma tumor cell lines were applied. A rapid tiazolyl blue microtiter (MTT) assay was used to analyze radiosensitivity and cell kinetics and DNA ploidy of the cultured cells were determined by flow cytometry. The survival fraction after a dose of 2 Gy (SF2), parameters α and β of the linear quadratic model (LQ-model) and mean inactivation dose (D MID ) were also estimated. The DNA index of four cell lines equaled 1.0 and two cell lines equaled 1.5 and 1.6. Different mesothelioma cell lines showed a great variation in radiosensitivity. Mean survival fraction after a radiation dose of 2 Gy (SF2) was 0.60 and ranged from 0.36 to 0.81 and mean α value was 0.26 (range 0.48-0.083). The SF2 of the most sensitive diploid mesothelioma cell line was 0.36: Less than that of the foetal fibroblast cell line (0.49). The survival fractions (0.81 and 0.74) of the two most resistant cell lines, which also were aneuploid, were equal to that of the HeLa cell line (0.78). The α/β ratios of the most sensitive cell lines were almost an order of magnitude greater than those of the two most resistant cell lines. Radiation-induced delay of the most resistant aneuploid cell line was similar to that of HeLa cells but in the most sensitive (diploid cells) there was practically no entry into the G1 phase following the 2 Gy radiation dose during 36 h. (orig.)

Radiosensitivity of mesothelioma cell lines

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Haekkinen, A.M. [Dept. of Oncology, Univ. Central Hospital, Helsinki (Finland); Laasonen, A. [Dept. of Pathology, Central Hospital of Etelae-Pohjanmaa, Seinaejoki (Finland); Linnainmaa, K. [Dept. of Industrial Hygiene and Toxicology, Inst. of Occupational Health, Helsinki (Finland); Mattson, K. [Dept. Pulmonary Medicine, Univ. Central Hospital, Helsinki (Finland); Pyrhoenen, S. [Dept. of Oncology, Univ. Central Hospital, Helsinki (Finland)

1996-10-01

The present study was carried out in order to examine the radiosensitivity of malignant pleural mesothelioma cell lines. Cell kinetics, radiation-induced delay of the cell cycle and DNA ploidy of the cell lines were also determined. For comparison an HeLa and a human foetal fibroblast cell line were simultaneously explored. Six previously cytogenetically and histologically characterized mesothelioma tumor cell lines were applied. A rapid tiazolyl blue microtiter (MTT) assay was used to analyze radiosensitivity and cell kinetics and DNA ploidy of the cultured cells were determined by flow cytometry. The survival fraction after a dose of 2 Gy (SF2), parameters {alpha} and {beta} of the linear quadratic model (LQ-model) and mean inactivation dose (D{sub MID}) were also estimated. The DNA index of four cell lines equaled 1.0 and two cell lines equaled 1.5 and 1.6. Different mesothelioma cell lines showed a great variation in radiosensitivity. Mean survival fraction after a radiation dose of 2 Gy (SF2) was 0.60 and ranged from 0.36 to 0.81 and mean {alpha} value was 0.26 (range 0.48-0.083). The SF2 of the most sensitive diploid mesothelioma cell line was 0.36: Less than that of the foetal fibroblast cell line (0.49). The survival fractions (0.81 and 0.74) of the two most resistant cell lines, which also were aneuploid, were equal to that of the HeLa cell line (0.78). The {alpha}/{beta} ratios of the most sensitive cell lines were almost an order of magnitude greater than those of the two most resistant cell lines. Radiation-induced delay of the most resistant aneuploid cell line was similar to that of HeLa cells but in the most sensitive (diploid cells) there was practically no entry into the G1 phase following the 2 Gy radiation dose during 36 h. (orig.).

Cytotoxicity and effect on GJIC of SiO2 nanoparticles in HL-7702 cells

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Pan Tao; Jin Minghua; Liu Xiaomei; Du Zhongjun; Zhou Xianqing; Huang Peili; Sun Zhiwei

2013-01-01

Objective: To study the cytotoxicity and effect on gap junction intracellular communication (GJIC) of SiO 2 nanoparticles in HL-7702 cells, and to provide experimental basis for toxicity assessment and the security applications of SiO 2 nanoparticles. Methods: Transmission electron microscope (TEM) was used to characterize two kinds of SiO 2 nanoparticles, verifying their size, dispersion and shape; dynamic light scattering (DLS) method was used to analyze the water dispersion and culture medium dispersion of the SiO 2 nanoparticles; MTT assay was carried out to examine the cytotoxicities of the two sizes SiO 2 nanoparticles on the cells; lactate dehydrogenase (LDH) release assay was performed to examine the integrity nano of the cell membrane; Scrape-loading and dye transfer assay was performed to examine the effect of SiO 2 nanoparticles on GJIC. Results: Based on the result of TEM, two kinds of SiO 2 nanoparticles were spherically shaped, uniformly sized and sporadically dispersed; the statistical analysis results showed the diameters of the two nanoparticles were (447.60±20.78) nm and (67.42±5.69) nm, respectively, thus they could be categorized as submicron scale and nano scale. The DLS method results manifested that the hydration nanoparticle sizes of the two SiO 2 nanoparticles were (684.37±18.76) nm, (128.31±7.64) nm in high purity water and (697.02±19.57) nm, (133.74±8.97) nm in RPMI-1640 solution, all the two nanoparticles were well dispersed without aggregation. MTT assay indicated that 24 h after treatment of SiO 2 nanoparticles, the cell viabilities were affected by both the size and the dose of the SiO 2 nanoparticles; the higher the dose was, the less viability the cells exhibited. Moreover, the nano scale particles inflicted more damage to the cells. LDH release assay indicated that the SiO 2 particles could also damage the cell membrane in a dose-dependent and size-dependent way. Scrape-loading and dye transfer assay indicated that the nano

c-Myc over-expression in Ramos Burkitt's lymphoma cell line predisposes to iron homeostasis disruption in vitro

International Nuclear Information System (INIS)

Habel, Marie-Eve; Jung, Daniel

2006-01-01

Burkitt's lymphoma is an aggressive B-cell neoplasm resulting from deregulated c-myc expression. We have previously shown that proliferation of Burkitt's lymphoma cell lines such as Ramos is markedly reduced by iron treatment. It has been shown that iron induces expression of c-myc which, owing to its transcriptional regulatory functions, regulates genes involved in iron metabolism. Transient enhancement of c-myc expression by iron could increase the expression of genes involved in iron incorporation, which could lead to an accumulation of intracellular free iron. Here, we have investigated whether cells with a high basal level of c-Myc were more likely to accumulate free iron. Our results suggest that the basal level of c-Myc in Ramos cells is twofold higher than what is seen in HL-60 cells. Moreover, in Ramos cells, where c-Myc is expressed at a high level, H-ferritin expression is down-regulated, transferrin receptor (CD71) expression is increased, and ferritin translation is inhibited. These modifications in iron metabolism, resulting from the strong basal expression of c-Myc, and amplified by iron addition, could lead to a disruption in homeostasis and consequently to growth arrest

Growth-inhibitory effects of the red alga Gelidium amansii on cultured cells.

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Chen, Yue-Hwa; Tu, Ching-Jung; Wu, Hsiao-Ting

2004-02-01

The objective of this study was to investigate the effects of Gelidium amansii, an edible red agar cultivated off the northeast coast of Taiwan, on the growth of two lines of cancer cells, murine hepatoma (Hepa-1) and human leukemia (HL-60) cells, as well as a normal cell line, murine embryo fibroblast cells (NIH-3T3). The potential role of G. amansii on the induction of apoptosis was also examined. The results indicated that all extracts from G. amansii, including phosphate-buffered saline (PBS) and methanol extracts from dried algae as well as the dimethyl sulfoxide (DMSO) extract from freeze-dried G. amansii agar, inhibited the growth of Hepa-1 and NIH-3T3 cells, but not the growth of HL-60 cells. Annexin V-positive cells were observed in methanol and DMSO extract-treated, but not PBS extract-treated Hepa-1 and NIH-3T3 cells, suggesting that the lipid-soluble extracts of G. amansii induced apoptosis. In summary, extracts of G. amansii from various preparations exhibited antiproliferative effects on Hepa-1 and NIH-3T3 cells, and apoptosis may play a role in the methanol and DMSO extract-induced inhibitory effects. However, the antiproliferative effects of PBS extracts was not through apoptosis. Moreover, the growth-inhibitory effects of G. amansii were not specific to cancer cells.

Two new meroterpenoids produced by the endophytic fungus Penicillium sp. SXH-65.

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Sun, Xinhua; Kong, Xianglan; Gao, Huquan; Zhu, Tianjiao; Wu, Guangwei; Gu, Qianqun; Li, Dehai

2014-08-01

Two new meroterpenoids, arisugacins I (1) and J (2), together with five known meroterpenoids including arisugacin B (3), arisugacin F (4), arisugacin G (5), territrem B (6) and territrem C (7) were isolated from an endophytic fungus Penicillium sp. SXH-65. Their structures were determined by extensive spectroscopic experiments and comparison with literature data. Their cytotoxicities were evaluated against Hela, HL-60 and K562 cell lines, and only 3 and 4 exhibited weak cytotoxicities against Hela, HL-60 and K562 cell lines with IC50 values ranging from 24 to 60 μM.

Variations in Mre11/Rad50/Nbs1 status and DNA damage-induced S-phase arrest in the cell lines of the NCI60 panel

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Eastman Alan

2011-05-01

Full Text Available Abstract Background The Mre11/Rad50/Nbs1 (MRN complex is a regulator of cell cycle checkpoints and DNA repair. Defects in MRN can lead to defective S-phase arrest when cells are damaged. Such defects may elicit sensitivity to selected drugs providing a chemical synthetic lethal interaction that could be used to target therapy to tumors with these defects. The goal of this study was to identify these defects in the NCI60 panel of cell lines and identify compounds that might elicit selective cytotoxicity. Methods We screened the NCI60 panel in search of cell lines that express low levels of MRN proteins, or that fail to arrest in S-phase in response to the topisomerase I inhibitor SN38. The NCI COMPARE program was used to discover compounds that preferentially target cells with these phenotypes. Results HCT116 cells were initially identified as defective in MRN and S phase arrest. Transfection with Mre11 also elevated Rad50 and Nbs1, and rescued the defective S-phase arrest. Cells of the NCI60 panel exhibited a large range of protein expression but a strong correlation existed between Mre11, Rad50 and Nbs1 consistent with complex formation determining protein stability. Mre11 mRNA correlated best with protein level suggesting it was the primary determinant of the overall level of the complex. Three other cell lines failed to arrest in response to SN38, two of which also had low MRN. However, other cell lines with low MRN still arrested suggesting low MRN does not predict an inability to arrest. Many compounds, including a family of benzothiazoles, correlated with the failure to arrest in S phase. The activity of benzothiazoles has been attributed to metabolic activation and DNA alkylation, but we note several cell lines in which sensitivity does not correlate with metabolism. We propose that the checkpoint defect imposes an additional mechanism of sensitivity on cells. Conclusions We have identified cells with possible defects in the MRN complex

Variations in Mre11/Rad50/Nbs1 status and DNA damage-induced S-phase arrest in the cell lines of the NCI60 panel

International Nuclear Information System (INIS)

Garner, Kristen M; Eastman, Alan

2011-01-01

The Mre11/Rad50/Nbs1 (MRN) complex is a regulator of cell cycle checkpoints and DNA repair. Defects in MRN can lead to defective S-phase arrest when cells are damaged. Such defects may elicit sensitivity to selected drugs providing a chemical synthetic lethal interaction that could be used to target therapy to tumors with these defects. The goal of this study was to identify these defects in the NCI60 panel of cell lines and identify compounds that might elicit selective cytotoxicity. We screened the NCI60 panel in search of cell lines that express low levels of MRN proteins, or that fail to arrest in S-phase in response to the topisomerase I inhibitor SN38. The NCI COMPARE program was used to discover compounds that preferentially target cells with these phenotypes. HCT116 cells were initially identified as defective in MRN and S phase arrest. Transfection with Mre11 also elevated Rad50 and Nbs1, and rescued the defective S-phase arrest. Cells of the NCI60 panel exhibited a large range of protein expression but a strong correlation existed between Mre11, Rad50 and Nbs1 consistent with complex formation determining protein stability. Mre11 mRNA correlated best with protein level suggesting it was the primary determinant of the overall level of the complex. Three other cell lines failed to arrest in response to SN38, two of which also had low MRN. However, other cell lines with low MRN still arrested suggesting low MRN does not predict an inability to arrest. Many compounds, including a family of benzothiazoles, correlated with the failure to arrest in S phase. The activity of benzothiazoles has been attributed to metabolic activation and DNA alkylation, but we note several cell lines in which sensitivity does not correlate with metabolism. We propose that the checkpoint defect imposes an additional mechanism of sensitivity on cells. We have identified cells with possible defects in the MRN complex and S phase arrest, and a series of compounds that may

FGF-23 dysregulates calcium homeostasis and electrophysiological properties in HL-1 atrial cells.

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Kao, Yu-Hsun; Chen, Yao-Chang; Lin, Yung-Kuo; Shiu, Rong-Jie; Chao, Tze-Fan; Chen, Shih-Ann; Chen, Yi-Jen

2014-08-01

Fibroblast growth factor (FGF)-23 is a key regulator of phosphate homeostasis. Higher FGF-23 levels are correlated with poor outcomes in cardiovascular diseases. FGF-23 can produce cardiac hypertrophy and increase intracellular calcium, which can change cardiac electrical activity. However, it is not clear whether FGF-23 possesses arrhythmogenic potential through calcium dysregulation. Therefore, the purposes of this study were to evaluate the electrophysiological effects of FGF-23 and identify the underlying mechanisms. Patch clamp, confocal microscope with Fluo-4 fluorescence, and Western blot analyses were used to evaluate the electrophysiological characteristics, calcium homeostasis and calcium regulatory proteins in HL-1 atrial myocytes with and without FGF-23 (10 and 25 ng/mL) incubation for 24 h. FGF-23 (25 ng/mL) increased L-type calcium currents, calcium transient and sarcoplasmic reticulum Ca(2+) contents in HL-1 cells. FGF-23 (25 ng/mL)-treated cells (n = 14) had greater incidences (57%, 17% and 15%, P calcium/calmodulin-dependent protein kinase IIδ and phospholamban (PLB) at threonine 17 but had similar phosphorylation extents of PLB at serine 16, total PLB and sarcoplasmic reticulum Ca(2+) -ATPase protein. Moreover, the FGF receptor inhibitor (PD173074, 10 nM), calmodulin inhibitor (W7, 5 μM) and phospholipase C inhibitor (U73122, 1 μM) attenuated the effects of FGF-23 on calcium/calmodulin-dependent protein kinase II phosphorylation. FGF-23 increases HL-1 cells arrhythmogenesis with calcium dysregulation through modulating calcium-handling proteins. © 2014 Stichting European Society for Clinical Investigation Journal Foundation.

The cytokine-dependent MUTZ-3 cell line as an in vitro model for the screening of contact sensitizers

International Nuclear Information System (INIS)

Azam, Philippe; Peiffer, Jean-Luc; Chamousset, Delphine; Tissier, Marie-Helene; Bonnet, Pierre-Antoine; Vian, Laurence; Fabre, Isabelle; Ourlin, Jean-Claude

2006-01-01

Langerhans cells (LC) are key mediators of contact allergenicity in the skin. However, no in vitro methods exist which are based on the activation process of LC to predict the sensitization potential of chemicals. In this study, we have evaluated the performances of MUTZ-3, a cytokine-dependent human monocytic cell line, in its response to sensitizers. First, we compared undifferentiated MUTZ-3 cells with several standard human cells such as THP-1, KG-1, HL-60, K-562, and U-937 in their response to the strong sensitizer DNCB and the irritant SDS by monitoring the expression levels of HLA-DR, CD54, and CD86 by flow cytometry. Only MUTZ-3 and THP-1 cells show a strong and specific response to sensitizer, while other cell lines showed very variable responses. Then, we tested MUTZ-3 cells against a wider panel of sensitizers and irritants on a broader spectrum of cell surface markers (HLA-DR, CD40, CD54, CD80, CD86, B7-H1, B7-H2, B7-DC). Of these markers, CD86 proved to be the most reliable since it detected all sensitizers, including benzocaine, a classical false negative in local lymph node assay (LLNA) but not irritants. We confirmed the MUTZ-3 response to DNCB by real-time PCR analysis. Taken together, our data suggest that undifferentiated MUTZ-3 cells may represent a valuable in vitro model for the screening of potential sensitizers

Up-Regulation of P21 Inhibits TRAIL-Mediated Extrinsic Apoptosis, Contributing Resistance to SAHA in Acute Myeloid Leukemia Cells

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Xing Wu

2014-08-01

Full Text Available Background/Aim: P21, a multifunctional cell cycle-regulatory molecule, regulates apoptotic cell death. In this study we examined the effect of altered p21 expression on the sensitivity of acute myeloid leukemia cells in response to HDAC inhibitor SAHA treatment and investigated the underlying mechanism. Methods: Stably transfected HL60 cell lines were established in RPMI-1640 with supplementation of G-418. Cell viability was measured by MTT assay. Western blot was applied to assess the protein expression levels of target genes. Cell apoptosis was monitored by AnnexinV-PE/7AAD assay. Results: We showed HL60 cells that that didn't up-regulate p21 expression were more sensitive to SAHA-mediated apoptosis than NB4 and U937 cells that had increased p21 level. Enforced expression of p21 in HL60 cells reduced sensitivity to SAHA and blocked TRAIL-mediated apoptosis. Conversely, p21 silencing in NB4 cells enhanced SAHA-mediated apoptosis and lethality. Finally, we found that combined treatment with SAHA and rapamycin down-regulated p21 and enhanced apoptosis in AML cells. Conclusion: We conclude that up-regulated p21 expression mediates resistance to SAHA via inhibition of TRAIL apoptotic pathway. P21 may serve as a candidate biomarker to predict responsiveness or resistance to SAHA-based therapy in AML patients. In addition, rapamycin may be an effective agent to override p21-mediated resistance to SAHA in AML patients.

Novel 5-Fluorouracil Derivatives: Synthesis and Cytotoxic Activity of 2-Butoxy-4-Substituted 5-Fluoropyrimidines

International Nuclear Information System (INIS)

Sun, Jian; Zhou, Wei; Hu, Weixiao; Shan, Shang; Zhang, Shijie; Li, Haibo

2013-01-01

Twenty two new 5-fluorouracil (5-FU) derivatives, 2-butoxy-4-substituted 5-fluoropyrimidines, were synthesized and characterized by IR, 1 H NMR, MS, HRMS. All compounds were preliminarily evaluated by MTT assay on human liver BEL-7402 cancer cell line in vitro. Ten compounds were selected to test their cytotoxic activity against A549, HL-60 and MCF-7 cancer cell lines in vitro. These compounds were more sensitive to BEL-7402 than other cell lines, particularly, cytotoxic activity of compounds 6b, 6d-f, 6p, 6s-u were in sub-micromolar scale. The highest cytotoxic potency against A549, HL-60 and MCF-7 was shown by 2-butoxy-4-chloro-5-fluoropyrimidine (5) with IC 50 values of 0.10, 1.66 and 0.59 μM, respectively. Compounds 6d and 6e were effective against MCF-7 with IC 50 9.73 μM and HL-60 with IC 50 8.83 μM, respectively

Honey bee venom combined with 1,25-dihydroxyvitamin D3as a highly efficient inducer of differentiation in human acute myeloid leukemia cells.

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Mohseni-Kouchesfahani, Homa; Nabioni, Mohammad; Khosravi, Zahra; Rahimi, Maryam

2017-01-01

Most cancer cells exhibit a defect in their capacity to mature into nonreplicating adult cells and existing in a highly proliferating state. Differentiation therapy by agents such as 1,25-dihydroxyvitamin D3(1,25-(OH)2 VD3) represents a useful approach for the treatment of cancer including acute myeloid leukemia. Human myeloid leukemia cell lines are induced to terminal differentiation into monocyte lineage by 1,25-(OH)2 VD3. However, usage of these findings in the clinical trials is limited by calcemic effects of 1,25-(OH)2 VD3. Attempts to overcome this problem have focused on a combination of low concentrations 1,25-(OH)2 VD3 with other compounds to induce differentiation of HL-60 cells. In this study, the effect of honey bee venom (BV) and 1,25-(OH)2 VD3, individually and in combination, on proliferation and differentiation of human myeloid leukemia HL-60 cells were assayed. In this in vitro study, toxic and nontoxic concentrations of BV and 1,25-(OH)2 VD3 were tested using Trypan blue stained cell counting and (3[4, 5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay. In addition, differentiation of cells was assayed using a Wright-Giemsa staining and nitroblue tetrazolium reduction test. Data were analyzed by a one-way analysis of the variance test using SPSS software. Our findings showed that both the BV and 1,25-(OH)2 VD3, in a dose and time-dependent manner, caused cell death at high concentrations and inhibited cell proliferation at lower concentrations. About 5 nM of 1,25-(OH)2 VD3 induced differentiation of HL-60 cells to monocytes after 72 h. 2.5 μg/ml of BV suppressed proliferation of HL-60 cells but had not any effects on their differentiation, whereas in combination with 5 nM of 1,25-(OH)2 VD3, it enhanced antiproliferative and differentiation potency of 1,25-(OH)2 VD3. These results indicate that BV potentiates the 1,25-(OH)2 VD3-induced HL-60 cell differentiation into monocytes.

The role of a new CD44st in increasing the invasion capability of the human breast cancer cell line MCF-7

International Nuclear Information System (INIS)

Fang, Xin Jian; Jiang, Hua; Zhao, Xv Peng; Jiang, Wei Mei

2011-01-01

CD44, a hyaluronan (HA) receptor, is a multistructural and multifunctional cell surface molecule involved in cell proliferation, cell differentiation, cell migration, angiogenesis, presentation of cytokines, chemokines and growth factors to the corresponding receptors, and docking of proteases at the cell membrane, as well as in signaling for cell survival. The CD44 gene contains 20 exons that are alternatively spliced, giving rise to many CD44 isoforms, perhaps including tumor-specific sequences. Reverse transcriptase polymerase chain reaction (RT-PCR) and Western blotting were used to detect CD44st mRNA and CD44 protein in sensitive MCF-7, Lovo, K562 and HL-60 cell lines as well as their parental counterparts, respectively. The full length cDNA encoding CD44st was obtained from the total RNA isolated from MCF-7/Adr cells by RT-PCR, and subcloned into the pMD19-T vector. The CD44st gene sequence and open reading frame were confirmed by restriction enzyme analysis and nucleotide sequencing, and then inserted into the eukaryotic expression vector pcDNA3.1. The pcDNA3.1-CD44st was transfected into MCF-7 cells using Lipofectamine. After transfection, the positive clones were obtained by G418 screening. The changes of the MMP-2 and MMP-9 genes and protein levels were detected by RT-PCR and gelatin zymography, respectively. The number of the cells penetrating through the artificial matrix membrane in each group (MCF-7, MCF-7+HA, MCF-7/neo, MCF-7/neo+HA, MCF-7/CD44st, MCF-7/CD44st+HA and MCF-7/CD44st+Anti-CD44+HA) was counted to compare the change of the invasion capability regulated by the CD44st. Erk and P-Erk were investigated by Western blotting to approach the molecular mechanisms of MMP-2 and MMP-9 expression regulated by the CD44st. Sensitive MCF-7, Lovo, K562 and HL-60 cells did not contain CD44st mRNA and CD44 protein. In contrast, the multidrug resistance MCF-7/Adr, Lovo/Adr, K562/Adr and HL-60/Adr cells expressed CD44st mRNA and CD44 protein. The CD44st m

Simvastatin Inhibits IL-5-Induced Chemotaxis and CCR3 Expression of HL-60-Derived and Human Primary Eosinophils.

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Fu, Chia-Hsiang; Tsai, Wan-Chun; Lee, Ta-Jen; Huang, Chi-Che; Chang, Po-Hung; Su Pang, Jong-Hwei

2016-01-01

IL-5-induced chemotaxis of eosinophils is an important feature of allergic airway inflammatory diseases. Simvastatin, a lipid lowering agent, has been shown to exhibit anti-inflammatory and anti-allergic effects. Our aim was to investigate the effect of simvastatin on IL-5-induced eosinophil chemotaxis and its regulatory mechanisms. Eosinophils were derived by treating HL-60 clone 15 (HC15) cells with butyric acid (BA) in an alkaline condition or through direct isolation from human peripheral blood. The expressions of CC chemokine receptor 3 (CCR3) and interleukin (IL)-5 receptors (IL5Rα and β) were analyzed using RT/real-time PCR. The granular proteins were stained using fast green. Eotaxin-induced chemotaxis was measured using a transwell migration assay. CCR3 protein expression was revealed by immunocytochemistry. An animal model of allergic rhinitis was established by challenging Sprague-Dawley® rats repeatedly with ovalbumin. Butyric acid significantly increased the expression of IL5Rα and IL5Rβ, CCR3 and granular proteins in HC15 cells, indicating the maturation of eosinophils (BA-E cells). IL-5 further enhanced the CCR3 expression at both the mRNA and protein levels and the eotaxin-induced chemotaxis of BA-E cells. Simvastatin inhibited the effects of IL-5 on BA-E cells, but not in the presence of mevalonate. Similar results were also exhibited in human primary eosinophils. In vivo animal studies further confirmed that oral simvastatin could significantly suppress the infiltration of eosinophils into turbinate tissues of allergic rats. Therefore, simvastatin was demonstrated to inhibit IL-5-induced CCR3 expression and chemotaxis of eosinophils mediated via the mevalonate pathway. We confirmed that simvastatin also reduced eosinophilic infiltration in allergic rhinitis.

Establishment and characterization of a novel Hodgkin lymphoma cell line, AM-HLH, carrying the Epstein-Barr virus genome integrated into the host chromosome.

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Hayashida, Masahiko; Daibata, Masanori; Tagami, Erika; Taguchi, Takahiro; Maekawa, Fumiyo; Takeoka, Kayo; Fukutsuka, Katsuhiro; Shimomura, Daiki; Hayashi, Takamasa; Iwatani, Yoshinori; Ohno, Hitoshi

2017-12-01

We describe the establishment and characterization of a cell line, AM-HLH, obtained from a patient with Epstein-Barr virus-positive (EBV + ) nodular sclerosis-type Hodgkin lymphoma (HL). The cells were positive for CD2 and CD30 and negative for CD15. The immunoglobulin heavy- and κ light-chain genes were rearranged. The karyotype was of the triploid range. Southern blotting using the EBV terminal repeat probe detected 3 hybridizing bands that were identical to those of the parental HL material. The cells expressed EBV-encoded RNAs as well as latent genes (EBNA1, EBNA2, LMP1, and LMP2A) and lytic genes (BZLF1 and BALF2). Fluorescence in situ hybridization (FISH) with the cosmid pJB8 clone containing a fragment of EBV DNA as a probe revealed multiple hybridization signals at a marker chromosome. Additional FISH using whole chromosome painting and centromere probes in combination with multicolor FISH determined that multiple EBV copies were clustered within the chromosome 20 materials of the marker chromosome. Culture supernatants of AM-HLH contained IL-10 as measured by the bead-based immunoassay. It is possible that an integrated EBV genome and cellular genes on chromosome 20 were coamplified, leading to the enhanced expression of genes involved in cell growth control. The AM-HLH cell line will be useful to clarify the role of cytokines in the development of EBV + HL. Copyright © 2016 John Wiley & Sons, Ltd.

Anti-Leukemic Activity of Shikonin: Role of ERP57 in Shikonin Induced Apoptosis in Acute Myeloid Leukemia

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Rachana Trivedi

2016-07-01

Full Text Available Background/Aims: ER-Stress and activation of unfolded protein response belong to the major factors involved in chemoresistance in cancer cells. In this study we investigated the effect of shikonin on the survival of acute myeloid leukemia cells and the role of ER-stress protein ERP57, a protein disulfide isomerase, in improvement of chemotherapy. Methods: Using MTT assay we studied cytotoxic effects of shikonin on HL-60 cells. The flow cytometry was adopted to examine the shikonin induced mode of cell death in HL-60 cells. The overall protein expression alteration resulting from shikonin treatment was investigated using proteomics methods. Western blotting was performed to quantify the alteration in protein expression in HL-60 after shikonin treatment. Silencing and overexpression studies were carried out to highlight the therapeutic role of ERP57 in shikonin effect on AML cells. Results: Shikonin induces apoptosis in HL-60 cells without significant effect on Primary cells from healthy volunteers. The apoptotic effect was dose and time dependent and was accompanied by strong alteration in cell proteome. Among the proteins targeted by shikonin, ERP57 was significantly downregulated in HL-60 after treatment. Compared to healthy control ERP57 was found to be highly expressed in AML cell line HL60 and was downregulated after shikonin treatment. Overexpression of ERP57 protected HL-60 from shikonin induced apoptosis, whereas knockdown of ERP57 expression resulted in increase in shikonin induced apoptosis. Conclusions: Our results demonstrate that ERP57 plays a crucial role in resistance towards shikonin induced apoptosis in AML cells. Targeting of ERP57 might offer a new therapeutic option for the treatment of acute myeloid leukemia.

Cardenolides and bufadienolide glycosides from Kalanchoe tubiflora and evaluation of cytotoxicity.

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Huang, Hui-Chi; Lin, Ming-Kuem; Yang, Hsin-Ling; Hseu, You-Cheng; Liaw, Chih-Chuang; Tseng, Yen-Hsueh; Tsuzuki, Minoru; Kuo, Yueh-Hsiung

2013-09-01

Two new cardenolides, kalantubolide A (1) and kalantubolide B (2), and two bufadienolide glycosides, kalantuboside A (3) and kalantuboside B (4), as well as eleven known compounds were isolated and characterized from the EtOH extract of Kalanchoe tubiflora. The structures of compounds were assigned based on 1D and 2D NMR spectroscopic analyses including HMQC, HMBC, and NOESY. Biological evaluation indicated that cardenolides (1-2) and bufadienolide glycosides (3-7) showed strong cytotoxicity against four human tumor cell lines (A549, Cal-27, A2058, and HL-60) with IC50 values ranging from 0.01 µM to 10.66 µM. Cardenolides (1-2) also displayed significant cytotoxicity toward HL-60 tumor cell line. In addition, compounds 3, 4, 5, 6, and 7 blocked the cell cycle in the G2/M-phase and induced apoptosis in HL-60 cells. Georg Thieme Verlag KG Stuttgart · New York.

Human primary erythroid cells as a more sensitive alternative in vitro hematological model for nanotoxicity studies: Toxicological effects of silver nanoparticles.

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Rujanapun, Narawadee; Aueviriyavit, Sasitorn; Boonrungsiman, Suwimon; Rosena, Apiwan; Phummiratch, Duangkamol; Riolueang, Suchada; Chalaow, Nipon; Viprakasit, Vip; Maniratanachote, Rawiwan

2015-12-01

Although immortalized cells established from cancerous cells have been widely used for studies in nanotoxicology studies, the reliability of the results derived from immortalized cells has been questioned because of their different characteristics from normal cells. In the present study, human primary erythroid cells in liquid culture were used as an in vitro hematological cell model for investigation of the nanotoxicity of silver nanoparticles (AgNPs) and comparing the results to the immortalized hematological cell lines HL60 and K562. The AgNPs caused significant cytotoxic effects in the primary erythroid cells, as shown by the decreased cell viability and induction of intracellular ROS generation and apoptosis, whereas they showed much lower cytotoxic and apoptotic effects in HL60 and K562 cells and did not induced ROS generation in these cell lines. Scanning electron microcopy revealed an interaction of AgNPs to the cell membrane in both primary erythroid and immortalized cells. In addition, AgNPs induced hemolysis in the primary erythroid cells in a dose-dependent manner, and transmission electron microcopy analysis revealed that AgNPs damaged the erythroid cell membrane. Taken together, these results suggest that human primary erythroid cells in liquid culture are a more sensitive alternative in vitro hematological model for nanotoxicology studies. Copyright © 2015 Elsevier B.V. All rights reserved.

Rhein Elicits In Vitro Cytotoxicity in Primary Human Liver HL-7702 Cells by Inducing Apoptosis through Mitochondria-Mediated Pathway

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Guy-Armel Bounda

2015-01-01

Full Text Available Objective. To study rhein-induced apoptosis signaling pathway and to investigate its molecular mechanisms in primary human hepatic cells. Results. Cell viability of HL-7702 cells treated with rhein showed significant decrease in dose-dependent manner. Following rhein treatment (25 μM, 50 μM, and 100 μM for 12 h, the detection of apoptotic cells was significantly analyzed by flow cytometry and nuclear morphological changes by Hoechst 33258, respectively. Fatty degeneration studies showed upregulation level of the relevant hepatic markers (P < 0.01. Caspase activities expressed significant upregulation of caspase-3, caspase-9, and caspase-8. Moreover, apoptotic cells by rhein were significantly inhibited by Z-LEHD-FMK and Z-DEVD-FMK, caspase-9 inhibitor, and caspase-3 inhibitor, respectively. Overproduction of reactive oxygen species, lipid peroxidation, and loss of mitochondrial membrane potential were detected by fluorometry. Additionally, NAC, a ROS scavenger, significantly attenuated rhein-induced oxidative damage in HL-7702 cells. Furthermore, real-time qPCR results showed significant upregulation of p53, PUMA, Apaf-1, and Casp-9 and Casp-3 mRNA, with no significant changes of Fas and Cytochrome-c. Immunoblotting revealed significant Cytochrome-c release from mitochondria into cytosol and no change in Fas expression. Conclusion. Taken together, these observations suggested that rhein could induce apoptosis in HL-7702 cells via mitochondria-mediated signal pathway with involvement of oxidative stress mechanism.

Effects of 60Co administration on early placental cells

International Nuclear Information System (INIS)

Honda, Jin

1979-01-01

The effects of 60 Co administration on early placental cells were studied. Placental tissue and embryo obtained by induced abortion (6 - 13 weeks gestational age) were placed in the minimal essential medium (MEM) and irradiated with various doses of 60 Co. After irradiation, the villi were cultured in a CO 2 incubater at 37 0 C. Cell growth process was observed every day with the phase-contrast microscope. Between 1 and 5 days epitheloid cells were dominant, but from about 7th day on fibroblastic cells dominated the culture. In placental tissue irradiated with 100, 200, 500 rad, fibroblastic cells began to grow earlier than in non-treated. Over 3000 rad 60 Co inhibited the growth of cells and a culture was impossible. For each dose, the tissue was incubated for various periods of time, exposed to tritiated thymidine for the last hour and autoradiogram was prepared by the dipping method. The labeling index of irradiated trophoblasts showed a significant decrease compared with controls. A chromosome study was made in irradiated in vitro cell lines of fetus and placenta. There was no significant difference between the two cell lines concerning the frequency of chromosome aberration, which tended to increase as the chromosome becomes longer. It is concluded that the trophoblast is highly radiosensitive and that irradiation early in pregnancy may damage DNA synthesis in the trophoblast, and induce abortion. (author)

Preparation and characterization of a novel nanobody against T-cell immunoglobulin and mucin-3 (TIM-3).

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Homayouni, Vida; Ganjalikhani-Hakemi, Mazdak; Rezaei, Abbas; Khanahmad, Hossein; Behdani, Mahdi; Lomedasht, Fatemeh Kazemi

2016-11-01

As T-cell immunoglobulin and mucin domain 3 (TIM-3) is an immune regulatory molecule; its blocking or stimulating could alter the pattern of immune response towards a desired condition. Based on the unique features of nanobodies, we aimed to construct an anti-TIM-3 nanobody as an appropriate tool for manipulating immune responses for future therapeutic purposes. We immunized a camel with TIM-3 antigen and then, synthesized a VHH phage: mid library from its B cell's transcriptome using nested PCR. Library selection against TIM-3antigen was performed in three rounds of panning. Using phage-ELISA, the most reactive colonies were selected for sub-cloning in soluble protein expression vectors. The Nanobody was purified and confirmed with a nickel-nitrilotriacetic acid (Ni-NTA) column, SDS-PAGE and Western blotting. A flowcytometric analysis was performed to analyze the binding and biologic activities of theTIM-3 specific nanobody with TIM-3 expressing HL-60 and HEK cell lines. Specific 15kD band representing for nanobody was observed on the gel and confirmed with Western blotting. The nanobody showed significant specific immune-reactivity against TIM-3 with a relatively high binding affinity. The nanobody significantly suppressed the proliferation of TIM-3 expressing HL-60 cell line. Finally, we successfully prepared a functional anti-humanTIM-3 specific nanobody with a high affinity and an anti-proliferative activity on an AML cell line in vitro.

Characterization of the formyl peptide chemotactic receptor appearing at the phagocytic cell surface after exposure to phorbol myristate acetate

International Nuclear Information System (INIS)

Gardner, J.P.; Melnick, D.A.; Malech, H.L.

1986-01-01

The biochemistry and subcellular source of new formyl peptide chemotactic receptor appearing at the human neutrophil and differentiated HL-60 (d-HL-60) cell surface after stimulation with phorbol myristate acetate (PMA) were examined. Formyl peptide receptor was analyzed by affinity labeling with formyl-norleu-leu-phe-norleu- [ 125 I]iodotyr-lys and ethylene glycol bis(succinimidyl succinate) followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and densitometric analysis of autoradiographs. PMA, a specific granule secretagogue, increases affinity labeling of formyl peptide receptors on the neutrophil surface by 100%, and on d-HL-60, which lack specific granule markers, by 20%. Papain treatment markedly reduces surface labeling of formyl peptide receptor in both neutrophils and d-HL-60, and results in the appearance of a lower m.w. membrane-bound receptor fragment. PMA stimulation of papain-treated cells increases uncleaved surface receptor on neutrophils by 400%, and on D-HL-60 by only 45%. This newly appearing receptor is the same apparent m.w. (55,000 to 75,000 for neutrophils; 62,000 to 80,000 for d-HL-60) and yields the same papain cleavage product as receptor on the surface of unstimulated cells. These observations suggest that specific granule membranes contain large amounts of formyl peptide receptor, which is biochemically identical to that found on the cell surface and can be mobilized to the cell surface with appropriate stimulation

Prevalence, birth incidence, and penetrance of von Hippel-Lindau disease (vHL) in Denmark

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Binderup, Marie Louise Mølgaard; Galanakis, Michael Carter Bisgaard; Budtz-Jørgensen, Esben

2017-01-01

the international diagnostic criteria. We found an overall penetrance of 87% at age 60 years. When considering only vHL patients who have not attended surveillance, 20% will still be asymptomatic at age 60 years. This should be considered in the context of genetic counselling, especially when assessing the risk...... of vHL in asymptomatic adult first-degree relatives who are often not genetically tested.European Journal of Human Genetics advance online publication, 14 December 2016; doi:10.1038/ejhg.2016.173....

Delayed K562 cell apoptosis promoted by cleaved LyGDI after 60Co γ-rays irradiation

International Nuclear Information System (INIS)

Sun Huali; Duan Weiming; Shao Yanyan; Xiao Hainan; Zhou Xinwen

2010-01-01

Objective: To elucidate the function and regulatory mechanism of LyGDI involved delayed cell death in the human K562 cells and HL-60 cells induced by 60 Co γ-rays. Methods: Erythrosine B cells staining was used to count the apoptosis rate. PI staining and flow cytometry were applied to check the cell cycle. The expression of LYGDI and Rac1 was resolved by Western blot by using monoclonal antibody of LyGDI and Rac1. The distribution of Rac1 protein in cells was observed with immunofluorescence by using the confocal microscope. Results: The K562 cells showed G 2 /M phase arrest and the percent age was 71.3%. The apoptosis rate was very low at early post-irradiation stage in the K562 cells. The apoptosis rate was 14% in the K562 cells at 24 h post-irradiation with 8 Gy of γ-rays, and delayed cell apoptosis was present. LyGDI was cleaved in the K562 cells irradiated by 4 Gy 60 Co γ-rays after 24 hours post-irradiation. The expression of Rac1 protein was not altered at all, but the distribution was changed in the irradiated cells while the Rac1 protein moved to cell membrane and a little in cell nucleus. The Rac1 was activated with the losing the binding affinity with the LyGDI. Conclusion: LyGDI could promote the delayed cell apoptosis, which is through the activation of the Rac1. (authors)

The natural cytokinin 2OH3MeOBAR induces cell death by a mechanism that is different from that of the „classical“ cytokinin ribosides

Czech Academy of Sciences Publication Activity Database

Voller, Jiří; Béres, T.; Zatloukal, M.; Kaminski, P. A.; Niemann, P.; Doležal, K.; Džubák, P.; Hajdúch, M.; Strnad, Miroslav

2017-01-01

Roč. 136, APR (2017), s. 156-164 ISSN 0031-9422 R&D Projects: GA MŠk(CZ) LO1204; GA MŠk(CZ) LO1304; GA MŠk(CZ) LM2015064; GA MŠk LM2015063; GA ČR GA14-19590S Institutional support: RVO:61389030 Keywords : myeloid-leukemia cells * kinetin-riboside * hl-60 cells * cancer-cells * in-vitro * apoptosis * gene * rcl * identification * lines * Phytohormone * Cytokinin * Leukemia * Cancer * Apoptosis * NCI60 panel Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Plant sciences, botany Impact factor: 3.205, year: 2016

Discrimination of bromodeoxyuridine labelled and unlabelled mitotic cells in flow cytometric bromodeoxyuridine/DNA analysis

DEFF Research Database (Denmark)

Jensen, P O; Larsen, J K; Christensen, I J

1994-01-01

Bromodeoxyuridine (BrdUrd) labelled and unlabelled mitotic cells, respectively, can be discriminated from interphase cells using a new method, based on immunocytochemical staining of BrdUrd and flow cytometric four-parameter analysis of DNA content, BrdUrd incorporation, and forward and orthogonal...... light scatter. The method was optimized using the human leukemia cell lines HL-60 and K-562. Samples of 10(5) ethanol-fixed cells were treated with pepsin/HCl and stained as a nuclear suspension with anti-BrdUrd antibody, FITC-conjugated secondary antibody, and propidium iodide. Labelled mitoses could...

Preparation and characterization of a novel nanobody against T-cell immunoglobulin and mucin-3 (TIM-3

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Vida Homayouni

2016-11-01

Full Text Available Objective(s: As T-cell immunoglobulin and mucin domain 3 (TIM-3 is an immune regulatory molecule; its blocking or stimulating could alter the pattern of immune response towards a desired condition. Based on the unique features of nanobodies, we aimed to construct an anti-TIM-3 nanobody as an appropriate tool for manipulating immune responses for future therapeutic purposes. Materials and Methods:We immunized a camel with TIM-3 antigen and then, synthesized a VHH phagemid library from its B cell’s transcriptome using nested PCR. Library selection against TIM-3antigen was performed in three rounds of panning. Using phage-ELISA, the most reactive colonies were selected for sub-cloning in soluble protein expression vectors. The Nanobody was purified and confirmed with a nickel-nitrilotriacetic acid (Ni-NTA column, SDS-PAGE and Western blotting. A flowcytometric analysis was performed to analyze the binding and biologic activities of theTIM-3 specific nanobody with TIM-3 expressing HL-60 and HEK cell lines. Results:Specific 15kD band representing for nanobody was observed on the gel and confirmed with Western blotting. The nanobody showed significant specific immune-reactivity against TIM-3 with a relatively high binding affinity. The nanobody significantly suppressed the proliferation of TIM-3 expressing HL-60 cell line. Conclusion: Finally, we successfully prepared a functional anti-humanTIM-3 specific nanobody with a high affinity and an anti-proliferative activity on an AML cell line in vitro.

FLI-1 Flightless-1 and LET-60 Ras control germ line morphogenesis in C. elegans

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Dentler William L

2008-05-01

Full Text Available Abstract Background In the C. elegans germ line, syncytial germ line nuclei are arranged at the cortex of the germ line as they exit mitosis and enter meiosis, forming a nucleus-free core of germ line cytoplasm called the rachis. Molecular mechanisms of rachis formation and germ line organization are not well understood. Results Mutations in the fli-1 gene disrupt rachis organization without affecting meiotic differentiation, a phenotype in C. elegans referred to here as the germ line morphogenesis (Glm phenotype. In fli-1 mutants, chains of meiotic germ nuclei spanned the rachis and were partially enveloped by invaginations of germ line plasma membrane, similar to nuclei at the cortex. Extensions of the somatic sheath cells that surround the germ line protruded deep inside the rachis and were associated with displaced nuclei in fli-1 mutants. fli-1 encodes a molecule with leucine-rich repeats and gelsolin repeats similar to Drosophila flightless 1 and human Fliih, which have been shown to act as cytoplasmic actin regulators as well as nuclear transcriptional regulators. Mutations in let-60 Ras, previously implicated in germ line development, were found to cause the Glm phenotype. Constitutively-active LET-60 partially rescued the fli-1 Glm phenotype, suggesting that LET-60 Ras and FLI-1 might act together to control germ line morphogenesis. Conclusion FLI-1 controls germ line morphogenesis and rachis organization, a process about which little is known at the molecular level. The LET-60 Ras GTPase might act with FLI-1 to control germ line morphogenesis.

An oncogenic axis of STAT-mediated BATF3 upregulation causing MYC activity in classical Hodgkin lymphoma and anaplastic large cell lymphoma.

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Lollies, A; Hartmann, S; Schneider, M; Bracht, T; Weiß, A L; Arnolds, J; Klein-Hitpass, L; Sitek, B; Hansmann, M-L; Küppers, R; Weniger, M A

2018-01-01

Classical Hodgkin lymphoma (cHL) and anaplastic large cell lymphoma (ALCL) feature high expression of activator protein-1 (AP-1) transcription factors, which regulate various physiological processes but also promote lymphomagenesis. The AP-1 factor basic leucine zipper transcription factor, ATF-like 3 (BATF3), is highly transcribed in cHL and ALCL; however, its functional importance in lymphomagenesis is unknown. Here we show that proto-typical CD30 + lymphomas, namely cHL (21/30) and primary mediastinal B-cell lymphoma (8/9), but also CD30 + diffuse large B-cell lymphoma (15/20) frequently express BATF3 protein. Mass spectrometry and co-immunoprecipitation established interactions of BATF3 with JUN and JUNB in cHL and ALCL lines. BATF3 knockdown using short hairpin RNAs was toxic for cHL and ALCL lines, reducing their proliferation and survival. We identified MYC as a critical BATF3 target and confirmed binding of BATF3 to the MYC promoter. JAK/STAT signaling regulated BATF3 expression, as chemical JAK2 inhibition reduced and interleukin 13 stimulation induced BATF3 expression in cHL lines. Chromatin immunoprecipitation substantiated a direct regulation of BATF3 by STAT proteins in cHL and ALCL lines. In conclusion, we identified STAT-mediated BATF3 expression that is essential for lymphoma cell survival and promoted MYC activity in cHL and ALCL, hence we recognized a new oncogenic axis in these lymphomas.

Involvement of Src tyrosine kinase and protein kinase C in the expression of macrophage migration inhibitory factor induced by H{sub 2}O{sub 2} in HL-1 mouse cardiac muscle cells

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Rao, F. [Department of Cardiology, Guangdong General Hospital, Guangdong Cardiovascular Institute, Guangdong Academy of Medical Sciences, Guangzhou (China); Research Center of Medical Sciences, Guangdong General Hospital, Guangzhou (China); Guangdong Academy of Medical Sciences, Guangzhou (China); Deng, C.Y. [Research Center of Medical Sciences, Guangdong General Hospital, Guangzhou (China); Guangdong Academy of Medical Sciences, Guangzhou (China); Zhang, Q.H.; Xue, Y.M. [Department of Cardiology, Guangdong General Hospital, Guangdong Cardiovascular Institute, Guangdong Academy of Medical Sciences, Guangzhou (China); Guangdong Academy of Medical Sciences, Guangzhou (China); Xiao, D.Z.; Kuang, S.J.; Lin, Q.X.; Shan, Z.X.; Liu, X.Y.; Zhu, J.N. [Research Center of Medical Sciences, Guangdong General Hospital, Guangzhou (China); Guangdong Academy of Medical Sciences, Guangzhou (China); Yu, X.Y. [Department of Cardiology, Guangdong General Hospital, Guangdong Cardiovascular Institute, Guangdong Academy of Medical Sciences, Guangzhou (China); Research Center of Medical Sciences, Guangdong General Hospital, Guangzhou (China); Guangdong Academy of Medical Sciences, Guangzhou (China); Wu, S.L. [Department of Cardiology, Guangdong General Hospital, Guangdong Cardiovascular Institute, Guangdong Academy of Medical Sciences, Guangzhou (China); Guangdong Academy of Medical Sciences, Guangzhou (China)

2013-09-06

Macrophage migration inhibitory factor (MIF), a pleiotropic cytokine, plays an important role in the pathogenesis of atrial fibrillation; however, the upstream regulation of MIF in atrial myocytes remains unclear. In the present study, we investigated whether and how MIF is regulated in response to the renin-angiotensin system and oxidative stress in atrium myocytes (HL-1 cells). MIF protein and mRNA levels in HL-1 cells were assayed using immunofluorescence, real-time PCR, and Western blot. The result indicated that MIF was expressed in the cytoplasm of HL-1 cells. Hydrogen peroxide (H{sub 2}O{sub 2}), but not angiotensin II, stimulated MIF expression in HL-1 cells. H{sub 2}O{sub 2}-induced MIF protein and gene levels increased in a dose-dependent manner and were completely abolished in the presence of catalase. H{sub 2}O{sub 2}-induced MIF production was completely inhibited by tyrosine kinase inhibitors genistein and PP1, as well as by protein kinase C (PKC) inhibitor GF109203X, suggesting that redox-sensitive MIF production is mediated through tyrosine kinase and PKC-dependent mechanisms in HL-1 cells. These results suggest that MIF is upregulated by HL-1 cells in response to redox stress, probably by the activation of Src and PKC.

An in vitro based investigation of the cytotoxic effect of water extracts of the Chinese herbal remedy LD on cancer cells

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Jones Lucy A

2009-09-01

Full Text Available Abstract Background Long Dan Xie Gan Wan (LD, a Chinese herbal remedy formulation, is traditionally used to treat a range of conditions, including gall bladder diseases, hepatitis, hyperthyroidism, migraines but it is not used for the management or treatment of cancer. However some of its herbal constituents, specifically Radix bupleuri, Radix scutellariae and Rhizoma alismatis have been shown to inhibit the growth of cancer cells. Thus, the aim of the study was to investigate the impact of LD on cancer cells in vitro. Methods HL60 and HT29 cancer cell lines were exposed to water extracts of LD (1:10, 1:50, 1:100 and/or 1:1000 prepared from a 3 mg/30 ml stock and for both cell lines growth, apoptotic induction, alterations in cell cycle characteristics and genotoxicity were investigated. The specificity of the action of LD on these cancer cell lines was also investigated by determining its effect on human peripheral blood lymphocytes. Preliminary chemical analysis was carried out to identify cytotoxic constituents of LD using HPLC and LCMS. Results LD was significantly cytotoxic to, and induced apoptosis in, both cell lines. Apoptotic induction appeared to be cell cycle independent at all concentrations of LD used (1:10, 1:50 and 1:100 for the HL60 cell lines and at 1:10 for the HT29 cell line. At 1:50 and 1:100 apoptotic induction by LD appeared to be cell cycle dependent. LD caused significant genotoxic damage to both cell lines compared to their respective controls. The specificity study showed that LD exerted a moderate cytotoxic action against non-proliferating and proliferating blood lymphocytes but not apoptosis. Chemical analysis showed that a number of fractions were found to exert a significant growth inhibitory effect. However, the molecular weights of compounds within these fractions did not correspond to those from the herbal constituents of LD. Conclusion It is possible that LD may have some chemotherapeutic potential. However

Interaction of leukemic cells with proteins of the extracellular matrix Interações de células leucêmicas com proteínas da matriz extracelular

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Adriana Rodrigues-Anjos

2004-01-01

Full Text Available The interaction of neoplastic cells with basement membrane molecules is the first step for the dissemination of tumor cells in vivo. Leukemic cells have a great ability to spread in the host, since cells are released from the bone marrow to the circulation. In this study we analysed whether CEM, U937, K562 and HL-60 cells were able to attach to different concentrations of laminin and/or fibronectin and/or type IV collagen. Attachment to type IV collagen was low, but it increased with the addition of laminin and occurred in all four leukemic cell lines. On the other hand, attachment to fibronectin was higher, but it decreased with the addition of laminin in the assays using U937 and HL-60 cells. The combination of type IV collagen and fibronectin was a good substratum for cellular attachment. However, the addition of laminin to this substratum impaired its attachment activity in U937, HL-60 and K562. These data suggest that laminin may control cellular attachment to the extracellular matrix during leukemic dissemination in hosts in different ways.A interação de células neoplásicas com moléculas da membrana basal é a primeira etapa para a disseminação, in vivo, de células tumorais in vivo. As células leucêmicas possuem grande capacidade de espraiamento e disseminação no organismo uma vez que as mesmas são liberadas da medula óssea para a circulação. Neste trabalho avaliamos a capacidade das linhagens celulares CEM, U937, K562 and HL-60 em aderirem a uma matriz extracelular constituída por diferentes concentrações de laminina e,ou fibronectina e sobre colágeno IV. A adesão de todas a linhagens leucêmicas a colágeno IV foi baixa, mas aumentou com a associação à laminina. Por outro lado, as células U937 e HL-60 apresentaram alta ligação à fibronectina porém, foi reduzida com a adição de laminina. A associação de colágeno IV e fibronectina possibilitou um bom substrato para a adesão celular. Entretanto, a adi

Compound A398, a novel podophyllotoxin analogue: cytotoxicity and induction of apoptosis in human leukemia cells.

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Alethéia L Silveira

Full Text Available Despite advances in oncology research, cancer is one of the leading causes of death worldwide. Thus, there is a demand for the development of more selective and effective antitumor agents. This study showed that A398, a novel podophyllotoxin analogue, was cytotoxic to the HT-29, MCF-7, MOLT-4 and HL-60 tumor cell lines, being less active in human peripheral blood mononuclear cells and normal cell lines FGH and IEC-6. Tests using the HepG2 lineage indicated that its metabolites do not contribute to its cytotoxicity. In the HL-60 cells, A398 induced apoptosis in a time and concentration-dependent manner, promoting mitochondrial depolarization, inhibition of Bcl-2, phosphatidylserine exposure, activation of caspases -8, -9 and -3, and DNA fragmentation. The production of reactive oxygen species does not seem to be a crucial event for the apoptotic process. Pretreatment with specific inhibitors of kinases ERK1/2, JNK and p38 resulted in an increased percentage of death induced by A398. These results indicate that the compound induced apoptosis through activation of intrinsic and extrinsic death pathways with the mechanism involving the inhibition of the MAPKs and Bcl-2. Taken together, our findings suggest that A398 has an anticancer potential, proving itself to be a candidate for preclinical studies.

Outcome of patients older than 60 years with classical Hodgkin lymphoma treated with front line ABVD chemotherapy: frequent pulmonary events suggest limiting the use of bleomycin in the elderly.

Science.gov (United States)

Stamatoullas, Aspasia; Brice, Pauline; Bouabdallah, Reda; Mareschal, Sylvain; Camus, Vincent; Rahal, Ilhem; Franchi, Patricia; Lanic, Hélène; Tilly, Hervé

2015-07-01

There is no standard of care in elderly classical Hodgkin lymphoma (cHL) patients. ABVD (doxorubicin, bleomycin, vinblastine, dacarbazine), the standard chemotherapy for younger patients, is also used in elderly patients but little is known about toxicity and efficacy. We retrospectively analysed 147 patients aged 60 years and over treated with ABVD in three French haematological centres. Treatment regimen modification was applied in 56 patients for toxicity or HL progression. Bleomycin was removed or reduced in 53 patients, mainly for pulmonary toxicity. Neither initial characteristics nor treatment characteristics were found to correlate with lung toxicity. One hundred and seventeen patients achieved a complete remission, 6 a partial remission, 16 had refractory disease and 8 were non-evaluable. Five-year overall survival was estimated at 67%. With a median follow-up of 58 months, 51 patients died and 14% of deaths were related to lung toxicity. Our study confirms the efficacy of ABVD in elderly patients even if results are inferior to those obtained in younger patients with the same regimen. ABVD can be proposed as front-line chemotherapy in selected elderly cHL patients. The frequency of pulmonary events leads us to propose to either reduce the dose of bleomycin or to remove it from the regimen. © 2015 John Wiley & Sons Ltd.

Radiosensitivity variations in human tumor cell lines exposed in vitro to p(66)/Be neutrons or 60Co γ-rays

International Nuclear Information System (INIS)

Slabbert, J.P.; Theron, T.; Serafin, A.; Jones, D.T.L.; Boehm, L.; Schmitt, G.

1996-01-01

Neutron therapy should be beneficial to patients with tumor types which are resistant to photons but relatively sensitive to high-LET radiation. In this work the potential therapeutic gain of a clinical neutron beam is evaluated by quantifying the variations in radiosensitivity of different cell lines to neutrons and photons. Different cell lines were exposed in vitro to p(66)/Be neutrons or 60 Co γ-rays. Micronuclei frequencies in binucleated cells and surviving fractions were determined for each cell type. Following exposure to either 1 or 1.5 Gy neutrons, micronuclei frequencies were significantly correlated with that observed for 2 Gy photons. A weak but significant correlation between the variation in neutron RBE values, determined from survival curve inactivation parameters and the mean inactivation doses for photon exposures, was also established. It is concluded that although neutron and photon sensitivities are related, the use of this high energy neutron source may constitute a potential therapeutic gain for tumor types that can be identified as very resistant to photons. Considering that a definitive oxygen gain factor has been established for this neutron beam the observed therapeutic gain is expected to be further enhanced in tumors where hypoxia protects cells from conventional radiation damage. (orig.) [de

HL-LHC kicker magnet (MKI)

CERN Multimedia

Brice, Maximilien

2018-01-01

HL-LHC kicker magnet (MKI): last vacuum test, preparation for transport to LHC transfer line in underground tunnel.The LHC injection kicker systems (MKIs) generate fast field pulses to inject the clockwise rotating beam at Point 2 and the anti-clockwise rotating beam at Point 8: there are eight MKI magnets installed in total. Each MKI magnet contains a high purity alumina tube: if an MKI magnet is replaced this tube requires conditioning with LHC beam: until it is properly conditioned, there can be high vacuum pressure due to the beam. This high pressure can also cause electrical breakdowns in the MKI magnets. A special coating (Cr2O3) has been applied to the inside of the alumina tube of an upgraded MKI magnet – this is expected to greatly reduce the pressure rise with beam. In addition, HL-LHC beam would result in excessive heating of the MKI magnets: the upgraded design includes modifications that will reduce heating, and move the power deposition to parts that will be easier to cool. Experience during 2...

Cell survival of human tumor cells compared with normal fibroblasts following 60Co gamma irradiation

International Nuclear Information System (INIS)

Lloyd, E.L.; Henning, C.B.; Reynolds, S.D.; Holmblad, G.L.; Trier, J.E.

1982-01-01

Three tumor cell lines, two of which were shown to be HeLa cells, were irradiated with 60 Co gamma irradiation, together with two cell cultures of normal human diploid fibroblasts. Cell survival was studied in three different experiments over a dose range of 2 to 14 gray. All the tumor cell lines showed a very wide shoulder in the dose response curves in contrast to the extremely narrow shoulder of the normal fibroblasts. In addition, the D/sub o/ values for the tumor cell lines were somewhat greater. These two characteristics of the dose response curves resulted in up to 2 orders of magnitude less sensitivity for cell inactivation of HeLa cells when compared with normal cells at high doses (10 gray). Because of these large differences, the extrapolation of results from the irradiation of HeLa cells concerning the mechanisms of normal cell killing should be interpreted with great caution

HSP90 is essential for Jak-STAT signaling in classical Hodgkin lymphoma cells

Directory of Open Access Journals (Sweden)

Kube Dieter

2009-07-01

Full Text Available Abstract In classical Hodgkin lymphoma (cHL chemotherapeutic regimens are associated with stagnant rates of secondary malignancies requiring the development of new therapeutic strategies. We and others have shown that permanently activated Signal Transducer and Activator of Transcription (STAT molecules are essential for cHL cells. Recently an overexpression of heat-shock protein 90 (HSP90 in cHL cells has been shown and inhibition of HSP90 seems to affect cHL cell survival. Here we analysed the effects of HSP90 inhibition by geldanamycin derivative 17-AAG or RNA interference (RNAi on aberrant Jak-STAT signaling in cHL cells. Treatment of cHL cell lines with 17-AAG led to reduced cell proliferation and a complete inhibition of STAT1, -3, -5 and -6 tyrosine phosphorylation probably as a result of reduced protein expression of Janus kinases (Jaks. RNAi-mediated inhibition of HSP90 showed similar effects on Jak-STAT signaling in L428 cHL cells. These results suggest a central role of HSP90 in permanently activated Jak-STAT signaling in cHL cells. Therapeutics targeting HSP90 may be a promising strategy in cHL and other cancer entities associated with deregulated Jak-STAT pathway activation.

Synergistic cytotoxic action of vitamin C and vitamin K3.

Science.gov (United States)

Zhang, W; Negoro, T; Satoh, K; Jiang, Y; Hashimoto, K; Kikuchi, H; Nishikawa, H; Miyata, T; Yamamoto, Y; Nakano, K; Yasumoto, E; Nakayachi, T; Mineno, K; Satoh, T; Sakagami, H

2001-01-01

We investigated the combination effect of sodium ascorbate (vitamin C) and menadione (vitamin K3) on the viability of various cultured cells. Human oral squamous cell carcinoma (HSC-2, HSC-3) and human promyelocytic leukemia (HL-60) cells were more sensitive to these vitamins as compared to normal cells (human gingival fibroblast HGF, human periodontal ligament fibroblast HPLF, human pulp cell HPC). The combination of vitamin C and vitamin K3 produced synergistic cytotoxicity against all these 6 cell lines. Treatment with vitamin C or vitamin K3, or their combination, induced internucleosomal DNA fragmentation only in HL-60 cells, but not in the oral tumor cell lines (HSC-2, HSC-3, HSG). ESR spectroscopy showed that vitamins C and K3 produce radicals under alkaline conditions and that the combination of these two vitamins synergistically enhanced their respective radical intensities.

Power supply for plasma generator of HL-1M neutral beam injector

International Nuclear Information System (INIS)

Wang Detai; Qian Jiamei; Lei Guangjiu; Shun Mengda; Jiang Shaofeng; Wang Enyao; Lu Xuejun; Yang Tiehai; Wang Xuehua; Zhao Zhimin; Hao Ming; Huang Jianrong; Yu Yanqiu; Cheng Baoqiang; Wu Zhige; Sheng Ning; Hu Qingtao

1999-01-01

The diagram of the HL-1M Neutral Beam Injector (NBI) and the power supply (PS) system is shown. The NBI consists of ion source, beam line and power supply system etc. The ion source includes plasma generator and three-electrode extraction system. The power supply for plasma generator consists of a filament PS, an arc PS and gas valve PS. Testing has shown that the PS for plasma generator of the HL-1M NBI has excellent stability and obtain good plasma heating effect

Design, Synthesis and Cytotoxic Activities of Novel Aliphatic Amino-Substituted Flavonoids

Directory of Open Access Journals (Sweden)

Guannan Liu

2013-11-01

Full Text Available A series of flavonoids 9a–f, 13b, 13d, 13e and 14a–f bearing diverse aliphatic amino moieties were designed, synthesized and evaluated for their cytotoxic activities against the ECA-109, A-549, HL-60, and PC-3 cancer cell lines. Most of the compounds exhibited moderate to good activities. The structure-activity relationships were studied, revealing that the chalcone skeleton is the most preferable for cytotoxic activities. Chalcone 9d was the most promising compound due to its high potency against the examined cancer cell lines (its IC50 values against ECA-109, A549, HL-60 and PC-3 cells were 1.0, 1.5, 0.96 and 3.9 μM, respectively.

Development of a USB-based multi-channel time division scaler for HL-2A

International Nuclear Information System (INIS)

Liang Ping

2008-01-01

HL-2A is China's first Tokamak device with divertor configuration. Mastering the process and plasma parameter changes with time are of great significance to achieve controlled nuclear fusion. In the recent upgrading of HL-2A, for which a higher and faster electronic equipment was required, we developed a new type USB multi-channel time division scaler for HL-2A including functions: USB interface, PC graphical user interface, simultaneously calibrating more than five channel signals, optional time division spacing 2-50 ms, count rate up to over 2 MHz, accessing all the information 20 s after a activated signal, and processing data and displaying off-line. (authors)

Hans Pöhl - Estlandssvenskarnas hövding = Hans Pöhl - rannarootslaste eestvõitleja / Torkel Jansson

Index Scriptorium Estoniae

Jansson, Torkel, 1947-

2011-01-01

Raamatututvustus: Hans Pöhl - Estlandssvenskarnas hövding : en biografi över Hans Pöhl (1876-1930), estlandssvenskarnas främste företrädare och ledare = Hans Pöhl - rannarootslaste eestvõitleja : Hans Pöhli (1876-1930), Eesti rootslaste vaimse liidri ja valgustaja elulugu. (Stockholm ; Tallinn, 2010)

The preparation and biological characterization of a new HL91-derivative for hypoxic imaging on stroke mice

Energy Technology Data Exchange (ETDEWEB)

Hsia, C.-C. [Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, Taiwan (China); Institute of Nuclear Energy Research, Taiwan (China); Huang, F.-L.; Lin, C.-H.; Shen, L.-H. [Institute of Nuclear Energy Research, Taiwan (China); Wang, H.-E., E-mail: hewang@ym.edu.t [Department of Biomedical Imaging and Radiological Sciences, National Yang-Ming University, Taiwan (China)

2010-09-15

Aim: {sup 99m}Tc-HL91 (Prognox, GE-Healthcare) was the first nonnitro-aryl-based radiotracer for evaluating hypoxic fraction in neoplasm, stroke and myocardium infarction regions. However, the high hydrophilicity of {sup 99m}Tc-HL91 might hamper its penetration into cells. In this study, we prepared a new ligand 4,4,11,11-tetramethyl- 5,10-diazatetradecane- 3,12-dionedioxime (HL91-ET) with higher lipophilicity but structurally similar compared with that of HL91. The chemical and biological characterizations of {sup 99m}Tc-HL91-ET as a scintigraphic probe for hypoxia were performed with a stroke-bearing mouse model. Materials and Methods: HL91-ET was synthesized and formulated with stannous chloride and buffer to afford kits. After mixing with {sup 99m}Tc-pertechnetate, {sup 99m}Tc-HL91-ET can be prepared in high yield and high radiochemical purity (both >96%). The partition coefficient of {sup 99m}Tc-HL91-ET was determined in n-octanol/PBS system. Cellular uptake assays under normoxic and hypoxic conditions were performed in an oxygen-controlled CO{sub 2} incubator. Brain stroke in the mouse model was induced by the electrocautery of the middle cerebral artery. After intravenous injection of {sup 99m}Tc-HL91-ET into the Balb/c mouse suffering brain stroke, small-animal SPECT images were acquired at designated time points and autoradiography of the brain slides was conducted. Parallel studies of {sup 99m}Tc-HL91 were also conducted at the same conditions for comparison. Results: The higher partition coefficient of {sup 99m}Tc-HL91-ET (0.294{+-}0.007) indicated higher lipophlicity compared with that of {sup 99m}Tc-HL91 (0.089{+-}0.005). The {sup 99m}Tc-HL91-ET preparation was stable at ambient temperature for 24 h. Cellular uptake assay showed that {sup 99m}Tc-HL91-ET was less selectively retained in hypoxic cells than {sup 99m}Tc-HL91. The target-to-normal brain ratios derived from the autoradiograms of the brains of stroke mice were 1.31{+-}0.02 and 17

The preparation and biological characterization of a new HL91-derivative for hypoxic imaging on stroke mice

International Nuclear Information System (INIS)

Hsia, C.-C.; Huang, F.-L.; Lin, C.-H.; Shen, L.-H.; Wang, H.-E.

2010-01-01

Aim: 99m Tc-HL91 (Prognox, GE-Healthcare) was the first nonnitro-aryl-based radiotracer for evaluating hypoxic fraction in neoplasm, stroke and myocardium infarction regions. However, the high hydrophilicity of 99m Tc-HL91 might hamper its penetration into cells. In this study, we prepared a new ligand 4,4,11,11-tetramethyl- 5,10-diazatetradecane- 3,12-dionedioxime (HL91-ET) with higher lipophilicity but structurally similar compared with that of HL91. The chemical and biological characterizations of 99m Tc-HL91-ET as a scintigraphic probe for hypoxia were performed with a stroke-bearing mouse model. Materials and Methods: HL91-ET was synthesized and formulated with stannous chloride and buffer to afford kits. After mixing with 99m Tc-pertechnetate, 99m Tc-HL91-ET can be prepared in high yield and high radiochemical purity (both >96%). The partition coefficient of 99m Tc-HL91-ET was determined in n-octanol/PBS system. Cellular uptake assays under normoxic and hypoxic conditions were performed in an oxygen-controlled CO 2 incubator. Brain stroke in the mouse model was induced by the electrocautery of the middle cerebral artery. After intravenous injection of 99m Tc-HL91-ET into the Balb/c mouse suffering brain stroke, small-animal SPECT images were acquired at designated time points and autoradiography of the brain slides was conducted. Parallel studies of 99m Tc-HL91 were also conducted at the same conditions for comparison. Results: The higher partition coefficient of 99m Tc-HL91-ET (0.294±0.007) indicated higher lipophlicity compared with that of 99m Tc-HL91 (0.089±0.005). The 99m Tc-HL91-ET preparation was stable at ambient temperature for 24 h. Cellular uptake assay showed that 99m Tc-HL91-ET was less selectively retained in hypoxic cells than 99m Tc-HL91. The target-to-normal brain ratios derived from the autoradiograms of the brains of stroke mice were 1.31±0.02 and 17.47±0.10 (n=3), respectively, at 2 h post injection of 99m Tc-HL91-ET and 99m Tc-HL91

Ageing studies of resistive Micromegas detectors for the HL-LHC

CERN Document Server

Galán, J; Ferrer-Ribas, E; Giganon, A; Giomataris, I; Herlant, S; Jeanneau, F; Peyaud, A; Schune, Ph; Alexopoulos, T; Byszewski, M; Iakovidis, G; Iengo, P; Ntekas, K; Leontsinis, S; de Oliveira, R; Tsipolitis, Y; Wotschack, J

2013-01-01

Resistive-anode Micromegas detectors are in development since several years, in an effort to solve the problem of sparks when working in high flux and high radiations environment like in the HL-LHC (ten times the luminosity of the LHC). They have been chosen as one of the technologies that will be part of the ATLAS New Small Wheel project (forward muon system). An ageing study is mandatory to assess their capabilities to handle the HL-LHC environment on a long-term period. A prototype has been exposed to several types of irradiations (X-rays, cold neutrons, 60 Co gammas) up to an equivalent HL-LHC time of more than five years without showing any degradation of the performances in terms of gain and energy resolution. Beam test studies took place in October 2012 to assess the tracking performances (efficiency, spatial resolution,...). Results of ageing studies and beam test performances are reported in this paper.

Radiation recall and radiosensitisation with alkylating agents

Energy Technology Data Exchange (ETDEWEB)

Kellie, S.J.; Plowman, P.N.; Malpas, J.S.

1987-05-16

This brief note records the results of experiments with Adriamycin, 1,2:5,6 dianhydrodulcit and hydroxyurea combined with an endogenous substance inhibiting myeloid proliferation selectively against target cells taken from normal rat, mouse or human haemopoietic lines, spontaneous human leukaemias and the HL-60 established promyelocytic cell line.

In vitro response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells exposed to 60Co at single fraction

Directory of Open Access Journals (Sweden)

Lídia Maria Andrade

2005-10-01

Full Text Available Radiotherapy using gamma rays is a common modality of breast cancer treatment. The aim of this research is to investigate the biological response of the human breast cancer cell line MDAMB-231 and human peripheral blood mononuclear cells (PBMC exposed in vitro to 60 Co irradiation at a single fraction of 10 Gy, 25 Gy and 50 Gy doses at 136,4 cGy.min-1 rate. Cells were irradiated at room temperature by the Theratron 80 radiotherapy system. Biological response was evaluated through cellular viability using MTT assay and nucleus damages visualized by Propidium Iodide assay and electrophoresis agarose gel after gamma irradiation. Nucleus damages induced by 60Co irradiation were compared to damage caused by cell exposure to 10% methanol. The 50 Gy dose of irradiation did not stimulate nuclus damages at the same level as that affected by 10% methanol induction in the MDAMB-231. Further studies are necessary to understand these mechanisms in the MDAMB-231 human breast carcinoma cell line.Radioterapia utilizando radiação gama é uma modalidade comum no tratamento do câncer de mama. A proposta deste estudo é investigar a resposta biológica in vitro da linhagem celular MDAMB-231 de câncer de mama humano e células do sangue periférico humano (PBMC expostas à irradiação pelo Co60 em frações simples de 10Gy, 25Gy e 50Gy e 136,4cGy min-1 rate. As células foram irradiadas a temperatura ambiente usando o equipamento de radioterapia Theratron 80 radiotherapy system. A resposta biológica, após irradiação gama, foi avaliada através do ensaio do MTT para viabilidade celular e o do ensaio com Iodeto de Propídio para visualização do dano nuclear, além da eletroforese em gel de agarose. Os danos nucleares induzidos pelo Co60 foram comparados aos danos causados pela exposição das células à solução de metanol a 10%. Nós observamos que a dose de 50Gy não estimulou a mesma quantidade de danos nucleares que a solução de metanol a 10% nas c

The CNGRC-GG-D(KLAKLAK)2 peptide induces a caspase-independent, Ca2+-dependent death in human leukemic myeloid cells by targeting surface aminopeptidase N/CD13

OpenAIRE

Bouchet, Sandrine; Tang, Ruoping; Fava, Fanny; Legrand, Ollivier; Bauvois, Brigitte

2015-01-01

The CD13 antigen's binding site for the Asn-Gly-Arg (NGR) motif enables NGR-containing chemotherapeutic drugs to be delivered to CD13-positive tumours. Human CD13-positive acute myeloid leukemia (AML) cells proliferate abnormally and escape death. Here, we show that the CNGRC-GG-D(KLAKLAK)2 peptide induces death in AML cell lines (U937, THP-1, NB4, HL-60) and primary blood cells from AML patients. Cell death was characterized as a caspase-independent mechanism, without DNA fragmentation, but ...

Cytotoxic effect of betulinic acid and betulinic acid acetate isolated ...

African Journals Online (AJOL)

Cytotoxic effect of betulinic acid and betulinic acid acetate isolated from Melaleuca cajuput on human myeloid leukemia (HL-60) cell line. ... The cytotoxic effect of betulinic acid (BA), isolated from Melaleuca cajuput a Malaysian plant and its four synthetic derivatives were tested for their cytotoxicity in various cell line or ...

Antioxidant Activities and Anti-Cancer Cell Proliferation Properties of Natsuhaze (Vaccinium oldhamii Miq.), Shashanbo (V. bracteatum Thunb.) and Blueberry Cultivars.

Science.gov (United States)

Tsuda, Hirotoshi; Kunitake, Hisato; Kawasaki-Takaki, Ryoko; Nishiyama, Kazuo; Yamasaki, Masao; Komatsu, Haruki; Yukizaki, Chizuko

2013-02-15

Antioxidants are abundant in blueberries, and while there are many studies concerning the bioactive compound of fruit, it is only recently that the wild Vaccinium species has attracted attention for their diverse and abundant chemical components. The aim of this study was to investigate the bioactive compounds of blueberry cultivars and wild species found in Japan. Among the five extracts of the Vaccinium species, Natsuhaze (Vaccinium oldhamii Miq.) was found to be the most effective at inhibiting the growth of HL-60 human leukemia cells in vitro. Although all ethanol extracts showed a growth inhibitory effect on HL-60 cells, the degree of the effects differed among the species. The extract of Natsuhaze induced apoptotic bodies and nucleosomal DNA fragmentation in the HL-60 cells. Of the extracts tested, that of Natsuhaze contained the largest amount of total polyphenols and showed the greatest antioxidant activity, but the anthocyanin content of Natsuhaze was similar to that of rabbiteye blueberry (V. virgatum Ait.). The results showed that total polyphenols contributed to the high antioxidant activity and growth inhibitory effect on HL-60 human leukemia cells of Natsuhaze extract.

Antioxidant Activities and Anti-Cancer Cell Proliferation Properties of Natsuhaze (Vaccinium oldhamii Miq., Shashanbo (V. bracteatum Thunb. and Blueberry Cultivars

Directory of Open Access Journals (Sweden)

Hirotoshi Tsuda

2013-02-01

Full Text Available Antioxidants are abundant in blueberries, and while there are many studies concerning the bioactive compound of fruit, it is only recently that the wild Vaccinium species has attracted attention for their diverse and abundant chemical components. The aim of this study was to investigate the bioactive compounds of blueberry cultivars and wild species found in Japan. Among the five extracts of the Vaccinium species, Natsuhaze (Vaccinium oldhamii Miq. was found to be the most effective at inhibiting the growth of HL-60 human leukemia cells in vitro. Although all ethanol extracts showed a growth inhibitory effect on HL-60 cells, the degree of the effects differed among the species. The extract of Natsuhaze induced apoptotic bodies and nucleosomal DNA fragmentation in the HL-60 cells. Of the extracts tested, that of Natsuhaze contained the largest amount of total polyphenols and showed the greatest antioxidant activity, but the anthocyanin content of Natsuhaze was similar to that of rabbiteye blueberry (V. virgatum Ait.. The results showed that total polyphenols contributed to the high antioxidant activity and growth inhibitory effect on HL-60 human leukemia cells of Natsuhaze extract.

Apoptotic effect of a novel kefir product, PFT, on multidrug-resistant myeloid leukemia cells via a hole-piercing mechanism

Science.gov (United States)

GHONEUM, MAMDOOH; GIMZEWSKI, JAMES

2014-01-01

We examined the apoptotic effect of a novel Probiotics Fermentation Technology (PFT) kefir grain product; PFT is a natural mixture composed primarily of Lactobacillus kefiri P-IF, a specific strain of L. kefiri with unique growth characteristics. The aim of this study was to examine the apoptotic effect of PFT on human multidrug-resistant (MDR) myeloid leukemia (HL60/AR) cells in vitro and explore the mechanistic approach underlying its effect. HL60/AR cells were cultured with PFT (0.6–5.0 mg/ml) for 3 days. The apoptotic effect of PFT was assessed through examination of percent apoptosis, caspase 3 activation, Bcl-2 expression levels and changes in mitochondrial membrane potential (MMP). PFT induced apoptosis in HL60/AR cells in a dose-dependent manner which was maximal at 67.5% for 5 mg/ml. Induction of apoptosis was associated with activation of caspase 3, decreased expression of Bcl-2 and decreased polarization of MMP. In addition, PFT showed a unique characteristic of piercing holes in HL60/AR cells, as indicated by AFM studies. This hole induction may be responsible for the apoptotic effect on cancer cells. These results suggest that PFT may act as a potential therapy for the treatment of MDR leukemia. PMID:24430613

Pilot Peter Hoag and HL-10

Science.gov (United States)

1969-01-01

Air Force Major Peter Hoag stands in front of the HL-10 Lifting Body. Maj. Hoag joined the HL-10 program in 1969 and made his first glide flight on June 6, 1969. He made a total of 8 flights in the HL-10. They included the fastest lifting-body flight, which reached Mach 1.861 on Feb. 18, 1970. The HL-10 was one of five heavyweight lifting-body designs flown at NASA's Flight Research Center (FRC--later Dryden Flight Research Center), Edwards, California, from July 1966 to November 1975 to study and validate the concept of safely maneuvering and landing a low lift-over-drag vehicle designed for reentry from space. Northrop Corporation built the HL-10 and M2-F2, the first two of the fleet of 'heavy' lifting bodies flown by the NASA Flight Research Center. The contract for construction of the HL-10 and the M2-F2 was $1.8 million. 'HL' stands for horizontal landing, and '10' refers to the tenth design studied by engineers at NASA's Langley Research Center, Hampton, Va. After delivery to NASA in January 1966, the HL-10 made its first flight on Dec. 22, 1966, with research pilot Bruce Peterson in the cockpit. Although an XLR-11 rocket engine was installed in the vehicle, the first 11 drop flights from the B-52 launch aircraft were powerless glide flights to assess handling qualities, stability, and control. In the end, the HL-10 was judged to be the best handling of the three original heavy-weight lifting bodies (M2-F2/F3, HL-10, X-24A). The HL-10 was flown 37 times during the lifting body research program and logged the highest altitude and fastest speed in the Lifting Body program. On Feb. 18, 1970, Air Force test pilot Peter Hoag piloted the HL-10 to Mach 1.86 (1,228 mph). Nine days later, NASA pilot Bill Dana flew the vehicle to 90,030 feet, which became the highest altitude reached in the program. Some new and different lessons were learned through the successful flight testing of the HL-10. These lessons, when combined with information from it's sister ship, the M2

HL-A27 and anterior uveitis.

Science.gov (United States)

Woodrow, J C; Mapstone, R; Anderson, J; Usher, N

1975-09-01

HL-A types were determined in 90 successive patients with non-granulomatous uveitis. Fifty-one were HL-A27 positive (55.7%) compared to 8.2% of controls. Of 16 patients with ankylosing spondylitis, 13 were HL-A27 positive, as were two patients with a history of Reiter's syndrome. Twenty-eight patients were HL-A27 positive but had no evidence of rheumatic disease. The findings are discussed in relation to the possible pathogenesis of uveitis.

Genipin-induced inhibition of uncoupling protein-2 sensitizes drug-resistant cancer cells to cytotoxic agents.

Directory of Open Access Journals (Sweden)

Ryan J Mailloux

2010-10-01

Full Text Available Uncoupling protein-2 (UCP2 is known to suppress mitochondrial reactive oxygen species (ROS production and is employed by drug-resistant cancer cells to mitigate oxidative stress. Using the drug-sensitive HL-60 cells and the drug-resistant MX2 subline as model systems, we show that genipin, a UCP2 inhibitor, sensitizes drug-resistant cells to cytotoxic agents. Increased MX2 cell death was observed upon co-treatment with genipin and different doses of menadione, doxorubicin, and epirubicin. DCFH-DA fluorimetry revealed that the increase in MX2 cell death was accompanied by enhanced cellular ROS levels. The drug-induced increase in ROS was linked to genipin-mediated inhibition of mitochondrial proton leak. State 4 and resting cellular respiratory rates were higher in the MX2 cells in comparison to the HL-60 cells, and the increased respiration was readily suppressed by genipin in the MX2 cells. UCP2 accounted for a remarkable 37% of the resting cellular oxygen consumption indicating that the MX2 cells are functionally reliant on this protein. Higher amounts of UCP2 protein were detected in the MX2 versus the HL-60 mitochondria. The observed effects of genipin were absent in the HL-60 cells pointing to the selectivity of this natural product for drug-resistant cells. The specificity of genipin for UCP2 was confirmed using CHO cells stably expressing UCP2 in which genipin induced an ∼22% decrease in state 4 respiration. These effects were absent in empty vector CHO cells expressing no UCP2. Thus, the chemical inhibition of UCP2 with genipin sensitizes multidrug-resistant cancer cells to cytotoxic agents.

Hypoxia-induced redox alterations and their correlation with 99mTc-MIBI and 99mTc-HL-91 uptake in colon cancer cells

International Nuclear Information System (INIS)

Abrantes, Ana Margarida; Silva Serra, Maria Elisa; Goncalves, Ana Cristina; Rio, Joana; Oliveiros, Barbara; Laranjo, Mafalda; Rocha-Gonsalves, Antonio Manuel; Sarmento-Ribeiro, Ana Bela; Filomena Botelho, Maria

2010-01-01

Colorectal cancer is one of the most common malignancies in the Western world and is an example of a solid tumour in which hypoxia is a common feature and develops because of the inability of the vascular system to supply adequate amounts of oxygen to growing tumours. Hypoxia effects on tumour cell biology can be detected and characterized using different methods. The use of imaging with γ-emitting radionuclides to detect hypoxic tissue was first suggested by Chapman in 1979 [N Engl J Med 301 (1979) 1429-1432]. 99m Tc-4,9-diaza-3,3,10,10-tetramethyldodecan-2,11-dione dioxime, also known as 99m Tc-HL-91, has been among the most studied hypoxia markers. The objective of this study was to correlate the uptake of 99m Tc-HL-91 and 99m Tc-MIBI in colon cancer cells under normoxic and hypoxic conditions and to compare this information with some parameters such as oxidative stress and mitochondrial dysfunction of the cells analyzed by flow cytometry. Our results show that the in vitro 99m Tc-HL-91 uptake is higher in hypoxic conditions, which is confirmed by the decreased uptake of 99m Tc-MIBI. Flow cytometry results demonstrate that hypoxic conditions used are not enough to induce cellular death, but are responsible for the alterations in the intracellular redox environment, namely, increase of ROS production, proteic pimonidazol-derived adduct formation and alteration in the mitochondrial membrane permeability. Therefore, these results confirm that 99m Tc-HL-91 is a radiopharmaceutical with favourable characteristics for detecting hypoxia.

Selections from 2016: Gaps in HL Tau's Protoplanetary Disk

Science.gov (United States)

Kohler, Susanna

2016-12-01

Editors note:In these last two weeks of 2016, well be looking at a few selections that we havent yet discussed on AAS Nova from among the most-downloaded paperspublished in AAS journals this year. The usual posting schedule will resume after the AAS winter meeting.Gas Gaps in the Protoplanetary Disk Around the Young Protostar HL TauPublished March 2016The dust (left) and gas (right) emission from HL Tau show that the gaps in its disk match up. [Yen et al. 2016]Main takeaway:At the end of last year, the Atacama Large Millimeter/Submillimeter Array released some of its first data including a spectacular observation of a dusty protoplanetary disk around the young star HL Tau. In this follow-up study, a team led by Hsi-Wei Yen (Academia Sinica Institute of Astronomy and Astrophysics, Taiwan) analyzed the ALMA data and confirmed the presence of two gaps in the gas of HL Taus disk, at radii of 28 and 69 AU.Why its interesting:The original ALMA image of HL Taus disk suggests the presence of gaps in disk, but scientists werent sure if they were caused by effects like gravitational instabilities or dust clumping, or if the gaps were created by the presence of young planets. Yen and collaborators showed that gaps in the disks gas line up with gaps in its dust, supporting the model in which these gaps have been carved out by newly formed planets.Added intrigue:The evidence for planets in this disk came as a bit of a surprise, since it was originally believed that it takes tens of millions of years to form planets from the dust of protoplanetary disks but HL Tau is only a million years old. These observations therefore suggest that planets start to form much earlier than we thought.CitationHsi-Wei Yen et al 2016 ApJL 820 L25. doi:10.3847/2041-8205/820/2/L25

An ageing study of resistive micromegas for the HL-LHC environment

CERN Document Server

Galan, J.; Ferrer-Ribas, E.; Giganon, A.; Giomataris, I.; Herlant, S.; Jeanneau, F.; Peyaud, A.; Schune, Ph; Alexopoulos, T.; Byszewski, M.; Iakovidis, G.; Iengo, P.; Ntekas, K.; Leontsinis, S.; de Oliveira, R.; Tsipolitis, Y.; Wotschack, J.

2013-01-01

Resistive-anode micromegas detectors are in development since several years, in an effort to solve the problem of sparks when working at high flux and high ionizing radiation like in the HL-LHC (up to ten times the luminosity of the LHC). They have been chosen as one of the technologies that will be part of the ATLAS New Small Wheel project (forward muon system). An ageing study is mandatory to assess their capabilities to handle the HL-LHC environment on a long-term period. A prototype has been exposed to several types of irradiation (X-rays, cold neutrons, $^{60}$Co gammas and alphas) above the equivalent charge produced at the detector in five HL-LHC running years without showing any degradation of the performances in terms of gain and energy resolution. This study has been completed with the characterization of the tracking performances in terms of efficiency and spatial resolution, verifying the compatibility of results obtained with both resistive micromegas detectors, irradiated and non-irradiated one.

Production, Structural Elucidation, and In Vitro Antitumor Activity of Trehalose Lipid Biosurfactant from Nocardia farcinica Strain.

Science.gov (United States)

Christova, Nelly; Lang, Siegmund; Wray, Victor; Kaloyanov, Kaloyan; Konstantinov, Spiro; Stoineva, Ivanka

2015-04-01

The objective of this study was to isolate and identify the chemical structure of a biosurfactant produced by Nocardia farcinica strain BN26 isolated from soil, and evaluate its in vitro antitumor activity on a panel of human cancer cell lines. Strain BN26 was found to produce glycolipid biosurfactant on n-hexadecane as the sole carbon source. The biosurfactant was purified using medium-pressure liquid chromatography and characterized as trehalose lipid tetraester (THL) by nuclear magnetic resonance spectroscopy and mass spectrometry. Subsequently, the cytotoxic effects of THL on cancer cell lines BV-173, KE-37 (SKW-3), HL-60, HL-60/DOX, and JMSU-1 were evaluated by MTT assay. It was shown that THL exerted concentration-dependent antiproliferative activity against the human tumor cell lines and mediated cell death by the induction of partial oligonucleosomal DNA fragmentation. These findings suggest that THL could be of potential to apply in biomedicine as a therapeutic agent.

6-Nitro-2-(3-hydroxypropyl-1H-benz[de]isoquinoline-1,3-dione, a potent antitumor agent, induces cell cycle arrest and apoptosis

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Singh Shashank K

2010-12-01

Full Text Available Abstract Background Anticancer activities of several substituted naphthalimides (1H-benz[de]isoquinoline-1,3-diones are well documented. Some of them have undergone Phase I-II clinical trials. Presently a series of ten N-(hydroxyalkyl naphthalimides (compounds 1a-j were evaluated as antitumor agents. Methods Compounds 1a-j were initially screened in MOLT-4, HL-60 and U-937 human tumor cell lines and results were compared with established clinical drugs. Cytotoxicities of compounds 1d and 1i were further evaluated in a battery of human tumor cell lines and in normal human peripheral blood mononuclear cells. Cell cycle analysis of compound 1i treated MOLT-4 cells was studied by flow cytometry. Its apoptosis inducing effect was carried out in MOLT-4 and HL-60 cells by flow cytometry using annexin V-FITC/PI double staining method. The activities of caspase-3 and caspase-6 in MOLT-4 cells following incubation with compound 1i were measured at different time intervals. Morphology of the MOLT-4 cells after treatment with 1i was examined under light microscope and transmission electron microscope. 3H-Thymidine and 3H-uridine incorporation in S-180 cells in vitro following treatment with 8 μM concentration of compounds 1d and 1i were studied. Results 6-Nitro-2-(3-hydroxypropyl-1H-benz[de]isoquinoline-1,3-dione (compound 1i, has exhibited maximum activity as it induced significant cytotoxicity in 8 out of 13 cell lines employed. Interestingly it did not show any cytotoxicity against human PBMC (IC50 value 273 μM. Cell cycle analysis of compound 1i treated MOLT-4 cells demonstrated rise in sub-G1 fraction and concomitant accumulation of cells in S and G2/M phases, indicating up-regulation of apoptosis along with mitotic arrest and/or delay in exit of daughter cells from mitotic cycle respectively. Its apoptosis inducing effect was confirmed in flow cytometric study in MOLT-4 and the action was mediated by activation of both caspase 3 and 6. Light and

Synthesis and Cytotoxic Evaluation of a Series of 2-Amino-Naphthoquinones against Human Cancer Cells

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Thiago A. P. de Moraes

2014-08-01

Full Text Available The cytotoxicity of a series of aminonaphthoquinones resulting from the reaction of suitable aminoacids with 1,4-naphthoquinone was assayed against SF-295 (glioblastoma, MDAMB-435 (breast, HCT-8 (colon, HCT-116 (colon, HL-60 (leukemia, OVCAR-8 (ovarian, NCI-H358M (bronchoalveolar lung carcinoma and PC3-M (prostate cancer cells and also against PBMC (peripheral blood mononuclear cells. The results demonstrated that all the synthetic aminonaphthoquinones had relevant cytotoxic activity against all human cancer lines used in this experiment. Five of the compounds showed high cytotoxicity and selectivity against all cancer cell lines tested (IC50 = 0.49 to 3.89 µg·mL−1. The title compounds were less toxic to PBMC, since IC50 was 1.5 to eighteen times higher (IC50 = 5.51 to 17.61 µg·mL−1 than values shown by tumour cell lines. The mechanism of cell growth inhibition and structure–activity relationships remains as a target for future investigations.

Establishment and conventional cytogenetic characterization of three gastric cancer cell lines.

Science.gov (United States)

Leal, Mariana Ferreira; Martins do Nascimento, José Luiz; da Silva, Carla Elvira Araújo; Vita Lamarão, Maria Fernanda; Calcagno, Danielle Queiroz; Khayat, André Salim; Assumpção, Paulo Pimentel; Cabral, Isabel Rosa; de Arruda Cardoso Smith, Marília; Burbano, Rommel Rodríguez

2009-11-01

Gastric cancer is the fourth most frequent type of cancer and the second most frequent cause of cancer mortality worldwide. Only a modest number of gastric carcinoma cell lines have been isolated thus far. Here we describe the establishment and cytogenetic characterization of three new gastric cancer cell lines obtained from primary gastric adenocarcinoma (ACP02 and ACP03) and cancerous ascitic fluid (AGP01) of individuals from northern Brazil. ACP02, ACP03, and AGP01 cell lines are presently in the 60th passage. The cell lines grew in a disorganized single layer with some agglomerations and heterogeneous divisions (bipolar and multipolar). All cell lines exhibited a composite karyotype with several clonal chromosome alterations. Trisomy 8 was the most frequent alteration. Chromosome 8 aneusomy was confirmed by fluorescence in situ hybridization. All cell lines also exhibited trisomy 7 and deletion of chromosome arm 17p. These results suggest that, although frequent chromosome alterations are commonly observed due to culture process, the ACP02, ACP03, and AGP01 cell lines and primary gastric cancer from individuals of northern Brazil share genetic alterations, supporting use of these cell lines as a model of gastric carcinogenesis in this population.

KHYG-1, a model for the study of enhanced natural killer cell cytotoxicity.

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Suck, Garnet; Branch, Donald R; Smyth, Mark J; Miller, Richard G; Vergidis, Joanna; Fahim, Soad; Keating, Armand

2005-10-01

To compare the cytotoxicity of KHYG-1 with other natural killer (NK)/NK T-cell lines and identify molecules that may be associated with enhanced cytotoxicity, thereby eventually leading to improved NK cell-mediated cancer immunotherapy. NK/NK T-cell lines KHYG-1, NK-92, YT, and SNT-8 were compared with a novel flow cytometric cytotoxicity assay under different culture conditions. Transcription, expression, and phosphorylation studies were performed using polymerase chain reaction sequence-specific primers, reverse transcription polymerase chain reaction, immunoblotting, and flow cytometry. KHYG-1 is a highly cytotoxic cell line, exceeding the cytolytic capacity of the other cell lines against K562. KHYG-1 is also highly cytotoxic against the leukemia cell lines EM2, EM3, and HL60. The novel activation receptor NKp44 and its adaptor, DAP12, NKG2D, and constitutively phosphorylated ERK2 may be associated with the enhanced cytotoxicity of KHYG-1. This cell line most likely mediates cytolysis by granzyme M (but not granzymes A and B) together with perforin, which is constitutively fully cleaved to the 60-kD form, in contrast to the other cell lines. KHYG-1 is a valuable model for the study of enhanced cytotoxicity by NK cells. In addition to the activation of NKp44, KHYG-1 may induce apoptosis of tumor cells by the newly described granzyme M/perforin pathway. Targeted modifications of effector molecules demonstrated in this model could generate NK cells with even greater killing ability that may be particularly attractive for clinical application. Moreover, our demonstration of greater cytotoxicity of KHYG-1 versus NK-92 cells, already in clinical trials, suggests a direct therapeutic role for KHYG-1.

Esters of Quinoxaline 1ˏ4-Di-N-oxide with Cytotoxic Activity on Tumor Cell Lines Based on NCI-60 Panel

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Rivera, Gildardo; Ahmad Shah, Syed Shoaib; Arrieta-Baez, Daniel; Palos, Isidro; Mongue, Antonio; Sánchez-Torres, Luvia Enid

2017-01-01

Quinoxalines display diverse and interesting pharmacological activities as antibacterial, antiviral, antiparasitic and anticancer agents. Particularly, their 1ˏ4-di-N-oxide derivatives have proved to be cytotoxic agents that are active under hypoxic conditions as that of solid tumours. A new series of quinoxaline 1ˏ4-di-N-oxide substitutes at 7-position with esters group were synthetized and characterized by infrared (IR), proton nuclear magnetic resonance (1H-NMR), spectroscopy, and elemental analysis. Seventeen derivatives (M1-M3, E1-E8, P1-P3 and DR1-DR3) were selected and evaluated for antitumor activities using the NCI-60 human tumor cell lines screen. Results showed that E7, P3 and E6 were the most active compounds against the cell lines tested. Substitutions at 7-position with esters group not necessarily affect the biological activity, but the nature of the esters group could exert an influence on the selectivity. Additionally, substitutions at 2-position influenced the cytotoxic activity of the compounds. PMID:29201086

Synthesis and Antitumor Activity of 3-Methyl-4-oxo-3,4-dihydroimidazo [5,1-d][1,2,3,5]tetrazine-8-carboxylates and -carboxamides

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Lin-Xiang Zhao

2010-12-01

Full Text Available Seventeen novel 3-methyl-4-oxo-3,4-dihydroimidazo[5,1-d][1,2,3,5]tetrazine-8-carboxylate and -carboxamide derivatives were synthesized and evaluated for their growth inhibition in seven human solid tumor and a human leukemia HL-60 cell lines. Compound IVa showed more activity than the other compounds and the positive control temozolomide. In the presence of 40 mg/mL of IVa, the survival rate of all tested tumor cells was less than 10%. Esters displayed more potent antitumour activity than amides and temozolomide against HL-60 cells. These compounds also exhibited considerably enhanced water-solubility.

Curcumin and Its Analogue Induce Apoptosis in Leukemia Cells and Have Additive Effects with Bortezomib in Cellular and Xenograft Models

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L. I. Nagy

2015-01-01

Full Text Available Combination therapy of bortezomib with other chemotherapeutics is an emerging treatment strategy. Since both curcumin and bortezomib inhibit NF-κB, we tested the effects of their combination on leukemia cells. To improve potency, a novel Mannich-type curcumin derivative, C-150, was synthesized. Curcumin and its analogue showed potent antiproliferative and apoptotic effects on the human leukemia cell line, HL60, with different potency but similar additive properties with bortezomib. Additive antiproliferative effects were correlated well with LPS-induced NF-κB inhibition results. Gene expression data on cell cycle and apoptosis related genes, obtained by high-throughput QPCR, showed that curcumin and its analogue act through similar signaling pathways. In correlation with in vitro results similar additive effect could be obsereved in SCID mice inoculated systemically with HL60 cells. C-150 in a liposomal formulation given intravenously in combination with bortezomib was more efficient than either of the drugs alone. As our novel curcumin analogue exerted anticancer effects in leukemic cells at submicromolar concentration in vitro and at 3 mg/kg dose in vivo, which was potentiated by bortezomib, it holds a great promise as a future therapeutic agent in the treatment of leukemia alone or in combination.

Allium compounds, dipropyl and dimethyl thiosulfinates as antiproliferative and differentiating agents of human acute myeloid leukemia cell lines

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Faten Merhi

2008-08-01

Full Text Available Faten Merhi1, Jacques Auger2, Francine Rendu1, Brigitte Bauvois11UMR 7131 UPMC Paris Universitas/CNRS, Groupe Hospitalier Broussais-HEGP, Paris, France; 2University F. Rabelais, IRBI, UPRESA CNRS 6035, Tours, FranceAbstract: Epidemiologic studies support the premise that Allium vegetables may lower the risk of cancers. The beneficial effects appear related to the organosulfur products generated upon processing of Allium. Leukemia cells from patients with acute myeloid leukemia (AML display high proliferative capacity and have a reduced capacity of undergoing apoptosis and maturation. Whether the sulfur-containing molecules thiosulfinates (TS, diallyl TS (All2TS, dipropyl TS (Pr2TS and dimethyl TS (Me2TS, are able to exert chemopreventative activity against AML is presently unknown. The present study was an evaluation of proliferation, cytotoxicity, differentiation and secretion of AML cell lines (U937, NB4, HL-60, MonoMac-6 in response to treatment with these TS and their related sulfides (diallylsulfide, diallyl disulfide, dipropyl disulfide, dimethyl disulfide. As assessed by flow cytometry, ELISA, gelatin zymogaphy and RT-PCR, we showed that Pr2TS and Me2TS, but not All2TS and sulfides, 1 inhibited cell proliferation in dose- and time-dependent manner and this process was neither due to cytotoxicity nor apoptosis, 2 induced macrophage maturation, and 3 inhibited the levels of secreted MMP-9 (protein and activity and TNF-α protein, without altering mRNA levels. By establishing for the first time that Pr2TS and Me2TS affect proliferation, differentiation and secretion of leukemic cell lines, this study provides the opportunity to explore the potential efficiency of these molecules in AML.Keywords: acute myeloid leukemia, thiosulfinate, proliferation, differentiation, matrix metalloproteinase-9

Electronic transitions and band offsets in C60:SubPc and C60:MgPc on MoO3 studied by modulated surface photovoltage spectroscopy

International Nuclear Information System (INIS)

Fengler, S.; Dittrich, Th.; Rusu, M.

2015-01-01

Electronic transitions at interfaces between MoO 3 layers and organic layers of C 60 , SubPc, MgPc, and nano-composite layers of SubPc:C 60 and MgPc:C 60 have been studied by modulated surface photovoltage (SPV) spectroscopy. For all systems, time dependent and modulated SPV signals pointed to dissociation of excitons at the MoO 3 /organic layer interfaces with a separation of holes towards MoO 3 . The highest occupied molecular orbital (HOMO)-lowest unoccupied molecular orbital (LUMO) gaps (E HL ) of C 60 , SubPc, and MgPc and the effective E HL of SubPc:C 60 and MgPc:C 60 were measured. The offsets between the LUMO (ΔE L ) or HOMO (ΔE H ) bands were obtained with high precision and amounted to 0.33 or 0.73 eV for SubPc:C 60 , respectively, and to −0.33 or 0.67 eV for MgPc:C 60 , respectively. Exponential tails below E HL and most pronounced sub-bandgap transitions were characterized and ascribed to disorder and transitions from HOMO bands to unoccupied defect states

Derivation of Huntington Disease affected Genea046 human embryonic stem cell line

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Biljana Dumevska

2016-03-01

Full Text Available The Genea046 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, carrying HTT gene CAG expansion of 45 repeats, indicative of Huntington Disease. Following ICM outgrowth on inactivated human feeders, karyotype was confirmed as 46, XX by CGH and STR analysis demonstrated a female Allele pattern. The hESC line had pluripotent cell morphology, 85% of cells expressed Nanog, 92% Oct4, 75% Tra1–60 and 99% SSEA4 and demonstrated Alkaline Phosphatase activity. The cell line was negative for Mycoplasma and visible contamination.

The dichloromethane extract of the ethnomedicinal plant Neurolaena lobata inhibits NPM/ALK expression which is causal for anaplastic large cell lymphomagenesis.

Science.gov (United States)

Unger, Christine; Popescu, Ruxandra; Giessrigl, Benedikt; Laimer, Daniela; Heider, Susanne; Seelinger, Mareike; Diaz, Rene; Wallnöfer, Bruno; Egger, Gerda; Hassler, Melanie; Knöfler, Martin; Saleh, Leila; Sahin, Emine; Grusch, Michael; Fritzer-Szekeres, Monika; Dolznig, Helmut; Frisch, Richard; Kenner, Lukas; Kopp, Brigitte; Krupitza, Georg

2013-01-01

The present study investigates extracts of Neuolaena lobata, an anti-protozoan ethnomedicinal plant of the Maya, regarding its anti-neoplastic properties. Firstly, extracts of increasing polarity were tested in HL-60 cells analyzing inhibition of cell proliferation and apoptosis induction. Secondly, the most active extract was further tested in anaplastic large cell lymphoma (ALCL) cell lines of human and mouse origin. The dichloromethane extract inhibited proliferation of HL-60, human and mouse ALCL cells with an IC50 of ~2.5, 3.7 and 2.4 µg/ml, respectively and arrested cells in the G2/M phase. The extract induced the checkpoint kinases Chk1 and Chk2 and perturbed the orchestrated expression of the Cdc25 family of cell cycle phosphatases which was paralleled by the activation of p53, p21 and downregulation of c-Myc. Importantly, the expression of NPM/ALK and its effector JunB were drastically decreased, which correlated with the activation of caspase 3. Subsequently also platelet derived growth factor receptor β was downregulated, which was recently shown to be transcriptionally controlled by JunB synergizing with ALK in ALCL development. We show that a traditional healing plant extract downregulates various oncogenes, induces tumor suppressors, inhibits cell proliferation and triggers apoptosis of malignant cells. The discovery of the 'Active Principle(s)' is warranted.

A new high sensitivity far-infrared laser interferometer for the HL-2A tokamak

Science.gov (United States)

Li, Y. G.; Zhou, Y.; Li, Y.; Deng, Z. C.; Wang, H. X.; Yi, J.

2017-08-01

A new four-chord Michelson-type formic acid (HCOOH, λ = 432.5 μm) laser interferometer has been successfully commissioned on the HL-2A tokamak to measure the electron density and density fluctuations. Due to the employment of the two-laser heterodyne technique, the time resolution of the interferometer reached 1.0 microseconds (μs). Four chords of line electron densities with a line-averaged density resolution 2 × 1016/m3 were obtained in a recent HL-2A experimental campaign, and detailed electron density fluctuations, caused by events such as edge localized mode, sawtooth precursor-oscillations, and energetic particle driven instabilities, were distinctly measured. In particular, the high-frequency electron density fluctuations (up to 500 kHz) caused by the reversed shear Alfvénic eigenmode were observed by the internal two interferometry channels, and their fluctuation location could be approximately identified from the spectra characteristics of multi-chord line electron densities.

Development of buffalo (Bubalus bubalis embryonic stem cell lines from somatic cell nuclear transferred blastocysts

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Syed Mohmad Shah

2015-11-01

Full Text Available We developed buffalo embryonic stem cell lines from somatic cell nuclear transfer derived blastocysts, produced by hand-guided cloning technique. The inner cell mass of the blastocyst was cut mechanically using a Microblade and cultured onto feeder cells in buffalo embryonic stem (ES cell culture medium at 38 °C in a 5% CO2 incubator. The stem cell colonies were characterized for alkaline phosphatase activity, karyotype, pluripotency and self-renewal markers like OCT4, NANOG, SOX2, c-Myc, FOXD3, SSEA-1, SSEA-4, TRA-1-60, TRA-1-81 and CD90. The cell lines also possessed the capability to differentiate across all the three germ layers under spontaneous differentiation conditions.

HL-LHC Accelerator

CERN Document Server

Zimmermann, F

2013-01-01

The tentative schedule, key ingredients, as well as progress of pertinent R&D and component prototypes for the LHC luminosity upgrade, "HL-LHC," are reviewed. Also alternative scenarios based on performance-improving consolidations (PICs) instead of a full upgrade are discussed. Tentative time schedules and expected luminosity evolutions for the different scenarios are sketched. The important role of HL-LHC development as a step towards a future HE-LHC or VHE-LHC is finally highlighted. Presented at "Higgs & Beyond" Conference Tohoku University, Sendai 7 June 2013.

HL-LHC parameter space and scenarios

International Nuclear Information System (INIS)

Bruning, O.S.

2012-01-01

The HL-LHC project aims at a total integrated luminosity of approximately 3000 fb -1 over the lifetime of the HL-LHC. Assuming an exploitation period of ca. 10 years this goal implies an annual integrated luminosity of approximately 200 fb -1 to 300 fb -1 per year. This paper looks at potential beam parameters that are compatible with the HL-LHC performance goals and discusses briefly potential variation in the parameter space. It is shown that the design goal of the HL-LHC project can only be achieved with a full upgrade of the injector complex and the operation with β* values close to 0.15 m. Significant margins for leveling can be achieved for β* values close to 0.15 m. However, these margins can only be harvested during the HL-LHC operation if the required leveling techniques have been demonstrated in operation

Induction of apoptosis by plumbagin through reactive oxygen species-mediated inhibition of topoisomerase II

International Nuclear Information System (INIS)

Kawiak, Anna; Piosik, Jacek; Stasilojc, Grzegorz; Gwizdek-Wisniewska, Anna; Marczak, Lukasz; Stobiecki, Maciej; Bigda, Jacek; Lojkowska, Ewa

2007-01-01

Reactive oxygen species (ROS) have been recognized as key molecules, which can selectively modify proteins and therefore regulate cellular signalling including apoptosis. Plumbagin, a naphthoquinone exhibiting antitumor activity, is known to generate ROS and has been found to inhibit the activity of topoisomerase II (Topo II) through the stabilization of the Topo II-DNA cleavable complex. The objective of this research was to clarify the role of ROS and Topo II inhibition in the induction of apoptosis mediated by plumbagin. As determined by the comet assay, plumbagin induced DNA cleavage in HL-60 cells, whereas in a cell line with reduced Topo II activity-HL-60/MX2, the level of DNA damage was significantly decreased. The onset of DNA strand break formation in HL-60 cells was delayed in comparison with the generation of intracellular ROS. In HL-60/MX2 cells, ROS were generated at a similar rate, whereas a significant reduction in the level of DNA damage was detected. The pretreatment of cells with N-acetylcysteine (NAC) attenuated plumbagin-induced DNA damage, pointing out to the involvement of ROS generation in cleavable complex formation. These results suggest that plumbagin-induced ROS does not directly damage DNA but requires the involvement of Topo II. Furthermore, experiments carried out using light spectroscopy indicated no direct interactions between plumbagin and DNA. The induction of apoptosis was significantly delayed in HL-60/MX2 cells indicating the involvement of Topo II inhibition in plumbagin-mediated apoptosis. Thus, these findings strongly suggest ROS-mediated inhibition of Topo II as an important mechanism contributing to the apoptosis-inducing properties of plumbagin

Induction of apoptosis by 3-amino-6-(3-aminopropyl)-5,6-dihydro-5,11-dioxo-11H-indeno[1,2-c]isoquinoline via modulation of MAPKs (p38 and c-Jun N-terminal kinase) and c-Myc in HL-60 human leukemia cells.

Science.gov (United States)

Park, Eun-Jung; Kiselev, Evgeny; Conda-Sheridan, Martin; Cushman, Mark; Pezzuto, John M

2012-03-23

Recently, we reported that 3-amino-6-(3-aminopropyl)-5,6-dihydro-5,11-dioxo-11H-indeno[1,2-c]isoquinoline (AM6-36), sharing structural similarity with naturally occurring isoquinolines, induced activities mediated by retinoid X receptor (RXR) response element accompanied by antiproliferative effects on breast cancer cells. To further characterize the biologic potential of AM6-36, we currently report studies conducted with HL-60 human leukemia cells. AM6-36 significantly inhibited cellular proliferation in a dose- and time-dependent manner with an IC(50) value of 86 nM. When evaluated at low test concentrations (≤0.25 μM), AM6-36 induced arrest in the G2/M phase of the cell cycle. At higher concentrations (1 and 2 μM), the response shifted to apoptosis, which was consistent with the effect of AM6-36 on other apoptotic signatures including an increase of apoptotic annexin V(+) 7-AAD(-) cells, loss of mitochondrial membrane potential, induction of poly(ADP-ribose) polymerase cleavage, and activation of several caspases. These apoptotic effects are potentially due to up-regulation of p38 MAPK and JNK phosphorylation and down-regulation of c-Myc oncogene expression. Taken together, AM6-36 might serve as an effective anticancer agent by inducing G2/M cell cycle arrest and apoptosis through the activation of MAPKs and inhibition of c-Myc.

Difference in membrane repair capacity between cancer cell lines and a normal cell line

DEFF Research Database (Denmark)

Frandsen, Stine Krog; McNeil, Anna K.; Novak, Ivana

2016-01-01

repair was investigated by disrupting the plasma membrane using laser followed by monitoring fluorescent dye entry over time in seven cancer cell lines, an immortalized cell line, and a normal primary cell line. The kinetics of repair in living cells can be directly recorded using this technique...... cancer cell lines (p immortalized cell line (p

Rannarootslaste suurmees Hans Pöhl / Olev Liivik

Index Scriptorium Estoniae

Liivik, Olev, 1975-

2011-01-01

Arvustus: Estlandssvenskarnas hövding. En biografi över Hans Pöhl (1876-1930), estlandssvenskarnas främste företrädare och ledare = Hans Pöhl - rannarootslaste eestvõileja : Hans Pöhl (1876-1930), Eesti rootslaste vaimse liidri ja valgustaja elulugu. Stockholm, 2010

Studies of 99mTc-BnAO (HL-91): a non-nitroaromatic compound for hypoxic cell detection

International Nuclear Information System (INIS)

Zhang, X.; Melo, T.; Ballinger, J.R.; Rauth, A.M.

1998-01-01

Purpose: Solid tumours of similar type and stage can vary widely in their hypoxic cell fraction. Such cells may be prognostic for aggressive, metastatic, and radiation-resistant disease. A 99m technetium ( 99m Tc)-labelled non-nitroaromatic agent, butyleneamine oxime ( 99m Tc-BnAO) or HL-91 (Amersham International, Inc., Amersham, UK) has been evaluated both in vitro and in vivo for its possible efficacy as a noninvasive marker for the clinical detection of hypoxic cells in solid tumours. Materials and Methods: Suspension cultures of Chinese hamster ovary (CHO) cells under controlled levels of oxygen were used to measure the oxygen dependency of 99m Tc-BnAO accumulation. V79 cells grown as multilayers on a semipermeable membrane served as an in vitro model for drug penetration through the extravascular space of the tumour. C3H mice bearing KHT-C leg tumours were the in vivo models for selective drug accumulation as a function of time after i.v. administration of 99m Tc-BnAO. Results: 99m Tc accumulated selectively in hypoxic vs. aerobic cells, resulting in a 9 ± 2-fold differential in radioactivity per cell at 4 h. The k m for this selective accumulation was 20 ppm of oxygen. The labelled drug was equally effective in penetrating the cellular multilayer under aerobic or hypoxic conditions. In vivo measurements indicated favourable labelling of solid tumours containing hypoxic cells with 1% of the total activity per g of tumour, a tumour-to-blood ratio of 1.2, and a tumour-to-muscle ratio of 4.6 at 4 to 6 h after drug administration. In contrast to more lipophilic 99m Tc- labelled compounds, excretion was primarily via the urinary tract. Nitro-L-arginine selectively increased solid tumour labelling over normal tissue. Conclusions: 99m Tc-BnAO or HL-91 is a promising agent for clinical studies of tumour hypoxia, although the mechanism of its selective hypoxic cell accumulation remains unexplained

HL7 Messaging Engine with Customizable Translation System

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PRODAN, R.

2010-05-01

Full Text Available This paper introduces a new client-server messaging engine used to exchange clinical data between various medical software applications. Our portal uses the HL7 (Health Level Seven messaging standard to provide translated clinical data to HL7 and non-HL7 client applications. We used HL7 because this standard is worldwide used to facilitate the communication between clinical applications.

Human metastatic melanoma cell lines express high levels of growth hormone receptor and respond to GH treatment

Energy Technology Data Exchange (ETDEWEB)

Sustarsic, Elahu G. [Edison Biotechnology Institute, 1 Watertower Drive, Athens, OH (United States); Department of Biological Sciences, Ohio University, Athens, OH (United States); Junnila, Riia K. [Edison Biotechnology Institute, 1 Watertower Drive, Athens, OH (United States); Kopchick, John J., E-mail: kopchick@ohio.edu [Edison Biotechnology Institute, 1 Watertower Drive, Athens, OH (United States); Department of Biological Sciences, Ohio University, Athens, OH (United States); Department of Biomedical Sciences, Heritage College of Osteopathic Medicine, Ohio University, Athens, OH (United States)

2013-11-08

Highlights: •Most cancer types of the NCI60 have sub-sets of cell lines with high GHR expression. •GHR is highly expressed in melanoma cell lines. •GHR is elevated in advanced stage IV metastatic tumors vs. stage III. •GH treatment of metastatic melanoma cell lines alters growth and cell signaling. -- Abstract: Accumulating evidence implicates the growth hormone receptor (GHR) in carcinogenesis. While multiple studies show evidence for expression of growth hormone (GH) and GHR mRNA in human cancer tissue, there is a lack of quantification and only a few cancer types have been investigated. The National Cancer Institute’s NCI60 panel includes 60 cancer cell lines from nine types of human cancer: breast, CNS, colon, leukemia, melanoma, non-small cell lung, ovarian, prostate and renal. We utilized this panel to quantify expression of GHR, GH, prolactin receptor (PRLR) and prolactin (PRL) mRNA with real-time RT qPCR. Both GHR and PRLR show a broad range of expression within and among most cancer types. Strikingly, GHR expression is nearly 50-fold higher in melanoma than in the panel as a whole. Analysis of human metastatic melanoma biopsies confirmed GHR gene expression in melanoma tissue. In these human biopsies, the level of GHR mRNA is elevated in advanced stage IV tumor samples compared to stage III. Due to the novel finding of high GHR in melanoma, we examined the effect of GH treatment on three NCI60 melanoma lines (MDA-MB-435, UACC-62 and SK-MEL-5). GH increased proliferation in two out of three cell lines tested. Further analysis revealed GH-induced activation of STAT5 and mTOR in a cell line dependent manner. In conclusion, we have identified cell lines and cancer types that are ideal to study the role of GH and PRL in cancer, yet have been largely overlooked. Furthermore, we found that human metastatic melanoma tumors express GHR and cell lines possess active GHRs that can modulate multiple signaling pathways and alter cell proliferation. Based on

Radiosensitization of C225 on human non-small cell lung cancer cell line H-520

International Nuclear Information System (INIS)

Zhang Yingdong; Wang Junjie; Liu Feng; Zhao Yong

2008-01-01

Objective: To investigate the efficacy of C225 (cetuximab), a chimeric human-mouse anti-epithelial growth factor receptor monoclonal antibody, combined with 60 Co gamma irradiation against human non-small cell lung cancer cell line H-520. Methods: H-520 cells were treated either with different dose of 60 Co irradiation (1,2,4,6,8 and 10 Gy)alone or together with C225 (100 nmol/L). Colony forming capacity was determined to create the survival curve 10 days after the treatment. Cells in different groups were harvested 72 hours after irradiation for apoptosis analysis or 48 hours after irradiation for cell cycle analysis by flow cytometry assay. Results: The clone number in combinational treatment group was less than that in irradiation only group, which suggested that the cell survival rate in the combinational treatment group was significantly decreased comparing with irradiation only group (F=6.36, P O + G 1 phases for C225 treatment, in G 2 + M phases for 60 Co irradiation, and in both G 0 + G 1 and G 2 + M phases for C225 in combination with 60 Co irradiation. Conclusions: C225 has radiosensitizing effects on H-520 cells, which may through the enhancement of 60 Co irradiation-induced cell death and cell cycle arrest. This study provides a supportive evidence for clinical treatment in non-small cell lung cancer. (authors)

Hypoxia-induced redox alterations and their correlation with {sup 99m}Tc-MIBI and {sup 99m}Tc-HL-91 uptake in colon cancer cells

Energy Technology Data Exchange (ETDEWEB)

Abrantes, Ana Margarida [Biophysics/Biomathematics Institute, IBILI, Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Centre of Investigation on Environment Genetics and Oncobiology (CIMAGO), Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal)], E-mail: mabrantes@ibili.uc.pt; Silva Serra, Maria Elisa [Chemistry Department, Faculty of Sciences and Technology, University of Coimbra, 3000-535 Coimbra (Portugal); Goncalves, Ana Cristina [Biochemistry Institute, Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Centre of Investigation on Environment Genetics and Oncobiology (CIMAGO), Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Rio, Joana; Oliveiros, Barbara; Laranjo, Mafalda [Biophysics/Biomathematics Institute, IBILI, Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Rocha-Gonsalves, Antonio Manuel [Chemistry Department, Faculty of Sciences and Technology, University of Coimbra, 3000-535 Coimbra (Portugal); Centre of Investigation on Environment Genetics and Oncobiology (CIMAGO), Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Sarmento-Ribeiro, Ana Bela [Biochemistry Institute, Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Centre of Investigation on Environment Genetics and Oncobiology (CIMAGO), Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Filomena Botelho, Maria [Biophysics/Biomathematics Institute, IBILI, Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal); Centre of Investigation on Environment Genetics and Oncobiology (CIMAGO), Faculty of Medicine, University of Coimbra, 3000-548 Coimbra (Portugal)

2010-02-15

Colorectal cancer is one of the most common malignancies in the Western world and is an example of a solid tumour in which hypoxia is a common feature and develops because of the inability of the vascular system to supply adequate amounts of oxygen to growing tumours. Hypoxia effects on tumour cell biology can be detected and characterized using different methods. The use of imaging with {gamma}-emitting radionuclides to detect hypoxic tissue was first suggested by Chapman in 1979 [N Engl J Med 301 (1979) 1429-1432]. {sup 99m}Tc-4,9-diaza-3,3,10,10-tetramethyldodecan-2,11-dione dioxime, also known as {sup 99m}Tc-HL-91, has been among the most studied hypoxia markers. The objective of this study was to correlate the uptake of {sup 99m}Tc-HL-91 and {sup 99m}Tc-MIBI in colon cancer cells under normoxic and hypoxic conditions and to compare this information with some parameters such as oxidative stress and mitochondrial dysfunction of the cells analyzed by flow cytometry. Our results show that the in vitro {sup 99m}Tc-HL-91 uptake is higher in hypoxic conditions, which is confirmed by the decreased uptake of {sup 99m}Tc-MIBI. Flow cytometry results demonstrate that hypoxic conditions used are not enough to induce cellular death, but are responsible for the alterations in the intracellular redox environment, namely, increase of ROS production, proteic pimonidazol-derived adduct formation and alteration in the mitochondrial membrane permeability. Therefore, these results confirm that {sup 99m}Tc-HL-91 is a radiopharmaceutical with favourable characteristics for detecting hypoxia.

Derivation of NEM2 affected human embryonic stem cell line Genea079

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Biljana Dumevska

2016-03-01

Full Text Available The Genea079 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, carrying compound heterozygous mutations in the NEB gene, exon 55 deletion & c.15110dupA, indicative of Nemaline Myopathy Type 2 (NEM2. Following ICM outgrowth on inactivated human feeders, karyotype was confirmed as 46, XY and STR analysis demonstrated a male Allele pattern. The hESC line had pluripotent cell morphology, 86% of cells expressed Nanog, 95% Oct4, 54% Tra1-60 and 98% SSEA4 and gave a PluriTest Pluripotency score of 30.25, Novelty of 1.21. The cell line was negative for Mycoplasma and visible contamination.

78 FR 13688 - Proposed Collection; 60-Day Comment Request: Request for Human Embryonic Stem Cell Line To Be...

Science.gov (United States)

2013-02-28

... Comment Request: Request for Human Embryonic Stem Cell Line To Be Approved for Use in NIH Funded Research... Embryonic Stem Cell Line to be Approved for Use in NIH Funded Research. OMB No. 0925-0601-- Expiration Date... and Use of Information Collection: The form is used by applicants to request that human embryonic stem...

An experimental study on the low-dose radiosensitivity of tumor cell lines

International Nuclear Information System (INIS)

Kim, Min Sook; Koh, Kwang Joon

1994-01-01

The purpose of this study was to aid in the radiation therapy of head and neck cancer patients. For this study, radiation survival curves were generated for B16, MG-63 and YAC-1 cell lines using semiautomated MTT assay and Dye Exclusion Assay. Irradiation of 2, 4, 6, 8, 10 Gy were delivered at room temperature at a dose rate of 210.2 cGy/min using 60 COγ-ray irradiator ALDORADO 8. The viable cells were determined for each radiation dose and compared to control values. The obtained results were as follows: 1. The was significantly different absorbance at 10 Gy on B16 cell line in MTT assay (P<0.05). 2. There was significantly different absorbance at 4, 6, 8, 10 Gy on MG-63 cell line in MTT assay (P<0.05). 3. YAC-1 cell line was more sensitive than B16 or MG-63 cell line to all doses of radiation (P<0.05). 4. There was significantly different absorbance among all tumor cell lines except between B16 and MG-63 cell line at 2 Gy in MTT assay (P<0.05). 5. Good correlation was obtained between MTT assay and DEA (P<0.05). The efficient of correlation of B16, MG-63 and YAC-1 cell line was 0.845-0.824 and 0.906, respectively.

Von Hippel-Lindau disease (vHL)

DEFF Research Database (Denmark)

Binderup, Marie Louise Mølgaard; Bisgaard, Søs Marie Luise; Harbud, Vibeke

2013-01-01

for vHL which is comprised of Danish doctors and specialists interested in vHL. The recommendations are based on longstanding clinical experience, Danish original research, and extensive review of the international literature. vHL is a hereditary multi-tumour disease caused by germline mutations....../MRI of the abdomen, e) annual plasma-metanephrine, plasma-normetanephrine, and plasma-chromogranin A tests, and f) annual hearing examination at a department of audiology. It is advised that one doctor takes on the responsibility of coordination of and referral to the many examinations, and the communication...

Alterations in polyamine levels induced by phorbol diesters and other agents that promote differentiation in human promyelocytic leukemia cells

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Huberman, E.; Weeks, C.; Herrmann, A.; Callaham, M.; Slaga, T.

1981-02-01

Polyamine levels were evaluated in human HL-60 promyelocytic leukemia cells after treatment with inducers of terminal differentiation. Differentiation in these cells was determined by increases in the percentage of morphologically mature cells and in lysozyme activity. Treatment of the HL-60 cells with phorbol 12-myristate-13-acetate (PMA), phorbol 12,13-didecanoate or other inducers of terminal differentiation such as dimethylsulfoxide and retinoic acid resulted in increased levels of putrescine. However, no increase in putrescine could be detected after PMA treatment of a HL-60 cell variant that exhibited a decreased susceptibility to PMA-induced terminal differentiation. Similarly, no increase in putrescine was observed with two nontumor-promoters (phorbol 12,13-diacetate and 4-O-methyl-PMA) or with anthralin, a non-phorbol tumor promoter. In addition to enhancing putrescine levels, PMA also increased the amount of spermidine and decreased the amount of spermine. The increase in putrescine and spermidine preceded the expression of the various differentiation markers. Unlike the changes observed in the polyamine levels after PMA treatment, the activities of ornithine and S-adenosylmethionine decarboxylases, which are polyamine biosynthetic enzymes, did not significantly change. ..cap alpha..-Methylornithine and ..cap alpha..-difluoromethylornithine and methylglyoxal bis(guanylhydrazone), which are inhibitors of the polyamine biosynthetic enzymes, did not affect differentiation in control or PMA-treated cells. Because of these observations, we suggest that the change in polyamine levels involve biochemical pathways other than the known biosynthetic ones. By-products of these pathways may perhaps be the controlling factors involved in the induction of terminal differentiation in the HL-60 and other cell types as well.

Concentration-dependent metabolic effects of metformin in healthy and Fanconi anemia lymphoblast cells.

Science.gov (United States)

Ravera, Silvia; Cossu, Vanessa; Tappino, Barbara; Nicchia, Elena; Dufour, Carlo; Cavani, Simona; Sciutto, Andrea; Bolognesi, Claudia; Columbaro, Marta; Degan, Paolo; Cappelli, Enrico

2018-02-01

Metformin (MET) is the drug of choice for patients with type 2 diabetes and has been proposed for use in cancer therapy and for treating other metabolic diseases. More than 14,000 studies have been published addressing the cellular mechanisms affected by MET. However, several in vitro studies have used concentrations of the drug 10-100-fold higher than the plasmatic concentration measured in patients. Here, we evaluated the biochemical, metabolic, and morphologic effects of various concentrations of MET. Moreover, we tested the effect of MET on Fanconi Anemia (FA) cells, a DNA repair genetic disease with defects in energetic and glucose metabolism, as well as on human promyelocytic leukemia (HL60) cell lines. We found that the response of wild-type cells to MET is concentration dependent. Low concentrations (15 and 150 µM) increase both oxidative phosphorylation and the oxidative stress response, acting on the AMPK/Sirt1 pathway, while the high concentration (1.5 mM) inhibits the respiratory chain, alters cell morphology, becoming toxic to the cells. In FA cells, MET was unable to correct the energetic/respiratory defect and did not improve the response to oxidative stress and DNA damage. By contrast, HL60 cells appear sensitive also at 150 μM. Our findings underline the importance of the MET concentration in evaluating the effect of this drug on cell metabolism and demonstrate that data obtained from in vitro experiments, that have used high concentrations of MET, cannot be readily translated into improving our understanding of the cellular effects of metformin when used in the clinical setting. © 2017 Wiley Periodicals, Inc.

The in vivo fate of a 211At labelled monoclonal antibody with known specificity in a murine system

International Nuclear Information System (INIS)

Vaughan, A.T.M.; Bateman, W.J.; Fisher, D.R.

1982-01-01

A monoclonal antibody reactive against the human transferrin receptor has been labelled with the alpha and X ray emitting isotope Astatine 211. The labelling procedure does not affect the ability of the product to bind to the transferrin receptor on the human leukemic cell line HL60. Using a direct binding assay, 211 At labelled antibody can be specifically inhibited from binding to its target cells by excess unlabelled antibody. Furthermore, the binding inhibition demonstrated in this system correlates to enhanced clonogenic survival of these cells, indicating that very few atoms of 211 At/cell are required for cell death. Data obtained from labelled antibody injected into mice show that the labelled product in serum retains the ability to bind to HL60 cells in vitro, although tissue distributions of the injected activity implies that some of the radiolabel is lost from the protein. Despite this loss of label, preliminary experiments on the localization of labelled antibody to HL60 cells growing s/c in nude mice show that tumor tissue has a higher specific activity than all other tissues, other than blood, after 12 hours. This suggests that further work on the nature of label degradation in vivo is warranted in the context of potential therapeutic and diagnostic studies

Synthesis and cytotoxicity evaluation of thiosemicarbazones and their thiazole derivatives

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Saulo Feheiberg Pinto Braga

Full Text Available ABSTRACT The aims of this study were to synthesize a series of thiosemicarbazones and their thiazole derivatives, to investigate their cytotoxic activity against three human cancers and normal (Vero cells cell lines, and to evaluate the pro-apoptotic potential of the most active compounds. Materials and Methods: The thiosemicarbazones were obtained by reacting an aromatic aldehyde with thiosemicarbazide (yield 71-96%, which were subjected to a cyclization with α-bromoacetophenone to yield the required thiazole heterocycles (yield 63-100%. All the synthesized compounds were screened at 50 µM concentration against three cell lines representing HL60 (promyelocytic leukemia, Jurkat (acute lymphoblastic leukemia, and MCF-7 (breast cancer. The pro-apoptotic effect was measured by flow cytometry as the percentage of cells with hypodiploid DNA. Results: Three thiazole compounds showed activity against at least one tumor cell line (IC50 = 43-76 µM and low cytotoxicity against Vero cells (IC50 > 100 M. The most active compound of this series induced 91% and 51% DNA fragmentation in HL60 and MCF-7 cell lines, respectively, suggesting that this compound triggered apoptosis in these cells. Conclusion: Among the synthesized compounds, one in particular was found to exert antiproliferative and pro-apoptotic activity on tumor cells and can be considered promising as a lead molecule for the design of new analogues with improved activity.

Characterization of miRNomes in Acute and Chronic Myeloid

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Qian Xiong

2014-04-01

Full Text Available Myeloid leukemias are highly diverse diseases and have been shown to be associated with microRNA (miRNA expression aberrations. The present study involved an in-depth miRNome analysis of two human acute myeloid leukemia (AML cell lines, HL-60 and THP-1, and one human chronic myeloid leukemia (CML cell line, K562, via massively parallel signature sequencing. mRNA expression profiles of these cell lines that were established previously in our lab facilitated an integrative analysis of miRNA and mRNA expression patterns. miRNA expression profiling followed by differential expression analysis and target prediction suggested numerous miRNA signatures in AML and CML cell lines. Some miRNAs may act as either tumor suppressors or oncomiRs in AML and CML by targeting key genes in AML and CML pathways. Expression patterns of cell type-specific miRNAs could partially reflect the characteristics of K562, HL-60 and THP-1 cell lines, such as actin filament-based processes, responsiveness to stimulus and phagocytic activity. miRNAs may also regulate myeloid differentiation, since they usually suppress differentiation regulators. Our study provides a resource to further investigate the employment of miRNAs in human leukemia subtyping, leukemogenesis and myeloid development. In addition, the distinctive miRNA signatures may be potential candidates for the clinical diagnosis, prognosis and treatment of myeloid leukemias.

An experimental study of 99Tcm-PnAO-nitroimidazole and 99Tcm-HL91 in detection of myocardial hypoxia

International Nuclear Information System (INIS)

Yao Zhiming; Liu Xiujie; Shi Rongfang; Guo Feng; Liu Yunzhong; Wang Qi; Wei Hongxing; Zhu Lin

1999-01-01

Objective: To investigate the biological characteristics of two hypoxic-avid imaging agents, 99 Tc m -PnAO-nitroimidazole and 99 Tc m -HL91, and the experimental myocardial hypoxia, low perfusion and ischemic reperfusion were performed. Methods: Isolated rat hearts were retrograde perfused with Kerbs-Henscleit buffer (KH) in four phantoms: control, low perfusion, ischemic reperfusion and hypoxia. Control hearts were perfused with 95% O 2 and 5% CO 2 balanced KH at 9∼10 mL/min rate. Low perfusion hearts were perfused with the same KH at 1 mL/min rate, and ischemic reperfusion rat hearts were discontinued perfusion for 20 min then reperfused with the same KH. Hypoxic hearts were perfused with 100% N 2 balanced KH. Results: 1) At 30 and 60 min, the radioactivity ratio of liver to heart (L/H) was 11.6 and 6.9 ( 99 Tc m -PnAO-nitroimidazole), and 5.7 and 6.2 ( 99 Tc m -HL91), respectively. The lung to heart ratios at 60 min of 99 Tc m -PnAO-nitroimidazole and 99 Tc m -HL91 were 1.8 and 2.1, respectively 2) The accumulation of 99 Tc m -PnAo-nitroimidazole in the hypoxic, low perfusion and ischemic hearts was 4.8, 3.7 and 2.8 times that in the control hearts. At 60 min after starting washout, the retentions of 99 Tc m -PnAO-nitroimidazole in hypoxic, low flow and ischemic reperfusion myocardium were much higher than that of control hearts [(23.7 +- 9.1)%]. The accumulation of 99 Tc m -HL91 in the hypoxic and low perfusion hearts was also significantly higher than that in the control and ischemic reperfusion hearts, P 99 Tc m -HL91 in the low perfusion heart was 5.34, 3.43 and 1.23 times those of control, ischemic reperfusion and hypoxic hearts, respectively. Conclusions: 99 Tc m -PnAO-nitroimidazole can be accumulated in low perfusion, ischemic reperfusion and hypoxic hearts, while 99 Tc m -HL91 is mainly accumulated in hypoxic, specially low perfusion hearts. Since there is a high L/H in perfusions with either agent, the detecting of inferoposterior wall may be

Cell lines authentication and mycoplasma detection as minimun quality control of cell lines in biobanking.

Science.gov (United States)

Corral-Vázquez, C; Aguilar-Quesada, R; Catalina, P; Lucena-Aguilar, G; Ligero, G; Miranda, B; Carrillo-Ávila, J A

2017-06-01

Establishment of continuous cell lines from human normal and tumor tissues is an extended and useful methodology for molecular characterization of cancer pathophysiology and drug development in research laboratories. The exchange of these cell lines between different labs is a common practice that can compromise assays reliability due to contamination with microorganism such as mycoplasma or cells from different flasks that compromise experiment reproducibility and reliability. Great proportions of cell lines are contaminated with mycoplasma and/or are replaced by cells derived for a different origin during processing or distribution process. The scientific community has underestimated this problem and thousand of research experiment has been done with cell lines that are incorrectly identified and wrong scientific conclusions have been published. Regular contamination and authentication tests are necessary in order to avoid negative consequences of widespread misidentified and contaminated cell lines. Cell banks generate, store and distribute cell lines for research, being mandatory a consistent and continuous quality program. Methods implementation for guaranteeing both, the absence of mycoplasma and authentication in the supplied cell lines, has been performed in the Andalusian Health System Biobank. Specifically, precise results were obtained using real time PCR detection for mycoplasma and 10 STRs identification by capillary electrophoresis for cell line authentication. Advantages and disadvantages of these protocols are discussed.

Heterogeneity of osteosarcoma cell lines led to variable responses in reprogramming.

Science.gov (United States)

Choong, Pei Feng; Teh, Hui Xin; Teoh, Hoon Koon; Ong, Han Kiat; Choo, Kong Bung; Sugii, Shigeki; Cheong, Soon Keng; Kamarul, Tunku

2014-01-01

Four osteosarcoma cell lines, Saos-2, MG-63, G-292 and U-2 OS, were reprogrammed to pluripotent state using Yamanaka factors retroviral transduction method. Embryonic stem cell (ESC)-like clusters started to appear between 15 to 20 days post transduction. Morphology of the colonies resembled that of ESC colonies with defined border and tightly-packed cells. The reprogrammed sarcomas expressed alkaline phosphatase and pluripotency markers, OCT4, SSEA4, TRA-1-60 and TRA-1-81, as in ESC up to Passage 15. All reprogrammed sarcomas could form embryoid body-like spheres when cultured in suspension in a low attachment dish for up to 10 days. Further testing on the directed differentiation capacity of the reprogrammed sarcomas showed all four reprogrammed sarcoma lines could differentiate into adipocytes while reprogrammed Saos-2-REP, MG-63-REP and G-292-REP could differentiate into osteocytes. Among the 4 osteosarcoma cell lines, U-2 OS reported the highest transduction efficiency but recorded the lowest reprogramming stability under long term culture. Thus, there may be intrinsic differences governing the variable responses of osteosarcoma cell lines towards reprogramming and long term culture effect of the reprogrammed cells. This is a first report to associate intrinsic factors in different osteosarcoma cell lines with variable reprogramming responses and effects on the reprogrammed cells after prolonged culture.

Heterogeneity of Osteosarcoma Cell Lines Led to Variable Responses in Reprogramming

Science.gov (United States)

Choong, Pei Feng; Teh, Hui Xin; Teoh, Hoon Koon; Ong, Han Kiat; Choo, Kong Bung; Sugii, Shigeki; Cheong, Soon Keng; Kamarul, Tunku

2014-01-01

Four osteosarcoma cell lines, Saos-2, MG-63, G-292 and U-2 OS, were reprogrammed to pluripotent state using Yamanaka factors retroviral transduction method. Embryonic stem cell (ESC)-like clusters started to appear between 15 to 20 days post transduction. Morphology of the colonies resembled that of ESC colonies with defined border and tightly-packed cells. The reprogrammed sarcomas expressed alkaline phosphatase and pluripotency markers, OCT4, SSEA4, TRA-1-60 and TRA-1-81, as in ESC up to Passage 15. All reprogrammed sarcomas could form embryoid body-like spheres when cultured in suspension in a low attachment dish for up to 10 days. Further testing on the directed differentiation capacity of the reprogrammed sarcomas showed all four reprogrammed sarcoma lines could differentiate into adipocytes while reprogrammed Saos-2-REP, MG-63-REP and G-292-REP could differentiate into osteocytes. Among the 4 osteosarcoma cell lines, U-2 OS reported the highest transduction efficiency but recorded the lowest reprogramming stability under long term culture. Thus, there may be intrinsic differences governing the variable responses of osteosarcoma cell lines towards reprogramming and long term culture effect of the reprogrammed cells. This is a first report to associate intrinsic factors in different osteosarcoma cell lines with variable reprogramming responses and effects on the reprogrammed cells after prolonged culture. PMID:25170299

Experiment study on the relationship between radiotherapy effect and 99Tcm-HL-91 and 18F-FDG imaging in S180 mouse

International Nuclear Information System (INIS)

Yang Aimin; Deng Huixing; Li Jie; Yu Yan; Li Xu; Chen Wei; Luo Wei

2010-01-01

To investigate the relationship between radiotherapy effect and 99 Tc m - HL-91 and 18 F-FDG imaging in S180 mouse. Methods: Animal: twenty male Kunming mice (27±3 g), obtained from animal center of medical school of Xi'an Jiaotong university, were randomly divided into two groups of radiotherapy and non-radiotherapy control group. Cells: S180 cell lines, obtained from molecular and biology center of medical school of Xi'an Jiaotong university, were thawed injected into peritoneal cavity of the 5 mouse. When the S180 tumor liquid developed, l ml liquid were dripped and dilute to the suspension solution of 2 × 106 cells. Then, 0.2 ml of it was injected into the hippo of right rear leg of mouse. The mouse model was used to experiment while the tumor dimension developed to 1-1.5 cm. 99 Tc m -HL-91 imaging: 37 MBq 99 Tc m -HL-91, obtained from Guangdong Xiai Radio-pharmaceutical Center, was injected into mouse models by tail vein. After for 4h, SPECT imaging were taken before and after radiotherapy at the time of 1 h, 2 d and 10 d. GE Hawkeye VG SPECT, equipped with low energy collimator, matrix 128 × 128, zoom 1.3, was used to acquire images for 150 seconds. Radiotherapy: Two groups mouse was irradiated to 0 Gy and 8 Gy X-ray after the first 99 Tc m -HL-91 and 18 F-FDG imaging. Images analysis: the ROI region, in tumor and lung site, was drawn for calculating the UR (uptake ratio). Results: After 1 h, 2 d and 10 d of radiation exposure, the UR values in 99 Tc m -HL-91 imaging were 3.53±1.62, 3.41±1.42, 2.5% 1.57 and 1.26±0.03, respectively, while the UR values were 3.62±1.65. 3.02±1.94, 4.10±1.48 and 2.96±2.02 in control group. This revealed that tumors hypoxic level was decreased after radiation and suggested that tumors develop re-oxygenation. After 11 d of radiation exposure, the UR values in 18 F-FDG imaging were 2.49±1.29 and 1.49±0.56, while the UR values were 2.22±0.45 and 1.89±0.08, expressing a coincident trend with 99 Tc m -HL-91 imaging. This

Quercus Suber L. Cork Extracts Induce Apoptosis in Human Myeloid Leukaemia HL-60 Cells.

Science.gov (United States)

Bejarano, Ignacio; Godoy-Cancho, Belén; Franco, Lourdes; Martínez-Cañas, Manuel A; Tormo, María A

2015-08-01

Quercus suber L. cork contains a diversity of phenolic compounds, mostly low molecular weight phenols. A rising number of reports support with convergent findings that polyphenols evoke pro-apoptotic events in cancerous cells. However, the literature related to the anti-cancer bioactivity of Q. suber L. cork extractives (QSE) is still limited. Herein, we aim to describe the antitumor potential displayed by cork extractives obtained by different extraction methods in the human promyelocytic leukaemia cells. In order to quantify the effects of QSE on cancer cells viability, phosphatidylserine exposure, caspase-3 activity, mitochondrial membrane potential and cell cycle were evaluated. The results indicated that the QSE present a time-dependent and dose-dependent cytotoxicity in the human promyelocytic leukaemia cells. Such a noxious effect leads these leukaemia cells to their death through apoptotic processes by altering the mitochondrial outer membrane potential, activating caspase-3 and externalizing phosphatidylserine. However, cells cycle progression was not affected by the treatments. This study contributes to open a new way to use this natural resource by exploiting its anti-cancer properties. Moreover, it opens new possibilities of application of cork by-products, being more efficient in the sector of cork-based agriculture. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

[The characters and specific features of new human embryonic stem cells lines].

Science.gov (United States)

Krylova, T A; Kol'tsova, A M; Zenin, V V; Gordeeva, O F; Musorina, A S; Goriachaia, T S; Shlykova, S A; Kamenetskaia, Iu K; Pinaev, G P; Polianskaia, G G

2009-01-01

Four continuous human embryonic stem cell lines (SC1, SC2, SC3 and SC4), derived from the blastocysts has been described. The cell lines were cultivated on mitotically inactivated human feeder cells. The cell lines SC1 and SC2 have passed through 150 population doublings and the cell lines SC3 and SC4 -- near 120 populations doublings, which exceeds Hayflick limit sufficiently. These cell lines maintain high activity of alkaline phosphatase, expression of transcription factor OCT-4 and cell surface antigens (SSEA-4, TRA-1-60 and TRA-1-81), confirming their ESC status and human specificity. Immunofluorescent detection of antigens, characteristic of ectoderm, endoderm and mesoderm confirms the ability of these cells to retain their pluripotency under in vitro condition. PCR analysis revealed expression of six genes specific for pluripotent cells (OCT-4, NANOG, DPPA3/STELLA, TDGF/CRIPTO and LEFTYA). Correlation between the level of proliferative activity and the character of DNA-bound fluorescent staining was found. Fluorescent dyes, Hoechst 33342 and PI, produced diffuse staining of the nuclei in slowly proliferating cells of the SC1 and SC2 lines. In contrast, in actively proliferating cells of the SC3 and SC4 lines, the clear staining of the nuclei was observed. Upon changing the cultivation condition, proliferative activity of SC3 and SC4 lines decreased and became similar to that of SC1 and SC2 lines. The character of the fluorescent staining of all these lines was also shown to be similar. These results show that quality of the fluorescent staining with Hoechst 33342 and PI reflects the level of proliferation. Possible causes and mechanisms of this feature of human ESC are discussed.

Radiation sensitivity of Merkel cell carcinoma cell lines

Energy Technology Data Exchange (ETDEWEB)

Leonard, J.H.; Ramsay, J.R.; Birrell, G.W. [Queensland Institute of Medical Research (Australia)] [and others

1995-07-30

Merkel cell carcinoma (MCC), being a small cell carcinoma, would be expected to be sensitive to radiation. Clinical analysis of patients at our center, especially those with macroscopic disease, would suggest the response is quite variable. We have recently established a number of MCC cell lines from patients prior to radiotherapy, and for the first time are in a position to determine their sensitivity under controlled conditions. Some of the MCC lines grew as suspension cultures and could not be single cell cloned; therefore, it was not possible to use clonogenic survival for all cell lines. A tetrazolium based (MTT) assay was used for these lines, to estimate cell growth after {gamma} irradiation. Control experiments were conducted on lymphoblastoid cell lines (LCL) and the adherent MCC line, MCC13, to demonstrate that the two assays were comparable under the conditions used. We have examined cell lines from MCC, small cell lung cancer (SCLC), malignant melanomas, Epstein Barr virus (EBV) transformed lymphocytes (LCL), and skin fibroblasts for their sensitivity to {gamma} irradiation using both clonogenic cell survival and MTT assays. The results show that the tumor cell lines have a range of sensitivities, with melanoma being more resistant (surviving fraction at 2 Gy (SF2) 0.57 and 0.56) than the small cell carcinoma lines, MCC (SF2 range 0.21-0.45, mean SF2 0.30, n = 8) and SCLC (SF2 0.31). Fibroblasts were the most sensitive (SF2 0.13-0.20, mean 0.16, n = 5). The MTT assay, when compared to clonogenic assay for the MCC13 adherent line and the LCL, gave comparable results under the conditions used. Both assays gave a range of SF2 values for the MCC cell lines, suggesting that these cancers would give a heterogeneous response in vivo. The results with the two derivative clones of MCC14 (SF2 for MCC14/1 0.38, MCC14/2 0.45) would further suggest that some of them may develop resistance during clonogenic evolution. 25 refs., 3 figs., 1 tab.

Radiation sensitivity of Merkell cell carcinoma cell lines

International Nuclear Information System (INIS)

Leonard, J. Helen; Ramsay, Jonathan R.; Kearsley, John H.; Birrell, Geoff W.

1995-01-01

Purpose: Merkel cell carcinoma (MCC), being a small cell carcinoma, would be expected to be sensitive to radiation. Clinical analysis of patients at our center, especially those with macroscopic disease, would suggest the response is quite variable. We have recently established a number of MCC cell lines from patients prior to radiotherapy, and for the first time are in a position to determine their sensitivity under controlled conditions. Methods and Materials: Some of the MCC lines grew as suspension cultures and could not be single cell cloned; therefore, it was not possible to use clonogenic survival for all cell lines. A tetrazolium based (MTT) assay was used for these lines, to estimate cell growth after γ irradiation. Control experiments were conducted on lymphoblastoid cell lines (LCL) and the adherent MCC line, MCC13, to demonstrate that the two assays were comparable under the conditions used. Results: We have examined cell lines from MCC, small cell lung cancer (SCLC), malignant melanomas, Epstein Barr virus (EBV) transformed lymphocytes (LCL), and skin fibroblasts for their sensitivity to γ irradiation using both clonogenic cell survival and MTT assays. The results show that the tumor cell lines have a range of sensitivities, with melanoma being more resistant (surviving fraction at 2 Gy (SF2) 0.57 and 0.56) than the small cell carcinoma lines, MCC (SF2 range 0.21-0.45, mean SF2 0.30, n = 8) and SCLC (SF2 0.31). Fibroblasts were the most sensitive (SF2 0.13-0.20, mean 0.16, n = 5). The MTT assay, when compared to clonogenic assay for the MCC13 adherent line and the LCL, gave comparable results under the conditions used. Conclusion: Both assays gave a range of SF2 values for the MCC cell lines, suggesting that these cancers would give a heterogeneous response in vivo. The results with the two derivative clones of MCC14 (SF2 for MCC14/1 0.38, MCC14/2 0.45) would further suggest that some of them may develop resistance during clonogenic evolution

Single-dose and fractionated irradiation of four human lung cancer cell lines in vitro

International Nuclear Information System (INIS)

Brodin, O.; Lennartsson, L.; Nilsson, S.

1991-01-01

Four established human lung cancer cell lines were exposed to single-dose irradiation. The survival curves of 2 small cell lung carcinomas (SCLC) were characterized by a limited capacity for repair with small and moderate shoulders with extrapolation numbers (n) of 1.05 and 1.60 respectively. Two non-small cell lung carcinoma (NSCLC) cell lines, one squamous cell (SQCLC) and one large cell (LCLC) had large shoulders with n-values of 73 and 15 respectively. The radiosensitivity when measured as D 0 did not, however, differ as much from cell line to cell line, with values from 1.22 to 1.65. The surviving fraction after 2 Gy (SF2) was 0.24 and 0.42 respectively in the SCLC cell lines and 0.90 and 0.88 respectively in the NSCLC cell lines. Fractionated irradiation delivered according to 3 different schedules was also investigated. All the schedules delivered a total dose of 10 Gy in 5 days and were applied in 1, 2 and 5 Gy dose fractions respectively. Survival followed the pattern found after single-dose irradiation; it was lowest in the SCLC cell line with the lowest SF and highest in the two NSCLC cell lines. In the SCLC cell lines all schedules were approximately equally efficient. In the LCLC and in the SQCLC cell lines, the 5 Gy schedule killed more cells than the 1 and 2 Gy schedules. The results indicate that the size of the shoulder of the survival curve is essential when choosing the most tumoricidal fractionation schedule. (orig.)

Efficacy of an adapted granzyme B-based anti-CD30 cytolytic fusion protein against PI-9-positive classical Hodgkin lymphoma cells in a murine model

International Nuclear Information System (INIS)

Schiffer, S; Hansen, H P; Hehmann-Titt, G; Huhn, M; Fischer, R; Barth, S; Thepen, T

2013-01-01

Tumors develop when infiltrating immune cells contribute growth stimuli, and cancer cells are selected to survive within such a cytotoxic microenvironment. One possible immune-escape mechanism is the upregulation of PI-9 (Serpin B9) within cancer cells. This serine proteinase inhibitor selectively inactivates apoptosis-inducing granzyme B (GrB) from cytotoxic granules of innate immune cells. We demonstrate that most classical Hodgkin lymphoma (cHL)-derived cell lines express PI-9, which protects them against the GrB attack and thereby renders them resistant against GrB-based immunotherapeutics. To circumvent this disadvantage, we developed PI-9-insensitive human GrB mutants as fusion proteins to target the Hodgkin-selective receptor CD30. In contrast to the wild-type GrB, a R201K point-mutated GrB construct most efficiently killed PI-9-positive and -negative cHL cells. This was tested in vitro and also in vivo whereby a novel optical imaging-based tumor model with HL cell line L428 was applied. Therefore, this variant, as part of the next generation immunotherapeutics, also named cytolytic fusion proteins showing reduced immunogenicity, is a promising molecule for (targeted) therapy of patients with relapsing malignancies, such as cHL, and possibly other PI-9-positive malignancies, such as breast or lung carcinoma

The CMS HGCAL detector for HL-LHC upgrade

CERN Document Server

Martelli, Arabella

2017-01-01

The High Luminosity LHC (HL-LHC) will integrate 10 times more luminosity than the LHC, posing significant challenges for radiation tolerance and event pileup on detectors, especially for forward calorimetry, and hallmarks the issue for future colliders. As part of its HL-LHC upgrade program, the CMS collaboration is designing a High Granularity Calorimeter to replace the existing endcap calorimeters. It features unprecedented transverse and longitudinal segmentation for both electromagnetic (ECAL) and hadronic (HCAL) compartments. This will facilitate particle-flow calorimetry, where the fine structure of showers can be measured and used to enhance pileup rejection and particle identification, whilst still achieving good energy resolution. The ECAL and a large fraction of HCAL will be based on hexagonal silicon sensors of 0.5 - 1 cm$^2$ cell size, with the remainder of the HCAL based on highly-segmented scintillators with SiPM readout. The intrinsic high-precision timing capabilities of the silicon sensors wi...

The CMS High Granularity Calorimeter for HL-LHC

CERN Document Server

Mastrolorenzo, Luca

2017-01-01

The High Luminosity LHC (HL-LHC) will integrate 10 times more luminosity than the LHC, posing significant challenges for radiation tolerance and event pileup on detectors, especially for forward calorimetry, and hallmarks the issue for future colliders. As part of its HL-LHC upgrade program, the CMS collaboration is designing a High Granularity Calorimeter to replace the existing endcap calorimeters. It features unprecedented transverse and longitudinal segmentation for both electromagnetic (ECAL) and hadronic (HCAL) compartments. This will facilitate particle-flow calorimetry, where the fine structure of showers can be measured and used to enhance pileup rejection and particle identification, whilst still achieving good energy resolution. The ECAL and a large fraction of HCAL will be based on hexagonal silicon sensors of 0.5 - 1 cm$^2$ cell size, with the remainder of the HCAL based on highly-segmented scintillators with SiPM readout. The intrinsic high-precision timing capabilities of the silicon sensors wi...

Coldbox installation for HL-LHC crab cavity test in the SPS tunnel (BA6)

CERN Document Server

Ordan, Julien Marius

2018-01-01

The Cold-box, produced by Linde Kryotechnik for the SPS crab-cavity test stand for HL-LHC, on its arrival at CERN. It wll be transported into the tunnel in horizontal position and then will have to be tilted in its operational position, on its vertical axis, before connecting it to cryogenics lines.

Cell Line Data Base: structure and recent improvements towards molecular authentication of human cell lines.

Science.gov (United States)

Romano, Paolo; Manniello, Assunta; Aresu, Ottavia; Armento, Massimiliano; Cesaro, Michela; Parodi, Barbara

2009-01-01

The Cell Line Data Base (CLDB) is a well-known reference information source on human and animal cell lines including information on more than 6000 cell lines. Main biological features are coded according to controlled vocabularies derived from international lists and taxonomies. HyperCLDB (http://bioinformatics.istge.it/hypercldb/) is a hypertext version of CLDB that improves data accessibility by also allowing information retrieval through web spiders. Access to HyperCLDB is provided through indexes of biological characteristics and navigation in the hypertext is granted by many internal links. HyperCLDB also includes links to external resources. Recently, an interest was raised for a reference nomenclature for cell lines and CLDB was seen as an authoritative system. Furthermore, to overcome the cell line misidentification problem, molecular authentication methods, such as fingerprinting, single-locus short tandem repeat (STR) profile and single nucleotide polymorphisms validation, were proposed. Since this data is distributed, a reference portal on authentication of human cell lines is needed. We present here the architecture and contents of CLDB, its recent enhancements and perspectives. We also present a new related database, the Cell Line Integrated Molecular Authentication (CLIMA) database (http://bioinformatics.istge.it/clima/), that allows to link authentication data to actual cell lines.

Oxygen enhancement ratio (OER) to Neutron and Co-60 γ ray

International Nuclear Information System (INIS)

Kim, Mi Sook; Ji, Young Hoon; Lee, Yong Min; Kim Kyeoung Jung

1997-01-01

Experiments in vitro, using human cell lines was carried out in order to establish whether or not there was a difference between oxygen enhancement ratio (OER) of neutron and Co-60 γ-ray and to determine OER dependence on radiation dose. MG-63 cell line and H-460 cell line were defined as the most sensitive cell line to neutron among our laboratory holding cell lines through preliminary study. Anoxia as was produced in glove box. The box was flushed for one hour with a mixture of 5 % CO 2 in ultrapure N 2 (total oxygen concentration < 10 ppm) and irradiated with neutron and Co-60 γ-ray. Oxic condition was same as anoxic condition except being irradiated in general air condition. The lower OER was observed in neutron than in Co-60 γ-ray. The dose dependence of OER was observed in neutron and Co-60 γ-ray all. But the dose dependence of the OER is somewhat larger for Co-60 γ-ray than for neutron. In the range of 1 to 8 Gy, the OER for photon and neutron range from 1.54 to 1.94 and 1.23 to 1.26 in MG-63 cell line. In case of H-460 the OER for Co-60 γ-ray and neutron range from 1.24 to 1.60 and 1.06 to 1.07 respectively. (author). 19 refs., 5 tabs., 5 figs

Influence of high-frequency electromagnetic fields on different modes of cell death and gene expression.

Science.gov (United States)

Port, M; Abend, M; Römer, B; Van Beuningen, D

2003-09-01

International thresholds for exposure to non-ionizing radiation leading to non-thermal effects were conservatively set by the International Commission on Non-Ionizing Radiation Protection (ICNIRP). The aim of this study was to examine whether biological effects such as different modes of cell death and gene expression modifications related to tumorgenesis are detectable above the threshold defined. Human leukaemia cells (HL-60) grown in vitro were exposed to electromagnetic fields (EMF; t 1/2(r) about 1 ns; field strength about 25 times higher than the ICNIRP reference levels for occupational exposure) leading to non-thermal effects using a high-voltage-improved GTEM cell 5302 (EMCO) connected to a pulse generator NP20 (C = 1 nF, U(Load) = 20kV). HL-60 cells were harvested at 0, 24, 48 and 72 h after radiation exposure. Micronuclei, apoptosis and abnormal cells (e.g. necrosis) were determined using morphological criteria. In parallel, the expression of 1176 genes was measured using Atlas Human 1.2. Array. Based on high data reproducibility calculated from two independent experiments (> 99%), array analysis was performed. No significant change in apoptosis, micronucleation, abnormal cells and differential gene expression was found. Exposure of HL-60 cells to EMFs 25 times higher than the ICNIRP reference levels for occupational exposure failed to induce any changes in apoptosis, micronucleation, abnormal morphologies and gene expression. Further experiments using EMFs above the conservatively defined reference level set by the ICNIRP may be desirable.

Preliminary results of MHD stability in HL-1 tokamak

International Nuclear Information System (INIS)

Zheng Yongzhen; Ma Tengcai; Xiao Zhenggui Cai Renfang

1987-01-01

In this paper, MHD activities of HL-1 tokamak plasma are studied with Fourier transform and correlatio analysis. The poloidal modes m = 1, 2, 3,4 and toroidal modes n of MHD magnetic fluctuation signals are detected. Methods for suppressing MHD instabilities are suggested and tested, after MHD instabilities are studied in HL-1. The effects of MHD characteristics in the beginning stage of discharge on the whole process of discharge are analyzed. The disruption, in HL-1 device could be divided into three kinds: internal disruption, minor disruption and major disruption. The result shows that HL-1 will have a better operation condition if internal disruption appears. In is end, the stable operation region of HL-1 tokamak is also given

Pulse discharge cleaning of the vacuum vessel of HL-1 tokamak

International Nuclear Information System (INIS)

Li Guodong; Zhu Yukun; Xiao Zhenggui; Sun Shouqi; Ze Mingrui

1986-01-01

The HL-1 Tokamak was test-operated on September 21, 1984. During the period of vacuum conditioning, including 60 hours of baking up to 200 deg C and 7 x 10 4 shots of pulse discharge cleaning, the calculated quantities of carbon and oxygen removed are equivalent to 24 and 6 monolayers, respectively. Then, 124 shots of tokamak discharge were performed with low level plasma parameters. The plasma current and pulse length achieved were 60 kA and 85 ms at the toroidal magnetic field of 15 kG. This paper described the techniques used and the effect on discharge characteristics of bakeout and pulse discharge cleaning of the vacuum vessel

Therapeutic Effects of Myeloid Cell Leukemia-1 siRNA on Human Acute Myeloid Leukemia Cells

Directory of Open Access Journals (Sweden)

Hadi Karami

2014-05-01

Full Text Available Purpose: Up-regulation of Mcl-1, a known anti-apoptotic protein, is associated with the survival and progression of various malignancies including leukemia. The aim of this study was to explore the effect of Mcl-1 small interference RNA (siRNA on the proliferation and apoptosis of HL-60 acute myeloid leukemia (AML cells. Methods: siRNA transfection was performed using Lipofectamine™2000 reagent. Relative mRNA and protein expressions were quantified by quantitative real-time PCR and Western blotting, respectively. Trypan blue assay was performed to assess tumor cell proliferation after siRNA transfection. The cytotoxic effect of Mcl-1 siRNA on leukemic cells was measured using MTT assay. Apoptosis was detected using ELISA cell death assay. Results: Mcl-1 siRNA clearly lowered both Mcl-1 mRNA and protein levels in a time-dependent manner, leading to marked inhibition of cell survival and proliferation. Furthermore, Mcl-1 down-regulation significantly enhanced the extent of HL-60 apoptotic cells. Conclusion: Our results suggest that the down-regulation of Mcl-1 by siRNA can effectively trigger apoptosis and inhibit the proliferation of leukemic cells. Therefore, Mcl-1 siRNA may be a potent adjuvant in AML therapy.

CLO : The cell line ontology

NARCIS (Netherlands)

Sarntivijai, Sirarat; Lin, Yu; Xiang, Zuoshuang; Meehan, Terrence F.; Diehl, Alexander D.; Vempati, Uma D.; Schuerer, Stephan C.; Pang, Chao; Malone, James; Parkinson, Helen; Liu, Yue; Takatsuki, Terue; Saijo, Kaoru; Masuya, Hiroshi; Nakamura, Yukio; Brush, Matthew H.; Haendel, Melissa A.; Zheng, Jie; Stoeckert, Christian J.; Peters, Bjoern; Mungall, Christopher J.; Carey, Thomas E.; States, David J.; Athey, Brian D.; He, Yongqun

2014-01-01

Background: Cell lines have been widely used in biomedical research. The community-based Cell Line Ontology (CLO) is a member of the OBO Foundry library that covers the domain of cell lines. Since its publication two years ago, significant updates have been made, including new groups joining the CLO

Nanostructured gold microelectrodes for extracellular recording from electrogenic cells.

Science.gov (United States)

Brüggemann, D; Wolfrum, B; Maybeck, V; Mourzina, Y; Jansen, M; Offenhäusser, A

2011-07-01

We present a new biocompatible nanostructured microelectrode array for extracellular signal recording from electrogenic cells. Microfabrication techniques were combined with a template-assisted approach using nanoporous aluminum oxide to develop gold nanopillar electrodes. The nanopillars were approximately 300-400 nm high and had a diameter of 60 nm. Thus, they yielded a higher surface area of the electrodes resulting in a decreased impedance compared to planar electrodes. The interaction between the large-scale gold nanopillar arrays and cardiac muscle cells (HL-1) was investigated via focused ion beam milling. In the resulting cross-sections we observed a tight coupling between the HL-1 cells and the gold nanostructures. However, the cell membranes did not bend into the cleft between adjacent nanopillars due to the high pillar density. We performed extracellular potential recordings from HL-1 cells with the nanostructured microelectrode arrays. The maximal amplitudes recorded with the nanopillar electrodes were up to 100% higher than those recorded with planar gold electrodes. Increasing the aspect ratio of the gold nanopillars and changing the geometrical layout can further enhance the signal quality in the future.

Nanostructured gold microelectrodes for extracellular recording from electrogenic cells

Energy Technology Data Exchange (ETDEWEB)

Brueggemann, D; Wolfrum, B; Maybeck, V; Mourzina, Y; Jansen, M; Offenhaeusser, A, E-mail: a.offenhaeusser@fz-juelich.de [Institute of Complex Systems and Peter Gruenberg Institute: Bioelectronics (ICS8/PGI8), Forschungszentrum Juelich GmbH, Leo-Brandt-Strasse, 52428 Juelich (Germany); Juelich-Aachen Research Alliance-Fundamental of Future Information Technology (JARA-FIT) (Germany)

2011-07-01

We present a new biocompatible nanostructured microelectrode array for extracellular signal recording from electrogenic cells. Microfabrication techniques were combined with a template-assisted approach using nanoporous aluminum oxide to develop gold nanopillar electrodes. The nanopillars were approximately 300-400 nm high and had a diameter of 60 nm. Thus, they yielded a higher surface area of the electrodes resulting in a decreased impedance compared to planar electrodes. The interaction between the large-scale gold nanopillar arrays and cardiac muscle cells (HL-1) was investigated via focused ion beam milling. In the resulting cross-sections we observed a tight coupling between the HL-1 cells and the gold nanostructures. However, the cell membranes did not bend into the cleft between adjacent nanopillars due to the high pillar density. We performed extracellular potential recordings from HL-1 cells with the nanostructured microelectrode arrays. The maximal amplitudes recorded with the nanopillar electrodes were up to 100% higher than those recorded with planar gold electrodes. Increasing the aspect ratio of the gold nanopillars and changing the geometrical layout can further enhance the signal quality in the future.

Molecular analysis of urothelial cancer cell lines for modeling tumor biology and drug response.

Science.gov (United States)

Nickerson, M L; Witte, N; Im, K M; Turan, S; Owens, C; Misner, K; Tsang, S X; Cai, Z; Wu, S; Dean, M; Costello, J C; Theodorescu, D

2017-01-05

The utility of tumor-derived cell lines is dependent on their ability to recapitulate underlying genomic aberrations and primary tumor biology. Here, we sequenced the exomes of 25 bladder cancer (BCa) cell lines and compared mutations, copy number alterations (CNAs), gene expression and drug response to BCa patient profiles in The Cancer Genome Atlas (TCGA). We observed a mutation pattern associated with altered CpGs and APOBEC-family cytosine deaminases similar to mutation signatures derived from somatic alterations in muscle-invasive (MI) primary tumors, highlighting a major mechanism(s) contributing to cancer-associated alterations in the BCa cell line exomes. Non-silent sequence alterations were confirmed in 76 cancer-associated genes, including mutations that likely activate oncogenes TERT and PIK3CA, and alter chromatin-associated proteins (MLL3, ARID1A, CHD6 and KDM6A) and established BCa genes (TP53, RB1, CDKN2A and TSC1). We identified alterations in signaling pathways and proteins with related functions, including the PI3K/mTOR pathway, altered in 60% of lines; BRCA DNA repair, 44%; and SYNE1-SYNE2, 60%. Homozygous deletions of chromosome 9p21 are known to target the cell cycle regulators CDKN2A and CDKN2B. This loci was commonly lost in BCa cell lines and we show the deletions extended to the polyamine enzyme methylthioadenosine (MTA) phosphorylase (MTAP) in 36% of lines, transcription factor DMRTA1 (27%) and antiviral interferon epsilon (IFNE, 19%). Overall, the BCa cell line genomic aberrations were concordant with those found in BCa patient tumors. We used gene expression and copy number data to infer pathway activities for cell lines, then used the inferred pathway activities to build a predictive model of cisplatin response. When applied to platinum-treated patients gathered from TCGA, the model predicted treatment-specific response. Together, these data and analysis represent a valuable community resource to model basic tumor biology and to study

Indole alkaloids from leaves and twigs of Rauvolfia verticillata.

Science.gov (United States)

Zhang, Bing-Jie; Peng, Lei; Wu, Zhi-Kun; Bao, Mei-Fen; Liu, Ya-Ping; Cheng, Gui-Guang; Luo, Xiao-Dong; Cai, Xiang-Hai

2013-01-01

Seven new indole alkaloids, rauverines A-G (1-7), and 19 known indole alkaloids were isolated from the leaves and twigs of Rauvolfia verticillata. All compounds showed no cytotoxicity against five human cancer cell lines, human myeloid leukemia (HL-60), hepatocellular carcinoma (SMMC-7721), lung cancer (A-549), breast cancer (MCF-7), and colon cancer (SW480) cells.

Higher brightness beams from the SPS for the HL-LHC era

CERN Document Server

AUTHOR|(CDS)2085448; Bracco, Chiara (CERN)

The need to push the LHC beyond its limits and increase the deliverable luminosity to the experiments by about one order of magnitude has driven the ongoing injector and HL-LHC upgrades. The higher luminosity requires to increase the beam brightness, which directly translates in the need to adapt the different machine protection systems. Among all the foreseen upgrades, the transfer line collimators (TCDI) and the LHC injection protection systems will be revised. In particular, the guaranteed protection is evaluated in this Ph D work, together with the specification for the minimum shielded aperture in case of injection failures. A detailed model is also developed which insures a more reliable and efficient procedure for the validation of the TCDI setup within the required accuracy. The physics beyond colliders will also be pushed over its current limits in the HL-LHC era. SHiP, a new proposed fixed target experiment served by the SPS is under study. The unprecedented level of requested protons on target per ...

Transcriptional up-regulation of restin by all-trans retinoic acid through STAT1 in cancer cell differentiation process

International Nuclear Information System (INIS)

Fu Haiyan; Yang Guodong; Lu Fan; Wang Ruihua; Yao Libo; Lu Zifan

2006-01-01

RESTIN, a member of the melanoma-associated antigen superfamily, is a nuclear protein induced by atRA (all-trans retinoic acid) in HL60 cells. HeLa cells stably transfected with restin results in G1 cell cycle arrest. How this gene is regulated by atRA in the cell differentiation process is still unclear. In this study, we observed that up-regulation of restin was present during the atRA-induced HL60 cell differentiation process, suggesting the functional relevance between RESTIN and atRA-induced cellular effects. In order to further define the transcriptional regulation of restin by atRA, we analyzed the promoter region of restin. About 2.1 kb 5' flanking sequence of this gene was cloned into vector pGL3 and its core promoter region was identified through systemic deletions. Interestingly, restin promoter containing several potential consensus-binding sites of STAT-1α was activated by atRA in ER + MCF-7 cells but not in ER - MDA-MB-231 cells, over-expression of STAT-1α in latter rescued the activation effect of restin promoter in response to atRA and IFNγ. Our evidence supported that STAT-1α plays an important role in the atRA-induced transcriptional up-regulation of restin, which was associated with the atRA-induced HL60 cell differentiation and potentially mediated the downstream effects of atRA signal pathway via STAT-1α in some cancer cells

Spontaneous pyrogen production by mouse histiocytic and myelomonocytic tumor cell lines in vitro.

Science.gov (United States)

Bodel, P

1978-05-01

Tumor-associated fever occurs commonly in acute leukemias and lymphomas. We investigated the capacity for in vitro production of pyrogen by three mouse histiocytic lymphoma cell lines (J-774, PU5-1.8, p 388 D1), one myelomonoyctic line (WEHI-3), and tow lymphoma-derived lines, RAW-8 and R-8. Pyrogen was released spontaneously into the culture medium during growth by all cell lines with macrophage or myeloid characteristics including lysozyme production; R-8 cells, of presumed B-lymphocyte origin, did not produce pyrogen. When injected into mice, the pyrogens gave fever curves typical of endogenous pyrogen, were inactived by heating to 56 degrees C and by pronase digestion, and appeared to be secreted continuously by viable cells. Two pyrogenic molecular species produced by H-774 cells were identified by Sephadex filtration, one of mol wt approximately equal to 30,000, and the other greater than or equal to 60,000. By contrast, three carcinoma cell lines of human origin and SV-40 3T3 mouse fibroblasts did not produce pyrogen in vitro. These results suggest that some malignant cells derived from phagocytic cells of bone marrow origin retain their capacity for pyrogen production, and may spontaneously secrete pyrogen during growth.

Authentication of M14 melanoma cell line proves misidentification of MDA‐MB‐435 breast cancer cell line