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Sample records for jurkat cells effects

  1. Normal Untreated Jurkat Cells

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Biomedical research offers hope for a variety of medical problems, from diabetes to the replacement of damaged bone and tissues. Bioreactors, which are used to grow cells and tissue cultures, play a major role in such research and production efforts. The objective of the research was to define a way to differentiate between effects due to microgravity and those due to possible stress from non-optimal spaceflight conditions. These Jurkat cells, a human acute T-cell leukemia was obtained to evaluate three types of potential experimental stressors: a) Temperature elevation; b) Serum starvation; and c) Centrifugal force. The data from previous spaceflight experiments showed that actin filaments and cell shape are significantly different for the control. These normal cells serve as the baseline for future spaceflight experiments.

  2. [Effect of phenylhexyl isothiocyanate on Wnt/beta-catenin signaling pathway in Jurkat cell line].

    PubMed

    Lin, Juan; Huang, Yi-Qun; Ma, Xu-Dong

    2013-04-01

    This study was purposed to investigate the effect of phenylhexyl isothiocyanate (PHI) on Wnt/β-catenin signaling pathway, histone acetylation, histone methylation and cell apoptosis in Jurkat cell line. The viability of Jurkat cells after treatment with PHI was tested by MTT. Apoptotic rate of Jurkat cells was measured by flow cytometry. The levels of Wnt/β-catenin related proteins including β-catenin, TCF, c-myc, and cyclinD1, histone acetylated H3 and H4, histone methylated H3K9 and H3K4 were detected by Western blot. The results showed that PHI inhibited the cell growth and induced apoptosis in Jurkat cells in time-and dose-dependent manners. Its IC50 at 48 h was about 20 µmol/L. Expression of histone acetylated H3, H4 and histone methylated H3k4 increased after exposure to PHI for 3 h, while histone methylated H3K9 decreased. Expression of β-catenin was not changed after exposure to PHI for 3 h, but expression of β-catenin, and its cell cycle-related genes such as TCF, c-myc and cyclinD1 decreased after exposure to PHI for 7 h. It is concluded that PHI regulates acetylation and methylation of histone, inhibits Wnt/β-catenin signal pathway, and is able to induce apoptosis and inhibits growth of Jurkat cells. PMID:23628033

  3. Protective effects of certain pharmaceutical compounds against abrin induced cell death in Jurkat cell line.

    PubMed

    Saxena, Nandita; Rao, P V L; Bhaskar, A S B; Bhutia, Y D

    2014-08-01

    Abrin is a plant glycoprotein toxin from the seeds of Abrus precatorius, and shares the structure and properties with ricin. Abrin is highly toxic, with an estimated human fatal dose of 0.1-1 μg/kg, causing death after accidental and intentional poisoning. It is a potent toxin warfare agent. There are no antidotes available for abrin intoxication. It is becoming increasingly important to develop countermeasures for abrin by developing pre- and post-exposure medical therapy. The present study involves the screening of certain pharmaceutical agents for their potential to counter abrin toxicity in Jurkat T lymphocytes and the probable mechanism of action of the compounds with protective effect. The compounds studied are: Prednisolone, Minocycline, Amifostine, DRDE-07 (amifostine analog), Melatonin, Ebselen, N-Acetyl-l-cysteine (NAC) and Trolox. Among them, only NAC and trolox were found to confer significant protection in Jurkat cells by restoring antioxidant enzymes depleted by abrin treatment. Abrin also shown to increase in stress factor associated proteins SAPK/JNK, c-fos and c-jun levels which were effectively suppressed by NAC and trolox. In addition to this, both compounds significantly inhibit abrin induced inflammation and caspase-3 activity. These data suggest that NAC and trolox may serve as potential candidates for management of abrin-induced poisoning. PMID:24938881

  4. Effect of loop structure of bovine lactoferricin on apoptosis in Jurkat cells.

    PubMed

    Zhang, Tie-nan; Yang, Wei; Liu, Ning

    2010-06-01

    Bovine lactoferricin (LfcinB) is a cationic peptide that selectively induces apoptosis in Jurkat cells. However less is known about the influence of this kind of apoptosis on the intra-cellular ceramide metabolism and the structure-function relationship between the loop structure of LfcinB and its action of inducing apoptosis in Jurkat cells. In the present study, the artificially synthesized LfcinB and LfcinB-derived peptide (Cys 19 residue in LfcinB was replaced by Ala) was added in Jurkat cells, the nucleolus shape was observed by fluorescent microscopy, the ceramide concentration in Jurkat cells was determined by reversed phase high performance liquid chromatography (RP-HPLC). The results of MTT assay showed that LfcinB inhibited proliferation of Jurkat cells, and the inhibition rate was approximately 18.90%. Moreover, the inhibition rate of LfcinB together with MAPP was upto approximately 59.89%. The RP-HPLC result showed that LfcinB improved the ceramide level in Jurkat cells. By using the DNA fragmentation assay and observing the nucleolus shape, the result displayed deficiency of the loop structure could cause LfcinB losing the biological activity of inducing apoptosis in Jurkat cells. PMID:20237822

  5. Effects of Zizyphus lotus L. (Desf.) polyphenols on Jurkat cell signaling and proliferation.

    PubMed

    Abdoul-Azize, Souleymane; Bendahmane, Malika; Hichami, Aziz; Dramane, Gado; Simonin, Anne-Marie; Benammar, Chahid; Sadou, Hassimi; Akpona, Simon; El Boustani, Es-Saddik; Khan, Naim A

    2013-02-01

    We assessed the effects of Zizyphus lotus L. (Desf.) polyphenols (ZLP) on T-cell signaling and proliferation. Our results showed that ZLP exerted no effect on the increases in intracellular free calcium concentrations, [Ca(2+)]i, in human Jurkat T-cells. However, ZLP modulated the thapsigargin-induced increases in [Ca(2+)]i in these cells. ZLP treatment was found to decrease the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2). In addition, ZLP induced a rapid (t1/2=33s) and dose-dependent decrease in intracellular pH (pHi) in human Jurkat T-cells. Furthermore, ZLP significantly curtailed T-cell proliferation by diminishing their progression from S to G2/M phase of cell cycle, and the expression of interleukin-2 (IL-2) mRNA. Taken together, the results of the present study demonstrate that ZLP modulate cell signaling and exert immunosuppressive effects in human T-cells. PMID:23219580

  6. Probing Mechanical Properties of Jurkat Cells under the Effect of ART Using Oscillating Optical Tweezers

    PubMed Central

    2015-01-01

    Acute lymphoid leukemia is a common type of blood cancer and chemotherapy is the initial treatment of choice. Quantifying the effect of a chemotherapeutic drug at the cellular level plays an important role in the process of the treatment. In this study, an oscillating optical tweezer was employed to characterize the frequency-dependent mechanical properties of Jurkat cells exposed to the chemotherapeutic agent, artesunate (ART). A motion equation for a bead bound to a cell was applied to describe the mechanical characteristics of the cell cytoskeleton. By comparing between the modeling results and experimental results from the optical tweezer, the stiffness and viscosity of the Jurkat cells before and after the ART treatment were obtained. The results demonstrate a weak power-law dependency of cell stiffness with frequency. Furthermore, the stiffness and viscosity were increased after the treatment. Therefore, the cytoskeleton cell stiffness as the well as power-law coefficient can provide a useful insight into the chemo-mechanical relationship of drug treated cancer cells and may serve as another tool for evaluating therapeutic performance quantitatively. PMID:25928073

  7. Effects of nanosecond pulsed electrical fields (nsPEFs) on the cell cycle of CHO and Jurkat cells

    NASA Astrophysics Data System (ADS)

    Mahlke, Megan A.; Navara, Christopher; Ibey, Bennett L.

    2014-03-01

    Exposure to nano-second pulsed electrical fields (nsPEFs) can cause poration of external and internal cell membranes, DNA damage, and disassociation of cytoskeletal components, all of which are capable of disrupting a cell's ability to replicate. Variations between cell lines in membrane and cytoskeletal structure as well as in survival of nsPEF exposure should correspond to unique line-dependent cell cycle effects. Additionally, phase of cell cycle during exposure may be linked to differential sensitivities to nsPEFs across cell lines, as DNA structure, membrane elasticity, and cytoskeletal structure change dramatically during the cell cycle. Populations of Jurkat and Chinese Hamster Ovary (CHO) cells were examined post-exposure (10 ns pulse trains at 150kV/cm) by analysis of DNA content via propidium iodide staining and flow cytometric analysis at various time points (1, 6, and 12h post-exposure) to determine population distribution in cell cycle phases. Additionally, CHO and Jurkat cells were synchronized in G1/S and G2/M phases, pulsed, and analyzed to evaluate role of cell cycle phase in survival of nsPEFs. CHO populations recovered similarly to sham populations postnsPEF exposure and did not exhibit a phase-specific change in response. Jurkat cells exhibited considerable apoptosis/necrosis in response to nsPEF exposure and were unable to recover and proliferate in a manner similar to sham exposed cells. Additionally, Jurkat cells appear to be more sensitive to nsPEFs in G2/M phases than in G1/S phases. Recovery of CHO populations suggests that nsPEFs do not inhibit proliferation in CHO cells; however, inhibition of Jurkat cells post-nsPEF exposure coupled with preferential cell death in G2/M phases suggest that cell cycle phase during exposure may be an important factor in determining nsPEF toxicity in certain cell lines. Interestingly, CHO cells have a more robust and rigid cytoskeleton than Jurkat cells which is thought to contribute to their ability to

  8. Effect of light irradiation and sex hormones on jurkat T cells: 17beta-estradiol but not testosterone enhances UVA-induced cytotoxicity in Jurkat lymphocytes.

    PubMed

    Cohly, Hari H P; Graham-Evans, Barbara; Ndebele, Kenneth; Jenkins, John K; McMurray, Robert; Yan, Jian; Yu, Hongtao; Angel, Michael F

    2005-04-01

    In Eastern cultures, such as India, it is traditionally recommended that women but not men cover their heads while working in the scorching sun. The purpose of this pilot study was to determine whether there was any scientific basis for this cultural tradition. We examined the differential cytotoxic effects of ultraviolet A light (UVA) on an established T cell line treated with female and male sex hormones. CD4+ Jurkat T cells were plated in 96 well plates at 2 x 106 cells/ml and treated with 17beta-estradiol (EST) or testosterone (TE). These cells were irradiated by UVA light with an irradiance of 170 J/cm2 for 15min at a distance of 6 cm from the surface of the 96-well plate. Controls included cells not treated with hormones or UVA. The effects of EST and TE were investigated between 1 and 20 ng/mL. Cytotoxicity by fluorescein-diacetate staining and COMET assay generating single strand DNA cleavage, tail length and tail moment measurements were examined. The effect of estrogen (5ng/mL) on apoptosis and its mediators was further studied using DNA laddering and western blotting for bcl-2 and p53. We found that EST alone, without UVA, enhanced Jurkat T cell survival. However, EST exhibited a dose-related cytotoxicity in the presence of UVA; up to 28% at 20 ng/ml. TE did not alter UVA-induced cytotoxicity. Since TE did not alter cell viability in the presence of UVA further damaging studies were not performed. COMET assay demonstrated the harmful effects of EST in the presence of UVA while EST without UVA. had no significant effect on the nuclear damage. Apoptosis was not present as indicated by the absence of DNA laddering on agarose gel electrophoresis at 5ng/ml EST or TE +/- UVA. Western blot showed that estrogen down regulated bcl-2 independently of UVA radiation while p53 was down regulated in the presence of UVA treatment. EST and TE have differential effects on UVA-induced cytotoxicity in Jurkat T-lymphocyte which suggested that women may be more susceptible

  9. Cytotoxic effect of inositol hexaphosphate and its Ni(II) complex on human acute leukemia Jurkat T cells.

    PubMed

    de Lima, Eliane May; Kanunfre, Carla Cristine; de Andrade, Lucas Ferrari; Granato, Daniel; Rosso, Neiva Deliberali

    2015-12-01

    Inositol hexaphosphate (InsP6) is present in cereals, legumes, nuts and seed oils and is biologically active against some tumor and cancer cells. Herein, this study aimed at evaluating the cellular toxicity, antiproliferative activity and effects on cell cycle progression of free InsP6 and InsP6-Ni(II) of leukemic T (Jurkat) and normal human cells. Treatments with InsP6 at concentrations between 1.0 and 4.0mM significantly decreased the viability of Jurkat cells, but showed no cytotoxic effect on normal human lymphocytes. Treatment with InsP6-Ni(II) complex at concentrations between 0.05 and 0.30 mM showed an anti-proliferative dose and a time-dependent effect, with significantly reduced cell viability of Jurkat cells but showed no cytotoxic effect on normal human lymphocytes as compared to the control. Ni(II) free ion was toxic to normal cells while InsP6-Ni(II) had no cytotoxic effect. The InsP6-Ni(II) complex potentiated (up to 10×) the antiproliferative effect of free InsP6 on Jurkat cells. The cytometric flow assay showed that InsP6 led to an accumulation of cells in the G0/G1 phase of the cell cycle, accompanied by a decrease in the number of cells in S and G2/M phases, whereas InsP6-Ni(II) has led to an accumulation of cells in the S and G2/M phases. Our findings showed that InsP6-Ni(II) potentiates cytotoxic effects of InsP6 on Jurkat cells and may be a potential adjuvant in the treatment of cancer. PMID:26335902

  10. Effect of sphingosine on Ca2+ entry and mitochondrial potential of Jurkat T cells--interaction with Bcl2.

    PubMed

    Dangel, Georg Richard; Lang, Florian; Lepple-Wienhues, Albrecht

    2005-01-01

    Triggers of Jurkat T cell apoptosis include sphingosine and ceramide. Sphingosine and ceramide further inhibit capacitative Ca2+ entry (ICRAC), an effect leading to inactivation but not death of Jurkat T cells. Mitochondria are key organelles in the machinery leading to apoptosis and on the other hand have been shown to participate in the regulation of Ca2+ entry. The present experiments were performed to explore whether treatment of Jurkat T cells with sphingosine leads to apoptosis and reduced Ca2+ entry and whether those effects are sensitive to expression of the antiapoptotic protein Bcl2, localized in the outer mitochondrial membrane. Exposure of Jurkat T cells to 10 microM spingosine was according to DiOC6 fluorescence followed by mitochondrial depolarization and according to Fura-red/Fluo-3 fluorescence followed by decreased capacitative Ca2+ entry. Mitochondrial depolarization was significantly delayed in cells overexpressing wild type Bcl2 or Bcl2 targeted to the mitochondrial membrane, whereas no significant influence on mitochondrial depolarization was observed in cells expressing Bcl2 lacking the membrane targeting motif or Bcl2 targeted to the endoplasmatic reticulum. In contrast to mitochondrial potential, the blunting of capacitative Ca2+ entry following sphingosine treatment was not sensitive to mitochondrial Bcl2 expression. In conclusion sphingosine exposure leads to both, mitochondrial depolarization and inhibition of capacitative Ca2+ entry. Mitochondrial Bcl2 reverses the effect on mitochondria but not on Ca2+ entry and thus leads to dissociation of those two sequelae of sphingosine treatment. PMID:16121028

  11. Differential effects of mercury, lead, and cadmium on IL-2 production by Jurkat T cells.

    PubMed

    Colombo, Myrian; Hamelin, Claudine; Kouassi, Edouard; Fournier, Michel; Bernier, Jacques

    2004-06-01

    Mercury, lead, and cadmium are widespread and highly toxic pollutants. The aim of this study was to determine the effects of sublethal doses of CH(3)HgCl, CdCl(2), and PbCl(2) on IL-2 production by T lymphocytes. Jurkat T cells were stimulated by triggering CD3 and CD28 molecules before, in conjunction with, or following heavy metal exposure. Heavy metals, individually or mixed together at equimolar concentrations, were used. Results demonstrated that low, noncytotoxic doses of metals induce tyrosine phosphorylation. Mercury and lead (1 microM) inhibit IL-2 production regardless of the state of T cell activation. Cadmium stimulated IL-2 production only in preactivated T cells. Surprisingly, a mixture of these three metals had no effect. We subsequently determined the effects of heavy metals on NFAT (nuclear factors of activated T cells) activity. When cells were stimulated by potent stimulation involving the CD3 and CD28 molecules, an increased NFAT activation was noted when the cells were exposed to mercury and to the metal mixture. Activation with PMA/calcium ionophores indicated that the target of heavy metals is located downstream from PKC and calcium mobilization. These results suggest that the state and mode of T cell activation are important parameters to consider in heavy metal toxicity. PMID:15183152

  12. Effects of in vitro Brevetoxin Exposure on Apoptosis and Cellular Metabolism in a Leukemic T Cell Line (Jurkat)

    PubMed Central

    Walsh, Catherine J.; Leggett, Stephanie R.; Strohbehn, Kathryn; Pierce, Richard H.; Sleasman, John W.

    2008-01-01

    Harmful algal blooms (HABs) of the toxic dinoflagellate, Karenia brevis, produce red tide toxins, or brevetoxins. Significant health effects associated with red tide toxin exposure have been reported in sea life and in humans, with brevetoxins documented within immune cells from many species. The objective of this research was to investigate potential immunotoxic effects of brevetoxins using a leukemic T cell line (Jurkat) as an in vitro model system. Viability, cell proliferation, and apoptosis assays were conducted using brevetoxin congeners PbTx-2, PbTx-3, and PbTx-6. The effects of in vitro brevetoxin exposure on cell viability and cellular metabolism or proliferation were determined using trypan blue and MTT (1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan), respectively. Using MTT, cellular metabolic activity was decreased in Jurkat cells exposed to 5 – 10 μg/ml PbTx-2 or PbTx-6. After 3 h, no significant effects on cell viability were observed with any toxin congener in concentrations up to 10 μg/ml. Viability decreased dramatically after 24 h in cells treated with PbTx-2 or -6. Apoptosis, as measured by caspase-3 activity, was significantly increased in cells exposed to PbTx-2 or PbTx-6. In summary, brevetoxin congeners varied in effects on Jurkat cells, with PbTx-2 and PbTx-6 eliciting greater cellular effects compared to PbTx-3. PMID:18728729

  13. Effect of UV irradiation on the apoptosis and necrosis of Jurkat cells using UV LEDs

    NASA Astrophysics Data System (ADS)

    Inada, Shunko A.; Amano, Hiroshi; Akasaki, Isamu; Morita, Akimichi; Kobayashi, Keiko

    2009-02-01

    Phototherapy is a very effective method for treating most of the incurable skin diseases. A fluorescent light bulb is used as a conventional UV light source for this type of therapy. However, infrared radiation from the light source sometimes causes serious problems on patient's health. In addition, the normal part of the skin is irradiated when a large fluorescent light bulb is used. Moreover, a conventional UV irradiation system is heavy and has a short lifetime and a high electrical power consumption. Therefore, a new UV light source for solving the problems of phototherapy is required. To realize low-power-consumption, lightweight and long-lifetime systems, group III nitride-based UV-A1 light-emitting diodes (LEDs) were investigated. We examined the UV LED irradiation of Jurkat cell, which is a tumor cell and more sensitive to UV light than a healthy cell. The numbers of apoptotic and necrotic cells were confirmed to be the same using a UV LED and a conventional lamp system. The UV LED showed the possibility of realizing a new UV light source for phototherapy.

  14. Effect of silencing HOXA5 gene expression using RNA interference on cell cycle and apoptosis in Jurkat cells

    PubMed Central

    HUANG, HUI-PING; LIU, WEN-JUN; GUO, QU-LIAN; BAI, YONG-QI

    2016-01-01

    Acute lymphocytic leukemia (ALL) is a common malignant tumor with a high morbidity rate among children, accounting for approximately 80% of leukemia cases. Although there have been improvements in the treatment of patients frequent relapse lead to a poor prognosis. The aim of the present study was to determine whether HOXA5 may be used as a target for gene therapy in leukemia in order to provide a new treatment. Mononuclear cells were extracted from the bone marrow according to the clinical research aims. After testing for ALL in the acute stage, the relative mRNA and protein expression of HOXA5 was detected in the ALL remission groups (n=25 cases per group) and the control group [n=20 cases, immune thrombocytopenia (ITP)]. Gene silencing by RNA interference (RNAi) was used to investigate the effect of silencing HOXA5 after small interfering RNA (siRNA) transfection to Jurkat cells. The HOXA5-specific siRNA was transfected to Jurkat cells using lipofectamine. The experiment was divided into the experimental group (liposomal transfection of HOXA5 targeting siRNA), the negative control group (liposomal transfection of cells with negative control siRNA) and the control group (plus an equal amount of cells and culture media only). Western blotting and quantitative fluorescent polymerase chain reaction (QF-PCR) were used to detect the relative HOXA5 mRNA expression and protein distribution in each cell group. Cell distribution in the cell cycle and the rate of cells undergoing apoptosis were determined using flow cytometry. The expression of HOXA5 at the mRNA and protein levels in the acute phase of ALL was significantly higher than that in ALL in the remission and control groups. In cells transfected with HOXA5-specific siRNA, the expression of HOXA5 at the mRNA and protein levels decreased significantly (P<0.05). The distribution of cells in the cell cycle was also altered. Specifically, more cells were present in the G0/G1 phase compared to the S phase (P<0.05). In

  15. Pro-apoptotic effect of Persea americana var. Hass (avocado) on Jurkat lymphoblastic leukemia cells.

    PubMed

    Bonilla-Porras, Angelica R; Salazar-Ospina, Andrea; Jimenez-Del-Rio, Marlene; Pereañez-Jimenez, Andres; Velez-Pardo, Carlos

    2013-11-01

    Abstract Context: Therapy for leukemia has a limited efficacy. There is a need to search for alternative anti-leukemia therapies. Persea americana Mill var. Hass (Lauraceae) is a tropical fruit (avocado) that might be used against cancer. Objective: To investigate whether P. americana induces death in Jurkat lymphoblastic leukemia cells. Materials and methods: Four ethanol extracts (0.1, 0.5, 1, 2 and 5 mg/mL) from avocado fruit (endocarp, whole seed, seed and leaves) were analyzed against Jurkat cells. Hydrogen peroxide generation by oxidation of 2',7'-dichlorodihydrofluorescein diacetate to the fluorescent compound 2',7'-dichlorfluorescein assay, acridine orange/ethidium bromide staining, flow cytometry analysis of annexin-V/7-amino-actinomycin, mitochondrial membrane potential and immunocytochemistry detection of transcription factor p53, caspase-3 and apoptosis-inducing factor (AIF) were evaluated. Results: Endocarp, seed, whole seed, and leaf (0.1 mg/mL) extracts induced significant apoptosis in Jurkat cells (p < 0.001) in an oxidative stress-dependent fashion via mitochondrial membrane depolarization (52-87%), activation of transcription factor p53 (6.3-25.4%), protease caspase-3 (8.3-20%) and predominance of AIF reactivity (20.6-36%) in all extracts. Similar results were obtained with 0.5 mg/mL extracts. However, extract ≥1 mg/mL concentration induced necrosis (100%). Conclusions: P. americana extracts function as a pro-apoptotic compound. Leukemic cells are eliminated through an oxidative stress mechanism. This study contributes to the understanding of the molecular mechanism of the avocado and its therapeutic action on leukemia. PMID:24188375

  16. Effects of valproic acid and pioglitazone on cell cycle progression and proliferation of T-cell acute lymphoblastic leukemia Jurkat cells

    PubMed Central

    Jazi, Marie Saghaeian; Mohammadi, Saeed; Yazdani, Yaghoub; Sedighi, Sima; Memarian, Ali; Aghaei, Mehrdad

    2016-01-01

    Objective(s): T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignant tumor. Administration of chemical compounds influencing apoptosis and T cell development has been discussed as promising novel therapeutic strategies. Valproic acid (VPA) as a recently emerged anti-neoplastic histone deacetylase (HDAC) inhibitor and pioglitazone (PGZ) as a high-affinity peroxisome proliferator-activated receptor-gamma (PPARγ) agonist have been shown to induce apoptosis and cell cycle arrest in different studies. Here, we aimed to investigate the underlying molecular mechanisms involved in anti-proliferative effects of these compounds on human Jurkat cells. Materials and Methods: Treated cells were evaluated for cell cycle progression and apoptosis using flowcytometry and MTT viability assay. Real-time RT-PCR was carried out to measure the alterations in key genes associated with cell death and cell cycle arrest. Results: Our findings illustrated that both VPA and PGZ can inhibit Jurkat E6.1 cells in vitro after 24 hr; however, PGZ 400 μM presents the most anti-proliferative effect. Interestingly, treated cells have been arrested in G2/M with deregulated cell division cycle 25A (Cdc25A) phosphatase and cyclin-dependent kinase inhibitor 1B (CDKN1B or p27) expression. Expression of cyclin D1 gene was inhibited when DNA synthesis entry was declined. Cell cycle deregulation in PGZ and VPA-exposed cells generated an increase in the proportion of aneuploid cell population, which has not reported before. Conclusion: These findings define that anti-proliferative effects of PGZ and VPA on Jurkat cell line are mediated by cell cycle deregulation. Thus, we suggest PGZ and VPA may relieve potential therapeutic application against apoptosis-resistant malignancies.

  17. The synergistic repressive effect of NF-κB and JNK inhibitor on the clonogenic capacity of Jurkat leukemia cells.

    PubMed

    Liu, Xinli; Zhang, Jun; Li, Jing; Volk, Andrew; Breslin, Peter; Zhang, Jiwang; Zhang, Zhou

    2014-01-01

    Deregulation of Nuclear Transcription Factor-κB (NF-κB) and Jun N-terminal kinase (JNK) signaling is commonly detected in leukemia, suggesting an important role for these two signaling pathways in the pathogenesis of leukemia. In this study, using Jurkat cells, an acute T-lymphoblastic leukemia (T-ALL) cell line, we evaluated the effects of an NF-κB inhibitor and a JNK inhibitor individually and in combination on the proliferation, survival and clonogenic capacity of leukemic cells. We found that leukemic stem/progenitor cells (LSPCs) were more sensitive to NF-κB inhibitor treatment than were healthy hematopoietic stem/progenitor cells (HSPCs), as shown by a reduction in the clonogenic capacity of the former. Inactivation of NF-κB leads to the activation of JNK signaling in both leukemic cells and healthy HSPCs. Interestingly, JNK inhibitor treatment enhanced the repressive effects of NF-κB inhibitor on LSPCs but prevented such repression in HSPCs. Our data suggest that JNK signaling stimulates proliferation/survival in LSPCs but is a death signal in HSPCs. The combination of NF-κB inhibitor and JNK inhibitor might provide a better treatment for T-ALL leukemia by synergistically killing LSPCs while simultaneously preventing the death of normal HPCs. PMID:25526629

  18. The Synergistic Repressive Effect of NF-κB and JNK Inhibitor on the Clonogenic Capacity of Jurkat Leukemia Cells

    PubMed Central

    Liu, Xinli; Zhang, Jun; Li, Jing; Volk, Andrew; Breslin, Peter; Zhang, Jiwang; Zhang, Zhou

    2014-01-01

    Deregulation of Nuclear Transcription Factor-κB (NF-κB) and Jun N-terminal kinase (JNK) signaling is commonly detected in leukemia, suggesting an important role for these two signaling pathways in the pathogenesis of leukemia. In this study, using Jurkat cells, an acute T-lymphoblastic leukemia (T-ALL) cell line, we evaluated the effects of an NF-κB inhibitor and a JNK inhibitor individually and in combination on the proliferation, survival and clonogenic capacity of leukemic cells. We found that leukemic stem/progenitor cells (LSPCs) were more sensitive to NF-κB inhibitor treatment than were healthy hematopoietic stem/progenitor cells (HSPCs), as shown by a reduction in the clonogenic capacity of the former. Inactivation of NF-κB leads to the activation of JNK signaling in both leukemic cells and healthy HSPCs. Interestingly, JNK inhibitor treatment enhanced the repressive effects of NF-κB inhibitor on LSPCs but prevented such repression in HSPCs. Our data suggest that JNK signaling stimulates proliferation/survival in LSPCs but is a death signal in HSPCs. The combination of NF-κB inhibitor and JNK inhibitor might provide a better treatment for T-ALL leukemia by synergistically killing LSPCs while simultaneously preventing the death of normal HPCs. PMID:25526629

  19. Minerval induces apoptosis in Jurkat and other cancer cells

    PubMed Central

    Llado, Victoria; Gutierrez, Antonio; Martínez, Jordi; Casas, Jesús; Terés, Silvia; Higuera, Mónica; Galmés, Antonio; Saus, Carles; Besalduch, Joan; Busquets, Xavier; Escribá, Pablo V

    2010-01-01

    Abstract Minerval is an oleic acid synthetic analogue that impairs lung cancer (A549) cell proliferation upon modulation of the plasma membrane lipid structure and subsequent regulation of protein kinase C localization and activity. However, this mechanism does not fully explain the regression of tumours induced by this drug in animal models of cancer. Here we show that Minerval also induced apoptosis in Jurkat T-lymphoblastic leukaemia and other cancer cells. Minerval inhibited proliferation of Jurkat cells, concomitant with a decrease of cyclin D3 and cdk2 (cyclin-dependent kinase2). In addition, the changes that induced on Jurkat cell membrane organization caused clustering (capping) of the death receptor Fas (CD95), caspase-8 activation and initiation of the extrinsic apoptosis pathway, which finally resulted in programmed cell death. The present results suggest that the intrinsic pathway (associated with caspase-9 function) was activated downstream by caspase-8. In a xenograft model of human leukaemia, Minerval also inhibited tumour progression and induced tumour cell death. Studies carried out in a wide variety of cancer cell types demonstrated that apoptosis was the main molecular mechanism triggered by Minerval. This is the first report on the pro-apoptotic activity of Minerval, and in part explains the effectiveness of this non-toxic anticancer drug and its wide spectrum against different types of cancer. PMID:19413889

  20. Galectin-1 and Galectin-3 induce mitochondrial apoptotic pathway in Jurkat cells

    NASA Astrophysics Data System (ADS)

    Vasil'eva, O. A.; Isaeva, A. V.; Prokhorenko, T. S.; Zima, A. P.; Novitsky, V. V.

    2016-08-01

    Cellular malignant transformation is often accompanied by increased gene expression of low-molecular proteins of lectins family-galectins. But it is unknown how galectins promote tumor growth and malignization. Galectins-1 and galectin-3 are thought to be possible immunoregulators exerting their effects by regulating the balance of CD4+ lymphocytes. In addition it is known that tumor cells overexpressing galectins are capable of escaping immunological control, causing apoptosis of lymphocytes. The aim of the study is to investigate the role of galectin-1 and galectin-3 in the implementation of mitochondrial apoptotic pathway in Jurkat cells. Methods: Jurkat cells were used as a model for the study of T-lymphocytes. Jurkat cells were activated with antibodies to CD3 and CD28 and cultured with recombinant galectin-1 and -3. Apoptosis of Jurkat cells and depolarization of the mitochondrial membrane were assessed by flow cytometry. It was found that galectin-1 and galectin-3 have a dose-dependent pro-apoptotic effect on Jurkat cells in vitro and enlarge the number of cells with decreased mitochondrial membrane potential compared with intact cells.

  1. Escin sodium induces apoptosis of human acute leukemia Jurkat T cells.

    PubMed

    Zhang, Zhenzhen; Gao, Jian; Cai, Xueting; Zhao, Youlong; Wang, Yafei; Lu, Wuguang; Gu, Zhenhua; Zhang, Shuangquan; Cao, Peng

    2011-12-01

    Escin sodium has been used in the clinic as an antioedematous, antiexudative and vasoprotective agent for many years and has shown excellent tolerability. However, little is known about its anticancer activity. This is a report for the first time that escin sodium exerts a cytotoxic effect on human acute leukemia Jurkat T cells via the induction of apoptosis rather than cell cycle arrest. Escin sodium activated the initiator caspase-8, -9, and the effector caspase-3, degraded poly (ADP-ribose) polymerase (PARP) and attenuated the expression of Bcl-2. In addition, escin sodium inhibited the growth of cancer cells in a selective manner with Jurkat cells most sensitive to it. Taken together, the data show that escin sodium possesses potent apoptogenic activity toward human acute leukemia Jurkat T cells. PMID:21452372

  2. On cell signalling mechanism of Mycobacterium leprae soluble antigen (MLSA) in Jurkat T cells.

    PubMed

    Joshi, Beenu; Khedouci, Sihem; Dagur, Pradeep Kumar; Hichami, Aziz; Sengupta, Utpal; Khan, Naim Akhtar

    2006-07-01

    We investigated the role of Mycobaterium leprae soluble antigen (MLSA) in the modulation of calcium signalling, phosphorylation of mitogen-activated protein (MAP) kinases and IL-2 mRNA expression in human Jurkat T cells. We observed that MLSA induced an increase in free intracellular calcium concentrations, [Ca2+]i, via opening CRAC (Ca2+-release activated- Ca2+) channels. Furthermore, MLSA failed to potentiate both thapsigargin- and anti-CD3 antibodies-induced capacitative calcium influx in Jurkat T cells. We observed that MLSA failed to affect the degree of phosphorylation of two MAP kinases, i.e., ERK1/ERK2, stimulated by anti-CD3 antibodies alone or phorbol 12-myristate 13-acetate (PMA) alone. In order to mimic co-stimulation of T cells, we stimulated them by both PMA and anti-CD3 antibodies. MLSA significantly curtailed the phosphorylation of ERK1/ERK2, stimulated by both PMA and anti-CD3 antibodies in Jurkat T cells. Also MLSA was found to reduce the transcription of IL-2 gene in PMA plus anti-CD3 antibodies-activated Jurkat T cells. Our finding demonstrates that Ca2+ influx via CRAC channels, inhibition of ERK1/ERK2 phosphorylation and IL-2 gene transcription may be implicated in immunosuppressive effects of MLSA antigen. PMID:16583135

  3. GENOTOXIC MECHANISMS OF ARSENIC TRIOXIDE IN HUMAN JURKAT T-LYMPHOMA CELLS.

    PubMed

    Yedjou, Clement; Sutton, La'mont; Tchounwou, Paul

    2008-01-01

    Arsenic trioxide (As(2)O(3)) has cytotoxic effects on several cancer cell lines. However, the molecular mechanisms of action remain to be elucidated. Hence, the aim of the present study was to evaluate the cytotoxicity and genotoxicity induced by As(2)O(3) in a human Jurkat T-lymphoma cell line using the trypan blue exclusion test and alkaline single cell gel electrophoresis (Comet) assays, respectively. Jurkat T-cells were treated with different doses of As(2)O(3) for 24 and 48 h prior to cytogenetic assessment. Data obtained from the trypan blue exclusion test indicated that As(2)O(3) significantly (p < 0.05) reduced the viability of Jurkat T-cells in a dose and time-dependent manner. Data generated from the comet assay also indicated a significant dose and time-dependent increase in DNA damage in Jurkat T-cells associated with As(2)O(3) exposure. We observed a significant increase (P < 0.05) in comet tail-length, tail arm and tail moment, as well as in percentages of DNA cleavage at all doses tested, showing an evidence As(2)O(3) -induced genotoxic damage in Jurkat T-cells. This study confirms that the comet assay is a sensitive and effective method to detect DNA damage caused by heavy metals such as arsenic. Taken together, our findings suggest that As(2)O(3) exposure significantly (p < 0.05) reduces cellular viability and induces DNA damage in human Jurkat T-lymphoma cells. PMID:21796259

  4. Antiproliferative Activity of T. welwitschii Extract on Jurkat T Cells In Vitro

    PubMed Central

    Moyo, Batanai; Mukanganyama, Stanley

    2015-01-01

    Triumfetta welwitschii is a plant used traditionally for the treatment of fever and diarrhoea. Previous work has shown that T. welwitschii has antibacterial activity. The purpose of this study was to investigate T. welwitschii extract for anticancer activity against Jurkat T cells. The Jurkat T cell line is used to study acute T cell leukaemia. An antiproliferation assay, determination of induction of apoptosis, the determination of the effect of the combination of the extract and GSH, and effects of the extract on DNA leakage were conducted. T. welwitschii was found to decrease cell viability in a dose- and time-dependent manner. T. welwitschii caused apoptosis in the Jurkat T cells as shown by DNA fragmentation. When T. welwitschii was combined with reduced GSH, it was found that the growth of the Jurkat T cells was significantly reduced compared to untreated cells after 72 h of treatment. This was unexpected, as cancer cells have elevated levels of GSH compared to normal cells. The results of this study show that T. welwitschii is a potential source of compounds that may serve as leads for anticancer compounds. PMID:26557698

  5. RHAMNAZIN INHIBITS PROLIFERATION AND INDUCES APOPTOSIS OF HUMAN JURKAT LEUKEMIA CELLS IN VITRO.

    PubMed

    Philchenkov, A A; Zavelevych, M P

    2015-01-01

    Antiproliferative and apoptogenic effects of rhamnazin, a dimethoxylated derivative of quercetin, were studied in human acute lymphoblastic leukemia Jurkat cells. The cytotoxicity and apoptogenic activity of rhamnazin in vitro are inferior to that of quercetin. The apoptogenic activity of rhamnazin is realized via mitochondrial pathway and associated with activation of caspase-9 and -3. The additive apoptogenic effect of rhamnazin and suboptimal doses of etoposide, a DNA topoisomerase II inhibitor, is demonstrated. Therefore, methylation of quercetin modifies its biological effects considerably. PMID:27025066

  6. Constituents of French Marigold (Tagetes patula L.) Flowers Protect Jurkat T-Cells against Oxidative Stress.

    PubMed

    Chkhikvishvili, Irakli; Sanikidze, Tamar; Gogia, Nunu; Enukidze, Maia; Machavariani, Marine; Kipiani, Nana; Vinokur, Yakov; Rodov, Victor

    2016-01-01

    The flowers of French marigold (Tagetes patula L.) are widely used in folk medicine, in particular for treating inflammation-related disorders. However, cellular mechanisms of this activity demand further investigation. In the present work, we studied the potential of T. patula compounds to alleviate the oxidative stress in hydrogen peroxide-challenged human lymphoblastoid Jurkat T-cells. Crude extracts of marigold flowers and purified fractions containing flavonoids patuletin, quercetagetin, and quercetin and their derivatives, as well as the carotenoid lutein, were brought in contact with Jurkat cells challenged with 25 or 50 μM H2O2. Hydrogen peroxide caused oxidative stress in the cells, manifested as generation of superoxide and peroxyl radicals, reduced viability, arrested cell cycle, and enhanced apoptosis. The stress was alleviated by marigold ingredients that demonstrated high radical-scavenging capacity and enhanced the activity of antioxidant enzymes involved in neutralization of reactive oxygen species. Flavonoid fraction rich in quercetin and quercetagetin showed the highest cytoprotective activity, while patuletin in high dose exerted a cytotoxic effect associated with its anticancer potential. T. patula compounds enhanced the production of anti-inflammatory and antioxidant interleukin-10 (IL-10) in Jurkat cells. Both direct radical-scavenging capacity and stimulation of protective cellular mechanisms can underlay the anti-inflammatory properties of marigold flowers. PMID:27433287

  7. Constituents of French Marigold (Tagetes patula L.) Flowers Protect Jurkat T-Cells against Oxidative Stress

    PubMed Central

    Chkhikvishvili, Irakli; Sanikidze, Tamar; Gogia, Nunu; Enukidze, Maia; Machavariani, Marine; Kipiani, Nana; Vinokur, Yakov; Rodov, Victor

    2016-01-01

    The flowers of French marigold (Tagetes patula L.) are widely used in folk medicine, in particular for treating inflammation-related disorders. However, cellular mechanisms of this activity demand further investigation. In the present work, we studied the potential of T. patula compounds to alleviate the oxidative stress in hydrogen peroxide-challenged human lymphoblastoid Jurkat T-cells. Crude extracts of marigold flowers and purified fractions containing flavonoids patuletin, quercetagetin, and quercetin and their derivatives, as well as the carotenoid lutein, were brought in contact with Jurkat cells challenged with 25 or 50 μM H2O2. Hydrogen peroxide caused oxidative stress in the cells, manifested as generation of superoxide and peroxyl radicals, reduced viability, arrested cell cycle, and enhanced apoptosis. The stress was alleviated by marigold ingredients that demonstrated high radical-scavenging capacity and enhanced the activity of antioxidant enzymes involved in neutralization of reactive oxygen species. Flavonoid fraction rich in quercetin and quercetagetin showed the highest cytoprotective activity, while patuletin in high dose exerted a cytotoxic effect associated with its anticancer potential. T. patula compounds enhanced the production of anti-inflammatory and antioxidant interleukin-10 (IL-10) in Jurkat cells. Both direct radical-scavenging capacity and stimulation of protective cellular mechanisms can underlay the anti-inflammatory properties of marigold flowers. PMID:27433287

  8. Effects of parathyroid hormone-related protein and macrophage inflammatory protein-1α in Jurkat T-cells on tumor formation in vivo and expression of apoptosis regulatory genes in vitro

    PubMed Central

    Shu, Sherry T.; Dirksen, Wessel P.; Lanigan, Lisa G.; Martin, Chelsea K.; Thudi, Nanda K.; Werbeck, Jillian L.; Fernandez, Soledad A.; Hildreth, Blake E.; Rosol, Thomas J.

    2012-01-01

    Parathyroid hormone-related protein (PTHrP) and macrophage inflammatory protein-1α (MIP-1α) have been implicated in the pathogenesis of adult T-cell leukemia/lymphoma, but their effects on T-cells have not been well studied. Here we analyzed the functions of PTHrP and MIP-1α on T-cell growth and death both in vitro and in vivo by overexpressing either factor in human Jurkat T-cells. PTHrP or MIP-1α did not affect Jurkat cell growth in vitro, but PTHrP increased their sensitivity to apoptosis. Importantly, PTHrP and MIP-1α decreased both tumor incidence and growth in vivo. To investigate possible mechanisms, polymerase chain reaction (PCR) arrays and real-time reverse transcription (RT)-PCR assays were performed. Both PTHrP and MIP-1α increased the expression of several factors including signal transducer and activator of transcription 4, tumor necrosis factor α, receptor activator of nuclear factor κB ligand and death-associated protein kinase 1, and decreased the expression of inhibitor of DNA binding 1, interferon γ and CD40 ligand in Jurkat cells. In addition, MIP-1α also increased the expression of transcription factor AP-2α and PTHrP increased expression of the vitamin D3 receptor. These data demonstrate that PTHrP and MIP-1α exert a profound antitumor effect presumably by increasing the sensitivity to apoptotic signals through modulation of transcription and apoptosis factors in T-cells. PMID:21942940

  9. Umbelliprenin from Ferula szowitsiana Activates both Intrinsic and Extrinsic Pathways of Apoptosis in Jurkat T-CLL cell line.

    PubMed

    Gholami, Omid; Jeddi-Tehrani, Mahmood; Iranshahi, Mehrdad; Zarnani, Amir Hassan; Ziai, Seyed Ali

    2013-01-01

    Umbelliprenin is a prenylated compound, which belongs to the class of sesquiterpene coumarins. It is extracted from dried roots of Ferula szwitsiana collected from the mountains of Golestan forest (Golestan Province, north of Iran). Induction of apoptosis in Jurkat T-CLL cells has been previously shown. In this study, effect of umbelliprenin on proapoptotic caspases (caspase-8 and -9) and antiapoptotic Bcl-2 family protein was studied. Jurkat cells were incubated with umbelliprenin. Cells were then lysed and activation of proteins was studied by Western blot analysis. In this study, we showed that umbelliprenin activates intrinsic and extrinsic pathways of apoptosis by the activation of caspase-8 and -9 respectively. Inhibition of Bcl-2 was also shown. In conclusion, umbelliprenin induced apoptosis in Jurkat cells through caspase-dependent apoptosis pathway. PMID:24250644

  10. Umbelliprenin from Ferula szowitsiana Activates both Intrinsic and Extrinsic Pathways of Apoptosis in Jurkat T-CLL cell line

    PubMed Central

    Gholami, Omid; Jeddi-Tehrani, Mahmood; Iranshahi, Mehrdad; Zarnani, Amir Hassan; Ziai, Seyed Ali

    2013-01-01

    Umbelliprenin is a prenylated compound, which belongs to the class of sesquiterpene coumarins. It is extracted from dried roots of Ferula szwitsiana collected from the mountains of Golestan forest (Golestan Province, north of Iran). Induction of apoptosis in Jurkat T-CLL cells has been previously shown. In this study, effect of umbelliprenin on proapoptotic caspases (caspase-8 and -9) and antiapoptotic Bcl-2 family protein was studied. Jurkat cells were incubated with umbelliprenin. Cells were then lysed and activation of proteins was studied by Western blot analysis. In this study, we showed that umbelliprenin activates intrinsic and extrinsic pathways of apoptosis by the activation of caspase-8 and -9 respectively. Inhibition of Bcl-2 was also shown. In conclusion, umbelliprenin induced apoptosis in Jurkat cells through caspase-dependent apoptosis pathway. PMID:24250644

  11. Carvacrol induces mitochondria-mediated apoptosis in HL-60 promyelocytic and Jurkat T lymphoma cells.

    PubMed

    Bhakkiyalakshmi, Elango; Suganya, Natarajan; Sireesh, Dornadula; Krishnamurthi, Kannan; Saravana Devi, Sivanesan; Rajaguru, Palanisamy; Ramkumar, Kunka Mohanram

    2016-02-01

    The aim of the present study was to investigate the effect of carvacrol, a phenolic monoterpenoid on the induction of apoptosis in HL-60 (Human acute promyelocytic leukemia cells) and Jurkat (human T lymphocyte cells) cells. Carvacrol showed a potent cytotoxic effect on both cells with dose-dependent increase in the level of free radical formation as measured by an oxidation sensitive fluorescent dye, 2,7-dichlorodihydrofluorescein diacetate (H2DCFDA) levels. The reduction in the level of antioxidants such as catalase (CAT) and superoxide dismutase (SOD) (P<0.05) was observed in carvacrol-treated cells. The major cytotoxic effect appears to be intervened by the induction of apoptotic cell death as assessed by annexin-V labeling assay using flow cytometry. Western blot analysis showed that Bax expression was increased, whereas Bcl-2 expression was significantly decreased in carvacrol exposed HL-60 cells and Jurkat cells. Further studies revealed that the dissipation of mitochondrial membrane potential of intact cells was accompanied by the activation of caspase-3. Our results found that the potential mechanism of cellular apoptosis induced by carvacrol is mediated by caspase-3 and is associated with the collapse of mitochondrial membrane potential, generation of free radicals, and depletion of the intracellular antioxidant pool. PMID:26724845

  12. Inhibition of CXCL12-mediated chemotaxis of Jurkat cells by direct immunotoxicants.

    PubMed

    Shao, Jia; Stout, Inge; Volger, Oscar L; Hendriksen, Peter J M; van Loveren, Henk; Peijnenburg, Ad A C M

    2016-07-01

    Directional migration of cells to specific locations is required in tissue development, wound healing, and immune responses. Immune cell migration plays a crucial role in both innate and adaptive immunity. Chemokines are small pro-inflammatory chemoattractants that control the migration of leukocytes. In addition, they are also involved in other immune processes such as lymphocyte development and immune pathology. In a previous toxicogenomics study using the Jurkat T cell line, we have shown that the model immunotoxicant TBTO inhibited chemotaxis toward the chemokine CXCL12. In the present work, we aimed at assessing a novel approach to detecting chemicals that affect the process of cell migration. For this, we first evaluated the effects of 31 chemicals on mRNA expression of genes that are known to be related to cell migration. With this analysis, seven immunotoxicants were identified as potential chemotaxis modulators, of which five (CoCl2 80 µM, MeHg 1 µM, ochratoxin A 10 µM, S9-treated ochratoxin A 10 µM, and TBTO 100 nM) were confirmed as chemotaxis inhibitor in an in vitro trans-well chemotaxis assay using the chemokine CXCL12. The transcriptome data of the five compounds together with previously obtained protein phosphorylation profiles for two out of five compounds (i.e., ochratoxin A and TBTO) revealed that the mechanisms behind the chemotaxis inhibition are different for these immunotoxicants. Moreover, the mTOR inhibitor rapamycin had no effect on the chemotaxis of Jurkat cells, indicating that the mTOR pathway is not involved in CXCL12-mediated chemotaxis of Jurkat cells, which is opposite to the findings on human primary T cells (Munk et al. in PLoS One 6(9):e24667, 2011). Thus, the results obtained from the chemotaxis assay conducted with Jurkat cells might not fully represent the results obtained with human primary T cells. Despite this difference, the present study indicated that some compounds may exert their immunotoxic effects through

  13. Andrographolide inhibits growth of human T-cell acute lymphoblastic leukemia Jurkat cells by downregulation of PI3K/AKT and upregulation of p38 MAPK pathways.

    PubMed

    Yang, Tingfang; Yao, Shuluan; Zhang, Xianfeng; Guo, Yan

    2016-01-01

    T-cell acute lymphoblastic leukemia (T-ALL) as a prevalent hematologic malignancy is one of the most common malignant tumors worldwide in children. Andrographolide (Andro), the major active component from Andrographis paniculata, has been shown to possess antitumor activities in several types of cancer cells. However, whether Andro would inhibit T-ALL cell growth remains unclear. In this study, we investigated the cytotoxic effect of Andro on human T-ALL Jurkat cells and explored the mechanisms of cell death. Cell apoptosis was assayed by flow cytometry, and the signaling transduction for Andro was analyzed by Western blotting. The results indicated 10 μg/mL Andro could significantly induce Jurkat cells' apoptosis, depending on the inhibition of PI3K/AKT pathway. Moreover, Andro-induced apoptosis is enhanced by AKT-selective inhibitor LY294002. ERK- or JNK-selective inhibitors PD98059 and SP600125 had no effect on Andro-induced apoptosis. In addition, p38 inhibitor SB203580 could reverse Andro-induced apoptosis in Jurkat cells. We also found that the protein expression of p-p53 and p-p38 were increased after Andro treatments. The result of an in vivo study also demonstrated Andro's dose-dependent inhibition in subcutaneous Jurkat xenografts. In conclusion, our findings explained a novel mechanism of drug action by Andro in Jurkat cells and suggested that Andro might be developed into a new candidate therapy for T-ALL patients in the coming days. PMID:27114702

  14. A synthesized nostocionone derivative potentiates programmed cell death in human T-cell leukemia Jurkat cells through mitochondria via the release of endonuclease G.

    PubMed

    Itoh, Tomohiro; Muramatsu, Yuji; Masu, Masayo; Tsuge, Ayaka; Taniguchi, Masaki; Ninomiya, Masayuki; Ando, Masashi; Tsukamasa, Yasuyuki; Koketsu, Mamoru

    2014-01-01

    Nostocionone (Nost), a compound isolated from Nostoc commune, and its synthesized derivatives (NostDs) were evaluated for in vitro cytotoxicity against human T-cell leukemia Jurkat cells. NostD3 [(1E,4E)-1-(3,4-dihydroxyphenyl)-5-(2,6,6-trimethylcyclohex-1-enyl)penta-1,4-dien-3-one] inhibited cell growth more potently than Nost. To elucidate the mechanisms of NostD3-induced cell death, we examined changes in cell morphology, the loss of mitochondrial membrane potential (MMT), and DNA fragmentation. From these results, the cytotoxic effects of NostD3 were found to be mainly due to Type I programmed cell death (PCDI; i.e., apoptosis). Using caspase inhibitors, we further found that NostD-3-induced PCDI occurred through a caspase-independent pathway. Moreover, NostD3 decreased MMT and modulated multiple signaling molecules (MAPKs, Akt, Bcl-2, Bax, and c-Myc) in Jurkat cells, thereby inducing the release of endonuclease G (Endo-G) from mitochondria. The level of intracellular reactive oxygen species (ROS) in cells treated with NostD3 was elevated up to 1 h after the treatment. However, suppression of ROS by N-acetyl-l-cysteine restored Jurkat cell growth. Taken together, our data suggested that ROS production acted as a trigger in NostD3-induced PCDI in Jurkat cells through release of Endo-G from the mitochondria. PMID:25333640

  15. Vimentin Is Involved in Peptidylarginine Deiminase 2-Induced Apoptosis of Activated Jurkat Cells

    PubMed Central

    Hsu, Pei-Chen; Liao, Ya-Fan; Lin, Chin-Li; Lin, Wen-Hao; Liu, Guang-Yaw; Hung, Hui-Chih

    2014-01-01

    Peptidylarginine deiminase type 2 (PADI2) deiminates (or citrullinates) arginine residues in protein to citrulline residues in a Ca2+-dependent manner, and is found in lymphocytes and macrophages. Vimentin is an intermediate filament protein and a well-known substrate of PADI2. Citrullinated vimentin is found in ionomycin-induced macrophage apoptosis. Citrullinated vimentin is the target of anti-Sa antibodies, which are specific to rheumatoid arthritis, and play a critical role in the pathogenesis of the disease. To investigate the role of PADI2 in apoptosis, we generated a Jurkat cell line that overexpressed the PADI2 transgene from a tetracycline-inducible promoter, and used a combination of 12-O-tetradecanoylphorbol-13-acetate and ionomycin to activate Jurkat cells. We found that PADI2 overexpression reduced the cell viability of activated Jurkat cells in a dose- and time-dependent manner. The PADI2-overexpressed and -activated Jurkat cells presented typical manifestations of apoptosis, and exhibited greater levels of citrullinated proteins, including citrullinated vimentin. Vimentin overexpression rescued a portion of the cells from apoptosis. In conclusion, PADI2 overexpression induces apoptosis in activated Jurkat cells. Vimentin is involved in PADI2-induced apoptosis. Moreover, PADI2-overexpressed Jurkat cells secreted greater levels of vimentin after activation, and expressed more vimentin on their cell surfaces when undergoing apoptosis. Through artificially highlighting PADI2 and vimentin, we demonstrated that PADI2 and vimentin participate in the apoptotic mechanisms of activated T lymphocytes. The secretion and surface expression of vimentin are possible ways of autoantigen presentation to the immune system. PMID:24850148

  16. Retama monosperma n-hexane extract induces cell cycle arrest and extrinsic pathway-dependent apoptosis in Jurkat cells

    PubMed Central

    2014-01-01

    Background Retama monosperma L. (Boiss.) or Genista monosperma L. (Lam.), locally named as “R’tam”, is an annual and spontaneous plant belonging to the Fabaceae family. In Morocco, Retama genus is located in desert regions and across the Middle Atlas and it has been widely used in traditional medicine in many countries. In this study, we show that Retama monosperma hexane extract presents significant anti-leukemic effects against human Jurkat cells. Methods Human Jurkat cells, together with other cell lines were screened with different concentrations of Retama monosperma hexane extract at different time intervals. Growth inhibition was determined using luminescent-based viability assays. Cell cycle arrest and apoptosis were measured by flow cytometry analysis. Combined caspase 3 and 7 activities were measured using luminometric caspase assays and immunoblots were performed to analyze expression of relevant pro- and anti-apoptotic proteins. GC-MS were used to determine the chemical constituents of the active extract. Results Retama monosperma hexane extract (Rm-HE) showed significant cytotoxicity against Jurkat cells, whereas it proved to be essentially ineffective against both normal mouse fibroblasts (NIH3T3) and normal lymphocytes (TK-6). Cytometric analysis indicated that Rm-HE promoted cell cycle arrest and apoptosis induction accompanied by DNA damage induction indicated by an increase in p-H2A.X levels. Rm-HE induced apoptosis was partially JNK-dependent and characterized by an increase in Fas-L levels together with activation of caspases 8, 3, 7 and 9, whereas neither the pro-apoptotic nor anti-apoptotic mitochondrial membrane proteins analyzed were significantly altered. Chemical identification analysis indicated that α-linolenic acid, campesterol, stigmasterol and sitosterol were the major bioactive components within the extract. Conclusions Our data suggest that bioactive compounds present in Rm-HE show significant anti leukemic activity inducing

  17. Osmocenyl-tamoxifen derivatives target the thioredoxin system leading to a redox imbalance in Jurkat cells.

    PubMed

    Scalcon, Valeria; Top, Siden; Lee, Hui Zhi Shirley; Citta, Anna; Folda, Alessandra; Bindoli, Alberto; Leong, Weng Kee; Salmain, Michèle; Vessières, Anne; Jaouen, Gérard; Rigobello, Maria Pia

    2016-07-01

    The synthesis and the biological effects of two ferrocifen analogs in the osmium series, namely the monophenolic complex 1, the tamoxifen-like complex 2 and their oxidized quinone methide (QM) derivatives, 1-QM and 2-QM, are reported. Inhibition of purified thioredoxin reductase (TrxR) is observed with 1 and 2 only after their enzymatic oxidation by the hydrogen peroxide/horseradish peroxidase (H2O2/HRP) system with IC50 of 2.4 and 1.2μM respectively. However, this inhibition is larger than that obtained with the corresponding quinone methides (IC50=5.4μM for 1-QM and 3.6μM for 2-QM). The UV-Vis spectra of 1 or 2 incubated in the presence of H2O2/HRP show that the species generated is not a quinone methide, but probably the corresponding cation. In Jurkat cells, 2 shows high toxicity (IC50=7.4μM), while 1 is less effective (IC50=42μM). Interestingly, a significant inhibition of TrxR activity is observed in cells incubated with 2 (about 70% inhibition with 15μM) while the inhibition induced by 1 is much weaker (about 30% inhibition with 50μM). This strong inhibition of TrxR by 2 leads to accumulation of thioredoxin and peroxiredoxin 3 in oxidized form and to a decrease of the mitochondrial membrane potential (MMP). These results show that cytotoxicity of the osmocifens depends on their oxidation within the cell and that inhibition of thioredoxin reductase by oxidized species is a key factor in rationalizing the cytotoxicity of these complexes on Jurkat cells. PMID:27130146

  18. Combined treatment with fenretinide and indomethacin induces AIF-mediated, non-classical cell death in human acute T-cell leukemia Jurkat cells

    SciTech Connect

    Hojka-Osinska, Anna; Ziolo, Ewa; Rapak, Andrzej

    2012-03-16

    Highlights: Black-Right-Pointing-Pointer The combination of fenretinide and indomethacin induces a high level of cell death. Black-Right-Pointing-Pointer Apoptotic pathway is caspase-independent. Black-Right-Pointing-Pointer Jurkat cells undergo AIF-mediated cell death. -- Abstract: Currently used cytotoxic drugs in cancer therapy have a similar mechanism of action and low specificity. Applied simultaneously, they show an additive effect with strong side effects. Clinical trials with the use of different agents in cancer therapy show that the use of these compounds alone is not very effective in fighting cancer. An alternative solution could be to apply a combination of these agents, because their combination has a synergistic effect on some cancer cells. Therefore, in our investigations we examined the effects of a synthetic retinoid-fenretinide when combined with a non-steroidal anti-inflammatory drug-indomethacin on the process of apoptosis in the acute human T-cell leukemia cell line Jurkat. We demonstrate that treatment with the combination of the tested compounds induces the death of cells, that is peculiar and combines features of apoptosis as well as non-apoptotic cell death. In detail we observed, cell membrane permeabilization, phosphatydylserine exposure, no oligonucleosomal DNA fragmentation, no caspase-3 activation, but apoptosis inducing factor (AIF) nuclear translocation. Taken together these results indicate, that Jurkat cells after treatment with a combination of fenretinide and indomethacin undergo AIF-mediated programmed cell death.

  19. Gamma radiation alters cell cycle and induces apoptosis in p53 mutant E6.1 Jurkat cells.

    PubMed

    Ahmadianpour, Mohammad Reza; Abdolmaleki, Parviz; Mowla, Seyed Javad; Hosseinkhani, Saman

    2013-01-01

    This study aimed at investigating the effect of gamma radiation with 1.5, 3.0 and 7.5 Gy doses on apoptosis induction, cell cycle alteration and increment of amount of p-ATM (phosphorylated ATM) and p-E2F1 (phosphorylated E2F1) proteins in Jurkat T-lymphoblastoid E6.1 cells. Exposure of human p53 mutant Jurkat cells to gamma radiation resulted in apoptosis, which was detected by luminometric and flow cytometric analysis. Also, phosphorylated ATM (ataxia telangiectasia mutated) and E2F1 (elongation factor) proteins were detected by western blot analysis. Based on luminescence detection data the lethal dose of 7.5 Gy induced cell death 12 h after exposure (p<0.05) while sub-lethal doses of 1.5 and 3.0 Gy induced apoptosis 18 h after exposure (p<0.05). Flow cytometric analysis revealed a G2 arrest 24h after exposure to 3.0 and 7.5 Gy. This arrest was accompanied by cell death with an increasing rate of occurrence up to 72 h after exposure. Western blot analysis showed that 1 h after cell irradiation by 1.5, 3.0 and 7.5 Gy, the amount of p-ATM increased to its maximum rate and remained constant up to 6 h, and then it decreased. Moreover, the amount of phosphorylated E2F1 (Ser-31) increased 2 h after exposure to the same doses and remained constant up to 12 h after irradiation. Survival and cell division of treated Jurkat cells showed a decrease compared to the control group. We believe that ionizing radiation-induced DNA damage activates a p53-independent apoptosis pathway via back-up systems in which the phosphorylation of ATM and E2F1 proteins was involved. Thus, gamma radiation can induce apoptosis and cell cycle alteration in Jurkat cells via a P53-independent pathway. PMID:23079488

  20. Lithium Induces ER Stress and N-Glycan Modification in Galactose-Grown Jurkat Cells

    PubMed Central

    Kátai, Emese; Yahiro, Rikki K. K.; Poór, Viktor S.; Montskó, Gergely; Zrínyi, Zita; Kovács, Gábor L.; Miseta, Attila

    2013-01-01

    We previously reported that lithium had a significant impact on Ca2+ regulation and induced unfolded protein response (UPR) in yeast cells grown on galactose due to inhibition of phosphoglucomutase (PGM), however the exact mechanism has not been established yet. In this study, we analysed lithium's effect in galactose-fed cells to clarify whether these ER-related changes are the result of a relative hypoglycemic state. Furthermore, we investigated whether the alterations in galactose metabolism impact protein post-translational modifications. Thus, Jurkat cells were incubated in glucose or galactose containing media with or without lithium treatment. We found that galactose-fed and lithium treated cells showed better survivability than fasting cells. We also found higher UDP-Hexose and glycogen levels in these cells compared to fasting cells. On the other hand, the UPR (X-box binding protein 1 mRNA levels) of galactose-fed and lithium treated cells was even greater than in fasting cells. We also found increased amount of proteins that contained N-linked N-acetyl-glucosamine, similar to what was reported in fasting cells by a recent study. Our results demonstrate that lithium treatment of galactose-fed cells can induce stress responses similar to hypoglycemia, however cell survival is still secured by alternative pathways. We propose that clarifying this process might be an important addition toward the better understanding of the molecular mechanisms that regulate ER-associated stress response. PMID:23894652

  1. Fe2O3 nanoparticles suppress Kv1.3 channels via affecting the redox activity of Kvβ2 subunit in Jurkat T cells

    NASA Astrophysics Data System (ADS)

    Yan, Li; Liu, Xiao; Liu, Wei-Xia; Tan, Xiao-Qiu; Xiong, Fei; Gu, Ning; Hao, Wei; Gao, Xue; Cao, Ji-Min

    2015-12-01

    Superparamagnetic iron oxide nanoparticles (SPIONs) are promising nanomaterials in medical practice due to their special magnetic characteristics and nanoscale size. However, their potential impacts on immune cells are not well documented. This study aims to investigate the effects of Fe2O3 nanoparticles (Fe2O3-NPs) on the electrophysiology of Kv1.3 channels in Jurkat T cells. Using the whole-cell patch-clamp technique, we demonstrate that incubation of Jurkat cells with Fe2O3-NPs dose- and time-dependently decreased the current density and shifted the steady-state inactivation curve and the recovery curve of Kv1.3 channels to a rightward direction. Fe2O3-NPs increased the NADP level but decreased the NADPH level of Jurkat cells. Direct induction of NADPH into the cytosole of Jurkat cells via the pipette abolished the rightward shift of the inactivation curve. In addition, transmission electron microscopy showed that Fe2O3-NPs could be endocytosed by Jurkat cells with relatively low speed and capacity. Fe2O3-NPs did not significantly affect the viability of Jurkat cells, but suppressed the expressions of certain cytokines (TNFα, IFNγ and IL-2) and interferon responsive genes (IRF-1 and PIM-1), and the time courses of Fe2O3-NPs endocytosis and effects on the expressions of cytokines and interferon responsive genes were compatible. We conclude that Fe2O3-NPs can be endocytosed by Jurkat cells and act intracellularly. Fe2O3-NPs decrease the current density and delay the inactivation and recovery kinetics of Kv1.3 channels in Jurkat cells by oxidizing NADPH and therefore disrupting the redox activity of the Kvβ2 auxiliary subunit, and as a result, lead to changes of the Kv1.3 channel function. These results suggest that iron oxide nanoparticles may affect T cell function by disturbing the activity of Kv1.3 channels. Further, the suppressing effects of Fe2O3-NPs on the expressions of certain inflammatory cytokines and interferon responsive genes suggest that iron

  2. [Comparison of protein expression profiles between bortezomib-resistant JurkatB cells with PSMB5 mutation and their parent cells].

    PubMed

    Lü, Shu-Qing; Yang, Jian-Min; Huang, Chong-Mei; Xu, Xiao-Qian; Zhou, Hong; Song, Ning-Xia; Wang, Jian-Min

    2011-08-01

    This study was purposed to investigate the differences of cyto biological characteristics and protein expression levels between bortezomib-resistant T-lymphoblastic lymphoma/leukemia cell lines JurkatB containing PSMB5 G322A mutation and their parent cell line Jurkat, The cytotoxicities of bortezomib and chemotherapeutic drugs to JurkatB5 cells (end selection concentration of bortezomib was 500 nmol/L), JurkatB8 (end selection concentration 800 nmol/L) and Jurkat cells were analyzed. The cell growth curves were drawn with viable cell counts by trypan blue assay, the colony formation rate were assayed by soft-agar colony culture, and the cell distributions in cell cycle were analyzed by flow cytometry, mRNA expression levels of multidrug resistance (MDR) genes MDR1, LRP and MRP were measured by real-time fluorescence quantitative RT-PCR, the differences of protein expression levels were detected by SpringBio antibody microarray containing 720 proteins. The results showed that the drug resistance multiples for 48 hours of JurkatB5 and JurkatB8 cells (relative to Jurkat) to bortezomib were increased by 33.52 and 39.04 times, respectively. JurkatB5 and JurkatB8 cells did not display significant cross-resistance to daunorubicin, adriamycin, vindesine, and etoposide after exposure for 48 hours. There were no significant differences in the cell growth curve, colony formation rate and cell distributions in cell cycle between JurkatB5, JurkatB8 and Jurkat cells (p > 0.05). There were no significant differences of mRNA expression levels of MDR1, LRP, MRP between JurkatB5 and Jurkat cells (p > 0.05). There were 264 analyzable expression points detected by antibody microarray. Among them, 252 protein expression levels were not significantly different between JurkatB5, JurkatB8 and Jurkat cells (< 2-fold), including 15 drug resistance-related proteins. 12 proteins were detected at higher or lower expression levels in JurkatB5 or JurkatB8 cells then that in Jurkat cells (cell

  3. Andrographolide inhibits growth of human T-cell acute lymphoblastic leukemia Jurkat cells by downregulation of PI3K/AKT and upregulation of p38 MAPK pathways

    PubMed Central

    Yang, Tingfang; Yao, Shuluan; Zhang, Xianfeng; Guo, Yan

    2016-01-01

    T-cell acute lymphoblastic leukemia (T-ALL) as a prevalent hematologic malignancy is one of the most common malignant tumors worldwide in children. Andrographolide (Andro), the major active component from Andrographis paniculata, has been shown to possess antitumor activities in several types of cancer cells. However, whether Andro would inhibit T-ALL cell growth remains unclear. In this study, we investigated the cytotoxic effect of Andro on human T-ALL Jurkat cells and explored the mechanisms of cell death. Cell apoptosis was assayed by flow cytometry, and the signaling transduction for Andro was analyzed by Western blotting. The results indicated 10 μg/mL Andro could significantly induce Jurkat cells’ apoptosis, depending on the inhibition of PI3K/AKT pathway. Moreover, Andro-induced apoptosis is enhanced by AKT-selective inhibitor LY294002. ERK- or JNK-selective inhibitors PD98059 and SP600125 had no effect on Andro-induced apoptosis. In addition, p38 inhibitor SB203580 could reverse Andro-induced apoptosis in Jurkat cells. We also found that the protein expression of p-p53 and p-p38 were increased after Andro treatments. The result of an in vivo study also demonstrated Andro’s dose-dependent inhibition in subcutaneous Jurkat xenografts. In conclusion, our findings explained a novel mechanism of drug action by Andro in Jurkat cells and suggested that Andro might be developed into a new candidate therapy for T-ALL patients in the coming days. PMID:27114702

  4. Comprehensive RNA dataset of AGO2 associated RNAs in Jurkat cells following miR-21 over-expression.

    PubMed

    Carissimi, Claudia; Colombo, Teresa; Azzalin, Gianluca; Cipolletta, Emanuela; Laudadio, Ilaria; Macino, Giuseppe; Fulci, Valerio

    2016-06-01

    We set out to identify miR-21 targets in Jurkat cells using a high-throughput biochemical approach (10.1016/j.biochi.2014.09.021[1]). Using a specific monoclonal antibody raised against AGO2, RISC complexes were immunopurified in Jurkat cells over-expressing miR-21 following lentiviral trasduction as well as in Jurkat control cells lines. A parallel immunoprecipitation using isotype-matched rat IgG was performed as a control. AGO2 associated mRNAs were profiled by microarray (GEO: GSE37212). AGO2 bound miRNAs were profiled by RNA-seq. PMID:27054165

  5. Ceramide inhibits PKCθ by regulating its phosphorylation and translocation to lipid rafts in Jurkat cells.

    PubMed

    Hage-Sleiman, Rouba; Hamze, Asmaa B; El-Hed, Aimée F; Attieh, Randa; Kozhaya, Lina; Kabbani, Sarah; Dbaibo, Ghassan

    2016-08-01

    Protein kinase C theta (PKCθ) is a novel, calcium-independent member of the PKC family of kinases that was identified as a central player in T cell signaling and proliferation. Upon T cell activation by antigen-presenting cells, PKCθ gets phosphorylated and activated prior to its translocation to the immunological synapse where it couples with downstream effectors. PKCθ may be regulated by ceramide, a crucial sphingolipid that is known to promote differentiation, growth arrest, and apoptosis. To further investigate the mechanism, we stimulated human Jurkat T cells with either PMA or anti-CD3/anti-CD28 antibodies following induction of ceramide accumulation by adding exogenous ceramide, bacterial sphingomyelinase, or Fas ligation. Our results suggest that ceramide regulates the PKCθ pathway through preventing its critical threonine 538 (Thr538) phosphorylation and subsequent activation, thereby inhibiting the kinase's translocation to lipid rafts. Moreover, this inhibition is not likely to be a generic effect of ceramide on membrane reorganization. Other lipids, namely dihydroceramide, palmitate, and sphingosine, did not produce similar effects on PKCθ. Addition of the phosphatase inhibitors okadaic acid and calyculin A reversed the inhibition exerted by ceramide, and this suggests involvement of a ceramide-activated protein phosphatase. Such previously undescribed mechanism of regulation of PKCθ raises the possibility that ceramide, or one of its derivatives, and may prove valuable in novel therapeutic approaches for disorders involving autoimmunity or excessive inflammation-where PKCθ plays a critical role. PMID:26798039

  6. Enhanced histamine production through the induction of histidine decarboxylase expression by phorbol ester in Jurkat cells.

    PubMed

    Nagashima, Yusuke; Kako, Koichiro; Kim, Jun-Dal; Fukamizu, Akiyoshi

    2012-11-01

    Histamine (HA), a mediator of inflammation, type I allergic responses and neurotransmission, is synthesized from L-histidine, the reaction of which is catalyzed by histidine decarboxylase (HDC). HDC has been reported to be induced by various stimuli, not only in mast cells and basophils, but also in T lymphocytes and macrophages. Although its mRNA has been shown to be increased in Jurkat cells when treated with phorbol 12-myristate 13-acetate (TPA), little is known concerning the induced production of HA by HDC. The present study quantified the trace amounts of intracellular HA using ultra-high liquid chromatography in combination with the 6-aminoquinoline carbamate-derivatization technique. To test whether the cellular level of HA is elevated by the induction of HDC in Jurkat cells treated with TPA, the peak corresponding to authentic HA in the cell lysate was fractioned and its molecular weight determined by matrix-assisted laser desorption/ionization quadrupole ion trap time-of-flight mass spectrometry. The results of this study show that the HA level is increased by the induction of HDC expression by TPA in Jurkat cells. Therefore, this method is useful in elucidating the physiological significance of HA production. PMID:22940786

  7. Static magnetic field of 6 mT induces apoptosis and alters cell cycle in p53 mutant Jurkat cells.

    PubMed

    Ahmadianpour, Mohammad Reza; Abdolmaleki, Parviz; Mowla, Seyed Javad; Hosseinkhani, Saman

    2013-03-01

    This study aimed to investigate the effect of 6 milliTesla (mT) static magnetic field (SMF) on apoptosis induction and cell cycle alteration in T-lymphoblastoid Jurkat E6.1 cells. Exposure of human p53 mutant Jurkat cells to 6 mT SMF resulted in apoptosis, which was detected by luminometric and flow cytometric analysis also, phosphorylated ATM and E2F1 proteins were detected by western blot analysis. Based on luminescence detection data, apoptosis initiated 36 h after exposure to 6 mT SMF. Apoptosis also reached its maximum rate 48 h after treatment. Flow cytometric analysis revealed a temporary G2 arrest after exposure to 6 mT SMF. Indeed, cellular population of S and G2 phases was increased. Based on reports of other investigations on the effect of magnetic fields on Ca2+flux changes in cell membranes and the effect of MFs on free radical formation, it can be suggested that the magnetic fields may induce the apoptosis and alter the cell population in different cell cycle phases of Jurkat cells via changing the Ca2+fluxes through cell membranes and playing a role in free radical formation. Western blot analysis showed that the amount of phosphorylated ATM and E2F1 proteins were increased in treated cells. The results of luminometric and flow cytometric detection did not show a significant difference in the apoptosis rate between 6 h-treated and 24 h-treated cells by 6 mT SMF. Thus, 6 mT SMF can induce apoptosis and alter cell cycle in Jurkat cells via a p53-independent pathway. PMID:23320430

  8. Diversity and Complexity of Ceramide Generation After Exposure of Jurkat Leukemia Cells to Irradiation

    SciTech Connect

    Ardail, Dominique Maalouf, Mira; Boivin, Anthony; Chapet, Olivier; Bodennec, Jacques; Rousson, Robert; Rodriguez-Lafrasse, Claire

    2009-03-15

    Purpose: To define which intracellular pools of sphingomyelin and ceramide are involved in the triggering of apoptosis of Jurkat leukemia cells in response to {gamma}-ray exposure. Methods and Materials: We examined the kinetics of ceramide generation at the whole-cell level and in different subcellular compartments (plasma membrane rafts, mitochondria, and endoplasmic reticulum) after irradiation with photons. Ceramide was measured by high-performance liquid chromatography or after pulse labeling experiments, and the presence of sphingomyelinase within mitochondria was assessed by electron microscopy. Results: Irradiation of Jurkat leukemia cells resulted in the sequential triggering of sphingomyelin hydrolysis, followed by de novo synthesis that led to a late ceramide response (from 24 h) correlated with the triggering of apoptosis. At the subcellular level, pulse-label experiments, using [{sup 3}H]-palmitate as a precursor, strengthened the involvement of the radiation-induced sphingomyelin breakdown and revealed a very early peak (15 min) of ceramide in plasma membrane rafts. A second peak in mitochondria was measured 4 h after irradiation, resulting from an increase of the sphingomyelin content relating to the targeting of acid sphingomyelinase toward this organelle. Conclusion: These data confirm that ceramide is a major determinant in the triggering of radiation-induced apoptosis and highlight the complexity of the sequential compartment-specific ceramide-mediated response of Jurkat leukemia cells to {gamma}-rays.

  9. Catharanthus roseus Aqueous Extract is Cytotoxic to Jurkat Leukaemic T-cells but Induces the Proliferation of Normal Peripheral Blood Mononuclear Cells

    PubMed Central

    Ahmad, Nor Hazwani; Rahim, Rohanizah Abdul; Mat, Ishak

    2010-01-01

    Research on natural products has been widely used as a strategy to discover new drugs with potential for applications in complementary medicines because they have fewer side effects than conventional drugs. The aim of the present study was to evaluate the in vitro cytotoxic effects of crude aqueous Catharanthus roseus extract on Jurkat cells and normal peripheral blood mononuclear cells (PBMCs). The aqueous extract was standardised to vinblastine by high performance liquid chromatography (HPLC) and was used to determine cytotoxicity by the MTS [3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay. DNA fragmentation assay was employed to determine if cell death was due to apoptosis. The results showed that the aqueous extract induced cell death of Jurkat cells at 24, 48 and 72 hours post-treatment in a time- and dose-dependent manner. However, cells treated at 48 and 72 hours produced higher cytotoxic effects with half maximal inhibitory concentration (IC50) values of 2.55 μg/ml and 2.38 μg/ml, respectively. In contrast, the extract induced normal PBMC proliferation, especially after 24 hours treatment with 1000 μg/ml. This result indicates that the C. roseus crude aqueous extract showed differential effects of inhibiting the proliferation of the Jurkat cell line and promoting the growth of PBMCs. These data suggest that the extract may be applicable for modulating the normal and transformed immune cells in leukaemia patients. PMID:24575203

  10. Microarray dataset of Jurkat cells following miR-93 over-expression.

    PubMed

    Gioiosa, Silvia; Verduci, Lorena; Azzalin, Gianluca; Carissimi, Claudia; Fulci, Valerio; Macino, Giuseppe

    2016-09-01

    The dataset presented here represents a microarray experiment of Jurkat cell line over-expressing miR-93 after lentiviral transgenic construct transduction. Three biological replicates have been performed. We further provide normalized and processed data, log2 Fold Change based ranked list and GOterms resulting table. The raw microarray data are available in the ArrayExpress database (www.ebi.ac.uk/arrayexpress) under accession number ArrayExpress: E-MTAB-4588. PMID:27408928

  11. Rosmarinic Acid-Rich Extracts of Summer Savory (Satureja hortensis L.) Protect Jurkat T Cells against Oxidative Stress

    PubMed Central

    Chkhikvishvili, Irakli; Sanikidze, Tamar; Gogia, Nunu; Mchedlishvili, Tamar; Enukidze, Maia; Machavariani, Marine; Vinokur, Yakov

    2013-01-01

    Summer savory (Satureja hortensis L., Lamiaceae) is used in several regions of the world as a spice and folk medicine. Anti-inflammatory and cytoprotective effects of S. hortensis and of its rosmarinic acid-rich phenolic fraction have been demonstrated in animal trials. However, previous studies of rosmarinic acid in cell models have yielded controversial results. In this study, we investigated the effects of summer savory extracts on H2O2-challenged human lymphoblastoid Jurkat T cells. LC-MS analysis confirmed the presence of rosmarinic acid and flavonoids such as hesperidin and naringin in the phenolic fraction. Adding 25 or 50 µM of H2O2 to the cell culture caused oxidative stress, manifested as generation of superoxide and peroxyl radicals, reduced cell viability, G0/G1 arrest, and enhanced apoptosis. This stress was significantly alleviated by the ethanolic and aqueous extracts of S. hortensis and by the partially purified rosmarinic acid fraction. The application of an aqueous S. hortensis extract doubled the activity of catalase and superoxide dismutase in the cells. The production of IL-2 and IL-10 interleukins was stimulated by H2O2 and was further enhanced by the addition of the S. hortensis extract or rosmarinic acid fraction. The H2O2-challenged Jurkat cells may serve as a model for investigating cellular mechanisms of cytoprotective phytonutrient effects. PMID:24349613

  12. Clausmarin A, Potential Immunosuppressant Revealed by Yeast-Based Assay and Interleukin-2 Production Assay in Jurkat T Cells

    PubMed Central

    Suauam, Pitipreya; Yingyongnarongkul, Boon-ek; Palaga, Tanapat; Miyakawa, Tokichi; Yompakdee, Chulee

    2015-01-01

    Small-molecule inhibitors of Ca2+-signaling pathways are of medicinal importance, as exemplified by the immunosuppressants FK506 and cyclosporin A. Using a yeast-based assay devised for the specific detection of Ca2+-signaling inhibitors, clausmarin A, a previously reported terpenoid coumarin, was identified as an active substance. Here, we investigated the likely mechanism of clausmarin A action in yeast and Jurkat T-cells. In the presence of 100 mM CaCl2 in the growth medium of Ca2+-sensitive Δzds1 strain yeast, clausmarin A exhibited a dose-dependent alleviation of various defects due to hyperactivation of Ca2+ signaling, such as growth inhibition, polarized bud growth and G2 phase cell-cycle arrest. Furthermore, clausmarin A inhibited the growth of Δmpk1 (lacking the Mpk1 MAP kinase pathway) but not Δcnb1 (lacking the calcineurin pathway) strain, suggesting that clausmarin A inhibited the calcineurin pathway as presumed from the synthetic lethality of these pathways. Furthermore, clausmarin A alleviated the serious defects of a strain expressing a constitutively active form of calcineurin. In the human Jurkat T-cell line, clausmarin A exhibited a dose-dependent inhibition of IL-2 production and IL-2 gene transcription, as well as an inhibition of NFAT dephosphorylation. The effects of clausmarin A observed in both yeast and Jurkat cells are basically similar to those of FK506. Our study revealed that clausmarin A is an inhibitor of the calcineurin pathway, and that this is probably mediated via inhibition of calcineurin phosphatase activity. As such, clausmarin A is a potential immunosuppressant. PMID:26313553

  13. Gene expression profile of Jurkat cells exposed to high power terahertz radiation

    NASA Astrophysics Data System (ADS)

    Grundt, Jessica E.; Roth, Caleb C.; Rivest, Benjamin D.; Doroski, Michael L.; Payne, Jason; Ibey, Bennett L.; Wilmink, Gerald J.

    2011-03-01

    Terahertz (THz) radiation sources are now being used in a host of military, defense, and medical applications. Widespread employment of these applications has prompted concerns regarding the health effects associated with THz radiation. In this study, we examined the gene expression profile of mammalian cells exposed to THz radiation. We hypothesized that if THz radiation couples directly to cellular constituents, then exposed cells may express a specific gene expression profile indicative of ensuing damage. To test this hypothesis, Jurkat cells were irradiated with a molecular gas THz laser (2.52 THz, 636 mWcm-2, durations: 5, 10, 20, 30, 40, or 50 minutes). Viability was assessed 24 h post-exposure using MTT assays, and gene expression profiles were evaluated 4 h post-exposure using mRNA microarrays. Comparable analyses were also performed for hyperthermic positive controls (44°C for 40 minutes). We found that cellular temperatures increased by ~6 °C during THz exposures. We also found that cell death increased with exposure duration, and the median lethal dose (LD50) was calculated to be ~44 minutes. The microarray data showed that THz radiation induced the transcriptional activation of genes associated with cellular proliferation, differentiation, transcriptional activation, chaperone protein stabilization, and apoptosis. For most genes, we found that the magnitude of differential expression was comparable for both the THz and thermal exposure groups; however, several genes were specifically activated by the THz exposure. These results suggest that THz radiation may elicit effects that are not exclusively due to the temperature rise created during THz exposures (i.e. thermal effects). In future work, we plan to verify the results of our microarray experiments using qPCR techniques.

  14. Stimulation of CD28 triggers an association between CD28 and phosphatidylinositol 3-kinase in Jurkat T cells.

    PubMed

    Truitt, K E; Hicks, C M; Imboden, J B

    1994-03-01

    The T cell surface molecule CD28 can provide costimulatory signals that permit the full activation of T cells. Here we demonstrate that stimulation of CD28, either by B7, its natural ligand, or by the anti-CD28 monoclonal antibody 9.3, induces an association between CD28 and phosphatidylinositol 3-kinase (PI3-K) in Jurkat T cells, raising the possibility that an interaction with PI3-K contributes to CD28-mediated signaling. To examine the mechanism of the association, we synthesized tyrosine-phosphorylated oligopeptides corresponding to each of the four tyrosines in the CD28 cytoplasmic domain. When added to lysates of B7-stimulated Jurkat cells, the oligopeptide corresponding to Tyr 173 inhibits the coimmunoprecipitation of PI3-K with CD28; the other oligopeptides have no effect. Tyr 173 is contained within the sequence YMNM, a motif that is also found in the platelet-derived growth factor receptor and that, when phosphorylated, forms a high affinity binding site for the p85 subunit of PI3-K. These observations suggest that phosphorylation of Tyr 173 may mediate the interaction between CD28 and PI3-K. However, because CD28 is not known to be phosphorylated, it remains possible that CD28 interacts with PI3-K through a mechanism independent of tyrosine phosphorylation. PMID:7509360

  15. Changes in protein expression of U937 and Jurkat cells exposed to nanosecond pulsed electric fields

    NASA Astrophysics Data System (ADS)

    Moen, Erick K.; Roth, Caleb C.; Cerna, Caesar; Estalck, Larry; Wilmink, Gerald; Ibey, Bennett L.

    2013-02-01

    Application of nanosecond pulsed electric fields (nsPEF) to various biological cell lines has been to shown to cause many diverse effects, including poration of the plasma membrane, depolarization of the mitochondrial membrane, blebbing, apoptosis, and intracellular calcium bursts. The underlying mechanism(s) responsible for these diverse responses are poorly understood. Of specific interest in this paper are the long-term effects of nsPEF on cellular processes, including the regulation of genes and production of proteins. Previous studies have reported transient activation of select signaling pathways involving mitogen-activated protein kinases (MAPKs), protein phosphorylation and downstream gene expression following nsPEF application. We hypothesize that nsPEF represents a unique stimulus that could be used to externally modulate cellular processes. To validate our hypothesis, we performed a series of cuvette-based exposures at 10 and 600ns pulse widths using a custom Blumlien line pulser system. We measured acute changes in the plasma membrane structure using flow cytometry by tracking phosphatidylserine externalization via FITC-Annexin V labeling and poration via propidium iodide uptake. We then compared these results to viability of the cells at 24 hours post exposure using MTT assay and changes in the MAPK family of proteins at 8 hours post-exposure using Luminex assay. By comparing exposures at 10 and 600ns duration, we found that most MAPK family-protein expression increased in Jurkat and U937 cell lines following exposure and compared well with drops in viability and changes in plasma membrane asymmetry. What proved interesting is that some MAPK family proteins (e.g. p53, STAT1), were expressed in one cell line, but not the other. This difference may point to an underlying mechanism for observed difference in cellular sensitivity to nsPEFinduced stresses.

  16. Transcriptome analysis of the human T lymphocyte cell line Jurkat and human peripheral blood mononuclear cells exposed to deoxynivalenol (DON): New mechanistic insights

    SciTech Connect

    Katika, Madhumohan R.; Hendriksen, Peter J.M.; Shao, Jia; Loveren, Henk van; Peijnenburg, Ad

    2012-10-01

    Deoxynivalenol (DON) or vomitoxin is a commonly encountered type-B trichothecene mycotoxin, produced by Fusarium species predominantly found in cereals and grains. DON is known to exert toxic effects on the gastrointestinal, reproductive and neuroendocrine systems, and particularly on the immune system. Depending on dose and exposure time, it can either stimulate or suppress immune function. The main objective of this study was to obtain a deeper insight into DON-induced effects on lymphoid cells. For this, we exposed the human T-lymphocyte cell line Jurkat and human peripheral blood mononuclear cells (PBMCs) to various concentrations of DON for various times and examined gene expression changes by DNA microarray analysis. Jurkat cells were exposed to 0.25 and 0.5 μM DON for 3, 6 and 24 h. Biological interpretation of the microarray data indicated that DON affects various processes in these cells: It upregulates genes involved in ribosome structure and function, RNA/protein synthesis and processing, endoplasmic reticulum (ER) stress, calcium-mediated signaling, mitochondrial function, oxidative stress, the NFAT and NF-κB/TNF-α pathways, T cell activation and apoptosis. The effects of DON on the expression of genes involved in ER stress, NFAT activation and apoptosis were confirmed by qRT-PCR. Other biochemical experiments confirmed that DON activates calcium-dependent proteins such as calcineurin and M-calpain that are known to be involved in T cell activation and apoptosis. Induction of T cell activation was also confirmed by demonstrating that DON activates NFATC1 and induces its translocation from the cytoplasm to the nucleus. For the gene expression profiling of PBMCs, cells were exposed to 2 and 4 μM DON for 6 and 24 h. Comparison of the Jurkat microarray data with those obtained with PBMCs showed that most of the processes affected by DON in the Jurkat cell line were also affected in the PBMCs. -- Highlights: ► The human T cell line Jurkat and human

  17. N-Farnesyloxy-norcantharimide inhibits progression of human leukemic Jurkat T cells through regulation of mitogen-activated protein kinase and interleukin-2 production

    PubMed Central

    Chang, Ming-Che; Wu, Jin-Yi; Liao, Hui-Fen; Chen, Yu-Jen

    2015-01-01

    This study investigated the anticancer effects of N-farnesyloxy-norcantharimide (NOC15), a newly synthesized norcantharidin (NCTD) analogue, on human leukemic Jurkat T cells and the signaling pathway underlying its effects. We found that the half maximal inhibitory concentration (IC50) of NOC15 on Jurkat T cells is 1.4 μmol/l, which is 11.14-fold (=15.6÷1.4) smaller than the 15.6 μmol/l of NCTD on Jurkat T cells, whereas the IC50 of NOC15 on human normal lymphoblast (HNL) is 207.9 μmol/l, which is 8.17-fold (=1698.0÷207.8) smaller than the 1698.0 μmol/l of NCTD on HNL cells. These results indicated that NOC15 exerts a higher anticancer effect on Jurkat T cells and has higher toxicity toward HNL cells than NCTD. Thus, NOC15 is 1.36-fold (=11.14÷8.17) beneficial as an anticancer agent toward Jurkat T cells compared with NCTD. Moreover, NOC15 can increase the percentage of cells in the sub-G1 phase and reduce the cell viability of Jurkat T cells, stimulate p38 and extracellular signal-regulated protein kinase 1/2 (ERK1/2) of mitogen-activated protein kinases (MAPKs) signaling pathway, and inhibit calcineurin expression and interleukin-2 (IL-2) production. However, NOC15 exerted no effects on the Jun-N-terminal kinase 1/2 (JNK1/2) signaling pathway, the production of IL-8, and tumor necrosis factor-α. We conclude that the anticancer activity of the newly synthesized NOC15 is 1.36-fold beneficial than NCTD as an anticancer agent and that NOC15 can increase the percentage of cells in the sub-G1 phase through the stimulation of p38 and ERK1/2 of the MAPK signaling pathway and the inhibition of calcineurin expression and IL-2 production. The NOC15 may have the potential of being developed into an anticancer agent in the future. PMID:26288134

  18. Endoplasmic reticulium protein profiling of heat-stressed Jurkat cells by one dimensional electrophoresis and liquid chromatography tandem mass spectrometry.

    PubMed

    Zhang, Xiulian; Kuramitsu, Yasuhiro; Ma, Aiguo; Zhang, Hui; Nakamura, Kazuyuki

    2016-08-01

    Proteomic study on membrane-integrated proteins in endoplasmic reticulum (ER) fractions was performed. In this study, we examined the effects of heat stress on Jurkat cells. The ER fractions were highly purified by differential centrifugation with sodium carbonate washing and acetone methanol precipitations. The ER membrane proteins were separated by one dimensional electrophoresis (1-DE), and some of the protein bands changed their abundance by heat stress, 12 of the 14 bands containing 40 and 60 ribosomal proteins whose expression level were decreased, on the contrary, 2 of the 14 bands containing ubiquitin and eukaryotic translation initiation factor 3 were increased. Heat treatment of human Jurkat cells led to an increase in the phosphorylation of PERK and eIF2α within 30 min of exposure. This was followed by an increase in the expression of the GRP78. Protein ubiquitination and subsequent degradation by the proteasome are important mechanisms regulating cell cycle, growth and differentiation, the result showed that heat stress enhanced ubiquitination modification of the microsomal proteins. The data of this study strongly suggest that heat treatment led to a significant reduction in protein expression and activated UPR, concomitant with protein hyperubiqutination in ER. PMID:25976506

  19. Use of the mitochondria toxicity assay for quantifying the viable cell density of microencapsulated jurkat cells.

    PubMed

    Werner, M; Biss, K; Jérôme, V; Hilbrig, F; Freitag, R; Zambrano, K; Hübner, H; Buchholz, R; Mahou, R; Wandrey, C

    2013-01-01

    The mitochondria toxicity assay (MTT assay) is an established method for monitoring cell viability based on mitochondrial activity. Here the MTT assay is proposed for the in situ quantification of the living cell density of microencapsulated Jurkat cells. Three systems were used to encapsulate the cells, namely a membrane consisting of an interpenetrating polyelectrolyte network of sodium cellulose sulphate/poly(diallyldimethylammonium chloride) (NaCS/PDADMAC), a calcium alginate hydrogel covered with poly(L-lysine) (Ca-alg-PLL), and a novel calcium alginate-poly(ethylene glycol) hybrid material (Ca-alg-PEG). MTT results were correlated to data obtained by the trypan blue exclusion assay after release of the cells from the NaCS/PDADMAC and Ca-alg-PLL capsules, while a resazurin-based assay was used for comparison in case of the Ca-alg-PEG material. Analysis by MTT assay allows quick and reliable determination of viable cell densities of encapsulated cells independent of the capsule material. The assay is highly reproducible with inter-assay relative standard deviations below 10%. PMID:23636962

  20. MUC1 (CD227) interacts with lck tyrosine kinase in Jurkat lymphoma cells and normal T cells.

    PubMed

    Mukherjee, P; Tinder, T L; Basu, G D; Gendler, S J

    2005-01-01

    MUC1 (CD227) is a large transmembrane epithelial mucin glycoprotein, which is aberrantly overexpressed in most adenocarcinomas and is a target for immune therapy for epithelial tumors. Recently, MUC1 has been detected in a variety of hematopoietic cell malignancies including T and B cell lymphomas and myelomas; however, its function in these cells is not clearly defined. Using the Jurkat T cell lymphoma cell line and normal human T cells, we demonstrate that MUC1 is not only expressed in these cells but is also phosphorylated upon T cell receptor (TCR) ligation and associates with the Src-related T cell tyrosine kinase, p56lck. Upon TCR-mediated activation of Jurkat cells, MUC1 is found in the low-density membrane fractions, where linker of T cell activation is contained. Abrogation of MUC1 expression in Jurkat cells by MUC1-specific small interfering RNA resulted in defects in TCR-mediated downstream signaling events associated with T cell activation. These include reduction in Ca2+ influx and extracellular signal-regulated kinase 1/2 phosphorylation, leading to a decrease in CD69 expression, proliferation, and interleukin-2 production. These results suggest a regulatory role of MUC1 in modulating proximal signal transduction events through its interaction with proteins of the activation complex. PMID:15513966

  1. Vitamin E synthetic derivate-TPGS-selectively induces apoptosis in jurkat t cells via oxidative stress signaling pathways: implications for acute lymphoblastic leukemia.

    PubMed

    Ruiz-Moreno, Cristian; Jimenez-Del-Rio, Marlene; Sierra-Garcia, Ligia; Lopez-Osorio, Betty; Velez-Pardo, Carlos

    2016-09-01

    D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) is a water-soluble derivative of natural vitamin E commonly used as a drug delivery agent. Recently, TPGS alone has been reported to induce cell death in lung, breast and prostate cancer. However, the effect of TPGS on cancer cell viability remains unclear. Thus, this study was aimed to evaluate the cytotoxic effect of TPGS on human periphral blood lymphocytes (PBL) and on T cell acute lymphocytic leukemia (ALL) Jurkat clone E6-1 cells and its possible mechanism of action. PBL and Jurkat cells were treated with TPGS (10, 20, 40, 60, and 80 μM), and morphological changes in the cell nucleus, mitochondrial membrane potential (ΔΨm), and intracellular reactive oxygen species levels were determined by immune-fluorescence microscopy and flow cytometry. Cellular apoptosis markers were also evaluated by immunocytochemistry. In this study, TPGS induced apoptotic cell death in Jurkat cells, but not in PBL, in a dose-response manner with increasing nuclear DNA fragmentation, increasing cell cycle arrest, and decreasing ΔΨm. Additionally, TPGS increased dichlorofluorescein fluorescence intensity, indicative of H2O2 production, in a dose-independent fashion. TPGS increased DJ-1 Cys(106)-sulfonate, as a marker of intracellular stress and induced the activation of NF-κB, p53 and c-Jun transcription factors. Additionally, it increased the expression of apoptotic markers Bcl-2 related pro-apoptotic proteins Bax and PUMAand activated caspase-3. The antioxidant N-acetyl-L-cysteine and known pharmacological inhibitors protected the cells from the TPGS induced effects. In conclusion, TPGS selectively induces apoptosis in Jurkat cells through two independent but complementary H2O2-mediated signaling pathways. Our findings support the use of TPGS as a potential treatment for ALL. PMID:27364951

  2. Successful validation of genomic biomarkers for human immunotoxicity in Jurkat T cells in vitro.

    PubMed

    Schmeits, Peter C J; Shao, Jia; van der Krieken, Danique A; Volger, Oscar L; van Loveren, Henk; Peijnenburg, Ad A C M; Hendriksen, Peter J M

    2015-07-01

    Previously, we identified 25 classifier genes that were able to assess immunotoxicity using human Jurkat T cells. The present study aimed to validate these classifiers. For that purpose, Jurkat cells were exposed for 6 h to subcytotoxic doses of nine immunotoxicants, five non-immunotoxicants and four compounds for which human immunotoxicity has not yet been fully established. RNA was isolated and subjected to Fluidigm quantitative real time (qRT)-PCR analysis. The sensitivity, specificity and accuracy of the screening assay as based on the nine immunotoxicants and five non-immunotoxicants used in this study were 100%, 80% and 93%, respectively, which is better than the performance in our previous study. Only one compound was classified as false positive (benzo-e-pyrene). Of the four potential (non-)immunotoxicants, chlorantraniliprole and Hidrasec were classified immunotoxic and Sunset yellow and imidacloprid as non-immunotoxic. ToxPi analysis of the PCR data provided insight in the molecular pathways that were affected by the compounds. The immunotoxicants 2,3-dichloro-propanol and cypermethrin, although structurally different, affected protein metabolism and cholesterol biosynthesis and transport. In addition, four compounds, i.e. chlorpyrifos, aldicarb, benzo-e-pyrene and anti-CD3, affected genes in cholesterol metabolism and transport, protein metabolism and transcription regulation. qRT-PCR on eight additional genes coding for similar processes as defined in ToxPi analyzes, supported these results. In conclusion, the 25 immunotoxic classifiers performed very well in a screening with new non-immunotoxic and immunotoxic compounds. Therefore, the Jurkat screening assay has great promise to be applied within a tiered approach for animal free testing of human immunotoxicity. PMID:25424538

  3. Polysaccharide Isolated from Zizyphus jujuba ( Hóng Zǎo) Inhibits Interleukin-2 Production in Jurkat T Cells.

    PubMed

    Hsu, Bo-Yang; Kuo, Yuh-Chi; Chen, Bing-Huei

    2014-04-01

    Zizyphus jujuba ( Hóng Zǎo), a traditional Chinese herb widely used in many Asian countries, has been shown to possess vital biological activities such as anti-cancer activity. The objective of this study was to evaluate the immunomodulatory effect of deproteinated polysaccharide (DP) isolated from Z. jujuba. The DP isolated from Z. jujuba consisted of two polysaccharide fractions and their molecular weights (MWs) were found to be 143,108 and 67,633 Da, respectively. The DP could significantly decrease interleukin (IL)-2 production in phytohemagglutinin (PHA)-activated Jurkat T cells in a dose-dependent manner after 48 h of incubation, with the inhibition being 47.5%, 61.2%, and 81.7% for DP concentrations of 0.75, 1.75, and 2.5 mg/ml, respectively. Thus, our study showed that DP isolated from Z. jujuba may possess anti-inflammatory activity as it could significantly reduce IL-2 production in activated Jurkat T cells. PMID:24860737

  4. Exposure of Jurkat cells to bis (tri-n-butyltin) oxide (TBTO) induces transcriptomics changes indicative for ER- and oxidative stress, T cell activation and apoptosis

    SciTech Connect

    Katika, Madhumohan R.; Hendriksen, Peter J.M.; Loveren, Henk van; Peijnenburg, Ad

    2011-08-01

    Tributyltin oxide (TBTO) is an organotin compound that is widely used as a biocide in agriculture and as an antifouling agent in paints. TBTO is toxic for many cell types, particularly immune cells. The present study aimed to identify the effects of TBTO on the human T lymphocyte cell line Jurkat. Cells were treated with 0.2 and 0.5 {mu}M TBTO for 3, 6, 12 and 24 h and then subjected to whole genome gene expression microarray analysis. The biological interpretation of the gene expression profiles revealed that endoplasmic reticulum (ER) stress is among the earliest effects of TBTO. Simultaneously or shortly thereafter, oxidative stress, activation of NFKB and NFAT, T cell activation, and apoptosis are induced. The effects of TBTO on genes involved in ER stress, NFAT pathway, T cell activation and apoptosis were confirmed by qRT-PCR. Activation and nuclear translocation of NFATC1 and the oxidative stress response proteins NRF2 and KEAP1 were confirmed by immunocytology. Taking advantage of previously published microarray data, we demonstrated that the induction of ER stress, oxidative stress, T cell activation and apoptosis by TBTO is not unique for Jurkat cells but does also occur in mouse thymocytes both ex vivo and in vivo and rat thymocytes ex vivo. We propose that the induction of ER stress leading to a T cell activation response is a major factor in the higher sensitivity of immune cells above other types of cells for TBTO. - Research Highlights: > The human T lymphocyte cell line Jurkat was exposed to TBTO. > Whole-genome microarray experiments were performed. > Data analysis revealed the induction of ER stress and activation of NFAT and NFKB. > Exposure to TBTO also led to T cell activation, oxidative stress and apoptosis.

  5. Functional and genomic analyses of FOXP3-transduced Jurkat-T cells as regulatory T (Treg)-like cells

    SciTech Connect

    Kim, Joon-Young Kim, Han-Jong; Hurt, Elaine M.; Chen, Xin; Howard, O.M. Zack; Farrar, William L.

    2007-10-12

    FOXP3, a forkhead transcription factor is essential for the development and function of CD4{sup +}CD25{sup +} regulatory T cells (Tregs). In contrast to conversion of murine naive T cells to Tregs by transduction of Foxp3, it is controversial whether ectopic expression of FOXP3 in human CD4{sup +} T cells is sufficient for acquisition of suppressive activity. Here, we show that retroviral transduction of FOXP3 induces a Treg phenotype in human leukemic CD4{sup +} Jurkat-T cells, evidenced by increased expression of Treg-associated cell surface markers as well as inhibition of cytokine production. Furthermore, FOXP3-transduced Jurkat-T cells suppress the proliferation of human CD4{sup +}CD25{sup -} T cells. Additionally, DNA microarray analysis identifies Treg-related genes regulated by FOXP3. Our study demonstrates that enforced expression of FOXP3 confers Treg-like properties on Jurkat-T cells, which can be a convenient and efficient Treg-like cell model for further study to identify Treg cell surface markers and target genes regulated by FOXP3.

  6. Mössbauer Study and Modeling of Iron Import and Trafficking in Human Jurkat Cells

    PubMed Central

    Jhurry, Nema D.; Chakrabarti, Mrinmoy; McCormick, Sean P.; Gohil, Vishal M.; Lindahl, Paul A.

    2014-01-01

    The Fe content of Jurkat cells grown on transferrin-bound iron (TBI) and FeIII citrate (FC) was characterized using Mössbauer, EPR, and UV-vis spectroscopies, electron microscopy, and ICP-MS. Isolated mitochondria were similarly characterized. Fe-limited cells contained ∼ 100 μM of essential Fe, mainly as mitochondrial Fe and non-mitochondrial nonheme high-spin (NHHS) FeII. Fe-replete cells also contained ferritin-bound Fe and FeIII oxyhydroxide nanoparticles. Only 400 ± 100 Fe ions were loaded per ferritin complex, regardless of the growth medium Fe concentration. Ferritin regulation thus appears more complex than is commonly assumed. The magnetic/structural properties of Jurkat nanoparticles differed from those in yeast mitochondria. They were smaller and may be located in the cytosol. The extent of nanoparticle formation scaled nonlinearly with the concentration of Fe in the medium. Nanoparticle formation was not strongly correlated with ROS damage. Cells could utilize nanoparticle Fe, converting such aggregates into essential Fe forms. Cells grown on galactose rather than glucose respired faster, grew slower, exhibited more ROS damage and generally contained more nanoparticles. Cells grown with TBI rather than FC contained lower Fe concentrations, more ferritin and fewer nanoparticles. Cells in which transferrin receptor expression was increased contained more ferritin Fe. Frataxin-deficient cells contained more nanoparticles than comparable WT cells. Data were analyzed by a chemically-based mathematical model. Although simple, it captured essential features of Fe import, trafficking and regulation. TBI import was highly regulated but FC import was not. Nanoparticle formation was not regulated but the rate was third-order in cytosolic Fe. PMID:24180611

  7. Opium induces apoptosis in Jurkat cells via promotion of pro-apoptotic and inhibition of anti-apoptotic molecules

    PubMed Central

    Arababadi, Mohammad Kazemi; Asadikaram, Gholamreza

    2016-01-01

    Objective(s): The aim of this study was to determine the important molecules involved in apoptosis induction by opium in Jurkat cell line. Materials and Methods: Jurkat cells were incubated 48 hrs with 2.86×10-5 g/ml concentration of opium and apoptosis as well as expression levels of related molecules were measured. Results: Our results demonstrated that 50.3±0.2 percent of opium treated Jurkat cells were revealed apoptotic features. The levels of mRNA of several pro-apoptotic and anti-apoptotic molecules were increased and decreased, respectively, in the opium treated cells. The results also demonstrated that expression levels of BCL2, DFFA and NOL3 as anti-apoptotic molecules were increased in the opium treated cells. Conclusion: It seems that opium induces apoptosis in Jurkat cells via both intrinsic and extrinsic pathways. Although opium induces apoptosis in the cells but increased expression of some anti-apoptotic molecules may be a normal resistance of the cell for death. PMID:27081468

  8. Lectin interactions with the Jurkat leukemic T-cell line: quantitative binding studies and interleukin-2 production

    SciTech Connect

    Dupuis, G.; Bastin, B.

    1988-03-01

    Phytohemagglutinin (PHA), concanavalin A (Con A), pea lectin, and wheat germ agglutinin (WGA) have been used to investigate their binding properties to Jurkat 77 6.8 leukemic human T cells and their ability to induce these cells to produce interleukin-2 (IL-2). Binding studies showed that the Jurkat cells fixed 0.82 +/- 0.11 microgram pea lectin, 2.02 +/- 0.17 micrograms Con A, 1.85 +/- 0.07 micrograms PHA and 8.88 +/- 0.61 micrograms WGA. Scatchard plots were linear, indicating that the binding process was homogeneous with respect to the binding constant. PHA and Con A bound with the highest affinity (Kass (apparent) approximately equal to 9 x 10(9) M-1), followed by pea lectin and WGA (Kass (apparent) approximately equal to 3 x 10(9) M-1). The number of lectin binding sites was in agreement with the results of saturation experiments. We also evaluated the effect of the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the binding process. Results show that there were no gross alterations in the value of (apparent) Kass in the case of PHA and WGA. In contrast, the presence of TPA decreased the affinity of Con A and modified the Scatchard profile for pea lectin, which was curvilinear with a concavity turned upward. In this case, data were (apparent) K1 = 17.7 x 10(9) M-1 (high-affinity sites) and (apparent) K2 = 2.6 x 10(9) M-1 (low-affinity sites). The four lectins shared the ability to stimulate Jurkat 77 6.8 cells to secrete IL-2. Optimal lectin concentrations were 20 micrograms/ml (PHA) and 50 micrograms/ml (WGA and Con A). Pea lectin failed to display a dose-response relationship, and IL-2 production increased proportionally with lectin concentration. Con A was the most efficient stimulator (250 U/ml), followed by WGA (160 U/ml) and PHA (108 U/ml).

  9. The pan-ErbB tyrosine kinase inhibitor canertinib induces caspase-mediated cell death in human T-cell leukemia (Jurkat) cells

    SciTech Connect

    Trinks, Cecilia; Severinsson, Emelie A.; Holmlund, Birgitta; Green, Anna; Green, Henrik; Joensson, Jan-Ingvar; Hallbeck, Anna-Lotta; Walz, Thomas M.

    2011-07-08

    Highlights: {yields} Canertinib induces caspase-mediated apoptosis in T-cell leukemia cells in vitro. {yields} Canertinib mediates activation of the intrinsic apoptotic pathway. {yields} Canertinib induces apoptosis in an ErbB receptor independent manner. {yields} Lymphocyte specific proteins as well as survival kinases are inhibited. {yields} Canertinib may act as a multi-kinase inhibiting drug in human T-cell malignancies. -- Abstract: Canertinib is a novel ErbB-receptor inhibitor currently in clinical development for the treatment of solid tumors overexpressing ErbB-receptors. We have recently demonstrated that canertinib displays anti-proliferative and pro-apoptotic effects in human myeloid leukemia cells devoid of ErbB-receptors. The mechanism mediating these effects are however unknown. In this study, we show that canertinib is able to act as a multi-kinase inhibitor by inhibition of several intracellular kinases involved in T-cell signaling such as Akt, Erk1/2 and Zap-70, and reduced Lck protein expression in the human T-cell leukemia cell line Jurkat. Treatment with canertinib at a concentration of 2 {mu}M caused accumulation of Jurkat cells in the G{sub 1} cell cycle phase and increased doses induced apoptosis in a time-dependent manner. Apoptotic signs of treated cells were detected by Annexin V staining and cleavage of PARP, caspase-3, -8, -9, -10 and Bid. A subset of the pro-apoptotic signals mediated by canertinib could be significantly reduced by specific caspase inhibitors. Taken together, these results demonstrate the dual ability of canertinib to downregulate important signaling pathways and to activate caspase-mediated intrinsic apoptosis pathway in human T-cell leukemia cells.

  10. Caspase-dependent inhibition of store-operated Ca{sup 2+} entry into apoptosis-committed Jurkat cells

    SciTech Connect

    Onopiuk, Marta; Wierzbicka, Katarzyna; Brutkowski, Wojciech; Szczepanowska, Joanna; Zablocki, Krzysztof

    2010-08-20

    Activation of T-cells triggers store-operated Ca{sup 2+} entry, which begins a signaling cascade leading to induction of appropriate gene expression and eventually lymphocyte proliferation and differentiation. The simultaneous enhancement of Fas ligand gene expression in activated cells allows the immune response to be limited by committing the activated cells to apoptosis. In apoptotic cells the store-operated calcium entry is significantly inhibited. It has been documented that moderate activation of Fas receptor may cause reversible inhibition of store-operated channels by ceramide released from hydrolyzed sphingomyelin. Here we show that activation of Fas receptor in T-cells results in caspase-dependent decrease of cellular STIM1 and Orai1 protein content. This effect may be responsible for the substantial inhibition of Ca{sup 2+} entry into Jurkat cells undergoing apoptosis. In turn, this inhibition might prevent overloading of cells with calcium and protect them against necrosis. -- Research highlights: {yields} Fas activation reduces STIM1 and Orai1 protein content in caspase dependent manner. {yields} Fas activation partially reduces mitochondrial potential in caspase dependent manner. {yields} Fas stimulation inhibits of store-operated Ca{sup 2+} entry in caspase dependent manner. {yields} Inhibition of Ca{sup 2+} entry in apoptotic cells may protect them from secondary necrosis.

  11. Bovine lactoferricin causes apoptosis in Jurkat T-leukemia cells by sequential permeabilization of the cell membrane and targeting of mitochondria

    SciTech Connect

    Mader, Jamie S.; Richardson, Angela; Salsman, Jayme; Top, Deniz; Antueno, Roberto de; Duncan, Roy; Hoskin, David W. . E-mail: d.w.hoskin@dal.ca

    2007-07-15

    Bovine lactoferricin (LfcinB) is a cationic antimicrobial peptide that kills Jurkat T-leukemia cells by the mitochondrial pathway of apoptosis. However, the process by which LfcinB triggers mitochondria-dependent apoptosis is not well understood. Here, we show that LfcinB-induced apoptosis in Jurkat T-leukemia cells was preceded by LfcinB binding to, and progressive permeabilization of the cell membrane. Colloidal gold electron microscopy revealed that LfcinB entered the cytoplasm of Jurkat T-leukemia cells prior to the onset of mitochondrial depolarization. LfcinB was not internalized by endocytosis because endocytosis inhibitors did not prevent LfcinB-induced cytotoxicity. Furthermore, intracellular delivery of LfcinB via fusogenic liposomes caused the death of Jurkat T-leukemia cells, as well as normal human fibroblasts. Collectively, these findings suggest that LfcinB caused damage to the cell membrane that allowed LfcinB to enter the cytoplasm of Jurkat T-leukemia cells and mediate cytotoxicity. In addition, confocal microscopy showed that intracellular LfcinB co-localized with mitochondria in Jurkat T-leukemia cells, while flow cytometry and colloidal gold electron microscopy showed that LfcinB rapidly associated with purified mitochondria. Furthermore, purified mitochondria treated with LfcinB rapidly lost transmembrane potential and released cytochrome c. We conclude that LfcinB-induced apoptosis in Jurkat T-leukemia cells resulted from cell membrane damage and the subsequent disruption of mitochondrial membranes by internalized LfcinB.

  12. Nanobody-based chimeric receptor gene integration in Jurkat cells mediated by PhiC31 integrase

    SciTech Connect

    Iri-Sofla, Farnoush Jafari; Rahbarizadeh, Fatemeh; Ahmadvand, Davoud; Rasaee, Mohammad J.

    2011-11-01

    The crucial role of T lymphocytes in anti-tumor immunity has led to the development of novel strategies that can target and activate T cells against tumor cells. Recombinant DNA technology has been used to generate non-MHC-restricted chimeric antigen receptors (CARs). Here, we constructed a panel of recombinant CAR that harbors the anti-MUC1 nanobody and the signaling and co-signaling moieties (CD3{zeta}/CD28) with different spacer regions derived from human IgG3 with one or two repeats of the hinge sequence or the hinge region of Fc{gamma}RII. The PhiC31 integrase system was employed to investigate if the recombination efficiency could be recruited for high and stable expression of T cell chimeric receptor genes. The effect of nuclear localization signal (NLS) and two different promoters (CMV and CAG) on efficacy of PhiC31 integrase in human T cell lines was evaluated. The presence of integrase in combination with NLS, mediated up to 7.6 and 8.5 fold increases in CAR expression in ZCHN-attB and ZCHHN-attB cassette integrated T cells, respectively. Our results showed that highly efficient and stable transduction of the Jurkat cell line by PhiC31 integrase is a feasible modality for generating anti-cancer chimeric T cells for use in cancer immunotherapy.

  13. Involvement of sphingosine in mitochondria-dependent Fas-induced apoptosis of type II Jurkat T cells.

    PubMed

    Cuvillier, O; Edsall, L; Spiegel, S

    2000-05-26

    Exposure to anti-Fas antibody in Jurkat cells (type II cells), which are characterized by a weak caspase-8 activation at the death-inducing signaling complex (DISC), induced a biphasic increase in ceramide levels. The early generation of ceramide preceded transient activation of acidic ceramidase and subsequent production of sphingosine, followed by cytochrome c release, activation of caspases-2, -3, -6, -7, -8, and -9, Bid cleavage, and a later sustained ceramide accumulation. The caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone inhibited early increases of ceramide and sphingosine, whereas overexpression of Bcl-x(L) had no effect, and both prevented the later sustained ceramide accumulation. Exogenous sphingosine, as well as cell-permeable C(2)-ceramide, induced cytochrome c release from mitochondria in a caspase-independent fashion leading to activation of caspase-9 and executioner caspases and, surprisingly, activation of the initiator caspase-8 and processing of its substrate Bid. These effects were also completely abolished by Bcl-x(L) overexpression. Our results suggest that sphingosine might also be involved in the mitochondria-mediated pathway of Fas-induced cell death in type II cells. PMID:10747891

  14. 17{alpha}-Estradiol arrests cell cycle progression at G{sub 2}/M and induces apoptotic cell death in human acute leukemia Jurkat T cells

    SciTech Connect

    Jun, Do Youn; Park, Hae Sun; Kim, Jun Seok; Kim, Jong Sik; Park, Wan; Song, Bang Ho; Kim, Hee-Sook; Taub, Dennis; Kim, Young Ho

    2008-09-15

    A pharmacological dose (2.5-10 {mu}M) of 17{alpha}-estradiol (17{alpha}-E{sub 2}) exerted a cytotoxic effect on human leukemias Jurkat T and U937 cells, which was not suppressed by the estrogen receptor (ER) antagonist ICI 182,780. Along with cytotoxicity in Jurkat T cells, several apoptotic events including mitochondrial cytochrome c release, activation of caspase-9, -3, and -8, PARP degradation, and DNA fragmentation were induced. The cytotoxicity of 17{alpha}-E{sub 2} was not blocked by the anti-Fas neutralizing antibody ZB-4. While undergoing apoptosis, there was a remarkable accumulation of G{sub 2}/M cells with the upregulatoin of cdc2 kinase activity, which was reflected in the Thr56 phosphorylation of Bcl-2. Dephosphorylation at Tyr15 and phosphorylation at Thr161 of cdc2, and significant increase in the cyclin B1 level were underlying factors for the cdc2 kinase activation. Whereas the 17{alpha}-E{sub 2}-induced apoptosis was completely abrogated by overexpression of Bcl-2 or by pretreatment with the pan-caspase inhibitor z-VAD-fmk, the accumulation of G{sub 2}/M cells significantly increased. The caspase-8 inhibitor z-IETD-fmk failed to influence 17{alpha}-E{sub 2}-mediated caspase-9 activation, but it markedly reduced caspase-3 activation and PARP degradation with the suppression of apoptosis, indicating the contribution of caspase-8; not as an upstream event of the mitochondrial cytochrome c release, but to caspase-3 activation. In the presence of hydroxyurea, which blocked the cell cycle progression at the G{sub 1}/S boundary, 17{alpha}-E{sub 2} failed to induce the G{sub 2}/M arrest as well as apoptosis. These results demonstrate that the cytotoxicity of 17{alpha}-E{sub 2} toward Jurkat T cells is attributable to apoptosis mainly induced in G{sub 2}/M-arrested cells, in an ER-independent manner, via a mitochondria-dependent caspase pathway regulated by Bcl-2.

  15. DNA Electrophoretic Migration Patterns Change after Exposure of Jurkat Cells to a Single Intense Nanosecond Electric Pulse

    PubMed Central

    Romeo, Stefania; Zeni, Luigi; Sarti, Maurizio; Sannino, Anna; Scarfì, Maria Rosaria; Vernier, P. Thomas; Zeni, Olga

    2011-01-01

    Intense nanosecond pulsed electric fields (nsPEFs) interact with cellular membranes and intracellular structures. Investigating how cells respond to nanosecond pulses is essential for a) development of biomedical applications of nsPEFs, including cancer therapy, and b) better understanding of the mechanisms underlying such bioelectrical effects. In this work, we explored relatively mild exposure conditions to provide insight into weak, reversible effects, laying a foundation for a better understanding of the interaction mechanisms and kinetics underlying nsPEF bio-effects. In particular, we report changes in the nucleus of Jurkat cells (human lymphoblastoid T cells) exposed to single pulses of 60 ns duration and 1.0, 1.5 and 2.5 MV/m amplitudes, which do not affect cell growth and viability. A dose-dependent reduction in alkaline comet-assayed DNA migration is observed immediately after nsPEF exposure, accompanied by permeabilization of the plasma membrane (YO-PRO-1 uptake). Comet assay profiles return to normal within 60 minutes after pulse delivery at the highest pulse amplitude tested, indicating that our exposure protocol affects the nucleus, modifying DNA electrophoretic migration patterns. PMID:22164287

  16. Pandemic Influenza A (H1N1) Virus Infection Increases Apoptosis and HIV-1 Replication in HIV-1 Infected Jurkat Cells

    PubMed Central

    Wang, Xue; Tan, Jiying; Biswas, Santanu; Zhao, Jiangqin; Devadas, Krishnakumar; Ye, Zhiping; Hewlett, Indira

    2016-01-01

    Influenza virus infection has a significant impact on public health, since it is a major cause of morbidity and mortality. It is not well-known whether influenza virus infection affects cell death and human immunodeficiency virus (HIV)-1 replication in HIV-1-infected patients. Using a lymphoma cell line, Jurkat, we examined the in vitro effects of pandemic influenza A (H1N1) virus (pH1N1) infection on cell death and HIV-1 RNA production in infected cells. We found that pH1N1 infection increased apoptotic cell death through Fas and Bax-mediated pathways in HIV-1-infected Jurkat cells. Infection with pH1N1 virus could promote HIV-1 RNA production by activating host transcription factors including nuclear factor kappa-light-chain-enhancer of activated B cells (NF-ĸB), nuclear factor of activated T-cells (NFAT) and activator protein 1 (AP-1) through mitogen-activated protein kinases (MAPK) pathways and T-cell antigen receptor (TCR)-related pathways. The replication of HIV-1 latent infection could be reactivated by pH1N1 infection through TCR and apoptotic pathways. These data indicate that HIV-1 replication can be activated by pH1N1 virus in HIV-1-infected cells resulting in induction of cell death through apoptotic pathways. PMID:26848681

  17. Regulation of heat shock protein message in Jurkat cells cultured under serum-starved and gravity-altered conditions

    NASA Technical Reports Server (NTRS)

    Lewis, M. L.; Hughes-Fulford, M.

    2000-01-01

    Although our understanding of effects of space flight on human physiology has advanced significantly over the past four decades, the potential contribution of stress at the cellular and gene regulation level is not characterized. The objective of this ground-based study was to evaluate stress gene regulation in cells exposed to altered gravity and environmentally suboptimal conditions. We designed primers to detect message for both the constitutive and inducible forms of the heat shock protein, HSP-70. Applying the reverse transcriptase-polymerase chain reaction (RT-PCR), we probed for HSP-70 message in human acute T-cell leukemia cells, Jurkat, subjected to three types of environmental stressors: (1) altered gravity achieved by centrifugation (hypergravity) and randomization of the gravity vector in rotating bioreactors, (2) serum starvation by culture in medium containing 0.05% serum, and (3) temperature elevation (42 degrees C). Temperature elevation, as the positive control, significantly increased HSP-70 message, while centrifugation and culture in rotating bioreactors did not upregulate heat shock gene expression. We found a fourfold increase in heat shock message in serum-starved cells. Message for the housekeeping genes, actin and cyclophilin, were constant and comparable to unstressed controls for all treatments. We conclude that gravitational perturbations incurred by centrifugal forces, exceeding those characteristic of a Space Shuttle launch (3g), and culture in rotating bioreactors do not upregulate HSP-70 gene expression. In addition, we found RT-PCR useful for evaluating stress in cultured cells. Copyright 2000 Wiley-Liss, Inc.

  18. Role of intracellular labile iron, ferritin, and antioxidant defence in resistance of chronically adapted Jurkat T cells to hydrogen peroxide

    PubMed Central

    Al-Qenaei, Abdullah; Yiakouvaki, Anthie; Reelfs, Olivier; Santambrogio, Paolo; Levi, Sonia; Hall, Nick D.; Tyrrell, Rex M.; Pourzand, Charareh

    2014-01-01

    To examine the role of intracellular labile iron pool (LIP), ferritin (Ft), and antioxidant defence in cellular resistance to oxidative stress on chronic adaptation, a new H2O2-resistant Jurkat T cell line “HJ16” was developed by gradual adaptation of parental “J16” cells to high concentrations of H2O2. Compared to J16 cells, HJ16 cells exhibited much higher resistance to H2O2-induced oxidative damage and necrotic cell death (up to 3 mM) and had enhanced antioxidant defence in the form of significantly higher intracellular glutathione and mitochondrial ferritin (FtMt) levels as well as higher glutathione-peroxidase (GPx) activity. In contrast, the level of the Ft H-subunit (FtH) in the H2O2-adapted cell line was found to be 7-fold lower than in the parental J16 cell line. While H2O2 concentrations higher than 0.1 mM fully depleted the glutathione content of J16 cells, in HJ16 cells the same treatments decreased the cellular glutathione content to only half of the original value. In HJ16 cells, H2O2 concentrations higher than 0.1 mM increased the level of FtMt up to 4-fold of their control values but had no effect on the FtMt levels in J16 cells. Furthermore, while the basal cytosolic level of LIP was similar in both cell lines, H2O2 treatment substantially increased the cytosolic LIP levels in J16 but not in HJ16 cells. H2O2 treatment also substantially decreased the FtH levels in J16 cells (up to 70% of the control value). In contrast in HJ16 cells, FtH levels were not affected by H2O2 treatment. These results indicate that chronic adaptation of J16 cells to high concentrations of H2O2 has provoked a series of novel and specific cellular adaptive responses that contribute to higher resistance of HJ16 cells to oxidative damage and cell death. These include increased cellular antioxidant defence in the form of higher glutathione and FtMt levels, higher GPx activity, and lower FtH levels. Further adaptive responses include the significantly reduced

  19. The human T-cell leukemia virus type 1 Rex regulatory protein exhibits an impaired functionality in human lymphoblastoid Jurkat T cells.

    PubMed Central

    Hamaia, S; Cassé, H; Gazzolo, L; Duc Dodon, M

    1997-01-01

    The Rex protein of human T-cell leukemia virus type 1 (HTLV-1) intervenes in the posttranscriptional regulation of proviral gene expression. Its binding to the Rex response element (XRE) present in the 3' long terminal repeat ensures the coordinate cytoplasmic accumulation of spliced and unspliced forms of viral messengers. Consequently, synthesis of viral structural and enzymatic proteins is strictly dependent on the Rex posttranscriptional activity. Here we report that synthesis of HTLV-1 envelope glycoproteins by Jurkat T cells could be detected only when they were regulated in a Rex-independent manner. Indeed, Jurkat T cells transfected with a Rex-dependent env expression vector (encompassing both the env and pX open reading frames) do not produce significant levels of envelope glycoproteins despite the production of significant amounts of Rex protein. The analysis of levels and distribution patterns of the unspliced env and of the singly spliced tax/rex transcripts suggests that the failure in envelope glycoprotein synthesis may be ascribed to a deficiency of Rex in mediating the nucleocytoplasmic transport of unspliced env RNAs in these cells. Furthermore, despite the synthesis of regulatory proteins, HTLV-1 structural proteins were not detected in Jurkat T cells transfected with an HTLV-1 infectious provirus. Conversely, and as expected, structural proteins were produced by Jurkat cells transfected by a human immunodeficiency virus type 1 (HIV-1) infectious provirus. This phenotype appeared to be linked to a specific dysfunction of Rex, since the functionally equivalent Rev protein of HIV-1 was shown to be fully efficient in promoting the synthesis of HTLV-1 envelope glycoproteins in Jurkat cells. Therefore, it seems likely that the block to Rex function in these lymphoblastoid T cells is determined by inefficient Rex-XRE interactions. These observations suggest that the acquisition of this Rex-deficient phenotype by in vivo-infected HTLV-1 T cells may

  20. Modulation of Cytokine Production and Transcription Factors Activities in Human Jurkat T Cells by Thymol and Carvacrol

    PubMed Central

    Gholijani, Nasser; Gharagozloo, Marjan; Kalantar, Fathollah; Ramezani, Amin; Amirghofran, Zahra

    2015-01-01

    Purpose: Thymol and carvacrol, two main components of thyme, have shown anti-inflammatory effects. The aim of this study was to assess the effects of these components on Jurkat leukemia cells as an in vitro T cell model and their molecular mechanisms of activity. Methods: Cells were cultured in the presence of components and subsequently stimulated with phorbol-12-myristate-13-acetate (PMA)/calcium ionophore for evaluating interleukin (IL)-2 and interferon (IFN)-γ production. The activation of T cell transcription factors that included nuclear factors of activated T cells (NFATs), activator protein-1 (AP-1; c-Jun/c-Fos), and nuclear factor (NF)-κB were examined by Western blot analysis. Results: Thymol and carvacrol at 25 µg/ml significantly reduced IL-2 levels from 119.4 ± 8pg/ml in control cells treated only with PMA/Calcium ionophore and the solvent to 66.9 ± 6.4pg/ml (thymol) and 32.3 ± 3.6pg/ml (carvacrol) and IFN-γ from 423.7 ± 19.7pg/ml in control cells to 311.9 ± 11.6pg/ml (thymol) and 293.5 ± 16.7pg/ml (carvacrol). Western blot analyses of nuclear extracts showed that the same concentrations of components significantly reduced NFAT-2 to 44.2 ± 2.7% (thymol) and 91.4 ± 2.3% (carvacrol) of the control (p<0.05), and c-Fos to 31.2 ± 6.2% (thymol) and 27.6 ± 3.1% (carvacrol) of the control (p<0.01). No effects on NFAT-1, c-Jun and phospho-NF-κBp65 levels were observed. Conclusion: Thymol and carvacrol could contribute to modulation of T cell activity by reducing IL-2 and IFN-γ production possibly through down regulation of AP-1 and NFAT-2 transcription factors suggesting their potential usefulness for reduction of T cell overactivity in immune-mediated diseases. PMID:26793612

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

    SciTech Connect

    Reich, Charles F.; Pisetsky, David S.

    2009-03-10

    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 {beta}-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.

  2. Comparative assessment of therapeutic safety of norcantharidin, N-farnesyloxy-norcantharimide, and N-farnesyl-norcantharimide against Jurkat T cells relative to human normal lymphoblast

    PubMed Central

    Chang, Ming-Che; Wu, Jin-Yi; Liao, Hui-Fen; Chen, Yu-Jen; Kuo, Cheng-Deng

    2016-01-01

    Abstract The therapeutic safety of an anticancer drug is one of the most important concerns of the physician treating the cancer patient. Half maximal inhibitory concentration (IC50) and hillslope are usually used to represent the strength and sensitivity of an anticancer drug on cancer cells. The therapeutic safety of the anticancer drug can be assessed by comparing the IC50 and hillslope of anticancer drugs on cancer cells relative to normal cells. Since there are situations where “more anticancer activity” implies “more toxicity,” the safety of an anticancer drug in these situations is hard to evaluate by using IC50 and hillslope alone. In a previous study, the “net effect” index was devised to represent the net therapeutic effects of one anticancer drug relative to the other. However, the therapeutic safety of one specific anticancer drug alone was not defined in the “net effect” index. This study introduced the “safety index (SI)” to quantify the degree of safety of an anticancer drug by using 4-parameter logistic model on cancer cells relative to normal cells. The therapeutic safety of norcantharidin (NCTD), N-farnesyloxy-norcantharimide (NOC15), and N-farnesyl-norcantharimide (NC15) in the treatment of Jurkat T cells relative to human normal lymphoblast was compared using the newly defined SI. We found that the SI of NOC15 and NC15 was significantly higher than that of NCTD, suggesting that both NOC15 and NC15 can damage more cancer cells and less normal cells than NCTD. We conclude that both NOC15 and NC15 are safer anticancer drugs than NCTD in the treatment of Jurkat T cells relative to human normal lymphoblast. The SI can be further applied to the screening, developments, and applications of anticancer drugs in the future. PMID:27495082

  3. Comparative proteomics of exosomes secreted by tumoral Jurkat T cells and normal human T cell blasts unravels a potential tumorigenic role for valosin-containing protein.

    PubMed

    Bosque, Alberto; Dietz, Lisa; Gallego-Lleyda, Ana; Sanclemente, Manuel; Iturralde, María; Naval, Javier; Alava, María Angeles; Martínez-Lostao, Luis; Thierse, Hermann-Josef; Anel, Alberto

    2016-05-17

    We have previously characterized that FasL and Apo2L/TRAIL are stored in their bioactive form inside human T cell blasts in intraluminal vesicles present in multivesicular bodies. These vesicles are rapidly released to the supernatant in the form of exosomes upon re-activation of T cells. In this study we have compared for the first time proteomics of exosomes produced by normal human T cell blasts with those produced by tumoral Jurkat cells, with the objective of identify proteins associated with tumoral exosomes that could have a previously unrecognized role in malignancy. We have identified 359 and 418 proteins in exosomes from T cell blasts and Jurkat cells, respectively. Interestingly, only 145 (around a 40%) are common. The major proteins in both cases are actin and tubulin isoforms and the common interaction nodes correspond to these cytoskeleton and related proteins, as well as to ribosomal and mRNA granule proteins. We detected 14 membrane proteins that were especially enriched in exosomes from Jurkat cells as compared with T cell blasts. The most abundant of these proteins was valosin-containing protein (VCP), a membrane ATPase involved in ER homeostasis and ubiquitination. In this work, we also show that leukemic cells are more sensitive to cell death induced by the VCP inhibitor DBeQ than normal T cells. Furthermore, VCP inhibition prevents functional exosome secretion only in Jurkat cells, but not in T cell blasts. These results suggest VCP targeting as a new selective pathway to exploit in cancer treatment to prevent tumoral exosome secretion. PMID:27086912

  4. Acute changes in cellular zinc alters zinc uptake rates prior to zinc transporter gene expression in Jurkat cells.

    PubMed

    Holland, Tai C; Killilea, David W; Shenvi, Swapna V; King, Janet C

    2015-12-01

    A coordinated network of zinc transporters and binding proteins tightly regulate cellular zinc levels. Canonical responses to zinc availability are thought to be mediated by changes in gene expression of key zinc transporters. We investigated the temporal relationships of actual zinc uptake with patterns of gene expression in membrane-bound zinc transporters in the human immortalized T lymphocyte Jurkat cell line. Cellular zinc levels were elevated or reduced with exogenous zinc sulfate or N,N,N',N-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), respectively. Excess zinc resulted in a rapid 44 % decrease in the rate of zinc uptake within 10 min. After 120 min, the expression of metallothionein (positive control) increased, as well as the zinc exporter, ZnT1; however, the expression of zinc importers did not change during this time period. Zinc chelation with TPEN resulted in a rapid twofold increase in the rate of zinc uptake within 10 min. After 120 min, the expression of ZnT1 decreased, while again the expression of zinc importers did not change. Overall, zinc transporter gene expression kinetics did not match actual changes in cellular zinc uptake with exogenous zinc or TPEN treatments. This suggests zinc transporter regulation may be the initial response to changes in zinc within Jurkat cells. PMID:26420239

  5. High ACSL5 Transcript Levels Associate with Systemic Lupus Erythematosus and Apoptosis in Jurkat T Lymphocytes and Peripheral Blood Cells

    PubMed Central

    2011-01-01

    Background Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease in which increased apoptosis and decreased apoptotic cells removal has been described as most relevant in the pathogenesis. Long-chain acyl-coenzyme A synthetases (ACSLs) have been involved in the immunological dysfunction of mouse models of lupus-like autoimmunity and apoptosis in different in vitro cell systems. The aim of this work was to assess among the ACSL isoforms the involvement of ACSL2, ACSL4 and ACSL5 in SLE pathogenesis. Findings With this end, we determined the ACSL2, ACSL4 and ACSL5 transcript levels in peripheral blood mononuclear cells (PBMCs) of 45 SLE patients and 49 healthy controls by quantitative real time-PCR (q-PCR). We found that patients with SLE had higher ACSL5 transcript levels than healthy controls [median (range), healthy controls = 16.5 (12.3–18.0) vs. SLE = 26.5 (17.8–41.7), P = 3.9×10 E-5] but no differences were found for ACSL2 and ACSL4. In in vitro experiments, ACSL5 mRNA expression was greatly increased when inducing apoptosis in Jurkat T cells and PBMCs by Phorbol-Myristate-Acetate plus Ionomycin (PMA+Io). On the other hand, short interference RNA (siRNA)-mediated silencing of ACSL5 decreased induced apoptosis in Jurkat T cells up to the control levels as well as decreased mRNA expression of FAS, FASLG and TNF. Conclusions These findings indicate that ACSL5 may play a role in the apoptosis that takes place in SLE. Our results point to ACSL5 as a potential novel functional marker of pathogenesis and a possible therapeutic target in SLE. PMID:22163040

  6. Evaluation of the Genetic Response of U937 and Jurkat Cells to 10-Nanosecond Electrical Pulses (nsEP).

    PubMed

    Roth, Caleb C; Glickman, Randolph D; Tolstykh, Gleb P; Estlack, Larry E; Moen, Erick K; Echchgadda, Ibtissam; Beier, Hope T; Barnes, Ronald A; Ibey, Bennett L

    2016-01-01

    Nanosecond electrical pulse (nsEP) exposure activates signaling pathways, produces oxidative stress, stimulates hormone secretion, causes cell swelling and induces apoptotic and necrotic death. The underlying biophysical connection(s) between these diverse cellular reactions and nsEP has yet to be elucidated. Using global genetic analysis, we evaluated how two commonly studied cell types, U937 and Jurkat, respond to nsEP exposure. We hypothesized that by studying the genetic response of the cells following exposure, we would gain direct insight into the stresses experienced by the cell and in turn better understand the biophysical interaction taking place during the exposure. Using Ingenuity Systems software, we found genes associated with cell growth, movement and development to be significantly up-regulated in both cell types 4 h post exposure to nsEP. In agreement with our hypothesis, we also found that both cell lines exhibit significant biological changes consistent with mechanical stress induction. These results advance nsEP research by providing strong evidence that the interaction of nsEPs with cells involves mechanical stress. PMID:27135944

  7. Evaluation of the Genetic Response of U937 and Jurkat Cells to 10-Nanosecond Electrical Pulses (nsEP)

    PubMed Central

    Glickman, Randolph D.; Tolstykh, Gleb P.; Estlack, Larry E.; Moen, Erick K.; Echchgadda, Ibtissam; Beier, Hope T.; Barnes, Ronald A.; Ibey, Bennett L.

    2016-01-01

    Nanosecond electrical pulse (nsEP) exposure activates signaling pathways, produces oxidative stress, stimulates hormone secretion, causes cell swelling and induces apoptotic and necrotic death. The underlying biophysical connection(s) between these diverse cellular reactions and nsEP has yet to be elucidated. Using global genetic analysis, we evaluated how two commonly studied cell types, U937 and Jurkat, respond to nsEP exposure. We hypothesized that by studying the genetic response of the cells following exposure, we would gain direct insight into the stresses experienced by the cell and in turn better understand the biophysical interaction taking place during the exposure. Using Ingenuity Systems software, we found genes associated with cell growth, movement and development to be significantly up-regulated in both cell types 4 h post exposure to nsEP. In agreement with our hypothesis, we also found that both cell lines exhibit significant biological changes consistent with mechanical stress induction. These results advance nsEP research by providing strong evidence that the interaction of nsEPs with cells involves mechanical stress. PMID:27135944

  8. Liquid Chromatography-Mass Spectrometry Metabolic and Lipidomic Sample Preparation Workflow for Suspension-Cultured Mammalian Cells using Jurkat T lymphocyte Cells

    PubMed Central

    Ulmer, Candice Z.; Yost, Richard A.; Chen, Jing; Mathews, Clayton E.; Garrett, Timothy J.

    2015-01-01

    Metabolomics is the comprehensive study of metabolism as it pertains to an organism or biological system. Lipidomics, a subset discipline of metabolomics, encompasses the study of cellular lipid functions: including pathways, networks, and interactions. The abundance of metabolites and lipids, along with their contribution to health and disease, makes metabolomics a valuable tool for biomarker research. Disease biomarker identification requires a reproducible, sensitive, and accurate analytical platform. Although transcriptomic and proteomic areas have well-established protocols for sample preparation and data processing, the metabolomics field is still developing comparable standardized conventions. Furthermore, of the few comparative LC-MS metabolomic studies that have been applied to mammalian cell cultures, most are targeted to adherent cell lines. The purpose of this work was to optimize a sample preparation workflow for the cellular metabolomic analysis of suspension-cultured mammalian cells using commercially available Jurkat T lymphocytes as a model system. The current investigation evaluated commonly used sample preparation techniques for reproducibility, accuracy, and applicability to untargeted biomarker discovery. Results show ammoniated cell rinsing solutions to be an effective means to remove extracellular components present in the media without causing ion suppression or affecting the integrity of the cellular membrane. Additionally, a novel workflow was designed to allow for the combined analysis of metabolites and lipids from mammalian suspension cells from a single cell pellet. The Folch lipid extraction protocol was coupled to an 80% MeOH metabolite isolation to ensure high extraction efficiency for phospholipids and triacylglycerides. While the workflow was tailored to cells in suspension, it could also be applied to adherent cell lines. PMID:26401069

  9. Potentiation of the store-operated calcium entry (SOCE) induces phytohemagglutinin-activated Jurkat T cell apoptosis.

    PubMed

    Djillani, Alaeddine; Doignon, Isabelle; Luyten, Tomas; Lamkhioued, Bouchaib; Gangloff, Sophie C; Parys, Jan B; Nüße, Oliver; Chomienne, Christine; Dellis, Olivier

    2015-08-01

    Store-operated Ca(2+) entry (SOCE) is the main Ca(2+) entry pathway of non-excitable cells. In the past decade, the activation of this entry has been unveiled, with STIM1, a protein of the endoplasmic reticulum able to sense the intraluminal Ca(2+) content, and Orai1, the pore-forming unit of the Ca(2+) release activated Ca(2+) (CRAC) channels. When Ca(2+) ions are released from the endoplasmic reticulum, STIM1 proteins oligomerize and directly interact with Orai1 proteins, allowing the opening of the CRAC channels and a massive Ca(2+) ion influx known as SOCE. As Ca(2+) is involved in various cellular processes, the discovery of new drugs acting on the SOCE should be of interest to control the cell activity. By testing analogs of 2-aminoethyl diphenylborinate (2-APB), a well known, though not so selective effector of the SOCE, we identified methoxy diethylborinate (MDEB), a molecule able to potentiate the SOCE in three leukocyte and two breast cancer cell lines by increasing the Ca(2+) influx amplitude. Unlike 2-APB, MDEB does not affect the Ca(2+) pumps or the Ca(2+) release from the endoplasmic reticulum. MDEB could therefore represent the first member of a new group of molecules, specifically able to potentiate SOCE. Although not toxic for non-activated Jurkat T cells, it could induce the apoptosis of phytohemagglutinin-stimulated cells. PMID:25963393

  10. Benzo[a]pyrene affects Jurkat T cells in the activated state via the antioxidant response element dependent Nrf2 pathway leading to decreased IL-2 secretion and redirecting glutamine metabolism

    SciTech Connect

    Murugaiyan, Jayaseelan; Rockstroh, Maxie; Wagner, Juliane; Baumann, Sven; Schorsch, Katrin; Trump, Saskia; Lehmann, Irina; Bergen, Martin von; Tomm, Janina M.

    2013-06-15

    There is a clear evidence that environmental pollutants, such as benzo[a]pyrene (B[a]P), can have detrimental effects on the immune system, whereas the underlying mechanisms still remain elusive. Jurkat T cells share many properties with native T lymphocytes and therefore are an appropriate model to analyze the effects of environmental pollutants on T cells and their activation. Since environmental compounds frequently occur at low, not acute toxic concentrations, we analyzed the effects of two subtoxic concentrations, 50 nM and 5 μM, on non- and activated cells. B[a]P interferes directly with the stimulation process as proven by an altered IL-2 secretion. Furthermore, B[a]P exposure results in significant proteomic changes as shown by DIGE analysis. Pathway analysis revealed an involvement of the AhR independent Nrf2 pathway in the altered processes observed in unstimulated and stimulated cells. A participation of the Nrf2 pathway in the change of IL-2 secretion was confirmed by exposing cells to the Nrf2 activator tBHQ. tBHQ and 5 μM B[a]P caused similar alterations of IL-2 secretion and glutamine/glutamate metabolism. Moreover, the proteome changes in unstimulated cells point towards a modified regulation of the cytoskeleton and cellular stress response, which was proven by western blotting. Additionally, there is a strong evidence for alterations in metabolic pathways caused by B[a]P exposure in stimulated cells. Especially the glutamine/glutamate metabolism was indicated by proteome pathway analysis and validated by metabolite measurements. The detrimental effects were slightly enhanced in stimulated cells, suggesting that stimulated cells are more vulnerable to the environmental pollutant model compound B[a]P. - Highlights: • B[a]P affects the proteome of Jurkat T cells also at low concentrations. • Exposure to B[a]P (50 nM, 5 μM) did not change Jurkat T cell viability. • Both B[a]P concentrations altered the IL-2 secretion of stimulated cells.

  11. Caspase-dependant activation of chymotrypsin-like proteases mediates nuclear events during Jurkat T cell apoptosis

    SciTech Connect

    O'Connell, A.R.; Lee, B.W.; Stenson-Cox, C. . E-mail: catherine.stenson@nuigalway.ie

    2006-06-30

    Apoptosis involves a cascade of biochemical and morphological changes resulting in the systematic disintegration of the cell. Caspases are central mediators of this process. Supporting and primary roles for serine proteases as pro-apoptotic mediators have also been highlighted. Evidence for such roles comes largely from the use of pharmacological inhibitors; as a consequence information regarding their apoptotic function and biochemical properties has been limited. Here, we circumvented limitations associated with traditional serine protease inhibitors through use of a fluorescently labelled inhibitor of serine proteases (FLISP) that allowed for analysis of the specificity, regulation and positioning of apoptotic serine proteases within a classical apoptotic cascade. We demonstrate that staurosporine triggers a caspase-dependant induction of chymotrypsin-like activity in the nucleus of apoptotic Jurkat T cells. We show that serine protease activity is required for the generation of late stage nuclear events including condensation, fragmentation and DNA degradation. Furthermore, we reveal caspase-dependant activation of two chymotrypsin-like protein species that we hypothesize mediate cell death-associated nuclear events.

  12. The influence of 8-prenylnaringenin on the activity of voltage-gated Kv1.3 potassium channels in human Jurkat T cells.

    PubMed

    Gąsiorowska, Justyna; Teisseyre, Andrzej; Uryga, Anna; Michalak, Krystyna

    2012-12-01

    Using the whole-cell patch-clamp technique, we investigated the influence of 8-prenylnaringenin on the activity of the voltage-gated Kv1.3 potassium channels in the human leukemic T lymphocyte cell line Jurkat. 8-prenylnaringenin is a potent plant-derived phytoestrogen that has been found to inhibit cancer cell proliferation. The results show that it inhibited the Kv1.3 channels in a concentration-dependent manner. Complete inhibition occurred at concentrations higher than 10 μM. The inhibitory effect of 8-prenylnaringenin was reversible. It was accompanied by a significant acceleration of channel inactivation without any pronounced change in the activation rate. Of the naringenin derivatives tested to date, 8-prenylnaringenin is the most potent inhibitor of the Kv1.3 channels. The potency of the inhibition may be due to the presence of a prenyl group in the molecule of this flavonoid. The inhibition of the Kv1.3 channels might be involved in the antiproliferative and pro-apoptotic effects of 8-prenylnaringenin that have been observed in cancer cell lines expressing these channels. PMID:22933043

  13. Characterization of the EBV/C3d receptor on the human Jurkat T cell line: evidence for a novel transcript.

    PubMed

    Sinha, S K; Todd, S C; Hedrick, J A; Speiser, C L; Lambris, J D; Tsoukas, C D

    1993-06-15

    EBV is a human herpes virus that causes mononucleosis and is associated with various tumors. EBV infects cells via the CR2 that was previously thought to be expressed only on the surface of B cells and certain epithelial cells. Recent findings in our laboratory and those of others, however, have shown that the EBV receptor is also present on T cells. Our study shows that Jurkat human T cells have a molecule that reacts with both anti-CR2 antibodies and the third component of complement, C3. Furthermore, the data indicate that this molecule binds EBV detected by incubation with biotin-conjugated virus and streptavidin phycoerythrin. Viral binding is specific, as it is inhibited by nonconjugated virus, with anti-CR2 antibodies, and with an antibody reactive with the glycoprotein (gp350) that EBV uses to bind CR2. In addition, EBV variably infects Jurkat cells as demonstrated by the presence of transcripts of Epstein Barr nuclear Ag (EBNA-1) using the polymerase chain reaction. Immunoprecipitation experiments with anti-CR2 antibodies and SDS-PAGE analysis reveal a protein with an apparent molecular mass of 155 kDa which is higher than the one seen in B cells. The size of this molecule is reduced to 119 kDa upon endoglycosidase F treatment. Northern blot analysis of Jurkat poly(A)+ RNA shows a transcript of 4.7 kb upon probing with the B cell CR2 cDNA. This size is consistent with that of B cell CR2 mRNA. Two cDNA clones were identified upon screening of a Jurkat cell cDNA library with the B cell CR2 cDNA. One of the clones possesses an identical nucleotide sequence to the one corresponding to B cell CR2, whereas the other represents a differentially spliced transcript which lacks the exon 8b of B cell CR2. Analysis of Jurkat and Raji mRNA by PCR demonstrated the presence of this novel splice variant in both cell lines. PMID:8390533

  14. Inhibitors of MAPK Pathway ERK1/2 or p38 Prevent the IL-1β-induced Up-regulation of SRP72 Autoantigen in Jurkat Cells*

    PubMed Central

    Arana-Argáez, Victor E.; Delgado-Rizo, Vidal; Pizano-Martínez, Oscar E.; Martínez-Garcia, Erika A.; Martín-Márquez, Beatriz T.; Muñoz-Gómez, Andrea; Petri, Marcelo H.; Armendáriz-Borunda, Juan; Espinosa-Ramírez, Guillermo; Zúñiga-Tamayo, Diego A.; Herrera-Esparza, Rafael; Vázquez-Del Mercado, Mónica

    2010-01-01

    Phosphorylation is the most important post-translational event at a cellular level that is regulated by protein kinases. MAPK is a key player in the important cellular signaling pathway. It has been hypothesized that phosphorylation might have a role in the induction of break tolerance against some autoantigens such as SRP72. The aim of this study was to explore the pathways of phosphorylation and overexpression of the SRP72 polypeptide, using an in vitro model of Jurkat cells stimulated by recombinant human (rh)IL-1β in the presence of MAPK inhibitors. We used Jurkat cells as a substrate stimulated with rhIL-1β in the presence of MAPK inhibitors at different concentrations in a time course in vitro experiment by immunoprecipitation, immunoprecipitation-Western blotting, and real time PCR. Our results showed that rhIL-1β causes up-regulation of protein expression and phosphorylation of SRP72 in Jurkat cells. Inhibitors of the MAPK pathway ERK1/2 or p38α/β down-regulate the expression of SRP72 autoantigen in Jurkat cells stimulated by rhIL-1β. Our results highlight the importance of studying the pathways of activation and overexpression of autoantigens. It will be necessary to perform careful research on various kinases pathways, including MAPK in dermatomyositis and other rheumatic diseases, to help to explain the routes of activation and inhibition of autoantigens. The understanding of this process may help to develop new therapies to prevent and control the loss of tolerance toward own normal proteins. PMID:20729213

  15. ETS1 transactivates the human GM-CSF promoter in Jurkat T cells stimulated with PMA and ionomycin.

    PubMed

    Thomas, R S; Tymms, M J; Seth, A; Shannon, M F; Kola, I

    1995-11-16

    Activation of T helper cells results in coordinate expression of a number of cytokines involved in differentiation, proliferation and activation of the haematopoietic system. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is one such cytokine whose increased expression results partly from increases in transcription. Cis-acting elements with NF kappa B, AP-1 and ETS-like motifs have been identified in the promoter region of the GM-CSF gene, which are important for transcriptional activity following PMA and ionomycin stimulation. A number of the ETS family of transcription factors are expressed in T cells, including ETS1 and ELF1. Here we describe the ability of these factors to interact with a site (GM5), located within the CLE0 element, -47 to -40 upstream of the GM-CSF transcription initiation site. Exogenous ETS1, but not ELF1, can transactivate GM-CSF, through the GM5 site, in a PMA/ionomycin dependent manner. Other unidentified ETS-like factors present in Jurkat cells are also capable of binding GM5. Mutation of the core ETS binding site from -GGAA- to -GGAT- prevents the binding of ETS-like factors with the exception of ETS1. The GM-CSF promoter, modified in this way to be ETS1 specific, is fully responsive to PMA/ionomycin induction, in addition to ETS1 transactivation in the presence of PMA and ionomycin. Together these data suggest that ETS1 may be involved in mediating the increased GM-CSF production associated with T cell activation. PMID:7478534

  16. Apoptogenic activity of auraptene of Zanthoxylum schinifolium toward human acute leukemia Jurkat T cells is associated with ER stress-mediated caspase-8 activation that stimulates mitochondria-dependent or -independent caspase cascade.

    PubMed

    Jun, Do Y; Kim, Jun S; Park, Hae S; Han, Cho R; Fang, Zhe; Woo, Mi H; Rhee, In K; Kim, Young H

    2007-06-01

    To isolate pharmacologically safe compounds that can induce apoptosis of tumor cells, leaves of an aromatic plant (Zanthoxylum schinifolium), which are widely used as a food flavor and herbal medicine in Korea and Japan, were sequentially extracted by organic solvents. An apoptogenic ingredient in the methylene chloride extract was further purified by silica gel column chromatography and identified as auraptene (AUR). The IC(50) value of AUR against Jurkat T cells was 16.5 microg/ml. After the treatment of Jurkat T cells with AUR, the endoplasmic reticulum (ER) stress-mediated activation of caspase-12 and -8 and subsequent apoptotic events including c-Jun N-terminal kinase (JNK) activation, cleavage of FLICE inhibitory protein and Bid, mitochondrial cytochrome c release, activation of caspase-9 and -3, degradation of poly (ADP-ribose) polymerase and apoptotic DNA fragmentation were induced in a dose-dependent manner. The cytotoxicity of AUR was not blocked by the anti-Fas neutralizing antibody ZB-4. The AUR-induced cytotoxicity and apoptotic events were abrogated by ectopic over-expression of Bcl-xL or addition of the pan-caspase inhibitor z-VAD-fmk. The individual or simultaneous addition of the m-calpain inhibitor (E64d), JNK inhibitor (SP600125) and mitochondrial permeability transition pore inhibitor (CsA) failed to prevent apoptotic events including caspase-8 activation and Bid cleavage, unless the caspase-8 inhibitor (z-IETD-fmk) was combined, whereas AUR-induced caspase-12 activation was sustained even in the concomitant presence of z-IETD-fmk. These results demonstrated that the apoptotic effect of AUR on Jurkat T cells was exerted by the ER stress-mediated activation of caspase-8, and the subsequent induction of mitochondria-dependent or -independent activation of caspase cascade, which could be suppressed by Bcl-xL. PMID:17301064

  17. A novel 2,3-benzodiazepine-4-one derivative AMPA antagonist inhibits G2/M transition and induces apoptosis in human leukemia Jurkat T cell line.

    PubMed

    Parenti, S; Casagrande, G; Montanari, M; Espahbodinia, M; Ettari, R; Grande, A; Corsi, L

    2016-05-01

    It has been shown that the antagonism of glutamate receptors activity was able inhibit proliferation and induce apoptosis in several neuronal and non-neuronal cancer cell lines. In addition, it has been shown that glutamate might facilitate the spread and growth of leukemia T cells through interactions with AMPA receptors. The aim of the present study was to investigate the modulation of cell cycle elicited by a novel 2,3-benzodiazepine-4-one non-competitive AMPA antagonist derivative in the human leukemia Jurkat T cells. Our results indicated that the 1-(4-amino-3,5-dimethylphenyl)-3,5-dihydro-7,8-ethylenedioxy-4h-2,3-benzodiazepin-4-one, named 1g, exerted a significant growth inhibition of leukemia Jurkat T cells in a time and dose dependent manner, arresting the transition of G2/M phase through activation of Myt-1. The molecule also induced apoptosis through the enhanced expression of the pro-apoptotic p53, and the inhibition of Bcl-2, and Bcl-xl, followed by the activation of caspase-3. The results suggested that compound 1g might act mostly as a cytostatic rather than cytotoxic compound. Although further studies are necessary, in order to identify others specific pathways involved in the activity of the present molecule, the presented results identified a novel molecule acting on specific G2/M checkpoint regulation pathway. Finally, our data suggest that compound 1g might be a good molecule for future development in the cancer research. PMID:27178220

  18. Metabolic labelling of membrane microdomains/rafts in Jurkat cells indicates the presence of glycerophospholipids implicated in signal transduction by the CD3 T-cell receptor.

    PubMed Central

    Rouquette-Jazdanian, Alexandre K; Pelassy, Claudette; Breittmayer, Jean-Philippe; Cousin, Jean-Louis; Aussel, Claude

    2002-01-01

    Cell membranes contain sphingolipids and cholesterol, which cluster together in distinct domains called rafts. The outer-membrane leaflet of these peculiar membrane domains contains glycosylphosphatidylinositol-anchored proteins, while the inner leaflet contains proteins implicated in signalling, such as the acylated protein kinase p56(lck) and the palmitoylated adaptator LAT (linker for activation of T-cells). We present here an approach to study the lipid composition of rafts and its change upon T-cell activation. Our method is based on metabolic labelling of Jurkat T-cells with different precursors of glycerophospholipid synthesis, including glycerol and fatty acids with different lengths and degrees of saturation as well as phospholipid polar head groups. The results obtained indicate that lipid rafts isolated by the use of sucrose density-gradient centrifugation after Triton X-100 extraction in the cold, besides sphingolipids and cholesterol, contain unambiguously all classes of glycerophospholipids: phosphatidylserine, phosphatidylinositol, phosphatidylethanolamine and phosphatidylcholine. Fatty acid labelling shows that lipid rafts are labelled preferentially with saturated fatty acids while the rest of the plasma membrane incorporates mostly long-chained polyunsaturated fatty acids. To see whether the raft composition as measured by metabolic labelling of phospholipids is involved in T-cell activation, we investigated the production of sn-1,2-diacylglycerol (DAG) in CD3-activated cells. DAG production occurs within rafts, confirming previous demonstration of protein kinase C translocation into membrane microdomains. Our data demonstrate that raft disorganization by methyl-beta-cyclodextrin impairs both CD3-induced DAG production and changes in cytosolic Ca(2+) concentration. These lines of evidence support the conclusion that the major events in T-cell activation occur within or due to lipid rafts. PMID:11964165

  19. Mollugin induces apoptosis in human Jurkat T cells through endoplasmic reticulum stress-mediated activation of JNK and caspase-12 and subsequent activation of mitochondria-dependent caspase cascade regulated by Bcl-xL

    SciTech Connect

    Kim, Sun Mi; Park, Hae Sun; Jun, Do Youn; Woo, Hyun Ju; Woo, Mi Hee; Yang, Chae Ha; Kim, Young Ho

    2009-12-01

    Exposure of Jurkat T cells to mollugin (15-30 muM), purified from the roots of Rubia cordifolia L., caused cytotoxicity and apoptotic DNA fragmentation along with mitochondrial membrane potential disruption, mitochondrial cytochrome c release, phosphorylation of c-Jun N-terminal kinase (JNK), activation of caspase-12, -9, -7, -3, and -8, cleavage of FLIP and Bid, and PARP degradation, without accompanying necrosis. While these mollugin-induced cytotoxicity and apoptotic events including activation of caspase-8 and mitochondria-dependent activation of caspase cascade were completely prevented by overexpression of Bcl-xL, the activation of JNK and caspase-12 was prevented to much lesser extent. Pretreatment of the cells with the pan-caspase inhibitor (z-VAD-fmk), the caspase-9 inhibitor (z-LEHD-fmk), the caspase-3 inhibitor (z-DEVD-fmk) or the caspase-12 inhibitor (z-ATAD-fmk) at the minimal concentration to prevent mollugin-induced apoptosis appeared to completely block the activation of caspase-7 and -8, and PARP degradation, but failed to block the activation of caspase-9 and -3 with allowing a slight enhancement in the level of JNK phosphorylation. Both FADD-positive wild-type Jurkat clone A3 and FADD-deficient Jurkat clone I2.1 exhibited a similar susceptibility to the cytotoxicity of mollugin, excluding involvement of Fas/FasL system in triggering mollugin-induced apoptosis. Normal peripheral T cells were more refractory to the cytotoxicity of mollugin than were Jurkat T cells. These results demonstrated that mollugin-induced cytotoxicity in Jurkat T cells was mainly attributable to apoptosis provoked via endoplasmic reticulum (ER) stress-mediated activation of JNK and caspase-12, and subsequent mitochondria-dependent activation of caspase-9 and -3, leading to activation of caspase-7 and -8, which could be regulated by Bcl-xL.

  20. Evaluation of Synergetic Anticancer Activity of Berberine and Curcumin on Different Models of A549, Hep-G2, MCF-7, Jurkat, and K562 Cell Lines.

    PubMed

    Balakrishna, Acharya; Kumar, M Hemanth

    2015-01-01

    Ayurvedic system of medicine is using Berberis aristata and Curcuma longa herbs to treat different diseases including cancer. The study was performed to evaluate the synergetic anticancer activity of Berberine and Curcumin by estimating the inhibition of the cell proliferation by cytotoxicity assay using MTT method on specified human cell lines (A549, Hep-G2, MCF-7, Jurkat, and K562). All the cells were harvested from the culture and seeded in the 96-well assay plates at seeding density of 2.0 × 10(4) cells/well and were incubated for 24 hours. Test items Berberine with Curcumin (1 : 1), Curcumin 95% pure, and Berberine 95% pure were exposed at the concentrations of 1.25, 0.001, and 0.5 mg/mL, respectively, and incubated for a period of 48 hours followed by dispensing MTT solution (5 mg/mL). The cells were incubated at 37 ± 1°C for 4 hours followed by addition of DMSO for dissolving the formazan crystals and absorbance was read at 570 nm. Separate wells were prepared for positive control, controls (only medium with cells), and blank (only medium). The results had proven the synergetic anticancer activity of Berberine with Curcumin inducing cell death greater percentage of >77% when compared to pure curcumin with <54% and pure Berberine with <45% on average on all cell line models. PMID:26247019

  1. Evaluation of Synergetic Anticancer Activity of Berberine and Curcumin on Different Models of A549, Hep-G2, MCF-7, Jurkat, and K562 Cell Lines

    PubMed Central

    Balakrishna, Acharya; Kumar, M. Hemanth

    2015-01-01

    Ayurvedic system of medicine is using Berberis aristata and Curcuma longa herbs to treat different diseases including cancer. The study was performed to evaluate the synergetic anticancer activity of Berberine and Curcumin by estimating the inhibition of the cell proliferation by cytotoxicity assay using MTT method on specified human cell lines (A549, Hep-G2, MCF-7, Jurkat, and K562). All the cells were harvested from the culture and seeded in the 96-well assay plates at seeding density of 2.0 × 104 cells/well and were incubated for 24 hours. Test items Berberine with Curcumin (1 : 1), Curcumin 95% pure, and Berberine 95% pure were exposed at the concentrations of 1.25, 0.001, and 0.5 mg/mL, respectively, and incubated for a period of 48 hours followed by dispensing MTT solution (5 mg/mL). The cells were incubated at 37 ± 1°C for 4 hours followed by addition of DMSO for dissolving the formazan crystals and absorbance was read at 570 nm. Separate wells were prepared for positive control, controls (only medium with cells), and blank (only medium). The results had proven the synergetic anticancer activity of Berberine with Curcumin inducing cell death greater percentage of >77% when compared to pure curcumin with <54% and pure Berberine with <45% on average on all cell line models. PMID:26247019

  2. JS-K, a nitric oxide-releasing prodrug, modulates ß-catenin/TCF signaling in leukemic Jurkat cells: evidence of an S-nitrosylated mechanism.

    PubMed

    Nath, Niharika; Chattopadhyay, Mitali; Pospishil, Liliya; Cieciura, Lucyna Z; Goswami, Satindra; Kodela, Ravinder; Saavedra, Joseph E; Keefer, Larry K; Kashfi, Khosrow

    2010-12-01

    β-Catenin is a central player of the Wnt signaling pathway that regulates cell-cell adhesion and may promote leukemia cell proliferation. We examined whether JS-K, an NO-donating prodrug, modulates the Wnt/β-catenin/TCF-4 signaling pathway in Jurkat T-Acute Lymphoblastic Leukemia cells. JS-K inhibited Jurkat T cell growth in a concentration and time-dependent manner. The IC(50)s for cell growth inhibition were 14±0.7 and 9±1.2μM at 24 and 48h, respectively. Treatment of the cells with JS-K for 24h, caused a dose-dependent increase in apoptosis from 16±3.3% at 10μM to 74.8±2% at 100μM and a decrease in proliferation. This growth inhibition was also due, in part, to alterations in the different phases of the cell cycle. JS-K exhibited a dose-dependent cytotoxicity as measured by LDH release at 24h. However, between 2 and 8h, LDH release was less than 20% for any indicated JS-K concentration. The β-catenin/TCF-4 transcriptional inhibitory activity was reduced by 32±8, 63±5, and 93±2% at 2, 10, and 25μM JS-K, respectively, based on luciferase reporter assays. JS-K reduced nuclear β-catenin and cyclin D1 protein levels, but cytosolic β-catenin expression did not change. Based on a time-course assay of S-nitrosylation of proteins by a biotin switch assay, S-nitrsolyation of nuclear β-catenin was determined to precede its degradation. A comparison of the S-nitrosylated nuclear β-catenin to the total nuclear β-catenin showed that β-catenin protein levels were degraded at 24h, while S-nitrosylation of β-catenin occurred earlier at 0-6h. The NO scavenger PTIO abrogated the JS-K mediated degradation of β-catenin demonstrating the need for NO. PMID:20797387

  3. FasL-triggered death of Jurkat cells requires caspase 8-induced, ATP-dependent cross-talk between Fas and the purinergic receptor P2X(7).

    PubMed

    Aguirre, Adam; Shoji, Kenji F; Sáez, Juan C; Henríquez, Mauricio; Quest, Andrew F G

    2013-02-01

    Fas ligation via the ligand FasL activates the caspase-8/caspase-3-dependent extrinsic death pathway. In so-called type II cells, an additional mechanism involving tBid-mediated caspase-9 activation is required to efficiently trigger cell death. Other pathways linking FasL-Fas interaction to activation of the intrinsic cell death pathway remain unknown. However, ATP release and subsequent activation of purinergic P2X(7) receptors (P2X(7)Rs) favors cell death in some cells. Here, we evaluated the possibility that ATP release downstream of caspase-8 via pannexin1 hemichannels (Panx1 HCs) and subsequent activation of P2X(7)Rs participate in FasL-stimulated cell death. Indeed, upon FasL stimulation, ATP was released from Jurkat cells in a time- and caspase-8-dependent manner. Fas and Panx1 HCs colocalized and inhibition of the latter, but not connexin hemichannels, reduced FasL-induced ATP release. Extracellular apyrase, which hydrolyzes ATP, reduced FasL-induced death. Also, oxidized-ATP or Brilliant Blue G, two P2X(7)R blockers, reduced FasL-induced caspase-9 activation and cell death. These results represent the first evidence indicating that the two death receptors, Fas and P2X(7)R connect functionally via caspase-8 and Panx1 HC-mediated ATP release to promote caspase-9/caspase-3-dependent cell death in lymphoid cells. Thus, a hitherto unsuspected route was uncovered connecting the extrinsic to the intrinsic pathway to amplify death signals emanating from the Fas receptor in type II cells. PMID:22806078

  4. Effect of vibrational stress and spaceflight on regulation of heat shock proteins hsp70 and hsp27 in human lymphocytes (Jurkat)

    NASA Technical Reports Server (NTRS)

    Cubano, L. A.; Lewis, M. L.

    2001-01-01

    Heat shock protein levels are increased in cells as a result of exposure to stress. To determine whether heat shock protein regulation could be used to evaluate stress in cells during spaceflight, the response of Jurkat cells to spaceflight and simulated space shuttle launch vibration was investigated by evaluating hsp70 and hsp27 gene expression. Gene expression was assessed by reverse transcription-polymerase chain reaction using mRNA extracted from vibrated, nonvibrated, space-flown, and ground control cells. Results indicate that mechanical stresses of vibration and low gravity do not up-regulate the mRNA for hsp70, although the gene encoding hsp27 is up-regulated by spaceflight but not by vibration. In ground controls, the mRNA for hsp70 and hsp27 increased with time in culture. We conclude that hsp70 gene expression is a useful indicator of stress related to culture density but is not an indicator of the stresses of launch vibration or microgravity. Up-regulation of hsp27 gene expression in microgravity is a new finding.

  5. Modulating Heat Shock Proteins 70 and 90 Expression by Low Power Laser Irradiation (635nm and 780nm) in Jurkat E6.1 T-lymphocyte Leukemia Cell Line

    PubMed Central

    Ad’hiah, Ali Hussein; Al-Ameri, Layla Mohammed Hassan; Maki, Amel Mustfa; Wang, Qiuyu; ALQaisi, Mayada Hameed

    2015-01-01

    Introduction: Heat shock proteins (HSPs) are molecular chaperones involved in protein folding, stability and turnover, and due to their role in cancer progression, the effect of low power laser irradiation (LPLI) on the expression of HSP70 and HSP90 in Jurkat E6.1 T-lymphocyte leukemia (JELT) cell line was investigated in vitro. Methods: JETL cells were irradiated with LPLI at 635nm and 780m wavelengths (energy density 9.174 J/cm2), and assessed for the expression of HSP70 and HSP90 by flow cytometry after 24, 48 and 72 incubation time periods (ITPs). Results: At 24 hours ITP post-irradiation, control cultures showed that 10.7% of cells expressed HSP70, while LPLI cultures at 635nm and 780nm manifested a higher expression (32.1and 21.3%, respectively), and the difference was significant (P ≤ 0.05). However, at 48 hours ITP, the three means were decreased but approximated (5.6, 4.9 and 6.2%, respectively), while at 72 hours ITP, they were markedly increased (45.2, 76.5 and 66.7%, respectively). In contrast, HSP90 responded differently to LPLI. At 24 hours ITP, control cultures and 780nm cultures showed a similar expression (55.9 and 55.9%, respectively), but both means were significantly higher than that of 635nm cultures (24.0%). No such difference was observed at 48 hours ITP, and at 72 hours ITP, control cultures and 635nm cultures shared approximated means (31.7 and 35.6%, respectively); but both means were significantly higher than the observed mean in 780nm cultures (15.2%). Conclusion: The results highlighted that HSP70 and HSP90 expression responded differently to LPLI in JETL cells; an observation that may pave the way for further investigations in malignant cells PMID:25699163

  6. Effects of teicoplanin on cell number of cultured cell lines

    PubMed Central

    Kashkolinejad-Koohi, Tahere; Saadat, Iraj

    2015-01-01

    Teicoplanin is a glycopeptide antibiotic with a wide variation in human serum half-life. It is also a valuable alternative of vancomycin. There is however no study on its effect on cultured cells. The aim of the present study was to test the effect of teicoplanin on cultured cell lines CHO, Jurkat E6.1 and MCF-7. The cultured cells were exposed to teicoplanin at final concentrations of 0–11000 μg/ml for 24 hours. To determine cell viability, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test was performed. At low concentrations of teicoplanin the numbers of cultured cells (due to cell proliferation) were increased in the three cell lines examined. The maximum cell proliferation rates were observed at concentrations of 1000, 400, and 200 μg/ml of teicoplanin for CHO, MCF-7 and Jurkat cell lines, respectively. Cell toxicity was observed at final concentrations over 2000, 6000, and 400 μg/ml of teicoplanin for CHO, MCF-7 and Jurkat cell lines, respectively. A dose-dependent manner of cell toxicity was observed. Our present findings indicated that teicoplanin at clinically used concentrations induced cell proliferation. It should therefore be used cautiously, particularly in children, pregnant women and patients with cancer.

  7. Fas/APO-1 protein is increased in spaceflown lymphocytes (Jurkat)

    NASA Technical Reports Server (NTRS)

    Cubano, L. A.; Lewis, M. L.

    2000-01-01

    Human lymphocytes flown on the Space Shuttle respond poorly to mitogen stimulation and populations of the lymphoblastoid T cell line, Jurkat, manifest growth arrest, increase in apoptosis and time- and microgravity-dependent increases in the soluble form of the cell death factor, Fas/APO-1 (sFas). The potential role of apoptosis in population dynamics of space-flown lymphocytes has not been investigated previously. We flew Jurkat cells on Space Transportation System (STS)-80 and STS-95 to determine whether apoptosis and the apparent microgravity-related release of sFas are characteristic of lymphocytes in microgravity. The effects of spaceflight and ground-based tests simulating spaceflight experimental conditions, including high cell density and low serum concentration, were assessed. Immunofluorescence microscopy showed increased cell associated Fas in flown cells. Results of STS-80 and STS-95 confirmed increase in apoptosis during spaceflight and the release of sFas as a repeatable, time-dependent and microgravity-related response. Ground-based tests showed that holding cells at 1.5 million/ml in medium containing 2% serum before launch did not increase sFas. Reports of increased Fas in cells of the elderly and the increases in spaceflown cells suggest possible similarities between aging and spaceflight effects on lymphocytes.

  8. Highly Oxygenated Sesquiterpene Lactones from Cousinia aitchisonii and their Cytotoxic Properties: Rhaserolide Induces Apoptosis in Human T Lymphocyte (Jurkat) Cells via the Activation of c-Jun n-terminal Kinase Phosphorylation.

    PubMed

    Iranshahy, Milad; Tayarani-Najaran, Zahra; Kasaian, Jamal; Ghandadi, Morteza; Emami, Seyed Ahmad; Asili, Javad; Chandran, Jima N; Schneider, Bernd; Iranshahi, Mehrdad

    2016-02-01

    Infrared-guided chromatographic fractionation of sesquiterpene lactones from the extracts of Cousinia aitchisonii and Cousinia concolor led to the isolation of five pure compounds. A new sesquiterpene lactone, namely, aitchisonolide, and two known sesquiterpene lactones (desoxyjanerin and rhaserolide) were isolated from C. aitchisonii and two known lignans (arctiin and arctigenin) from C. concolor. The structures of these compounds were elucidated by one-dimensional and two-dimensional nuclear magnetic resonance techniques, as well as high-resolution mass spectrometry. The purified and characterized compounds were subjected to cytotoxicity assay. The sesquiterpene lactones desoxyjanerin and rhaserolide showed significant cytotoxic activities against five different cancer cell lines and the normal human embryonic kidney cell line. Rhaserolide was chosen to evaluate the possible mechanism of action. Western blot analysis revealed that rhaserolide could induce apoptosis in Jurkat cells via the activation of c-Jun n-terminal kinase phosphorylation. PMID:26581585

  9. Challenge of human Jurkat T-cells with the adenylate cyclase activator forskolin elicits major changes in cAMP phosphodiesterase (PDE) expression by up-regulating PDE3 and inducing PDE4D1 and PDE4D2 splice variants as well as down-regulating a novel PDE4A splice variant.

    PubMed Central

    Erdogan, S; Houslay, M D

    1997-01-01

    cells. Forskolin treatment led to a marked decrease of this novel PDE4A species and allowed the detection of a strong signal for an approximately 67 kDa PDE4D species, suggested to be PDE4D1, but did not induce PDE4B and PDE4C isoforms. Elevation of intracellular cAMP concentrations in Jurkat T-cells thus exerts a highly selective effect on the transcriptional activity of the genes encoding the various PDE4 isoforms. This leads to the down-regulation of a novel PDE4A splice variant and the induction of PDE4D1 and PDE4D2 splice variants, leading to a net increase in the total PDE4 activity of Jurkat T-cells. PMID:9003416

  10. Cytotoxicity of diacetoxyscirpenol is associated with apoptosis by activation of caspase-8 and interruption of cell cycle progression by down-regulation of cdk4 and cyclin B1 in human Jurkat T cells

    SciTech Connect

    Jun, Do Youn; Kim, Jun Seok; Park, Hae Sun; Song, Woo Sun; Bae, Young Seuk; Kim, Young Ho . E-mail: ykim@knu.ac.kr

    2007-07-15

    To understand the mechanism underlying T-cell toxicity of diacetoxyscirpenol (DAS) from Fusarium sambucinum, its apoptogenic as well as growth retardation activity was investigated in human Jurkat T cells. Exposure to DAS (0.01-0.15 {mu}M) caused apoptotic DNA fragmentation along with caspase-8 activation, Bid cleavage, mitochondrial cytochrome c release, activation of caspase-9 and caspase-3, and PARP degradation, without any alteration in the levels of Fas or FasL. Under these conditions, necrosis was not accompanied. The cytotoxicity of DAS was not blocked by the anti-Fas neutralizing antibody ZB-4. Although the DAS-induced apoptotic events were completely prevented by overexpression of Bcl-xL, the cells overexpressing Bcl-xL were unable to divide in the presence of DAS, resulting from the failure of cell cycle progression possibly due to down-regulation in the protein levels of cdk4 and cyclin B1. The DAS-mediated apoptosis and activation of caspase-8, -9, and -3 were abrogated by either pan-caspase inhibitor (z-VAD-fmk) or caspase-8 inhibitor (z-IETD-fmk). While the DAS-mediated apoptosis and activation of caspase-9 and caspase-3 were slightly suppressed by the mitochondrial permeability transition pore inhibitor (CsA), both caspase-8 activation and Bid cleavage were not affected by CsA. The activated normal peripheral T cells possessed a similar susceptibility to the cytotoxicity of DAS. These results demonstrate that the T-cell toxicity of DAS is attributable to not only apoptosis initiated by caspase-8 activation and subsequent mitochondrion-dependent or -independent activation of caspase cascades, which can be regulated by Bcl-xL, but also interruption of cell cycle progression caused by down-regulation of cdk4 and cyclin B1 proteins.

  11. Prometaphase arrest-dependent phosphorylation of Bcl-2 family proteins and activation of mitochondrial apoptotic pathway are associated with 17α-estradiol-induced apoptosis in human Jurkat T cells.

    PubMed

    Han, Cho Rong; Jun, Do Youn; Kim, Yoon Hee; Lee, Ji Young; Kim, Young Ho

    2013-10-01

    In Jurkat T cell clone (JT/Neo), G2/M arrest, apoptotic sub-G1 peak, mitochondrial membrane potential (Δψm) loss, and TUNEL-positive DNA fragmentation were induced following exposure to 17α-estradiol (17α-E2), whereas none of these events (except for G2/M arrest) were induced in Jurkat cells overexpressing Bcl-2 (JT/Bcl-2). Under these conditions, phosphorylation at Thr161 and dephosphorylation at Tyr15 of Cdk1, upregulation of cyclin B1 level, histone H1 phosphorylation, Cdc25C phosphorylation at Thr-48, Bcl-2 phosphorylation at Thr-56 and Ser-70, Mcl-1 phosphorylation, and Bim phosphorylation were detected in the presence of Bcl-2 overexpression. However, the 17α-E2-induced upregulation of Bak levels, activation of Bak, activation of caspase-3, and PARP degradation were abrogated by Bcl-2 overexpression. In the presence of the G1/S blocking agent hydroxyurea, 17α-E2 failed to induce G2/M arrest and all apoptotic events including Cdk1 activation and phosphorylation of Bcl-2, Mcl-1 and Bim. The 17α-E2-induced phosphorylation of Bcl-2 family proteins and mitochondrial apoptotic events were suppressed by a Cdk1 inhibitor but not by aurora A and aurora B kinase inhibitors. Immunofluorescence microscopic analysis showed that an aberrant bipolar microtubule array, incomplete chromosome congression at the metaphase plate, and prometaphase arrest, which was reversible, were the underlying factors for 17α-E2-induced mitotic arrest. The in vitro microtubule polymerization assay showed that 17α-E2 could directly inhibit microtubule formation. These results show that the apoptogenic activity of 17α-E2 was due to the impaired mitotic spindle assembly causing prometaphase arrest and prolonged Cdk1 activation, the phosphorylation of Bcl-2, Mcl-1 and Bim, and the activation of Bak and mitochondria-dependent caspase cascade. PMID:23707954

  12. microRNA let-7c is essential for the anisomycin-elicited apoptosis in Jurkat T cells by linking JNK1/2 to AP-1/STAT1/STAT3 signaling.

    PubMed

    Zhou, Zhiwei; Lu, Xijian; Wang, Jin; Xiao, Jia; Liu, Jing; Xing, Feiyue

    2016-01-01

    Anisomycin, an antibiotic produced by Streptomyces griseolus, strongly induces apoptosis in various tumor cells in vitro, superior dramatically to adriamycin. The present study aims to elucidate its detailed mechanistic process. The results showed that anisomycin sufficiently promoted the apoptosis in human leukemic Jurkat T cells at a quite low dose. microRNA let-7c (let-7c) contributed to the anisomycin-induced apoptosis, which could be abrogated by the inactivation of JNK signaling. The let-7c over-expression and the addition of its mimics facilitated the activation of AP-1, STAT1 and Bim by linking JNK1/2 to AP-1/STAT1, but rather inhibited the activation of STAT3 and Bcl-xL by connecting JNK1/2 to STAT3, followed by the augmented apoptosis in the cells. The let-7c deficiency reduced the AP-1, STAT1 and Bim activities, and enhanced the STAT3 and Bcl-xL, alleviating the anisomycin-induced apoptosis. The knockdown of the bim gene repressed the anisomycin-boosted apoptosis through the attenuation of the active Bak and Bax. The findings indicate for the first time that miR let-7c is essential for the anisomycin-triggered apoptosis by linking JNK1/2 to AP-1/STAT1/STAT3/Bim/Bcl-xL/Bax/Bak signaling. This provides a novel insight into the mechanism by which anisomycin leads to the tumor cell apoptosis, potentially laying the foundations for its development and clinical application. PMID:27087117

  13. microRNA let-7c is essential for the anisomycin-elicited apoptosis in Jurkat T cells by linking JNK1/2 to AP-1/STAT1/STAT3 signaling

    PubMed Central

    Zhou, Zhiwei; Lu, Xijian; Wang, Jin; Xiao, Jia; Liu, Jing; Xing, Feiyue

    2016-01-01

    Anisomycin, an antibiotic produced by Streptomyces griseolus, strongly induces apoptosis in various tumor cells in vitro, superior dramatically to adriamycin. The present study aims to elucidate its detailed mechanistic process. The results showed that anisomycin sufficiently promoted the apoptosis in human leukemic Jurkat T cells at a quite low dose. microRNA let-7c (let-7c) contributed to the anisomycin-induced apoptosis, which could be abrogated by the inactivation of JNK signaling. The let-7c over-expression and the addition of its mimics facilitated the activation of AP-1, STAT1 and Bim by linking JNK1/2 to AP-1/STAT1, but rather inhibited the activation of STAT3 and Bcl-xL by connecting JNK1/2 to STAT3, followed by the augmented apoptosis in the cells. The let-7c deficiency reduced the AP-1, STAT1 and Bim activities, and enhanced the STAT3 and Bcl-xL, alleviating the anisomycin-induced apoptosis. The knockdown of the bim gene repressed the anisomycin-boosted apoptosis through the attenuation of the active Bak and Bax. The findings indicate for the first time that miR let-7c is essential for the anisomycin-triggered apoptosis by linking JNK1/2 to AP-1/STAT1/STAT3/Bim/Bcl-xL/Bax/Bak signaling. This provides a novel insight into the mechanism by which anisomycin leads to the tumor cell apoptosis, potentially laying the foundations for its development and clinical application. PMID:27087117

  14. Resveratrol-3-O-glucuronide and resveratrol-4’-O-glucuronide reduce DNA strand breakage but not apoptosis in Jurkat T cells treated with camptothecin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Resveratrol has been reported to inhibit or induce DNA damage depending upon the type of cell and experimental conditions. Dietary resveratrol is present in the body mostly as metabolites and little is known about the activities of these metabolic products. We evaluated physiologically obtainable ...

  15. Ethanol promotes T cell apoptosis through the mitochondrial pathway

    PubMed Central

    Kapasi, Aditi A; Patel, Geeta; Goenka, Anuj; Nahar, Nilay; Modi, Neeraj; Bhaskaran, Madhu; Reddy, Krishna; Franki, Nicholas; Patel, Jaimita; Singhal, Pravin C

    2003-01-01

    Clinical reports suggest that acute ethanol intoxication is often associated with lymphopenia. Previously, ethanol was reported to invoke thymocyte apoptosis. We studied the effect of ethanol on T cell apoptosis. In addition, we evaluated the molecular mechanism of ethanol-induced T cell apoptosis. Human T cells harvested from healthy subjects after an alcohol drinking binge showed enhanced T cell apoptosis (before, 0·4 ± 0·2% versus after, 19·6 ± 2·5% apoptotic lymphocytes/field; P < 0·001). In in vitro studies, ethanol in a concentration of 50 mm and higher enhanced the apoptosis of Jurkat cells. DNA isolated from ethanol-treated Jurkat cells displayed integer multiples of 180 base pairs. Ethanol decreased Jurkat cell expression of Bcl-2, whereas ethanol increased Jurkat cell expression of Bax. Jurkat cells treated with ethanol also showed translocation of cytochrome C into cytosol. Moreover, a caspase-9 inhibitor partially inhibited ethanol-induced Jurkat cell apoptosis. In in vivo studies, after binge drinking, T cell expression of Bcl-2 also decreased. In addition, binge drinking induced the cleavage of caspase-3, suggesting activation of caspase-3 in T cells. These results suggest that ethanol promotes T cell apoptosis through the activation of intrinsic or mitochondrial pathway. PMID:12603597

  16. Shutdown of HIV-1 Transcription in T Cells by Nullbasic, a Mutant Tat Protein

    PubMed Central

    Jin, Hongping; Li, Dongsheng; Sivakumaran, Haran; Lor, Mary; Rustanti, Lina; Cloonan, Nicole; Wani, Shivangi

    2016-01-01

    ABSTRACT Nullbasic is a derivative of the HIV-1 transactivator of transcription (Tat) protein that strongly inhibits HIV-1 replication in lymphocytes. Here we show that lentiviral vectors that constitutively express a Nullbasic-ZsGreen1 (NB-ZSG1) fusion protein by the eEF1α promoter led to robust long-term inhibition of HIV-1 replication in Jurkat cells. Although Jurkat-NB-ZSG1 cells were infected by HIV-1, no virus production could be detected and addition of phorbol ester 12-myristate 13-acetate (PMA) and JQ1 had no effect, while suberanilohydroxamic acid (SAHA) modestly stimulated virus production but at levels 300-fold lower than those seen in HIV-1-infected Jurkat-ZSG1 cells. Virus replication was not recovered by coculture of HIV-1-infected Jurkat-NB-ZSG1 cells with uninfected Jurkat cells. Latently infected Jurkat latent 6.3 and ACH2 cells treated with latency-reversing agents produced measurable viral capsid (CA), but little or none was made when they expressed NB-ZSG1. When Jurkat cells chronically infected with HIV-1 were transduced with lentiviral virus-like particles conveying NB-ZSG1, a >3-log reduction in CA production was observed. Addition of PMA increased virus CA production but at levels 500-fold lower than those seen in nontransduced Jurkat cells. Transcriptome sequencing analysis confirmed that HIV-1 mRNA was strongly inhibited by NB-ZSG1 but indicated that full-length viral mRNA was made. Analysis of HIV-1-infected Jurkat cells expressing NB-ZSG1 by chromatin immunoprecipitation assays indicated that recruitment of RNA polymerase II (RNAPII) and histone 3 lysine 9 acetylation were inhibited. The reduction of HIV-1 promoter-associated RNAPII and epigenetic changes in viral nucleosomes indicate that Nullbasic can inhibit HIV-1 replication by enforcing viral silencing in cells. PMID:27381288

  17. Spaceflight alters microtubules and increases apoptosis in human lymphocytes (Jurkat)

    NASA Technical Reports Server (NTRS)

    Lewis, M. L.; Reynolds, J. L.; Cubano, L. A.; Hatton, J. P.; Lawless, B. D.; Piepmeier, E. H.

    1998-01-01

    Alteration in cytoskeletal organization appears to underlie mechanisms of gravity sensitivity in space-flown cells. Human T lymphoblastoid cells (Jurkat) were flown on the Space Shuttle to test the hypothesis that growth responsiveness is associated with microtubule anomalies and mediated by apoptosis. Cell growth was stimulated in microgravity by increasing serum concentration. After 4 and 48 h, cells filtered from medium were fixed with formalin. Post-flight, confocal microscopy revealed diffuse, shortened microtubules extending from poorly defined microtubule organizing centers (MTOCs). In comparable ground controls, discrete microtubule filaments radiated from organized MTOCs and branched toward the cell membrane. At 4 h, 30% of flown, compared to 17% of ground, cells showed DNA condensation characteristic of apoptosis. Time-dependent increase of the apoptosis-associated Fas/ APO-1 protein in static flown, but not the in-flight 1 g centrifuged or ground controls, confirmed microgravity-associated apoptosis. By 48 h, ground cultures had increased by 40%. Flown populations did not increase, though some cells were cycling and actively metabolizing glucose. We conclude that cytoskeletal alteration, growth retardation, and metabolic changes in space-flown lymphocytes are concomitant with increased apoptosis and time-dependent elevation of Fas/APO-1 protein. We suggest that reduced growth response in lymphocytes during spaceflight is linked to apoptosis.

  18. Protein kinase C inhibits the transplasma membrane influx of Ca2+ triggered by 4-aminopyridine in Jurkat T lymphocytes.

    PubMed

    Barbar, Elie; Rola-Pleszczynski, Marek; Payet, Marcel D; Dupuis, Gilles

    2003-07-23

    4-aminopyridine (4AP) is a general blocker of voltage-dependent K+ channels. This pyridine derivative has also been shown to inhibit T cell proliferation, to modulate immune responses and to alleviate some of the symptoms associated with neurological disorders such as multiple sclerosis, myasthenia gravis and Alzheimer's disease. 4AP triggers a Ca2+ response in lymphocytes, astrocytes, neurons and muscle cells but little is known about the regulation of the 4AP response in these cells. We report that 4AP induced a non-capacitative transplasma membrane influx of Ca2+ in Jurkat T lymphocytes. The influx of Ca2+ was not affected by activation or inhibition of protein kinase A (PKA). In contrast, activation of protein kinase C (PKC) by phorbol myristyl acetate (PMA), mezerein or 1-oleoyl-2-acetyl-sn-glycerol (OAG) inhibited the influx of Ca2+ triggered by 4AP. The inhibitory effect of PKC could be prevented by prior exposure of the cells to the PKC inhibitor GF 109203X. Under these conditions, mezerein and OAG no longer inhibited the 4AP-dependent Ca2+ response. Inhibition of serine and threonine protein phosphatases PP1 and PP2A by treating the cells with calyculin A (CalA) reduced the Ca2+ response to 4AP. Okadaic acid (OA) had no effect, suggesting an involvement of PP1. A combination of CalA and OAG (or PMA) abolished the influx of Ca2+ induced by 4AP, adding further evidence to the importance of protein phosphorylation in the modulation of the 4AP response. Our data suggest that the transplasma membrane influx of Ca2+ triggered by 4AP in Jurkat T cells can be modulated by the opposite actions of PKC and protein serine and threonine phosphatase(s). PMID:12880946

  19. In vitro evaluation of the effects of perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) on IL-2 production in human T-cells

    PubMed Central

    Midgett, Kristin; Peden-Adams, Margie M.; Gilkeson, Gary S.; Kamen, Diane L.

    2014-01-01

    Perfluorinated compounds, such as perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA), have been shown to alter various immune functions suggesting they are immunotoxic. This study assessed the effects of PFOS and PFOA on interleukin (IL)-2 production in the human Jurkat T-cell line and PFOS in healthy human primary T cells. Jurkat cells were stimulated with phytohemagglutinin (PHA)/phorbol myristate acetate (PMA), anti CD-3/anti CD-28, or anti CD-3, and dosed with 0, 0.05, 0.1, 0.5, 1, 5, 10, 50, 75, or 100 μg ml−1 PFOS or 0, 0.005, 0.01, 0.05, 0.1, 0.5, 1, 5, or 10 μg ml−1 PFOA. Jurkat cells stimulated with PHA/PMA or anti CD-3 exhibited decreased IL-2 production beginning at 50 μg PFOS ml−1 and 5 μg PFOS ml−1 respectively, but stimulation with anti-CD3/anti-CD28 resulted in no changes compared with the control. Addition of the PPAR-alpha antagonist GW6471 to PFOS-dosed cells stimulated with PHA/PMA resulted in decreases in IL-2 production starting at 50 μg PFOS ml−1, which suggests PFOS affected T-cell IL-2 production via PPAR-alpha-independent mechanisms. Exposure to PFOA, PFOA + GW6471, or PFOS + PFOA in Jurkat cells resulted in no significant differences in IL-2 production. In vitro dosing studies using healthy primary human CD4+ T cells were consistent with the Jurkat results. These data demonstrated that PFOA did not impact IL-2 production, but PFOS suppressed IL-2 production in both a human cell line and human primary cells at dose levels within the high end of the human exposure range. A decrease in IL-2 production is characteristic of autoimmune diseases such as systemic lupus erythematosus and should be further investigated. PMID:25056757

  20. The Effects of Intense Submicrosecond Electrical Pulses on Cells

    PubMed Central

    Deng, Jingdong; Schoenbach, Karl H.; Buescher, E. Stephen; Hair, Pamela S.; Fox, Paula M.; Beebe, Stephen J.

    2003-01-01

    A simple electrical model for living cells predicts an increasing probability for electric field interactions with intracellular substructures of both prokaryotic and eukaryotic cells when the electric pulse duration is reduced into the sub-microsecond range. The validity of this hypothesis was verified experimentally by applying electrical pulses (durations 100 μs–60 ns, electric field intensities 3–150 kV/cm) to Jurkat cells suspended in physiologic buffer containing propidium iodide. Effects on Jurkat cells were assessed by means of temporally resolved fluorescence and light microscopy. For the longest applied pulses, immediate uptake of propidium iodide occurred consistent with electroporation as the cause of increased surface membrane permeability. For nanosecond pulses, more delayed propidium iodide uptake occurred with significantly later uptake of propidium iodide occurring after 60 ns pulses compared to 300 ns pulses. Cellular swelling occurred rapidly following 300 ns pulses, but was minimal following 60 ns pulses. These data indicate that submicrosecond pulses achieve temporally distinct effects on living cells compared to microsecond pulses. The longer pulses result in rapid permeability changes in the surface membrane that are relatively homogeneous across the cell population, consistent with electroporation, while shorter pulses cause surface membrane permeability changes that are temporally delayed and heterogeneous in their magnitude. PMID:12668479

  1. Cytosine arabinoside potentiates the apoptotic effect of bendamustine on several B- and T-cell leukemia/lymphoma cells and cell lines.

    PubMed

    Castegnaro, Silvia; Visco, Carlo; Chieregato, Katia; Bernardi, Martina; Albiero, Elena; Zanon, Cristina; Madeo, Domenico; Rodeghiero, Francesco

    2012-11-01

    Bendamustine and cytosine arabinoside (ara-c) are commonly used cytotoxic agents with unique mechanisms of action. We have previously reported a striking additive cytotoxic effect of the consecutive combination of bendamustine and ara-c in mantle cell lymphoma (MCL) cell lines. In the present study, cell lines of follicular lymphoma (DOHH-2), chronic lymphocytic leukemia/lymphoma (EHEB), diffuse large B-cell lymphoma (SU-DHL-4), T-cell leukemia/lymphoma (JURKAT and KARPAS-299) and MCL (JEKO-1 and GRANTA-519) were exposed to the two single drugs or the drugs combined, given simultaneously and consecutively. Peripheral blood chronic lymphocytic leukemia (CLL) B-cells from five patients were also analyzed. Apoptosis, cell proliferation/metabolic activity and mitochondrial damage were evaluated. The combination index (CI) was used to assess synergy between the drugs. Bendamustine exhibited a relevant cytotoxic effect that was dose- and time-dependent, except for SU-DHL-4 and T-cell lymphoma cells. The addition of ara-c after bendamustine significantly potentiated the single-drug cytotoxic effect of bendamustine on all cell lines, including 17p - CLL B-cells, JURKAT and SU-DHL-4, the latter presenting the highest synergism (CI < 0.01). Bendamustine and ara-c are highly synergistic on T- and B-cell lymphoma cells and cell lines, similar to MCL, overcoming resistance to the single agents. PMID:22530665

  2. Anti-tumoral effect of arsenic compound, sodium metaarsenite (KML001), in non-Hodgkin's lymphoma: an in vitro and in vivo study.

    PubMed

    Yoon, Jin Sun; Hwang, Deok Won; Kim, Eun Shil; Kim, Jung Soon; Kim, Sujong; Chung, Hwa Jin; Lee, Sang Kook; Yi, Jun Ho; Uhm, Jieun; Won, Young Woong; Park, Byeong Bae; Choi, Jung Hye; Lee, Young Yiul

    2016-02-01

    Arsenic compounds have been used in traditional medicine for several centuries. KML001 (sodium metaarsenite; NaAsO2) is an orally bio-available arsenic compound with potential anti-cancer activity. However, the effect of KML001 has not been studied in lymphoid neoplasms. The aim of this study is to evaluate the anti-proliferative effect of KML001 in non-Hodgkin's lymphoma and to compare its efficacy with As2O3. KML001 inhibited cellular proliferation in all tested lymphoma cell lines as well as JurkatR cells (adriamycin-resistant Jurkat cells) in a dose-dependent manner, while As2O3 was not effective. Cell cycle regulatory protein studies have suggested that KML001 induces G1 arrest via p27-induced inhibition of the kinase activities of CDK2, 4, and 6. Treatment of KML001 induced apoptosis in Jurkat and JurkatR cells. The apoptotic process was associated with down-regulation of Bcl-2 (antiapoptotic molecule), up-regulation of Bax (proapoptotic molecule), and inhibition of caspase-3, -8, and -9. In addition, cell signaling including the STAT, PI3K/Akt, MAPK, and NF-κB signal pathways were inhibited in KML001-treated Jurkat and JurkatR cells. Furthermore, targeting the telomere by KML001 was observed in the Jurkat and JurkatR cells. The In vivo anti-tumoral activity of KML001 was confirmed in a xenograft murine model. Interestingly, partial responses were seen in two lymphoma patients treated with 10 mg/day (follicular lymphoma for 16 weeks and mantle cell lymphoma for 24 weeks) without severe toxicities. These findings suggest that KML001 may be a candidate agent for the treatment of de novo, refractory, and relapsed non-Hodgkin's lymphoma patients. PMID:26581399

  3. Cholesteryl butyrate solid lipid nanoparticles as a butyric acid pro-drug: effects on cell proliferation, cell-cycle distribution and c-myc expression in human leukemic cells.

    PubMed

    Serpe, Loredana; Laurora, Stefano; Pizzimenti, Stefania; Ugazio, Elena; Ponti, Renata; Canaparo, Roberto; Briatore, Federica; Barrera, Giuseppina; Gasco, Maria Rosa; Bernengo, Maria Grazia; Eandi, Mario; Zara, Gian Paolo

    2004-06-01

    Cholesteryl butyrate solid lipid nanoparticles (chol-but SLN) have been proposed as a pro-drug to deliver butyric acid. We compared the effects on cell growth, cell-cycle distribution and c-myc expression of chol-but SLN and sodium butyrate (Na-but) in the human leukemic cell lines Jurkat, U937 and HL-60. In all the cell lines 0.5 and 1.0 mM chol-but SLN provoked a complete block of cell growth. Cell-cycle analysis demonstrated in Jurkat cells that 0.25 mM chol-but SLN caused a pronounced increase of G2/M cells and a decrease of G0/G1 cells, whereas in U937 and HL-60 cells chol-but SLN led to a dose-dependent increase of G0/G1 cells, with a decrease of G2/M cells. In Jurkat and HL-60 cells 0.5 mM chol-but SLN induced a significant increase of sub-G0/G1 apoptotic cells. Cell growth and cell-cycle distribution were unaffected by the same concentrations of Na-but. A concentration of 0.25 mM chol-but SLN was able to cause a rapid and transient down-regulation of c-myc expression in all the cell lines, whereas 1 mM Na-but caused a slight reduction of c-myc expression only in U937 cells. The results show how chol-but SLN affects the proliferation pattern of both myeloid and lymphoid cells to an extent greater than the natural butyrate. PMID:15166628

  4. Effect of selected flavones on cancer and endothelial cells.

    PubMed

    Pilátová, Martina; Stupáková, Viktória; Varinská, Lenka; Sarisský, Marek; Mirossay, Ladislav; Mirossay, Andrej; Gál, Peter; Kraus, Vladimír; Dianisková, Katarína; Mojzis, Ján

    2010-06-01

    In our study we used quercetin (3,3 ,4 ,5,7-pentahydroxyflavone) as the reference standard to compare antiproliferative and antiangiogenic effects of chrysin (5,7-dihydroxyflavone) and 3-hydroxyflavone. Our data indicates that chrysin and 3-hydroxyflavone showed significantly higher cytotoxic effect than reference standard quercetin. These tested agents significantly decreased cell survival with the efficacy of 65-85% at the concentration 100 micromol/l for HUVEC, lung carcinoma and leukemic cells being the most sensitive. Cell cycle analysis indicates that quercetin and 3-hydroxyflavone might affect the cell cycle of Jurkat cells by a similar or the same mechanism of action which lead to G2/M arrest as well as to an increase in sub-G0/G1 fraction. Treatment of Jurkat cells with chrysin resulted only increase in the fraction of cells with sub-G0/G1 DNA content, which is considered to be a marker of apoptotic cell death. Apoptosis was confirmed by DNA fragmentation and by staining with annexin V. All three tested flavones inhibited endothelial cell migration after 24 h of incubation at a concentration 100 micromol/l. At a lower concentration (10 micromol/l) only quercetin significantly inhibited migration of endothelial cells. Furthermore, in our experiments decreased secretion of matrix metalloproteinases (MMP-2 and MMP-9) was observed after a 72 h treatment with quercetin. No decrease in secretion of MMP-2 and MMP-9 was seen after chrysin and 3-hydroxyflavone treatment. On the other hand, our results showed that none of three flavonoids blocked microcapillary tube formation. Further studies are necessary to investigate the mechanism of action and to find out the relationship between the structure, character and position of substituents of natural substances and their biological activities. PMID:20577025

  5. Inhibition of Cancer Cell Proliferation and Antiradical Effects of Decoction, Hydroalcoholic Extract, and Principal Constituents of Hemidesmus indicus R. Br.

    PubMed

    Statti, Giancarlo; Marrelli, Mariangela; Conforti, Filomena; Spagnoletti, Antonella; Tacchini, Massimo; Fimognari, Carmela; Brognara, Eleonora; Gambari, Roberto; Sacchetti, Gianni; Guerrini, Alessandra

    2015-06-01

    Indian Sarsaparilla (Hemidesmus indicus R. Br.) is widely used in Indian traditional medicine. In the present work, we explored the effects of decoction, traditional Ayurvedic preparation, and hydroalcoholic extract, a phytocomplex more traditionally studied and commercialized as food supplement in western medicine, from the roots as possible source of chemicals with new functional potential linked to their nutritional uses. The antiproliferative and antioxidant properties were assayed. To test antiproliferative affects, different cancer cell lines, growing both as monolayers (CaCo2, MCF-7, A549, K562, MDA-MB-231, Jurkat, HepG2, and LoVo) and in suspension (K562 and Jurkat) were used. The decoction showed strong activity on HepG2 cells, while the hydroalcoholic extracts were active on HepG2, LoVo, MCF-7, K562, and Jurkat cell lines. Weak inhibition of cancer cell proliferation was observed for the principal constituents of the preparations: 2-hydroxy-4-methoxybenzaldehyde, 2-hydroxy-4-methoxybenzoic acid, and 3-hydroxy-4-methoxybenzaldehyde that were tested alone. The antiradical activity was tested with 2,2-diphenyl-1-picrylhydrazyl and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt tests and inhibition of nitric oxide production in lipopolysaccharide-stimulated RAW 264.7 macrophages. Interesting result has also been obtained for hydroalcoholic extract regarding genoprotective potential (58.79% of inhibition at 37.5 µg/mL). PMID:25753739

  6. In Vitro Cytotoxic Potential of Essential Oils of Eucalyptus benthamii and Its Related Terpenes on Tumor Cell Lines

    PubMed Central

    Döll-Boscardin, Patrícia Mathias; Sartoratto, Adilson; Sales Maia, Beatriz Helena Lameiro de Noronha; Padilha de Paula, Josiane; Nakashima, Tomoe; Farago, Paulo Vitor; Kanunfre, Carla Cristine

    2012-01-01

    Eucalyptus L. is traditionally used for many medicinal purposes. In particular, some Eucalyptus species have currently shown cytotoxic properties. Local Brazilian communities have used leaves of E. benthamii as a herbal remedy for various diseases, including cancer. Considering the lack of available data for supporting this cytotoxic effect, the goal of this paper was to study the in vitro cytotoxic potential of the essential oils from young and adult leaves of E. benthamii and some related terpenes (α-pinene, terpinen-4-ol, and γ-terpinene) on Jurkat, J774A.1 and HeLa cells lines. Regarding the cytotoxic activity based on MTT assay, the essential oils showed improved results than α-pinene and γ-terpinene, particularly for Jurkat and HeLa cell lines. Terpinen-4-ol revealed a cytotoxic effect against Jurkat cells similar to that observed for volatile oils. The results of LDH activity indicated that cytotoxic activity of samples against Jurkat cells probably involved cell death by apoptosis. The decrease of cell DNA content was demonstrated due to inhibition of Jurkat cells proliferation by samples as a result of cytotoxicity. In general, the essential oils from young and adult leaves of E. benthamii presented cytotoxicity against the investigated tumor cell lines which confirms their antitumor potential. PMID:22645627

  7. Immune Suppressive Effect of Cinnamaldehyde Due to Inhibition of Proliferation and Induction of Apoptosis in Immune Cells: Implications in Cancer

    PubMed Central

    Gomez-Casado, Cristina; Diaz-Perales, Araceli; Oida, Kumiko; Singer, Josef; Kinaciyan, Tamar; Fuchs, Heidemarie C.; Jensen-Jarolim, Erika

    2014-01-01

    Background Besides its anti-inflammatory effects, cinnamaldehyde has been reported to have anti-carcinogenic activity. Here, we investigated its impact on immune cells. Methods Activation of nuclear factor-κB by cinnamaldehyde (0–10 µg/ml) alone or in combination with lipopolysaccharide was assessed in THP1XBlue human monocytic cell line and in human peripheral blood mononuclear cells (PBMCs). Proliferation and secretion of cytokines (IL10 and TNFα) was determined in primary immune cells and the human cell lines (THP1, Jurkat E6-1 and Raji cell lines) stimulated with cinnamaldehyde alone or in conjunction with lipopolysaccharide. Nitric oxide was determined in mouse RAW264.7 cells. Moreover, different treated PBMCs were stained for CD3, CD20 and AnnexinV. Results Low concentrations (up to 1 µg/ml) of cinnamaldehyde resulted in a slight increase in nuclar factor-kB activation, whereas higher concentrations led to a dose-dependent decrease of nuclear factor-kB activation (up to 50%) in lipopolysachharide-stimulated THP1 cells and PBMCs. Accordingly, nitric oxide, interleukin 10 secretion as well as cell proliferation were reduced in lipopolysachharide-stimulated RAW264.7 cells, PBMCs and THP1, Raji and Jurkat-E6 immune cells in the presence of cinnamaldehyde in a concentration-dependent manner. Flow cytometric analysis of PBMCs revealed that CD3+ were more affected than CD20+ cells to apopotosis by cinnamaldehyde. Conclusion We attribute the anti-inflammatory properties of cinnamaldehyde to its ability to block nuclear factor-κB activation in immune cells. Treatment with cinnamaldehyde led to inhibition of cell viability, proliferation and induced apoptosis in a dose-dependent manner in primary and immortalized immune cells. Therefore, despite its described anti-carcinogenic property, treatment with cinnamaldehyde in cancer patients might be contraindicated due to its ability to inhibit immune cell activation. PMID:25271635

  8. Interactions between endothelial cells and T cells modulate responses to mixed neutron/gamma radiation.

    PubMed

    Cary, Lynnette H; Noutai, Daniel; Salber, Rudolph E; Williams, Margaret S; Ngudiankama, Barbara F; Whitnall, Mark H

    2014-06-01

    Detonation of an improvised nuclear device near a population center would cause significant casualties from the acute radiation syndrome (ARS) due to exposure to mixed neutron/gamma fields (MF). The pathophysiology of ARS involves inflammation, microvascular damage and alterations in immune function. Interactions between endothelial cells (EC) and hematopoietic cells are important not only for regulating immune cell traffic and function, but also for providing the microenvironment that controls survival, differentiation and migration of hematopoietic stem and progenitor cells in blood-forming tissues. Endothelial cells/leukocyte interactions also influence tumor progression and the results of anticancer therapies. In this study, we hypothesized that irradiation of endothelial cells would modulate their effects on hematopoietic cells and vice versa. Human umbilical vein endothelial cells (HUVEC) and immortalized T lymphocytes (Jurkat cells) were cultured individually and in co-culture after exposure to mixed fields. Effects of nonirradiated cells were compared to effects of irradiated cells and alterations in signaling pathways were determined. Mitogen-activated protein kinases (MAPKs) p38 and p44/42 (ERK1/2) in HUVEC exhibited higher levels of phosphorylated protein after exposure to mixed field radiation. IL-6, IL-8, G-CSF, platelet derived growth factor (PDGF) and angiopoietin 2 (ANG2) protein expression were upregulated in HUVEC by exposure to mixed field radiation. PCR arrays using HUVEC mRNA revealed alterations in gene expression after exposure to mixed fields and/or co-culture with Jurkat cells. The presence of HUVEC also influenced the function of Jurkat cells. Nonirradiated Jurkat cells showed an increase in proliferation when co-cultured with nonirradiated HUVEC, and a decrease in proliferation when co-cultured with irradiated HUVEC. Additionally, nonirradiated Jurkat cells incubated in media from irradiated HUVEC exhibited upregulation of activated

  9. Effect of extremely low frequency magnetic fields on cell proliferation and gene expression.

    PubMed

    Lee, Hyung Chul; Hong, Mi-Na; Jung, Seung Hee; Kim, Bong Cho; Suh, Young Ju; Ko, Young-Gyu; Lee, Yun-Sil; Lee, Byeong-Yoon; Cho, Yeun-Gyu; Myung, Sung-Ho; Lee, Jae-Seon

    2015-10-01

    Owing to concerns regarding possible effects of extremely low frequency magnetic fields (ELF-MF) on human health, many studies have been conducted to elucidate whether ELF-MF can induce modifications in biological processes. Despite this, controversies regarding effects of ELF-MF are still rife. In this study, we investigated biological effects of ELF-MF on MCF10A, MCF7, Jurkat, and NIH3T3 cell lines. ELF-MF with a magnetic flux density of 1 mT at 60 Hz was employed to stimulate cells for 4 or 16 h, after which the effects of ELF-MF on cell proliferation, cell death, cell viability, and DNA synthesis rates were assessed. Whereas Jurkat and NIH3T3 cells showed no consistent variation in cell number, cell viability, and DNA synthesis rate, MCF10A and MCF7 cells showed consistent and significant decreases in cell number, cell viability, and DNA synthesis rates. However, there was no effect of ELF-MF on cell death in any of tested cell lines. Next, to investigate the effect of ELF-MF on gene expression, we exposed MCF7 cells to 2 mT at 60 Hz for 16 h and examined transcriptional responses by using gene expression array. We found a gene, PMAIP1, that exhibited statistically significant variation using two-fold cut-off criteria and certified its expression change by using semi-quantitative and quantitative reverse transcription polymerase chain reaction. From these results, we concluded that ELF-MF could induce the delay of cell cycle progression in MCF7 and MCF10A cells in a cell context-specific manner and could up-regulate PMAIP1 in MCF7 cells. PMID:26239017

  10. Proteomic Analysis of Terminalia chebula Extract-Dependent Changes in Human Lymphoblastic T Cell Protein Expression

    PubMed Central

    Das, Nando Dulal; Jung, Kyoung Hwa; Park, Ji Hyun; Choi, Mi Ran; Lee, Hyung Tae; Kim, Moo Sung; Lee, Sang Rin

    2012-01-01

    Abstract Terminalia chebula is a native plant from southern Asia to southwestern China that is used in traditional medicine for the treatment of malignant tumors and diabetes. This plant also has antibacterial and immunomodulatory properties. The present study assessed T. chebula extract-dependent protein expression changes in Jurkat cells. Matrix-assisted laser desorption/ionization–time-of-flight mass spectrometry and Ingenuity Pathways Analysis (IPA) were performed to assess protein expression and networks, respectively. A comparative proteomic profile was determined in T. chebula extract (50 μg/mL)-treated and control cells; the expressions of β-tubulin, ring finger and CHY zinc finger domain containing 1, and insulin-like growth factor 1 receptor kinase were significantly down-regulated in T. chebula extract-treated Jurkat cells. Moreover, the molecular basis for the T. chebula extract-dependent protein expression changes in Jurkat cells was determined by IPA. Treatment with the T. chebula extract significantly inhibited nuclear factor-κB activity and affected the proteomic profile of Jurkat cells. The molecular network signatures and functional proteomics obtained in this study may facilitate the evaluation of potential antitumor therapeutic targets and elucidate the molecular mechanism of T. chebula extract-dependent effects in Jurkat cells. PMID:22471968

  11. Reversal effect of vitamin D on different multidrug-resistant cells.

    PubMed

    Yan, M; Nuriding, H

    2014-01-01

    We investigated the reversal effect of vitamin D on the multidrug-resistant leukemic Jurkat/ADR and K562/ADR cell lines and conducted a preliminary investigation of its reversal mechanism. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method was used to detect the reversal effect of vitamin D on multidrug-resistant cells. Real-time polymerase chain reaction was used to determine the effect of vitamin D on intracellular expression of mRNA of the multidrug-resistant gene (MDRI) and the multidrug-resistance-related gene (MRP1). A protein quantitative analysis method was used to determine the effect of vitamin D on intracellular glutathione content. After treatment of Jurkat/ADR and K562/ADR cells with vitamin D, multidrug resistance was reversed in a dose-dependent manner, which may have reduced mRNA expression of the MDR1 and MRP1 genes, the P-glycoprotein content on the cell surface, and the intracellular glutathione level. Different concentrations of vitamin D showed varying reversal effects on different multidrug-resistant cells. The resistance mechanism may be related to the inhibition of the expression of MDR1 and MRP1 genes. PMID:25158250

  12. In Vitro Effects of Propranolol on T Helper Type 1 Cytokine Profile in Human Leukemic T Cells

    PubMed Central

    Hajighasemi, Fatemeh; Mirshafiey, Abbas

    2016-01-01

    Introduction: Cytokines are a large group of proteins play a key role in inflammation. Down-regulation of pro-inflammatory cytokines has beneficial effect on heart function. Propranolol, as a non selective beta-adrenergic blocker, has been extensively used for treatment of many cardiovascular problems such as arrhythmias and heart malfunction. In addition anti-inflammatory effects of propranolol have been revealed. In this study the propranolol effect on T helper type 1 cytokine profile in human leukemic T cells has been assessed in vitro. Materials and methods: Human leukemic T cells (Molt-4 and Jurkat) were cultured in complete RPMI medium. The cells were then incubated with different concentrations of propranolol (0.03- 30 µM) in the presence or absence of PHA (10 µg/ml) for 48 hours. The supernatants of cell culture media were collected and used for cytokines assay. Results: Propranolol significantly decreased the T helper type 1 cytokine profile [Interleukin-2 (IL-2) and Interferon- γ (IFN-γ)] production in PHA stimulated Molt-4 and Jurkat cells, after 48 hour of incubation time, dose-dependently compared to untreated control cells. Conclusion: Our data showed a dose dependent inhibitory effect of propranolol on the IL-2 and IFN-γ production in human leukemic Molt-4 and Jurkat cells. The anti- inflammatory effect of propranolol reported by other investigators may be in part due to its suppressive effect on production of inflammatory cytokines such as IL-2 and IFN-γ. So, propranolol along with its chronic long-term usage in cardiovascular problems may have potential implication in treatment of inflammatory-based disorders. PMID:27252810

  13. The effect of rotation on function and signal transduction in immune cell

    NASA Astrophysics Data System (ADS)

    Song, J. P.; Zhong, P.; Li, Y. H.; Yang, F.

    Objective Both spaceflight and modeled weightlessness on ground could compromise immune function especially cellular immunity In turn astrouants would not resist to external pathogen effectually the health status and work ability of astrounants were perhaps affected but the cellular and molecular mechanisms by which spaceflight alters human immune functions are poorly understood The aim this trial was to using high aspect rotation vessal HARV investigate the functional changes of immune cell rotated for virous time period in vitro and explore mechanisms in which space weightlessness affect immune function through cell signal transduction Methods Using high aspect rotation vessal HARV as simulated weightlessness model mouse splenic lymphocyte and Jurkat E6 1 as cell model the effects of rotation on cell proliferation cytokine secretion expression and activation of signal molecule ZAP-70 were studied Results After rotation T lymphocytic proliferation in mouse splenocyte were inhibited and the concentration of IL-2 and IFN- A secreted were reduced markly and all this happen within 6 hours after T cell were activated The activity of ZAP-70 in Jurkat cell were repressed significantly Conclusion Incapable activation of ZAP-70 might be one cause of depressed lymphocyte function under weightlessness

  14. Pyrvinium selectively induces apoptosis of lymphoma cells through impairing mitochondrial functions and JAK2/STAT5.

    PubMed

    Xiao, Meifang; Zhang, Liming; Zhou, Yizheng; Rajoria, Pasupati; Wang, Changfu

    2016-01-15

    Targeting mitochondrial respiration has emerged as an attractive therapeutic strategy in blood cancer due to their unique metabolic dependencies. In this study, we show that pyrvinium, a FDA-approved anthelmintic drug, selectively targets lymphoma T-cells though inhibition of mitochondrial functions and JAK2/STAT5. Pyrvinium induces apoptosis of malignant T-cell line Jurkat and primary T-cells from lymphoma patients while sparing T-cells from healthy donors. Increased level of active caspase-3 and decreased levels of Bcl-2 and Mcl-1 were also observed in Jurkat and lymphoma T-cells but not normal T-cells treated with pyrvinium. In addition, pyrvinium impairs mitochondrial functions by inhibit mitochondrial respiration, suppressing mitochondrial respiratory complex I activity, increasing ROS and decreasing ATP levels. However, the effects of pyrvinium were abolished in mitochondrial respiration-deficient Jurkat ρ(0) cells, confirming that pyrvinium acts on lymphoma T-cells via targeting mitochondrial respiration. We further show that lymphoma T-cells derived from patients depend more on mitochondrial respiration than normal T-cells, and this explains the selective toxicity of pyrvinium in lymphoma versus normal T-cells. Finally, we demonstrate that pyrvinium also suppresses JAK2/STAT5 signaling pathway in Jurkat cells. Our study suggests that pyrvinium is a useful addition to T-cell lymphoma treatment, and emphasizes the potential therapeutic value of the differences in the mitochondrial characteristics between malignant and normal T-cells in blood cancer. PMID:26707639

  15. Oxidative phenomena are implicated in human T-cell stimulation.

    PubMed Central

    Sekkat, C; Dornand, J; Gerber, M

    1988-01-01

    Phytohaemagglutinin (PHA), phorbol myristate acetate (PMA) and PHA + PMA stimulation of T-enriched peripheral blood lymphocytes (PBL) and the Jurkat malignant T-cell line leads to oxidative-product formation, as evaluated by flow cytofluorometric studies, an increase in K+ flux across the membrane, cGMP production and a depolarization of the cell membrane. Irradiation (20 Gy), which enhances IL-2 synthesis by activated T-enriched PBL and Jurkat cells, also increases oxidative product formation, K+ flux, cGMP production, and induces cell membrane depolarization. Conversely, irradiation does not produce a rise in intracellular free Ca2+, as measured in PHA-stimulated Jurkat cells. PMA is also without effect on intracellular free Ca2+, added before or after PHA stimulation. Thus, except for the rise in intracellular free Ca2+, irradiation and stimulation exert similar effects on some of the events observed in IL-2-producing Jurkat cells, but these effects are not additive. Stimulation and irradiation effects are shown to be additive or synergistic only for cGMP production. It is proposed that irradiation may increase IL-2 synthesis by participating in an additional signal related to the oxidative metabolism of arachidonic acid (AA). PMID:3258279

  16. Oxidative phenomena are implicated in human T-cell stimulation.

    PubMed

    Sekkat, C; Dornand, J; Gerber, M

    1988-03-01

    Phytohaemagglutinin (PHA), phorbol myristate acetate (PMA) and PHA + PMA stimulation of T-enriched peripheral blood lymphocytes (PBL) and the Jurkat malignant T-cell line leads to oxidative-product formation, as evaluated by flow cytofluorometric studies, an increase in K+ flux across the membrane, cGMP production and a depolarization of the cell membrane. Irradiation (20 Gy), which enhances IL-2 synthesis by activated T-enriched PBL and Jurkat cells, also increases oxidative product formation, K+ flux, cGMP production, and induces cell membrane depolarization. Conversely, irradiation does not produce a rise in intracellular free Ca2+, as measured in PHA-stimulated Jurkat cells. PMA is also without effect on intracellular free Ca2+, added before or after PHA stimulation. Thus, except for the rise in intracellular free Ca2+, irradiation and stimulation exert similar effects on some of the events observed in IL-2-producing Jurkat cells, but these effects are not additive. Stimulation and irradiation effects are shown to be additive or synergistic only for cGMP production. It is proposed that irradiation may increase IL-2 synthesis by participating in an additional signal related to the oxidative metabolism of arachidonic acid (AA). PMID:3258279

  17. Specific inhibition of Wee1 kinase and Rad51 recombinase: A strategy to enhance the sensitivity of leukemic T-cells to ionizing radiation-induced DNA double-strand breaks

    SciTech Connect

    Havelek, Radim; Cmielova, Jana; Kralovec, Karel; Bruckova, Lenka; Bilkova, Zuzana; Fousova, Ivana; Sinkorova, Zuzana; Vavrova, Jirina; Rezacova, Martina

    2014-10-24

    Highlights: • Pre-treatment with the inhibitors increased the sensitivity of Jurkat cells to irradiation. • Combining both inhibitors together resulted in a G2 cell cycle arrest abrogation in Jurkat. • Jurkat cells pre-treated with inhibitors were positive for γH2AX foci 24 h upon irradiation. • Pre-treatment with Rad51 RI-1 had no effect on apoptosis induction in MOLT-4 cells. • When dosed together, the combination decreased MOLT-4 cell survival. - Abstract: Present-day oncology sees at least two-thirds of cancer patients receiving radiation therapy as a part of their anticancer treatment. The objectives of the current study were to investigate the effects of the small molecule inhibitors of Wee1 kinase II (681641) and Rad51 (RI-1) on cell cycle progression, DNA double-strand breaks repair and apoptosis following ionizing radiation exposure in human leukemic T-cells Jurkat and MOLT-4. Pre-treatment with the Wee1 681641 or Rad51 RI-1 inhibitor alone increased the sensitivity of Jurkat cells to irradiation, however combining both inhibitors together resulted in a further enhancement of apoptosis. Jurkat cells pre-treated with inhibitors were positive for γH2AX foci 24 h upon irradiation. MOLT-4 cells were less affected by inhibitors application prior to ionizing radiation exposure. Pre-treatment with Rad51 RI-1 had no effect on apoptosis induction; however Wee1 681641 increased ionizing radiation-induced cell death in MOLT-4 cells.

  18. Mechanisms of Dihydroartemisinin and Dihydroartemisinin/Holotransferrin Cytotoxicity in T-Cell Lymphoma Cells

    PubMed Central

    Zhao, Xindong; Zhao, Chunting; Zhao, Hongguo; Huo, Lanfen

    2015-01-01

    The validated therapeutic effects of dihydroartemisinin (DHA) in solid tumors have encouraged us to explore its potential in treating T-cell lymphoma. We found that Jurkat cells (a T-cell lyphoma cell line) were sensitive to DHA treatment with a IC50 of dihydroartemisinin. The cytotoxic effect of DHA in Jurkat cells showed a dose- and time- dependent manner. Interestingly, the cytotoxic effect of DHA was further enhanced by holotransferrin (HTF) due to the high expression of transferrin receptors in T-cell lymphoma. Mechanistically, DHA significantly increased the production of intracellular reactive oxygen species, which led to cell cycle arrest and apoptosis. The DHA treatment also inhibited the expression of protumorgenic factors including VEGF and telomerase catalytic subunit. Our results have proved the therapeutic effect of DHA in T-cell lymphoma. Especially in combination with HTF, DHA may provide a novel efficient approach in combating the deadly disease. PMID:26502166

  19. Mechanisms of Dihydroartemisinin and Dihydroartemisinin/Holotransferrin Cytotoxicity in T-Cell Lymphoma Cells.

    PubMed

    Wang, Qiuyan; Wu, Shaoling; Zhao, Xindong; Zhao, Chunting; Zhao, Hongguo; Huo, Lanfen

    2015-01-01

    The validated therapeutic effects of dihydroartemisinin (DHA) in solid tumors have encouraged us to explore its potential in treating T-cell lymphoma. We found that Jurkat cells (a T-cell lyphoma cell line) were sensitive to DHA treatment with a IC50 of dihydroartemisinin. The cytotoxic effect of DHA in Jurkat cells showed a dose- and time- dependent manner. Interestingly, the cytotoxic effect of DHA was further enhanced by holotransferrin (HTF) due to the high expression of transferrin receptors in T-cell lymphoma. Mechanistically, DHA significantly increased the production of intracellular reactive oxygen species, which led to cell cycle arrest and apoptosis. The DHA treatment also inhibited the expression of protumorgenic factors including VEGF and telomerase catalytic subunit. Our results have proved the therapeutic effect of DHA in T-cell lymphoma. Especially in combination with HTF, DHA may provide a novel efficient approach in combating the deadly disease. PMID:26502166

  20. The cathepsin B inhibitor, z-FA-CMK is toxic and readily induced cell death in human T lymphocytes

    SciTech Connect

    Liow, K.Y.; Chow, S.C.

    2013-11-01

    The cathepsin B inhibitor, benzyloxycarbonyl-phenylalanine-alanine-chloromethylketone (z-FA-CMK) was found to be toxic and readily induced cell death in the human T cell line, Jurkat, whereas two other analogs benzyloxycarbonyl-phenylalanine-alanine-fluoromethylketone (z-FA-FMK) and benzyloxycarbonyl-phenylalanine-alanine-diazomethylketone (z-FA-DMK) were not toxic. The toxicity of z-FA-CMK requires not only the CMK group, but also the presence of alanine in the P1 position and the benzyloxycarbonyl group at the N-terminal. Dose–response studies showed that lower concentrations of z-FA-CMK induced apoptosis in Jurkat T cells whereas higher concentrations induced necrosis. In z-FA-CMK-induced apoptosis, both initiator caspases (-8 and -9) and effector caspases (-3, -6 and -7) were processed to their respective subunits in Jurkat T cells. However, only the pro-form of the initiator caspases were reduced in z-FA-CMK-induced necrosis and no respective subunits were apparent. The caspase inihibitor benzyloxycarbonyl-valine-alanine-aspartic acid-(O-methyl)-fluoromehylketone (z-VAD-FMK) inhibits apoptosis and caspase processing in Jurkat T cells treated with low concentration of z-FA-CMK but has no effect on z-FA-CMK-induced necrosis and the loss of initiator caspases. This suggests that the loss of initiator caspases in Jurkat T cells during z-FA-CMK-induced necrosis is not a caspase-dependent process. Taken together, we have demonstrated that z-FA-CMK is toxic to Jurkat T cells and induces apoptosis at low concentrations, while at higher concentrations the cells die of necrosis. - Highlights: • z-FA-CMK is toxic and induce cell death in the human T cells. • z-FA-CMK toxicity requires the CMK group, alanine and the benzyloxycarbonyl group. • z-FA-CMK induced apoptosis at low concentration and necrosis at high concentration.

  1. Effect Of Simulated Microgravity On Activated T Cell Gene Transcription

    NASA Technical Reports Server (NTRS)

    Morrow, Maureen A.

    2003-01-01

    Studies of T lymphocytes under the shear stress environment of clinorotation have demonstrated an inhibition of activation in response to TCR mediated signaling. These results mimic those observed during space flight. This work investigates the molecular signaling events of T lymphocyte activation with clinorotation. Purified human T lymphocytes and the T cell clone Jurkat exhibit an uncoupling of signaling as mediated through the TCR. Activation of the transcription factor AP-1 is inhibited while activation of NFAT occurs. NFAT dephosphorylation and activation is dependent on sustained Ca(++) influx. Alternatively, AP-1, which consists of two transcription factors, jun and fos, is activated by PKC and Ras mediated pathways. TCR signaling is known to be dependent on cytoskeletal rearrangements, in particular, raft aggregation is critical. Raft aggregation, as mediated through GM, crosslinking, overcomes the inhibition of T lymphocyte activation with clinorotation, indicating that the block is occurring upstream of raft aggregation. Clinorotation is shown to have an effect similar to a weak TCR signal.

  2. Effect of substrate mechanical properties on T cell activation

    NASA Astrophysics Data System (ADS)

    Hui, King; Upadhyaya, Arpita

    2013-03-01

    T cell activation is a key process in cell-mediated immunity, and engagement of T cell receptors by peptides on antigen presenting cells leads to activation of signaling cascades as well as cytoskeletal reorganization and large scale membrane deformations. While significant advances have been made in understanding the biochemical signaling pathways, the effects imposed by the physical environment and the role of mechanical forces on cell activation are not well understood. In this study, we have used anti-CD3 coated elastic polyacrylamide gels as stimulatory substrates to enable the spreading of Jurkat T cells and the measurement of cellular traction forces. We have investigated the effect of substrate stiffness on the dynamics of T cell spreading and cellular force generation. We found that T cells display more active and sustained edge dynamics on softer gels and that they exert increased traction stresses with increasing gel stiffness. A dynamic actin cytoskeleton was required to maintain the forces generated during activation, as inferred from small molecule inhibition experiments. Our results indicate an important role for physical properties of the antigen presenting cell as well as cytoskeleton-driven forces in signaling activation.

  3. Chitosan-modified cobalt oxide nanoparticles stimulate TNF-α-mediated apoptosis in human leukemic cells.

    PubMed

    Chattopadhyay, Sourav; Dash, Sandeep Kumar; Kar Mahapatra, Santanu; Tripathy, Satyajit; Ghosh, Totan; Das, Balaram; Das, Debasis; Pramanik, Panchanan; Roy, Somenath

    2014-03-01

    The objective of this study was to develop chitosan-based delivery of cobalt oxide nanoparticles to human leukemic cells and investigate their specific induction of apoptosis. The physicochemical properties of the chitosan-coated cobalt oxide nanoparticles were characterized using transmission electron microscopy, dynamic light scattering, X-ray diffraction, and Fourier transform infrared spectroscopy. The solubility of chitosan-coated cobalt oxide nanoparticles was higher at acidic pH, which helps to release more cobalt ions into the medium. Chitosan-coated cobalt oxide nanoparticles showed good compatibility with normal cells. However, our results showed that exposure of leukemic cells (Jurkat cells) to chitosan-coated cobalt oxide nanoparticles caused an increase in reactive oxygen species generation that was abolished by pretreatment of cells with the reactive oxygen species scavenger N-acetyl-L-cysteine. The apoptosis of Jurkat cells was confirmed by flow-cytometric analysis. Induction of TNF-α secretion was observed from stimulation of Jurkat cells with chitosan-coated cobalt oxide nanoparticles. We also tested the role of TNF-α in the induction of Jurkat cell death in the presence of TNF-α and caspase inhibitors. Treatment of leukemic cells with a blocker had a greater effect on cancer cell viability. From our findings, oxidative stress and caspase activation are involved in cancer cell death induced by chitosan-coated cobalt oxide nanoparticles. PMID:24445996

  4. Resveratrol‑4‑O‑D‑(2'‑galloyl)‑glucopyranoside exerts an anticancer effect on leukemia cells via inducing apoptosis.

    PubMed

    Chen, Pu; Wang, Beili; Pan, Baishen; Guo, Wei

    2016-03-01

    The aim of the present study was to investigate the anticancer effects of resveratrol‑4‑O‑D-(2'‑galloyl)-glucopyranoside (REG) on leukemia and the mechanism underlying its effects. Three leukemia cell lines (HL‑60, Jurkat and U937) were used in this study. A Cell Counting kit‑8 assay was performed to evaluate the anti‑proliferative activity of REG on leukemia cell lines, and flow cytometric analysis was used to detect REG‑induced apoptosis. In addition, western blot analysis was conducted to detect the levels of apoptosis‑related proteins including, cytochrome c, cleaved (c)‑caspases‑3 and ‑9, B‑cell lymphoma 2 (Bcl‑2) and Bcl‑2‑associated protein x (Bax). Finally, a HL‑60 cell xenograft model in nude mice was used to evaluate the antitumor effect of REG on leukemia in vivo. The present results indicated that REG can significantly inhibit the proliferation of HL‑60, Jurkat and U937 cell lines in a concentration‑ and time‑dependent manner. The half maximal inhibitory concentration values were 38.4, 49.1 and 48.2 µg/ml for HL‑60, Jurkat and U937 cells, respectively. Furthermore, flow cytometric analysis demonstrated that REG can induce the apoptosis of HL‑60 cells, as well as increase the levels of cytochrome c, c‑caspases‑3 and ‑9, and Bax, as well as downregulate the expression of Bcl‑2. In vivo, REG was found to possess a marked anticancer effect on leukemia. In combination, the present results indicated that REG exerts significant anticancer effects on leukemia in vivo and in vitro through the induction of apoptosis. PMID:26781500

  5. Low toxic and high soluble camptothecin derivative 2-47 effectively induces apoptosis of tumor cells in vitro.

    PubMed

    Zhou, Yao; Zhao, Hong-Ye; Jiang, Du; Wang, Lu-Yao; Xiang, Cen; Wen, Shao-Peng; Fan, Zhen-Chuan; Zhang, Yong-Min; Guo, Na; Teng, Yu-Ou; Yu, Peng

    2016-04-01

    The cytotoxic activity of camptothecin derivatives is so high that these compounds need to be further modified before their successful application as anti-cancer agents clinically. In this study, we reported the synthesis and biological evaluation of a novel camptothecin derivative called compound 2-47. The changes in structure did not reduce its activity to inhibit DNA topoisomerase I. Compound 2-47 induced apoptosis of many tumor cells including leukemia cells K562, Jurkat, HL-60, breast cancer cell BT-549, colon cancer cell HT-29 and liver cancer cell HepG2 with a half maximal inhibitory concentration (IC50) of 2- to 3-fold lower than HCPT as a control. In particular, 2-47 inhibited the proliferation of Jurkat cells with an IC50 of as low as 40 nM. By making use of Jurkat cell as a model, following treatment of Jurkat cells, compound 2-47 activated caspase-3 and PARP, resulting in a decreased Bcl-2/Bax ratio. These data showed that compound 2-47 induces Jurkat cell death through the mitochondrial apoptotic pathway. In addition, compound 2-47 showed a decreased cytotoxic activity against normal cells and an improved solubility in low-polar solvent. For example, compound 2-47 solutes in CHCl3 130-fold higher than HCPT. Taken together, our data demonstrated that camptothecin derivative 2-47 notably inhibits the tumor cell proliferation through mitochondrial-mediated apoptosis in vitro. PMID:26879138

  6. The protein pheromone Er-1 of the ciliate Euplotes raikovi stimulates human T-cell activity: Involvement of interleukin-2 system

    SciTech Connect

    Cervia, Davide; Catalani, Elisabetta; Belardinelli, Maria Cristina; Perrotta, Cristiana; Picchietti, Simona; Alimenti, Claudio; Casini, Giovanni; Fausto, Anna Maria; Vallesi, Adriana

    2013-02-01

    Water-soluble protein signals (pheromones) of the ciliate Euplotes have been supposed to be functional precursors of growth factors and cytokines that regulate cell–cell interaction in multi-cellular eukaryotes. This work provides evidence that native preparations of the Euplotes raikovi pheromone Er-1 (a helical protein of 40 amino acids) specifically increases viability, DNA synthesis, proliferation, and the production of interferon-γ, tumor necrosis factor-α, interleukin (IL)-1β, IL-2, and IL-13 in human Jurkat T-cells. Also, Er-1 significantly decreases the mRNA levels of the β and γ subunits of IL-2 receptor (IL-2R), while the mRNA levels of the α subunit appeared to be not affected. Jurkat T-cell treatments with Er-1 induced the down-regulation of the IL-2Rα subunit by a reversible and time-dependent endocytosis, and increased the levels of phosphorylation of the extracellular signal-regulated kinases (ERK). The cell-type specificity of these effects was supported by the finding that Er-1, although unable to directly influence the growth of human glioma U-373 cells, induced Jurkat cells to synthesize and release factors that, in turn, inhibited the U-373 cell proliferation. Overall, these findings imply that Er-1 coupling to IL-2R and ERK immuno-enhances T-cell activity, and that this effect likely translates to an inhibition of glioma cell growth. -- Highlights: ► Euplotes pheromone Er-1 increases the growth of human Jurkat T-cells. ► Er-1 increases the T-cell production of specific cytokines. ► Er-1 activates interleukin-2 receptor and extracellular signal-regulated kinases. ► The immuno-enhancing effect of Er-1 on Jurkat cells translates to an inhibition of human glioma cell growth.

  7. Sphingosine 1-phosphate inhibits activation of caspases that cleave poly(ADP-ribose) polymerase and lamins during Fas- and ceramide-mediated apoptosis in Jurkat T lymphocytes.

    PubMed

    Cuvillier, O; Rosenthal, D S; Smulson, M E; Spiegel, S

    1998-01-30

    Ceramide, a sphingolipid generated by the hydrolysis of membrane-associated sphingomyelin, appears to play a role as a gauge of apoptosis. A further metabolite of ceramide, sphingosine 1-phosphate (SPP), prevents ceramide-mediated apoptosis, and it has been suggested that the balance between intracellular ceramide and SPP levels may determine the cell fate (Cuvillier, O., Pirianov, G, Kleuser, B., Vanek, P. G., Coso, O. A., Gutkind, J. S., and Spiegel, S. (1996) Nature 381, 800-803). Here, we investigated the role of SPP and the protein kinase C activator, phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA), in the caspase cascade leading to the proteolysis of poly(ADP-ribose) polymerase (PARP) and lamins. In Jurkat T cells, Fas ligation or addition of exogenous C2-ceramide induced activations of caspase-3/CPP32 and caspase-7/Mch3 followed by PARP cleavage, effects that can be blocked either by SPP or TPA. Furthermore, both SPP and TPA inhibit the activation of caspase-6/Mch2 and subsequent lamin B cleavage. Ceramide, in contrast to Fas ligation, did not induce activation of caspase-8/FLICE and neither SPP nor TPA were able to prevent this activation. Thus, SPP, likely generated via protein kinase C-mediated activation of sphingosine kinase, suppresses the apoptotic pathway downstream of FLICE but upstream of the executioner caspases, caspase-3, -6, and -7. PMID:9446602

  8. Freezing and post-thaw apoptotic behaviour of cells in the presence of palmitoyl nanogold particles

    NASA Astrophysics Data System (ADS)

    Thirumala, Sreedhar; Forman, Julianne M.; Monroe, W. Todd; Devireddy, Ram V.

    2007-05-01

    The aim of this study was to evaluate the freezing response of HeLa and Jurkat cells in the presence of commercially available nanoparticles, NPs (Palmitoyl Nanogold®, Nanoprobes). The cells were incubated with NPs for either 5 min or 3 h, and a calorimeter technique was then used to generate the volumetric shrinkage response during freezing at 20 °C min-1. Concomitantly, we also examined the effect of a commonly used cryoprotectant, dimethylsulfoxide, DMSO (10% v/v ratio) on the freezing response of HeLa and Jurkat cells. By fitting a model of water transport to the experimentally determined volumetric shrinkage data, the reference hydraulic conductivity, Lpg, (μm/min-atm) and activation energy, ELp, (kcal mol-1) were obtained. For HeLa cells, the values of Lpg ranged from 0.08 to 0.23 µm/min-atm, while ELp ranged from 10.9 to 37.4 kcal mol-1. For Jurkat cells these parameter values ranged from 0.05 to 0.16 µm/min-atm and 9.5 to 35.9 kcal mol-1. A generic optimal cooling rate equation was then used to predict the optimal rates of freezing HeLa and Jurkat cells in the presence and absence of DMSO and NPs. The post-thaw viability and apoptotic response of HeLa and Jurkat cells was further investigated by cooling cells at three rates in the presence and absence of DMSO and NPs using a commercially available controlled rate freezer. Jurkat cells treated in this manner demonstrated an increase in their adhesive properties after 18 h incubation and adhered strongly to the bottom of the culture plate. This observation prevented further analysis of Jurkat apoptotic and necrotic post-thaw responses. There was no significant effect of NPs or DMSO alone on HeLa cell viability prior to freezing. The post-thaw results from HeLa cells show that the NPs increased the measured post-freeze apoptotic response when cooled at 1 °C min-1, suggesting a possible therapeutic use of NPs in cryodestructive procedures.

  9. Impact of non-thermal plasma treatment on MAPK signaling pathways of human immune cell lines.

    PubMed

    Bundscherer, Lena; Wende, Kristian; Ottmüller, Katja; Barton, Annemarie; Schmidt, Anke; Bekeschus, Sander; Hasse, Sybille; Weltmann, Klaus-Dieter; Masur, Kai; Lindequist, Ulrike

    2013-10-01

    In the field of wound healing research non-thermal plasma (NTP) increasingly draws attention. Next to its intensely studied antibacterial effects, some studies already showed stimulating effects on eukaryotic cells. This promises a unique potential in healing of chronic wounds, where effective therapies are urgently needed. Immune cells do play an important part in the process of wound healing and their reaction to NTP treatment has yet been rarely examined. Here, we studied the impact of NTP treatment using the kinpen on apoptotic and proliferative cell signaling pathways of two human immune cell lines, the CD4(+)T helper cell line Jurkat and the monocyte cell line THP-1. Depending on NTP treatment time the number of apoptotic cells increased in both investigated cell types according to a caspase 3 assay. Western blot analysis pointed out that plasma treatment activated pro-apoptotic signaling proteins like p38 mitogen-activated protein kinase (p38 MAPK) and c-Jun N-terminal kinase 1 and 2 (JNK 1/2) in both cell types. Stronger signals were detected in Jurkat cells at comparable plasma treatment times. Intriguingly, exposure of Jurkat and THP-1 cells to plasma also activated the pro-proliferative signaling molecules extracellular signal-regulated kinase 1/2 (ERK 1/2) and MAPK/ERK kinase 1 and 2 (MEK 1/2). In contrast to Jurkat cells, the anti-apoptotic heat shock protein 27 (HSP27) was activated in THP-1 cells after plasma treatment, indicating a possible mechanism how THP-1 cells may reduce programmed cell death. In conclusion, several signaling cascades were activated in the examined immune cell lines after NTP treatment and in THP-1 monocytes a possible defense mechanism against plasma impacts could be revealed. Therefore, plasma might be a treatment option for wound healing. PMID:23735483

  10. Timed, sequential administration of paclitaxel improves its cytotoxic effectiveness in a cell culture model.

    PubMed

    Fisi, Viktória; Kátai, Emese; Bogner, Péter; Miseta, Attila; Nagy, Tamás

    2016-05-01

    Paclitaxel (taxol) is a chemotherapeutic agent frequently used in combination with other anti-neoplastic drugs. It is most effective during the M phase of the cell-cycle and tends to cause synchronization in malignant cells lines. In this study, we investigated whether timed, sequential treatment based on the cell-cycle characteristics could be exploited to enhance the cytotoxic effect of paclitaxel. We characterized the cell-cycle properties of a rapidly multiplying cell line (Sp2, mouse myeloma cells) by propidium-iodide DNA staining such as the lengths of various cell cycle phases and population duplication time. Based on this we designed a paclitaxel treatment protocol that comprised a primary and a secondary, timed treatment. We found that the first paclitaxel treatment synchronized the cells at the G2/M phase but releasing the block by stopping the treatment allowed a large number of cells to enter the next cell-cycle by a synchronized manner. The second treatment was most effective during the time when these cells approached the next G2/M phase and was least effective when it occurred after the peak time of this next G2/M phase. Moreover, we found that after mixing Sp2 cells with another, significantly slower multiplying cell type (Jurkat human T-cell leukemia) at an initial ratio of 1:1, the ratio of the two different cell types could be influenced by timed sequential paclitaxel treatment at will. Our results demonstrate that knowledge of the cell-cycle parameters of a specific malignant cell type could improve the effectivity of the chemotherapy. Implementing timed chemotherapeutic treatments could increase the cytotoxicity on the malignant cells but also decrease the side-effects since other, non-malignant cell types will have different cell-cycle characteristic and be out of synch during the treatment. PMID:27104236

  11. Timed, sequential administration of paclitaxel improves its cytotoxic effectiveness in a cell culture model

    PubMed Central

    Fisi, Viktória; Kátai, Emese; Bogner, Péter; Miseta, Attila; Nagy, Tamás

    2016-01-01

    ABSTRACT Paclitaxel (taxol) is a chemotherapeutic agent frequently used in combination with other anti-neoplastic drugs. It is most effective during the M phase of the cell-cycle and tends to cause synchronization in malignant cells lines. In this study, we investigated whether timed, sequential treatment based on the cell-cycle characteristics could be exploited to enhance the cytotoxic effect of paclitaxel. We characterized the cell-cycle properties of a rapidly multiplying cell line (Sp2, mouse myeloma cells) by propidium-iodide DNA staining such as the lengths of various cell cycle phases and population duplication time. Based on this we designed a paclitaxel treatment protocol that comprised a primary and a secondary, timed treatment. We found that the first paclitaxel treatment synchronized the cells at the G2/M phase but releasing the block by stopping the treatment allowed a large number of cells to enter the next cell-cycle by a synchronized manner. The second treatment was most effective during the time when these cells approached the next G2/M phase and was least effective when it occurred after the peak time of this next G2/M phase. Moreover, we found that after mixing Sp2 cells with another, significantly slower multiplying cell type (Jurkat human T-cell leukemia) at an initial ratio of 1:1, the ratio of the two different cell types could be influenced by timed sequential paclitaxel treatment at will. Our results demonstrate that knowledge of the cell-cycle parameters of a specific malignant cell type could improve the effectivity of the chemotherapy. Implementing timed chemotherapeutic treatments could increase the cytotoxicity on the malignant cells but also decrease the side-effects since other, non-malignant cell types will have different cell-cycle characteristic and be out of synch during the treatment. PMID:27104236

  12. CD31 promotes beta1 integrin-dependent engulfment of apoptotic Jurkat T lymphocytes opsonized for phagocytosis by fibronectin.

    PubMed

    Vernon-Wilson, Elizabeth F; Auradé, Frédéric; Brown, Simon B

    2006-06-01

    Phagocyte integrins, by binding "bridging" molecules, mediate the ingestion of late apoptotic cells and apoptotic bodies by mechanisms that remain obscure. We recently reported that human monocyte-derived macrophages capture viable and apoptotic human leukocytes through homophilic interactions involving CD31 and that CD31 then promotes the engulfment of apoptotic cells or the detachment of viable cells. We now report that CD31 homophilic interactions between phagocyte and target cells lead to activation of phagocyte alpha5beta1 integrin and the engulfment of apoptotic Jurkat T lymphocytes via a fibronectin (Fn) "bridge." Although Fn and serum served as an opsonin for beta1 integrin-dependent phagocytosis of apoptotic leukemic T cells, they failed to do so for neutrophils. Given the complexities and inherent variability of working with primary cells, we have refined our model to show that ligation of CD31 on THP-1 macrophages also regulates beta1 integrin-dependent phagocytosis of Fn-coated Latex beads. Thus, selective "tethering" of apoptotic leukocytes by phagocyte CD31 not only discriminates dying from viable cells but also selectively activates phagocyte integrins for the engulfment of apoptotic cells. PMID:16551678

  13. Specific Anti-Leukemic Activity of the Peptide Warnericin RK and Analogues and Visualization of Their Effect on Cancer Cells by Chemical Raman Imaging.

    PubMed

    Loiseau, Clémence; Augenstreich, Jacques; Marchand, Adrienne; Harté, Etienne; Garcia, Martine; Verdon, Julien; Mesnil, Marc; Lecomte, Sophie; Berjeaud, Jean-Marc

    2016-01-01

    Antimicrobial peptides can be used as therapeutic agents against cancer cells. Warnericin RK and derivatives (WarnG20D and WarnF14V) were tested on various, solid tumor or leukemia, cancer cells. These peptides appeared to be cytotoxic on all the cell types tested, cancerous as well healthy, but very interestingly displayed no deleterious effect on healthy mononuclear cells. The mode of action of the peptide was proposed to be membranolytic, using chemical Raman imaging. Addition of peptide induced a large disorganization of the membrane leading to the loss of the content of inner compartments of Jurkat cell, whereas no effect was observed on the healthy mononuclear cells. The less hemolytic peptides WarnG20D and WarnF14V could be good candidates for the leukemia treatment. PMID:27598770

  14. Protective Role of Hsp27 Protein Against Gamma Radiation-Induced Apoptosis and Radiosensitization Effects of Hsp27 Gene Silencing in Different Human Tumor Cells

    SciTech Connect

    Aloy, Marie-Therese Hadchity, Elie; Bionda, Clara; Diaz-Latoud, Chantal; Claude, Line; Rousson, Robert; Arrigo, Andre-Patrick; Rodriguez-Lafrasse, Claire

    2008-02-01

    Purpose: The ability of heat shock protein 27 (Hsp27) to protect cells from stressful stimuli and its increased levels in tumors resistant to anticancer therapeutics suggest that it may represent a target for sensitization to radiotherapy. In this study, we investigate the protective role of Hsp27 against radiation-induced apoptosis and the effect of its attenuation in highly expressing radioresistant cancer cell lines. Methods and Materials: We examined clonogenic death and the kinetics of apoptotic events in different tumor cell lines overexpressing or underexpressing Hsp27 protein irradiated with photons. The radiosensitive Jurkat cell line, which does not express Hsp27 constitutively or in response to {gamma}-rays, was stably transfected with Hsp27 complementary DNA. Attenuation of Hsp27 expression was accomplished by antisense or RNAi (interfering RNA) strategies in SQ20B head-and-neck squamous carcinoma, PC3 prostate cancer, and U87 glioblastoma radioresistant cells. Results: We measured concentration-dependent protection against the cytotoxic effects of radiation in Jurkat-Hsp27 cells, which led to a 50% decrease in apoptotic cells at 48 hours in the highest expressing cells. Underlying mechanisms leading to radiation resistance involved a significant increase in glutathione levels associated with detoxification of reactive oxygen species, a delay in mitochondrial collapse, and caspase activation. Conversely, attenuation of Hsp27 in SQ20B cells, characterized by their resistance to apoptosis, sensitizes cells to irradiation. This was emphasized by increased apoptosis, decreased glutathione basal level, and clonogenic cell death. Sensitization to irradiation was confirmed in PC3 and U87 radioresistant cells. Conclusion: Hsp27 gene therapy offers a potential adjuvant to radiation-based therapy of resistant tumors.

  15. Synthesis of N-methylarylnitrones derived from alkyloxybenzaldehydes and antineoplastic effect on human cancer cell lines.

    PubMed

    Costa, Débora S S; Martino, Thiago; Magalhães, Fernanda C; Justo, Graça; Coelho, Marsen G P; Barcellos, Julio C F; Moura, Victor B; Costa, Paulo R R; Sabino, Kátia C C; Dias, Ayres G

    2015-05-01

    New O-isoprenylated-N-methylarylnitrones derived from isomeric o, m and p-hydroxybenzaldehydes have been prepared and the antineoplastic effects on human cancer cell lines were evaluated. The O-geranylated nitrone LQB-278 (1b) and its isomers 2b and 3b inhibited the NO production, but the anti-leukemic activity was drastically dependent on nitrone isomer, with the 1b being the most effective one (IC₅₀ of 6.7 μM) on Jurkat leukemia cell, by MTT assay. In addition, 1b up-regulated p21CIP1/WAF1/Sdi1 protein expression (flow cytometry), a cell cycle inhibitor, reduced cell growth, and induced DNA fragmentation (increased sub-G1 phase cells) and phosphatidylserine externalization in plasmatic membrane (increased annexin V positive cells). Finally, the 1b up-regulation of p21 expression and apoptosis induction seem to be the mechanisms by which it promotes its anti-leukemic effects, making this new molecular architecture a promising prototype for leukemia intervention. PMID:25813896

  16. Impairment of Na(+),K(+)-ATPase in CD95(APO-1)-induced human T-cell leukemia cell apoptosis mediated by glutathione depletion and generation of hydrogen peroxide.

    PubMed

    Yin, W; Cheng, W; Shen, W; Shu, L; Zhao, J; Zhang, J; Hua, Z-C

    2007-08-01

    Human T-cell leukemia is a malignant disease that needs various regimens of cytotoxic chemotherapy to overcome drug resistance. Recently, Na(+),K(+)-ATPase has emerged as a potential target for cancer therapy. However, its exact signaling pathway in human T-cell leukemia cell death has not been well defined. In the current study, we found CD95(APO-1) was able to trigger the internalization of plasma membrane Na(+),K(+)-ATPase in Jurkat cells or primary T cells as a mechanism to suppress its activity. This internalization was closely relevant to intracellular glutathione (GSH) depletion in Jurkat cells downstream of Fas-associated death domain protein (FADD) and caspase 8. GSH depletion in Fas L-treated Jurkat cells induced the generation of hydrogen peroxide (H(2)O(2)), which subsequently increased the serine phosphorylation of Na(+),K(+)-ATPase alpha1 subunit. Exogenous H(2)O(2) even mimicked the effect of Fas L to upregulate the serine phosphorylation of Na(+),K(+)-ATPase alpha1 subunit and suppress Na(+),K(+)-ATPase activity. Overall, our results indicate that CD95(APO-1) induces the FADD- and caspase 8-dependent internalization of Na(+),K(+)-ATPase through intracellular GSH loss, and the subsequent generation of H(2)O(2)-mediated serine phosphorylation of Na(+),K(+)-ATPase alpha1 subunit. Taken together, this study presents a novel regulatory mechanism of Na(+),K(+)-ATPase in CD95(APO-1)-mediated human T-leukemia cell apoptosis. PMID:17554377

  17. Ultrasound-mediated structural changes in cells revealed by FTIR spectroscopy: A contribution to the optimization of gene and drug delivery

    NASA Astrophysics Data System (ADS)

    Grimaldi, Paola; Di Giambattista, Lucia; Giordani, Serena; Udroiu, Ion; Pozzi, Deleana; Gaudenzi, Silvia; Bedini, Angelico; Giliberti, Claudia; Palomba, Raffaele; Congiu Castellano, Agostina

    2011-12-01

    Ultrasound effects on biological samples are gaining a growing interest concerning in particular, the intracellular delivery of drugs and genes in a safe and in a efficient way. Future progress in this field will require a better understanding of how ultrasound and acoustic cavitation affect the biological system properties. The morphological changes of cells due to ultrasound (US) exposure have been extensively studied, while little attention has been given to the cells structural changes. We have exposed two different cell lines to 1 MHz frequency ultrasound currently used in therapy, Jurkat T-lymphocytes and NIH-3T3 fibroblasts, both employed as models respectively in the apoptosis and in the gene therapy studies. The Fourier Transform Infrared (FTIR) Spectroscopy was used as probe to reveal the structural changes in particular molecular groups belonging to the main biological systems. The genotoxic damage of cells exposed to ultrasound was ascertained by the Cytokinesis-Block Micronucleus (CBMN) assay. The FTIR spectroscopy results, combined with multivariate statistical analysis, regarding all cellular components (lipids, proteins, nucleic acids) of the two cell lines, show that Jurkat cells are more sensitive to therapeutic ultrasound in the lipid and protein regions, whereas the NIH-3T3 cells are more sensitive in the nucleic acids region; a meaningful genotoxic effect is present in both cell lines only for long sonication times while in the Jurkat cells also a significant cytotoxic effect is revealed for long times of exposure to ultrasound.

  18. Grassypeptolides As Natural Inhibitors of Dipeptidyl Peptidase 8 and T-Cell Activation

    PubMed Central

    Kwan, Jason C.; Liu, Yanxia; Ratnayake, Ranjala; Hatano, Ryo; Kuribara, Akiko; Morimoto, Chiko; Ohnuma, Kei; Paul, Valerie J.; Ye, Tao

    2014-01-01

    Natural products made by marine cyanobacteria are often highly modified peptides and depsipeptides that have the potential to act as inhibitors for proteases. In the interest of finding novel protease inhibition activity and selectivity grassypeptolide A (1) was screened against a panel of proteases and found to selectively inhibit DPP8 over DPP4. Grassypeptolides were also found to inhibit IL-2 production and proliferation in activated T-cells, consistent with a putative role of DPP8 in the immune system. These effects were also observed in Jurkat cells, and DPP activity in Jurkat cell cytosol was shown to be inhibited by grassypeptolides. In silico docking suggests two possible binding modes of grassypeptolides – both at the active site of DPP8 and at one of the entrances to the internal cavity. Collectively these results suggest that grassypeptolides may be useful tool compounds in the study of DPP8 function. PMID:24591193

  19. Nitric oxide and bcl-2 mediated the apoptosis induced by nickel(II) in human T hybridoma cells

    SciTech Connect

    Guan Fuqin; Zhang Dongmei; Wang Xinchang; Chen Junhui . E-mail: jhchen@nju.edu.cn

    2007-05-15

    Although effects of nickel(II) on the immune system have long been recognized, little is known about the effects of nickel(II) on the induction of apoptosis and related signaling events in T cells. In the present study, we investigated the roles and signaling pathways of nickel(II) in the induction of apoptosis in a human T cell line jurkat. The results showed that the cytotoxic effects of Ni involved significant morphological changes and chromosomal condensation (Hoechst 33258 staining). Analyses of hypodiploid cells and FITC-Annexin V and PI double staining showed significant increase of apoptosis in jurkat cells 6, 12 and 24 h after nickel(II) treatment. Flow cytometry analysis also revealed that the loss of mitochondrial membrane potential (MMP) occurred concomitantly with the onset of NiCl{sub 2}-induced apoptosis. Induction of apoptotic cell death by nickel was mediated by reduction of bcl-2 expression. Furthermore, nickel stimulated the generation of nitric oxide (NO). These results suggest that nickel(II) chloride induces jurkat cells apoptosis via nitric oxide generation, mitochondrial depolarization and bcl-2 suppression.

  20. Cytotoxicity of recombinant tamapin and related toxin-like peptides on model cell lines.

    PubMed

    Ramírez-Cordero, Belén; Toledano, Yanis; Cano-Sánchez, Patricia; Hernández-López, Rogelio; Flores-Solis, David; Saucedo-Yáñez, Alma L; Chávez-Uribe, Isabel; Brieba, Luis G; del Río-Portilla, Federico

    2014-06-16

    The scorpion toxin tamapin displays the most potent and selective blockage against KCa2.2 channels known to date. In this work, we report the biosynthesis, three-dimensional structure, and cytotoxicity on cancer cell lines (Jurkat E6-1 and human mammary breast cancer MDA-MB-231) of recombinant tamapin and five related peptides bearing mutations on residues (R6A,R7A, R13A, R6A-R7A, and GS-tamapin) that were previously suggested to be important for tamapin's activity. The indicated cell lines were used as they constitutively express KCa2.2 channels. The studied toxin-like peptides displayed lethal responses on Jurkat T cells and breast cancer cells; their effect is dose- and time-dependent with IC50 values in the nanomolar range. The order of potency is r-tamapin>GS-tamapin>R6A>R13A>R6A-R7A>R7A for Jurkat T cells and r-tamapin>R7A for MDA-MB-231 breast cancer cells. Our structural determination by NMR demonstrated that r-tamapin preserves the folding of the αKTx5 subfamily and that neither single nor double alanine mutations affect the three-dimensional structure of the wild-type peptide. In contrast, our activity assays show that changes in cytotoxicity are related to the chemical nature of certain residues. Our results suggest that the toxic activity of r-tamapin on Jurkat and breast cancer cells could be mediated by the interaction of charged residues in tamapin with KCa2.2 channels via the apoptotic cell death pathway. PMID:24821061

  1. Polyalkoxybenzenes from plants. 5. Parsley seed extract in synthesis of azapodophyllotoxins featuring strong tubulin destabilizing activity in the sea urchin embryo and cell culture assays.

    PubMed

    Semenova, Marina N; Kiselyov, Alex S; Tsyganov, Dmitry V; Konyushkin, Leonid D; Firgang, Sergei I; Semenov, Roman V; Malyshev, Oleg R; Raihstat, Mikhail M; Fuchs, Fabian; Stielow, Anne; Lantow, Margareta; Philchenkov, Alex A; Zavelevich, Michael P; Zefirov, Nikolay S; Kuznetsov, Sergei A; Semenov, Victor V

    2011-10-27

    A series of 4-azapodophyllotoxin derivatives with modified rings B and E have been synthesized using allylpolyalkoxybenzenes from parsley seed oil. The targeted molecules were evaluated in vivo in a phenotypic sea urchin embryo assay for antimitotic and tubulin destabilizing activity. The most active compounds identified by the in vivo sea urchin embryo assay featured myristicin-derived ring E. These molecules were determined to be more potent than podophyllotoxin. Cytotoxic effects of selected molecules were further confirmed and evaluated by conventional assays with A549 and Jurkat human leukemic T-cell lines including cell growth inhibition, cell cycle arrest, cellular microtubule disruption, and induction of apoptosis. The ring B modification yielded 6-OMe substituted molecule as the most active compound. Finally, in Jurkat cells, compound induced caspase-dependent apoptosis mediated by the apical caspases-2 and -9 and not caspase-8, implying the involvement of the intrinsic caspase-9-dependent apoptotic pathway. PMID:21916509

  2. Umbelliprenin Induces Apoptosis in CLL Cell Lines

    PubMed Central

    Ziai, Seyed Ali; Gholami, Omid; Iranshahi, Mehrdad; Zamani, Amir Hassan; Jeddi-Tehrani, Mahmood

    2012-01-01

    Chronic lymphocytic leukemia (CLL) remains an incurable disease that requires innovative new approaches to improve therapeutic outcome. Many Ferula species, including F. asa-foetida, synthesize terpenyloxy coumarins. One of these coumarins is umbelliprenin, which has been implicated with induction of apoptosis in some cancer cell lines. In this study induction of apoptosis by umbelliprenin on Jurkat T-CLL and Raji B-CLL cell lines was studied. In this regard, cells were incubated with various concentrations of umbelliprenin in-vitro for different times and assayed for apoptosis with annexin V–FITC/PI double staining flowcytometry method. Results showed that umbelliprenin induced apoptosis in leukemic cells in a dose- and time-dependent manner and that CLL cells were more susceptible to umbelliprenin induced cell death than normal peripheral blood mononuclear cell (PBMCs). Moreover, we study the induction of apoptosis in Jurkat cells by umbelliprenin in the presence of interleukin 4 (IL-4) as an agent that causes resistance to apoptosis in CLL cells, was also student. We showed that IL-4 can not reduce apoptotic effect of umbelliprenin. The preferential toxicity of umbelliprenin for CLL cells, supports the hypothesis that oral administration of umbelliprenin in the form of foods or folk medicines containing this coumarin, might enhance protection against the development of CLL in man with little side effects. In conclusion, umbelliprenin may be an effective therapeutic agent in the treatment of CLL, and thus clinical studies with umbelliprenin may be appropriate. PMID:24250490

  3. Assessment of the effect of sphingosine kinase inhibitors on apoptosis,unfolded protein response and autophagy of T-cell acute lymphoblastic leukemia cells; indications for novel therapeutics.

    PubMed

    Evangelisti, Cecilia; Evangelisti, Camilla; Teti, Gabriella; Chiarini, Francesca; Falconi, Mirella; Melchionda, Fraia; Pession, Andrea; Bertaina, Alice; Locatelli, Franco; McCubrey, James A; Beak, Dong Jae; Bittman, Robert; Pyne, Susan; Pyne, Nigel J; Martelli, Alberto M

    2014-09-15

    Sphingosine 1-phosphate (S1P) is a bioactive lipid that is formed by the phosphorylation of sphingosine and catalysed by sphingosine kinase 1 (SK1) or sphingosine kinase 2 (SK2). Sphingosine kinases play a fundamental role in many signaling pathways associated with cancer, suggesting that proteins belonging to this signaling network represent potential therapeutic targets. Over the last years, many improvements have been made in the treatment of T-cell acute lymphoblastic leukemia (T-ALL); however, novel and less toxic therapies are still needed, especially for relapsing and chemo-resistant patients. Here, we analyzed the therapeutic potential of SKi and ROMe, a sphingosine kinase 1 and 2 inhibitor and SK2-selective inhibitor, respectively. While SKi induced apoptosis, ROMe initiated an autophagic cell death in our in vitro cell models. SKi treatment induced an increase in SK1 protein levels in Molt-4 cells, whereas it activated the endoplasmic reticulum (ER) stress/unfolded protein response (UPR) pathway in Jurkat and CEM-R cells as protective mechanisms in a sub-population of T-ALL cells. Interestingly, we observed a synergistic effect of SKi with the classical chemotherapeutic drug vincristine. In addition, we reported that SKi affected signaling cascades implicated in survival, proliferation and stress response of cells. These findings indicate that SK1 or SK2 represent potential targets for treating T-ALL. PMID:25226616

  4. A novel 2,6-diisopropylphenyl-docosahexaenoamide conjugate induces apoptosis in T cell acute lymphoblastic leukemia cell lines

    SciTech Connect

    Altenburg, Jeffrey D.; Harvey, Kevin A.; McCray, Sharon; Xu, Zhidong; Siddiqui, Rafat A.

    2011-07-29

    Highlights: {yields} 2,6-Diisopropylphenyl-docosahexaenoamide conjugates (DIP-DHA) inhibits the proliferation of T-cell leukemic cell lines. {yields} DIP-DHA resulted in increased activation of caspase-3, and caspase-7. {yields} DIP-DHA significantly downregulated CXCR4 surface expression. -- Abstract: We have previously characterized the effects of 2,6-diisopropylphenyl-docosahexaenoamide (DIP-DHA) conjugates and their analogs on the proliferation and progression of breast cancer cell lines. For this study, we investigated the effects of the DIP-DHA conjugate on 2 representative T cell acute lymphoblastic leukemia (T-ALL) cell lines: CEM and Jurkat. Treatment of both cell lines with DIP-DHA resulted in significantly greater inhibition of proliferation and induction of apoptosis than that of parent compounds, 2,6-diisopropylphenol (DIP) or docosahexaenoate (DHA). Treatment of the cells with DIP-DHA resulted in increased activation of caspase-3, and caspase-7. Furthermore, induction of apoptosis in both cell lines was reversed in the presence of a caspase family inhibitor. Treatment with DIP-DHA reduced mitochondrial membrane potential. These observations suggest that the effects are driven by intrinsic apoptotic pathways. DIP-DHA treatment also downregulated surface CXCR4 expression, an important chemokine receptor involved in cancer metastasis that is highly expressed in both CEM and Jurkat cells. In conclusion, our data suggest that the DIP-DHA conjugate exhibits significantly more potent effects on CEM and Jurkat cells than that of DIP or DHA alone. These conjugates have potential use for treatment of patients with T cell acute lymphoblastic leukemia.

  5. Potent Suppression of Kv1.3 Potassium Channel and IL-2 Secretion by Diphenyl Phosphine Oxide-1 in Human T Cells

    PubMed Central

    Zhao, Ning; Dong, Qian; Du, Li-Li; Fu, Xiao-Xing; Du, Yi-Mei; Liao, Yu-Hua

    2013-01-01

    Diphenyl phosphine oxide-1 (DPO-1) is a potent Kv1.5 channel inhibitor that has therapeutic potential for the treatment of atrial fibrillation. Many other Kv1.5 channel blockers also potently inhibit the Kv1.3 channel, but whether DPO-1 blocks Kv1.3 channels has not been investigated. The Kv1.3 channel is highly expressed in activated T cells, which is considered a favorable target for immunomodulation. Accordingly, we hypothesized that DPO-1 may exert immunosuppressive and anti-inflammatory effects by inhibiting Kv1.3 channel activity. In this study, DPO-1 blocked Kv1.3 current in a voltage-dependent and concentration-dependent manner, with IC50 values of 2.58 µM in Jurkat cells and 3.11 µM in human peripheral blood T cells. DPO-1 also accelerated the inactivation rate and negatively shifted steady-state inactivation. Moreover, DPO-1 at 3 µM had no apparent effect on the Ca2+ activated potassium channel (KCa) current in both Jurkat cells and human peripheral blood T cells. In Jurkat cells, pre-treatment with DPO-1 for 24 h decreased Kv1.3 current density, and protein expression by 48±6% and 60±9%, at 3 and 10 µM, respectively (both p<0.05). In addition, Ca2+ influx to Ca2+-depleted cells was blunted and IL-2 production was also reduced in activated Jurkat cells. IL-2 secretion was also inhibited by the Kv1.3 inhibitors margatoxin and charybdotoxin. Our results demonstrate for the first time that that DPO-1, at clinically relevant concentrations, blocks Kv1.3 channels, decreases Kv1.3 channel expression and suppresses IL-2 secretion. Therefore, DPO-1 may be a useful treatment strategy for immunologic disorders. PMID:23717641

  6. NF-kappa B activity in T cells stably expressing the Tax protein of human T cell lymphotropic virus type I

    SciTech Connect

    Lacoste, J.; Cohen, L.; Hiscott, J. )

    1991-10-01

    The effect of constitutive Tax expression on the interaction of NF-{kappa} B with its recognition sequence and on NF-{kappa} B-dependent gene expression was examined in T lymphoid Jurkat cell lines (19D and 9J) stably transformed with a Tax expression vector. Tax expressing T cell lines contained a constitutive level of NF-{kappa} B binding activity, detectable by mobility shift assay and uv cross-linking using a palindromic NF-{kappa} B probe homologous to the interferon beta PRDII site. In Jurkat and NC2.10 induction with phorbol esters resulted in the appearance of new DNA binding proteins of 85, 75, and 54 kDa, whereas in Tax expressing cells the 85-kDa protein and a 92-kDa DNA binding protein were constitutively induced. Expression of Tax protein in 19D and 9J resulted in transcription of the endogenous NF-kappa B-dependent granulocyte-macrophage colony stimulating factor gene and increased basal level expression of transfected NF-kappa B-regulated promoters. Nonetheless transcription of both the endogenous and the transfected gene was inducible by PMA treatment. Tax expression in Jurkat T cells may alter the stoichiometry of NF-kappa B DNA binding proteins and thus change the expression of NF-kappa B-regulated promoters.

  7. Study of antitumor effect of selected vanadium and molybdenum organometallic complexes in human leukemic T-cells.

    PubMed

    Šebestová, Lucie; Havelek, Radim; Řezáčová, Martina; Honzíček, Jan; Kročová, Zuzana; Vinklárek, Jaromír

    2015-12-01

    This work describes cytotoxic effect of non-platinum metal-based compounds on the human T-leukemic cells with different p53 status (p53 wild-type MOLT-4 and p53-deficient Jurkat cells). The cytotoxic and apoptosis-inducing effect of the vanadium complex [(η(5)-C5H5)2V(5-NH2-phen)]OTf (V1) and molybdenum complex [(η(3)-C3H5)Mo(CO)2(phen)Cl] (Mo1) were studied using flow cytometry, spectrophotometry and Western blotting. We found that the cytotoxic effect of both tested complexes after 24 h is higher against the both examined cell lines than that of cis-platin (cis-DDP). At later investigated time intervals of 48 and 72 h, the cytotoxic effect of the cis-DDP increased but the values of the cytotoxicity of the tested V1 and Mo1 complexes remained unchanged, with the cytotoxicity of V1 comparable to that of cis-DDP. Furthermore we observed that the apoptotic process was induced by the activation of the caspases 9 (intrinsic pathway) and 8 (extrinsic pathway) in cells exposed to evaluated complexes. In case of the p53 wild-type MOLT-4 cells, the expression of the tumor-suppressor protein p53 and its form phosphorylated at the serine 15 increased after both V1 and Mo1 treatment, similar to the effect of cis-DDP. PMID:26391003

  8. Spaceflight and clinorotation cause cytoskeleton and mitochondria changes and increases in apoptosis in cultured cells

    NASA Technical Reports Server (NTRS)

    Schatten, H.; Lewis, M. L.; Chakrabarti, A.

    2001-01-01

    The cytoskeleton is a complex network of fibers that is sensitive to environmental factors including microgravity and altered gravitational forces. Cellular functions such as transport of cell organelles depend on cytoskeletal integrity; regulation of cytoskeletal activity plays a role in cell maintenance, cell division, and apoptosis. Here we report cytoskeletal and mitochondria alterations in cultured human lymphocyte (Jurkat) cells after exposure to spaceflight and in insect cells of Drosophila melanogaster (Schneider S-1) after exposure to conditions created by clinostat rotation. Jurkat cells were flown on the space shuttle in Biorack cassettes while Schneider S-1 cells were exposed to altered gravity forces as produced by clinostat rotation. The effects of both treatments were similar in the different cell types. Fifty percent of cells displayed effects on the microtubule network in both cell lines. Under these experimental conditions mitochondria clustering and morphological alterations of mitochondrial cristae was observed to various degrees after 4 and 48 hours of culture. Jurkat cells underwent cell divisions during exposure to spaceflight but a large number of apoptotic cells was also observed. Similar results were obtained in Schneider S-1 cells cultured under clinostat rotation. Both cell lines displayed mitochondria abnormalities and mitochondria clustering toward one side of the cells which is interpreted to be the result of microtubule disruption and failure of mitochondria transport along microtubules. The number of mitochondria was increased in cells exposed to altered gravity while cristae morphology was severely affected indicating altered mitochondria function. These results show that spaceflight as well as altered gravity produced by clinostat rotation affects microtubule and mitochondria organization and results in increases in apoptosis. Grant numbers: NAG 10-0224, NAG2-985. c 2001. Elsevier Science Ltd. All rights reserved.

  9. Laccase purified from Cerrena unicolor exerts antitumor activity against leukemic cells

    PubMed Central

    MATUSZEWSKA, ANNA; KARP, MARTA; JASZEK, MAGDALENA; JANUSZ, GRZEGORZ; OSIŃSKA-JAROSZUK, MONIKA; SULEJ, JUSTYNA; STEFANIUK, DAWID; TOMCZAK, WALDEMAR; GIANNOPOULOS, KRZYSZTOF

    2016-01-01

    Chronic lymphocytic leukemia (CLL) is the most commonly observed adult hematological malignancy in Western countries. Despite the fact that recent improvements in CLL treatment have led to an increased percentage of complete remissions, CLL remains an incurable disease. Cerrena unicolor is a novel fungal source of highly active extracellular laccase (ex-LAC) that is currently used in industry. However, to the best of our knowledge, no reports regarding its anti-leukemic activity have been published thus far. In the present study, it was hypothesized that C. unicolor ex-LAC may possess cytotoxic activity against leukemic cell lines and CLL primary cells. C. unicolor ex-LAC was separated using anion exchange chromatography on diethylaminoethyl cellulose-Sepharose and Sephadex G-50 columns. The cytotoxic effects of ex-LAC upon 24- and 48-h treatment on HL-60, Jurkat, RPMI 8226 and K562 cell lines, as well as CLL primary cells of nine patients with CLL, were evaluated using 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assay. Annexin V/propidium iodide staining of Jurkat cells treated with ex-LAC was used to investigate apoptosis via flow cytometry. Ex-LAC induced changes in Jurkat and RPMI 8226 cells, as visualized by fluorescence and scanning electron microscopy (SEM). The XTT assay revealed high cytotoxic rates following treatment with various concentrations of ex-LAC on all the cell lines and CLL primary cells analyzed, with a half maximal inhibitory concentration ranging from 0.4 to 1.1 µg/ml. Fluorescence microscopy and SEM observations additionally revealed apoptotic changes in Jurkat and RPMI 8226 cells treated with ex-LAC, compared with control cells. These results were in agreement with the apoptosis analysis of Jurkat cells on flow cytometry. In conclusion, C. unicolor ex-LAC was able to significantly induce cell apoptosis, and may represent a novel therapeutic agent for the treatment of various hematological neoplasms. PMID

  10. Specific inhibition of Wee1 kinase and Rad51 recombinase: a strategy to enhance the sensitivity of leukemic T-cells to ionizing radiation-induced DNA double-strand breaks.

    PubMed

    Havelek, Radim; Cmielova, Jana; Kralovec, Karel; Bruckova, Lenka; Bilkova, Zuzana; Fousova, Ivana; Sinkorova, Zuzana; Vavrova, Jirina; Rezacova, Martina

    2014-10-24

    Present-day oncology sees at least two-thirds of cancer patients receiving radiation therapy as a part of their anticancer treatment. The objectives of the current study were to investigate the effects of the small molecule inhibitors of Wee1 kinase II (681641) and Rad51 (RI-1) on cell cycle progression, DNA double-strand breaks repair and apoptosis following ionizing radiation exposure in human leukemic T-cells Jurkat and MOLT-4. Pre-treatment with the Wee1 681641 or Rad51 RI-1 inhibitor alone increased the sensitivity of Jurkat cells to irradiation, however combining both inhibitors together resulted in a further enhancement of apoptosis. Jurkat cells pre-treated with inhibitors were positive for γH2AX foci 24h upon irradiation. MOLT-4 cells were less affected by inhibitors application prior to ionizing radiation exposure. Pre-treatment with Rad51 RI-1 had no effect on apoptosis induction; however Wee1 681641 increased ionizing radiation-induced cell death in MOLT-4 cells. PMID:25285634

  11. Dynamic effects and applications for nanosecond pulsed electric fields in cells and tissues

    NASA Astrophysics Data System (ADS)

    Beebe, Stephen J.; Blackmore, Peter F.; Hall, Emily; White, Jody A.; Willis, Lauren K.; Fauntleroy, Laura; Kolb, Juergen F.; Schoenbach, Karl H.

    2005-04-01

    Nanosecond, high intensity pulsed electric fields [nsPEFs] that are below the plasma membrane [PM] charging time constant have decreasing effects on the PM and increasing effects on intracellular structures and functions as the pulse duration decreases. When human cell suspensions were exposed to nsPEFs where the electric fields were sufficiently intense [10-300ns, <=300 kV/cm.], apoptosis signaling pathways could be activated in several cell models. Multiple apoptosis markers were observed in Jurkat, HL-60, 3T3L1-preadipocytes, and isolated rat adipocytes including decreased cell size and number, caspase activation, DNA fragmentation, and/or cytochrome c release into the cytoplasm. Phosphatidylserine externalization was observed as a biological response to nsPEFs in 3T3-L1 preadipocytes and p53-wildtype and -null human colon carcinoma cells. B10.2 mouse fibrosarcoma tumors that were exposed to nsPEFs ex vivo and in vivo exhibited DNA fragmentation, elevated caspase activity, and reduced size and weight compared to contralateral sham-treated control tumors. When nsPEF conditions were below thresholds for apoptosis and classical PM electroporation, non-apoptotic responses were observed similar to those initiated through PM purinergic receptors in HL-60 cells and thrombin in human platelets. These included Ca2+ mobilization from intracellular stores [endoplasmic reticulum] and subsequently through store-operated Ca2+ channels in the PM. In addition, platelet activation measured as aggregation responses were observed in human platelets. Finally, when nsPEF conditions followed classical electroporation-mediated transfection, the expression intensity and number of GFP-expressing cells were enhanced above cells exposed to electroporation conditions alone. These studies demonstrate that application of nsPEFs to cells or tissues can modulate cell-signaling mechanisms with possible applications as a new basic science tool, cancer treatment, wound healing, and gene therapy.

  12. Role of ascorbate in the activation of NF-kappaB by tumour necrosis factor-alpha in T-cells.

    PubMed Central

    Muñoz, E; Blázquez, M V; Ortiz, C; Gomez-Díaz, C; Navas, P

    1997-01-01

    The first product of ascorbate oxidation, the ascorbate free radical (AFR), acts in biological systems mainly as an oxidant, and through its role in the plasma membrane redox system exerts different effects on the cell. We have investigated the role of ascorbate, AFR and dehydroascorbate (DHA) in the activation of the NF-kappaB transcription factor in Jurkat T-cells stimulated by tumour necrosis factor-alpha (TNF-alpha). Here we show, by electrophoretic mobility shift assays, that ascorbate increases the binding of NF-kappaB to DNA in TNF-alpha-stimulated Jurkat cells. The ability of ascorbate to enhance cytoplasmic inhibitory IkBalpha protein degradation correlates completely with its capacity to induce NF-kappaB binding to DNA and to potentiate NF-kappaB-mediated transactivation of the HIV-1 long terminal repeat promoter in TNF-alpha-stimulated Jurkat cells but not in cells stimulated with PMA plus ionomycin. AFR behaves like ascorbate, while DHA and ascorbate phosphate do not affect TNF-alpha-mediated NF-kappaB activation. These results provide new evidence for a possible relationship between the activation of the electron-transport system at the plasma membrane by ascorbate or its free radical and redox-dependent gene transcription in T-cells. PMID:9224625

  13. Chemical sporulation and germination: cytoprotective nanocoating of individual mammalian cells with a degradable tannic acid-FeIII complex.

    PubMed

    Lee, Juno; Cho, Hyeoncheol; Choi, Jinsu; Kim, Doyeon; Hong, Daewha; Park, Ji Hun; Yang, Sung Ho; Choi, Insung S

    2015-12-01

    Individual mammalian cells were coated with cytoprotective and degradable films by cytocompatible processes maintaining the cell viability. Three types of mammalian cells (HeLa, NIH 3T3, and Jurkat cells) were coated with a metal-organic complex of tannic acid (TA) and ferric ion, and the TA-Fe(III) nanocoat effectively protected the coated mammalian cells against UV-C irradiation and a toxic compound. More importantly, the cell proliferation was controlled by programmed formation and degradation of the TA-Fe(III) nanocoat, mimicking the sporulation and germination processes found in nature. PMID:26528931

  14. Inhibition of proliferation by agricultural plant extracts in seven human adult T-cell leukaemia (ATL)-related cell lines.

    PubMed

    Kai, Hisahiro; Akamatsu, Ena; Torii, Eri; Kodama, Hiroko; Yukizaki, Chizuko; Sakakibara, Yoichi; Suiko, Masahito; Morishita, Kazuhiro; Kataoka, Hiroaki; Matsuno, Koji

    2011-07-01

    Adult T-cell leukaemia (ATL) is caused by human T-cell leukaemia virus type I (HTLV-I) infection and is resistant to conventional chemotherapy. We evaluated the inhibitory effects of agricultural plants on the proliferation of seven ATL-related human leukaemia cells, using three ATL cell lines (ED, Su9T01 and S1T), two human T-cell lines transformed by HTLV-I infection (HUT-102 and MT-2) and two HTLV-I-negative human T-cell acute lymphoblastic leukaemia cell lines (Jurkat and MOLT-4). A total of 52 samples of 80% ethanol extracts obtained from 30 types of agricultural plants were examined. On the basis of IC(50) values, we selected samples with greater activity than genistein, which was used as a positive control. The highest inhibitory effect was observed with extracts from leaves of Vaccinium virgatum Aiton (blueberry) on four cell lines (ED, Su9T01, HUT-102 and Jurkat); seeds of Momordica charantia L. (bitter gourd) exhibited the second highest activity. The bitter gourd seeds suppressed the proliferation of three cell lines (Su9T01, HUT-102 and Jurkat). The extracts from edible parts of Ipomea batatas LAM. (sweet potato), edible parts of Colocasia esculenta (L.) Schott (taro), skin of taro and seeds of Prunus mume Sieb. et Zucc. (mume) showed markedly greater inhibitory effects on Su9T01 than genistein. These findings suggest that ATL-preventative bioactive compounds may exist in these agricultural plants, which are considered to be functional foods. PMID:21293936

  15. The effect of hypogravity and hypergravity on cells of the immune system

    NASA Technical Reports Server (NTRS)

    Cogoli, A.

    1993-01-01

    This article reviews the gravity effects discovered in T lymphocytes and other cells of the immune system. The strong depression of mitogenic activation first observed in an experiment conducted in Spacelab 1 in 1983 triggered several other investigations in space and on the ground in the clinostat and in the centrifuge in the past 10 years. During this period, great progress was made in our knowledge of the complex mechanism of T cell activation as well as the technology to analyze the lymphokines produced during stimulation. Nevertheless, several aspects of the steps leading to activation are not yet clear. Studies in hypogravity and hypergravity may contribute to answering some of the questions. A recent investigation in the U.S. Spacelab SLS-1, based on a new technology in which leukocytes are attached to microcarrier beads, showed that the strong inhibition of activation in microgravity is due to a malfunction of monocytes acting as accessory cells. In fact, interleukin-1 production is nearly nil in resuspended monocytes, whereas T cell activation is doubled in attached cells. In hypergravity, but not at 1g, concanavalin A bound to erythrocytes activates B lymphocytes in addition to T cells. The activation of Jurkat cells is also severely impaired in space. These recent results have raised new questions that have to be answered in experiments to be conducted in space and on Earth in this decade. The experimental system, based on the mitogenic activation of T lymphocytes and accessory cells attached to microcarriers, offers an optimum model for studying basic biological mechanisms of the cell to assess the immunological fitness of humans in space and to test the feasibility of bioprocesses in space as well as on Earth.

  16. HER Specific TKIs Exert Their Antineoplastic Effects on Breast Cancer Cell Lines through the Involvement of STAT5 and JNK

    PubMed Central

    Gschwantler-Kaulich, Daphne; Grunt, Thomas W.; Muhr, Daniela; Wagner, Renate; Kölbl, Heinz; Singer, Christian F.

    2016-01-01

    Background HER-targeted tyrosine kinase inhibitors (TKIs) have demonstrated pro-apoptotic and antiproliferative effects in vitro and in vivo. The exact pathways through which TKIs exert their antineoplastic effects are, however, still not completely understood. Methods Using Milliplex assays, we have investigated the effects of the three panHER-TKIs lapatinib, canertinib and afatinib on signal transduction cascade activation in SKBR3, T47D and Jurkat neoplastic cell lines. The growth-inhibitory effect of blockade of HER and of JNK and STAT5 signaling was measured by proliferation- and apoptosis-assays using formazan dye labeling of viable cells, Western blotting for cleaved PARP-1 and immunolabeling for active caspase 3, respectively. Results All three HER-TKIs clearly inhibited proliferation and increased apoptosis in HER2 overexpressing SKBR3 cells, while their effect was less pronounced on HER2 moderately expressing T47D cells where they exerted only a weak antiproliferative and essentially no pro-apoptotic effect. Remarkably, phosphorylation/activation of JNK and STAT5A/B were inhibited by HER-TKIs only in the sensitive, but not in the resistant cells. In contrast, phosphorylation/activation of ERK/MAPK, STAT3, CREB, p70 S6 kinase, IkBa, and p38 were equally affected by HER-TKIs in both cell lines. Moreover, we demonstrated that direct pharmacological blockade of JNK and STAT5 abrogates cell growth in both HER-TKI-sensitive as well as -resistant breast cancer cells, respectively. Conclusion We have shown that HER-TKIs exert a HER2 expression-dependent anti-cancer effect in breast cancer cell lines. This involves blockade of JNK and STAT5A/B signaling, which have been found to be required for in vitro growth of these cell lines. PMID:26735495

  17. Cytotoxicity and cell cycle effects of novel indolo[2,3-b]quinoline derivatives.

    PubMed

    Humeniuk, Rita; Kaczmarek, Lukasz; Peczyńska-Czoch, Wanda; Marcinkowska, Ewa

    2003-01-01

    Cellular effects of novel indolo[2,3-b]quinoline derivatives were studied. These compounds are synthetic analogs of plant alkaloid neocryptolepine (5-methyl-5H-indolo[2,3-b]quinoline) present in extracts from Cryptolepis sanguinolenta. They are traditionally used in natural medicine in Central and West Africa. Previous molecular and computational studies indicated that these compounds were DNA intercalators and inhibitors of topoisomerase II. We have extended our studies on their mode of action to the cellular level. Past experiments have shown that these compounds were active in vitro against cell lines derived from solid tumors, so for the present studies we selected leukemic cell lines. Jurkat acute T cell, CCRF-CEM T lymphoblastoid, THP-1 acute monocytic, HL-60 acute promyelocytic leukemias, and HL-60/MX2 subline with reduced expression of topoisomerase II were used. We evaluated the cytotoxicity and cell cycle effects of the indolo[2,3-b]quinoline compounds. We also tested if these compounds were able to induce apoptosis in the cells. Our studies revealed that novel indolo[2,3-b]quinoline derivatives were more cytotoxic to all cell lines than etoposide (used as a reference topoisomerase II inhibitor), and that their cytotoxicity depended on the substituents introduced to the indolo[2,3-b]quinoline core. Surprisingly, our studies have shown that HL-60/MX2 cell line and also THP-1 cell line, resistant to etoposide, were susceptible to methyl- and methoxy-substituted indolo[2,3-b]quinoline derivatives. In parallel to the evaluation of cytotoxicity we studied cell cycle effects of these compounds. Treatment of HL-60 cells with etoposide in subcytotoxic concentrations resulted in a massive accumulation of the cells in the G2/M phase of the cell cycle. When we used subcytotoxic concentrations of our novel indolo[2,3-b]quinoline derivatives the cell cycle progression of HL-60 cells was not affected. Moreover, the cell cycle of HL-60/MX2 cells was not influenced by

  18. High expression of IMPACT protein promotes resistance to indoleamine 2,3-dioxygenase-induced cell death.

    PubMed

    Habibi, Darya; Jalili, Reza B; Forouzandeh, Farshad; Ong, Christopher J; Ghahary, Aziz

    2010-10-01

    Indoleamine 2,3-dioxygenase (IDO), a tryptophan degrading enzyme, is a potent immunomodulatory factor. IDO expression in fibroblasts selectively induces apoptosis in immune cells but not in primary skin cells. However, the mechanism(s) of this selective effect of IDO-induced low tryptophan environment is not elucidated. The aim of present study was to investigate whether the activity of general control non-derepressible-2(GCN2) kinase stress-responsive pathway and its known inhibitor, protein IMPACT homolog, in immune and skin cells are differentially regulated in response to IDO-induced low tryptophan environment. IDO-expressing human fibroblasts were co-cultured with Jurkat cells, human T cells, fibroblasts, or keratinocytes. Activation of GCN2 pathway was significantly higher in immune cells exposed to IDO-expressing environment relative to that of skin cells. In contrast, IMPACT was highly and constitutively expressed in skin cells while its expression was very low in stimulated T cells and undetectable in Jurkat cells. A significant IDO-induced suppressive as well as apoptotic effect was demonstrated in IMPACT knocked down fibroblasts co-cultured with IDO-expressing fibroblasts. Proliferation of Jurkat cells, stably transduced with IMPACT-expressing vector, was rescued significantly in tryptophan-deficient but not IDO-expressing environment. This may be due to the ability of IMPACT to recover the effects of IDO-mediated tryptophan depletion (GCN2 dependent) but not the effects of IDO-generated cytotoxic metabolites. These findings collectively suggest for the first time that high expression of protein IMPACT homolog in non-immune cells such as skin cells acts as a protective mechanism against IDO-induced GCN2 activation, therefore, makes them resistant to the amino acid-deprived environment caused by IDO. PMID:20648630

  19. Cadmium-coordinated supramolecule suppresses tumor growth of T-cell leukemia in mice

    PubMed Central

    Zhou, Xiaoping; Koizumi, Yukio; Zhang, Muxin; Natsui, Miyuki; Koyota, Souichi; Yamada, Manabu; Kondo, Yoshihiko; Hamada, Fumio; Sugiyama, Toshihiro

    2015-01-01

    Cadmium is a toxic pollutant with occupational and environmental significance, due to its diverse toxic effects. Supramolecules that conjugate and decontaminate toxic metals have potential for use in treatment of cadmium intoxication. In addition, metal-coordinating ability has been postulated to contribute to the cytotoxic effects of anti-tumor agents such as cisplatin or bleomycin. Thiacalixarenes, cyclic oligomers of p-alkylphenol bridged by sulfur atoms, are supramolecules known to have potent coordinating ability to metal ions. In this study, we show that cadmium-coordinated thiacalix[4]arene tetrasulfate (TC4ATS-Cd) exhibits an anti-proliferative effect against T-cell leukemia cells. Cadmium exhibited cytotoxicity with IC50 values ranging from 36 to 129 μM against epithelia-derived cancer cell lines, while TC4ATS-Cd elicited no significant cytotoxicity (IC50 > 947 μM). However, a number of T-cell leukemia cell lines exhibited marked sensitivity to TC4ATS-Cd. In Jurkat cells, toxicity of TC4ATS-Cd occurred with an IC50 of 6.9 μM, which is comparable to that of 6.5 μM observed for cadmium alone. TC4ATS-Cd induced apoptotic cell death through activation of caspase-3 in Jurkat cells. In a xenograft model, TC4ATS-Cd (13 mg/kg) treatment significantly suppressed the tumor growth of Jurkat cells in mice. In addition, TC4ATS-Cd-treated mice exhibited significantly less cadmium accumulation in liver and kidney compared to equimolar cadmium-treated mice. These results suggest that cadmium-coordinated supramolecules may have therapeutic potential for treatment of T-cell leukemia. PMID:25735932

  20. Real-time analysis of imatinib- and dasatinib-induced effects on chronic myelogenous leukemia cell interaction with fibronectin.

    PubMed

    Obr, Adam; Röselová, Pavla; Grebeňová, Dana; Kuželová, Kateřina

    2014-01-01

    Attachment of stem leukemic cells to the bone marrow extracellular matrix increases their resistance to chemotherapy and contributes to the disease persistence. In chronic myelogenous leukemia (CML), the activity of the fusion BCR-ABL kinase affects adhesion signaling. Using real-time monitoring of microimpedance, we studied in detail the kinetics of interaction of human CML cells (JURL-MK1, MOLM-7) and of control BCR-ABL-negative leukemia cells (HEL, JURKAT) with fibronectin-coated surface. The effect of two clinically used kinase inhibitors, imatinib (a relatively specific c-ABL inhibitor) and dasatinib (dual ABL/SRC family kinase inhibitor), on cell binding to fibronectin is described. Both imatinib and low-dose (several nM) dasatinib reinforced CML cell interaction with fibronectin while no significant change was induced in BCR-ABL-negative cells. On the other hand, clinically relevant doses of dasatinib (100 nM) had almost no effect in CML cells. The efficiency of the inhibitors in blocking the activity of BCR-ABL and SRC-family kinases was assessed from the extent of phosphorylation at autophosphorylation sites. In both CML cell lines, SRC kinases were found to be transactivated by BCR-ABL. In the intracellular context, EC50 for BCR-ABL inhibition was in subnanomolar range for dasatinib and in submicromolar one for imatinib. EC50 for direct inhibition of LYN kinase was found to be about 20 nM for dasatinib and more than 10 µM for imatinib. Cells pretreated with 100 nM dasatinib were still able to bind to fibronectin and SRC kinases are thus not necessary for the formation of cell-matrix contacts. However, a minimal activity of SRC kinases might be required to mediate the increase in cell adhesivity induced by BCR-ABL inhibition. Indeed, active (autophosphorylated) LYN was found to localize in cell adhesive structures which were visualized using interference reflection microscopy. PMID:25198091

  1. Bioorthogonal Click Chemistry-Based Synthetic Cell Glue.

    PubMed

    Koo, Heebeom; Choi, Myunghwan; Kim, Eunha; Hahn, Sei Kwang; Weissleder, Ralph; Yun, Seok Hyun

    2015-12-22

    Artificial methods of cell adhesion can be effective in building functional cell complexes in vitro, but methods for in vivo use are currently lacking. Here, a chemical cell glue based on bioorthogonal click chemistry with high stability and robustness is introduced. Tetrazine (Tz) and trans-cyclooctene (TCO) conjugated to the cell surface form covalent bonds between cells within 10 min in aqueous conditions. Glued, homogeneous, or heterogeneous cell pairs remain viable and stably attached in a microfluidic flow channel at a shear stress of 20 dyn cm(-2) . Upon intravenous injection of assembled Jurkat T cells into live mice, fluorescence microscopy shows the trafficking of cell pairs in circulation and their infiltration into lung tissues. These results demonstrate the promising potential of chemically glued cell pairs for various applications ranging from delivering therapeutic cells to studying cell-cell interactions in vivo. PMID:26768353

  2. Dichloromethane fraction of Melissa officinalis induces apoptosis by activation of intrinsic and extrinsic pathways in human leukemia cell lines.

    PubMed

    Ebrahimnezhad Darzi, Salimeh; Amirghofran, Zahra

    2013-06-01

    Various components from medicinal plants are currently used in cancer therapy because of their apoptosis-inducing effects. The present study has aimed to investigate the growth inhibitory and apoptotic effects of Melissa officinalis on tumor cells. We prepared different fractions of this plant to investigate their inhibitory effects on two leukemia cell lines, Jurkat and K562. Fractions with the highest inhibitory effects were examined for induction of apoptosis by the annexin V/propidium iodide assay and cell cycle changes by flow cytometry. Real-time polymerase chain reaction evaluated the changes in expression of apoptosis-related genes. Among different fractions, dichloromethane and n-hexane dose-dependent showed the strongest inhibitory effects on both K562 and Jurkat cells. The dichloromethane fraction significantly induced apoptosis at concentration of 50 µg/ml on Jurkat (85.66 ± 4.9%) and K562 cells (65.04 ± 0.93%) at 24 h after treatment (p < 0.002). According to cell cycle analysis, more than 70% of the cells accumulated in the sub-G1 phase when cultured in the presence of the dichloromethane fraction. This fraction up-regulated Fas and Bax mRNA expression as well as the Bax/Bcl-2 ratio according to cell type, showing its effect on the activation of both extrinsic and intrinsic pathways of apoptosis. The expression of apoptosis-related genes did not significantly change following treatment with the n-hexane fraction. These data indicated that the dichloromethane fraction of M. officinalis had the ability to induce apoptosis and change apoptosis-related gene expression in leukemia cells. PMID:23432355

  3. Delivery of anticancer drugs and antibodies into cells using ultrasound

    NASA Astrophysics Data System (ADS)

    Wu, Junru; Pepe, Jason; Rincon, Mercedes

    2005-04-01

    It has been shown experimentally in cell suspensions that pulsed ultrasound (2.0 MHz) could be used to deliver an anti-cancer drug (Adriamycin hydrochloride) into Jurkat lymphocytes and antibodies (goat anti rabbit IgG and anti mouse IgD) into human peripheral blood mononuclear (PBMC) cells and Jurkat lymphocytes assisted by encapsulated microbubbles (Optison). When Adriamycin hydrochloride (ADR) was delivered, the delivery efficiency reached 4.80% and control baseline (no ultrasound and no ADR) was 0.17%. When anti-rabbit IgD was delivered, the efficiencies were 34.90% (control baseline was 1.33%) and 32.50% (control baseline was 1.66%) respectively for Jurkat cells and PBMC. When goat anti rabbit IgG was delivered, the efficiencies were 78.60% (control baseline was 1.60%) and 57.50% (control baseline was 11.30%) respectively for Jurkat cells and PBMC.

  4. Chemical sporulation and germination: cytoprotective nanocoating of individual mammalian cells with a degradable tannic acid-FeIII complex

    NASA Astrophysics Data System (ADS)

    Lee, Juno; Cho, Hyeoncheol; Choi, Jinsu; Kim, Doyeon; Hong, Daewha; Park, Ji Hun; Yang, Sung Ho; Choi, Insung S.

    2015-11-01

    Individual mammalian cells were coated with cytoprotective and degradable films by cytocompatible processes maintaining the cell viability. Three types of mammalian cells (HeLa, NIH 3T3, and Jurkat cells) were coated with a metal-organic complex of tannic acid (TA) and ferric ion, and the TA-FeIII nanocoat effectively protected the coated mammalian cells against UV-C irradiation and a toxic compound. More importantly, the cell proliferation was controlled by programmed formation and degradation of the TA-FeIII nanocoat, mimicking the sporulation and germination processes found in nature.Individual mammalian cells were coated with cytoprotective and degradable films by cytocompatible processes maintaining the cell viability. Three types of mammalian cells (HeLa, NIH 3T3, and Jurkat cells) were coated with a metal-organic complex of tannic acid (TA) and ferric ion, and the TA-FeIII nanocoat effectively protected the coated mammalian cells against UV-C irradiation and a toxic compound. More importantly, the cell proliferation was controlled by programmed formation and degradation of the TA-FeIII nanocoat, mimicking the sporulation and germination processes found in nature. Electronic supplementary information (ESI) available: Experimental details, LSCM images, and SEM and TEM images. See DOI: 10.1039/c5nr05573c

  5. Am80 inhibits stromal cell-derived factor-1-induced chemotaxis in T-cell acute lymphoblastic leukemia cells.

    PubMed

    Matsumoto, Taichi; Jimi, Shiro; Hara, Shuuji; Takamatsu, Yasushi; Suzumiya, Junji; Tamura, Kazuo

    2010-03-01

    C-X-C motif chemokine receptor 4 (CXCR4) and stromal cell-derived factor-1 (SDF-1) play a potent role in metastasis and infiltration of many types of tumors, including T-cell acute lymphoblastic leukemia (T-ALL), into the central nervous system or lymph nodes. Although higher levels of CXCR4 expression have been shown to correlate with shorter survival of patients, effective drugs affecting cell surface CXCR4 expression are still unknown. In the present study, we examined the effects of a synthetic retinoid Am80 on CXCR4 expression of cultured T-ALL cells, such as Jurkat. Am80 inhibited surface CXCR4 expression and SDF-1-induced chemotaxis by the acceleration of CXCR4 internalization via activation of conventional PKC. Am80 may be an effective drug to inhibit the extramedullary infiltration of T-ALL cells. PMID:20141446

  6. Mechanisms of T-cell protection from death by IRX-2: a new immunotherapeutic

    PubMed Central

    Czystowska, Malgorzata; Szczepanski, Miroslaw J.; Szajnik, Marta; Quadrini, Karen; Brandwein, Harvey; Hadden, John W.

    2013-01-01

    Objectives IRX-2 is a novel immunotherapeutic containing physiologic quantities of several cytokines which protects human T lymphocytes from tumor-induced or drug-induced apoptosis. Here, we investigate the mechanisms responsible for IRX-2-mediated protection of T lymphocytes exposed to tumor-derived microvesicles (TMV). Methods Jurkat cells or primary human T cells ± IRX-2 were co-incubated with TMV and then examined by flow cytometry or Western blots for expression of molecules regulating cell survival (FLIP, Bcl-2, Bcl-xL, Mcl-1) or death (Fas, caspase 8, caspase 9, Bax, Bid). ANX V binding, caspase activation or cytochrome c release were also measured ± cycloheximide (CHX) or ± the Akt-specific inhibitor. Jurkat cells transfected with the cFLIP gene were used to evaluate the role of cFLIP in IRX-2-mediated protection. Effects of CHX on IRX-2-mediated protection and activation of NF-κB upon the TMV/IRX-2 treatment were also measured. Results IRX-2 protected T cells from apoptosis by preventing Fas overexpression induced by TMV and blocking caspase 8 activation by up-regulating cFLIP. Jurkat cells overexpressing cFLIP were more resistant to TMV-induced apoptosis than the mock-transfected cells (p < 0.02). Signaling via the PI3K/Akt pathway, IRX-2 corrected the imbalance of pro- versus anti-apoptotic proteins induced by TMV and promoted NF-κB translocation to the nucleus. CHX abolished IRX-2-mediated protection in T cells, suggesting that IRX-2 induces de novo synthesis of one or more proteins that are required for protection. Conclusions This biologic may be therapeutically useful for protection of activated T cells from tumor-induced immune suppression and death. PMID:21181158

  7. Investigation of a direct effect of nanosecond pulse electric fields on mitochondria

    NASA Astrophysics Data System (ADS)

    Estlack, Larry E.; Roth, Caleb C.; Cerna, Cesario Z.; Wilmink, Gerald J.; Ibey, Bennett L.

    2014-03-01

    The unique cellular response to nanosecond pulsed electric field (nsPEF) exposure, as compared to longer pulse exposure, has been theorized to be due to permeabilization of intracellular organelles including the mitochondria. In this investigation, we utilized a high-throughput oxygen and pH sensing system (Seahorse® XF24 extracellular flux analyzer) to assess the mitochondrial activity of Jurkat and U937 cells after nsPEF. The XF Analyzer uses a transient micro-chamber of only a few μL in specialized cell culture micro-plates to enable oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) to be monitored in real-time. We found that for nsPEF exposures of 10 pulses at 10-ns pulse width and at 50 kV/cm e-field, we were able to cause an increase in OCR in both U937 and Jurkat cells. We also found that high pulse numbers (>100) caused a significant decrease in OCR. Higher amplitude 150 kV/cm exposures had no effect on U937 cells and yet they had a deleterious effect on Jurkat cells, matching previously published 24 hour survival data. These results suggest that the exposures were modulating metabolic activity in cells possibly due to direct effects on the mitochondria themselves. To validate this hypothesis, we isolated mitochondria from U937 cells and exposed them similarly and found no significant change in metabolic activity for any pulse number. In a final experiment, we removed calcium from the buffer solution that the cells were exposed in and found that no significant enhancement in metabolic activity was observed. These results suggest that direct permeabilization of the mitochondria is unlikely a primary effect of nsPEF exposure and calcium-mediated intracellular pathway activation is likely responsible for observed pulse-induced mitochondrial effects.

  8. Scorpion venom component III inhibits cell proliferation by modulating NF-κB activation in human leukemia cells

    PubMed Central

    SONG, XIANGFENG; ZHANG, GUOJUN; SUN, AIPING; GUO, JIQIANG; TIAN, ZHONGWEI; WANG, HUI; LIU, YUFENG

    2012-01-01

    Scorpion venom contains various groups of compounds that exhibit anticancer activity against a variety of malignancies through a poorly understood mechanism. While the aberrant activation of nuclear factor κB (NF-κB) has been linked with hematopoietic malignancies, we hypothesized that scorpion venom mediates its effects by modulating the NF-κB signaling pathway. In the present study, we examined the effects of scorpion venom component III (SVCIII) on the human leukemia cell lines THP-1 and Jurkat and focused on the NF-κB signaling pathway. Our results showed that SVCIII inhibited cell proliferation, caused cell cycle arrest at G1 phase and inhibited the expression of cell cycle regulatory protein cyclin D1 in a dose-dependent manner in THP-1 and Jurkat cells. SVCIII also suppressed the constitutive NF-κB activation through inhibition of the phosphorylation and degradation of IκBα. NF-κB luciferase reporter activity was also inhibited by SVCIII. Our data suggest that SVCIII, a natural compound, may exert its antiproliferative effects by inhibiting the activation of NF-κB and, thus, has potential use in the treatment of hematopoietic malignancies, alone or in combination with other agents. PMID:23060939

  9. Effects of the tumor inhibitory triterpenoid avicin G on cell integrity, cytokinesis, and protein ubiquitination in fission yeast

    PubMed Central

    Gutterman, Jordan U.; Lai, Hong T.; Yang, Peirong; Haridas, Valsala; Gaikwad, Amos; Marcus, Stevan

    2005-01-01

    Avicins comprise a class of triterpenoid compounds that exhibit tumor inhibitory activity. Here we show that avicin G is inhibitory to growth of the fission yeast Schizosaccharomyces pombe. S. pombe cells treated with a lethal concentration of avicin G (20 μM) exhibited a shrunken morphology, indicating that avicin G adversely affects cell integrity. Cells treated with a sublethal concentration of avicin G (6.5 μM) exhibited a strong cytokinesis-defective phenotype (multiseptated cells), as well as cell morphology defects. These phenotypes bear resemblance to those resulting from loss of Rho1 GTPase function in S. pombe. Indeed, Rho1-deficient S. pombe cells were strongly hypersensitive to avicin G, suggesting that the compound may perturb Rho1-dependent processes. Consistent with previously observed effects in human Jurkat T cells, avicin G treatment resulted in hyperaccumulation of ubiquitinated proteins in S. pombe cells. Interestingly, proteasome-defective S. pombe mutants were not markedly hypersensitive to avicin G, whereas an anaphase-promoting complex (mitotic ubiquitin ligase) mutant exhibited avicin G resistance, suggesting that the increase in levels of ubiquitinated proteins resulting from avicin G treatment may be due to increased protein ubiquitination, rather than inhibition of 26S proteasome activity. Mutants defective in the cAMP/PKA pathway also exhibited resistance to avicin G. Our results suggest that S. pombe will be a useful model organism for elucidating molecular targets of avicin G and serve as a guide to clinical application where dysfunctional aspects of Rho and/or ubiquitination function have been demonstrated as in cancer, fibrosis, and inflammation. PMID:16118282

  10. Acetaminophen induces a caspase-dependent and Bcl-XL sensitive apoptosis in human hepatoma cells and lymphocytes.

    PubMed

    Boulares, A Hamid; Zoltoski, Anna J; Stoica, Bogdan A; Cuvillier, Olivier; Smulson, Mark E

    2002-01-01

    Acetaminophen is a widely used analgesic and antipyretic drug that exhibits toxicity at high doses to the liver and kidneys. This toxicity has been attributed to cytochrome P-450-generated metabolites which covalently modify target proteins. Recently, acetaminophen, in its unmetabolized form, has been shown to affect a variety of cells and tissues, for instance, testicular and lymphoid tissues and lymphocyte cell lines. The effects on cell viability of acetaminophen at a concentration comparable to that achieved in plasma during acetaminophen toxicity have now been examined with a hepatoma cell line SK-Hep1, primary human peripheral blood lymphocytes and human Jurkat T cells. Acetaminophen reduced cell viability in a time-dependent manner. Staining of cells with annexin-V also revealed that acetaminophen induced, after 8 hr of treatment, a loss of the asymmetry of membrane phospholipids, which is an early event associated with apoptosis. Acetaminophen triggered the release of cytochrome c from mitochondria into the cytosol, activation of caspase-3, 8, and 9, cleavage of poly(ADP-ribose) polymerase, and degradation of lamin B1 and DNA. Whereas cleavage of DNA into internucleosomal fragments was apparent in acetaminophen treated SK-Hep1 and primary lymphocytes, DNA was only degraded to 50-kb fragments in treated Jurkat cells. Overexpression of the antiapoptotic protein Bcl-XL prevented these various apoptotic events induced by acetaminophen in Jurkat cells. Caspase-8 activation was a postmictochondrial event and occurred in a Fas-independent manner. These results demonstrate that acetaminophen induces caspases-dependent apoptosis with mitochondria as a primary target. These results also reiterate the potential role of apoptosis in acetaminophen hepatic and extrahepatic toxicity. PMID:12005112

  11. Cell death induced by Bothrops asper snake venom metalloproteinase on endothelial and other cell lines.

    PubMed

    Brenes, Oscar; Muñóz, Eduardo; Roldán-Rodríguez, Raquel; Díaz, Cecilia

    2010-06-01

    Two adherent cell lines, BAEC and HeLa, and non-adherent Jurkat, were treated with snake venom metalloproteinase BaP1 to determine whether cytotoxicity, previously reported for this toxin, could be mediated by the process of anoikis. It was observed that there was no correlation between the ability of this toxin to induce loss of adherence, and the cytotoxic effect, since concentrations that do not induce loss of adherence (3-6 microg/mL), were able to trigger 50% of cytotoxicity in BAEC. In the case of HeLa, where toxicity was very low (less than 20% at maximun concentrations and times of exposure), significant detachment and no toxicity was observed at concentrations of 1.5 microg/mL, showing also no correlation between both events. We also observed differences between BAEC toxicity measured by XTT reduction and DNA fragmentation determined by flow cytometry (as an indicator of apoptosis), since concentrations that induce 100% of cytotoxicity barely showed any DNA fragmentation (12% at 24h), suggesting that if apoptosis was involved, DNA damage is still not present, although chromatin condensation, another indicator of apoptosis, is observed in 40% of the cells. Inhibition of BAEC cytotoxicity by caspase inhibitors indicate that apoptosis is playing a role in this process, but other mechanisms of cell death could be participating also. Another way to determine whether the mechanism of cell death was related to anoikis was using a non-adherent cell line, which should show substrate independence. We determined by TUNEL that at 50 microg/ml BaP1 triggered 50% of apoptosis at 96 h, an effect that was seen earlier, suggesting also that if this toxin was inducing apoptosis in a non-adherent cell line, the mechanism could not be related to loss of attachment. Cell cycle arrest in S phase was also observed in Jurkat cells, an effect that could be leading to apoptosis. In conclusion, since there was no correlation between cell detachment and cytotoxicity (and apoptosis

  12. Immunotherapy of murine sarcomas with interleukin 2. II. Activation of killer cells by human recombinant IL-2.

    PubMed

    Indrová, M; Bubeník, J; Toulcová, A

    1986-01-01

    Highly purified human recombinant interleukin 2 induced cytotoxicity in mouse spleen cells against mouse sarcoma cells when added during the 51Cr microcytotoxicity assay. It elicited similar levels of killer cell activation as did human lymphoid (Jurkat leukaemia-derived) or mouse lymphoid (EL-4 leukaemia-derived) IL-2 preparations. The susceptibility of six MC-induced mouse sarcomas to the cytolytic effect of lymphokine-activated killer cells was compared. Five (MC11, MC13, MC14, MC15, MC16) of six mouse sarcoma cell lines examined were sensitive in vitro to the LAK cell effect, whereas one cell line (MC12) was resistant. Since the sensitive and resistant target cell lines had been induced with the same carcinogen and in mice of the same genotype, they represent a very useful model for investigation of target cell structures responsible for the sensitivity to the LAK cell effect. PMID:3492397

  13. Investigation of cellular mechanisms involved in apoptosis induced by a synthetic naphthylchalcone in acute leukemia cell lines.

    PubMed

    Maioral, Mariana Franzoni; Moraes, Ana Carolina Rabello de; Sgambatti, Karen Ristau; Mascarello, Alessandra; Chiaradia-Delatorre, Louise Domeneghini; Yunes, Rosendo Augusto; Nunes, Ricardo José; Santos da Silva, Maria Cláudia

    2016-09-01

    We have previously reported the cytotoxic effects of chalcone A1, derived from 1-naphthaldehyde, in leukemia cell lines. On the basis of these findings, the main aim of this study was to elucidate some of the molecular mechanisms involved in apoptosis induced by chalcone A1 toward K562 and Jurkat cells. In both cell lines, chalcone A1 decreased the mitochondrial membrane potential, increased the expression of Bax proapoptotic protein, and decreased the expression of Bcl-2 antiapoptotic protein (resulting in the inversion of the Bcl-2/Bax ratio), which indicates the involvement of the intrinsic pathway. In addition, chalcone A1 increased the expression of FasR in Jurkat cells, which also indicates the involvement of the extrinsic pathway in this cell line. The results also showed an increased expression of effector caspase-3 and cleaved PARP-1 and a decreased expression of IAP protein survivin, which are consistent with apoptotic cell death. The decreased expression of Ki67 suggests that the mechanism involved in cell death induced by chalcone A1 also involves a decrease in cell proliferation. In ex-vivo experiments, chalcone A1 reduced the cell viability of blast cells collected from eight patients with different types of acute leukemia, confirming the cytotoxicity results found in vitro. The results obtained so far are very promising and further studies need to be carried out so that chalcone A1 can be used as a prototype for the development of new antileukemia agents. PMID:27337110

  14. Pre-clinical antitumour evaluation of Biphosphinic Palladacycle Complex in human leukaemia cells.

    PubMed

    Oliveira, Carlos R; Barbosa, Christiano M V; Nascimento, Fábio D; Lanetzki, Camilla S; Meneghin, Marília B; Pereira, Flávia E G; Paredes-Gamero, Edgar J; Ferreira, Alice T; Rodrigues, Tiago; Queiroz, Mary L S; Caires, Antonio C F; Tersariol, Ivarne L S; Bincoletto, Claudia

    2009-02-12

    Previous studies reported by our group have introduced a new antitumoural drug called Biphosphinic Palladacycle Complex (BPC). In this paper we show that BPC causes apoptosis in leukaemia cells (HL60 and Jurkat), but not in normal human lymphocytes. IC(50) values obtained for both cell lines using the MTT and trypan blue exclusion assays 5h after BPC treatment were lower than 8.0 microM. Using metachromatic fluorophore, acridine orange, we observed that BPC elicited lysosomal rupture of leukaemic cells. Furthermore, BPC triggered caspase-3 and caspase-6 activation and apoptosis in cell lines, inducing chromatin condensation, apoptotic bodies, and DNA fragmentation. Interestingly, the lysosomal cathepsin B inhibitor CA074 markedly decreased BPC-induced caspase-3 and caspase-6 activation as well as cell death. Lysosomal BPC-induced membrane destabilisation was not dependent on reactive oxygen species generation, which was consistent with the absence of cellular HL60 and Jurkat membrane lipid peroxidation. We conclude that, following BPC treatment, lysosomal membrane rupture precedes cell death and the apoptotic signalling pathway is initiated by the release of cathepsin B in the cytoplasm of leukaemia cells. As no toxic effects for human lymphocytes were observed, we suggest that BPC is more selective for transformed cells, mainly due to their exacerbated lysosome expression. PMID:19026616

  15. Arsenic, cadmium, and manganese levels in shellfish from Map Ta Phut, an industrial area in Thailand, and the potential toxic effects on human cells.

    PubMed

    Rangkadilok, Nuchanart; Siripriwon, Pantaree; Nookabkaew, Sumontha; Suriyo, Tawit; Satayavivad, Jutamaad

    2015-01-01

    Map Ta Phut Industrial Estate is a major industrial area in Thailand for both petrochemical and heavy industries. The release of hazardous wastes and other pollutants from these industries increases the potential for contamination in foods in the surrounding area, especially farmed shellfish. This study determined the arsenic (As), cadmium (Cd), and manganese (Mn) concentrations in the edible flesh of farmed shellfish, including Perna viridis, Meretrix meretrix, and Scapharca inaequivalvis, around the Map Ta Phut area using inductively coupled plasma mass spectrometry. The results showed that shellfish samples contained high levels of total As [1.84-6.42 mg kg(-1) wet weight (ww)]. High Mn concentrations were found in P. viridis and M. meretrix, whereas S. inaequivalis contained the highest Cd. Arsenobetaine (AsB) was found to be the major As species in shellfish (>45% of total As). The in vitro cytotoxicity of these elements was evaluated using human cancer cells (T47D, A549, and Jurkat cells). An observed decrease in cell viability in T47D and Jurkat cells was mainly caused by exposure to inorganic As (iAs) or Mn but not to AsB or Cd. The combined elements (AsB+Mn+Cd) at concentrations predicted to result from the estimated daily intake of shellfish flesh by the local people showed significant cytotoxicity in T47D and Jurkat cells. PMID:24986306

  16. 2-Deoxy-D-glucose Sensitizes Cancer Cells to Barasertib and Everolimus by ROS-independent Mechanism(s).

    PubMed

    Zhelev, Zhivko; Ivanova, Donika; Aoki, Ichio; Saga, Tsuneo; Bakalova, Rumiana

    2015-12-01

    The aim of the present study was to investigate: (i) the possibility of sensitizing cancer cells to anticancer drugs using the redox modulator 2-deoxy-D-glucose (2-DDG); (ii) to find such combinations with synergistic cytotoxic effect; (iii) and to clarify the role of reactive oxygen species (ROS) for induction of apoptosis and cytotoxicity through these combinations. The study covers 15 anticancer drugs--both conventional and new-generation. Four parameters were analyzed simultaneously in Jurkat leukemia cells, treated by drugs or 2-DDG (separately or in combination): cell viability, induction of apoptosis, levels of ROS, and level of protein-carbonyl products. Very well-expressed synergistic cytotoxic effects were found after 48-h treatment of Jurkat cells with 2-DDG in combination with: palbociclib, everolimus, lonafarnib, bortezomib, and barasertib. The synergistic cytotoxic effect of everolimus with 2-DDG was accompanied by very strong induction of apoptosis in cells, but a very strong reduction of ROS level. Changes in the levels of protein-carbonyl products were not detected. The synergistic cytotoxic effect of barasertib with 2-DDG was accompanied by very strong induction of apoptosis in cells, without any increase of ROS levels, but with an enhancement of protein-carbonyl products. PMID:26637878

  17. Cell stimulus and lysis in a microfluidic device with segmented gas-liquid flow.

    PubMed

    El-Ali, Jamil; Gaudet, Suzanne; Günther, Axel; Sorger, Peter K; Jensen, Klavs F

    2005-06-01

    We describe a microfluidic device with rapid stimulus and lysis of mammalian cells for resolving fast transient responses in cell signaling networks. The device uses segmented gas-liquid flow to enhance mixing and has integrated thermoelectric heaters and coolers to control the temperature during cell stimulus and lysis. Potential negative effects of segmented flow on cell responses are investigated in three different cell types, with no morphological changes and no activation of the cell stress-sensitive mitogen activated protein kinases observed. Jurkat E6-1 cells are stimulated in the device using alpha-CD3, and the resulting activations of ERK and JNK are presented for different time points. Stimulation of cells performed on chip results in pathway activation identical to that of conventionally treated cells under the same conditions. PMID:15924398

  18. Nanosecond pulsed electric fields induce poly(ADP-ribose) formation and non-apoptotic cell death in HeLa S3 cells

    SciTech Connect

    Morotomi-Yano, Keiko; Akiyama, Hidenori; Yano, Ken-ichi

    2013-08-30

    Highlights: •Nanosecond pulsed electric field (nsPEF) is a new and unique means for life sciences. •Apoptosis was induced by nsPEF exposure in Jurkat cells. •No signs of apoptosis were detected in HeLa S3 cells exposed to nsPEFs. •Formation of poly(ADP-ribose) was induced in nsPEF-exposed HeLa S3 cells. •Two distinct modes of cell death were activated by nsPEF in a cell-dependent manner. -- Abstract: Nanosecond pulsed electric fields (nsPEFs) have recently gained attention as effective cancer therapy owing to their potency for cell death induction. Previous studies have shown that apoptosis is a predominant mode of nsPEF-induced cell death in several cell lines, such as Jurkat cells. In this study, we analyzed molecular mechanisms for cell death induced by nsPEFs. When nsPEFs were applied to Jurkat cells, apoptosis was readily induced. Next, we used HeLa S3 cells and analyzed apoptotic events. Contrary to our expectation, nsPEF-exposed HeLa S3 cells exhibited no molecular signs of apoptosis execution. Instead, nsPEFs induced the formation of poly(ADP-ribose) (PAR), a hallmark of necrosis. PAR formation occurred concurrently with a decrease in cell viability, supporting implications of nsPEF-induced PAR formation for cell death. Necrotic PAR formation is known to be catalyzed by poly(ADP-ribose) polymerase-1 (PARP-1), and PARP-1 in apoptotic cells is inactivated by caspase-mediated proteolysis. Consistently, we observed intact and cleaved forms of PARP-1 in nsPEF-exposed and UV-irradiated cells, respectively. Taken together, nsPEFs induce two distinct modes of cell death in a cell type-specific manner, and HeLa S3 cells show PAR-associated non-apoptotic cell death in response to nsPEFs.

  19. Mechanistic Study of the Phytocompound, 2- β -D-Glucopyranosyloxy-1-hydroxytrideca-5,7,9,11-tetrayne in Human T-Cell Acute Lymphocytic Leukemia Cells by Using Combined Differential Proteomics and Bioinformatics Approaches.

    PubMed

    Shiau, Jeng-Yuan; Yin, Shu-Yi; Chang, Shu-Lin; Hsu, Yi-Jou; Chen, Kai-Wei; Kuo, Tien-Fen; Feng, Ching-Shan; Yang, Ning-Sun; Shyur, Lie-Fen; Yang, Wen-Chin; Wen, Tuan-Nan

    2015-01-01

    Bidens pilosa, a medicinal herb worldwide, is rich in bioactive polyynes. In this study, by using high resolution 2-dimensional gel electrophoresis coupled with mass spectrometry analysis, as many as 2000 protein spots could be detected and those whose expression was specifically up- or downregulated in Jurkat T cells responsive to the treatment with 2-β-D-glucopyranosyloxy-1-hydroxytrideca-5,7,9,11-tetrayne (GHTT) can be identified. GHTT treatment can upregulate thirteen proteins involved in signal transduction, detoxification, metabolism, energy pathways, and channel transport in Jurkat cells. Nine proteins, that is, thioredoxin-like proteins, BH3 interacting domain death agonist (BID protein involving apoptosis), methylcrotonoyl-CoA carboxylase beta chain, and NADH-ubiquinone oxidoreductase, were downregulated in GHTT-treated Jurkat cells. Further, bioinformatics tool, Ingenuity software, was used to predict signaling pathways based on the data obtained from the differential proteomics approach. Two matched pathways, relevant to mitochondrial dysfunction and apoptosis, in Jurkat cells were inferred from the proteomics data. Biochemical analysis further verified both pathways involving GHTT in Jurkat cells. These findings do not merely prove the feasibility of combining proteomics and bioinformatics methods to identify cellular proteins as key players in response to the phytocompound in Jurkat cells but also establish the pathways of the proteins as the potential therapeutic targets of leukemia. PMID:26557148

  20. Mechanistic Study of the Phytocompound, 2- β -D-Glucopyranosyloxy-1-hydroxytrideca-5,7,9,11-tetrayne in Human T-Cell Acute Lymphocytic Leukemia Cells by Using Combined Differential Proteomics and Bioinformatics Approaches

    PubMed Central

    Shiau, Jeng-Yuan; Yin, Shu-Yi; Chang, Shu-Lin; Hsu, Yi-Jou; Chen, Kai-Wei; Kuo, Tien-Fen; Feng, Ching-Shan; Yang, Ning-Sun; Shyur, Lie-Fen; Yang, Wen-Chin; Wen, Tuan-Nan

    2015-01-01

    Bidens pilosa, a medicinal herb worldwide, is rich in bioactive polyynes. In this study, by using high resolution 2-dimensional gel electrophoresis coupled with mass spectrometry analysis, as many as 2000 protein spots could be detected and those whose expression was specifically up- or downregulated in Jurkat T cells responsive to the treatment with 2-β-D-glucopyranosyloxy-1-hydroxytrideca-5,7,9,11-tetrayne (GHTT) can be identified. GHTT treatment can upregulate thirteen proteins involved in signal transduction, detoxification, metabolism, energy pathways, and channel transport in Jurkat cells. Nine proteins, that is, thioredoxin-like proteins, BH3 interacting domain death agonist (BID protein involving apoptosis), methylcrotonoyl-CoA carboxylase beta chain, and NADH-ubiquinone oxidoreductase, were downregulated in GHTT-treated Jurkat cells. Further, bioinformatics tool, Ingenuity software, was used to predict signaling pathways based on the data obtained from the differential proteomics approach. Two matched pathways, relevant to mitochondrial dysfunction and apoptosis, in Jurkat cells were inferred from the proteomics data. Biochemical analysis further verified both pathways involving GHTT in Jurkat cells. These findings do not merely prove the feasibility of combining proteomics and bioinformatics methods to identify cellular proteins as key players in response to the phytocompound in Jurkat cells but also establish the pathways of the proteins as the potential therapeutic targets of leukemia. PMID:26557148

  1. Inhibition of type I insulin-like growth factor receptor tyrosine kinase by picropodophyllin induces apoptosis and cell cycle arrest in T lymphoblastic leukemia/lymphoma

    PubMed Central

    Huang, Zhiwei; Fang, Zhijia; Zhen, Hong; Zhou, Li; Amin, Hesham M.; Shi, Ping

    2015-01-01

    It has been recently shown that IGF-IR contributes significantly to the survival of T lymphoblastic leukemia/lymphoma (T-LBL) cells, and it was therefore suggested that IGF-IR could represent a legitimate therapeutic target in this aggressive disease. Picropodphyllin (PPP) is a potent, selective inhibitor of IGF-IR that is currently used with notable success in clinical trials that include patients with aggressive types of epithelial tumors. In the present study, we tested the effects of PPP on Jurkat and Molt-3 cells; two prototype T-LBL cell lines. Our results demonstrate that PPP efficiently induced apoptotic cell death and cell cycle arrest of these two cells. These effects were attributable to alterations of downstream target proteins. By using proteomic analysis, 7 different proteins were found to be affected by PPP treatment of Jurkat cells. These proteins are involved in various aspects of cellular metabolism, cytoskeleton organization, and signal transduction pathways. The results suggest that PPP affects multiple signaling molecules and inhibits fundamental pathways that control cell growth and survival. Our study also provides novel evidence that PPP could be potentially utilized for the treatment of the aggressive T-LBL. PMID:24206093

  2. Cytotoxicity Mediated by the Fas Ligand (FasL)-activated Apoptotic Pathway in Stem Cells*

    PubMed Central

    Mazar, Julia; Thomas, Molly; Bezrukov, Ludmila; Chanturia, Alexander; Pekkurnaz, Gulcin; Yin, Shurong; Kuznetsov, Sergei A.; Robey, Pamela G.; Zimmerberg, Joshua

    2009-01-01

    Whereas it is now clear that human bone marrow stromal cells (BMSCs) can be immunosuppressive and escape cytotoxic lymphocytes (CTLs) in vitro and in vivo, the mechanisms of this phenomenon remain controversial. Here, we test the hypothesis that BMSCs suppress immune responses by Fas-mediated apoptosis of activated lymphocytes and find both Fas and FasL expression by primary BMSCs. Jurkat cells or activated lymphocytes were each killed by BMSCs after 72 h of co-incubation. In comparison, the cytotoxic effect of BMSCs on non-activated lymphocytes and on caspase-8(−/−) Jurkat cells was extremely low. Fas/Fc fusion protein strongly inhibited BMSC-induced lymphocyte apoptosis. Although we detected a high level of Fas expression in BMSCs, stimulation of Fas with anti-Fas antibody did not result in the expected BMSC apoptosis, regardless of concentration, suggesting a disruption of the Fas activation pathway. Thus BMSCs may have an endogenous mechanism to evade Fas-mediated apoptosis. Cumulatively, these data provide a parallel between adult stem/progenitor cells and cancer cells, consistent with the idea that stem/progenitor cells can use FasL to prevent lymphocyte attack by inducing lymphocyte apoptosis during the regeneration of injured tissues. PMID:19531476

  3. Quantitative Effect of Suboptimal Codon Usage on Translational Efficiency of mRNA Encoding HIV-1 gag in Intact T Cells

    PubMed Central

    Ngumbela, Kholiswa C.; Ryan, Kieran P.; Sivamurthy, Rohini; Brockman, Mark A.; Gandhi, Rajesh T.; Bhardwaj, Nina; Kavanagh, Daniel G.

    2008-01-01

    Background The sequences of wild-isolate strains of Human Immunodeficiency Virus-1 (HIV-1) are characterized by low GC content and suboptimal codon usage. Codon optimization of DNA vectors can enhance protein expression both by enhancing translational efficiency, and by altering RNA stability and export. Although gag codon optimization is widely used in DNA vectors and experimental vaccines, the actual effect of altered codon usage on gag translational efficiency has not been quantified. Methodology and Principal Findings To quantify translational efficiency of gag mRNA in live T cells, we transfected Jurkat cells with increasing doses of capped, polyadenylated synthetic mRNA corresponding to wildtype or codon-optimized gag sequences, measured Gag production by quantitative ELISA and flow cytometry, and estimated the translational efficiency of each transcript as pg of Gag antigen produced per µg of input mRNA. We found that codon optimization yielded a small increase in gag translational efficiency (approximately 1.6 fold). In contrast when cells were transfected with DNA vectors requiring nuclear transcription and processing of gag mRNA, codon optimization resulted in a very large enhancement of Gag production. Conclusions We conclude that suboptimal codon usage by HIV-1 results in only a slight loss of gag translational efficiency per se, with the vast majority of enhancement in protein expression from DNA vectors due to altered processing and export of nuclear RNA. PMID:18523584

  4. PKCθ and HIV-1 Transcriptional Regulator Tat Co-exist at the LTR Promoter in CD4+ T Cells

    PubMed Central

    López-Huertas, María Rosa; Li, Jasmine; Zafar, Anjum; Rodríguez-Mora, Sara; García-Domínguez, Carlota; Mateos, Elena; Alcamí, José; Rao, Sudha; Coiras, Mayte

    2016-01-01

    PKCθ is essential for the activation of CD4+ T cells. Upon TCR/CD28 stimulation, PKCθ is phosphorylated and migrates to the immunological synapse, inducing the activation of cellular transcription factors such as NF-κB and kinases as ERK that are critical for HIV-1 replication. We previously demonstrated that PKCθ is also necessary for HIV-1 replication but the precise mechanism is unknown. Efficient HIV-1 transcription and elongation are absolutely dependent on the synergy between NF-κB and the viral regulator Tat. Tat exerts its function by binding a RNA stem-loop structure proximal to the viral mRNA cap site termed TAR. Besides, due to its effect on cellular metabolic pathways, Tat causes profound changes in infected CD4+ T cells such as the activation of NF-κB and ERK. We hypothesized that the aberrant upregulation of Tat-mediated activation of NF-κB and ERK occurred through PKCθ signaling. In fact, Jurkat TetOff cells with stable and doxycycline-repressible expression of Tat (Jurkat-Tat) expressed high levels of mRNA for PKCθ. In these cells, PKCθ located at the plasma membrane was phosphorylated at T538 residue in undivided cells, in the absence of stimulation. Treatment with doxycycline inhibited PKCθ phosphorylation in Jurkat-Tat, suggesting that Tat expression was directly related to the activation of PKCθ. Both NF-κB and Ras/Raf/MEK/ERK signaling pathway were significantly activated in Jurkat-Tat cells, and this correlated with high transactivation of HIV-1 LTR promoter. RNA interference for PKCθ inhibited NF-κB and ERK activity, as well as LTR-mediated transactivation even in the presence of Tat. In addition to Tat-mediated activation of PKCθ in the cytosol, we demonstrated by sequential ChIP that Tat and PKCθ coexisted in the same complex bound at the HIV-1 LTR promoter, specifically at the region containing TAR loop. In conclusion, PKCθ-Tat interaction seemed to be essential for HIV-1 replication in CD4+ T cells and could be used as a

  5. miRNA-149* promotes cell proliferation and suppresses apoptosis by mediating JunB in T-cell acute lymphoblastic leukemia.

    PubMed

    Fan, Sheng-Jin; Li, Hui-Bo; Cui, Gang; Kong, Xiao-Lin; Sun, Li-Li; Zhao, Yan-Qiu; Li, Ying-Hua; Zhou, Jin

    2016-02-01

    MicroRNA-149* (miRNA-149*) functions as an oncogenic regulator in human melanoma. However, the effect of miRNA-149* on T-cell acute lymphoblastic leukemia (T-ALL) is unclear. Here we aimed to analyze the effects of miRNA-149* on in vitro T-ALL cells and to uncover the target for miRNA-149* in these cells. The miRNA-149* level was determined in multiple cell lines and bone marrow cells derived from patients with T-ALL, B acute lymphoblastic leukemia (B-ALL), acute myelocytic leukemia (AML), and healthy donors. We found that miRNA-149* was highly expressed in T-ALL cell lines and T-ALL patients' bone marrow samples. JunB was identified as a direct target of miR-149*. miRNA-149* mimics downregulated JunB levels in Molt-4 and Jurkat cells, while miRNA-149* inhibitors dramatically upregulated JunB expression in these cells. miRNA-149* mimics promoted proliferation, decreased the proportion of cells in G1 phase, and reduced cell apoptosis in T-ALL cells, while miRNA-149* inhibitors prevented these effects. miRNA-149* mimics downregulated p21 and upregulated cyclinD1, 4EBP1, and p70s6k in Molt-4 and Jurkat cells. Again, inhibitors prevented these effects. Our findings demonstrate that miRNA-149* may serve as an oncogenic regulator in T-ALL by negatively regulating JunB. PMID:26725775

  6. [Comparative analysis of natural and synthetic antimutagens as regulators of gene expression in human cells under exposure to ionizing radiation].

    PubMed

    Mikhailov, V F; Shishkina, A A; Vasilyeva, I M; Shulenina, L V; Raeva, N F; Rogozhin, E A; Startsev, M I; Zasukhina, G D; Gromov, S P; Alfimov, M V

    2015-02-01

    This paper studies the effect of plant peptides of thionine Ns-W2 extracted from seeds of fennel flower (Nigella sativa) and β-purothionine from wheat germs (Triticum kiharae), as well as a synthetic antimutagen (crown-compound), on the expression of several genes involved in the.control of cellular homeostasis, processes of carcinogenesis, and radiation response in human rhabdomyosarcoma cells (RD cells), T-lymphoblastoid cell line Jurkat, and blood cells. All of these agents acted as antimutagens-anticarcinogens, reducing the expression of genes involved in carcinogenesis (genes of families MMP, TIMP, and IAP and G-protein genes) in a tumor cell. A pronounced reduction in the mRNA level of these genes was caused by thionine Ns-W2, and the least effect was demonstrated by β-purothionine. Antimutagens had very little effect on the mRNA levels of the several studied genes in normal blood cells. PMID:25966580

  7. Leukemia cell microvesicles promote survival in umbilical cord blood hematopoietic stem cells

    PubMed Central

    Razmkhah, Farnaz; Soleimani, Masoud; Mehrabani, Davood; Karimi, Mohammad Hossein; Kafi-abad, Sedigheh Amini

    2015-01-01

    Microvesicles can transfer their contents, proteins and RNA, to target cells and thereby transform them. This may induce apoptosis or survival depending on cell origin and the target cell. In this study, we investigate the effect of leukemic cell microvesicles on umbilical cord blood hematopoietic stem cells to seek evidence of apoptosis or cell survival. Microvesicles were isolated from both healthy donor bone marrow samples and Jurkat cells by ultra-centrifugation and were added to hematopoietic stem cells sorted from umbilical cord blood samples by magnetic associated cell sorting (MACS) technique. After 7 days, cell count, cell viability, flow cytometry analysis for hematopoietic stem cell markers and qPCR for P53 gene expression were performed. The results showed higher cell number, higher cell viability rate and lower P53 gene expression in leukemia group in comparison with normal and control groups. Also, CD34 expression as the most important hematopoietic stem cell marker, did not change during the treatment and lineage differentiation was not observed. In conclusion, this study showed anti-apoptotic effect of leukemia cell derived microvesicles on umbilical cord blood hematopoietic stem cells. PMID:26862318

  8. Effect of p40tax trans-activator of human T cell lymphotropic virus type I on expression of autoantigens.

    PubMed

    Banki, K; Ablonczy, E; Nakamura, M; Perl, A

    1994-03-01

    The possibility of a retroviral etiology has long been raised in a number of autoimmune disorders. More recently, Sjögren's syndrome and rheumatoid arthritis were noted in transgenic mice carrying the tax gene of human T cell leukemia virus type I (HTLV-I). To evaluate the involvement of HTLV-I Tax in autoimmunity, its effect on expression of autoantigens was investigated. A metallothionein promoter-driven p40tax expression plasmid, pMAXRHneo-1, was stably transfected into Molt4 and Jurkat cells and the p40tax protein was induced with CdCl2. trans-Activation or trans-repression of autoantigens by HTLV-I Tax was studied by Western blot analysis utilizing autoantigen-specific murine monoclonal and rabbit polyvalent antibodies as well as sera from 161 autoimmune patients. Induction of p40tax of HTLV-I had no significant effect on levels of expression of common autoantigens U1 snRNP, Sm, Ro, La, HSP-70, topoisomerase I/Scl70, PCNA, and HRES-1. Expression of two potentially novel autoantigens, 44 and 46 kDa, was induced by p40tax as detected by sera of progressive systemic sclerosis patients, BAK and VAR. By contrast, expression of 24- and 34-kDa proteins was suppressed in response to induction of p40tax as detected by sera of systemic lupus erythematosus patients PUS and HOR. Because none of these patients were infected by HTLV-I, a protein functionally similar to p40tax may be involved in eliciting autoantigen expression and a subsequent autoantibody response in a minority of patients with PSS and SLE. Sera of autoimmune patients may also be utilized to detect novel proteins trans-activated or trans-repressed by p40tax of HTLV-I. PMID:8018391

  9. Synthesis of ZnPc loaded poly(methyl methacrylate) nanoparticles via miniemulsion polymerization for photodynamic therapy in leukemic cells.

    PubMed

    Feuser, Paulo Emilio; Gaspar, Pamela Cristina; Jacques, Amanda Virtuoso; Tedesco, Antônio Claudio; dos Santos Silva, Maria Claudia; Ricci-Júnior, Eduardo; Sayer, Claudia; de Araújo, Pedro Henrique Hermes

    2016-03-01

    The goal of this work was to synthesize and characterize ZnPc loaded poly(methyl methacrylate) (PMMA) nanoparticles (NPs) by miniemulsion polymerization. Biocompatibility assays were performed in murine fibroblast (L929) cells and human peripheral blood lymphocytes (HPBL). Finally, photobiological assays were performed in two leukemic cells: chronic myeloid leukemia in blast crisis (K562) and acute lymphoblastic leukemia (Jurkat). ZnPc loaded PMMA NPs presented an average diameter of 97±2.5 nm with a low polydispersity index and negative surface charge. The encapsulation efficiency (EE %) of ZnPc PMMA NPs was 87%±2.12. The release of ZnPc from PMMA NPs was slow and sustained without the presence of burst effect, indicating homogeneous drug distribution in the polymeric matrix. NP biocompatibility was observed on the treatment of peripheral blood lymphocytes and L929 fibroblast cells. Phototoxicity assays showed that the ZnPc loaded in PMMA NPs was more phototoxic than ZnPc after activation with visible light at 675 nm, using a low light dose of 2J/cm(2) in both leukemic cells (Jurkat and K562). The results from fluorescence microscopy (EB/OA) and DNA fragmentation suggest that the ZnPc loaded PMMA NPs induced cell death by apoptosis. Based on presented results, our study suggests that PDT combined with the use of polymeric NPs, may be an excellent alternative for leukemia treatment. PMID:26706552

  10. Lapatinib induces autophagic cell death and differentiation in acute myeloblastic leukemia

    PubMed Central

    Chen, Yu-Jen; Fang, Li-Wen; Su, Wen-Chi; Hsu, Wen-Yi; Yang, Kai-Chien; Huang, Huey-Lan

    2016-01-01

    Lapatinib is an oral-form dual tyrosine kinase inhibitor of epidermal growth factor receptor (EGFR or ErbB/Her) superfamily members with anticancer activity. In this study, we examined the effects and mechanism of action of lapatinib on several human leukemia cells lines, including acute myeloid leukemia (AML), chronic myeloid leukemia (CML), and acute lymphoblastic leukemia (ALL) cells. We found that lapatinib inhibited the growth of human AML U937, HL-60, NB4, CML KU812, MEG-01, and ALL Jurkat T cells. Among these leukemia cell lines, lapatinib induced apoptosis in HL-60, NB4, and Jurkat cells, but induced nonapoptotic cell death in U937, K562, and MEG-01 cells. Moreover, lapatinib treatment caused autophagic cell death as shown by positive acridine orange staining, the massive formation of vacuoles as seen by electronic microscopy, and the upregulation of LC3-II, ATG5, and ATG7 in AML U937 cells. Furthermore, autophagy inhibitor 3-methyladenine and knockdown of ATG5, ATG7, and Beclin-1 using short hairpin RNA (shRNA) partially rescued lapatinib-induced cell death. In addition, the induction of phagocytosis and ROS production as well as the upregulation of surface markers CD14 and CD68 was detected in lapatinib-treated U937 cells, suggesting the induction of macrophagic differentiation in AML U937 cells by lapatinib. We also noted the synergistic effects of the use of lapatinib and cytotoxic drugs in U937 leukemia cells. These results indicate that lapatinib may have potential for development as a novel antileukemia agent. PMID:27499639

  11. Lapatinib induces autophagic cell death and differentiation in acute myeloblastic leukemia.

    PubMed

    Chen, Yu-Jen; Fang, Li-Wen; Su, Wen-Chi; Hsu, Wen-Yi; Yang, Kai-Chien; Huang, Huey-Lan

    2016-01-01

    Lapatinib is an oral-form dual tyrosine kinase inhibitor of epidermal growth factor receptor (EGFR or ErbB/Her) superfamily members with anticancer activity. In this study, we examined the effects and mechanism of action of lapatinib on several human leukemia cells lines, including acute myeloid leukemia (AML), chronic myeloid leukemia (CML), and acute lymphoblastic leukemia (ALL) cells. We found that lapatinib inhibited the growth of human AML U937, HL-60, NB4, CML KU812, MEG-01, and ALL Jurkat T cells. Among these leukemia cell lines, lapatinib induced apoptosis in HL-60, NB4, and Jurkat cells, but induced nonapoptotic cell death in U937, K562, and MEG-01 cells. Moreover, lapatinib treatment caused autophagic cell death as shown by positive acridine orange staining, the massive formation of vacuoles as seen by electronic microscopy, and the upregulation of LC3-II, ATG5, and ATG7 in AML U937 cells. Furthermore, autophagy inhibitor 3-methyladenine and knockdown of ATG5, ATG7, and Beclin-1 using short hairpin RNA (shRNA) partially rescued lapatinib-induced cell death. In addition, the induction of phagocytosis and ROS production as well as the upregulation of surface markers CD14 and CD68 was detected in lapatinib-treated U937 cells, suggesting the induction of macrophagic differentiation in AML U937 cells by lapatinib. We also noted the synergistic effects of the use of lapatinib and cytotoxic drugs in U937 leukemia cells. These results indicate that lapatinib may have potential for development as a novel antileukemia agent. PMID:27499639

  12. Modulation of HIV transcription by CD8+ cells is mediated via multiple elements of the long terminal repeat

    PubMed Central

    Maslove, D M; Ni, L W; Hawley-Foss, N C; Badley, A D; Copeland, K F T

    2001-01-01

    HIV replication and LTR-mediated gene expression can be modulated by CD8+ cells in a cell type-dependent manner. We have previously shown that supernatant fluids of activated CD8+ cells of HIV-infected individuals suppress long terminal repeat (LTR)-mediated transcription of HIV in T cells while enhancing transcription in monocytic cells. Here, we have examined the effect of culture of T cells and monocytic cells with CD8+ supernatant fluids, and subsequent binding of transcription factors to the HIV-1 LTR. In transfections using constructs in which NFκB or NFAT-1 sites were mutated, the LTR retained the ability to respond positively to culture with CD8 supernatant fluid in monocytic cells. Nuclear extracts prepared from both Jurkat T cells and U38 monocytic cells cultured with CD8+ cell supernatant fluid demonstrated increased binding to the HIV-1 LTR at an AP-1 site which overlapped the chicken ovalbumin upstream promoter (COUP) site. In monocytic cells, increased binding activity was observed at the NFκB sites of the LTR. In contrast, an inhibition in binding at the NFκB sites was observed in Jurkat cells. Examination of two NFAT-1 sites revealed enhanced binding at −260 to −275 bp in U38 cells which was reduced by cellular activation. PMA and ionomycin-induced binding at a second NFAT-1 site (− 205 to −216 bp) was abrogated by CD8+ cell supernatant fluid in T cells. These results, taken together, suggest that factors present in CD8+ supernatant fluids may act through several sites of the LTR to modulate transcription in a cell type-dependant manner. PMID:11472432

  13. Endothelial cell micropatterning: Methods, effects, and applications

    PubMed Central

    Anderson, Deirdre E.J.; Hinds, Monica T.

    2012-01-01

    The effects of flow on endothelial cells have been widely examined for the ability of fluid shear stress to alter cell morphology and function; however, the effects of endothelial cell morphology without flow have only recently been observed. An increase in lithographic techniques in cell culture spurred a corresponding increase in research aiming to confine cell morphology. These studies lead to a better understanding of how morphology and cytoskeletal configuration affect the structure and function of the cells. This review examines endothelial cell micropatterning research by exploring both the many alternative methods used to alter endothelial cell morphology and the resulting changes in cellular shape and phenotype. Micropatterning induced changes in endothelial cell proliferation, apoptosis, cytoskeletal organization, mechanical properties, and cell functionality. Finally, the ways these cellular manipulation techniques have been applied to biomedical engineering research, including angiogenesis, cell migration, and tissue engineering, is discussed. PMID:21761242

  14. The effect of dendritic cells on the retinal cell transplantation

    SciTech Connect

    Oishi, Akio; Nagai, Takayuki; Mandai, Michiko Takahashi, Masayo; Yoshimura, Nagahisa

    2007-11-16

    The potential of bone marrow cell-derived immature dendritic cells (myeloid iDCs) in modulating the efficacy of retinal cell transplantation therapy was investigated. (1) In vitro, myeloid iDCs but not BMCs enhanced the survival and proliferation of embryonic retinal cells, and the expression of various neurotrophic factors by myeloid iDCs was confirmed with RT-PCR. (2) In subretinal transplantation, neonatal retinal cells co-transplanted with myeloid iDCs showed higher survival rate compared to those transplanted without myeloid iDCs. (3) CD8 T-cells reactive against donor retinal cells were significantly increased in the mice with transplantation of retinal cells alone. These results suggested the beneficial effects of the use of myeloid iDCs in retinal cell transplantation therapy.

  15. Molecular Cytotoxic Mechanisms of 1-(3,4,5-Trihydroxyphenyl)-dodecylbenzoate in Human Leukemia Cell Lines

    PubMed Central

    Maioral, M. F.; Bubniak, L.D.S.; Marzarotto, M.A.L.; De Moraes, A.C.R.; Leal, P.; Nunes, R.; Yunes, R. A.; Santos-Silva, M. C.

    2016-01-01

    Recent studies have shown that gallic acid and its alkylesters induce apoptosis in different cell lines. Since new compounds with biological activity and less cytotoxicity to normal cells are necessary for cancer therapy, the aim of this study was to evaluate the cytotoxic effect of 1-(3,4,5-trihydroxyphenyl)-dodecylbenzoate on human acute myeloid leukemia K562 cells and on human acute lymphoblastic leukemia Jurkat cells. The cell viability was determined by MTT method. The apoptosis induction was assessed by bromide and acridine orange staining and by Annexin V-FITC Apoptosis Detection kit. The cell cycle analysis was carried out by flow cytometry using propidium iodide. Cytometric analysis was also performed to evaluate the expression of the following proteins: AIF, p53, Bcl-2 and Bax. The mitochondrial potential was also assessed by flow cytometry using MitoView633 kit. The results showed that the compound significantly reduced the cell viability of K562 and Jurkat cells in a concentration and time dependent manner (IC50 of 30 μM). The compound induced cell cycle arrest in G0/G1phase and significantly increased the proportion of cells in the sub-G0/G1phase. Apoptosis was confirmed by the sight of morphological characteristics of apoptosis and by phosphatidylserine externalization (73.47±5.71% of cells expressing annexin). The results also showed that the compound promotes a modification in Bax:Bcl-2 ratio and increases p53 expression. Thus, it is possible to conclude that 1-(3,4,5-trihydroxyphenyl)-dodecylbenzoate induces apoptosis by inhibiting the antiapoptotic protein Bcl-2 and by increasing the release of AIF, Bax and p53. In addition, it blocks the cell cycle at G0/G1, stopping cell proliferation. So far, the results suggest that this compound may have a potential therapeutic effect against leukemia cells. PMID:27168690

  16. Substrate & Cell Compliance Effects on Cell Spreading and Differentiation

    NASA Astrophysics Data System (ADS)

    Discher, Dennis; Sheehan, Maureen; Engler, Adam

    2004-03-01

    The stiffness of the substrate that a cell adheres to is emerging as a critically important physical factor in the response of many cell types. The effects are seen with cells on gels as well as cells on cells - highlighting implications for organism development. The basis for the effects lies in the fact that cells literally 'feel' their substrate. Like other anchorage dependent cells, muscle cells feel their substrate and are found in our studies to spread more and organize their cytoskeleton and focal adhesions much more so on rigid glass and stiff substrates than on soft gels. Such spreading is not necessarily physiological or conducive to biological function. Collagen density certainly factors into cell on gel adhesive spreading, with minimal spreading on very low collagen and a weak maximum in cell spreading on intermediate collagen densities. Bell-shaped curves are readily modeled to highlight the coupling between ligand density and substrate stiffness. Most surprising, however, spreading on soft gels is found to be almost independent of adhesive ligand density: even with high collagen densities, the minimal spreading of cells cannot be over-ridden. Remarkably, muscle cells show the strongest tendency to differentiate and striate their acto-myosin on gels that have a stiffness similar to relaxed muscle. Cells gown on top of other cells also show a very strong tendency to striate, even if the underlying cells do not striate; elasticity measurements appear to unify all of the effects. The implications for organismal development as well as cell biological studies can be very important.

  17. Immunosuppressive effects of apoptotic cells

    NASA Astrophysics Data System (ADS)

    Voll, Reinhard E.; Herrmann, Martin; Roth, Edith A.; Stach, Christian; Kalden, Joachim R.; Girkontaite, Irute

    1997-11-01

    Apoptotic cell death is important in the development and homeostasis of multicellular organisms and is a highly controlled means of eliminating dangerous, damaged or unnecessary cells without causing an inflammatory response or tissue damage,. We now show that the presence of apoptotic cells during monocyte activation increases their secretion of the anti-inflammatory and immunoregulatory cytokine interleukin 10 (IL-10) and decreases secretion of the proinflammatory cytokines tumour necrosis factor-α (TNF-α), IL-1 and IL-12. This may inhibit inflammation and contribute to impaired cell-mediated immunity in conditions associated with increased apoptosis, such as viral infections, pregnancy, cancer and exposure to radiation.

  18. Paracrine effects of haematopoietic cells on human mesenchymal stem cells

    PubMed Central

    Zhou, Shuanhu

    2015-01-01

    Stem cell function decline during ageing can involve both cell intrinsic and extrinsic mechanisms. Bone and blood formation are intertwined in bone marrow, therefore haematopoietic cells and bone cells could be extrinsic factors for each other. In this study, we assessed the paracrine effects of extrinsic factors from haematopoietic cells on human mesenchymal stem cells (MSCs). Our data showed that haematopoietic cells stimulate proliferation, osteoblast differentiation and inhibit senescence of MSCs; TNF-α, PDGF-β, Wnt1, 4, 6, 7a and 10a, sFRP-3 and sFRP-5 are dominantly expressed in haematopoietic cells; the age-related increase of TNF-α in haematopoietic cells may perform as a negative factor in the interactions of haematopoietic cells on MSCs via TNF-α receptors and then activating NF-κB signaling or Wnt/β-catenin signaling to induce senescence and reduce osteoblast differentiation in MSCs. In conclusion, our data demonstrated that there are paracrine interactions of haematopoietic cells on human MSCs; immunosenescence may be one of the extrinsic mechanisms by which skeletal stem cell function decline during human skeletal ageing. PMID:26030407

  19. Apoptosis and modulation of cell cycle control by bile acids in human leukemia T cells.

    PubMed

    Fimognari, Carmela; Lenzi, Monia; Cantelli-Forti, Giorgio; Hrelia, Patrizia

    2009-08-01

    Depending on the nature of chemical structures, different bile acids exhibit distinct biological effects. Their therapeutic efficacy has been widely demonstrated in various liver diseases, suggesting that they might protect hepatocytes against common mechanisms of liver damage. Although it has been shown to prevent apoptotic cell death in certain cell lines, bile acids significantly inhibited cell growth and induced apoptosis in cancer cells. To better understand the pharmacological potential of bile acids in cancer cells, we investigated and compared the effects of deoxycholic acid (DCA), ursodeoxycholic acid (UDCA), and their taurine-derivatives [taurodeoxycholic acid (TDCA) and tauroursodeoxycholic acid (TUDCA), respectively] on the induction of apoptosis and inhibition of cell proliferation of a human T leukemia cell line (Jurkat cells). All the bile acids tested induced a delay in cell cycle progression. Moreover, DCA markedly increased the fraction of apoptotic cells. The effects of TDCA, UDCA, and TUDCA were different from those observed for DCA. Their primary effect was the induction of necrosis. These distinctive features suggest that the hydrophobic properties of DCA play a role in its cytotoxic potential and indicate that it is possible to create new drugs useful for cancer therapy from bile acid derivatives as lead compounds. PMID:19723064

  20. Anticancer activity of the new photosensitizers: dose and cell type dependence

    NASA Astrophysics Data System (ADS)

    Gyulkhandanyan, Grigor V.; Ghambaryan, Sona S.; Amelyan, Gayane V.; Ghazaryan, Robert K.; Haroutiunian, Samvel G.; Gyulkhandanyan, Aram G.; Gasparyan, Gennadi H.

    2005-04-01

    The necessity of researches of antitumor efficiency of new photosensitizers (PS) is explained by the opportunity of their application in photodynamic therapy of tumors. PS, selectively accumulated in cancer cells and activated by the light, generate the active oxygen species that cause apoptosis. Earlier, it was shown that PS chlorin e6 (0.3-0.5 μg/ml) induces rat embryo fibroblast-like cell apoptosis. In present work antitumor activity of the new photosensitizers, water-soluble cationic porphyrins and their metal complexes, is investigated. The dose-dependent destruction of cancer cells was shown on PC-12 (pheochromocytoma, rat adrenal gland) and Jurkat (human lymphoma) cell lines. Meso-tetra-[4-N-(2 `- oxyethyl) pyridyl] porphyrin (TOEPyP) and chlorin e6 possessed the same toxicity at LD50 dose on PC-12 cell line, whereas phototoxicity of TOEPyP was 3 times less compared to chlorin e6(LD50=0.2 and 0.075 μg/ml accordingly). The results have shown weak photosensitizing effect of Zn-and Ag-derivatives of TOEPyP on PC-12 cell line. TOEPyP and Zn-TOEPyP (0.1 - 50 μg/ml) were non-toxic for Jurkat cell line, whereas Ag-TOEPyP was toxic at 10 μg/ml (LD90). TOEPyP and chlorin e6 have shown phototoxic effect in the same dose range (LD50=0.5 and 0.3 μg/ml accordingly). The investigation of toxic and phototoxic effects of the new porphyrins revealed significantly different sensitivity of various cell lines to PSs.

  1. LDEF solar cell radiation effects analysis

    NASA Technical Reports Server (NTRS)

    Rives, Carol J.; Azarewicz, Joseph L.; Massengill, Lloyd

    1993-01-01

    Because of the extended time that the Long Duration Exposure Facility (LDEF) mission stayed in space, the solar cells on the satellite experienced greater environments than originally planned. The cells showed an overall degradation in performance that is due to the combined effects of the various space environments. The purpose of this analysis is to calculate the effect of the accumulated radiation on the solar cells, thereby helping Marshall Space Flight Center (MSFC) to unravel the relative power degradation from the different environments.

  2. The Interaction of CD154 with the α5β1 Integrin Inhibits Fas-Induced T Cell Death.

    PubMed

    Bachsais, Meriem; Naddaf, Nadim; Yacoub, Daniel; Salti, Suzanne; Alaaeddine, Nada; Aoudjit, Fawzi; Hassan, Ghada S; Mourad, Walid

    2016-01-01

    CD154, a critical regulator of the immune response, is usually associated with chronic inflammatory, autoimmune diseases as well as malignant disorders. In addition to its classical receptor CD40, CD154 is capable of binding other receptors, members of the integrin family, the αIIbβ3, αMβ2 and α5β1. Given the role attributed to integrins and particularly the β1 integrins in inhibiting apoptotic events in normal as well as malignant T cells, we were highly interested in investigating the role of the CD154/α5β1 interaction in promoting survival of malignant T cells contributing as such to tumor development and/or propagation. To support our hypothesis, we first show that soluble CD154 binds to the T-cell acute lymphoblastic leukemia cell line, Jurkat E6.1 in a α5β1-dependent manner. Binding of soluble CD154 to α5β1 integrin of Jurkat cells leads to the activation of key survival proteins, including the p38 and ERK1/2 mitogen-activated protein kinases (MAPKs), phosphoinositide 3 kinase (PI-3K), and Akt. Interestingly, soluble CD154 significantly inhibits Fas-mediated apoptosis in T cell leukemia-lymphoma cell lines, Jurkat E6.1 and HUT78 cells, an important hallmark of T cell survival during malignancy progression. These anti-apoptotic effects were mainly mediated by the activation of the PI-3K/Akt pathway but also involved the p38 and the ERK1/2 MAPKs cascades. Our data also demonstrated that the CD154-triggered inhibition of the Fas-mediated cell death response was dependent on a suppression of caspase-8 cleavage, but independent of de novo protein synthesis or alterations in Fas expression on cell surface. Together, our results highlight the impact of the CD154/α5β1 interaction in T cell function/survival and identify novel targets for the treatment of malignant disorders, particularly of T cell origin. PMID:27391025

  3. The Interaction of CD154 with the α5β1 Integrin Inhibits Fas-Induced T Cell Death

    PubMed Central

    Yacoub, Daniel; Salti, Suzanne; Alaaeddine, Nada; Aoudjit, Fawzi; Hassan, Ghada S.; Mourad, Walid

    2016-01-01

    CD154, a critical regulator of the immune response, is usually associated with chronic inflammatory, autoimmune diseases as well as malignant disorders. In addition to its classical receptor CD40, CD154 is capable of binding other receptors, members of the integrin family, the αIIbβ3, αMβ2 and α5β1. Given the role attributed to integrins and particularly the β1 integrins in inhibiting apoptotic events in normal as well as malignant T cells, we were highly interested in investigating the role of the CD154/α5β1 interaction in promoting survival of malignant T cells contributing as such to tumor development and/or propagation. To support our hypothesis, we first show that soluble CD154 binds to the T-cell acute lymphoblastic leukemia cell line, Jurkat E6.1 in a α5β1-dependent manner. Binding of soluble CD154 to α5β1 integrin of Jurkat cells leads to the activation of key survival proteins, including the p38 and ERK1/2 mitogen-activated protein kinases (MAPKs), phosphoinositide 3 kinase (PI-3K), and Akt. Interestingly, soluble CD154 significantly inhibits Fas-mediated apoptosis in T cell leukemia-lymphoma cell lines, Jurkat E6.1 and HUT78 cells, an important hallmark of T cell survival during malignancy progression. These anti-apoptotic effects were mainly mediated by the activation of the PI-3K/Akt pathway but also involved the p38 and the ERK1/2 MAPKs cascades. Our data also demonstrated that the CD154-triggered inhibition of the Fas-mediated cell death response was dependent on a suppression of caspase-8 cleavage, but independent of de novo protein synthesis or alterations in Fas expression on cell surface. Together, our results highlight the impact of the CD154/α5β1 interaction in T cell function/survival and identify novel targets for the treatment of malignant disorders, particularly of T cell origin. PMID:27391025

  4. The Scaffolding Protein Dlg1 Is a Negative Regulator of Cell-Free Virus Infectivity but Not of Cell-to-Cell HIV-1 Transmission in T Cells

    PubMed Central

    Nzounza, Patrycja; Chazal, Maxime; Guedj, Chloé; Schmitt, Alain; Massé, Jean-Marc; Randriamampita, Clotilde; Pique, Claudine; Ramirez, Bertha Cecilia

    2012-01-01

    Background Cell-to-cell virus transmission of Human immunodeficiency virus type-1 (HIV-1) is predominantly mediated by cellular structures such as the virological synapse (VS). The VS formed between an HIV-1-infected T cell and a target T cell shares features with the immunological synapse (IS). We have previously identified the human homologue of the Drosophila Discs Large (Dlg1) protein as a new cellular partner for the HIV-1 Gag protein and a negative regulator of HIV-1 infectivity. Dlg1, a scaffolding protein plays a key role in clustering protein complexes in the plasma membrane at cellular contacts. It is implicated in IS formation and T cell signaling, but its role in HIV-1 cell-to-cell transmission was not studied before. Methodology/Principal Findings Kinetics of HIV-1 infection in Dlg1-depleted Jurkat T cells show that Dlg1 modulates the replication of HIV-1. Single-cycle infectivity tests show that this modulation does not take place during early steps of the HIV-1 life cycle. Immunofluorescence studies of Dlg1-depleted Jurkat T cells show that while Dlg1 depletion affects IS formation, it does not affect HIV-1-induced VS formation. Co-culture assays and quantitative cell-to-cell HIV-1 transfer analyses show that Dlg1 depletion does not modify transfer of HIV-1 material from infected to target T cells, or HIV-1 transmission leading to productive infection via cell contact. Dlg1 depletion results in increased virus yield and infectivity of the viral particles produced. Particles with increased infectivity present an increase in their cholesterol content and during the first hours of T cell infection these particles induce higher accumulation of total HIV-1 DNA. Conclusion Despite its role in the IS formation, Dlg1 does not affect the VS and cell-to-cell spread of HIV-1, but plays a role in HIV-1 cell-free virus transmission. We propose that the effect of Dlg1 on HIV-1 infectivity is at the stage of virus entry. PMID:22272285

  5. A passive-flow microfluidic device for imaging latent HIV activation dynamics in single T cells

    PubMed Central

    Gearhart, Larisa M.; Miller-Jensen, Kathryn

    2015-01-01

    Quantifying cell-to-cell variability in drug response dynamics is important when evaluating therapeutic efficacy. For example, optimizing latency reversing agents (LRAs) for use in a clinical “activate-and-kill” strategy to purge the latent HIV reservoir in patients requires minimizing heterogeneous viral activation dynamics. To evaluate how heterogeneity in latent HIV activation varies across a range of LRAs, we tracked drug-induced response dynamics in single cells via live-cell imaging using a latent HIV–GFP reporter virus in a clonal Jurkat T cell line. To enable these studies in suspension cells, we designed a simple method to capture an array of single Jurkat T cells using a passive-flow microfluidic device. Our device, which does not require external pumps or tubing, can trap hundreds of cells within minutes with a high retention rate over 12 hours of imaging. Using this device, we quantified heterogeneity in viral activation stimulated by transcription factor (TF) activators and histone deacetylase (HDAC) inhibitors. Generally, TF activators resulted in both faster onset of viral activation and faster rates of production, while HDAC inhibitors resulted in more uniform onset times, but more heterogeneous rates of production. Finally, we demonstrated that while onset time of viral gene expression and rate of viral production together predict total HIV activation, rate and onset time were not correlated within the same individual cell, suggesting that these features are regulated independently. Overall, our results reveal drug-specific patterns of noisy HIV activation dynamics not previously identified in static single-cell assays, which may require consideration for the most effective activate-and-kill regime. PMID:26138068

  6. Effect of peritoneal cells on tumors cells growth in vitro.

    PubMed

    Salwa, J

    1995-01-01

    The cytotoxic and cytostatic activity of PMA-treated macrophages, obtained from pristane-primed BALB/c mice, was analyzed in vitro. The activated macrophages were cytotoxic and cytostatic for YAC-1 lymphoma, P-388 leukemia and P-815 mastocytoma target cells. However, the RPC-5 plasmacytoma target cells appeared to be resistant to their cytotoxicity. The observed cytotoxic or cytostatic effects of macrophages in vitro were not correlated with their ability to produce the superoxide ion. Cytotoxic activity of NK cells, obtained from pristane-primed mice, was also studied. No differences in cytotoxicity of NK cells obtained from pristane-treated and untreated donors, were found. However, only the effector cells from untreated mice were able to respond to stimulatory effect of polyinosinic acid-polycytidylic acid-poly-L-lysine (poly ICLC). PMID:8744682

  7. Hyperglycemic Conditions Prime Cells for RIP1-dependent Necroptosis.

    PubMed

    LaRocca, Timothy J; Sosunov, Sergey A; Shakerley, Nicole L; Ten, Vadim S; Ratner, Adam J

    2016-06-24

    Necroptosis is a RIP1-dependent programmed cell death (PCD) pathway that is distinct from apoptosis. Downstream effector pathways of necroptosis include formation of advanced glycation end products (AGEs) and reactive oxygen species (ROS), both of which depend on glycolysis. This suggests that increased cellular glucose may prime necroptosis. Here we show that exposure to hyperglycemic levels of glucose enhances necroptosis in primary red blood cells (RBCs), Jurkat T cells, and U937 monocytes. Pharmacologic or siRNA inhibition of RIP1 prevented the enhanced death, confirming it as RIP1-dependent necroptosis. Hyperglycemic enhancement of necroptosis depends upon glycolysis with AGEs and ROS playing a role. Total levels of RIP1, RIP3, and mixed lineage kinase domain-like (MLKL) proteins were increased following treatment with high levels of glucose in Jurkat and U937 cells and was not due to transcriptional regulation. The observed increase in RIP1, RIP3, and MLKL protein levels suggests a potential positive feedback mechanism in nucleated cell types. Enhanced PCD due to hyperglycemia was specific to necroptosis as extrinsic apoptosis was inhibited by exposure to high levels of glucose. Hyperglycemia resulted in increased infarct size in a mouse model of brain hypoxia-ischemia injury. The increased infarct size was prevented by treatment with nec-1s, strongly suggesting that increased necroptosis accounts for exacerbation of this injury in conditions of hyperglycemia. This work reveals that hyperglycemia represents a condition in which cells are extraordinarily susceptible to necroptosis, that local glucose levels alter the balance of PCD pathways, and that clinically relevant outcomes may depend on glucose-mediated effects on PCD. PMID:27129772

  8. Mechanisms of JP-8 jet fuel toxicity. I. Induction of apoptosis in rat lung epithelial cells.

    PubMed

    Stoica, B A; Boulares, A H; Rosenthal, D S; Iyer, S; Hamilton, I D; Smulson, M E

    2001-03-01

    JP-8 is a kerosene-based fuel widely used by the U.S. military. Various models of human occupational and animal exposure to JP-8 have demonstrated the potential for local and systemic toxicity but the mechanisms involved are unknown. The purpose of our investigation was to study the molecular mechanisms of JP-8 toxicity by using an in vitro model. JP-8 exposure in a rat lung alveolar type II epithelial cell line (RLE-6TN) induces biochemical and morphological markers of apoptotic cell death: caspase-3 activation, poly(ADP-ribose) polymerase (PARP) cleavage, chromatin condensation, membrane blebbing, cytochrome c release from mitochondria, and genomic DNA cleavage into both oligonucleosomal (DNA ladder) and high-molecular-weight (HMW) fragments. The human histiocytic lymphoma cell line (U937) also responds to JP-8 with caspase-3 activation, cleavage of caspase substrates, including PARP, DNA-PK, and lamin B1, and degradation of genomic DNA with the production of HMW fragments. Caspase-3 activation and PARP cleavage also occur in the acute T-cell leukemia cell line (Jurkat) following treatment with JP-8. Furthermore, Jurkat cells stably transfected with a plasmid encoding the antiapoptotic protein Bcl-x(L) or pretreated with the pan-caspase inhibitor Boc-d-fmk, are relatively resistant to the cytotoxic effects of JP-8 compared to control cells. Finally, we demonstrate that PARP cleavage occurs in primary mouse thymocytes exposed to JP-8. In conclusion, our data support the hypothesis that apoptotic cell death is responsible at least partially for the cytotoxic effects of JP-8 and suggest that inhibition of the apoptotic cascade might reduce JP-8 toxicity. PMID:11222085

  9. Antiproliferative effect of catechin in GRX cells.

    PubMed

    Bragança de Moraes, Cristina Machado; Melo, Denizar Alberto da Silva; Santos, Roberto Christ Vianna; Bitencourt, Shanna; Mesquita, Fernanda Cristina; dos Santos de Oliveira, Fernanda; Rodriguez-Carballo, Edgardo; Bartrons, Ramon; Rosa, Jose Luis; Ventura, Francesc Pujol; Rodrigues de Oliveira, Jarbas

    2012-08-01

    The phenolic compounds present in cocoa seeds have been studied regarding health benefits, such as antioxidant and anti-inflammatory activities. Fibrosis is a wound healing response that occurs in almost all patients with chronic liver injury. A large number of cytokines and soluble intercellular mediators are related to changes in the behavior and phenotype of the hepatic stellate cell (HSC) that develop a fibrogenic and contractile phenotype leading to the development of fibrosis. The objective of this study was to assess the catechin effect in GRX liver cells in activities such as cell growth and inflammation. The GRX cells treatment with catechin induced a significant decrease in cell growth. This mechanism does not occur by apoptosis or even by autophagy because there were no alterations in expression of caspase 3 and PARP (apoptosis), and LC3 (autophagy). The expression of p27 and p53 proteins, regulators of the cell cycle, showed increased expression, while COX-2 and IL-6 mRNA showed a significant decrease in expression. This study shows that catechin decreases cell growth in GRX cells and, probably, this decrease does not occur by apoptosis or autophagy but through an anti-inflammatory effect and cell cycle arrest. Catechin also significantly decreased the production of TGF-β by GRX cells, showing a significant antifibrotic effect. PMID:22574829

  10. Group I PAK inhibitor IPA-3 induces cell death and affects cell adhesivity to fibronectin in human hematopoietic cells.

    PubMed

    Kuželová, Kateřina; Grebeňová, Dana; Holoubek, Aleš; Röselová, Pavla; Obr, Adam

    2014-01-01

    P21-activated kinases (PAKs) are involved in the regulation of multiple processes including cell proliferation, adhesion and migration. However, the current knowledge about their function is mainly based on results obtained in adherent cell types. We investigated the effect of group I PAK inhibition using the compound IPA-3 in a variety of human leukemic cell lines (JURL-MK1, MOLM-7, K562, CML-T1, HL-60, Karpas-299, Jurkat, HEL) as well as in primary blood cells. IPA-3 induced cell death with EC50 ranging from 5 to more than 20 μM. Similar range was found for IPA-3-mediated dephosphorylation of a known PAK downstream effector, cofilin. The cell death was associated with caspase-3 activation, PARP cleavage and apoptotic DNA fragmentation. In parallel, 20 μM IPA-3 treatment induced rapid and marked decrease of the cell adhesivity to fibronectin. Per contra, partial reduction of PAK activity using lower dose IPA-3 or siRNA resulted in a slight increase in the cell adhesivity. The changes in the cell adhesivity were also studied using real-time microimpedance measurement and by interference reflection microscopy. Significant differences in the intracellular IPA-3 level among various cell lines were observed indicating that an active mechanism is involved in IPA-3 transport. PMID:24664099

  11. Effects of stress waves on cells

    SciTech Connect

    Campbell, H L; Da Silva, L B; Visuri, S R

    1998-03-02

    Laser induced stress waves are being used in a variety of medical applications, including drug delivery and targeted tissue disruption. Stress waves can also be an undesirable side effect in laser procedures such as ophthalmology and angioplasty. Thus, a study of the effects of stress waves on a cellular level is useful. Thermoelastic stress waves were produced using a Q-switched frequency-doubled Nd:YAG laser (@.=532nm) with a pulse duration of 4 ns. The laser radiation was delivered to an absorbing media. A thermoelastic stress wave was produced in the absorbing media and propagated into plated cells. The energy per pulse delivered to a sample and the spot size were varied. Stress waves were quantified. We assayed for cell viability and damage using two methods. The laser parameters within which cells maintain viability were investigated and thresholds for cell damage were defined. A comparison of cell damage thresholds for different cell lines was made.

  12. Protein tyrosine kinase inhibition and cell proliferation: is the [3H]-thymidine uptake assay representative of the T-lymphocyte proliferation rate?

    PubMed

    Spinozzi, F; Pagliacci, M C; Agea, E; Migliorati, G; Riccardi, C; Bertotto, A; Nicoletti, I

    1995-01-01

    T-cell growth is controlled to a large degree by extracellular signals that bind to specific receptors on the surface of cells. A number of these receptors have intrinsic protein tyrosine kinase (PTK) activity. Their action on second messenger generation, and thus on cell proliferation, has been indirectly demonstrated by the decrease in [3H]-thymidine (TdR) uptake that follows co-stimulation of T-cells with mitogens and PTK inhibitors such as genistein (GEN). In this paper we report that the [3H]-TdR uptake assay is not a valid and reliable tool for investigating the proliferative activity of certain T-cell lines. In fact, a concomitant assessment of both [3H]-TdR uptake and cell cycle progression demonstrated that GEN is able to block G2/M progression of Jurkat T-lymphocytes even at doses (5 micrograms/ml) that do not influence [3H]-TdR uptake. Pretreatment with sodium o-vanadate (100 nM) could not reverse the GEN-related cell cycle perturbation, but was able to restore optimal [3H]-TdR uptake. Finally, GEN treatment was able to induce concentration-dependent apoptotic cell death of Jurkat T-cells. The control of cell activation, proliferation and programmed cell death is undoubtedly influenced by receptor-associated PTKs. The final effect on cell survival is almost entirely dependent on the activation state of the cell. The [3H]-TdR uptake assay seems to be inadequate for a correct interpretation of the expected results. PMID:7655707

  13. Rescue effects: irradiated cells helped by unirradiated bystander cells.

    PubMed

    Lam, R K K; Fung, Y K; Han, W; Yu, K N

    2015-01-01

    The rescue effect describes the phenomenon where irradiated cells or organisms derive benefits from the feedback signals sent from the bystander unirradiated cells or organisms. An example of the benefit is the mitigation of radiation-induced DNA damages in the irradiated cells. The rescue effect can compromise the efficacy of radioimmunotherapy (RIT) (and actually all radiotherapy). In this paper, the discovery and subsequent confirmation studies on the rescue effect were reviewed. The mechanisms and the chemical messengers responsible for the rescue effect studied to date were summarized. The rescue effect between irradiated and bystander unirradiated zebrafish embryos in vivo sharing the same medium was also described. In the discussion section, the mechanism proposed for the rescue effect involving activation of the nuclear factor κB (NF-κB) pathway was scrutinized. This mechanism could explain the promotion of cellular survival and correct repair of DNA damage, dependence on cyclic adenosine monophosphate (cAMP) and modulation of intracellular reactive oxygen species (ROS) level in irradiated cells. Exploitation of the NF-κB pathway to improve the effectiveness of RIT was proposed. Finally, the possibility of using zebrafish embryos as the model to study the efficacy of RIT in treating solid tumors was also discussed. PMID:25625514

  14. Rescue Effects: Irradiated Cells Helped by Unirradiated Bystander Cells

    PubMed Central

    Lam, R. K. K.; Fung, Y. K.; Han, W.; Yu, K. N.

    2015-01-01

    The rescue effect describes the phenomenon where irradiated cells or organisms derive benefits from the feedback signals sent from the bystander unirradiated cells or organisms. An example of the benefit is the mitigation of radiation-induced DNA damages in the irradiated cells. The rescue effect can compromise the efficacy of radioimmunotherapy (RIT) (and actually all radiotherapy). In this paper, the discovery and subsequent confirmation studies on the rescue effect were reviewed. The mechanisms and the chemical messengers responsible for the rescue effect studied to date were summarized. The rescue effect between irradiated and bystander unirradiated zebrafish embryos in vivo sharing the same medium was also described. In the discussion section, the mechanism proposed for the rescue effect involving activation of the nuclear factor κB (NF-κB) pathway was scrutinized. This mechanism could explain the promotion of cellular survival and correct repair of DNA damage, dependence on cyclic adenosine monophosphate (cAMP) and modulation of intracellular reactive oxygen species (ROS) level in irradiated cells. Exploitation of the NF-κB pathway to improve the effectiveness of RIT was proposed. Finally, the possibility of using zebrafish embryos as the model to study the efficacy of RIT in treating solid tumors was also discussed. PMID:25625514

  15. Downregulation of NPM-ALK by siRNA causes anaplastic large cell lymphoma cell growth inhibition and augments the anti cancer effects of chemotherapy in vitro.

    PubMed

    Hsu, Faye Yuan-yi; Zhao, Yi; Anderson, W French; Johnston, Patrick B

    2007-06-01

    The fusion protein, nucleophosmin-anaplastic lymphoma kinase (NPM-ALK), results from the chromosome translocation t(2;5)(p23;q25) and is present in 50-70 percent of anaplastic large-cell lymphomas (ALCLs). NPM-ALK is a constitutively activated kinase that transforms cells through stimulating several mitogenic signaling pathways. To examine if the NPM-ALK is a potential therapeutic target in ALCL, we used siRNA to specifically downregulate the expression of the NPM-ALK in ALCL cell lines. In this report, we demonstrated viability loss in t(2;5)-positive ALCL cell lines, SUDHL-1 and Karpas 299 cells, but not in lymphoma cell lines without the chromosome translocation, Jurkat and Granta 519 cells. Further study demonstrated that the downregulation of NPM-ALK resulted in decreased cell proliferation and increased cell apoptosis. When used in combination with chemotherapeutic agents, such as doxorubicin, the inhibition of the NPM-ALK augments the chemosensitivity of the tumor cells. These results revealed the importance of continuous expression of NPM-ALK in maintaining the growth of ALCL cells. Our data also suggested that the repression of the fusion gene might be a potential novel therapeutic strategy for NPM-ALK positive ALCLs. PMID:17612934

  16. Targeting TRPM2 Channels Impairs Radiation-Induced Cell Cycle Arrest and Fosters Cell Death of T Cell Leukemia Cells in a Bcl-2-Dependent Manner

    PubMed Central

    Klumpp, Dominik; Misovic, Milan; Szteyn, Kalina; Shumilina, Ekaterina; Rudner, Justine; Huber, Stephan M.

    2016-01-01

    Messenger RNA data of lymphohematopoietic cancer lines suggest a correlation between expression of the cation channel TRPM2 and the antiapoptotic protein Bcl-2. The latter is overexpressed in various tumor entities and mediates therapy resistance. Here, we analyzed the crosstalk between Bcl-2 and TRPM2 channels in T cell leukemia cells during oxidative stress as conferred by ionizing radiation (IR). To this end, the effects of TRPM2 inhibition or knock-down on plasma membrane currents, Ca2+ signaling, mitochondrial superoxide anion formation, and cell cycle progression were compared between irradiated (0–10 Gy) Bcl-2-overexpressing and empty vector-transfected Jurkat cells. As a result, IR stimulated a TRPM2-mediated Ca2+-entry, which was higher in Bcl-2-overexpressing than in control cells and which contributed to IR-induced G2/M cell cycle arrest. TRPM2 inhibition induced a release from G2/M arrest resulting in cell death. Collectively, this data suggests a pivotal function of TRPM2 in the DNA damage response of T cell leukemia cells. Apoptosis-resistant Bcl-2-overexpressing cells even can afford higher TRPM2 activity without risking a hazardous Ca2+-overload-induced mitochondrial superoxide anion formation. PMID:26839633

  17. Cell cycle-specific effects of lovastatin.

    PubMed Central

    Jakóbisiak, M; Bruno, S; Skierski, J S; Darzynkiewicz, Z

    1991-01-01

    Lovastatin (LOV), the drug recently introduced to treat hypercholesteremia, inhibits the synthesis of mevalonic acid. The effects of LOV on the cell cycle progression of the human bladder carcinoma T24 cell line expressing activated p21ras were investigated. At a concentration of 2-10 microM, LOV arrested cells in G1 and also prolonged--or arrested a minor fraction of cells in--the G2 phase of the cell cycle; at a concentration of 50 microM, LOV was cytotoxic. The cytostatic effects were reversed by addition of exogenous mevalonate. Cells arrested in the cycle by LOV were viable for up to 72 hr and did not show any changes in RNA or protein content or chromatin condensation, which would be typical of either unbalanced growth or deep quiescence. The expression of the proliferation-associated nuclear proteins Ki-67 and p105 in these cells was reduced by up to 72% and 74%, respectively, compared with exponentially growing control cells. After removal of LOV, the cells resumed progression through the cycle; they entered S phase asynchronously after a lag of approximately 6 hr. Because mevalonate is essential for the posttranslational modification (isoprenylation) of p21ras, which in turn allows this protein to become attached to the cell membrane, the data suggest that the LOV-induced G1 arrest may be a consequence of the loss of the signal transduction capacity of p21ras. Indeed, while exposure of cells to LOV had no effect on the cellular content of p21ras (detected immunocytochemically), it altered the intracellular location of this protein, causing its dissociation from the cell membrane and translocation toward the cytoplasm and nucleus. However, it is also possible that inhibition of isoprenylation of proteins other than p21ras (e.g., nuclear lamins) by LOV may be responsible for the observed suppression of growth of T24 cells. Images PMID:1673788

  18. Activated T cells exhibit increased uptake of silicon phthalocyanine Pc 4 and increased susceptibility to Pc 4-photodynamic therapy-mediated cell death.

    PubMed

    Soler, David C; Ohtola, Jennifer; Sugiyama, Hideaki; Rodriguez, Myriam E; Han, Ling; Oleinick, Nancy L; Lam, Minh; Baron, Elma D; Cooper, Kevin D; McCormick, Thomas S

    2016-06-01

    Photodynamic therapy (PDT) is an emerging treatment for malignant and inflammatory dermal disorders. Photoirradiation of the silicon phthalocyanine (Pc) 4 photosensitizer with red light generates singlet oxygen and other reactive oxygen species to induce cell death. We previously reported that Pc 4-PDT elicited cell death in lymphoid-derived (Jurkat) and epithelial-derived (A431) cell lines in vitro, and furthermore that Jurkat cells were more sensitive than A431 cells to treatment. In this study, we examined the effectiveness of Pc 4-PDT on primary human CD3(+) T cells in vitro. Fluorometric analyses of lysed T cells confirmed the dose-dependent uptake of Pc 4 in non-stimulated and stimulated T cells. Flow cytometric analyses measuring annexin V and propidium iodide (PI) demonstrated a dose-dependent increase of T cell apoptosis (6.6-59.9%) at Pc 4 doses ranging from 0-300 nM. Following T cell stimulation through the T cell receptor using a combination of anti-CD3 and anti-CD28 antibodies, activated T cells exhibited increased susceptibility to Pc 4-PDT-induced apoptosis (10.6-81.2%) as determined by Pc 4 fluorescence in each cell, in both non-stimulated and stimulated T cells, Pc 4 uptake increased with Pc 4 dose up to 300 nM as assessed by flow cytometry. The mean fluorescence intensity (MFI) of Pc 4 uptake measured in stimulated T cells was significantly increased over the uptake of resting T cells at each dose of Pc 4 tested (50, 100, 150 and 300 nM, p < 0.001 between 50 and 150 nM, n = 8). Treg uptake was diminished relative to other T cells. Cutaneous T cell lymphoma (CTCL) T cells appeared to take up somewhat more Pc 4 than normal resting T cells at 100 and 150 nm Pc 4. Confocal imaging revealed that Pc 4 localized in cytoplasmic organelles, with approximately half of the Pc 4 co-localized with mitochondria in T cells. Thus, Pc 4-PDT exerts an enhanced apoptotic effect on activated CD3(+) T cells that may be exploited in targeting T cell-mediated skin

  19. Aronia melanocarpa juice induces a redox-sensitive p73-related caspase 3-dependent apoptosis in human leukemia cells.

    PubMed

    Sharif, Tanveer; Alhosin, Mahmoud; Auger, Cyril; Minker, Carole; Kim, Jong-Hun; Etienne-Selloum, Nelly; Bories, Pierre; Gronemeyer, Hinrich; Lobstein, Annelise; Bronner, Christian; Fuhrmann, Guy; Schini-Kerth, Valérie B

    2012-01-01

    Polyphenols are natural compounds widely present in fruits and vegetables, which have antimutagenic and anticancer properties. The aim of the present study was to determine the anticancer effect of a polyphenol-rich Aronia melanocarpa juice (AMJ) containing 7.15 g/L of polyphenols in the acute lymphoblastic leukemia Jurkat cell line, and, if so, to clarify the underlying mechanism and to identify the active polyphenols involved. AMJ inhibited cell proliferation, which was associated with cell cycle arrest in G(2)/M phase, and caused the induction of apoptosis. These effects were associated with an upregulation of the expression of tumor suppressor p73 and active caspase 3, and a downregulation of the expression of cyclin B1 and the epigenetic integrator UHRF1. AMJ significantly increased the formation of reactive oxygen species (ROS), decreased the mitochondrial membrane potential and caused the release of cytochrome c into the cytoplasm. Treatment with intracellular ROS scavengers prevented the AMJ-induced apoptosis and upregulation of the expression of p73 and active caspase 3. The fractionation of the AMJ and the use of identified isolated compounds indicated that the anticancer activity was associated predominantly with chlorogenic acids, some cyanidin glycosides, and derivatives of quercetin. AMJ treatment also induced apoptosis of different human lymphoblastic leukemia cells (HSB-2, Molt-4 and CCRF-CEM). In addition, AMJ exerted a strong pro-apoptotic effect in human primary lymphoblastic leukemia cells but not in human normal primary T-lymphocytes. Thus, the present findings indicate that AMJ exhibits strong anticancer activity through a redox-sensitive mechanism in the p53-deficient Jurkat cells and that this effect involves several types of polyphenols. They further suggest that AMJ has chemotherapeutic properties against acute lymphoblastic leukemia by selectively targeting lymphoblast-derived tumor cells. PMID:22412883

  20. Aronia melanocarpa Juice Induces a Redox-Sensitive p73-Related Caspase 3-Dependent Apoptosis in Human Leukemia Cells

    PubMed Central

    Sharif, Tanveer; Alhosin, Mahmoud; Auger, Cyril; Minker, Carole; Kim, Jong-Hun; Etienne-Selloum, Nelly; Bories, Pierre; Gronemeyer, Hinrich; Lobstein, Annelise; Bronner, Christian; Fuhrmann, Guy; Schini-Kerth, Valérie B.

    2012-01-01

    Polyphenols are natural compounds widely present in fruits and vegetables, which have antimutagenic and anticancer properties. The aim of the present study was to determine the anticancer effect of a polyphenol-rich Aronia melanocarpa juice (AMJ) containing 7.15 g/L of polyphenols in the acute lymphoblastic leukemia Jurkat cell line, and, if so, to clarify the underlying mechanism and to identify the active polyphenols involved. AMJ inhibited cell proliferation, which was associated with cell cycle arrest in G2/M phase, and caused the induction of apoptosis. These effects were associated with an upregulation of the expression of tumor suppressor p73 and active caspase 3, and a downregulation of the expression of cyclin B1 and the epigenetic integrator UHRF1. AMJ significantly increased the formation of reactive oxygen species (ROS), decreased the mitochondrial membrane potential and caused the release of cytochrome c into the cytoplasm. Treatment with intracellular ROS scavengers prevented the AMJ-induced apoptosis and upregulation of the expression of p73 and active caspase 3. The fractionation of the AMJ and the use of identified isolated compounds indicated that the anticancer activity was associated predominantly with chlorogenic acids, some cyanidin glycosides, and derivatives of quercetin. AMJ treatment also induced apoptosis of different human lymphoblastic leukemia cells (HSB-2, Molt-4 and CCRF-CEM). In addition, AMJ exerted a strong pro-apoptotic effect in human primary lymphoblastic leukemia cells but not in human normal primary T-lymphocytes. Thus, the present findings indicate that AMJ exhibits strong anticancer activity through a redox-sensitive mechanism in the p53-deficient Jurkat cells and that this effect involves several types of polyphenols. They further suggest that AMJ has chemotherapeutic properties against acute lymphoblastic leukemia by selectively targeting lymphoblast-derived tumor cells. PMID:22412883

  1. Locally-Delivered T-Cell-Derived Cellular Vehicles Efficiently Track and Deliver Adenovirus Delta24-RGD to Infiltrating Glioma

    PubMed Central

    Balvers, Rutger K.; Belcaid, Zineb; van den Hengel, Sanne K.; Kloezeman, Jenneke; de Vrij, Jeroen; Wakimoto, Hiroaki; Hoeben, Rob C.; Debets, Reno; Leenstra, Sieger; Dirven, Clemens; Lamfers, Martine L.M.

    2014-01-01

    Oncolytic adenoviral vectors are a promising alternative for the treatment of glioblastoma. Recent publications have demonstrated the advantages of shielding viral particles within cellular vehicles (CVs), which can be targeted towards the tumor microenvironment. Here, we studied T-cells, often having a natural capacity to target tumors, for their feasibility as a CV to deliver the oncolytic adenovirus, Delta24-RGD, to glioblastoma. The Jurkat T-cell line was assessed in co-culture with the glioblastoma stem cell (GSC) line, MGG8, for the optimal transfer conditions of Delta24-RGD in vitro. The effect of intraparenchymal and tail vein injections on intratumoral virus distribution and overall survival was addressed in an orthotopic glioma stem cell (GSC)-based xenograft model. Jurkat T-cells were demonstrated to facilitate the amplification and transfer of Delta24-RGD onto GSCs. Delta24-RGD dosing and incubation time were found to influence the migratory ability of T-cells towards GSCs. Injection of Delta24-RGD-loaded T-cells into the brains of GSC-bearing mice led to migration towards the tumor and dispersion of the virus within the tumor core and infiltrative zones. This occurred after injection into the ipsilateral hemisphere, as well as into the non-tumor-bearing hemisphere. We found that T-cell-mediated delivery of Delta24-RGD led to the inhibition of tumor growth compared to non-treated controls, resulting in prolonged survival (p = 0.007). Systemic administration of virus-loaded T-cells resulted in intratumoral viral delivery, albeit at low levels. Based on these findings, we conclude that T-cell-based CVs are a feasible approach to local Delta24-RGD delivery in glioblastoma, although efficient systemic targeting requires further improvement. PMID:25118638

  2. Effects of cell-cell contact and oxygen tension on chondrogenic differentiation of stem cells.

    PubMed

    Cao, Bin; Li, Zhenhua; Peng, Rong; Ding, Jiandong

    2015-09-01

    While cell condensation has been thought to enhance chondrogenesis, no direct evidence so far confirms that cell-cell contact itself increases chondrogenic differentiation of stem cells, since the change of cell-cell contact is usually coupled with those of other cell geometry cues and soluble factors in cell culture. The present study semi-quantitatively examined the effect of cell-cell contact in a decoupled way. We fabricated two-dimensional micropatterns with cell-adhesive peptide arginine-glycine-aspartate (RGD) microdomains on a nonfouling poly(ethylene glycol) (PEG) hydrogel. Mesenchymal stem cells (MSCs) were well localized on the microdomains for a long time. Based on our micropattern design, single MSCs or cell clusters with given cell numbers (1, 2, 3, 6 and 15) and a similar spreading area per cell were achieved on the same substrate, thus the interference of soluble factor difference from cell autocrine and that of cell spreading area were ruled out. After 9-day chondrogenic induction, collagen II was stained to characterize the chondrogenic induction results; the mRNA expression levels of SOX9, collagen II, aggrecan, HIF-1α and collagen I were also detected. The statistics confirmed unambiguously that the extent of the chondrogenic differentiation increased with cell-cell contact, and even a linear relation between differentiation extent and contact extent was established within the examined range. The cell-cell contact effect worked under both hypoxia (5% O2) and normoxia (21% O2) conditions, and the hypoxia condition promoted the chondrogenic induction of MSCs on adhesive microdomains more efficiently than the normoxia condition under the same cell-cell contact extents. PMID:26113183

  3. Protease inhibitors effectively block cell-to-cell spread of HIV-1 between T cells

    PubMed Central

    2013-01-01

    Background The Human Immunodeficiency Virus type-1 (HIV-1) spreads by cell-free diffusion and by direct cell-to-cell transfer, the latter being a significantly more efficient mode of transmission. Recently it has been suggested that cell-to-cell spread may permit ongoing virus replication in the presence of antiretroviral therapy (ART) based on studies performed using Reverse Transcriptase Inhibitors (RTIs). Protease Inhibitors (PIs) constitute an important component of ART; however whether this class of inhibitors can suppress cell-to-cell transfer of HIV-1 is unexplored. Here we have evaluated the inhibitory effect of PIs during cell-to-cell spread of HIV-1 between T lymphocytes. Results Using quantitative assays in cell line and primary cell systems that directly measure the early steps of HIV-1 infection we find that the PIs Lopinavir and Darunavir are equally potent against both cell-free and cell-to-cell spread of HIV-1. We further show that a protease resistant mutant maintains its resistant phenotype during cell-to-cell spread and is transmitted more efficiently than wild-type virus in the presence of drug. By contrast we find that T cell-T cell spread of HIV-1 is 4–20 fold more resistant to inhibition by the RTIs Nevirapine, Zidovudine and Tenofovir. Notably, varying the ratio of infected and uninfected cells in co-culture impacted on the degree of inhibition, indicating that the relative efficacy of ART is dependent on the multiplicity of infection. Conclusions We conclude that if the variable effects of antiviral drugs on cell-to-cell virus dissemination of HIV-1 do indeed impact on viral replication and maintenance of viral reservoirs this is likely to be influenced by the antiviral drug class, since PIs appear particularly effective against both modes of HIV-1 spread. PMID:24364896

  4. Cytoprotective dibenzoylmethane derivatives protect cells from oxidative stress-induced necrotic cell death.

    PubMed

    Hegedűs, Csaba; Lakatos, Petra; Kiss-Szikszai, Attila; Patonay, Tamás; Gergely, Szabolcs; Gregus, Andrea; Bai, Péter; Haskó, György; Szabó, Éva; Virág, László

    2013-06-01

    Screening of a small in-house library of 1863 compounds identified 29 compounds that protected Jurkat cells from hydrogen peroxide-induced cytotoxicity. From the cytoprotective compounds eleven proved to possess antioxidant activity (ABTS radical scavenger effect) and two were found to inhibit poly(ADP-ribosyl)ation (PARylation), a cytotoxic pathway operating in severely injured cells. Four cytoprotective dibenzoylmethane (DBM) derivatives were investigated in more detail as they did not scavenge hydrogen peroxide nor did they inhibit PARylation. These compounds protected cells from necrotic cell death while caspase activation, a parameter of apoptotic cell death was not affected. Hydrogen peroxide activated extracellular signal regulated kinase (ERK1/2) and p38 MAP kinases but not c-Jun N-terminal kinase (JNK). The cytoprotective DBMs suppressed the activation of Erk1/2 but not that of p38. Cytoprotection was confirmed in another cell type (A549 lung epithelial cells), indicating that the cytoprotective effect is not cell type specific. In conclusion we identified DBM analogs as a novel class of cytoprotective compounds inhibiting ERK1/2 kinase and protecting from necrotic cell death by a mechanism independent of poly(ADP-ribose) polymerase inhibition. PMID:23523665

  5. Cytotoxicity effects of amiodarone on cultured cells.

    PubMed

    Golli-Bennour, Emna El; Bouslimi, Amel; Zouaoui, Olfa; Nouira, Safa; Achour, Abdellatif; Bacha, Hassen

    2012-07-01

    Amiodarone is a potent anti-arrhythmic drug used for the treatment of cardiac arrhythmias. Although, the effects of amiodarone are well characterized on post-ischemic heart and cardiomyocytes, its toxicity on extra-cardiac tissues is still poorly understood. To this aim, we have monitored the cytotoxicity effects of this drug on three cultured cell lines including hepatocytes (HepG2), epithelial cells (EAhy 926) and renal cells (Vero). We have investigated the effects of amiodarone on (i) cell viabilities, (ii) heat shock protein expressions (Hsp 70) as a parameter of protective and adaptive response and (iii) oxidative damage.Our results clearly showed that amiodarone inhibits cell proliferation, induces an over-expression of Hsp 70 and generates significant amount of reactive oxygen species as measured by lipid peroxidation occurrence. However, toxicity of amiodarone was significantly higher in renal and epithelial cells than in hepatocytes. Vitamin E supplement restores the major part of cell mortalities induced by amiodarone showing that oxidative damage is the predominant toxic effect of the drug.Except its toxicity for the cardiac system, our findings demonstrated that amiodarone can target other tissues. Therefore, kidneys present a high sensibility to this drug which may limit its use with subjects suffering from renal disorders. PMID:21093234

  6. Anticancer effect of arsenite on cell migration, cell cycle and apoptosis in human pancreatic cancer cells

    PubMed Central

    HORIBE, YOHEI; ADACHI, SEIJI; YASUDA, ICHIRO; YAMAUCHI, TAKAHIRO; KAWAGUCHI, JUNJI; KOZAWA, OSAMU; SHIMIZU, MASAHITO; MORIWAKI, HISATAKA

    2016-01-01

    The standard treatment for advanced pancreatic cancer is chemotherapy, but its clinical outcome remains unsatisfactory. Therefore, the development of novel treatments for this malignancy is urgently required. In the present study, the anticancer effect of arsenite on platelet-derived growth factor (PDGF)-BB-induced migration, cell cycle and apoptosis was investigated in pancreatic cancer cells (AsPC-1 and BxPC-3), and compared with the effect on normal pancreatic epithelial (PE) cells. In the cell migration assay, arsenite clearly inhibited PDGF-BB-induced cell migration in AsPC-1 cells, but not in BxPC-3 or PE cells. Arsenite also caused cell apoptosis in AsPC-1 cells, but not in BxPC-3 or PE cells. In AsPC-1 cells, the levels of cyclin D1 and phosphorylated retinoblastoma protein decreased following treatment with arsenite, but this was not observed in BxPC-3 cells. To further examine the differences between these two cell lines, the effect of arsenite on upstream p44/p42 mitogen-activated protein kinase (MAPK) and Akt was investigated. PDGF-BB caused phosphorylation of p44/p42 MAPK and Akt in both cell lines. Pretreatment with arsenite significantly suppressed PDGF-BB-induced phosphorylation of Akt, but not of p44/p42 MAPK in AsPC-1 cells. By contrast, arsenite did not affect these molecules in BxPC-3 cells. Since the inhibition of the Akt signaling pathway markedly reduced PDGF-BB-induced migration in AsPC-1 cells, the present results strongly suggest that arsenite inhibits PDGF-BB-induced migration by suppressing the Akt signaling pathway in AsPC-1 cells. Therefore, arsenite may be a useful tool for the treatment of patients with certain types of pancreatic cancer, without causing adverse effects on normal pancreatic cells. PMID:27347121

  7. Effect of chaetocin on renal cell carcinoma cells and cytokine-induced killer cells

    PubMed Central

    Rombo, Roman; Weiher, Hans; Schmidt-Wolf, Ingo G.H.

    2016-01-01

    We examined the cytotoxic effects of chaetocin on clear cell renal cell carcinoma (ccRCC) cells and the possibility to combine the effects of chaetocin with the effects of cytokine-induced killer cells (CIK) assayed by MTT assay and FACS analysis. Chaetocin is a thiodioxopiperazine produced by fungi belonging to the chaetomiaceae family. In 2007, it was first reported that chaetocin shows potent and selective ex vivo anti-cancer activity by inducing reactive oxygen species. CIK cells are generated from CD3+/CD56- T lymphocytes with double negative CD4-/CD8- phenotype that are isolated from human blood. The addition of distinct interleukins and antibodies results in the generation of CIK cells that are able to specifically target and destroy renal carcinoma cells. The results of this research state that the anti-ccRCC activity of chaetocin is weak and does not show a high grade of selectivity on clear cell renal cell carcinoma cells. Although the CIK cells show a high grade of selective anti-ccRCC activity, this effect could not be improved by the addition of chaetocin. So chaetocin seems to be no suitable agent for specific targeting ccRCC cells or for the combination therapy with CIK cells in renal cancer. PMID:27141211

  8. Upregulation of PTEN involved in scorpion venom-induced apoptosis in a lymphoma cell line.

    PubMed

    Gao, Fang; Li, Hao; Chen, Ya-Dong; Yu, Xiao-Ning; Wang, Ran; Chen, Xue-Liang

    2009-04-01

    We investigated whether the venom of the scorpion Buthus martensii Karsch (BmK) inhibited growth of human lymphoma cells by inducing apoptosis, and studied possible signal pathways involved in this cell death. BmK venom selectively reduced the viability of Raji and Jurkat cells, and had low toxicity to human peripheral blood lymphocytes. Flow cytometry showed that BmK venom-induced apoptosis and G(0)/G(1) cell cycle arrest in Raji and Jurkat cells. In Raji cells, BmK venom upregulated the expression of PTEN accompanied by decreased levels of Akt and Bad phosphorylation. Treatment with BmK venom and LY294002 (an inhibitor of Akt) synergistically enhanced apoptosis. The expression of p27 was increased in both PTEN-positive Raji and PTEN-negative Jurkat cells exposed to BmK venom. The results indicate that key regulators in BmK venom-induced apoptosis are PTEN, acting through downregulation of the PI3K/Akt signal pathway, in Raji cells and p27 in Jurkat cells. PMID:19373662

  9. Effects of Helicobacter suis γ-glutamyl transpeptidase on lymphocytes: modulation by glutamine and glutathione supplementation and outer membrane vesicles as a putative delivery route of the enzyme.

    PubMed

    Zhang, Guangzhi; Ducatelle, Richard; Pasmans, Frank; D'Herde, Katharina; Huang, Liping; Smet, Annemieke; Haesebrouck, Freddy; Flahou, Bram

    2013-01-01

    Helicobacter (H.) suis colonizes the stomach of the majority of pigs as well as a minority of humans worldwide. Infection causes chronic inflammation in the stomach of the host, however without an effective clearance of the bacteria. Currently, no information is available about possible mechanisms H. suis utilizes to interfere with the host immune response. This study describes the effect on various lymphocytes of the γ-glutamyl transpeptidase (GGT) from H. suis. Compared to whole cell lysate from wild-type H. suis, lysate from a H. suis ggt mutant strain showed a decrease of the capacity to inhibit Jurkat T cell proliferation. Incubation of Jurkat T cells with recombinantly expressed H. suis GGT resulted in an impaired proliferation, and cell death was shown to be involved. A similar but more pronounced inhibitory effect was also seen on primary murine CD4(+) T cells, CD8(+) T cells, and CD19(+) B cells. Supplementation with known GGT substrates was able to modulate the observed effects. Glutamine restored normal proliferation of the cells, whereas supplementation with reduced glutathione strengthened the H. suis GGT-mediated inhibition of proliferation. H. suis GGT treatment abolished secretion of IL-4 and IL-17 by CD4(+) T cells, without affecting secretion of IFN-γ. Finally, H. suis outer membrane vesicles (OMV) were identified as a possible delivery route of H. suis GGT to lymphocytes residing in the deeper mucosal layers. Thus far, this study is the first to report that the effects on lymphocytes of this enzyme, not only important for H. suis metabolism but also for that of other Helicobacter species, depend on the degradation of two specific substrates: glutamine and reduced glutatione. This will provide new insights into the pathogenic mechanisms of H. suis infection in particular and infection with gastric helicobacters in general. PMID:24147103

  10. Effects of Helicobacter suis γ- Glutamyl Transpeptidase on Lymphocytes: Modulation by Glutamine and Glutathione Supplementation and Outer Membrane Vesicles as a Putative Delivery Route of the Enzyme

    PubMed Central

    Zhang, Guangzhi; Ducatelle, Richard; Pasmans, Frank; D’Herde, Katharina; Huang, Liping; Smet, Annemieke; Haesebrouck, Freddy; Flahou, Bram

    2013-01-01

    Helicobacter (H.) suis colonizes the stomach of the majority of pigs as well as a minority of humans worldwide. Infection causes chronic inflammation in the stomach of the host, however without an effective clearance of the bacteria. Currently, no information is available about possible mechanisms H. suis utilizes to interfere with the host immune response. This study describes the effect on various lymphocytes of the γ-glutamyl transpeptidase (GGT) from H. suis. Compared to whole cell lysate from wild-type H. suis, lysate from a H. suis ggt mutant strain showed a decrease of the capacity to inhibit Jurkat T cell proliferation. Incubation of Jurkat T cells with recombinantly expressed H. suis GGT resulted in an impaired proliferation, and cell death was shown to be involved. A similar but more pronounced inhibitory effect was also seen on primary murine CD4+ T cells, CD8+ T cells, and CD19+ B cells. Supplementation with known GGT substrates was able to modulate the observed effects. Glutamine restored normal proliferation of the cells, whereas supplementation with reduced glutathione strengthened the H. suis GGT-mediated inhibition of proliferation. H. suis GGT treatment abolished secretion of IL-4 and IL-17 by CD4+ T cells, without affecting secretion of IFN-γ. Finally, H. suis outer membrane vesicles (OMV) were identified as a possible delivery route of H. suis GGT to lymphocytes residing in the deeper mucosal layers. Thus far, this study is the first to report that the effects on lymphocytes of this enzyme, not only important for H. suis metabolism but also for that of other Helicobacter species, depend on the degradation of two specific substrates: glutamine and reduced glutatione. This will provide new insights into the pathogenic mechanisms of H. suis infection in particular and infection with gastric helicobacters in general. PMID:24147103